Nano Materials Lectures

Web-based Selected Lecture Series
Nanomaterials and Bioapplications
Course Number: Bioengineering ECH 4657 and BCE 4003c and 4004c
Florida State University and Florida A&M University
Elective Course: EE 253 MP A&T University, Udaipur, Rajasthan, India
UNIT I: NANOMATERIALS AND PROPERTIES

UNIT II: NANOMATERIAL SYNTHESIS AND CHARACTERIZATION
UNIT III: BIOAPPLICATIONS IN MATERIAL SCIENCE
STATUS: PRIVATE MATERIAL ONLY FOR DISTRIBUTION TO STUDENTS IN COURSE
Lecture material compilation Editors:

Rakesh Sharma,Ph.D
Avdhesh Sharma,Ph.D
Sponsoring Institution: Innovations and Solutions Inc. USA(R)
Address: 1.Center of Nanoscience and Biotechnology,
Innovations and Solutions Inc.
901 West Jefferson Street, Tallahassee A26, Florida 32304
2.Center of Nanomagnetics and Biotechnology,
Florida State University, Tallahassee FL 32304
Copyright© 2009. The material here is a compilation from web-based information and
research articles solely for sharing and educational purposes. Prohibited material to use for
any profit making business without prior permission from sole editors or original authors.
Cited text and materials is from web sources: Sribd.com, Novapublishers.com, nsti.org.
ISSN: 1556-4002
Cited material sources for students to read:

Bio-Nanomaterials and Nanotechnology ISBN: 978-1-60876-105-0
Carbon Nanotubes:New Research ISBN:978-1-60692-236-1
NSTI 2007,2008 and 2009 Conference CDs

Free distributed material from scribd.com
Contact Email:
rksz2009@yahoo.com
avdhesh_2000@yahoo.com
Lecture Materials: Table of contents
___________________________________________________________________________
Topic of lecture page

Lecture 1: What are nanomaterials? Bioapplications 1
Lecture 2: Quantum Transport in Nanotubes 6
Lecture 3: Physical Characteristics of nanomaterials: Carbon nanotubes 10
Lecture 4: How Gas-Carbon nanotubes Interactions Do Happen? 35
Lecture 5: Carbon Nanotubes: Growth Kinetics and Functionalization with Silicon
Nanocrystals 93
Lecture 6: Carbon Nanotube/Polymer Composites: Interfacial Bonding Characteristics 112
Lecture 7: Carbon nanotubes: Molecular Dynamics and Mechanical Properties 150
Lecture 8: Engineered electrical and mechanical properties of nanomaterials;
carbon nanotube added SI3N4 nanocomposites 167
Lecture 9: Fluorinated Carbon nanotubes: How Much We Know About Them? 191
Lecture 10: Carbon Nanotubes: Applications in the development of analytical methods 221
Lecture 11: Carbon Nanotubes: Dispersion and field emission properties 256
Lecture 12: Carbon Nanotubes:Coagulation-Fragmentation Equations 285

Lecture 13: Artificial fossilization process: A shortcut to nanostructured materials
from natural substances 296
Lecture 14: Biomimetic mineralization and mesocrystals 312
Lecture 15: Nano-fabricated structures and biomolecular analysis 342
Lecture 16: Bionic Superhydrophobic Surfaces Based on Colloidal Crystal Technique 360
Lecture 17: Nanomaterials:Green synthesis 397
Lecture 18: Targeted nanoparticles in cancer therapeutics 446
Lecture 19: Lithographically-Structured,gripping devices 466

Lecture 20: Protein engineering tools for interfacing proteins and solid supports with
exquisite chemical control 481
Lecture 21: Bioinspired colloidal systems 503
Lecture 22: Bacilli, Green Algae, Diatoms and Red Blood Cells
How nanobiotechnological research inspires architecture? 519
Lecture 23: Biopolyelectrolyte multilayer microshells: Assembly,property and application 554
Lecture 24: Synthesis and Electron Field Emission from Different Morphology Carbon
Nanofibers 574
Lecture 25: Carbon Nanotubes: A New Alternative for Electrochemical Sensors 635
Concluding Remark 681

__________________________________________________________________
Topic page
Lecture 1: What are nanomaterials? Bioapplications 1
Lecture 2: Quantum Transport in Nanotubes 6
Lecture 3: Physical Characteristics of nanomaterials: Carbon nanotubes 10
Lecture 4: How Gas-Carbon nanotubes Interactions Do Happen? 35
Lecture 5: Carbon Nanotubes: Growth Kinetics and Functionalization with Silicon
Nanocrystals 93
Lecture 6: Carbon Nanotube/Polymer Composites: Interfacial Bonding Characteristics 112
Lecture 7: Carbon nanotubes: Molecular Dynamics and Mechanical Properties 150
Lecture 8: Engineered electrical and mechanical properties of nanomaterials;
carbon nanotube added SI3N4 nanocomposites 167
Lecture 9: Fluorinated Carbon nanotubes: How Much We Know About Them? 191
Lecture 10: Carbon Nanotubes: Applications in the development of analytical methods 221
Lecture 11: Carbon Nanotubes: Dispersion and field emission properties 256
Lecture 12: Carbon Nanotubes:Coagulation-Fragmentation Equations 285
THE FOLLOWING LECTURE MATERIAL IS COMPILED AND SELECTED FOR STUDENTS ONLY
STUDENTS IN COURSE: Bioengineering ECH 4657 and BCE 4003c and 4004c
Florida State University Blackboard site
STATUS: PRIVATE CIRCULATION FOR EDUCATION AND ACADEMIC PURPOSE ONLY

Nanomaterials and Bioapplications Editors: Rakesh Sharma and Avdhesh Sharma 1
Lecture Material 1
WHAT ARE NANOMATERIALS? BIOAPPLICATIONS

OBJECTIVE
Nano materials show optical, magnetic, electronic, structural unusual toxicity properties and
complicate safety in tissue. In optimized quantities with limited functionality these nanomaterial
cages act as drug delivery carriers. In recent past, carbon nanotubes (CNT) or fullerenes displayed
their targeting properties such as increased circulating time, and acceptable biofunctionality as
biocompatible properties for its bioapplications. Over years, researchers have analyzed physical
parameters of CNT size, mass, surface area and effect on macrophage action, inflammatory action
due to nanotoxicity and mechanisms of reactive oxygen (ROS) induced transepithelial resistance
decrease in airway physiological CNT concentrations. Several biological functions of biosys-
tems display different mechanisms of their unique applications in biochemical engineering, electro-
nics, surface chemistry, protein engineering, DNA damage, immune reactivity, bio-inspired use
and biosensors. In conclusion, nanomaterials show very high potential use in medical/nonmedical
bioapplications in occupational, health, industry, technology, bioscience but their nanotoxicity as
health concern is disputed. For example, carbon nanotubes, with their extraordinary mechanical
and electronic properties, have garnered much attention in the past five years. The nanomaterials
are important to learn for their use in nanoelectronics,composites,chemical biosensors, microscopy,
nanoelectromechanical systems and many more. The scientific community is more motivated than
ever to move beyond basic properties and explore the real issues associated with nanomaterials-
nanotubes, nanofibers, nanocomposites based applications.
INTRODUCTION AND LAYOUT OF LECTURES

In this present lecture series, we have attempted to gather information from different web
based sources available in public domain (Bio-Inspired Nanomaterials and Nanotechnology
ISBN: 978-1-60876-105-0 and Carbon Nanotubes: New Research ISBN:978 -1-60692-236-1;
Scribd.com, nsti.org conference series publications) useful for students and graduate
researchers in advanced standing.
Nanomaterials: 'Science and Applications' describes the various aspects, including
properties, growth, and processing techniques, while focusing on individual major application
areas. Authors present an overview on structures and properties in UNIT I, followed by
coverage of synthesis and characterization of nanomaterials in UNIT II. Applications become
the focal point in chapters on scanning probe microscopy, nanomaterial-based field emission,
and the development of chemical and physical biosensors, and composites in UNIT III. The
topics have been selected from standard course on undergraduate and graduate level syllabus
from different university resources including Caltec, University of Pennsylvania, Utah state
University, Rice university and other places. It is expected that students will be benefitted with
the contents of presented material. However, the new information and other available
knowledge is not limited incuding these growing concepts of nanomaterials.
The attempt is to present up-to-date literature citations that express the current state of
science, this lecture series based on the own experience of editors fully explores the
development phase of nanomaterials including carbon nanotube-based applications. It is
expected a valuable resource for engineers, scientists, researchers, and professionals in a wide
range of disciplines whose focus remains on the power and promise of carbon nanotubes. The
2 Nanomaterials and Bioapplications Editors: Rakesh Sharma and Avdhesh Sharma
focus of present and future compilation of lectures will strive for new information and
developments in the following known fields and other upcoming new researches in
nano-bioapplications:
Source: Wikipedia key hit: Nanomaterials
1. TEXTILE AND AUTOMOBILE
clothes: waterproof tear-resistant textiles
combat jackets: MIT is working on combat jackets that use carbon nanotubes as
ultrastrong fibers and to monitor the condition of the wearer. Cambridge University has
developed the fibres and given a license to a company.
concrete: In concrete, they increase the tensile strength, and halt crack propagation.
polyethylene: Researchers have found that adding them to polyethylene increases the
polymer's elastic modulus by 30%.
sports equipment: Stronger and lighter tennis rackets, bike parts, golf balls, golf clubs,
golf shaft and baseball bats.
space elevator: This will be possible only if tensile strengths of more than about 70 GPa
can be achieved.
synthetic muscles: Due to their giant elongations and contractions when a current is run
through them, CNTs are ideal for synthetic muscle.
high tensile strength fibers: A large number of research groups have spun fibers of single
wall carbon nanotubes embedded into a polymer. For example, fibers produced with
polyvinyl alcohol required 600 J/g to break In comparison, the bullet-resistant fiber Kevlar is
27–33 J/g.
bridges: Carbon nanotubes may be able to replace steel in suspension bridges.
ultrahigh-speed flywheels: The high strength/weight ratio enables very high speeds to be
achieved.
fire protection: covering material with a thin layer of buckypaper significantly improves
its fire resistance due to the efficient reflection of heat by the dense, compact layer of carbon
nanotubes or carbon fibers.
artificial muscles
buckypaper - a thin sheet made from nanotubes that are 250 times stronger than steel and
10 times lighter that could be used as a heat sink for chipboards, a backlight for LCD screens
or as a faraday cage to protect electrical devices/aeroplanes.
chemical nanowires: Carbon nanotubes additionally can also be used to produce nanowires
of other chemicals, such as gold or zinc oxide. These nanowires in turn can be used to cast
nanotubes of other chemicals, such as gallium nitride. These can have very different
properties from CNTs - for example, gallium nitride nanotubes are hydrophilic, while CNTs
are hydrophobic, giving them possible uses in organic chemistry that CNTs could not be used
for.
conductive films: A 2005 paper in Science notes that drawing transparent high strength
swathes of SWNT is a functional production technique. Additionally, Eikos Inc of Franklin,
Massachusetts
and Unidym Inc. of Silicon Valley, California are developing transparent,
electrically
conductive films of carbon nanotubes to replace indium tin oxide (ITO) in LCDs,
touch
screens, and photovoltaic devices. Nanotube films show promise for use in displays for
computers, cell phones, PDAs, and ATMs.

electric motor brushes: Conductive carbon nanotubes have been used for several years in

brushes for commercial electric motors. They replace traditional carbon black, which is

mostly impure spherical carbon fullerenes. The nanotubes improve electrical and thermal

conductivity because they stretch through the plastic matrix of the brush. This permits the
carbon
filler to be reduced from 30% down to 3.6%, so that more matrix is present in the
brush.
Nanotube composite motor brushes are better-lubricated (from the matrix), cooler-
running (both from better lubrication and superior thermal conductivity), less brittle (more
matrix, and fiber reinforcement), stronger and more accurately moldable (more matrix). Since
brushes are a critical failure point in electric motors, and also don't need much material, they
became economical before almost any other application.
light bulb filament: alternative to tungsten filaments in incandescent lamps.
magnets: MWNTs coated with magnetite
optical ignition: A layer of 29% iron enriched SWNT is placed on top of a layer of
explosive material such as PETN, and can be ignited with a regular camera flash.
ITO in some solar cells to act as a transparent conductive film in solar cells to allow light
to to theGE's
passcells:
solar carbon
active layersnanotube diodephotocurrent.
and generate has a photovoltaic effect. Nanotubes can replace
superconductor: Nanotubes have been shown to be superconducting at low temperatures.

ultracapacitors: MIT is researching the use of nanotubes bound to the charge plates of
capacitors in order to dramatically increase the surface area and therefore energy storage
ability.
displays: One use for nanotubes that has already been developed is as extremely fine
electron guns, which could be used as miniature cathode ray tubes in thin high-brightness
low-energy low-weight displays. This type of display would consist of a group of many tiny
CRTs, each providing the electrons to hit the phosphor of one pixel, instead of having one
giant CRT whose electrons are aimed using electric and magnetic fields. These displays are
known as field emission displays (FEDs).
Light weight automobile bodies: CNT in aviation use.
transistor: developed at Delft, IBM, and NEC.
2. ELECTROACOUSTIC
loudspeaker: In November 2008, researchers at the Tsinghua-Foxconn Nanotechnology
Research Centre in Beijing announced they had created loudspeakers from sheets of parallel
carbon nanotubes, generating sound in a manner similar to how lightning produces thunder.
Near-term commercial uses include replacing piezoelectric speakers in greeting cards.
3. CHEMICAL
air pollution filter: Future applications of nanotube membranes include filtering carbon
dioxide from power plant emissions.
biotech container: Nanotubes can be opened and filled with materials such as biological
molecules, raising the possibility of applications in biotechnology.
hydrogen storage: Research is currently being undertaken into the potential use of carbon
nanotubes for hydrogen storage. They have the potential to store between 4.2 and 65%
hydrogen by weight. This is an important area of research, since if they can be mass produced
economically there is potential to contain the same quantity of energy as a 50L gasoline tank
in 13.2L of nanotubes. See also, Hydrogen Economy.
water filter: Recently nanotube membranes have been developed for use in filtration. This
technique can purportedly reduce desalination costs by 75%. The tubes are so thin that small
particles (like water molecules) can pass through them, while larger particles (such as the
chloride ions in salt) are blocked.
4. MECHANICAL
oscillator: fastest known oscillators (> 50 GHz).
nanotube membrane: Liquid flows up to five orders of magnitude faster than predicted by
classical fluid dynamics.
A nanotube formed by joining nanotubes of two different diameters end to end can act as a
diode, suggesting the possibility of constructing electronic computer circuits entirely out of
nanotubes. Because of their good thermal properties, carbon nanotubes can also be used to
dissipate heat from tiny computer chips. The longest electricity conducting circuit is a fraction
of an inch long.[16]
Fabrication difficulties are major hurdles for carbon nanotubes to find prominent places in
circuits. The production of electrical circuits with carbon nanotubes are very different from
the traditional IC fabrication process. The IC fabrication process is somewhat like sculpture -
films are deposited onto a wafer and pattern-etched away. Because carbon nanotubes are
fundamentally different from films, carbon nanotube circuits can so far not be mass produced.
Researchers sometimes resort to manipulating nanotubes one-by-one with the tip of an
atomic force microscope in a painstaking, time-consuming process. Perhaps the best hope is
that carbon nanotubes can be grown through a chemical vapor deposition process from
patterned catalyst material on a wafer, which serve as growth sites and allow designers to
position one end of the nanotube. During the deposition process, an electric field can be
applied to direct the growth of the nanotubes, which tend to grow along the field lines from
negative to positive polarity. Another way for the self assembly of the carbon nanotube
transistors consist in using chemical or biological techniques to place the nanotubes from
solution to determinate place on a substrate.
Even if nanotubes could be precisely positioned, there remains the problem that, to this
date, engineers have been unable to control the types of nanotubes—metallic,
semiconducting, single-walled, multi-walled—produced. A chemical engineering solution is
needed if nanotubes are to become feasible for commercial circuits.
Interconnects
Metallic carbon nanotubes have aroused a lot of research interest in their applicability as
Very-large-scale integration (VLSI) interconnects of the future because of their desirable
properties of high thermal stability, high thermal conductivity and large current carrying
capacity. An isolated carbon nanotube can carry current densities in excess of 1000 MA/sq-
cm without any signs of damage even at an elevated temperature of 250 degrees C, thereby
eliminating electromigration reliability concerns that plague Cu interconnects. Recent
modeling work comparing the performance, power dissipation and thermal/reliability aspects
of carbon nanotube interconnect to scaled copper interconnects have shown that carbon
nanotube bundle interconnects can potentially offer advantages over copper. Additionally,
the concept of hybrid interconnects-employing carbon nanotube vias in tandem with copper
interconnects has been shown to offer advantages from a reliability/thermal-management
perspective.
slick surface: slicker than Teflon and waterproof.
Semiconducting CNTs have been used to fabricate field effect transistors (CNTFETs),
which show promise due to their superior electrical characteristics over silicon based
MOSFETs. Since the electron mean free path in SWCNTs can exceed 1 micrometer, long
channel CNTFETs exhibit near-ballistic transport characteristics, resulting in high speed
devices. In fact, CNT devices are projected to be operational in the frequency range of
hundreds of GHz. Recent work detailing the advantages and disadvantages of various forms
of CNTFETs have also shown that the tunneling based CNTFET offers better characteristics
compared to other CNTFET structures. This device has been found to be superior in terms of
subthreshold slope - a very important property for low power applications.
Nanotubes are usually grown on nanoparticles of magnetic metal (Fe, Co) that facilitates
production of electronic (spintronic) devices. In particular control of current through a field-
effect transistor by magnetic field has been demonstrated in such a single-tube nanostructure.
5. ELECTRONIC DESIGN AND DESIGN AUTOMATION

Although carbon nanotube devices and interconnects have been separately shown to be
promising in their own respects, there have been few efforts to successfully combine them in
a realistic circuit. Most CNTFET structures employ the silicon substrate as a back gate.
Applying different back gate voltages might become a concern when designing large circuits
out of these devices. Several top-gated structures have also been demonstrated, which can
alleviate this concern. Recently, a fully integrated logic circuit built on a single nanotube has
been reported. However, this circuit also employs a back-gate. Additionally, there are still
several process related challenges that need to be addressed before CNT-based devices and
interconnects can enter mainstream VLSI process. This makes it an exciting and open field
for research. Problems like purification, separation of carbon nanotubes, control over
nanotube length, chirality and desired alignment, low thermal budget as well as high contact
resistance are yet to be fully resolved. Although these are serious technological challenges,
innovative ideas have been proposed to build practical transistors out of nano-networks. Since
lack of control on chirality produces a mix of metallic as well as semi-conducting CNTs from
any fabrication process and it is difficult to control the growth direction of the CNTs, random
arrays of SWCNTs (that are easily produced) have been proposed to build thin film
transistors. This idea can be further exploited to build practical CNT based transistors and
circuits without the need for precise growth and assembly.
6. BIOLOGICAL APPLICATIONS
DRUG DELIVERY: Not much is known and still use remains to be established
BIOINSPIRED NANOTECHNOLOGY: Major focus of nanotubes in biomedicine
LIGHT-WEIGHT HIGH TENSIL STRENGTH BIOMATERIALS: In aeroplane design
CONCLUSION
Nanomaterials are discovered for variety of new characteristics with potentials of wider
applications. Specially, carbon nanotubes remained focus for the following nanobio
applications. 1.Fullerenes nanotubes showed free radical chemistry, attraction to electrons,
antioxidant properties. 2. Some Carbon-60 fullerenes bind to nucleotides, hamper self-repair
in double–strand DNA. 3. CNT display high electrical and thermal conductivity, high
strength, rigidity. Medical/nonmedical applications suggest occupational, accidental
exposure. 4. Fullerenes(cages), single wall nanotubes, multiwalled nanotubes show toxicity.
CNT produce superoxide anion, lipid peroxidation, cytotoxicity in plants and animals. 5.
Uncoated fullerenes in largemouth bass show lipid peroxidation in brain tissue and
glutathione depletion in gills. 6. C60 toxicity increases by Poly Vinyl Propylene due to stable
charge transfer complexes. 7. THF may pass through blood-brain barrier. Metal catalyst used
in nanotube fabrication. 8. More derivatized fullerenes were less toxic, due to low efficiency
in ROS generation. 9. CNT showed toxicity effects, dose dependent epitheloid granuloma. 10.
At optimized CNT single walled CNT concentrations, low Taxotere quantities encaged inside
may target breast tumor tissue more efficiently. 11. Cultured alveolar fibroblasts following
exposure of CNTs showed possibility of transplanting CNT encaged fibroblasts. The present
lecture series will cater the needs of undergraduate and graduate level course work in
nanobioscience and applied aspects.
Lecture Material 2
QUANTUM TRANSPORT IN CARBON NANOTUBES
OBJECTIVE
The Drude model relative to carrier drift mobility and conductivity is discussed in
relation to its applicability to explain quantum transport through both single-walled and
multiwalled carbon nanotubes. We show that the Drude model, suitable to describe
electrical conductivity in solids as, for instance, semiconductors, is certainly adequate to
discuss key aspects of quantum transport in multiwalled carbon nanotubes. In this
respect, conductance quantization is treated in relation to the model in question by
calculating carrier mobility and conductivity under ballistic regime. As a consequence, an
expression for the quantized conductance of a multiwalled carbon nanotube is derived.
Keywords: Drude model; carbon nanotubes; electrical conductivity; drift mobility;

ballistic regime.
INTRODUCTION
The well-known Drude model relative to electron transport in macroscopic systems is
extrapolable, under certain conditions, to nanoscopic structures [1-6]. In fact, by means of this
model, analytical approaches have been made to determine electron conductance through both
single-walled and multiwalled carbon nanotubes [3-5]. In this respect, we wish to remark the
significant importance of the theoretical determination of the electrical conductance in carbon
nanotubes. This conductance was found to be quantized [3-5, 7, 8] which constitutes a
relevant fact. Conductance quantization will be treated in the following in relation to ballistic
transport; at this point, let us remember that ballistic transport is a necessary condition (but
not sufficient) for conductance quantization (see, for example, ref.[7]). In other words,
conductance quantization implies ballistic regime but the reciprocal assertion is not true in
general. Therefore, if the carrier transport is diffusive, conductance is not quantized. Ballistic
transport through a nanotube, that is, when the electronic mean free path is much longer than
the tube length, is the regime under which, in fact, carbon nanotubes behave (see previous
references) so that scattering in these tubes should consist of elastic collisions (see, for
instance, ref.[7]). This communication is devoted to examine the Drude model within the
context of carbon nanotubes by emphasizing the main aspects related to ballistic regime and
conductance quantization.
THEORY
Let us consider the Drude model by which one has that the electrical conductivity is
given by eN where e is the absolute value of the electron charge, N is the spatial
carrier density, and denotes carrier drift mobility which, in turn, reads e m where
designates relaxation time (or momentum-scattering rate) and m stands for carrier
effective mass. Then, by replacing the second formula into the first one, it follows:
e2 N
(1)
m
Eq.(1) is standard in physics of semiconductors and constitutes a relevant element of

reference within the context of macroscopic condensed-matter systems. Under certain
conditions, this formula can be extrapolated to nanoscale systems (see, for instance, refs.[1-
6]). In particular, let us consider a multiwalled carbon nanotube (MWCNT) conceived as a
longitudinal quantum box [2, 3]; in such a tube, conductance is quantized according to the
fact that the involved quantum number coincides with the numbering of the layers of the tube
(conductance scales with the number of layers) [2-5, 7, 8]. Therefore, quantizing formula (1)
for a metallic MWCNT, one has for the conductivity due to the n layer (see, for instance,
th
refs.[3, 5, 6]):
e2 Nn
n
n
(2)
m
where n designates quantum number ( n 1,2,... ) and m has been replaced by the free-
electron mass denoted by m . In addition, now n is transit time or motion time (note that
transport is ballistic) so that n l v n where l is the length of the tube and vn stands for
the magnitude of the quantized Fermi velocity which, for n 1 , approaches the electron
velocity deduced from equating the quantized electron energy E n h 2 n 2 8ml 2
(corresponding to the electron confinement in the tube as a longitudinal ideal quantum box) to
1 2 mvn2 [6]. Hence, it follows:
hn
vn (3)
2ml
where h is the Planck constant.

We regard our MWCNT as a quasi-one-dimensional structure [2-5] so that A l
where A is the cross-sectional area of the tube. On the other hand, we will assume that the
number of electrons participating in the conduction process depends upon n in accordance
2
with the distribution of electrons in the atomic shells so that the above number equals 2n
[6]. Therefore, inserting the relation namely N n 2n 2 lA into formula (2), taking into
account that the quantized conductance is given by Gn n A l , and inserting into this
formula relations (2) and (3) with the fact that n l v n , one has finally that Gn 2G0 n
where G 0 2e 2 h is the fundamental conductance quantum (see ref.[6]). The above-
mentioned expression for the quantized conductance tells us that this conductance
approximately depends linearly on n which is acceptable in the quasi-classical case, that is,
for sufficiently large values of the quantum number. At any rate, a formula valid for every n
can be given. To get this, we use the following relationship concerning the energy-level
spacing induced by quantum confinement, namely [2, 4, 6]:
hv Fn
En 1 En (4)
2l
where v Fn is the magnitude of the quantized Fermi velocity.

By (4) and the expression for the quantized energy, one gets:
h 2n 1
v Fn (5)
4ml
Notice that the right-hand side of (3) coincides approximately with the right-hand side of
(5) when n 1 (quasi-classical case).
Repeating the calculation process in the light of the Drude model developed previously
by employing now formula (5) for the electron velocity, the quantized conductance can be
expressed as:
4kG0 n 2
Gn (6)
2n 1
where k is a phenomenological parameter such that 0 k 1 which is a measure of the
interwall (interlayer) coupling in the tube. We assume that k is a uniform continuous random
variable so its average or expected value is k 1 2 . In the quasi-classical case, that is,
when n 1, from relationship (6) it follows that Gn 2kG0 n which gives an expected
value Gn 2 k G0 n G0 n . This result agrees with experimental data [7].
For a single-walled carbon nanotube (SWCNT), we use formula (6) when n 1 (ground
state of our quantized system) [4] so that the corresponding conductance reads
G1 4kG0 3 . In this case, k belonging to the above range is invalid since now it is
obvious that speaking of interlayer coupling does not make sense. Consequently,
extrapolation of k to values outside the range in question is necessary. Given that, as it is
well-known, the conductance through an SWCNT is 2G0 , by equating this value to the
previous expression for G1 , one gets that k 3 2 which evidently is outside the open
interval 0 k 1 . However, this extrapolation cannot be derived from our formulation.
CONCLUSION
In essence, a mathematical relationship for the quantized conductance through an
MWCNT has been obtained in the light of the Drude model within a quantum-box approach.
The applicability of the Drude model to metallic tubes is acceptable in the quasi-classical
case. In this case, the conductance is approximately proportional to the number of quantized
modes (conducting channels). At any rate, eq.(6) is valid for all the values of n . The
interaction between consecutive layers has been specified by using the statistical parameter k
whose possible values have been discussed. In addition, considerations relative to SWCNTs
have been made. We emphasize the fact that our results agree well with experimental
observations [7]. Finally, we wish to remark the usefulness of certain approaches [9, 10] to
elucidate problems related to quantum transport in carbon nanotubes. We can also mention
potential-well based formulations [2, 3, 4, 6].
REFERENCES
[1] T. Dürkop, B.M. Kim, M.S. Fuhrer: Properties and applications of high-mobility
semiconducting nanotubes, J. Phys. Condensed Matter 16 (2004) R553-R580.
[2] M.A. Grado-Caffaro, M. Grado-Caffaro: A theoretical analysis on the Fermi level in
multiwalled carbon nanotubes, Mod. Phys. Lett. B 18 (2004) 501-503.
[3] M.A. Grado-Caffaro, M. Grado-Caffaro: Fractional conductance in multiwalled carbon
nanotubes: a semi-classical theory, Mod. Phys. Lett. B 18 (2004) 761-767.
[4] M.A. Grado-Caffaro, M. Grado-Caffaro: On the size of small single-walled carbon
nanotubes, Optik 116 (2005) 459-460.
[5] M.A. Grado-Caffaro, M. Grado-Caffaro: Theoretical characterization of quantum
ballistic conduction through multiwalled carbon nanotubes, Mod. Phys. Lett. B 19
(2005) 967-969.
[6] M.A. Grado-Caffaro, M. Grado-Caffaro: A potential-well based formulation to
calculate the quantized conductance of a one-atom constriction, Phys. Lett. A 372
(2008) 3573-3576.
[7] S. Frank, P. Poncharal, Z.L. Wang, W.A. de Heer: Carbon nanotube quantum resistors,
Science 280 (1998) 1744-1746.
[8] M.F. Lin, K.W.-K. Shung: Magnetoconductance of carbon nanotubes, Phys. Rev. B 51
(1995) 7592.
[9] N.B. Brandt, S.M. Chudinov, Ya.G. Ponomarev: Semimetals 1, Graphite and its
Compounds, vol.20.1 of Modern Problems in Condensed Matter Science (North-
Holland, Amsterdam, 1988) pp. 74-77.
[10] J. Chen, Z. Yang, J. Gu: Energy gap of the ―m etallic‖ single-walled carbon nanotubes,
Mod. Phys. Lett. B 18 (2004) 769-774.
Lecture Material 3
PHYSICAL CHARACTERISTICS OF CARBON

NANOTUBES
ABSTRACT
This chapter studies the physical characteristics of carbon nanotubes (CNTs) films
and Co-filled CNTs by using an electron cyclotron resonance chemical vapor deposition
(ECR-CVD) method. The results show that the optimum relative intensity ratio of the D
band to G band (i.e., I D / I G ) is 2 for the cases considered in this study. The effects of
different plasma powers of 200W, 300W, 400W and 500W, on the morphology, structure
and electrical properties of the CNTs film, are also studied. The surface density of the
vertical nanotubes decreases when the plasma power is higher than 200W. The Co-filled
CNTs grown at 300W and 400W have a current discharge at the applied voltages of 30 V
and 40 V, respectively. In addition, this chapter also develops a model that analyzes the
resonant frequency of the chiral single-walled carbon nanotubes (SWCNTs) subjected to
a thermal vibration by using Timoshenko beam model, including the effect of rotary
inertia and shear deformation. The frequency obtained by Timoshenko beam model is
lower than that calculated by Euler beam model. As the nanotube aspect ratio of length to
diameter decreases, the discrepancy is more obvious. As the effect of thermal vibration
increases, the frequency for chiral SWCNTs decreases. Furthermore, the oxidation
characteristics of TiN thin films by atomic force microscopy (AFM)
electrochemical nanolithography with carbon nanotube tip are investigated.
INTRODUCTION
Since carbons nanotubes (CNTs) were discovered in 1991 [1], they have attracted great
interest due to their exceptional exceptional mechanical, chemical, electronic, and thermal
properties [2-5]. Many applications of CNTs have been reported, such as probes in atomic
force microscope (AFM) [6,7], electron emitters for field emission display (FED)[8,9],
nanofillers for composite materials[10,11], electrodes for fuel cells[12,13], nanoscale
electronic devices for micro/nanoelectromechanical system(N/MEMS) [14].In addition,
multi-walled carbont nanotubes (MWCNTs) have the potential for the development of
frictionless nanomotors, nanoactuators, nanobearings and nanosprings [15]. Therefore, the
extensive research on CNTs may lead to new applications of other nanostructures and
nanoengineering [16-21].
The catalytic chemical vapor deposition (CVD) method is one of the most promising
approaches and has been widely used to growth CNTs on the substrate with the assistance of
metallic catalysts. The method includes that a gas flows over metallic catalytic particles such
as Fe, Co and Ni and a gas mixture containing a carbon precursor such as acetylene, carbon
monoxide, and methane. With the CVD method, CNTs can be grown at desired locations with
a specified direction. In recent years, different catalytic CVD methods for growing CNTs
have been developed, such as Merkulov et al. [22] and Cojocaru et al. [23] utilized the plasma
enhanced chemical vapor deposition (PE-CVD) growth of multi-walled carbon nanotubes
(MWCNTs) onto substrates coated with a suitable transition metal catalyst. Chen et al. [24]
and Wu et al. [25] utilized the electron cyclotron resonance chemical vapor deposition (ECR-
CVD) process is used to synthesize CNTs at low temperature. Fang et al. [26] studied the
effects of gas composition on the growth of MWCNTs and measured the surface topography
and thermal properties of MWCNTs using atomic force microscopy (AFM) and scanning
thermal microscopy (SThM).
In addition, a new type of palladium metal-filled carbon nanotubes (MF-CNTs) was
synthesized using a bias enhanced microwave plasma enhanced chemical vapor deposition
(BE-MPECVD)[27,28]. The MF-CNTs had a better radial rigidity than that of hollow
nanotubes [29]. The MF-CNTs provided higher mechanical and thermal stability than a short
cylinder. Therefore, MF-CNTs are one of the best candidates for use, not only in field
emission devices but also as scanning probe microscopy probe tips [30]. Recently, Fang et al.
[31] studied the growth characteristics of co-filled carbon nanotubes and examined the
surface morphology and the structure of the CNTs film using scanning electron microscopy
(SEM) and high-resolution field emission gun transmission electron microscopy (TEM).
Carbon nanotubes in terms of the chiral angle can be classified into three types: armchair,
zigzag and chiral. Numerous studies are available on the physical properties of armchair and
zigzag carbon nanotube in the literature [32-35]. However, only a limited portion of the
literature is concerned with the aspect of chiral carbon nanotubes. This is because the
geometric structure in chiral carbon nanotubes is complicated and difficult to analyze.
Recently, Leung et al. [36] proposed an energy-equivalent model to establish the
computational formula of Young‘s modulus for chiral single-walled carbon nanotubes and
found that Young‘s modulus of the chiral SWCNTs is affected both by its diameter and chiral
angle. Zhang et al. [37] performed the molecular dynamics simulations on SWCNTs to study
the effects of chirality on their buckling behavior under axial compression. Chen et al. [38]
studied the effects of the geometric structure and an electric field on the electronic and optical
properties of quasi-zero-dimensional finite carbon nanotubes by employing the tight-binding
model coupled with curvature effects.
In this chapter, the growth of the Co-filled CNTs was carried out using ECR-CVD. The
growth process of MWCNTs on a nickel coated silicon substrate using electron cyclotron
resonance chemical vapor deposition with the mixed gases of C3H8/ H2 is studied. The surface
morphology, micro Raman spectra, Energy dispersive X-ray spectroscopy (EDXS), TEM and
the current density (J)-electric field (E) properties of the CNTs are discussed. Furthermore,
this article proposes a thermal vibration model to analyze the resonance frequency of chiral
single-walled carbon nanotubes (SWCNTs) with simply supported ends using Timoshenko
beam theory. In addition, comparison of fundamental frequency of the nanotubes by Euler
beam and Timoshenko beam theories is also studied.
EXPERIMENTAL DETAILS
The nickel catalyst film with a thickness of 5 -10 nm was deposited on n-type silicon
wafer substrate of 10 mm × 10 mm in size by using ion beam sputtering at room temperature.
Then CNTs growth was carried out in an electron cyclotron resonance chemical vapor
deposition system with a base pressure below 10−6 Torr at the ECR power of 400 W and the
substrate temperature of 600 C for 5 min as shown in Figure 1[39]. During the growth
o
process, the propane ( C3 H 8 ) was chosen as the reactive gas mixed with hydrogen ( H 2 ), and
the different flow rates of C3 H 8 : H 2 were set that is 1:1, 1:2, 1:3, 1:4, and 2:3, respectively.
The morphology and microstructure of the CNTs were examined by field emission
scanning electron microscopy (FE-SEM, Hitachi S4200) and field emission gun transmission
electron microscopy (FEG-TEM, Philips Tecnai G2 F20). In addition, atomic force
microscopy (AFM, Veeco/TM CP-RII SPM), and scanning thermal microscopy (SThM, Topo
Metrix) are used to study the surface topography and thermal properties. The nanomechanical
characteristics of CNTs were also investigated using nanoindentation system. A nanoindenter
(Triboscope, Hysitron) was used to compress the nanotubes and generate force–displacement
curves. The electrical resistance of the samples was measured by probe station method.
In addition, the MF-CNTs film was grown using the ECR-CVD method. A negative bias
of 200 V was applied to the substrate holder to grow vertical-aligned MF-CNTs on the
substrate. Prior to the growth of the MF-CNTs, a thin film of Co metal was deposited onto Si
substrate surface. The substrate was then transferred into the growth chamber and kept in
vacuum for 30 min, and then nitrogen (N2) gas was fed into the chamber to maintain a
pressure of 3x10-4 Torr at a temperature of around 470 K. The microwave plasma powers of
200 to 500W were used. The feed gas, propane (C3H8), was introduced and the nitrogen gas
was adjusted to achieve the C3H8/ N2 ratio of 20 sccm/20 sccm at a total pressure of 5x10-3
Torr. Then, the MF-CNTs were grown at deposition times ranging from 5 min to 15 min.
The surface morphology and the structure of the MF-C NTs film were examined using
SEM and high-resolution field emission gun TEM. The micro Raman spectrum was measured
at room temperature using an Ar+ ion laser with a wavelength of 514.5 nm, and the signals
were separated by a monochromator.
Figure 1. The schematic diagram of the ECR-CVD system [26].

In the experiment the MF-CNTs film device made up of the MF-CNTs film serving as a
cathode and an aluminum plate serving as an anode was used to measure the electrical
properties. The two electrodes were isolated by a piece of glass. This device was placed in a
chamber, and the air was pumped out of the chamber to establish a high vacuum. The
voltages and current density were measured at room temperature. During the measurement,
the distance between the anode and cathode was kept at 0.1 mm.
ANALYSIS
In this analysis, the chiral SWCNTs with simply supported ends subjected to a thermal
vibration, has an equivalent Young‘s modulus E and the shear modulus G, is described as a
hollow cylindrical tube with cross-sectional area A, length L, inner diameter d and
thickness tc . The flexural vibration of the SWCNTs is a partial differential equation and its
transverse displacement Y is dependent on time t and the spatial coordinate X. The governing
equation of Timoshenko beam, including the effect of rotary inertia and shear deformation,
for the SWCNTs is [40]:
EI 4Y FT 2Y 2
Y I E 4
Y I 4
Y
1 0 (1)
A x4 A x2 t2 A KG 2 2
x t A KG t 4
where I is the moment of inertia of the nanotube. K is the shear coefficient of carbon
2(1 p )
nanotube, and its value is K and is Poisson‘s ratio [41]. FT denotes an
4 3
p
p
additional axial force and is dependent of temperature T and thermal expansion coefficient
of nanotube, the force can be expressed as FT T E A [42].
The corresponding boundary conditions are
d 2Y (0, t ) d 2Y ( L, t )
Y (0, t ) 0 (2) Y ( L, t ) 0 (3)
dx 2 dx 2
The boundary conditions given by Eqs. (2) and (3) correspond to conditions of zero
displacement and zero bending moment at X = 0 and X = L, respectively.
The solution of the differential equation given in Eq. (1) can be expressed as
Y x, t y x e i t
(4)
where is the fundamental frequency. The following dimensionless forms can be deduced
d4 2 d2 2 2
1 0 (5)
d 4 d 2
where , , , , and are dimensionless parameters and they are
y x I EI 2 A 2 L4 FT L2
, , , , , (6)
L L AL2 KGAL2 EI EI
where represents the effect of thermal vibration on the frequency of SWCNTs and can be
expressed as
4 T
2 2
(7)
D tc
L L
A general solution to Eq. (5) can be expressed as
c1 cos c2 sin c3 cosh c4 sinh (8)
where
1 4 2 2 2 2
2 4 (9)
2
1 2
(10)
2
The solution must be satisfied the boundary conditions of Eqs. (2) and (3). From the
conditions, we can yield the following constraint
sin 0 (11)
According to the Timoshenko beam model, the fundamental frequency of the SWCNTs
can be obtained from the equation given in Eq. (11). The frequency equation is
2
4
1 2
2
2
0 (12)
For the case of the Euler beam, 0 in Eq. (5) and then the fundamental
frequency equation can be reduced to
2
EI
2
(13)
L A
If the effect of thermal vibration is not taken into account (i.e., 0 ), the fundamental
frequency for the Euler beam is obtained and can be found from general textbooks of
vibration [43].
APPLICATION
The TiN thin films were deposited on a p-type Si(100) substrate by ALCVD. Before the
film‘s deposition, the Si wafer was cleaned in a HF solution to remove the native oxide SiO2
layer. TiCl4 and NH3 reaction gases were supplied sequentially during the deposition process
[44]. Ar was used as a purge gas to remove the chemical residue with no adsorption to the
surface. The amount of time each gas was sequentially supplied for TiCl4, Ar purge, NH3 and
Ar purge was 5, 4, 1 and 5 s, respectively. The possible reaction for the ALCVD using
precursors of TiCl4 and NH3 is as follows:
6TiCl4+8NH3 6TiN+24HCl+N2 (14)
The average surface roughness of the TiN films for the temperatures of 350, 400, 450 and
0
500 C were approximately 1.7, 0.7, 0.6 and 0.4 nm, respectively. The typical thickness of
TiN film is about 10 nm. The resistivity of the TiN films for the temperatures of 350, 400,
0
450 and 500 C were about 710, 325, 250 and 145 cm , respectively. As the
deposition temperature was increased, the surface roughness and the resistivity of the TiN
films decreased. In this study the TiN film with the best surface behavior was at the
0
deposition temperature of 500 C and it was chosen to explore its oxidation characteristics.
The substrate temperature during the deposition process was kept at 500℃ and the pressure
was at about 133 Nm−2.
The local oxidation experiments were performed using an AFM (NT-MDT Solver,
Russia) with a carbon nanotube probe (Nanoscience Instruments, USA) as shown in Figure 2.
The carbon nanotube probe consisted the multi-wall nanotube tip that was mounted onto a
commercial AFM etched silicon probe. The carbon nanotube diameter is about 30 nm. An Al
reflective layer of 30 nm was coated on the AFM cantilever backside. The cantilever length
and spring constant of the AFM probe were 125 m and 40 N/m, respectively. In this
technique, the oxide structures grew on the chemically reactive surfaces by the application of
a bias voltage between the surface and the AFM probe tip. Composition of the oxide and the
film was characterized by Auger electron spectroscopy (AES, VG Microlab 310D, USA)
with incident electron energy of 10 keV. The AFM probe was used as the cathode and the
adsorbed water created from an ambient humidity of 60 % was used as the electrolyte in non-
contact mode. When the electric field was greater than 2 x10 7 V/cm and the water meniscus
adsorbed on the specimen surface provided the oxyanions oxide structures were formed on
the surface.
Figure 2. Schematic of the electrochemical principle of AFM with carbon nanotube probe and SEM
image of the carbon nanotube probe [30].
RESULTS AND DISCUSSION

Figure 3 shows a SEM image of the Ni catalyst for growing the CNTs after ion beam
sputtering deposition. The Ni catalyst was aggregated after the film growth due to the surface
tension and the stress induced from the mismatch of the thermal expansion coefficients of the
silicon substrate and Ni film. The size of the Ni particle was about 5-10 nm. The Ni played an
important role in the promotion of CNTs growth. The growth on Ni catalyst was similar to
gas-solid interaction process to obtain a single CNT from every Ni particle.
Figure 3. A SEM image of the Ni catalyst.[26].
Figure 4(a) and (b) show the top view and the cross-section view, respectively, for SEM
images of CNTs deposited on with C3H8:H2 = 2:3 gas mixture at the ECR power of 400 W for
5 min. The SEM image appears the carbon nanotubes are approximately perpendicular to the
substrate surface and form an aligned carbon nanotube array. The nanotube array was grown
on the thin uniform catalyst layer.
(a) (b)
Figure 4. (a) Top view and (b) cross-section view SEM images of MWCNTs deposited on a Ni catalyst
film with C3H8:H2 = 2:3 gas mixture at the ECR power of 400 W for 5 min. [26].
Figure 5(a) shows a bundle of FEG-TEM images of MWCNTs deposited with C3H8:H2 =
1:4 gas mixture at the ECR power of 400 W for 5 min. The resolution of a TEM is generally
one order of magnitude higher than that an SEM. As shown in the figure, each MWCNT in
the bundle has a different length with a bamboo-like structure. It indicates that CNTs the
graphitic layers are not perfectly parallel to the tube axis and do not grow from the bottom to
the top of films. Figure 5(b) illustrates the closed-end tip images of MWCNTs shows that the
carbon nanotubes have hollow structure. The outer diameter and inner diameter of this
nanotube are about 20 and 5 nm, respectively. The outer diameter of CNT could be
determined by the diameter of the catalytic Ni particle. Amorphous carbon layer in the image
is observed and it indicates that the graphitization was not perfect due to the lower reaction
temperature.
(a) (b)
Figure 5. FEG-TEM images of MWCNTs deposited with C3H8:H2 = 1:4 gas mixture at the ECR power
of 400 W for 5 min. (a) The bundle of CNTs and (b) the closed-end tip [26].
Figure 6(a) shows the Raman spectra of the CNTs grown on the substrate for different
1
ratios of C3H8 to H2 and has two broadband peaks at about 1350 and 1580 cm , refer to the
D band and the G band, respectively. The D band is disorder-induced features and indicates
the existence of defective whereas the G band denotes original graphite sheet features.
(a)
Figure 6. (Continued).
(b)
Figure 6. Raman spectra of MWCNTs grown on Si substrates as (a) a function of the ratio of C3H8 to
H2. (b) Variation of I D / IG ratio with the flow ratio of H2 to C3H8.[26].
The role of atomic hydrogen that is responsible for the formation of nanoparticle nuclei,
and it is in a balance state during the deposition of nanotube film. Increasing levels of atomic
hydrogen could act to reduce the decomposition rate of propane on the catalyst surface. The
relative intensity ratio of the D band to G band (i.e., I D / I G ) varies with the flow ratio of H2
to C3H8 was shown in Figure 6(b). It can be seen that the optimum ratio for the cases of the
experiment is 2, and it indicates that the condition for the growth of CNTs is most suitable.
The images of SThM with resistance of RC = 21.3Ω for MWCNTs deposited at the ECR
power of 400 W with the gas mixture of C3H8L:H2 = 1:1 and C3H8:H2 = 1:2 are shown in
Figure 7(a) and 6(b), respectively. The average surface roughness of the films was about 7
~17 nm. This was due to the different vacancies and crystalline of the films. It can be found
from the figures that the surface roughness of MWCNTs is lower when the growth condition
of gas mixture of C3H8:H2 = 1:2 were used.
Figure 8(a) depicts the load-depth curve of nanoindentation test for MWCNTs at the
applied force of 250 μN as a function of the ratio of C3H8 to H2. The hardness and the stiffness
of the films were about 0.4~ 4 GPa and 10~70 kN/m, respectively. The higher hardness of the
samples were deposited at C3H8:H2 = 1:2. The result might indicate that the reduction of the
sp2 content increased the values of hardness. It can‘t seem to find the clear relationship
between the indentation depth and the ratio due to the film defects. However, the load-depth
curve with C3H8:H2 = 1:3 gas mixture appears that the indentation depth increased linearly
with increasing load as shown in Figure 8(b).
The surface images of the MF-CNTs film grown at different plasma powers of 200W,
300W, 400W and 500W are shown in Figure 9. The surface morphology differences among
the samples under different plasma powers are evident. The SEM images shows that the
carbon nanotubes formed as an amorphous carbon film and did not produce a hollow tube
shape when the film was grown at a plasma power of 200 W.
(a) (b)
Figure 7. SThM Images of with resistance of RC = 21.3Ω for MWCNTs deposited with the gas
mixture of (a) C3H8:H2 = 1:1 and (b) C3H8：H2 = 1:2 at the ECR power of 400 W for 5 min.[26].
When the plasma power was higher than 200W, the surface density of the vertical
nanotubes decreased and therefore the spacing among the tubes became larger. It can be seen
that on the surface the MF-CNTs are randomly oriented. When the plasma power of 300W
was used, the ends of the MF-CNTs became straighter and more uniform, although there still
were some tube tips protruding on the film‘s surface. When the plasma power of 500W was
used, the protruding tube tips increased and aggregated. This was due to the higher plasma
power providing higher kinetic energy to the carbon atoms which caused the tubes to
aggregate and also caused a lower adhesion so that the tube‘s flaked off the substrate. As the
deposition time was increased, the length of the MF-CNTs also increased. The surface islands
became smaller and the MF-CNTs film contained more amorphous carbon impurities. A
growth model that can explain the Co-filled CNTs and state as follows: After Co layer
deposition, the Co catalyst becomes fragmented into sphere-like nanoparticles by electrostatic
force and sintering thermodynamics [27]. When the CNTs are filled by the Co, the Co is
trapped in the CNTs from the basal side. Therefore, the growth of the Co-filled CNTs takes
place during the decomposition of the carbonaceous molecules on the Co nanoparticles
surface, and then the carbon atoms diffuse through the Co nanoparticle to produce the MF-
CNTs. The hollowness between the basal sides of the CNTs is due to the Co is not being
sufficient enough to fill the entire CNTs [27].
(a)
(b)
Figure 8. The load-depth curve of nanoindentation test for MWCNTs grown on Si substrates as (a)
a function of the ratio of C3H8 to H2 at F = 250μN and (b) a function of applied load with C3H8:H2
= 1:3 gas mixture.[26].
(a)
(b)
(c)
(d)
Figure 9. Co-filled CNTs film grown at different plasma powers; (a) 200W; (b) 300W; (c) 400W; and
(d) 500W.[27].
The micro Raman spectra of the MF-CNTs film at different plasma powers and a growth
time of 10 min are shown in Figure 10.
Figure 10. Raman patterns of the Co-filled CNTs film [27].
The Raman spectra consisted of two broad peaks located at 1355 cm-1 (the D peak)
and1590 cm-1 (the G peak), The D and G peaks are attributed to the small crystallite graphite
materials or the so-called disordered graphite and the in plain symmetric C–C stretching
(E2g), respectively. In these cases, the MF-CNTs film contained some amorphous carbon
inside the tube walls. As the plasma power was increased, the G-peak of the MF-CNTs film
became broader due to the large amount of amorphous carbon. The Raman shift indicated that
the structure changed from graphite to nanocrystalline graphite possible occurred by filling
Co metal inside the tubes. The Raman intensity ratio between the D and G bands (Id/Ig) of the
MF-CNTs was larger than that of CNTs in previous experiments [27].
A bright field image taken at low magnification showed that the Co-filled CNTs grew to
about 80 nm in diameter and about 800 nm in length as shown in Figs. 11(a) and 11(b). It can
be clearly seen that the upper part of the inner tube of the MF-CNTs was filled with a catalyst
metal Co. The metal filling almost completely covered the inside of the tube, and the
thickness of the layer is about 10 nm. After cooling, the solidification of Co led to the volume
change inside the tubes. The structure of the MF-CNTs wall did not exhibit clear graphene
sheets lattice fringes at all. The diffraction patterns taken only from the wall region of the
MF-CNTs indicated that the MF-CNTs were composed of poorly ordered graphene layers.
The strain was due to the absence of the ordered graphene sheets [29]. From Figure 11(b),
carbon shells (graphene layers) can be visible in high resolution TEM, and the (average)
number of graphene layers is about 40.
(a) (b)
Figure 11. (a)TEM image of the Co-filled CNTs and (b) its enlarged image [27].
Energy dispersive X-ray spectroscopy (EDXS) analyses were performed to identify the
composition of the material inside the CNTs as shown in Figure 12. The EDXS elemental
maps of Co and C measured at a solid area in the TEM image was shown in Figure 11. On the
basis of elemental analyses, it been seen not only Co, but also Cu and O elements. The
presence of Cu and O elements was confirmed from the EDXS spectrum incorporated into the
metal within the CNTs from the Cu-TEM mesh. Both the elemental map and the TEM image
clearly indicate a distribution of Co within the CNTs.
Figure 12. EDXS of the Co-filled CNTs [27].

Figure 13 shows the current density (J)-electric field (E) properties of the Co-filled CNTs
sample under the plasma powers of 200W, 300 W and 400 W. The field emission properties
are measured to evaluate the validity of the growth of carbon nanotubes. The Co-filled CNTs
was grown at a deposition time of 10 min. The results show, as the voltage was increased, the
current density increased. When the plasma power was increased, the current density
increased to a certain value. The tips of the Co-filled CNTs grown at 300W and 400 W had a
current discharge under the applied voltage of 30 V and 40 V, respectively. From the
experimental data it can be found that a turn-on electric field of 0.05 V / m was obtained at
2
an emission current density of 0.01 mA / cm for the deposited condition of 400 W and 0.1
V / m for the case of 300 W. An emission current density of 1 mA / cm2 was obtained at
1.1 V / m for the power of 300 W, which is higher than that of the 400W (0.3 V / m ).
The onset field of the 400 W sample is lower than that of the 300 W sample.
This article studies the resonant frequency of chiral single-walled carbon nanotubes. The
geometric and material parameters used in this analysis are as follows: L=50 nm, tc =0.34 nm,
2300kg / m3 , p =0.2. In addition, the young‘s modulus of the carbon nanotube is
dependent of the chiral angle and diameter and taken from the computational formula of
Leung et al.[36]. The computation of I, G, and A is based on the mechanics of material [45].
Figure 13. J-E properties of the Co-filled CNTs film [27].
The computational results of fundamental frequencies of carbon nanotube are shown in

Figure 14. In this figure, we illustrate the fundamental frequencies of chiral naotubes of
7.59 and 25.29 , excluding the effect of thermal vibration (i.e., 0 ), using
Euler beam model and Timoshenko beam model.
600
KHz)
500
Fundamental frequency,
400
300 Euler beam model
200
Timoshenko beam model

100
0 4 8 12 16 20
Nanotube aspect ratio, L / d
Figure 14. Comparison of fundamental frequencies of chiral carbon nanotube with 0 for different
aspect ratios by Euler beam model and Timoshenko beam model [40].
It can be seen that the frequency decreases with increasing the nanotube aspect ratio of
length to diameter, L/d. The trend is similar with those of armchair and zigzag carbon
nanotubes obtained by the previous study [46]. The frequency calculated by Timoshenko
beam model is lower than that obtained by Euler beam model. This is because the presence of
rotary inertia and shear deformation tends to make the nanotube less stiff. The effect of rotary
inertia and shear deformation on the frequency is significant as the value of L/d becomes low.
Furthermore, the chirality of nanotube has not obvious effect on the fundamental frequency.
The effect of thermal vibration on the resonant frequency of chiral carbon nanotube for
7.59 and 25.29 using Timoshenko beam model is listed in Table 1. In this
chapter, the effect of thermal vibration is represented by a parameter , which is a function
of the thermal expansion coefficient of the SWCNTs. Furthermore, the thermal expansion
coefficient depends on their diameter and chirality of the SWCNTs and is very
complicated [47]. For the case of 0.2 , it is equivalent to a temperature of about 500K for
6
1 10 1/ K and L/d=10. It can be found that the frequency of the SWCNTs for
7.59 and 25.29 decreases as the value of L/d increases. In addition, the frequency
also decreases as the value of increases. This can be seen from Eq.(7), which shows that
increasing the value of L/d increases the thermal effect .
The method developed in this chapter is used to analyze the resonant frequency of
SWCNTs with simply supported ends subjected a thermal vibration, which can also be
extended to apply to cantilevered ends and clamped ends.
Table 1. The fundamental frequency of chiral carbon nanotube using Timoshenko beam
model for various thermal effects.[40]
Chiral angle Aspect ratio Thermal effect Frequency

( ) (L/d) ( ) (GHz)
4.132 0.2 377.1
7.59 0.3 375.1

0.4 373.1
0.2 288
6.18 0.3 286.5
0.4 285
0.2 229.8
8.214 0.3 228.6
0.4 227.4
25.29 3.985 0.2 385.1
0.3 383.1
0.4 381
0.2 291.7
6.07 0.3 290.2
0.4 288.7
0.2 232.5
8.1 0.3 231.3
0.4 230.1
Figures 15(a) and 15(b) represent the oxide height and oxide width at anodization
voltages of 7~10 V and as a function of the anodization time. Figure 15(c) depicts a sequence
of AFM imaged oxide dots fabricated using the AFM- based oxidation method. The patterns
in Figure 15(c) were obtained by using the static voltages of 10 V at the different oxidation
times of 0.5, 1, 2, 5 and 10 s.
These experiments were carried out in an environment having a relative humidity of 60
%. It can be seen that the oxide height and width increased as the logarithm of the anodization
time increased and as the anodization voltage was increased. The observed relationship of the
oxide nanodot height to the probe tip‘s oxidation time can be explained by the field-assisted
oxidation theory of thin films [48]. The largest oxide nanodots corresponded to the longest
oxidation time. The oxide dots had not only grown along the vertical direction but also along
the horizontal direction.
The lateral resolution was determined by the oxide nanodot‘s width and was found to be
proportional to the oxide nanodot‘s height.
To fabricate a dot for a given size the anodization times should be shortened or lengthen
in relationship to the corresponding anodization voltages which would be increased or
decreased, respectively. This is to say when using a shorter anodization time a higher
anodization voltage should be use and when using a longer anodization time a lower
anodization voltage should be use to fabricate dots of equal mechanisms and proportion.
Figure 15(a). Oxide height of the oxidation process under different anodization times [30].
Figure 15(b). Oxide width of the oxidation process under different anodization times [30].
Figure 15 (c). AFM images of five oxide dots fabricated on the TiN surface at the different anodization
times of 0.5, 1, 2, 5 and 10 s at 10 V. [30].
To understand the oxide growth rate, experiments were performed to determine the
kinetics of the oxide process. The different size oxide dots were analyzed at the anodized
voltages of 7~10 V and between the anodization times of 0.005–100 s. During the AFM tip-
induced anodization process, the electric field enhanced the occurrence of the ion diffusion
mechanism. The relationships of the growth rate and the electric field strength at a relative
humidity of 60% are shown in Figure 16(a). In this figure it can be seen that the larger growth
rates were obtained and therefore a higher electric field strength occurs when the applied
anodization voltages was larger. The initial growth rate was on the order of ~100 nm/s at
applied voltages of 9 and 10V, and rapidly became smaller when the electric field strength
was smaller. When the applied voltage increased, the initial growth rate was increased. The
anodization process was enhanced when the electric field strength was at an order of ~2×10 7
V/cm.
As pointed out in Avouris et al. [48], an equation was used to describe the growth
kinetics as follows:
dH H
exp( ) (15)
dt lc
where H is the oxide height at time t, and lc is a characteristic length depending on the
anodization voltage. The relationships between the growth rate and the oxide height at four
different applied bias voltages of 7, 8, 9 and 10 V are plotted in Figure 16(b).
In this experiment it was observed that the larger the oxide height was the slower the
growth rate became. The applied bias voltage extended the electric field strength assisting the
oxidation mechanisms until the growth was limited by the diffusion.
Figure 16(a). Growth rate vs. electric field strength [30].
Figure 16(b). Growth rate vs. oxide height [30].
The oxide rate is not only a function of electric field strength but also appears to depend
on the bias voltage applied to the probe. The Cabrera-Mott theory [49] of field-induced
oxidation cannot account for this kinetics observed in this experiment. Attempts to explain the
differences between the kinetics of AFM-induced oxidation and the Cabrera-Mott field model
have invoked the thought that the possible reasons are such mechanism as: the mechanical
stress created and arisen within the oxide dots because of a large volume mismatch between
the specimen and the oxide dot‘s structure [50]; the space charge build-up within the oxide
dots [51]. In order to explore the growth behavior of the oxide, dot patterns were fabricated
by applying voltages with on/off pulse durations of 0.01 ~ 1 per second. The pulse voltage
had a square wave with a constant amplitude and a rest amplitude of 0 V. The relative
humidity was kept at 60% during the anodization process.
Figure 17 shows the dependence of the width of the oxide dots using different pulse
periods under anodization voltages of 7~10 V. It can be seen that a higher oxide structure was
created by the pulse voltage period than that of the static DC voltage and a shorter pulse
voltage caused a higher oxide growth rate.
This reason can be explained by the static DC voltage being applied to the AFM tip and
OH-ions drifting through the anode continuously inducing the growth of the oxide on the
sample‘s surface. If a pulse voltage is applied, the transportation of OH－ions will be broken
during the rest duration of the pulse voltage and will cause the oxidation growth to stop.
When the next pulse voltage is started, the transportation of the OH-ions will reinitialize. This
is due to the OH-ions traveling randomly during the rest period of the pulse. When the active
duration of the voltage is shorter, a stronger suppression of the oxide growth will be achieved
[51].
Figure 17. Oxide width of the oxidation process under different period of pulse voltage [30].
A moving carbon nanotube tip was used to create the nanowires. This method can easily
control the growth configuration and generation position of the TiN oxide nanowire. In order
to understand how the tip speed affects the oxide nanowire the static DC voltage operation
was used under different tip speeds. Figure 18 shows the relationship of the tip speeds on the
oxide height and width. The oxide nanowire generated at the applied voltages of 7, 8, 9 and
10V and at a relative humidity of 60%. When a higher speed was used to induce the oxide, a
slightly lower height of the nanowire was achieved at tip speeds of 0.1~10 m / s .
Figure 18. Oxide height and width at different scanning speeds [30].
Figure 19. Auger analysis on both local oxidized region and unmodified region of the TiN film [30].
Comparing the height of the nanowires between that of nanodots under the same voltage.
The height of the nanodots was found to be higher than that of the nanowire. This is due to a
higher tip speed during the oxide process and therefore the electric field being reduced and
the oxide time being shorter.
In order to understanding the composition of the local oxidized structure, Auger electron
2
microscopy analysis was conducted on an oxidized zone of 10 x 10 m . The Auger spectra
of the local oxidized region and the unmodified region are shown in Figure 19. It can find that
the clear emission peak of O with kinetic energy of about 502 eV. The oxygen to Ti intensity
ratio is greater in the local oxidized region than that in the unmodified region. This is
consistent with the result of the previous study [52]. They suggested that an enhanced
incorporated process of oxygen occurred and the weaker oxygen occurred in the unmodified
region originated from the native oxide [52].
Chemical analysis of the TiN is complicated by the fact the KL23L23 Auger electron
emission from nitrogen occurred at energy that overlaps the L3M23M23 transition from Ti at
the kinetic energy of about 385 eV [53]. Since there is no N Auger transitions above 400 eV,
the L3M23M45 transition of Ti at the kinetic energy of about 418 eV. This result showed that
the anodic oxidized TiN films contains the TiNxOy transition layer.
CONCLUSIONS
In this chapter, the synthesis and characterization of Co-based MF-CNTs and the effects
of gas composition on the MWCNTs films grown using ECR-CVD method are studied. The
result showed that the broadband peaks of the D and G bands are at about 1350 and
1
1580 cm , respectively. The optimum relative intensity ratio of the D band to G band
(i.e., I D / I G ) was 2 in the experiments. The images from FEG-TEM showed that each
MWCNT in the bundle has a different length with a bamboo-like structure. Observation by
TEM revealed that the MF-CNTs were composed of poorly ordered graphene layers and were
the same as those found in the micro Raman spectral results. In addition, an analytical
solution of fundamental frequency for chiral SWCNTs, which is subjected to a thermal
vibration, has been derived and a closed-form expression is yielded. The frequencies obtained
from Euler beam model and Timoshenko beam model are compared. Results showed that the
frequency calculated by Timoshenko beam model is lower than that obtained by Euler beam
model. The discrepancy was more pronounced as the nanotube aspect ratio of length to
diameter L/d decreased. The frequency for chiral SWCNTs decreased as the effect of thermal
vibration increased.
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Lecture Material 4
HOW GAS-CARBON NANOTUBES SURFACE

INTERACTIONS DO HAPPEN ?
OBJECTIVE
While numerous studies concerning the synthesis and materials science applications
of carbon nanotubes have been conducted, the data obtained at well-defined ultra-high
vacuum conditions by means of surface science techniques is still scarce. We present
specific examples of recent ultra-high vacuum surface science projects from our
laboratory conducted on clean carbon nanotubes with an emphasis on applications in
heterogeneous catalysis. In addition the related surface science literature is summarized
and future directions are outlined.
In particular, kinetics and molecular beam scattering (dynamics) results for alkanes,
alcohols, and thiophene are described. Alkanes perfectly allow for characterization of
different adsorption sites on carbon nanotube bundles, while alcohols are related to fuel
cell applications, i.e., to renewable energy production and a green (environmentally
friendly) chemistry approach. Thiophene is the probe molecule of choice to characterize
desulfurization catalysts which are important for the petroleum industry. Kinetics
experiments quantify the binding strength of these probe molecules on carbon nanotubes
and provide evidence for capture effects, which are one of the main advantages of
nanotubes in catalysis. Molecular beam scattering has characterized gas-surface energy
transfer processes and, again capture effects, i.e., the adsorption of the probe molecules in
carbon nanotubes.
In addition, a brief review of the related literature is included. The literature survey
focuses on experimental research conducted at high vacuum (< 10 -6 torr) where common
surface science techniques have been applied. A number of theoretical studies addressing
a structure activity relationship (SAR) are included too. However, electrochemistry or
pure catalysis studies (conducted at high pressure and high temperatures) are mostly
omitted.
Finally, an outline of future directions in surface science on CNTs is kindly
proposed.
ABBREVIATIONS
Materials
AC activated carbon
BNNT boron nitride nanotubes
BCNTs bamboo-like carbon nanotubes
Bucky paper t hick multi-wall CNTs layers
C60 Buckminsterfullerene
C70 C76 C84… Fullerenes
AP-CNTs as-prepared CNTs (soot)
CNTs carbon nanotubes
DWNT double-wall CNTs
SWCNTs single-wall carbon nanotubes (less common is C-SWNTs)
SWNTB single-wall carbon nanotube bundles
MWCNTs multi-wall carbon nanotubes (less common is C-MWNTs)
cut-CNTs CNTs cut by means of acidic treatments (not a very common term);
see o-CNTs
o-CNTs open-end CNTs
c-CNTs closed-end CNTs
f-CNTs functionalized CNTs
SC-CNTs semiconducting CNTs
s-CNTs semiconducting CNTs
CNFs cup-stacked carbon nanofibers
x@CNTs With x for a metal, e.g. Au@CNTs. Metal nanoparticles (Au)
functionalized CNTs
SWNHs single-wall carbon nanohorns
SDS sodium dodecyl sulfate
EG ethylene glycol
EG epitaxial graphene
HOPG highly oriented pyrolytic graphite
IF inorganic fullerene-like materials (non-carbon nanomaterials)
MeOH methanol
MCG micromechanical cleavaged graphene
Nanobud C60-CNT hybrid material
NT nanotubes
Peapods C60 inside NTs
TiNTs TiO2 nanotubes
Techniques
BSSE basis set superposition error

CVD chemical vapor deposition
CCVD catalytic CVD
DFT density functional theory
EDX energy-dispersive X-ray spectroscopy
FTIR Fourier transform infrared spectroscopy
GCMS Grand canonical Monte Carlo simulations
HF Hartree-Fock
HREELS high resolution electron energy loss spectroscopy
IR infrared spectroscopy
LEED low energy electron diffraction
LDA-DFT Local density approximation DFT
GGA-DFT Generalized-gradient approximation DFT

MAS NMR magic angle spinning NMR
MP2 Moller-Plesset perturbation theory
MBS molecular beam scattering
MM molecular mechanics simulations
MD molecular dynamics
NEXAFS near edge X-ray absorption fine structure
NMR nuclear magnetic resonance
RR retarded reflector (King and Wells) technique – adsorption
transients
RAIRS reflection absorption infrared spectroscopy
STM scanning tunneling microscopy
TDS / TPR thermal desorption spectroscopy/temperature programmed reactions
UVvis-NIR Ultra violet-visible-near infrared spectrophotometer
XPS x-ray photoelectron spectroscopy
Further Symbols and Acronyms
d diameter of CNTs
n, m chiral indices of CNTs, i.e. (n,m)-CNT
S0 initial adsorption probability
S( ) coverage dependent adsorption probability
coverage, surface particle density
L Langmuir (one sec gas exposure at 1x10-6 torr)
SAR structure activity relationship
NSF National Science Foundation
ASW amorphous solid water
CW clustered water
CW crystalline water
CoMoCAT s-CNTs (South West Nanotechnologies)
HV high-vacuum
HiPco high pressure CO disproportionation synthesis of CNTs
HDS hydrodesulfurisation
POAV -orbital axis vector (vector pointing in the direction of a -orbital)
Pyramidalization angle p = -0º (90º for a sp2 hybridized C
3
atom, 19.5º for sp -hybrid orbital, see e.g. ref.[25, 379])
Angle between and orbitals (90º for sp2, 109.47º for sp3
orbitals)
Misalignment angle of POAV of different carbon atoms [104]
UHV ultra-high vacuum (typically pressure <5x10-10 mbar)
INTRODUCTION
Recent UHV (see sect. 9 below for a definition of abbreviations) surface science projects
conducted in our group on clean CNTs are briefly summarized (sect. 5). In addition, a review
of related projects on CNTs conducted by other research groups as well as related data on
single crystal surfaces is presented (sect. 2-4). We focus on basic research conducted at
ultra/high vacuum (< 10-6 torr) and/or projects using common surface science techniques such
as TDS, IR, NEXAFS, STM, etc. Studies applying techniques such as cyclic voltammetry,
thermogravimetry, etc., or pure catalysis studies (at ambient/high pressure and high
temperatures) have not been included. Finally, an outline for future directions of possible
projects is proposed (sect. 6).
Motivation - Combining Nanotechnology and Surface Science
The combination of nanotechnology and surface science toward heterogeneous catalysis

is a promising challenge for both technical applications and fundamental research, which is
still in its infancy, even more than nano-electronics. However, successfully combining these
complementary research areas will be of enormous economic impact. One promising
application concerns carbon nanotubes as a component of catalysts for processes such as
alcohol oxidation (in direct liquid fuel cells [1, 2]), hydrogen-related technology, or Fischer-
Tropsch synthesis, [3-7] which would provide sustainable energy sources for the future.
Graphitic systems by themselves are not very catalytically active. However, using CNTs
(for a list of abbreviations see below) as supports is a promising approach. These samples are
models for porous materials and a rare experimental realization of one-dimensional systems.
Thus, these samples are pertinent for theoretical and applied studies. Metal-supported CNT
catalysts can be obtained by metal vapor deposition or by a variety of well-established ―w et-
chemistry‖ procedures. The first technique leads to ultra-clean samples pertinent for
mechanistic studies. The second approach, which can be up-scaled to synthesize bulk
quantities, leads typically to a larger dispersion of the nanometal clusters and more realistic
model systems for technical applications.
Expected Impact
The impact of studies including surface chemistry studies on, for example, CNTs, may be
summarized as follows:
(1) Heterogeneous catalysis is a multi-billion-dollar industry ($11.2 bil. in 2006) [41] of

great importance for the U.S.
(2) The strategic plan of the NSF [42] states that ―nanot echnology could become a $1
trillion/year industry by 2015‖
(3) Studies devoted to catalyst improvements, even in the long run, are part of the effort
toward ― Technology for a Sustainable Environment,‖ which is one of the new core
funding areas of U.S. funding agencies (see e.g. ref.[43, 44])
(4) Nanoscience is highly inter-disciplinary; therefore, the results obtained in surface

chemistry studies will be of great significance. For example, the gas-surface
interaction of O2 with nanostructured surfaces has been studied to design the next
generation of gas sensors [45] Therefore, we can safely assume that studies on
heterogeneous catalysis also will be significant for materials science studies
(5) We can expect that in the not-too-distant future mass production via nanotechnology
of samples tailored toward a given surface reaction will be achievable [46]. The
economic impact will be enormous
Advantages of Carbon Nanotubes as Supports in Heterogeneous Catalysis
Briefly, CNTs have the following advantages for applications in heterogeneous catalysis:
(1) Carbon materials (activated carbon, carbon black [1]) have already been used very
successfully in catalysis [1] The surface-to-volume ratio of CNTs can be even larger
than for activated carbon, i.e., all the advantages of conventionally used carbon are
preserved and enhanced [13]
(2) Supported CNTs have a higher throughput per unit volume than activated carbon, a
consequence of the larger dispersion and metal-support interactions
(3) The micro-porous structure1 of activated carbon leads to transport limitations for
surface reactions. Nanotubes have a meso-porous structure that minimizes this
limitation [49]
(4) The large diffusion coefficients for gas/liquid transport through nanotubes can
prevent catalyst poisoning [49]
(5) Functionalization of CNTs by metal nanoparticles allows for catalyst tailoring
(6) The variety in the crystal structures of CNTs may promote catalyst optimization
(7) The high electrical conductivity of CNTs is desirable for electro-chemical
applications such as fuel cells
(8) High purity avoids self poisoning of the catalyst
SAMPLE PREPARATION FOR SURFACE SCIENCE STUDIES
The following section may seem somewhat technical. However, first, the sample
preparation is pertinent for any subsequent experimental study; second, information provided
in the following is typically not included in very great detail in journal publications.
Monolayer CNT Samples: Drop-and-Dry Technique
Samples for surface science studies can be obtained based on CNT solutions using the so-
called drop-and-dry technique. The ― cooking recipe‖ outlined in the following is well tested
[53].
1
The following pore sizes, with d for the diameter, are typically used to classify different systems: micro (d < 2
nm), meso (2 to 50 nm), and macro (d>50 nm) porous materials [48].
Hydrophobic CNTs are insoluble in water, but amphiphilic surfactants such as SDS
(sodium dodecyl sulfate) can be used to obtain colloidal suspensions. In this procedure, one
micro spatula of CNTs powder was mixed with 2 ml of 1% w/v solution of SDS in deionized
water. The mixture was sonicated (using, for example, a sonication probe and an ice bath) and
centrifuged (at 100,000g for several hours). The sonication exfoliates the nanotube bundles.
The centrifugation removes the remaining bundles and other impurities including catalyst and
carbonaceous particles [53]. The supernatant consists of short isolated CNTs, as can be
confirmed by absorption and fluorescence spectroscopy as well as atomic force microscopy
[54].
A less expensive silicon wafer (10 by 10 mm) cleaned by sonicating in acetone, then
ethanol, and dried with compressed air, can be used as a support for the CNTs. The surface of
the wafer can be covered with the nanotube by simply dropping the solution on the wafer and
by allowing the wafer to dry for several hours. Several coatings may be added to increase the
CNT density. Two to 10 drop-and-dry cycles applying 50 m drops by means of a
micropipette typically leads to good samples using SDS-water as the solvent (Fig. 1). Much
smaller amounts of the CNT solution did not reveal clear nanotube effects in prior TDS and
beam scattering studies [47]. However, even if no perfectly closed CNT layer is formed, the
total surface area of the CNTs is much larger than the support surface area, i.e., possible
contributions of the support do not dominate results in kinetics experiments, as verified in
blind experiments.
Once dry, the wafer may be annealed under N2 (600 K for 30 min) in a tube furnace in
order to remove the SDS surfactant. In addition, annealing in UHV (~600 K) has been
applied. Under these conditions, the alkane backbone of the SDS molecule can be removed,
but Na2SO4 crystals remain on the surface of the sample, as can be determined by EDX and
AES [55]. According to ref., [56] annealing in an air ambient results in a loss of CNTs
starting at 900 K; however, annealing of CNT/silica samples in UHV above 600 K reduced
already the amount of adsorbed CNTs significantly, as seen by TDS [47]. It has been
confirmed by TDS experiments [47] that the CNTs thus prepared have open ends due to the
vigorous sonication that cuts the nanotubes and exfoliates the bundles. In addition, the
Na2SO4 impurities had little effect on kinetics parameters obtained, as judged by comparison
with data obtained from samples prepared by different procedures.
Variations in Sample Preparation Techniques for Surface Chemistry Studies
Solvents
Solvents such as methanol, [57] isopropanol, [58] DMF (dimethylformamide), [59] or
even water can be used, which may result in samples cleaner than those obtained with SDS.
However, it is more difficult to obtain a dense layer of CNTs with, for example, alcohol
solutions, due to the limited solubility of the CNTs. For example, 50 drop-and-dry cycles
were required for isopropanol-CNTs in order to obtain a closed CNT film on a gold foil [55].
In addition, the alcohol-CNT solution has to be resonicated for every drop-and-dry cycle.
Similarly, the sonication process will be affected by the solubility, leading mostly to bundled
CNTs. If ultra-clean samples are required, solvents such as alcohols may be preferred;
however, the sample preparation is somewhat inconvenient and mostly CNT bundles will be
formed. Recently, is has been proposed that NMP (N-methylpyrrolidone) would be the
ultimate solvent for CNTs. For a review of the solubility of CNTs, see ref.[24]
In studies applying optical spectroscopy techniques, CNTs embedded in a polymer
matrix, such as PMAOVE-water (poly – maleic acid/octyl vinyl ether), have been used [54].
Supports
Supports such as gold foil, [60] graphitized gold foil, [61] polycrystalline Al, [62] Au-
coated Ta foil, [60] or CaF2 (IR windows), [63, 64] have been used and may be advantageous
when charged probe particles are needed to analyze the samples. Pressing the CNT powder
directly in a metal grid has been used for IR experiments at ambient pressure [59, 65]. In this
case, typically thick (~1 mm) [65] CNT layers are used, i.e., interstitial sites form in contrast
to monolayer CNTs obtained by the drop-and-dry technique. Similarly, thick CNT layers
deposited (filtered) onto a membrane support and peeled off (bucky paper) have been studied
by surface science techniques [66-68] and are available commercially [69]. A passivation
procedure for silica allowing for keeping this support clean at ambient conditions is described
in ref.[70] Spin coating of CNT solution on silica has been used in ref. [71], which may
prevent the formation of CNT bundles more efficiently than the simple drop-and-dry
technique. In the same study it was concluded that the electronic structure of the CNTs is not
affected by the silica support, as determined by optical spectroscopy on solutions and
supported samples.
Sample Cleaning
The entry ports of CNTs can be blocked by functionalities (including oxygenated groups
such as C=O, C-OH as solvent residuals from the cleaning procedures used) [57, 72, 73]
which prevent the adsorption of gases inside the CNTs. These impurities can be flashed off by
high temperature annealing. In some cases extremely high annealing temperatures (~1300 K)
have been reported to clean CNT samples, [68, 74] a procedure known from cleaning of
HOPG samples; [75] in an IR study an annealing temperature of 873 K was sufficient [63].
However, to the best of our knowledge, modest annealing (< 600 K, 30 min) in UHV already
leads to fairly clean (SDS-CNT) samples [47]. In addition, annealing samples in a tube
furnace before mounting them in the UHV chamber appears to be very efficient. Much larger
annealing temperatures resulted in a destruction of CNT/silica samples. However, the
adhesion of CNTs using other supports such as graphitized Au foil is larger, allowing for
greater annealing temperatures and perhaps cleaner samples.
CNT Activation
A number of procedures have been tested to open the ends of CNTs as well as to generate
holes in the CNT side walls in order to enhance the catalytic activity of CNTs. The following
procedures have been used: wet-chemical etching (with H2SO4 and HNO), [76] etching with
gas phase ozone, [74, 77] Xe ion sputtering/etching, [78] mechanical treatments with
diamond particles, [79] annealing in CO2, [80] etc. Ozone treatments (in addition to other
procedures) have also been used to ―cl ose‖ the ends of CNTs in order to physically trap gases
in CNTs.[64] However, except for special applications and to the best of our knowledge, the
high purity raw CNT materials available in the meantime do not require this type of activation
procedure unless a functionalization of the CNTs is required. As received, CNTs showed
TDS features very similar to those of ozone or high-temperature ―act

ivated‖ CNTs (see e.g.
ref.[81]).
Experimental Pitfalls
Some questions and concerns which are typically raised are briefly addressed in the
following:
• A CNT/silica sample obtained as outlined above is typically well covered (Figure 1)

by the nanotubes applying a few drop-and-dry cycles. A closed layer of CNT is
formed. However, a SEM with a sufficient spatial resolution is essential to image the
CNT layer. Note also that dense and thick CNT layers are more difficult to image.
However, the procedure outlined above is well tested and works
• Catalytic effects of the entirely covered support (e.g. silica wafer, gold foil) are not
expected; however, pure silica samples (and other supports, e.g., gold [82]) have
been characterized [83, 84]. In addition, the total surface area of the CNTs exceeds
the support area by orders of magnitude
• Readsorption effects are not observed in UHV experiments on ―m onolayer‖ CNT
samples
• A convolution of diffusion and adsorption/desorption events, which may be observed
when studying powder samples or porous materials at high pressure, are not observed
for monolayer CNT samples in UHV experiments. The diffusion of probe molecules
is fast, as compared with the typical time scale of most surface chemistry measuring
techniques
• AES can be used [85] to calibrate the coverage of, for example, metal nanoparticles
on CNTs averaged over the spot size of the electron gun. These samples consist of a
monolayer of flat-lying CNTs. Thus, meaningful kinetics and dynamics (adsorption
probability) data will be obtained; AES can be applied
• The CNTs are randomly distributed on the support (when using the drop-and-dry
technique), but aligned CNT samples are not advantageous for kinetics studies
motivated by catalysis applications
• Depending on the amount of solution applied to the support, a ―m onolayer‖ of CNTs
is obtained. Thus, the amount of interstitial sites formed by this sample preparation
procedure is small (negligible), as judged by SEM
Figure 1. SEM of CNTs/silica prepared by the drop-and-dry technique. (top: ~50 g, bottom: ~500 g
powder in solution) [47].
Figure 2. Possible adsorption sites on CNT monolayers [50]. (The figure has been generated with the
Gaussian03 [51] software package. However, see e.g. ref.[52] for specialized graphics software
including free trial -demo- versions for graphing CNTs and nanoparticles).
Oriented (Lateral/Vertical) CNT Samples
Samples consisting of laterally ordered CNT structures have been synthesized, for
example, by means of templates such as mesoporous silica (see e.g. ref. [114] and references
therein), mostly motivated by applications in nanoelectronics and sensor design. Molecular
beam scattering on horizontally aligned CNT samples may reveal interesting anisotropy
effects (directing the beam along or perpendicular to the CNTs) which may lead to a
characterization of diffusion processes of probe molecules inside the CNTs in a unique way.
However, the CNT density on the support of this type of sample available so far would most
likely be too small in order to study them with standard kinetics/dynamics measuring
techniques; i.e., the data would be obscured by the effect of the template.
For a review of vertically aligned CNTs (membranes, etc.), see ref.[23]
Commercially Available CNT Materials
CNTs are commonly produced by 1) LASER ablation, 2) arc discharge, or 3) catalytic

chemical vapor deposition, as detailed in several reviews (see Table 1). A number of different
raw CNT powder materials are currently commercially available. For a fairly complete list of
vendors (~50), see, for example, ref.[46, 115, 116]
Table 1. Reviews and comprehensive publications about related topics can be identified
fast with this table
System Ref.
Graphite [8]
Graphene [9]
Graphite/CNTs and fuel cells [1, 10-12]
Graphite fibers [13, 14]
CNTs general introduction [15]
History of CNT discovery [16]
CNT synthesis [17, 18]
CNT growth mechanism [19-22]
Aligned CNTs (membranes etc.) [23]
Solubility of CNTs [24-26 ]
Hydrogen/CNTs [27-30]
Nanofluids and CNTs [31]
Alloy nanoparticles [32]
Electronic properties - fullerene [33]
Inorganic nanotubes (synthesis) [34-40]
Most of the early surface chemistry work (see e.g. refs.[62, 85]) on CNTs has used (ultra-
clean research-grade) SWCNTs from Carbon Nanotechnologies Inc.[117] (the company
originally founded by R. E. Smalley). To the best of our knowledge, these samples (HiPco -
high pressure CO disproportionation) [118] consist of mixed (⅓-metallic and ⅔-

semiconducting) single-wall CNTs dominated by (10,10)-CNTs (most probable length 3200
Å and outer diameter ~13.6 Å) [85].
Table 2. Studies of gas-surface interactions conducted at ultra-high vacuum conditions

and/or applying surface chemistry techniques on clean CNTs. A few theoretical studies
are also included., cf. Table 8
Adsorbates Techniques Ref.

Xe TDS, AES [60, 72, 85], [86]
H2, D2 TDS, XPS [87 58]
HF 88]
H2O IR, DFT, MD [65, 89]
H2O dissociation HF, DFT [90]
O2 TDS [45]
CO2 IR, DFT [63]
Ads. isotherms, HF [91]
IR [64]
CO DFT [92]
NO IR [93]
NO2 IR, TDS [94]
DFT [95, 96]
N2 XPS, ELS [97]
NH3, NO2 IR, TDS [98]
n-/iso-butane TDS, MBS [47]
n-nonane TDS [99]
n-pentane, n-nonane, [100]
2,2,4-trimethylpentane
n-hexane, n-pentane, n-butane [50]
n-heptane [101]
Thiophene TDS [81]
SF4 Adsorption isotherms [102]
SF4 IR [64]
CF4 Adsorption isotherms [102]
IR [59]
CCl4 TDS [61, 99]
Methanol (MeOH) TDS [103]
(alcohols): Me-, Et-, Pro-, But-, [50]
Pen-, Hex-, 2-Prop-OH
Benzene DFT-LDA [104]
DFT-GGA [105]
Benzene derivatives DFT [106]
Fused-ring aromatic compounds: NEXAFS [57]
xanthene, 9-phenanthrol, etc.
DMMP – dimethyl IR [62, 86]
methylphosphonate
Table 3. HOPG can be considered as a reference system for CNTs. Therefore, kinetics
parameters for some adsorbates are summarized. Low coverage binding energies, Ed,
(and corresponding TDS peak temperatures, Tp) are indicated. (Ed in kJ/mol, Tp in K.)
In most cases, for the monolayer adsorption regime, a first-order pre-exponential factor
of 1x1013/sec has been assumed. TDS* label systems where more complicated kinetics
(fractional order desorption, uncommon frequency factors, etc.) have been proposed;
see text and references for details. (CW: clustered water, ASW: amorphous solid water,
CW: crystalline water) (1eV/particle = 96.485 kJ/mol)
Adsorbate Ref. Technique HOPG

Ed Tp
Xe [67] TDS 21.9 60
MM 20.6
Oxygen [45] TDS 12 40
H/D [107] TDS* 57.9 445
91.7 490
Water [108] TDS 43.4 145
HREELS
[109] TDS* 132CW
RAIRS 39.9 146ASW
153CW
CCl4 [61] TDS 44.2 175
NH3 [110] TDS*, RAIS 23.2 94
Methanol [111] TDS*, RAIS 33 144
31 139, 157
n-butane [112] TDS* 40.8 ~130
n-hexane 63.0
n-octane 72.6 215
Benzene [113] TDS 46.2 150
Naphthalene TDS* 77.1 240
Coronene 125.4 380
Ovalene 212.3 480
The projects summarized in section 5 are conducted with HiPco powder-based

(purchased early in 2006) samples as well. HiPco are formed in a CVD process using an
unsupported iron (cluster) catalyst via CO disproportionation (2CO  C + CO2). The size
distribution of the Fe clusters determines the diameter and chiralities of the CNTs. [119].
Unfortunately, significant variations in the CNT diameter distribution may be present even
when using raw materials obtained from the same vendor. For the projects summarized in
section 5, HiPco CNTs (lot HPR120.3) have been used.
A few surface chemistry studies on s-SWCNTs from South West Nanotechnologies [120]
(CoMoCAT) [121] have been published (see e.g. ref.[122, 123]); projects with s-SWCNTs
are also on the way in our group. According to the vendor information (December 2007),
[120] these samples consist mostly of (6,5) and (7,5) semiconducting SWCNTs of small
diameter (~8 Å, purity 90%); near-armchair structures dominate, according to ref.[124] These
CNTs are formed using a silica-supported bimetallic Co-Mo catalyst; for details see ref.[124]
For a characterization of the CNTs‘ crystal structure (including HiPco and CoMoCAT),
mostly by optical spectroscopy, see, for example, ref.[122, 125-128] Solvent tests for
CoMoCAT can be found in ref.[26].
More specific CNTs such as pure metallic CNTs appear not to be available commercially
(March 2008) yet.
BRIEF LITERATURE SURVEY OF HOPG - HIGHLY ORDERED

PYROLYTIC GRAPHITE
A reference system for CNTs is HOPG. Therefore, a few studies on highly ordered
pyrolytic graphite (HOPG) are briefly summarized in the following (see Table 3).
Adsorption Kinetics on HOPG
Water and Hydrogen

The water adsorption/desorption kinetics follows pseudo 0th order kinetics independent of
coverage, i.e., the water-water interactions are stronger than the water-HOPG interactions
which typically also indicates formation of water clusters. The former has been studied by
TDS and the latter could be confirmed by HREELS [108]. Slightly different TDS data have
been obtained later [109]. In the latter study the monolayer and condensation peaks were
separated and the phase transition from ASW to CI resulted in a small TDS feature.
Furthermore, fractional (~0.26) order desorption has been determined and was related to the
effect of the hydrogen-bonded water network [109]. The discrepancy in these studies may be
related to different heating rates used for the TDS experiments, which can affect the
crystallization kinetics of the water layer. In addition, the adsorption process on surfaces
which do not form strong bonds with adsorbates is often dominated by surface defects which
are difficult to control experimentally.
The adsorption of atomic hydrogen/deuterium [107] and the coadsorption with water
[129] has been studied with TDS on HOPG. Accordingly, preadsorbed water blocks H/D
adsorption, but preadsorbed H/D has little effect on the water adsorption [129]. In addition,
H/D atoms cannot penetrate an adsorbed water film. Molecular adsorption of hydrogen
required extremely small temperatures (see ref.[130]), similarly to oxygen adsorption (see e.g.
ref.[68]).
Small Organic Molecules

Alkane adsorption kinetics has been studied extensively on HOPG; molecular and non-
activated adsorption is present. A dispute about the chain length dependence of the binding
energy and whether or not the pre-exponential depends on the molecular size of the alkanes is
present in the literature; see refs.[112, 135-137] for details.
Adsorption of alcohols on HOPG has been studied by TDS and IR; rather complicated
kinetics have been proposed [111].
Aromatic molecules are particularly interesting, since unexpected trends of the CNT tube
diameter on binding energies ( - interactions/stacking) have been proposed theoretically
(see sect. 6.3). Experiments on CNTs have not been conducted (to the best of my knowledge),
but experimental data on HOPG are available (see ref.[113]). Fairly large binding energies
have been obtained (Table 3). For the larger polyaromatic hydrocarbons, complicated
fractional order kinetics has been proposed. In addition, an increase in the pre-exponential
with molecular size was reported, similar to findings obtained for alkanes. The cleavage
energy of graphite (related to the exfoliation energy of CNT bundles) has been estimated.
Metal Clusters on HOPG

Extensive work has been conducted on HOPG supported metal clusters. Due to the weak
interaction of the clusters with the support, the growth morphology is dominated by surface
(support) defects. Just to mention one system as an example: Fe and Fe-oxide cluster growth
has been studied on HOPG, [138-142] carbon nanotubes, [143] Si(111), [144] sapphire, [145]
platinum single crystals, [146] and Fe [147, 148]/Fe oxide nanoparticles [149-152].
The binding energies for some probe molecules on HOPG are given in Table 3, which
can serve as a reference for studies on CNTs.
Adsorption Dynamics on HOPG
Although numerous kinetics studies about alkanes and other organic compounds (Table
3) adsorbed on graphite surfaces have been conducted, we are only aware of a few molecular
beam scattering projects (adsorption dynamics) conducted on HOPG. The oxidation and
patterning of HOPG by an effusive atomic oxygen beam, [153] methane decomposition on
platinum leading to a graphitic layer on the surface, [154] and effusive beam scattering of
large hydrocarbons [155] in order to address the growth of organics films have been studied.
Recently the adsorption dynamics of n-/iso-butane has been investigated on HOPG; [156] the
data could be explained by a simple hard-sphere model. More attention has been paid to
supersonic beam scattering with Xe for testing molecular dynamics simulations and to allow
for improvements on cube models (see e.g. refs.[157, 158]).
BRIEF LITERATURE OVERVIEW – UHV SURFACE SCIENCE STUDIES

ON CLEAN NANOTUBES
Except for a few studies conducted at high pressure, which indicated some catalytic
activity of carbon materials such as HOPG [159] towards adsorption/reaction of small
molecules, graphitic systems including CNTs by themselves are catalytically not very active.
In other words, the binding energies for small molecules on clean CNTs are very small, which
requires either very low adsorption temperatures or large pressures (high gas pressures) in
order to obtain detectable concentrations of these molecules on the surface of the CNTs.
However, a number of studies concerning the adsorption of small inorganic molecules which
applied surface science techniques have been conducted on clean CNTs (see Table 2). More
surface science projects on the adsorption of small organic molecules such as alkanes and
alcohols on clean CNTs have been performed. The UHV surface science literature on metal
functionalized CNTs is scarce (see sect. 6.1). The effect of the CNT crystal structure on
adsorption processes has, to the best of my knowledge, also not yet been studied
systematically by experimental surface science techniques at UHV conditions; a summary is
given in sect. 6. Similarly, chemical reactions such as CO oxidation, NO+CO reaction,
desulfurization, etc. have to the best of my knowledge not been considered in UHV surface
science studies on CNTs. However, see ref.[160] for a UHV project on a bimolecular surface
reaction on TiNTs. The following literature survey focuses on the adsorption kinetics of gas-
phase species at UHV conditions studied with surface science techniques, mostly using HiPco
(10,10)-CNTs.
Adsorption of Small Inorganic Molecules on CNTs at UHV Conditions
Hydrogen
Molecularly adsorbed hydrogen. The possible capture of hydrogen in CNTs was one of
the early ―hottopics‖ in the nanoscience community. However, the data base is still very
conflicting (see e.g. the discussion in ref.[130]); many reports have not been independently
verified. These difficulties are to a large extent related to ill-defined samples and measuring
conditions. Unfortunately, the binding energy of molecular hydrogen on clean CNTs is so
small (condensation temperature of ~14 K) that UHV surface chemistry techniques cannot
easily be applied. A binding energy of ~6 kJ/mol has been estimated for physisorbed
hydrogen in neutron scattering experiments on low grade SWCNTs with adsorption
temperatures as low as 25 K (Table 4) [130]. The energetics of hydrogen adsorption and
related reactions has been considered extensively theoretically (see Table 8).
Atomically adsorbed hydrogen. Hydrogen desorption at HV conditions on, for example,
NaOH-functionalized CNTs has been reported in HV-TDS experiments [161]. The etching
(adsorption) of atomic hydrogen/deuterium (obtained by dissociation of D2/H2 on a hot
filament) has been studied at quasi-HV conditions [58]. As expected, the structure of the
starting material breaks down due to hydrogen etching; desorption of H2 has been seen at high
temperatures. A non-destructive ―w et-chemistry‖ hydrogenation of CNTs is described in
ref.[162].
For reviews of hydrogen adsorption/storage, mostly studied at high pressure with typical
catalysis measuring techniques, see, for example ref.[27-30] and Table 1.
Rare Gases
The adsorption of Xe on CNTs has been studied at UHV conditions by TDS [60, 67, 72,
85] and IR [72] spectroscopy. Interestingly, condensation of Xe inside CNTs leading to 0th
order desorption kinetics has been seen. Xe forms a quasi one-dimensional phase confined in
the CNTs.[85] Depending on the cleanliness of the CNTs (varied by applying different
etching and annealing procedures) and their end termination (o-CNTs vs. c-CNTs), initial
adsorption probabilities, S0, within the range of 0.0006 to 0.0021 have been determined by
TDS.[72] Opening the tube ends increases the catalytically active surface area and results in
an increase in S0. Similar effects have later been seen in molecular beam scattering
experiments with alkanes on CNTs.[47].
In ref. [67] the adsorption of Xe on HOPG and CNTs (at low temperatures 50-170 K) is
directly compared by collecting TDS data. 25% larger binding energies of Xe adsorption on
the outer surface of CNT bundles (bucky paper) as compared with planar graphite (HOPG)
have been determined. Molecular dynamics calculations indicated adsorption of Xe in groove
sites of the CNT bundles, which leads to larger binding energies due to the larger
coordination of groove sites as compared with a planar surface.
Table 4. For a number of systems the assignment to specific adsorption sites on CNTs
(monolayer/bundles) is so far not possible or “solely” condensation of the gases inside
the CNTs has apparently been observed. This table summarizes kinetic parameters for
those systems. In most cases, low coverage binding energies, Ed, (and corresponding
experimental TDS peak temperatures, Tp,) determined for clean CNTs are given (Ed in
kJ/mol, Tp in K). In most cases for the monolayer adsorption regime a pre-exponential
factor of 1x1013/sec has been assumed. Mostly HiPco CNTs have been used as the raw
material. “TDS” refers to high pressure temperature programmed techniques similar to
UHV TDS experiments. “Ads. iso.” refers to adsorption isotherms typically obtained at
large pressures. (Ts) are calculated TDS peak temperatures using a Redhead equation
assuming first-order kinetics (1.0x1013/sec), and 1 K/sec as the heating ramp
Adsorbate ref. Technique CNT

Ed Tp
Ar [131] ads. iso. 15 (59)
H2 / D 2 [131] ads. iso. 7.5/8.8 (30)/(35)
O2 [131] ads. iso. 17.5 (69)
CO [92] DFT 49.2 (188)
(bent CNTs)
CO2 [91] ads. iso., HF 2.0 (9)
[131] ads. iso. 22.5 (88)
CH4 [131] ads. iso. ~18.8 (74)
Thiophene [81] TDS 60 (228)
Methanol [50] TDS 47.3 184
Ethanol 50.8 197
1-propanol 58.1 224
2-propanol 57.3 221
1-butanol 61.2 236
1-pentanol 63.7 245
1-hexanol 67.6 260
Acetone [132] XPS, TDS 102-249 378-858
NH3 [98] 106 (396)
NO2 [98] 114 (425)
NO-O2 [133] ―TDS‖ —- 400
CCl4 [61] TDS 44.2 (169)
Benzene [104] DFT-LDA 18.8-20.0 (71)
(79)
Dioxine [134] ―
TDS‖ 315 950
The adsorption of Xe on CNTs has also been studied extensively by theoretical methods
(see e.g. ref.[86]) and references therein. Adsorption of rare gases also has been studied
extensively at high pressure conditions by measuring adsorption isotherms; examples for
these kinds of studies can be found, for example, in refs.[102, 163, 164]. The surface area of
CNTs has also been determined precisely by the adsorption of rare gases; see e.g. refs.[85,
165]. However, these studies are mostly omitted here since we focus in this short literature
survey on UHV surface chemistry projects.
Oxygen
Motivation. The adsorption of oxygen has been investigated extensively by experimental
and theoretical techniques. This interest is motivated by applications as diverse as the design
of better sensors (based on a resistance change due to O2 adsorption on CNTs), [166, 167] and
the development of new cancer therapies (transport of O2 crossing cell membranes by means
of functionalized CNTs followed by production of radicals -using radiotherapy- which is
inducing cancer cell death) [30]. Conducting a literature search (February 2008), using
programs such as ― SciFinder‖ or ―W ebOfScience,‖ results in something on the order of 100
publications which are closely related to oxygen adsorption on CNTs. However, only a few
studies have been conducted under well-defined UHV conditions due to the small binding
energies of oxygen, which cause experimental difficulties for surface chemistry studies.
Physisorption of oxygen. Initially a dispute existed about the binding type of oxygen and
whether oxygen doping of CNTs is possible. According to a UHV TDS study by Hertel et al.
(adsorption temperatures as low as 28 K have been used), oxygen physisorbs on CNTs with
small binding energies (18.5 kJ/mol) which are, however, larger by 55% than those for HOPG
due to the larger coordination of oxygen adsorption sites on CNTs [45]. Nevertheless,
according to this study, the oxygen coverage on CNTs even at ambient (atmospheric) pressure
is negligible [68]. Chemisorbed or dissociative adsorption of oxygen has been ruled out
experimentally [68]. Most theoretical [168-171] and experimental studies agree that only a
weak physical interaction between oxygen and clean CNTs is present.
Defects and excited states. On the other hand, chemisorbed oxygen and the dissociation
of oxygen on defected HOPG has been seen in a STM study [172]. Recently the adsorption of
excited state oxygen (1 vs. 3 ground state) has been considered experimentally (UVvis-NIR
and fluorimetry), which led to surface oxide formation via 1,4-endoperoxide adsorption on
the other CNT surface. The latter is consistent with theoretical studies [54].
Small Nitrogen Containing Molecules (NO, NO2, NH3)

The adsorption of NO has been considered extensively on single crystal surfaces by
means of all standard surface chemistry techniques [173-175]. Often quite complicated results
with a number of competing decomposition pathways have been revealed; additionally the
formation of NO clusters is quite common. Therefore it may not be too surprising that the
formation of NO dimers has been observed by FTIR spectroscopy on (10,10)-SWCNTs (at
relatively large pressures of 0.001-2.85 torr) [93]. NO desorbs in the temperature range of
103-136 K, with a dissociation energy of the dimers of 15.1 kJ/mol. These studies are
motivated by the importance of CNT-based gas sensors: the electrical resistance of CNTs
changes distinctly and with short response times upon adsorption of nitrogen containing
species; see e.g. ref.[176].
The effect of CNT crystal structure on the adsorption of NO2 has been considered
theoretically by DFT calculations [96, 177]. Accordingly, metallic CNTs of small diameter
bind NO2 more strongly then semiconducting CNTs. NO2 and NH3 adsorption has been
studied by IR and TDS on c-CNTs; base pressure of the high-vacuum system used was in the
10-7 torr range [98]. Relatively small exposures (15 L) of those gases at room temperature
apparently allowed for collecting IR and TDS data. Ammonia adsorbs via the N atom and H
atoms on groove sites of bundled CNTs; NO2 adsorbs molecularly. Binding energies for NH3
and NO2 of 106 and 114 kJ/mol, respectively, have been determined. Low pressure TDS of
ammonia on oxygenated CNT powders is discussed in ref., [73] as well as NO-O adsorption
on CNTs in ref. [133].
Studies on N-doped CNTs [97] as well as theoretical projects on the formation of
polymeric nitrogen chains inside CNTs is motivated by the synthesis of, for example,
energetic materials (explosives) [178].
Carbon Dioxide
Even for traditional single crystal systems, rather few surface science studies179 deal with
the adsorption of CO2, which may be explained by the historic focus on CO as a probe
molecule in surface science. In addition, the binding energies of CO2 on metal surfaces are
quite small. On metal oxides, however, CO2 TDS allow us to distinguish pristine and surface
defects sites with rather large binding energies; see, for example ref.[180-182].
Only a few studies have characterized the adsorption kinetics, [91] adsorption structure
(theoretically [91, 177]/experimentally [63, 64, 183]), sorption, [91] and diffusion
(theoretically [184]) of CO2 on/in HiPco-CNTs. The main motivation is the capture of CO2
for air cleaning systems and the development of strategies to deal with the greenhouse gas
CO2 such as sequestering in coal mines. The pore structure of coal may be approximated by
materials such as CNTs which have well defined diameters [185]. Again, a number of studies
are devoted to (CO2) sensor design [186].
Physisorption of CO2 with extremely small binding energies has been obtained by
measuring high-pressure adsorption isotherms [91]. Therefore, it appears plausible that
apparently no UHV surface chemistry studies have been conducted so far. However, the
variation in experimental binding energies determined at high pressures is significant (see
Table 4).
A CO2 adsorption, side-on parallel to the (9,0)-CNT tube axis and adsorption via the C-
atom of the adsorbate above the center of the C6-CNT rings has been determined by HF
calculations; [91] somewhat larger binding energies have been obtained by DFT.[177]/ The
binding of CO2 shows little site specificity, with interstitial sites in CNT bundles being
slightly more reactive than grooves or internal sites [187].
Despite the small binding energies, the sorption capacity of CNT is twice as large as for
activated carbon [184]. The diffusivity of CO2 through CNTs increases with decreasing CNT
diameter, as expected due to the increasing smoothness of the potential-energy surfaces. In
addition, CO2 with an assumed spherical geometry has a larger diffusity than linear CO2, as
expected and concluded from a theoretical study [184]. Somewhat unexpected is that the
binding energies for CO2 are predicted theoretically to increase with increasing CNT tube
diameter; see HF calculations in ref.[177] (but no clear trend is evident in the DFT calculation
in ref.; [177] see Table 8 and sect. 5.3). The sorption and selectivity for CO2 adsorption on/in
CNTs in binary mixtures has been addressed theoretically; the selectivity of CNTs was
superior to that of activated carbon [185].
A number of IR studies have been conducted on CO2 adsorption; a typical base pressure
of the IR cell amounts to 10-8 torr, but high pressure gas dosing has been applied [63, 64,
183]. Commonly a red-shifting of IR peaks compared with gas-phase data is assigned to the
adsorption of the gas-phase species on internal sites, which has been nicely demonstrated by
―closing‖ the CNT tube ends after capturing the gas (see ref.[64]). According to LDS-DFT,
the IR peak shifts (softening of vibrational modes) are related to dipole-dipole coupling of the
adsorbates, curvature effects of the CNT potential, and dynamic image charges. Thus, CO2
adsorption sites can to some extent be assigned according to IR peak shifts.
Water
a) Motivation
The adsorption of water has been studied extensively on all traditional surface chemistry
model systems (see the reviews in ref.[188-190]); rather complicated multi-component
samples have been considered as well [191-193]. Questions about whether water dissociates,
how efficiently it wets different surfaces, the degree of crystallization, details of the water-ice
transition, and the cluster formation kinetics, remain the subject of controversy even on single
crystal surfaces. The traditional bilayer model proposed in the 1980s assumes molecular
water adsorption in the monolayer coverage range and formation of bulk-like ice bilayers of
water molecules hydrogen bonded to the monolayer prior to the condensation of ice
multilayers. At very low adsorption temperatures water monomers may be present, which
cluster at greater temperatures forming amorphous solid water (ASW). At even higher
temperatures, crystalline ice (CI) is formed.
From a fundamental point of view, water adsorption in CNTs is exciting, since new
structures of water clusters and unusual hydrogen bonding are expected theoretically due to
confinement effects which cannot be observed in bulk water. In addition, the weak interaction
of water with the CNT walls and the smoothness of the walls allows for fast water transport
through CNT membranes [31].
Studying water adsorption is also pertinent for a variety of applications including
catalysis, corrosion, solar energy conversion, [194] geochemistry, meteorology, etc. Most
studies on CNTs are motivated by biological applications, including drug delivery and
transport phenomena through biological membranes, [31] as well as separation of liquids, for
example, separation of water from ethanol in the processing of bioethanol [195].
A literature review, using the key words ―car bon nanotubes‖ and ―w ater,‖ results in
~1500 hits, with the number of publications increasing exponentially over time (Figure 3).
(Note that ~2100 papers including the key word ―C NT‖ alone have been published in
Physical Review B and Physical Review Letters since 1992.) However, again the literature
appears to still be dominated by theoretical studies (MD, MM, DFT, HF) and applied
research. Only a few surface science projects conducted under well-controlled measuring
conditions have been published; we are not aware of UHV experiments except those
presented in ref.[66]. Here water adsorption has been studied qualitatively on bucky paper by
TDS; a single peak at ~150 K (water condensation) has been seen [66]/ Unfortunately, the
data set is very limited, since a reactivity screening was apparently the motivation of this
study. Most studies as outlined in the following focus on the geometrical structure of ice
captured in CNTs.
Figure 3. Publications closely related to water adsorption on carbon nanotubes as retrieved in March
2008 by means of an online literature data base.
b) Structure of Water Ice in CNTs

The formation of ordered polygonal (n-gonal ring) ice cubes inside CNTs has been
concluded based on XRD experiments [196]/ These rings are stacked to ice cubes (140 Å
domain length) inside the CNTs with the distance of the ice layers determined by the
hydrogen bond length (2.8 Å). Ice rings of different diameters appear to be present, consistent
with the temperature dependence of XRD peak intensities, which indicate different (order-
disorder) phase transition temperatures. The water molecules do no interact strongly with
their containment, i.e., the CNT tube walls.
Shortly after these XRD studies, in IR experiments [89] conducted at ambient pressure, a
weak IR band could be assigned to water adsorbed in CNTs. The intensity of this IR feature
was dependent on water adsorption/desorption cycles; the IR band was absent when exposing
water at very low temperatures (due to kinetically hindered diffusion in interior sites), and
coadsorption experiments with alkanes revealed the common site blocking effects (see sect.
5.1). Water ring structures inside the CNTs, similar to those seen in the XRD experiments,
have been proposed with hydrogen bonding inside the plane of the ring (which keeps the ring
together) and hydrogen bonding between the stacked water rings. The number of water
molecules forming the ring inside the CNT simply depends on the CNT diameter. However,
for larger diameter CNTs, the coexistence of different water ring diameters is energetically
feasible, according to DFT and MD calculations [89]/ A somewhat different structure of
captured water has been proposed based on neutron-scattering experiments: an ice sheet
wrapped into the CNT (ice-shell) and a chain of water molecules inside the ice shell [197]. A
similar structure (ordered spiral of water – helical ice-sheets) has been proposed in an MD
simulation [198]/ The formation of chains inside CNTs has also been concluded for other
systems such as iodine atoms [199]. Thus, it appears that the final structural model for water
captured in CNTs has not yet been found.
Several NMR studies have been conducted at ambient pressure. In 1H NMR of
H2O/CNTs, incomplete filling of the NT with water has been concluded by means of
adsorption isotherms [200]/ By MAS 1H NMR differences in chemical shifts have been
assigned to water adsorption inside and outside the NT with apparently little effect on the
CNT chirality [201].
c) Possible Effect of Impurities and Dissociation of Water

The MD simulations in ref.[202] add a twist to the results and the models discussed so
far. Accordingly, clean CNTs are characterized by a negligible amount of water uptake at low
pressures and at 300 K, unless the CNTs are decorated by carbonyl groups [202]. Similar
effects have been proposed theoretically for hydrogen covered CNTs.[203]. However, no
direct experimental evidence for this effect appears to exist so far.
Most studies indicate molecular adsorption of water. However, dissociative adsorption
has also been considered in DFT/HF calculations [90]. Dissociative water adsorption at 300 K
(water exposure of 23 torr for two hours) has experimentally been reported in an IR study
(base pressure of the IR system 4x10-7 torr) as a minor adsorption pathway, probably related
to defect sites on the CNTs. Obviously these measuring conditions differ very much from the
traditional surface chemistry experiments described above.
Astonishingly no detailed UHV surface science studies, such as detailed TDS, XPS, UPS,
UHV IR, etc. experiments with water on CNTs have apparently been conducted.
Adsorption of Small Organic Molecules on CNTs at UHV Conditions
Small Chain Alkanes

The adsorption of small chain alkanes such as methane and ethane has been studied on
CNTs, for example, by NMR [205] and by measuring adsorption transients [206, 207].
Although a distinct enhancement (by 76%) of binding energies is present on CNTs as
compared with HOPG, [206] the binding energies are so small that extremely large pressures
or low temperatures are required to obtain a significant coverage of small chain alkanes on
the CNTs. Therefore, apparently no UHV surface chemistry experiments have yet been
conducted.
Longer Chain Alkanes

Yates et al., published a series of papers (see Table 2) related to the adsorption kinetics of
longer chain alkanes on clean HiPco (10,10)-CNTs studied mostly by TDS and IR.
Interestingly, TDS data provide ―f ingerprint‖ spectra to identify the adsorption sites of probe
molecules on CNTs in a simple way. For alkanes such as n-pentane, n-nonane, 2,2,4-
trimethylpentane, [100] and n-heptane [101], four distinct TDS peaks are seen which can be
assigned to adsorption on interior, groove, and exterior sites of the CNTs (Table 5).
Accordingly, internal sites are most reactive, followed by groove and external sites. This
assignment was based on determining gas-filling factors by means of a calibrated gas doser.
Related TDS and molecular beam scattering experiments with n-/iso-butane are reported in
ref. [47] (see sect. 5.1).
Table 5. Low coverage binding energies determined for CNTs in UHV surface chemistry
studies. Ed in kJ/mol, Tp in K. Adsorption sites: (A) internal (or endohedral sites), (B)
groove, (C) external (or exohedral), and (I) interstitial in CNT bundles. Note that TDS
features often overlap, making it difficult to separate different structures, which limits
the accuracy of the kinetics parameters. Also, differences in the cleanliness of samples
can lead to significant discrepancies in kinetics parameters. For most studies HiPco
CNTs have been used
Adsor- Tech- I: Interstitial A: Internal B: Groove C: External

batereference nique Ed Tp Ed Tp Ed Tp Ed Tp
Xe67 TDS 27 120
Xe67 MM 26 23.4
Xe72 TDS 26.8 120
Xe204 Calc. 22.6 8.7
oxygen45 TDS 18.5 60
oxygen45 MM 14.3 15.4 14.9
n-butane50 TDS 176 45 128 33 116 29
n-pentane50 189 49 154 39 136 35
Trimethyl- 229 59 183 47 167 43
pentane50
n-hexane50 211 55 172 44 160 41
Table 6. Initial adsorption probabilities, S0, as determined in UHV surface chemistry

studies. Mostly HiPco CNTs have been used
Adsorbate S0 Technique Ref.

c-CNTso-CNTs
O2 1.0 RR [45]
Xe 0.00060.0021 TDS [72]
n-butane 0.260.39 RR [47]
Iso-butane 0.310.42 RR
Similar adsorption site-specific TDS features have been detected for other non-polar
molecules such as CCl4 [61] but not for polar molecules such as alcohols [103]/ It appears
plausible that non-polar molecules interact strongly with non-polar graphitic systems, but that
the adsorption of polar molecules on non-polar CNTs is dominated by lateral interactions of
these molecules rather than by the interaction with the CNTs. Interestingly, coadsorption
experiments can be used to identify possible adsorption sites even if no distinct adsorbate
induced features are present in TDS [61, 81, 103].
Site-specific binding energies for a number of small molecules (CO, CO2, O2, etc.) in
CNT bundles have been calculated in ref. [187]. The computed sequence of binding energies
differs from the one seen for the alkanes: groove sites are more reactive than internal sites in
(10,10)-CNTs [187]/ However, alkanes have not been considered. More details are given
below in sect. 5.; see also Table 5 and Table 8.
Chlorine, Fluorine, and Phosphorus Containing Compounds
The adsorption properties of (rather) toxic (such as DMMP - (CH3O)2(CH3)P=O -
dimethyl methylphosphonate, [62, 86] dioxine - such as C12H8O2, [134] and anthrax [208])
and environmental hazardous compounds (e.g. CF4) on CNTs have been considered, typically
motivated by the capture of those compounds in CNTs [59, 61, 62, 86, 99, 102]. For example,
DMMP is a standard (and less toxic) probe molecule in studies on chemical warfare agents
[209]. The toxicity of dioxine-based compounds depends on the number and positions of the
chlorine atoms, which results in non-toxic or highly toxic chemicals. In most of these studies
no UHV surface science techniques have been used, i.e., only a very brief note is added here
for completeness.
Interestingly, in the case of CF4, internal and external adsorption sites on CNTs could be
distinguished by IR spectroscopy when comparing o-CNTs and c-CNTs.[59]. Non-polar CCl4
shows distinct adsorption site-specific features in TDS similarly to those seen for non-polar
alkanes [61]. Astonishingly, the sorption and binding energy of dioxine is significantly larger
on MWCNTs (315 kJ/mol) than on graphite (119 kJ/mol) powder (―act ivated carbon‖) based
sorbents [134]. Sugar-functionalized CNTs form aggregates with anthrax (the CNTs are
wrapped around the spores) which may lead to a strategy to prevent aerosol inhalation of
anthrax with the design of CNT based filter systems [208].
Alcohols
A detailed UHV study about alcohol adsorption kinetics on CNTs is summarized in sect.
5.2. The following sections compile what is known from traditional studies on single crystal
surfaces which serve as reference systems (see also the review given in ref. [210]).
The adsorption and decomposition of MeOH has been studied extensively on metal
surfaces, [210-212] alloys, [213] and metal oxides [214-216]. However, rather few surface
chemistry studies with methanol have been conducted on graphitic systems, [66, 217-219]. In
the monolayer adsorption regime, and according to IR and HREELS data on HOPG and metal
oxides, MeOH is adsorbed perpendicularly2 to the surface, with the O-atom pointing towards
the surface plane [111, 214]. A shift of TDS peaks to lower desorption temperatures has been
seen in the monolayer coverage range [213, 214, 220] and was assigned to repulsive lateral
interactions [214] of the alcohol molecules. No indications for bond activation has been seen
for (oxygen-free) HOPG, [111] silver, [221] NiO, [214] or Pt-Sn (ref. [213]) single crystals.
Several different adsorption phases of condensed methanol have been identified by IR
spectroscopy and TDS [111, 221, 222]. On HOPG, three distinct TDS peaks have been
observed and were assigned to a physisorbed monolayer and different multilayer structures
[111]. In most studies, a layered structure of frozen MeOH has been proposed. A mixed
crystalline/amorphous layer forms above a disordered buffer layer and a physisorbed
monolayer. The MeOH/HOPG systems have also been studied by x-ray diffraction [223] and
AFM, [219] as well as fluorinated alcohols by TDS [224]. The literature about longer chain
alcohols is scarce even for single crystal systems [66, 217-219]. For projects on CNTs, see
sect. 5.2.
2
In studies at ambient pressure with liquids of longer chain alcohols, parallel adsorption of the alcohols has been
concluded on HOPG.
Small Aromatic Molecules

The binding energy of benzene on CNTs amounts to (18-20) kJ/mol, i.e., amazingly it is
even smaller than for graphite (22 kJ/mol), according to DFT-LDA calculations, which may
be plausible considering the -stacking interactions involved; a diffusion activation energy of
15 meV has been estimated [104]. Thus, benzene physisorbs on CNTs, with little or no
disturbance of the electronic structure of the CNT. Somewhat unexpectedly, the binding
energy increases with increasing tube diameter (decreasing curvature); see sect. 6. For small
diameter CNTs, the most stable configuration is the one with the benzene molecule adsorbed
flat over a C-C bond of the CNT. Similar calculations have been conducted for azulene, [225]
pyrene, [225] cyclohexene, [105] a number of benzene derivatives (benzene with NO2, CH3,
NH2 functional groups), [106] and dichlorbenzene, [226] which lead mostly to a noncovalent
functionalization of CNTs. Very few experimental data are available, [57, 226] none at UHV
conditions, i.e., the theoretical predictions have not been verified yet.
Surface Science Studies on Other Nanostructured Catalysts at UHV

Conditions
To complement this literature survey, a few references are given in the following
concerning other nanostructured materials focusing on nanotubes.
Carbon-Based Systems
Other carbon-based nanostructured systems including fullerenes, fullerene-CNTs
combinations, bucky paper, graphene, etc. certainly can and have been studied with surface
science techniques. Not to extend this short review too much, only a few of these studies are
mentioned in the following; see also Table 7.
Table 7. A few examples for other carbon-based nanostructures studied with surface
science techniques are given here, however, not always at UHV conditions. Only a few
studies are included here, since this review focuses on CNTs
Support Adsorbate Technique Ref.

C60/Au(111) — STM, LEED [227]
C60/Ag(111) Xe LEED [228]
C60 Pentacene AFM [229]
Ammonia FTIR [230]
CH4, CD4, C2H4, C2H2, CH3OH, CH3OD FTIR [231]
CO, NO IR [232]
Bucky paper Reactivity screening TDS [66]
Xe TDS [67]
Oxygen TDS [68]
SWNHs Benzene/water adsorption [233]
Water transients [234]
Table 8. Theoretical prediction of a SAR (structure-activity relationship) for CNTs and

small molecules. Most studies considered the adsorption on the outer surface of the
CNTs (d: diameter of the CNTs)
(n,m)-CNT Probe molecules SAR Technique Ref.

(10,0)(5,5)(17.0) NO2,O2, H2O, Acceptor/donor models, DFT [177]
NH3, CH4,CO2, smaller d more active
H2,N2, Ar
(8,0), bent-(8,0) CO Regions of high DFT [92]
bent-(10,10) H curvature most active [350]
(n,0)-CNTs H2 Small d most active DFT-GGA [351]
(2,2) (3,3)…(6,6) H, F Inner surface more DFT, PM3, MM [352]
active then outer surface
(8,8)(10,10)(12,12) CO2 Efficient diffusion for [184]
(20,20)(40,40) small d CNTs
graphite, (9,0) CO2 Large d most active MP2 [91]
(6,6)(7,7) …(10,10) CO2/CH4 GCMC [185]
(4,4)…(17,0) Benzene DFT-LDA [104]
(8,0)-semiconducting O2 Metal-to-semiconductor DFT-GGA [353]
(6,6)-metal transition
(n,0)-CNTs H2 DFT-GGA [354]
Benzene/water adsorption has been studied on SWNHs (nanohorns – CNTs with cone-
shaped tips), however, at ambient conditions. SWNHs may be interesting for catalysis
applications, due to the rather large density of defects which may act as active sites for
surface reactions [233]. For a UHV TDS study about C60, see ref.[235] C60 supported on
silver single crystals has been studied by LEED in ref. [228] as well as C60/Au(111) by STM
in ref.[236] A reactivity screening of bucky paper (thick layer of CNTs) is summarized in
ref.[66] TDS of Xe on bucky paper is discussed in ref.; [67] for oxygen adsorption, see
ref.[68]
Non-Carbon Based Nanotubes – Inorganic Nanotubes

In the meantime, it is well known that nanotubes can be synthesized from a large variety
of materials. Non-carbon based nanotubes are typically referred to as inorganic nanotubes.
See refs.[34-40] for reviews, which mostly detail the synthesis of inorganic nanotubes. Two
inorganic NT systems will be mentioned briefly in the following.
a) TiO2 Nanotubes for Photocatalysis

TiO2 nanotubes are particularly promising for applications in photocatalysis [237-239]
Nitrogen doping of TiNTs has been used to adjust the band gap in order to efficiently harvest
sunlight [240-242]. In addition to new concepts for alternative energy generation with the
next generation of solar cells, improvements to catalytic processes for the decomposition of
chemical warfare agents and pesticides are urgently needed (cf., sect. 5.3.3). Most countries
have difficulty adhering to deadlines for reducing their arsenals of chemical weapons [243]
(25,000 tons in the U.S. alone) [7]. One promising strategy is catalytic oxidation using light,
heat, or both for the decomposition of these agents. Efficient catalysts useful in remote
locations would need to operate under ambient conditions, to require only air, and to be a
non-toxic material. A TiO2-based system is a promising candidate. The effect of TiNTs could
be two-fold: capture of toxic compounds during the night and photocatalytic decomposition
of the toxin in the daytime utilizing sunlight. TiO2 catalysts can also be coated on many
surfaces and can be used in air purification devices for vehicles and buildings.
A UHV surface science study about oxygen adsorption on TiO2 is given in refs.[244,
245] - a large number of photocatalytic reactions are oxidations, with the adsorption of
oxygen as the first elementary step in the reaction sequence. Interestingly, TiNTs were
intrinsically active towards oxygen adsorption, whereas the extensively studied rutile
TiO2(110) single crystal system [246] is catalytically not very active unless this single crystal
surface is heavily reduced.
In addition, TiNTs of different polymorphs (anatase, rutile, mixed systems) as well as
amorphous and polycrystalline TiNTs can be synthesized as thin film or powder samples [40].
This large variety in the material allows for catalyst tailoring and provides a unique
opportunity to compare the catalytic activity of the anatase and rutile polymorph of TiO2.
Whether anatase is catalytically more promising than rutile or not is a long-standing twist in
catalysis and surface science [247]. A kinetic structure activity relationship for the adsorption
of alkanes has indeed been seen in a UHV study on TiNTs.[245, 248].
The adsorption of CO and CO2 by TDS has been studied in ref.[245], in addition to the
adsorption kinetics of alkanes on TiNTs in ref.[245, 248].
Clean TiNTs were active at low temperatures for the CO oxidation reaction (see
ref.[160]). A few surface chemistry studies on metal supported TiNTs have already been
conducted; see ref.[249] and references therein.
b) MoS2 and WS2 Nanotubes for Desulfurization Catalysis

The active components of some HDS catalysts consist of MoS2 nanoparticles [250-252]
We are aware of a few high-pressure catalysis studies [252-254] where the adsorption of
thiophene on MoS2 nanotube/nanoparticle powder samples has been studied; the
nanocatalysts showed promising HDS characteristics. Furthermore, the catalytic activity of
MoS2 nanoparticles for thiophene HDS has been demonstrated in model studies under UHV
conditions [255]. High activity for methanation by MoS2 nanotubes has been reported [256].
The hydrogenation of olefins has been successfully studied on MoS2-based catalysts [257-
260].
The water-gas shift reaction is catalyzed by MoS2-based catalysts, too.[261] MoS2
nanotubes reversibly intercalate-deintercalate Li/Li+; excellent reversibility and stability have
been reported with capacities up to 180 mAh/g.[262]. Thus, MoS2 may be interesting for the
design of batteries. The MoS2 and WS2 nanotube/nanoparticle morphology has been
characterized by a variety of techniques, including Raman scattering, [263] SEM/TEM, [264]
EDX, voltametry, BET, [262] XRD, STM, [265, 266] AFM, [267] and thermogravimetry
[268]. Concerning the atomic structure of MoS2 and WS2 nano-particles/tubes see, for
example, refs.[39, 264] For a UHV TDS study of thiophene/WS2-IF, see ref.[269].
DETAILED EXAMPLES OF SPECIFIC SYSTEMS

Below a few examples are discussed in more detail based on recent work (CNTs [47, 50,
55, 81, 103, 270, 271], TiNTs [160, 244, 245, 248], WS2 nano-particles/tubes [269]) by our
group at NDSU.
Adsorption of Alkanes on CNTs
Motivation
Kinetics. Studies of the adsorption of hydrocarbons have a long-standing tradition in
surface chemistry due to their importance as building blocks in the petroleum industry.
Therefore, TDS studies characterizing the adsorption kinetics of alkanes have been conducted
for a large variety of single crystal surfaces such as graphite, [137, 272, 273] sapphire, [274]
ZnO, [275] Ru, [276] Cu, [277] surface alloys, [278] and silica [83]. At low exposures, the
alkanes form a monolayer, providing information about kinetics parameters which can reflect
specific adsorption sites, [181] and reveal the effect of lateral interactions [279]. At greater
exposures, multilayers grow with desorption temperatures independent of the surface and
adsorption site. As a unique feature of CNTs, the adsorption on grooves, external, and internal
sites (see Figure 2 and Figure 4) can be distinguished, with TDS providing ―f ingerprint
spectra.‖ In doing so, high energy binding sites [163] and gas adsorption in interior sites [100]
have been identified by TDS (see Table 5).
Dynamics. The adsorption dynamics of alkanes have been investigated by molecular
beam scattering on metal and metal oxide surfaces by means of adsorption probability
measurements mapping the potential energy surface (see e.g. refs.[275, 277, 280, 281]). For
most systems, molecular and precursor mediated adsorption is present, i.e., S( ) remains
approximately independent of the coverage, since the gas-phase species are trapped in
extrinsic (above occupied sites) and/or intrinsic (above clean sites) precursor states as a pre-
requisite for adsorption. At low adsorption temperatures and large impact energies, S( )
often increases with , i.e., adsorbates assist the adsorption of gas-phase species (adsorbate-
assisted adsorption) [275]. Without precursor states, S( ) decreases linearly with
(Langmuirian adsorption dynamics). For a number of metal surfaces, bond activation has
been observed (see e.g. ref.[281, 282]) and very recently also for a few metal oxides [283,
284]. The initial adsorption probability is typically independent of adsorption temperature and
decreases with increasing impact energy.
Kinetics of Alkane Adsorption on CNTs

As a typical example, Figure 4 depicts TDS data of n-pentane adsorption on a clean silica
support and on a CNT/silica sample, as a function of alkane exposure. These data sets look
entirely different, indicating that the CNTs indeed form a closed layer on the silica, i.e., the
CNT/silica data are not obscured by silica (support) effects.
For the silica support, two TDS peak are present (labeled as D and ). The leading edges
of the low temperature D-peaks, observed at large exposures, line up, indicating 0th-order
kinetics and a condensation of the alkane. The -peak is already present at low exposures and
shifts to lower desorption temperatures with increasing coverage.
Figure 4. TDS experiments of n-pentane on I) silica and II) HiPco CNTs/silica.[47] (A: internal, B:
groove, C: external adsoption sites, D: condensation structure; : monolayer adsorption on planar
catalyst).
Thus, first-order molecular adsorption/desorption kinetics and repulsive lateral

interactions are present. Both features are typically observed for alkane adsorption [275, 279].
In contrast, exposing n-pentane on the silica supported carbon nanotubes sample leads to
the detection of four distinct TDS structures (labeled as A through D). The D peak is easily
identified as a support-independent condensation peak; the leading edges line up. The C, B,
and A TDS structures have been seen before for a variety of alkanes including n-pentane.
These structures can be assigned to adsorption on interior (A), groove (B), and exterior (C)
sites of the carbon nanotubes (see sect. 5.1). Therefore, TDS provides fingerprint spectra,
allowing in a simple way to identify different adsorption sites on CNTs. In addition, these
data indicate that the CNTs‘ ends were open.
Similar experiments have been conducted for a number of linear and branched alkanes
(Table 5). In all cases adsorption site-specific peaks have been observed. Note, however, that
the CNT samples studied so far consist of a mixture of metallic and semiconducting CNTs;
also, the distribution of their diameter and length was rather large. This fact may explain the
rather large width of the TDS peaks, indicating a superposition of a number of kinetically
distinct adsorption sites belonging, however, to the same category of sites (i.e., internal sites,
external sites, etc.). This conclusion is consistent with recent kinetic Monte Carlo simulations
[271]. Related experiments with more specific CNT samples are underway in a number of
laboratories.
Figure 5. Coadsorption experiments can reveal adsorption sites. Here, a constant exposure of n-pentane
and a variable amount of methanol have been exposed on the monolayer HiPco CNT sample. As
evident, the methanol replaces the pentane from A sites into B sites and, further, with increasing
exposure, from B to C sites. At the largest exposures the methanol is pushing pentane even in
condensation (D) sites. However, no complete site blocking is seen, i.e., a mixed pentane-methanol
coadsorption phase is formed.
Adsorption Dynamics of Alkanes on CNTs

The adsorption dynamics of alkanes on CNTs, i.e., the gas surface energy transfer
processes, have been characterized by molecular beam scattering [47]. As a typical example,
Figure 6 shows the impact energy dependence of the initial adsorption probability for n-
butane adsorption on c-CNTs and o-CNTs. Note that one advantage of the King and Wells
technique, [285] which has been applied in this study for measuring adsorption probabilities,
is that it does not require any calibrations (e.g. flux, coverage) to obtain absolute S0 values
with high precision.
Figure 6. Initial adsorption probability of an alkane on open (annealed) and closed (as-prepared) HiPco
CNTs/silica as precisely measured by molecular beam scattering techniques [47].
Interestingly, opening the tube ends (accomplished in this case simply by annealing the
samples, leading to desorption of solvent residuals which initially blocked the tube ends)
results in an increase in S0. In simple terms, S0 is the ratio of adsorbed to exposed particles.
Thus, increasing the number of possible adsorption sites by opening the end of the
nanotubes should increase S0, as is indeed observed. This effect is the opposite of the
commonly seen site blocking effect in coadsorption studies where S0 decreases with the
decreasing number of available adsorption sites [286, 287]. Estimating the total number of
adsorbed molecules by King and Wells‘ adsorption transients also indicated that the tube ends
had been opened efficiently. In addition, in TDS data the A peak, characteristic of internal
adsorption sites, was present after annealing the c-CNTs.[47]/ An enhancement in S0 by
opening the tube ends has also been determined (by TDS) for Xe adsorption on o-CNTs (sect.
5.1) [85].
The decrease in S0 with increasing impact energy is consistent with smaller trapping
probabilities at large impact energies. This result has commonly been obtained for molecular
adsorption on planar catalysts [180]. At large Ei, more energy needs to be dissipated to the
catalyst in order to adsorb the gas-phase species, i.e., S0 decreases with Ei. S0 was independent
of adsorption temperature in agreement with non-activated molecular adsorption of the alkane
on CNTs.
Adsorption of Alcohols on CNTs

Motivation
Direct liquid fuel cells and alcohol synthesis can be regarded as green chemistry,
providing us with electricity based on renewable energy sources. The most promising fuel
cells are based on MeOH, which is safe, renewable, and easily storable [1, 2, 11, 288, 289].
Although ethanol has been suggested, its production is more expensive than methanol‘s, and
the practical energy density is smaller.2 Recently, formic acid fuel cells (and other fuels
[290]) have attracted some attention, since it appears that the anode-cathode fuel crossover is
reduced using standard fuel cell membranas [291, 292]. Crucial to the operation of fuel cells
is the adsorption of the fuel on the electrode/membrane surface. Therefore, it is very
important to characterize the adsorption process with all available experimental and
theoretical techniques. A major problem of direct liquid fuel cells is the anode-cathode
crossover of the liquid. Besides better membranes, a more efficient catalyst to oxidize more
liquid is one solution. CNT catalysts are promising; prototypes already show superior
performance [1, 10, 293, 294]. Although CNT fuel cells so far use mostly conventional
membranes, CNT membranas [295] have also already been developed.
Bifunctional vs. Ligand Model for Bimetallic and Alloy Catalysts

Pt/Ru catalysts have shown the best performance in prior catalysis/electrochemistry
studies on direct liquid fuel cells [12, 296-298]. ―Other bimetal catalysts have been tried, but
this 50:50 Pt/Ru combination seems to be hard to beat...‖ cited from ref.[2]. This
experimental fact has so far been explained by ―bi functional‖ or ―electronic effect/ligand‖
models (see, e.g., refs.[298-300]).
The bifunctional model predicts that increasing the number of Pt-Ru pair adsorption sites
on the fuel cell electrode improves their catalytic activity. Thus, a 1:1 atomic ratio of Pt and
Ru would indeed lead to the best results. It is believed that bi-component catalysts improve
the catalyst poisoning by reaction intermediates such as CO through hydroxyls adsorbed on
Ru, facilitating the removal of CO species poisoning Pt sites by oxidizing CO. Since CO and
hydroxyl molecularly adsorb a 1:1 ration of Pt and Ru, adsorption sites would work best, as
indeed is often observed.
The ligand mechanism assumes a modification of the electronic structure of the alloy as
compared with the pure metals, leading to weaker CO-metal bonds. Thus, CO would desorb
even at room temperature rather than poisoning Pt adsorption sites on the electrode surface.
Other models have been proposed that consider, for example, the effect of sample
morphology, including the crystallographic surface planes of the nanoclusters [298].
Recently, tri-component catalysts [301] apparently showed better performance than bi-
component systems, indicating that the mechanism may be more complex than assumed in the
bifunctional or ligand models.
Some colleagues may argue that ―ev erything‖ has already been done when it comes to
CNTs. Although this may hold true for materials science applications, only a few studies exist
that bridge nanoscience and heterogeneous catalysis by means of surface chemistry
techniques [61, 302-306]. In addition, one may argue that studying single crystal analogs such
as Pt and Ru surfaces as well as PtRu surface alloys would be more promising to obtain
mechanistic information. First of all, these systems have already been studied extensively.
Second, it is evident that nanoparticles behave quite differently from their single crystal
counterparts [307, 308]. Thus, it appears to be the next logical step to study metal
functionalized CNTs. However, to the best of our knowledge only very few detailed UHV
surface chemistry studies have so far been conducted on metal-on-CNT systems. XPS has
been applied to NO-Rh@CNTs, [309] ethanol-Mo2S@MWCNTS, [310] and methanol-

Pt@MWCNTs, [311] and DFT calculations are reported for H2-Pd4@(5,5)-CNTs [312].
Advantage of Direct Liquid Fuel Cells

This topic may be considered quite controversial [313-316]. What technology will finally
survive, hydrogen based vs. carbon based, will depend on a number of concerns, including
political and economic issues. Direct liquid fuel cells have the following advantages:
(1) Methanol is a liquid, therefore the transportation of the fuel is less critical than for a
gas such as hydrogen
(2) Methanol has a greater practical energy density than H2. The enthalpy for
combustion of H2 amounts to ~240 kJ/mol (or 120 kJ/g), which equals only 2 kJ/ml
at 200 atm. This compares with methanol‘s combustion enthalpy of ~776 kJ/mol (or
24 kJ/g), which interestingly equals 16 kJ/ml
(3) Methanol can be synthesized from coal, natural gas, or even biomass, i.e., it is
sustainable [313]
(4) CNT fuel cell prototypes are already superior to conventional systems, as stated in a
number of publications [1, 10, 293, 294] (See also refs.[3, 4, 6, 7] for patents on
CNT/Fischer-Tropsch catalysts.)
(5) Basically the same catalysts used for direct liquid fuel cells are also pertinent for
H2/O2 fuel cells
Alcohol Adsorption Kinetics on Clean CNTs

Aside from a reactivity screening of ―buckypaper‖ (thick multi-wall CNTs layers)
including (one) methanol TDS curve, [66] we are not aware of UHV surface chemistry
projects on alcohol-CNTs interactions. Before studying MeOH adsorption on supported
CNTs it is crucial to characterize clean CNTs in order to stimulate a mechanistic
understanding.
Figure 7 depicts TDS data of methanol on CNTs. At large exposures (not shown) the
TDS curves are dominated by the desorption of methanol from a condensed layer, as
identified by the low temperature edges of the TDS peaks which lined up, indicating 0th-order
kinetics. Unexpectedly, at small exposures only two structures are evident in methanol TDS
( 1 and 2 peak in Figure 7). Qualitatively similar results have been obtained with longer
chain linear and branched alcohols (Table 4).[50] Although the CNTs are open-ended and the
characteristic AC peaks have been seen in alkane TDS curves studied on the same sample,
no distinct TDS features are present for alcohol adsorption. The maximum size of methanol
amounts to 2.3 Å (according to a density functional-based geometry optimization with
Gaussian). Thus, this molecule would be small enough to fit in the HiPco (10,10) CNTs [63]
Coadsorption experiments indeed confirmed the adsorption of the alcohols on the interior
sites of the CNTs; see refs.[50, 103] for details.
The most plausible explanation for the lag of distinct TDS features for alcohols considers
dipole-dipole interactions. Graphitic surfaces such as CNTs and HOPG are hydrophobic.
Therefore the TDS data are dominated by hydrogen bonding [111, 223] within the adsorbed
alcohol mono-layer rather than by the alcohol-CNT interactions.
Thus, it appears plausible that no distinct features in alcohol/TDS have been observed
which could easily be assigned to different adsorption sites on the CNTs. In other words, non-
polar probe molecules such as alkanes and CCl4 lead to distinct (HiPco) CNT induced
features, but polar molecules such as alcohols and thiophene (see below) do not generate
distinct (HiPco) CNT induced structures while interacting with non-polar CNTs.
Figure 7. TDS of methanol adsorption on HiPco CNT/silica [103].
Indeed, the dipole-dipole interaction energies, Ed-d, are significant as compared with the
binding energies, assuming a standard distance dependence (Ed-d = -2 2/r3). For example, at a
binding distance of r = 2.4Å, which equals the dimensions of the graphite unit cell, Edd
amounts to more than 10% of the binding energy for MeOH. Thus, differences in the dipole
moments can affect the adsorption kinetics via lateral interaction. Molecular size effects are
unlikely since TDS data for a set of linear and branched alcohols did not show distinct CNT
induced features and the dipole moments decrease only slightly with increasing size of
alcohols.
An alternative explanation would consider adsorbate induced modifications of the
electronic structure of the CNTs. Related experiments on pure semiconducting/metallic CNTs
which should lead to an answer of this question are on the way.
Adsorption of Thiophene on CNTs
Motivation – Desulfurization Catalysis

One of the most serious challenges society is facing today is the utilization of clean and
environment-friendly energy production. The response to this challenge from the perspective
of a surface chemist, lies in the evaluation of advanced materials for catalysis. At present,
most of the world‘s energy supplies come from fossil sources such as coal, petroleum, and
natural gas. For this reason, the U.S. depends on petroleum imports and must accept all of the
inherent political and environmental (long and environmentally hazardous transportation)

shortcomings. Unfortunately, fossil sources will continue to be important, considering the
anticipated demand for energy in the near future, although very different concepts such as
catalytic water splitting appear to become feasible [194]. Efficiently harvesting domestic oil
in most cases requires very efficient desulfurization catalysts. Therefore, the development of
better hydrodesulfurisation catalysts for refining petroleum is urgently needed by the U.S.
economy to produce transportation fuels with ultra-low sulfur content that obey stiffer
legislative requirements. Furthermore, desulfurization catalysts are of pivotal importance for
the utilization of coal, such as coal gasification (syngas production), which is required for a
large number of applications, including methanol synthesis, Fischer-Trosch synthesis, and
fuel cells. Processing of biodiesel and related products also requires (in some cases)
desulfurization catalysts [317-319]. Furthermore, building the chemical feedstock for the
production of a variety of products (plastics, paint, etc.) requires hydrodesulfurization
catalysts. In addition to the environmental hazards caused by forming sulfur oxides while
combusting sulfur-contaminated fuel, [320, 321] sulfur-containing compounds are very
problematic for the processing of oil-derived bulk chemicals and lead to equipment corrosion.
According to ref., [322] hydrotreating catalysts represent about 10% of the total world market
for catalysis. In addition to the need for catalysts, new materials used for the desulfurization
of petroleum also have numerous applications in material science and energy storage.
Carbon Nanotubes for Desulfurization Catalysis?

A very recent application concerns CNTs for hydrodesulfurization of sulfur-containing
compounds catalyzed by metal supported CNTs [323-325]. Furthermore, preferential sorption
of sulfur compounds by CNTs has been observed [326]. According to studies on single
crystal surfaces [322, 327] defect sites as well as, according to projects on sulfided Mo
clusters,328 the rim of the metal nanoparticles are pertinent for HDS. Similarly the HDS
activity of hollow MoS2 nanoparticles was likened to an interplay of defect and confinement
effects.253 In another recent report, an enhancement for methanation activity in MoS2
nanotube powder catalysts has been observed [256] and was assigned to confinement effects.
Furthermore, inorganic nanotube powders [35, 36, 39] often consist of a mixture of
nanoparticles and nanotubes, which makes it difficult to isolate effects. Therefore, it is not
certain at this point if the curvature of the clean nanotube surface, confinement effects, and/or
defects in the tubular structure dominate the catalytic reactivity of nanotubes for HDS. The
final result of this controversy will affect the synthesis strategies for optimizing the catalytic
activity of nanotubes for HDS. To characterize confinement effects and gas-nanotube
interactions, it appears useful to consider first an expected less reactive, clean (without metal
clusters deposited) nanotube system with a small intrinsic defect density, such as carbon
nanotubes.
In HDS model studies, thiophene is the probe molecule of choice since it is among the
simplest sulfur-containing compounds present in natural petroleum. From studies on metal
[329] or metal oxide [330] surfaces an initially flat adsorption of thiophene has been
concluded. In addition, tilted molecules forming a compressed monolayer at larger coverages
are frequently observed. In addition to an amorphous phase, different crystalline phases are
documented for condensed films [331, 332]. At UHV condition, a crystalline phase has been
observed by IR spectroscopy [333].
Adsorption Kinetics of Thiophene on Clean Carbon Nanotubes

Kinetic studies of thiophene adsorption on CNTs have been conducted under UHV
conditions [81]. TDS experiments revealed molecular adsorption/desorption of thiophene on
clean CNTs. In the case of a bond activation including a desulfurization of thiophene, the
desorption of H2S, H2, and small chain alkanes are expected, which have not been observed.
The maximum molecular size of thiophene amounts to 4.6 Å. Therefore, adsorption on
interior sites of the (10,10)-CNTs is expected. Unfortunately, no distinct monolayer TDS
structures which could easily be assigned to CNT-specific adsorption sites have been
observed. Therefore, co-adsorption experiments with alkanes (similar to those shown in
Figure 5) have been conducted and have confirmed thiophene adsorption on interior sites. The
filling sequence of the different sites observed for thiophene (A and B  C  D) basically
follows the binding energy sequence seen for alkanes adsorbed on the different adsorption
sites. This suggests that thiophene adsorbs on the same sites as the alkanes with the same
sequence of binding energies. As already mentioned above, similar experiments have been
conducted with methanol and n-pentane [103]. In this case, the rather linear n-pentane
molecule could not be replaced as efficiently by the more spherical methanol from the linear
groove sites (sect. 5.1.2). Instead external sites were predominantly decorated by the alcohol,
especially at larger total exposures of both molecules [334]. Thiophene appears to adsorb
efficiently on groove sites while methanol does not. Thus, even in the absence of distinct TDS
features, coadsorption experiments can provide some insights into possible adsorption sites.
FUTURE DIRECTIONS
Two future directions for surface science research related to heterogeneous catalysis on
CNTs are obvious and in part already on the way in a number of laboratories. 1) Studying
CNTs of well-defined crystal structure (Figure 8, 9), and/or 2) characterizing CNTs
functionalized by, for example, metal-nano-clusters.
So far little is known about the effect of the crystal structure (metallic, semi-metallic,
semiconducting) on the catalytic activity of CNTs, which is a prerequisite to tailoring
catalysts. For example, ―...the performance of carbon support materials is largely influenced
by their electrical properties, morphology and crystallographic structure,‖ cited from an
electrochemistry study in ref., [335] which is related to fuel cell catalysts. Or, ―w e suspect
that the semiconducting and metallic SWCNTs have significantly different surface
properties‖ [337]. Although no systematic reactivity screening has been conducted at UHV
conditions, it appears plausible that chemical activity is related to curvature-induced strain of
the graphene sheet. Thus, the chemical activity should be related, for example, to the diameter
of the CNTs, with the smallest diameter nanotubes being the most reactive [338]. The
bottleneck to attack this problem from the perspective of a surface chemist is the availability
of high quality raw materials with well-defined crystal structure. The separation procedures
currently available are too complicated and time consuming for the synthesis and
characterization of the systems to be easily accomplished in a single research group, at least
in our opinion. Thus, interdisciplinary teams may be the solution to push the development
forward fast. To the best of our knowledge, today no detailed comparative UHV surface
science studies have been conducted on CNTs of specific crystal structure such as pure
metallic or semiconducting CNTs of a given diameter. However, comparative studies about
the CNTs‘ crystal structure, applying optical spectroscopy, have been conducted; see e.g.
refs.[71].
Figure 8. The selectivity problem in CNT synthesis involves a large number of parameters and levels of
specificity.
Figure 9. Inner diameter of CNTs as compared with the maximum size of small organic molecules
which are often used as probe molecules. Shown (solid lines) are the inner CNT tube diameters (left
scale) vs. index ―n‖ (bottom scale) and parametric in the index ―m‖ for a (n,m)-CNT. The outer
diameter, d, is given by (see e.g. ref.336); the inner diameter is smaller than d
by twice the van der Waals radius of carbon (2 x 1.7 Å). The size of a (10,10) CNT (HiPco) is labeled
as an example. The symbols refer to the maximum size of linear alcohols (solid circles) from methanol
to hexanol and the open squares to n-alkanes from butane to hexane. In this case, the left scale depicts
the maximum size of these molecules and the upper scale the number of carbon atoms of these probe
molecules. The molecular size of the probe molecules has been computed with the Gaussian 51 software
package using B3LYP/6-31G(d).
Metal functionalization of CNTs is well established and rather simple, but here also only
very few UHV surface chemistry studies are currently available [309]. The next two sections
summarize briefly the literature on the synthesis of CNTs with well-defined crystal structure
(sect. 6.2). In addition, some strategies to functionalize CNTs with nanometal clusters (sect.
6.1) are given.
Synthesis of Metal Functionalized CNTs
The functionalization of CNTs with metal clusters is, in the meantime, a developed
standard technique. The following strategies are common.
Thermal Evaporation (Physical Vapor Deposition)

Ultra-high vacuum metal evaporation commonly used in studies of model catalysts (see
e.g. refs. [142, 339, 340]) has also been applied to functionalize CNTs with metals [341].
Even high-melting-point metals can be evaporated easily by mean of resistively heating a thin
metal filament.
Commercial Materials
Some catalysis companies [342] offer a variety of unsupported nanoparticle catalysts.
These have been used successfully in ref.343 to attach metal clusters such as PtRu alloy
nanoparticles to CNTs from suspensions in ethanol. This approach appears experimentally to
be the simplest one and has additionally the advantage that an easily available standard
material with well-reproducible properties can be used.
Hydrothermal Method – Gas Solid Reduction of Precursors

In the so-called hydrothermal method, the CNTs are impregnated with, for example,
ethanol suspensions of commercially available precursors such as H2PtCl6, [11, 344]
RuCl3,[11], [344] Pd(NO3)2, [345] PtCl2, [346] etc. After drying the impregnated powders, the
precursor is decomposed/reduced in, for example, H2 ambient at high temperatures [11]. The
importance of the pH value of the suspension has been pointed out by some groups; for
example, NaOH can be added to the suspension to adjust the pH. [347]. For the reduction of
the precursor, the use of a tube furnace11 and different autoclave [344, 347] set-ups has been
reported. (The HT method has also been used to functionalize CNTs with halogen or sulfur
surface groups. [348]) A large number of variations of the hydrothermal technique have been
documented in the literature, including microwave heating [344]. Oxide formation has not
been reported for Pt, Ru, or PtRu alloy particles, but the reduction reaction with hydrogen
would also reduce eventually formed metal oxides, which has been demonstrated even for
metals such as Ni that are very reactive in an ambient of air [349]. Bimetallic Pt-Ru
functionalized CNTs have been obtained with the same procedure using a 1:1 mixture of the
Pt:Ru precursor solution [296].
Liquid-EG Solid Reductions of Precursors

Reduction of precursors such as H2PtCl6 and RuCl3 in ethylene glycol (EG)-water293
solution by means of, for example, a microwave system, is another widely used standard
technique to functionalize CNTs with metals. It has been pointed out that the pH value (i.e.,
the amount of water in the EG solution) allows for adjusting of the metal cluster particle sizes
[355]. Similarly, the same precursor can be reduced using sodium borohydride [301]. Other
precursors such as K2PtCl4 and K2RuCl5 have been used as well [356].
Other Techniques
Other techniques, including solution-phase approaches, [357] solid-state reactions, [358]
electro deposition, [359, 360] chemical vapor deposition, [346] fast evaporation, [361] and
galvanic displacement reactions, [362] have been developed. Despite a few convincing
examples, [345, 362] most techniques lead to the decoration of the outer surface of the CNTs
with metal clusters.
Activation and Cleaning Procedures

Before the functionalization process, sonication and centrifugation of the CNT
suspension were commonly applied to unfold CNT bundles and to remove catalyst particles
used to grow the CNTs (sect. 2) [47, 81, 103]. In addition, an activation procedure consisting
of etching the CNTs in KOH or H2SO4-HNO3 (typically at 350-400 K for several hours)
before impregnation with the metal particles is common. This procedure again removes
catalyst particles as well as forming oxygenated groups (such as -COOH, -OH, -COO, etc.) to
anchor the metal clusters with a large dispersion. It has convincingly been reported that the
few catalyst particles remaining from the CNT synthesis are encapsulated in graphite, i.e.,
they are catalytically inactive [345].
Synthesis of CNTs with Well Defined Crystal Structures - Separation

Strategies
Unfortunately, little attention has been paid so far to the effect of CNTs‘ crystal structure
on their catalytic behavior, at least in experimental UHV surface chemistry studies. This fact
is certainly related to the availability of specific samples. Spectroscopic techniques allow for
a precise characterization of the CNT crystal structure [126-128] (see also ref. [363] –
temperature programmed oxidation). However, the separation and specific synthesis of CNTs
are still a challenge.
Separation techniques may be classified as (1) “wet-chemistry” procedures, which are
typically based on a specific functionalization of nanotubes and differences in the solubility
of the functionalized tubes, (2) physical techniques taking advantage, for example, of
differences in electronic properties of CNTs, and (3) catalytic techniques used in the gas-
phase synthesis of specific CNTs.
(1) For example, left- and right-handed semiconducting CNTs have been separated by
functionalization with chiral probe molecules; the functionalized L-/R-CNTs differ
by their solubility [364]. The specific binding of porphyrins has been used to separate
metallic and semiconducting CNTs [162, 337], 365 Preferential solution-phase
ozonolysis of small diameter CNTs has been reported [366]. Very recently, so-called
molecular tweezers molecules have been considered to separate different crystal
structures of CNTs, a technique which may have potential for up-scaling [367].
(2) Dielectrophoresis has been used as well to separate different CNTs, however, in tiny
amounts [368]. More promising in obtaining bulk quantities may be the density-
gradient ultracentrifugation technique for metal-semiconductor separation, which is
based on differences in the buoyant densities (mass per volume) of CNTs [369]
(3) The size distribution of nanoparticles used to catalyze CNT formation in gas-phase
reactions determines the CNT tube diameters.119 Therefore, procedures to obtain
narrow particle size distribution of catalyst particles are another approach; see ref.
[370] Already commercialized is a catalytic technique (CoMoCAT) that allows for
specifically growing semiconducting CNTs; [124] for a sample characterization see,
e.g., ref. [371] The HiPco and CoMoCAT techniques are both based on CO gas-
phase disproportionation, but using different catalysts and a different catalyst pre-
processing. The HiPco technique leads to a broader diameter distribution of CNTs
than the CoMoCAT process; see ref. [124] for a comparison of HiPco and
CoMoCAT CNTs.
The theoretical perspective of CNT separation strategies is discussed, e.g., in ref.[96, 171,
372] as well as catalytic techniques in ref. [373] Considering the problem the other way
around: Separation techniques for CNTs may also lead to novel separation/cleaning
procedures for important compounds. For example, the separation of alkane mixtures by
CNTs has been considered theoretically [374]
A different approach concerning practical applications could be using so-called
Haeckelite carbon nanotubes which should have metallic character independent of tube
diameter and chirality, as theoretically predicted [18]. These tubes have, however, so far not
been synthesized.
Theoretical Predictions: Structure Activity Relationship - Effect of

Curvature/Chirality
Following simple reasoning, the catalytic activity of carbon-based nanostructures should

follow the trend (Figure 10):
fullerenes > metallic CNTs > s-CNTs > graphene
i.e., fullerenes are most reactive, graphene mostly inert, simply due to the decrease of the
curvature-induced strain (pyramidization) of the graphene sheet going from ―s pherical‖
fullerenes to planar graphene. The larger the curvature (pyramidalization angle; see Figure 11,
12), the larger the reactivity. Therefore, the reactivity of (the outer surface of) CNTs should
increase with decreasing diameter. Similarly, interior sites of CNTs provide more nearest-
neighbor sites for adsorbates and should be more reactive than exterior sites of CNTs.
Furthermore, metallic CNTs with comparable diameters/curvatures may be catalytically more
active than semiconducting CNTs. Detailed theoretical studies, however, predict different
trends and more subtle effects in some cases, which will briefly be discussed below. The
theoretical data base is quite extensive. Unfortunately, most theoretical studies have been
conducted on rather exotic systems which can most likely never be studied experimentally.
However, interesting trends have been seen and some concepts, including SAR rules, have
been proposed. Unfortunately, no detailed experimental surface science data are available yet.
Figure 10. Expected structure activity relationship based on the curvature of the nanostructures which
appears to be obeyed in the case of covalently bonded probe molecules.
Figure 11. The carbon atoms in planar graphite (HOPG) are well described by a sp 2-hydridization,
which is not necessarily the case for a bent graphene sheet (i.e. CNTs). The pydamidalization angle
allows quantifying the effect of CNT curvature on binding energies of probe molecules using molecular
orbital theory.
Donor-Acceptor Models
According to DFT calculations, [177] a most distinct SAR is expected for charge
acceptor molecules (such as O2, NO2) which significantly modify the density of states of the
CNTs, whereas charge donors (such as N2, H2O, CO2) would interact only weakly and non-
specifically with CNTs. [177] Variations in binding energies as large as 36 kJ/mol (for NO 2)
and 20 kJ/mol (for O2), depending on the diameter of the CNTs, have been proposed
theoretically. A clear trend for the binding energy on tube diameter –smaller diameter tubes
are more reactive – has been seen for NO2, O2, NH3, and CO2 but not for H2O, CH4, H2, N2,
and Ar. Although trends in binding energies are typically correctly predicted by DFT, some of
the theoretical binding energies appear not to be consistent with available experimental data.
POAV Misalignment – Noncovalent Interactions

Astonishingly for CO2, binding energies which increase with increasing CNT diameter
have been predicted theoretically; [91,185] similar trends have been proposed for benzene
physisorption [104]. Benzene adsorbed more strongly on HOPG than on CNTs, according to
theoretical predictions. Experimental data are not available.
The calculations predict a non-trivial dependence of binding energies on curvature and
chiral angle of CNTs. However, a simple SAR rule has been stated: ―t he smaller the orbital
axis vector (POAV) misalignment angle is, the larger the binding energy.‖[104] (See Figure
12.) Although, all C atoms in a CNT are equivalent, the C-C bonds are not. These bonds
differ by the direction of the orbitals (POAV) with respect to the -orbitals (the bond
directions) of the C atoms in the CNTs. Due to the CNT curvature, the POAV of neighbor C
atoms is misaligned. This misalignment depends on the tube diameter. In turn, a perhaps
unexpected dependence of the CNT reactivity (e.g. binding energy) on the diameter (and
curvature) of the NT is predicted theoretically [104] for noncovalent interactions of CNTs
(such as benzene adsorption on CNTs). However, experimental data confirming the
theoretical predictions are missing so far.
The POAV misalignment leads to kinetically distinct adsorption sites on the CNT
surface. In addition, the curvature-induced pyramidalization ( > 90º, for C60: p=11.6º, for
(5,5)-CNT: p(side wall)=6.0º, see Figure 12 and ref. [25]) of the C atoms leads to deviations
from the carbon sp2-hydridization seen for planar HOPG; the amount of s-p mixing has been
calculated in ref. [104] Both effects (POAV misalignment and pyramidalization) induce a
local strain in the CNTs. For the same reasons, the fullerene caps of c-CNTs should be more
reactive than the CNT walls [25]. The larger p, the larger the reactivity of the system. CNTs
may be considered ― cylindrical aromatic macromolecules‖ when applying MO-type theory to
understanding the adsorbate-CNT interactions, as discussed in ref., [25] when comparing the
electronic structure of fullerenes with CNTs.
In summary, for weakly interacting adsorbates (with dominating -orbitals) a trend of
increasing reactivity with increasing CNT tube diameter is predicted. Unexpectedly, large
diameter tubes are catalytically more active than small diameter CNTs.
Covalent Interactions
Probe molecules which result in a more covalent binding to CNTs show the opposite
trend: smaller diameter CNTs are more reactive; i.e., the binding energies increase with
decreasing diameter. Artificially bent CNTs have been considered in a number of theoretical
studies; in all cases a large curvature was related with high chemical activity [92, 350].
Figure 12. I) On a planar surface such as HOPG, the C atoms are sp 2-hybridized and the -orbitals of
different C-atoms point in the same direction, called the POAV- orbit axis vector. On a planar
surface, equivalent sets (on top sites, bridge sites, etc.) of adsorption sites exist, independent of the
given patch on the planar surface considered, i.e., all C-C bonds are equivalent. II) However, on a
curved surface, such as the CNT surface, the POAVs of different C-atoms are misaligned, as can be
quantified by the angle POAV misalignment angle . III) Thus, although all C-atoms on the CNT
surface are equivalent, having the same chemical environment, their bonds differ, which can result in
different binding energies of adsorbates depending on the CNT diameter, since certainly depends on
the diameter (curvature) of the CNTs. In addition, differs for different bonds on the CNT surface,
which can result in different binding energies for adsorbates along the surface of the CNTs.
For example, CO only physisorbs on unperturbed (8,0)-CNTs with basically repulsive

interaction (extremely small binding energy), but chemisorbs (with larger binding energies)
on bent CNTs. [92] Thus, regions of larger curvature of the CNTs are catalytically more
active — at least theoretically; experimental data is missing. Similar conclusions have been
drawn for other strongly bonded adsorbates: the smaller the diameter, the larger the binding
energies for hydrogen, [351, 352] fluorine, [352] and aluminum [351] adsorption on the other
CNT surfaces. Thus, it appears that the interactions of molecules with CNTs can be classified
according to the covalent character of the binding to the CNTs. For covalent binding, small
diameter CNTs are best; for non-covalent binding, large diameter CNTs are catalytically
more active.
Reactivity of the Inner and Outer Surface of CNTs

In ref. [352] the possible differences in the reactivity of the outer convex (excahedral)
surface and the inner concave (endohedral) surface of CNTs have been considered by
quantum chemical techniques in analogy to fullerene chemistry. The binding energies of H
and F atoms on (n,n)-CNTs have been calculated [352]. For these covalent interactions, the
binding energies for adsorption on the inner CNT surface decreases with increasing CNT
diameter. (Small diameter CNTs are ―bes t.‖) In contrast, the binding energies on the outer
surface are much smaller and rather independent of the CNT diameter, except for very narrow
CNTs, according to ref. [352]. In other words, most reactive for covalent bond formation is
the inner surface of small diameter CNTs. The trends observed in the binding energies for
small diameter (n,n)-CNTs (2<n<7) are related to the pyramidalization angle (angle between
and orbitals of the C atoms in the tube wall, Figure 12). The larger the pyramidalization
angle, the larger the binding energies on the interior surface of the CNTs, similarly to the
discussion in ref. [104] mentioned above. Unfortunately, it is doubtful that related
experiments with exotic (2,2)-CNT tubes and atomic hydrogen (sect. 3.1.1) will ever be
doable.
Kinetically Distinct Adsorption Sites in CNT Bundles and Isolated CNTs

Kinetically distinct adsorption sites have been seen for CNT robes/bundles in a number
of experimental projects (see Table 8, sect. 5.1); theoretical studies are rather rare [187, 375].
Accordingly, for most molecules interstitial sites have the largest reactivity, followed by
groove sites and internal sites [187]. Slightly different trends have been proposed in
experimental work, where the binding energies on internal sites were always larger than on
groove sites. However, these results certainly depend critically on the CNT diameter and the
geometrical arrangement of the CNT bundles.
Binding energies of small molecules on isolated CNTs have been calculated for a number
of different adsorbates including hydrogen [376] and benzene.104 For hydrogen, hollow sites
are energetically favored; benzene adopts bridge sites.
Metal-to-Semiconductor Transitions (vice versa)

A metal to semiconductor transition of metallic CNTs upon O2 adsorption has been
predicted theoretically, whereas semiconducting CNTs are less affected [353]. Related effects
have been discussed for the adsorption of hydrogen [354]. In this case, a massive
reconstruction of the CNTs was proposed theoretically; related effects have apparently been
seen experimentally [58] (see sect. 3.1.1).
IR and NMR Characterization of CNT SAR

Just as a brief note, since these techniques typically do not operate under UHV
conditions, we would like to mention that IR peaks shifts (see e.g. ref. [64]) as well as
chemical shifts in NMR, may be used to evaluate a SAR. For example, chemical shifts in
NMR increase systematically with CNT tube diameter, as theoretically determined for a
variety of probe molecules [377].
SAR of Diffusion Properties of CNT

The diffusion of molecules through CNTs is certainly dominated by surface effects.
Interestingly, it has theoretically predicted that, in addition to the tube diameter, the
crystallography of the CNTs affects the diffusion of, for example, water through CNTs (see
e.g. ref. [378]). Molecular dynamics calculations combined with calculated potential energy
surfaces predict that the diffusion of water is more efficient for zigzag CNTs as compared
with armchair CNTs. The CNT crystal structure affects the trajectories of water diffusion
inside the CNTs.
It appears the SAR of CNTs is more subtle than considering only the curvature of the
CNTs. However, most likely different classes of probe molecules (covalent vs. noncovalent
interactions) can be distinguished and SAR rules will be developed. As for now, basically no
UHV surface chemistry data are available which would characterize in detail the dependence
of binding energies on, for example, the CNT tube diameter.
SUMMARY
The surface science of CNTs studied at ultra-high vacuum conditions by traditional
surface chemistry techniques is still in its infancy (see Table 1). So far mostly mixed CNT
samples have been studied, which consist of a rather large distribution of diameters and tube
lengths. In addition, metal supported or otherwise functionalized CNT samples have, to the
best of our knowledge, not been studied yet in detail under UHV conditions with surface
chemistry techniques. The data base is even scarcer concerning inorganic nanotubes. A less-
than-optimal selectivity of catalysts which makes sophisticated separation and cleaning
procedures of the products pertinent is a major hassle in many industrial catalytic processes.
However, it appears to be just a matter of a few years before more efficient procedures will be
developed to synthesize CNTs of specific crystal structure. Most ―w et-chemistry‖ strategies
for the separation of CNTs (according to their crystal structure) take advantage of a structure-
activity relationship. Therefore, a better understanding of the SAR of CNTs is critical in this
endeavor. A number of theoretical studies (see Table 8) have already been conducted and
predict a SAR for CNTs, i.e., the adsorbate-CNT interactions depend on the CNT crystal
structure (curvature and/or chirality).
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Lecture Material 5
CARBON NANOTUBES: GROWTH KINETICS AND

FUNCTIONALIZATION WITH SILICON
NANOCRYSTALS
ABSTRACT
In this chapter the growth kinetics of most common carbon nanotubes (CNTs)
synthesis at industrial scale by catalyst assisted chemical vapor deposition (CVD) is
discussed. It is shown that the monitoring of CNTs growth at initial stages by using a
Tapered Element Oscillating Microbalance (TEOM) brings new insights into synthesis
and controllability of the CNTs properties. The high sensitivity of the TEOM technique
allows precisely determinates the crucial synthesis parameters. We argue that precise
TEOM control of the reaction temperature and the partial pressure allows evaluate the
order of the reaction kinetics and absolute reaction rate. Furthermore, CNTs solve some
challenges linked with connection and manipulation of silicon nanocrystals (Si-ncs) at
nanoscale level. Particularly, direct growth of CNTs on Si-ncs in the TEOM is performed
in order to connect single Si-nc. Compared to porous catalyst supports substantial
differences in CNTs growth kinetics are observed when the synthesis is performed on flat
Si-ncs surface. A model taking into accounts an associative and competitive adsorption of
ethane is used to interpret obtained results. The diameter of the CNTs depends on the size
of the Si-ncs, which remains connected on the tip of the CNTs. The wired Si-ncs keep
room temperature photoluminescence properties. It is shown that the CNT cavity, in
additionally, can serve as nano-reservoir for freestanding Si-ncs. Colloidal dispersion of
freestanding Si-ncs allows entering into CNT cavity by induced capillary force.
Alternatively, the shock waves generated during Si-ncs formation in transparent polymer
by nanosecond laser processing assure the filling as well. We believe that the present
findings might open new opportunities and situations in a development of new class of
nanodevices for the environmental friendly applications.
INTRODUCTION
There has been much interest in carbon nanotubes (CNTs) [1-4] last decade, especially
for potential use in microelectronic, photovoltaics and other terrestrial useful devices [5, 6].
The synthesis of CNTs can be performed by different kind of techniques. There are three
commonly used means by which to prepare CNTs. The first of these methods is laser ablation
[7]. A high power laser is applied across a carbon target and in the plasma plum the
generation of CNTs is achieved. The second is the Arc-discharge method synthesizes by
using a fairly low voltage power supply to strike an electrical arc between two carbon
electrodes. The CNTs form in the arc and collect on the anode electrode [8]. Chemical vapor
deposition (CVD) is the third most common way of CNTs synthesis [9, 10]. This means is the
most common synthesis at industrial scale and is achieved by taking a carbon species in the
gas phase. An energy source, such as plasma or a resistively heated coil, to impart energy to a
gaseous carbon molecule is used. Most often use gaseous carbon sources include methane,
ethane, carbon monoxide, and acetylene.
CVD growth of CNTs by decomposition of ethane is the means of synthesis that is of
interest for this chapter. CVD carbon nanotube synthesis is essentially a two-step process; (i)
a catalyst and (ii) actual synthesis of the CNTs [4]. Up to date, however, the mechanisms of
CVD kinetics of CNTs synthesis at early stage are under debate and not well understood yet.
The close relationship between nanotube properties and geometrical structure forcedly need
this understanding. In our knowledge, there are only few reports that investigate the kinetics
of CNTs growth in a methodical manner [11, 12]. In ccontrast to that the growths of carbon
nanofibers is better described and comprehend [13, 14]. Two methods are mainly used to in-
situ investigate the kinetic of the carbon nanofibers synthesis; (i) controlled atmosphere
electron microscopy [15], and (ii) thermogravimetric measurements [16-20]. One solution to
complete in-situ CNTs growth monitoring at early stages is accessible by using a tapered
element oscillating microbalance (TEOM) technique [21, 22]. This experimental set up
allows achieving homogeneity in contact with the catalyst and determining of absolute
reaction rate [23]. The growths of CNTs, either free standing (powder like) or solidly attached
on substrate, is achieved. At the same time an accurate reaction kinetic can be acquired [22].
The CNTs apart from being the best within the most of available one-dimensional (1D)
model systems show strong application potential as well. While some of the proposed
applications remain still a dream, others are close to technical realization. State-of-the-art
such dream is using CNTs in purpose to wire the single nanocrystals or quantum dots [24].
We like to emphasize that the excellent CNTs transport properties, with conductivity 1000
times the one of copper nowadays mostly technologically used, suggest them reach this goal.
The achievement allows solve some challenges linked with the manipulation and the device
fabrication at nanoscale level [24]. A silicon nanocrytal few nanometers in diameter (< 10
nm) is particularly attractive candidate for nanoscale device fabrication. Discovery of bright
photoluminescence (PL) in silicon nanocrystals (Si-nc) at room temperature increased hopes
in Si-ncs for many terrestrial applications [25-31]. The usefulness is amplified in a form
soluble freestanding and surfactant free Si-ncs [28, 30]. The development of reliable methods
for the connection of the CNTs with Si-ncs might provide an additional impetus towards
extending the scope of nanotubes application. The precise control of the CNTs synthesis by
TEOM on Si-ncs provides a route to connect Si-ncs with the characteristics as a will. All at
once offers the possibility to get better in manipulation and effective elaboration of Schottky
nanojunctions [24].
It is distinguished that the cavity of CNTs can serve as nano-reservoir for stabilization of
the nanoparticles and molecules [32-36]. The freestanding Si-ncs are soluble in almost any
liquid resulting to surface tension variation [37]. This allows an encapsulation of Si-ncs inside
CNTs cavity [38]. Contrary to the high surface tension of silicon that makes filling of CNT
cavity impossible [39], a dispersion of freestanding Si-ncs in organic solution can overcome
some related problems [38, 40]. Other technique that effectively varies Si-ncs surface tension
is laser processing in liquid [41, 42]. Direct laser processing of Si-ncs in liquid media (e.g.
transparent commercially available ethylpolysillicate polymer, water) [37, 42] decreases
surface tension and permits an introduction of Si-ncs within CNT cavity. In-situ CNT cavity
Si-ncs stabilization throughout laser processing prevents pollution, agglomeration of Si-ncs
andmight leads to original class 1D nano-composite fabrication at low cost [43].
In this chapter investigations the kinetics of CNTs growth in TEOM micro-reactor by
catalyst assisted CVD are described. We focus on influence of the reaction temperatures, gas
content and structure of catalyst support on the growing CNTs processes. The synthesis of
CNTs is studied on iron and on nickel catalyst. Catalyst is sited either on porous alumina or
flat Si-ncs support. The CNTs synthesis is assured by decomposition of ethane. The obtained
results clearly show that TEOM has unique capability precisely determinates reaction kinetic
order. Furthermore, connecting of single Si-nc in TEOM with conducting CNTs is regarded
to establish electrical contact and improved the localization of single Si-nc. After all, the
functionalization by assemblies of luminescent Si-ncs in carbon nanotube cavity is
overviewed. We briefly provide some highlight of the assemblies freestanding Si-ncs within
CNTs cavities.
EXPERIMENTAL
Experimental Set Up
The investigations of catalyst assisted CVD kinetics growth of CNTs are performed in a
commercially available TEOM from Rupprecht and Patashnick Co., Inc. Figure 1 (a) shows
the scheme of experimental set up used in this study. TEOM design permits a defined control
of the variety CNTs growth parameters. The TEOM is provided with two automatic heating
zones and gases flow control. The pre-heating zone 1 (Fig. 1(a)) controls the temperature of
the gas stream in the upper part of the sensor. The heat zone 2 controls temperature of the
tapered element and the micro-reactor where the CNTs synthesis takes place. An automatic
run of purge helium gas flows through the tapered element micro-reactor is managed. Catalyst
support in the microreactor between quartz cotton is positioned. In all presented experiments
the synthesis of CNTs is achieved with a gas mixture of ethane and hydrogen (C2H6:H2). The
vibration frequency of the tapered micro-reactor is monitored by an optical way. A transmitter
and a receiver are located on the opposite sides of the micro-reactor.
Figure 1. (a) Sketch of experimental set up of the Tapered Element Oscillating Microbalance (TEOM)
used for kinetics of CNTs growth studies. (b) One typical plot of the temperature variation in the micro-
reactor during the growth of CNTs by catalyst assisted CVD process.
The system resolute the mass uptake m = m(t) – m0 of the microreactor during the
synthesis times t and t = 0 using the following equation:
(1)
where f0 and f1 are the natural oscillating frequency at times t0 and t1, respectively, and K is a
constant that depends on the geometry of the experiment. At the beginning of the each novel
experiment a new frequency f0 is automatically settled after the catalyst with wool is placed
into the micro-reactor. Then the change in the frequency between f1 and f0, permits to
calculate the absolute values of the mass uptakes (Eq. 1.).
Figure 1 (b) displays typical variation of the temperature in the micro-reactor during the
synthesis and monitoring of CVD synthesis process. CNTs growth by introduction of the
catalyst on support and decomposition of ethane is assured. The temperature increases about
40 min under hydrogen flow from room temperature to 400 °C (673 K), and then this
temperature is maintained one hour to reduce the oxide form the catalyst precursor. The
temperature is then continuously increased up to reaction temperature. Figure 1 (b) represents
typical process of the CNTs growth at reaction temperature 750 °C (1023 K) that takes 5 min.
At this temperature, a mixture of ethane and hydrogen replaces the hydrogen flow. Helium is
used as a vector gas with a flow rate similar to the total flow rate of C2H6 and H2 (60 sccm).
In this chapter the similar process is applied. Whenever just one parameter for each
experiment i.e. reaction temperature, ratio of ethane and hydrogen, catalyst and catalyst
support is varied. When the influence of reaction temperature is investigated the temperature
varied from 873 to 1113 K and mixture of ethane and hydrogen (C2H6:H2 at 1:2 ratio) is kept
constant at 60 sccm for all series. For following experiments (e. g. variation of ethane and
hydrogen ratio, influence of the catalyst nature (Fe, Ni) and catalyst support (porous alumina,
Si-ncs)} the reaction temperature was 1023 K. Since the volume of the CNTs increases
markedly in the course of the synthesis, the growth time is consequently limited. Therefore
the synthesis for all cases not exceeds 5 min. After the synthesis the samples are cooled in
hydrogen atmosphere to room temperature for several hours.
Catalyst Supports
The CNTs growth by introduction of the catalyst on porous alumina and Si-ncs, supports
is promoted. Porous alumina support is a high surface area -Al2O3 (CK 300B Ketjen with a
surface area of 220 m2. g-1), which is mainly made up of a mesoporous network. Iron
deposited on alumina (Fe/Al2O3) is used as catalyst to insure the CNTs synthesis. The
alumina support is crushed and sieved. A fraction of 40 - 80 m is retained for catalyst
preparation. The catalyst is prepared by using an aqueous solution of (Fe(NO3)3 9 H2O), with
a Fe concentration fixed at 20 wt. % (20% Fe/Al2O3 catalyst). The wet solid is dried at 100 °C
and further calcined in air at 350 °C for 2 h in order to obtain an oxide form of the catalyst
precursor. Then 10 mg of the impregnated alumina support is placed in the micro-reactor of
TEOM between quartz wool (Figure 1 (a)).
For the purpose of the Si-ncs connection by CNTs the catalytic particles are deposited on
the freestanding Si-ncs prepared by electrochemical etching. The preparation of freestanding
and surfactant free Si-nc is described elsewhere [30]. Briefly, the Cz silicon, p-type boron
doped, <111>, 1 Ohm.cm is used as starting material to fabricate room temperature
photoluminescent Si-ncs. We compare CNTs growing process on Si-ncs coated either by iron
(Fe/Si-nc) or nickel (Ni/Si-nc). Those catalysts are prepared by following way. The Si-nc are
introduced into an aqueous solution of Fe (Fe(NO3)3 9 H2O) or nickel nitrade hexahydrate
(N2NiO6 6 H2O with the Fe (Ni) concentration fixed at 20 wt. %. This solution is then kept in
an ultrasonic bath for 30 min. The operation is repeated five times in order to ensure the
deposition of iron particles on the Si-nc surface. Then the solution is dried at 100 °C and then
calcined in air at 350 °C for 2 h. Similar to porous alumina, the catalyst reduction is
guaranteed in-situ in TEOM micro-reactor, before the CVD synthesis takes place, under a
hydrogen atmosphere at 450 °C.
CNTs Cavity Filling with Silicon Annotates
As reported elsewhere, basically, two means for opening of the CNTs ends are used. The
first one is thermal annealing [44–46] and the second one is chemical treatments [45-48]. In
order to open ends by thermal approach, the CNTs are thermally annealed in oxygen or air
atmosphere. The reactivity of graphite at the ends of the tube is higher than on the walls, due
to the stress-induced by the curvature [49]. In our case the opening temperature of CNTs has
been preliminary determined by thermogravimetric (TGA) analysis. The CNTs are annealed
at a rate of 5°C/min up to the temperature of the inflexion point deduced from TGA analyses
(T = 580°C). After use of this approach more than 80 % of observed carbon nanotubes were
opened [49]. Opened nanotubes are filled with together Si-ncs dispersed (0.01 wt.%) in
commercially available silicate based polymer [30].
For the CNTs cavity filling experiment by laser fragmentation same opened nanotubes
were used. The Si micrograins prepared by electrochemical etching serve as the source for Si-
ncs fabrication by pulsed laser fragmentation in transparent polymer [42]. In 30 ml of
polymer 0.01 mg of CNTs is homogenously dispersed. Then added 0.01 wt% of micrograins
and a 5 ml of such solution (micrograins/CNTs/polymer) is irradiated by a pulsed laser
(Nd:YAG, 355 nm, 30 Hz, 8 ns) at fluence 6 mJ/pulse for 2 hours at room temperature. The
laser beam on the liquid surface is focused and the glass container is rotated during laser
processing.
KINETCIS OF THE CARBON NANOTUBES GROWTH

Kinetics on Porous-Alumina Catalyst Support
Synthesis parameters, such as reaction temperature and partial pressure, heavily affect the
CNTs kinetics of growth. Therefore, in order to evaluate the reaction kinetics those
parameters are investigated and discussed in details in this section. In our TEOM
experimental configuration and used catalyst (iron on alumina support) the CNTs formation
starts at reaction temperature around ~ 963 K. In figure 2 (a-c) typical mass-uptakes in TEOM
micro-rector for three different temperatures are shown. It has to be noted that below < 933 K
an amorphous carbon is mainly formed. The mass uptake is associated to the initial mass m0
of catalyst (10 mg) at t = 0 min. It is observed that the mass uptake strongly increases with
temperature from 933 K up to 1083 K and then abruptly drops at 1113 K [50]. This is the
region of the temperatures where a significant weight changes in micro-reactor occur induced
by the growth of CNTs. Figure 2(b) shows mass uptake and growth rate when the formation
of CNTs takes place at temperature 1023 K. Two region indicated by red lines corresponding
two growth rates v1 and v2.
What is the origin of the difference in growth rates? In general it is accepted that the
growing rate vi (i = 1, 2) is defined as the initial rate of mass increase (t = 0) and the
subsequent linear mass increase described as follow [6]
Ea
v A exp , (2)
R0T
where R0 is the perfect gas constant (R0 = 8.31 J/K) and T (K) is reaction temperature. The
initial growth rate v1, is associated with the initial carbon uptake. This mass increase displays
the largest variations of the activation energy. At low reaction temperatures from 873 K to
993 K low activation energy of 12 kJ/mole is evaluated. Within 993K and 1023K fast
increase in activation energy is observed (146 kJ/mole). Above 1023K up to 1083K, however,
the grow rate is constant or slightly decreasing (– 12 kJ/mole). Finally, over the temperature >
1083 K the initial rate drops abruptly. Detailed kinetics analysis show that v1 can be assigned
to the formation of a dynamic equilibrium between carbon adsorption-bulk diffusion-
segregation on the iron catalytic nanoparticle, whatever the nature of this carbon is made up,
and the growth rate v2 to the specific formation of the CNTs. In addition, also Arhenius plot
show two growth rates; (i) v1 initial due to the adsorption /disorption C2H6:H2 on the catalyst,
and (ii) v2 rate attributed to the growing of the CNTs. Activation energies suggest the reaction
order toward a common kinetics [49].
Right panel in figure 2 shows corresponding SEM images of the samples after synthesis
at reaction temperatures 873, 1023 and 1113 K, respectively. With increased temperature the
CNTs are prolonged and diameter of the CNTs varies. Up to 993 K the outer diameter
increases and reaches of about 47 nm in average, then decreases to 20 nm. However to
evaluate the longer of CNTs is quite difficult because of forming bunches specially for higher
temperatures (> 850 K). The structural characterization by transmission electron microscopy
(TEM) clearly displays graphitic multiwalls around the tubule.
Figure 2. Left panel (a-c) shows time dependence of the weight mass uptake in the micro-reactor of
TEOM for CNTs grown 5 min at three different temperatures. Synthesis is ensured by decomposition of
a gas mixture of ethane and hydrogen C2H6:H2 at molar ratio 1:2 and 60 sccm. The growth rates v1 and
v2 are indicated for temperature 1023 K. Right panel shows corresponding SEM images.
In addition, the Raman spectra of the carbon deposit exhibit the two narrow bands D and
G at 1340 and 1570 cm-1 with a clear tailing around 1610 cm-1 [22, 49] These contributions
are quite characteristic of the presence of CNTs with some extent of defects as also evidenced
by the TEM image. Similar spectra are recorded on each sample when considerable uptake of
mass in TEOM was observed.
When we investigated an influence of the partial pressure on growing process we kept
constant the total flow D DC 2 H 6 DH 2 60 sccm and reaction temperature at 1023 K.
In this case the variable parameter of the synthesis is the ratio RM = (DC2H6 / DH2) = (PC2H6 /
PH2.). It is observed that besides of the reaction temperature also an increased content of
ethane substantially influence the initial growth rate. For instance, by decreasing of dilution
ratio of gas mixture from a 1:5 C2H6:H2 to a 1:1 C2H6:H2 the growth rate increased from
0.050 mg/mg(cat).min to 0.115 mg/mg(cat).min, respectively. With the ethane concentration
increases in the gas flow, sooner saturation of the mass uptake occurs [22]. In our particular
case, with a (1:5) C2H6:H2 gas mixture, saturation is not observed whereas with a 1:1 mixture
saturation starts after three minutes of the synthesis. This is in agreement with a promoted
coking process at higher hydrocarbon pressure, which is widely reported elsewhere [22, 50,
52].
In figure 3 the initial growth rate is plotted as a function of gas ratio. The linear behaviour
in a semi-logarithmic scale allows determining the power law of rate CNTs growth v2 versus
gaseous carbon sources mixture that can be expressed as follow
Ea
v kR x exp (3)
R0T
where k is a kinetic constant and Ea is the activation energy of the mass increase process.
From the linear extrapolation of the plot in figure 3
ln( v) x.ln( RM ) C (4)
an exponent x = 0.50 ± 0.03 can be evaluated. It is shown that the growth rate increases
linearly with the square root of the ethane-to-hydrogen ratio of partial pressures. Thus a first
order kinetics with regard to carbon concentration can be deduced from these results, whereas
hydrogen competes with hydrocarbon for the adsorption on the catalytic sites of the catalyst.
These results are coherent with a simple model where the dissociative adsorption of the
hydrocarbon is the rate-determining step of the overall process within the assumed limitations
that the carbon partial pressure is low compare to the hydrogen partial pressure. Thus the
corrected expression is
Ea
v k ( P C 2 H 6 / PH 2 ) 0.5 exp . (5)
R0T
It means that the kinetics of carbon mass increase is first order with regard to carbon
partial pressure and is prevented to the first order by hydrogen partial pressure. The fact that
hydrogen is competing with hydrocarbon source strongly supports a rate-limiting step due to
the adsorption on iron active catalytic sites. The carbon molar rate yield RM (%) calculated
according to expression (3) and (5) can be written as follows
RM = [R0 . T . k / 2 . D Mc . (PC2H6 / PH2)0.5] . exp(-EA / R0 .T), (6)
with D = 20 10-6 m3/min; Mc = 12 g/mole. The growing rates linear increases of allows
estimate the net molar rate of carbon deposition. In our case hydrocarbon flow fully contacts
the catalyst and therefore the initial mass increase is due to the pure carbon. We observe that
molar ratio increases linearly and for growth rates being rather similar slope for all series of
samples. This means that carbon is much easier to deposit on catalyst at higher temperature.
Carbon yields fall between 20% and 25% that means that one fourth to one fifth of the input
carbon is decomposed and used for the CNT growth. The beneficial effect of hydrogen is
explained by a probable hydrogenation of the coke deposited at the iron catalyst, leaving
some surface sites free for the reaction. These results together with reaction temperature
dependence growth corroborate and confirm that we deal with the reaction order toward a
common kinetics.
From structural viewpoint, as the concentration of ethane increases the quantity and
structural properties of synthesized CNTs altered as well. Ethane concentration results to the
synthesis with larger amount of CNTs and smaller diameter. For example, when the
ethane:hydrogen ratio is increased from 1:3 to 1:1, the average outer diameter of the CNTs
decreases from about 45 nm to 21 nm. It should be emphasized that this behaviour can
qualitatively be correlated with an increasing density of CNTs. New catalytic sites for CNTs
nucleation and growth are active with higher carbon content. As the density of tubes
significantly increase then less carbon is available for each catalytic particle and thinner
CNTs are grown.
Figure 3. Initial rate CNTs growth (G) as a function of the carbon molar rate yield (R M).
Flat Catalyst Support and Single Silicon Nanocrystal Connection
Subsequently direct functionalization of single Si-ncs by connecting with conducting

multiwalled CNTs is discussed. Figure 4 represents the average mass uptake in the micro-
reactor of TEOM at reaction temperature 1023 K. Iron and nickel catalysts sited onto Si-ncs
surface promote the CNTs synthesis. Open circles represent the iron (Fe/Si-nc) and full
squares the nickel (Ni/Si-nc) catalyst. The catalyst amount is the same (20 wt.%) for both
cases. The total mass uptakes increase linearly without saturation. Contrary to the porous
alumina, flat nanocrystal surface reduces inter porous diffusion and inhibit the mass uptake
saturation [22]. Then catalyst is assumed to be quite accessible to the ethane, whereas on high
surface area catalytic nanoparticles supported on alumina the accessibility perturbed. In
addition, on the flat surfaces, the growth rate is linear over the whole time range as it is
neither disturb by long-term poisoning of the catalyst, nor by the rapid initial transient steps
due to carbon saturation of the metal particle and nanotube nucleation. This infers that non-
steady state kinetics occur when encompassing a large time period of mass uptake over high
surface area catalysts. This non-stationary state has been explained by a selective coking of
the catalytic particles either located outside or inside the pores of the support [22]. Over a
long period of mass increase, the measured kinetics tends rapidly to saturation and the
determination of the kinetic order may be strongly changed. Indeed it is tempting to consider
that the initial rate at t = 0 would be the true rate corresponding to carbon incorporation.
However it was shown independently that the initial carbon uptake also involves the
irreversible carbon bulk saturation of the catalyst particle [32].
Figure 4. Time dependence of average mass increase in the micro-reactor of TEOM at temperature
1023 K. Two catalysts are supported into silicon nanocrystals surface. Red circles represent 20 wt.% of
iron and black squares represent 20 wt.% of nickel catalyst.
For both catalysts, after initial carbon absorption and nucleation at the interface between
the Si-ncs and the carbon nanotube the growth take place. It is observed that the growing on
the iron catalyst is more efficient compared to nickel. In iron case the growing rate reaching
the value of 1.21 + 0.02 mg/min with RM = 21.43 %. This value is very similar as iron
catalyst deposited on porous support. However, the growing rate on nickel is lower and
reaches 0.2096 + 0.046 mg/min with corresponding carbon yield RM = 3.71 %. This rather
low rate is assumed be assigned to low portion of the input carbon decomposition. A fast
carbon covering of the Ni particles cannot explain the deactivation of the nickel. If so then a
rapid initial mass increase followed by saturation would be observed. The low growth rate
can be explained by etching of the Si-ncs by hydrogen and formation of nickel silicide [22]
resulting lower mass uptake.
Figure 5 shows corresponding SEM images of CNTs obtained by CVD in TEOM when
nickel and iron catalyst is sited onto Si-ncs surface. From the SEM micrographs, it appears
that CNTs are grown with a random growth direction with lengths that exceed several
micrometers. It is observed that the diameters of CNTs grown on nickel catalyst are smaller
compared to iron. In the case of iron catalyst the average diameter is around 58 nm while for
nickel reaching of 29 nm.
Figure 5. Corresponding scanning electron microscopy (SEM) images of CNTs obtained by CVD in
TEOM when the catalyst is introduced into silicon nanocrystals; a) nickel b) iron.
As mentioned earlier this might be related the etching process of Si-ncs surface by
hydrogen that promotes nickel silicide formation leading to lower mass uptake (Fig. 4,
squares). Similar to the porous alumina support, no significant difference is observed in the
distribution in lengths and the purity. The length of the carbon nanotubes is several
micrometers and is rather similar for both catalysts and supports. In all cases the purity is
estimated to be larger than > 90%.
Detailed structural analysis reveals that the Si-nc stays stuck on tip of the CNT end. This
consequences direct nanotube connection to the Si-nc surface with a strong adhesion force. In
Figures 6 (a) a TEM image of a single Si-nc on top of a CNT is shown. In this case the iron
catalyst is coated on Si-ncs surface. It exhibits a size of about 50 nm fully capping the as-
grown carbon nanotube with a slightly smaller diameter. Apparently, the adhesion force is
strong enough to avoid the loss of the Si-nc from the tube tip. The CNTs direct growth on Si-
nc and a disappearance of the catalyst from the Si-nc surface is observed [24].
Figure 6. a) Typical TEM image of a single silicon nanocrystal (size around 50 nm) connected to a
carbon nanotube (CNT). b) Room temperature photoluminescence (PL) spectra of Si-ncs connected
with CNTs (full triangle) and CNTs (open circle) synthesized at the same conditions on porous alumina
support.
The carbon segregation from the catalyst phase localized on the surface of the Si-nc
allows the wetting of the rear face of the Si-nc and thus induces the formation of the CNT
with a diameter similar to that of the support i. e. Si-nc. It has to be emphasized that we check
that CNTs cannot be grown on Si-nc alone. This stresses that the catalyst plays a crucial role
in the growth of carbon nanotubes and connection of Si-ncs. The catalyst is a necessary
condition to allow the decomposition of the hydrocarbon ethane based precursor. From the
walls of the starting nanotubes, the interplanar distance characteristic of graphite can be
estimated to 0.338 nm. Detailed HR-TEM analysis revealed parallel stripes of Si-nc with a
diamond lattice same as the initial orientation of the starting material [24].
Next question that one can ask is whether the Si-ncs keep the luminescence properties
after CNTs synthesis and connection. We have investigated room temperature PL properties
of such Si-ncs wired with CNTs. The connected Si-ncs to CNTs showed visible room
temperature PL under YAG blue light illumination at wavelength 355 nm. Triangles in Figure
6 (b) represents PL spectrum of connected Si-ncs by CNTs where the iron catalyst promoted
the synthesis. The CNTs (open circles) grown on alumina support at same conditions showed
no visible PL in this spectral region at room temperature. In fact the PL peak maximum is
located at the same wavelength as for freestanding Si-ncs only [24, 30]. It has to be noticed
that the intensity is weaker more than 50 times compared to the intensity before CVD process.
Several causes are responsible for this decrease. When the Si-ncs are exposed to high
temperature treatment (> 600 °C) the changes of the surface states and the defect diffusion
induces a quenching of the PL intensity.
FILLING CARBON NANOTUBE CAVITY BY SILICON NANOCRYSTALS

As it is well known, the high surface tension of molten silicon disallow direct filling of
CNT cavity [39]. An immersion of CNTs into organic solutions containing of Si-ncs enables
to obtaining surface threshold values for nanotube wetting and introducing luminescent Si-ncs
into CNTs cavity [38, 40]. Here, we discuss the results of filling CNTs by two diverse
approaches based on different physical phenomena that promote the filling. Firstly, by
capillary forces induced by CNTs cavity, and secondly, by shock waves generated during
pulsed nanosecond laser irradiation of colloidal solution. In our explorations we have
investigated Si-ncs surface tension variation and cavity filling in environmental (e.g. water)
and technological (spin on glass) friendly solutions.
A figure 7 (a) shows the TEM image of CNTs cavity filling promoted by capillarity force
after immersion of CNTs in the Si-ncs/polymer solution. The solution is silicate-based
polymer containing homogenously dispersed Si-ncs. The image shows of a CNT with a
diameter of 50 nm, where a Si-ncs are observed in the core of the nanotube. The circles
indicate spots a size of around 3 nm and might correspond to a Si-nc. It has to stress that such
Si-ncs are found quite far, around 500 nm, from any end of the CNT [38]. This is in
agreement with the findings that most of the CNTs used here are not bamboo-like, without
internal carbon membranes that would stop the Si-nc incorporation [32, 40]. The high amount
of embedding amorphous polymer surrounding the Si-ncs does not allow resolving the Si
interplanar distances. In order to confirm Si-ncs presence, a selective diffraction analysis is
conducted on this area. A diffraction pattern taken in the tube is displayed in figure 7(b). Due
to the random Si-ncs orientation the rings were recorded and could be assigned to the silicon
lattice planes with diamond like structure.
Figure 7. (a) A plan-view HRTEM image of an open CNT and filled with a Si-nc/polymer solution. The
Si-ncs were prepared by electrochemical etching and homogeneously dispersed in organic based
polymer solution. (b) Corresponding diffraction pattern taken in the tube confirms the presence of
silicon nanocrystals with diamond like structure.
Recent development of fabrication of Si-ncs in liquid media by pulsed laser processing

allows preparation of Si-ncs with surface tension different to bulk silicon and unusual wetting
phenomena [41]. We have focused on introducing of freshly prepared Si-ncs by laser
processing in liquid transparent polymer. At the same times, as the Si-ncs are prepared the
shock waves are generated through the solution (polymer/CNTs/micrograins) [42, 43]. We
exploit shock waves as a principal force to fill CNTs cavities by Si-ncs freshly prepared in
polymer solution. To suppress the effect of the capillary forces the CNTs were prior to
introduce into solution. Detailed HR-TEM analysis is performed to confirm the presence of
Si-ncs within the CNTs cavity after laser processing. Figure 8 displays image of filled CNT
with inner diameter of 50 nm when the solution is irradiated at laser fluence of 6 mJ/pulse. It
is observed that nanotubes cavities are fully filled with Si-ncs/polymer composite.
Corresponding electron diffraction pattern (inset), reveals silicon diffraction rings that could
be assigned to the lattice planes <111>, <220>, <311> with cubic phase.
It is assumed that the filling occurred by following way. The shock waves propagating
through the solution generate reflux, which is the principal driving force to introduce the
freshly formed Si-ncs into CNT cavity. For generation of shock waves are responsible two
major effects. Firstly, the strong electric field generated by the laser causes the electron
avalanche near the silicon micrograin, and secondly, the breakdown of the polymer in liquid
phase. In our experiments, a bright spot near the surface level is observed and one could hear
explosion sounds due to the optical breakdown of the polymer. Either the collapse of
vaporized cavitations bubbles or the grains that can be accidentally diffused under the beam
cause these explosions. Even more, the shock waves are accelerated at large distances with
the explosion [43]. As a sign of a micrograins fragmentation and nanocrystals formation is
change of the solution color [42].
Figure 8. HR-TEM image of Si-ncs fragmented in silcate polymer and filled in CNTs cavity at laser
fluence of 6 mJ/pulse. Inset shows selected area electron diffraction pattern in the tube.
Also in our case the solution (polymer/CNTs/micrograins) when is irradiated with

enough high intensities (>4 mJ/pulse), loses its characteristic yellowish color and becomes
transparent. As the fragmentation occurs freshly-produced Si-ncs embedded in hydrophilic
polymer solution are introduced in the CNT cavity. The question arises what happens with the
CNTs during the pulsed laser irradiation. The CNTs itself at this dispersed concentrations and
used irradiation intensities have weak absorption at the wavelength at 355 nm [43]. Therefore,
mostly the micrograins fragmentation occur. However, at higher fluencies also broken CNTs
are found.
Figure 9. Normalized photoluminescence intensity as a function of wavelength of Si-nc/polymer/CNTs

composites. Full red squares represent CNTs filled with Si-ncs/polymer by capillary force. Open blue
circles correspond to filled CNTs by Si-ncs fabricated by laser fragmentation of the micrograins in
polymer solution at laser fluence 6 mJ/pulse.
It has to be stressed that both filling processes are achieved at room temperature (300 K).
This has several advantageous: (i) the inhibition of the emptying of the tubes [52], (ii) the
absence of defects formation in Si-ncs, and (iii) the limitation of the degradation of Si-ncs
based nanocomposite luminescence properties. Figure 9 compares the PL spectra of filled
CNTs by capillary force and shock waves generated during the laser processing. We found
that encapsulated Si-ncs keep their PL properties. Open circles represent filled CNTs with Si-
ncs fabricated by nanosecond pulsed laser fragmentation directly in the polymer solution. The
laser processing at laser irradiation at 6 mJ/pulse is performed. The composite shows rather
same characteristic as for Si-ncs formation in polymer only [42], leading to fabrication of
blue room temperature photoluminescent nanocomposite. The Si-ncs emit light with PL
maxima located at 450 nm. The filling CNTs by immersion leads to formation of
nanocomposites with typical red PL with maximum located at 680 nm. Laser fragmentation
process prepares nanocomposites that show significantly blue shift of the PL maximum (more
than 300 nm). Full squares represent the CNTs filled with Si-ncs/polyner by capillary force.
An encapsulated Si-ncs do not disturb PL properties and the intensity of PL is proportional to
the inner diameter of CNTs. However, neither a shift of the light nor a narrowing of the
emission band is observed [40]. This is explained because the inner diameter is generally
larger than the size of Si-ncs. In principle decreasing inner diameter of nanotubes would play
the role of a filter for promoting the insertion of smaller Si-nc. Then one can expect a
narrowing full width at half maximum of the PL signal. Moreover, after such a confinement
in CNTs, the Si-ncs with the lower mean sizes would emit light with a shift of the maximum
frequency due to quantum confinement size effect [31]. Both the PL intensity and diffraction
analysis indicate that the lower concentration of encapsulated Si-ncs in CNT cavity compared
to simple immersion is achieved by laser processing. For further improvement of the
processes to obtain the cavities with higher concentrations, it is necessary to find the
optimized conditions to get an efficient yield of filling Si-ncs/polymer. For instance, this
might be accomplished by laser elaboration of Si-ncs surface in different liquids that offers
fabrication of smaller and non-aggregated Si-ncs [43]. Those avoids capping of CNTs ends by
larger particles aggregates and set aside the higher concentration of Si-ncs inside of CNT
cavities. An assembly of luminescent Si-ncs inside CNTs cavity might lead to 1D
nanocomposite where the single Si-ncs are either randomly placed or display a chain-like
character which can be controlled by the diameter of the CNTs. In the case of blue
luminescent Si-ncs a superior operation and a possibility of higher integration could be
expected.
CONCLUSION
In this chapter, we discussed growth and functionalization of carbon nanotubes (CNTs)
prepared by catalyst assisted chemical vapor deposition (CVD) most commonly used at
industrial scale. An employment of Tapered Element Oscillating Microbalance (TEOM) for
CNTs reaction kinetics is explored. Hydrodynamic characteristics of the CVD process in
TEOM reactor, like a homogeneous and high degree of contact with catalyst allow accurate
monitoring of the growing process. The unique capacity of TEOM provides new physical
insight in CNTs reaction kinetics. By recording of small weight uptakes at initial stages of the
CNTs growth at different reaction temperatures, partial pressures on different catalyst

supports the reaction kinetics has been determined. We demonstrated that it is beneficial to
work in TEOM apparatus to describe more accurately the fundamental kinetics of the CNT
growth. Then morphology of the CNTs at the large-scale synthesis can be controlled via well-
established parameters determined from TEOM. The harvested knowledge might forcedly
lead to considerable improvement of CNTs production at industrial scale.
Moreover, better understanding of the kinetic processes on flat silicon nanocrystals (Si-
ncs) catalyst supports has been explored as well. Contrary to the porous alumina flat Si-ncs
surface reduce inter porous diffusion and do not allows to mass uptake saturation. It has been
showed that the catalyst coated on Si-ncs surface promote direct connection with CNTs. Fully
capped the as-grown carbon nanotube on Si-ncs are found with the adhesion force strong
enough to avoid the loss of the Si-nc from the tube tip. What's more ethylpolysillicate based
polymer naturally wets Si-nc surface and simply allows encapsulation of luminescent Si-ncs
in the CNT cavity by induced capillary force. In addition, a CNTs cavity filling by the blue
luminescent Si-ncs by nanosecond laser fragmentation was achieved. The shock waves
generated during the laser processing induce an entering of the fresh formed Si-ncs into the
CNTs cavities. Development of the concept of functionalization Si-ncs with CNTs by
connecting or by filling could opens up a wide range of new situations and potential
applications.
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Lecture Material 6
CARBON NANOTUBE/POLYMER COMPOSITES:

INTERFACIAL BONDING CHARACTERISTICS
ABSTRACT
Since the discovery of carbon nanotubes (CNTs) by Iijima in 1991, CNTs have
attracted great research interest due to their unique properties such as high electrical and
thermal conductivity, excellent stiffness against bending, and high tensile strength. Using
carbon nanotubes (CNTs) as nanofibers to enhance the mechanical, electrical, thermal,
and optical properties of composite materials has been pursued extensively. Molecular
mechanics (MM) and molecular dynamics (MD) simulations have become increasingly
popular in the theoretical investigations of reinforcement mechanisms in CNT-polymer
composite systems. This paper is dedicated to conduct theoretical study on the interfacial
characteristics of CNT reinforced polymer composites.
Firstly, force-field-based MD simulations are performed to study the interaction
between polymers and SWNTs. The ―w rapping‖ of nanotubes by polymer chains was
computed. The influence of temperature, nanotube radius and chirality on polymer
adhesion was investigated. Furthermore, the ―f illing‖ of nanotubes by polymer chains
was examined. The results show that the interaction between the SWNT and the polymer
is strongly influenced by the specific monomer structure such as aromatic rings, which
affect polymers‘ affinities for SWNTs significantly. The attractive interaction between
the simulated polymers and the SWNTs monotonically increases when the SWNT radius
is increased. The temperature influence is neglectable for PE and PP but strong for PS
and PANI.
Secondly, our simulations indicate that the adhesion energy between the SWNT and
the polymer strongly depends on the chirality. For SWNTs with similar molecular
weights, diameters and lengths, the armchair nanotube may be the best nanotube type for
reinforcement. The simulations of filling reveal that molecules of PE, PP and PS can fill
into a (10, 10) SWNT cavity due to the attractive van der Waals interactions. The
possible extension of polymers into SWNT cavities can be used to structurally bridge the
SWNTs and polymers to significantly improve the load transfer between them when
SWNTs are used to produce nanocomposites.
Finally, the influence of chemical functionalization on the interfacial bonding
characteristics of SWNTs reinforced polymer composites was investigated using MM and
MD simulations. The simulations show that functionalization of nanotubes at low
densities of functionalized carbon atoms drastically increase their interfacial bonding and
shear stress between the nanotubes and the polymer matrix. This indicates that increasing
the load transfer between SWNTs and a polymer matrix in a composite via chemisorption
may be an effective way and chemical attachment of nanotubes during processing may be
in part responsible for the enhanced stress transfer observed in some systems of the
nanotube-polymer composites. Furthermore, this suggests the possibility to use
functionalized nanotubes to effectively reinforce other kinds of polymer-based materials

as well.
The simulation results would be of important in the production of CNTs reinforced
polymer composites.
Keywords: Carbon nanotube polymer composites, Molecular mechanics, Molecular

dynamics, Interfacial bonding, Molecular interaction, Chemical functionalization.
INTRODUCTION AND BACKGROUND

Since the discovery of carbon nanotubes (CNTs) by Iijima in 1991 [1], CNTs have
attracted great research interest due to their unique properties such as high electrical and
thermal conductivity, excellent stiffness against bending, and high tensile strength [2]. Using
CNTs as nanofibers to enhance the mechanical [3-10], electricitrical [11-14], thermal [15-17],
and optical [18] properties of composite materials has been pursued extensively both in
experimental and theoretical studies.
Recently, experiments have shown remarkable enhancements in elastic modulus and
strength of polymer composites with an addition of small amounts of CNTs [19-22]. It is well
established, from the research on microfiber-reinforced composites over the past few decades,
that the structure and properties of the fiber-matrix interface play a major role in determining
mechanical performance and structural integrity of composite materials. However, due to
difficulties in devising experiments to study the CNT-polymer interface, molecular mechanics
(MM) and molecular dynamics (MD) simulations have become increasingly popular in the
investigations of reinforcement mechanisms in CNT-polymer composite systems [23]. Many
groups have investigated the interfaces in CNT-reinforced polymer composites using MD
simulations.
For example, Liao et al. [24] have studied the interfacial characteristics of a CNT-
reinforced Polystyrene (PS) composite system through MM simulations and elasticity
calculations. They found that the fiber/matrix adhesion comes from electrostatic, van der
Waals interaction, mismatch in the coefficients of thermal expansion and radial deformation
induced by atomic interactions. Frankland et al. [25] have investigated the influence of
chemical cross-links between a Single-walled nanotube (SWNT) and a polymer matrix on the
matrix-nanotube shear strength using MD simulations. The results suggest that load transfer
and modulus of nanotube-polymer composites can be effectively increased by deliberately
adding chemical cross-linking and inadvertent chemical bonding between nanotubes and
polymer matrices during processing may be in part responsible for the enhanced stress
transfer observed in some systems of this type. Wong et al. [26] have studied local fracture
morphologies of CNT/PS rod and CNT/epoxy film composites. Transmission and scanning
electron microscopy examinations showed that these polymers adhered well to CNT at the
nanometer scale. Some of the important interfacial characteristics that critically control the
performance of a composite material were quantified through MM simulations and elasticity
calculations. Multi-walled CNT morphology-related mechanical interlocking at nanometer
scale, thermal residual stresses, as well as relatively cavity free surface for polymer
adsorption are also believed to be contributing factors.
Gou et al. [27] investigated the interfacial bonding of SWNT reinforced epoxy
composites using a combination of computational and experimental methods. The interfacial
shear strength between the nanotube and the cured epoxy resin was calculated to be up to 75
MPa, indicating that there could be an effective stress transfer from the epoxy resin to the
nanotube. The following experimental results provided evidence of stress transfer in
agreement with the simulation results.
Yang et al. [28] has studied the interaction between polymers and CNTs using force-
field-based MD simulation. They found that the specific monomer structure plays a very
important role in determining the strength of interaction between nanotubes and polymers.
The polymers with a backbone containing aromatic rings are promising candidates for the
noncovalent binding of CNTs into composite structures, which can be used as building blocks
in amphiphilic copolymers to promote increased interfacial binding between the CNT and the
polymeric matrix.
Wei [29] has studied temperature dependent adhesion behavior and reinforcement in
CNT-polymer composite. They found that the interfacial shear stress through van der Waals
interactions increase linearly with applied tensile strains along the nanotube axis direction and
a lower bound value for the shear strength is found 46 MPa at low temperatures. Direct
stress-strain calculations show significant reinforcements in the composite in a wide
temperature range, with 00% increase in the Young‘s modulus when adding 6.5% volume
ratio of short CNTs.
However, most of the cited literatures considered the effect of only one or two factors on
the adhesion properties. In this study, we focus on the physisorption of polymers on SWNTs
and investigated the physical interactions between polymers and SWNTs in all conditions that
we could think about using MM and MD simulations.
EXPERIMENTAL
Computational Method
In this study, MM and MD simulations were conducted to explore the interfacial

characteristics between SWNTs and polymers, through which we could get useful
information for the development of nanotube-based polymeric composites. Here, MM and
MD simulations were carried out using a commercial software package called Materials
Studio developed by Accelrys Inc. The condensed phase optimization molecular potentials for
atomistic simulation studies (COMPASS) module in the Materials Studio software was used
to conduct force-field computations. The COMPASS was a parameterized, tested and
validated first ab initio force-field [30, 31], which enables an accurate prediction of various
gas-phase and condensed-phase properties of most of the common organic and inorganic
materials [32-34].
Force Field
The application of quantum mechanical techniques can accurately simulate a system of

interacting particles, but such techniques often cost too much time and are usually feasible
only in systems containing up to few hundreds of interacting particles. As we know, the main
goal of simulations of the systems containing a large number of particles is generally to
obtain the systems‘ bulk properties which are primarily controlled by the location of atomic
nuclei, so the knowledge of the electronic structure, provided by the quantum mechanic
techniques, is not critical. Thus, we could have a good insight into the behavior of a system if
a reasonable, physically-based approximation of the potential (force-field) can be obtained,
which can be used to generate a set of system configurations which are statistically consistent
with a fully quantum mechanical description. As stated above, a crucial point in the atomistic
simulations of multi-particle systems is the choice of the force-fields, a brief overview of
which is given in this section.
In general, the total potential energy of a molecular system includes the following terms
[35]:
Etotal Evalence Ecross term Enon bond (1)

Evalence Ebond Eangle Etorsion Eoop EUB (2)
Ecross term Ebond Eangle angle Ebond Eend Emiddle bond Eangle torsion
bond angle bond torsion torsion
(3)
Eangle angle torsion
Enon bond EvdW EColumb EH bond (4)
The valence energy generally includes a bond stretching term, Ebond , a two-bond angle
term, Eangle , a dihedral bond-torsion term, Etorsion, an inversion (or an out-of-plane
interaction) term, Eoop , and a Urey–Bradlay term (involves interactions between two atoms
bonded to a common atom), EUB .
The cross-term interacting energy, Ecross term , accounts for the effects such as bond
lengths and angles changes caused by the surrounding atoms and generally includes: stretch–
stretch interactions between two adjacent bonds, Ebond bond , bend–bend interactions between
two valence angles associated with a common vertex atom, Eangle angle , stretch–bend
interactions between a two-bond angle and one of its bonds, Ebond angle , stretch–torsion
interactions between a dihedral angle and one of its end bonds, Eend bond torsion , stretch–
torsion interactions between a dihedral angle and its middle bond, Emiddle bond torsion , bend-
torsion interactions between a dihedral angle and one of its valence angles, Eangle torsion , and
bend–bend–torsion interactions between a dihedral angle and its two valence angles,
Eangle-angle-torsion .
The non-bond interaction term, Enon bond , accounts for the interactions between non-
bonded atoms and includes the van der Waals energy, EvdW , the Coulomb electrostatic
energy, ECoulomb , and the hydrogen bond energy, E H bond .
The COMPASS force-field uses different expressions for various components of the
potential energy as follows [32, 33]:
2 3 4
Ebond [ K 2 b b0 K 3 b b0 K 4 b b0 ] (5)
b
2 3 4
Eangle [H 2 0 H3 0 H4 0 ] (6)
0 0 0
Etorsion [V1[1 cos( 1 )] V2 [1 cos( 2 2 )] V3[1 cos(3 3 )]] (7)
2
Eoop Kx (8)
x
Ebond bond Fbb' b b0 b' b0 ' (9)

b b'
Eangle angle F ' 0 ' 0 ' (10)

'
Ebond angle Fb b b0 0 (11)

b
Eend _ bond torsion Fb b b0 V1 cos V2 cos 2 V3 cos 3 (12)

b
Emiddle_ bond torsion Fb ' b' b0 ' b' b0 ' F1 cos F2 cos 2 F3 cos 3 (13)
b'
Eangle torsion F 0 V1 cos V2 cos 2 V3 cos 3 (14)
Eangle angle torsion K ' cos 0 ' 0 ' (15)

'
qi q j
ECoulomb (16)
i j rij
Aij Bij
EvDW 9
(17)
i j r ij rij6
where q is the atomic charge, is the dielectric constant, and rij is the i-j atomic separation
distance. b and b ' are the lengths of two adjacent bonds, is the two-bond angle, is the
dihedral torsion angle, and is the out of plane angle. b0 , ki (i 2 4) , 0 ,
0
H i (i 2 4) , i (i 1 3) , Vi (i 1 3) , Fbb' , b0 ' , F ' , 0 ' , Fb , Fb , Fb ' ,
Fi (i 1 3) , F , K ' , Aij , and Bij are fitted from quantum mechanics calculations and
are implemented into the Discover module of Materials Studio [36] , a powerful commercial
atomic simulation program used in this paper.
INVESTIGATION OF MOLECULAR INTERACTIONS BETWEEN SWNT

AND POLYETHYLENE/ POLYPROPYLENE/ POLYSTYRENE/
POLYANILINE MOLECULES
Molecular Model
Molecular Model of SWNT

In this study, we have used different kinds of SWNTs with diameters ranging from 7.83
to 27.12 Å and different chirality ranging from zigzag (10, 0) to armchair (10, 10). The
electronic structures of the all carbon atoms in the SWNT models were sp2 hybridization. We
have avoided the unsaturated boundary effect by adding hydrogen atoms at each end of CNT.
Each C–C bond length was 1.42 Å and C–H bond length was 1.14 Å. The hydrogen atoms
had charges of +0.1268 e and the carbon atoms connecting hydrogen atoms had charges of
−0.1268 e, thus the neutrally charged SWNTs were constructed. The computer graphics
picture of a (10, 10) SWNT model (with 400 carbon atoms and 40 hydrogen atoms) is shown
in Figure 1.
Molecular Model of the Investigated Polymers

The simulated polymers were polyethylene (PE), Polypropylene (PP), Polystyrene (PS),
Polyaniline (PANI). The chemical structure and molecular models are provided in Figures 2
and 3. The simulated polymers had comparable numbers of atoms and molecular weight (PE
38 atoms, 170, PP 38 atoms, 170, PS 34 atoms, 204, and PANI 26 atoms, 184), hence the
magnitude of the interaction energy gives a direct measure of the strength of their binding to
the SWNT. Since the number of atoms and monomers used is small (about 35 atoms per
molecule which corresponds to 6 monomers for PE, and 4 monomers for PP, 2 monomers for
PS, 2 monomers for PANI), the molecules are better described as oligomers than polymers.
However, we use the term ―pol ymer‖ throughout the paper for simplicity, with the
understanding that the results of our simulation describe the behavior of a small part (block)
of a ―long‖ polymer. PE is polymeric molecules with no side groups, whereas PP has side
groups of methyl. PS and PANI are both molecules with side groups of aromatic groups.
Figure 1. Molecular model of a (10, 10) SWNT.

Polyethylene Polypropylene Polystyrene Polyaniline
Figure 2. Chemical structure of the investigated polymers.
Polyethylene Polypropylene
Polystyrene Polyaniline
Figure 3. Molecular model of the investigated polymers under minimum energy.
Results and Discussion
Generally speaking, wrapping and filling are two typical phenomena which would take
place when the interactive process of the SWNT-polymer system is simulated [37]. We did a
constant NVT simulation with undefined boundary conditions, which implies that the
simulated volume is actually infinite. Also, it has to be noted that our simulation does not
explore all the states of the ensemble because of the short simulation time (several
nanoseconds). If the simulation time were long enough, then the polymer would move away
eventually and very likely never interact with the CNT again, which is a direct consequence
of the fact that we use infinite volume (no boundary conditions). However, it does not affect
the results within our simulation time because this ―es cape‖ event is extremely rare. We do
the simulation in a vacuum in order to simulate the effect of an ideal bad solvent and the
thermostat in our simulation can be thought of as mimicking the action of the bad solvent.
Here, the MD simulation for each case study was performed long enough to observe
several cycles of thermal vibration. The interval of each MD simulation step was typically 1
femtosecond. All calculations were carried out at the initial temperature of 400 K except a
special condition, using constant number of particles, constant volume and constant
temperature (NVT) ensembles.
Wrapping
Polymer wrapping of CNTs has received attention as a promising way for manipulating
and organizing CNTs into ordered structures. Recently, Chen et al. [38] found that a molecule
containing a planar phenyl group can irreversibly adsorb to the surface of a SWNT and
successfully immobilize proteins, DNA, and smaller biomolecules on the nanotube sidewalls.
Experiments for the wrapping of SWNTs with poly(m-phenyleneviny-co-2,5-dioctyloxy-p-
phenylenevinylene) have also been reported [39]. To simulate the interactions between
polymers and SWNTs, MD simulations were established with the polymers initially placed at
the side of the SWNTs within a distance of 9.5 Å, which is the cutoff distance of van der
Waals interactions in this study.
Interaction between Polymers and SWNTs

Firstly, an armchair (10, 10) CNT (Figure 1) and four polymer models are selected as the
representative elements for the simulation. Figures 4-7 show the snapshots of polymers and
SWNTs observed at different time steps of the simulation. Initially, the polymer chains were
put near the middle of SWNTs in a distance of about 8.5 Å. The simulation showed that all
the four molecular chains would stretch and move toward the nanotubes until they finally
wrapped on the surface of the helix of the nanotube and the equilibrium was achieved.
Particularly, it cost about 70 ps for the wrapping of PE, PP and PS, but only 5 ps for PANI,
which may be because of the strong polarity of PANI. We also noticed that the two aromatic
rings of PS and PANI molecules gradually orientated to align their ring planes parallel to the
SWNT surface during the dynamic interactions.
The dynamic behavior of the polymer molecules can be illustrated by tracking the
interaction energy of the SWNT-polymer molecules. Generally, the interaction energy is
estimated from the difference between the potential energy of the composites system and the
potential energies for the polymer molecules and the corresponding SWNTs as follows:
E Etotal ( ESW NT E polymer ) (18)
where Etotal is the total potential energy of the composite at the end of equilibration, ESW NT
is the energy of the nanotube without the polymer, and E polymer is the energy of the polymer
without the nanotube.
0ps 5ps 10ps
15ps 35ps 55ps
Figure 4. MD simulation snapshots of the SWNT–PE interactions.
0ps 5ps 10ps
15ps 35ps 70ps
Figure 5. MD simulation snapshots of the SWNT–PP interactions.
In the other words, the interaction energy can be calculated as the difference between the
minimum energy and the energy at an infinite separation of the nanotube and the polymer
matix [23, 27, 40].
0ps 5ps 10ps
30ps 35ps 65ps
Figure 6. MD simulation snapshots of the SWNT–PS interactions.

0ps 0.5ps 2.0ps
2.5ps 4.5ps 5ps

Figure 7. MD simulation snapshots of the SWNT–PANI interactions.
19200
19150
Potential energy (Kcal/mol)
19100
19050
19000
18950 BSWNT-PANI
18900 BSWNT-PS
BSWNT-PP
18850 BSWNT-PE
18800
18750
0 20 40 60 80 100
Time(ps)
Figure 8. Potential energy evolution for SWNT-Polymer composites during 100 ps of wrapping
simulation.
Figure 8 shows the potential energies during the simulation and we can find that the
potential energies of the four composites are almost the same during the simulations. Figure 9
shows the interactions during the wrapping process for PE, PP, PS, and PANI. Initially, for all
the polymers the interaction between SWNTs and polymer chains gradually decreases. For
PE and PP, the interaction decreases to -13 kcal/mol, while PS and PANI decreases to -20
kcal/mol, which is much stronger. Both PS and PANI are molecules with side groups of
aromatic rings, so the interaction energy between polymers and SWNTs may be influenced by
aromatic rings greatly.
0
Interaction energy (Kcal/mol)
SWNT-PANI SWNT-PS
-5 SWNT-PP SWNT-PE
-10
-15
-20
-25
0 20 40 60 80 100
Time(PS)
Figure 9. Interaction energy evolution for SWNT-Polymer composites during 100 ps of wrapping
simulation.
The Influence of Different Factors on Polymer Adhesion

Firstly, we put different polymer chains near the nanotube in order to save simulation
time, then the MD simulations were carried out to study the interactions between SWNTs and
individual molecules of PP, PE, PS, and PANI for 200 ps. After this, the systems were
minimized and the total interaction energies between the SWNTs and the polymers in
equilibrium were recorded.
Influence of the temperature on polymer adhesion. To assess the temperature dependence
of the adhesion energy between the polymers and the SWNTs, some MD simulations were
carried out at different temperatures, which varied from 300 K and 500 K in steps of 25 K.
Figure 10 shows the temperature dependence of the intermolecular interaction. It was shown
that the temperature influence is very small for all considered polymers except for PANI.
Influence of the nanotube radius on polymer adhesion. In order to determine the
influence of the nanotube radius on polymer adhesion, some MD simulations were carried out
on SWNTs with PE, PP, PS, and PANI, respectively. The SWNT radius was varied from 8.14
to 27.12 Å in these simulations. The simulations show that the attractive interaction between
the simulated polymers and the SWNTs monotonically increases when the SWNT radius is
increased. Especially, for PS and PANI , the interactions is much stronger and increases
rapidly than that of PE and PP. PS and PANI are both polymers with aromatic rings, which
are therefore expected to possess a strong attractive interaction with the surface of the
SWNTs and may play an important role in providing effective adhesion. When the SWNT
radius are increased, aromatic rings in polymers can well align parallel to the surface, and
thus the interactions between the aromatic rings and the SWNTs increases
significantly as shown in Figure 11.
-15
-20
-25
PE PP
PS PANI
-30
300 350 400 450 500
Temperature(K)
Figure 10. Intermolecular interaction as a function of temperature.
-15
-20
-25
-30
PE PP
PS PANI
10 15 20 25
-10
SCNT diameter(10 )m
Figure 11. Intermolecular interaction as a function of SWNT diameter.

Influence of the nanotube chirality on polymer adhesion. In this part of our study, the
initial atomic configurations of SWNTs are obtained by creating the planar hexagonal carbon
atom network corresponding to an (n, m) nanotube cut open axially. The corresponding chiral
angle and diameter Dn of a SWNT with (n, m) indices could be determined by using the
rolling grapheme model:
3m 3
arctan( ), Dn b (n 2 m2 nm) , (0 m n) (19)
2n m
where b is the C-C bond length23. Eleven types of SWNTs, with different chirality ranging
from zigzag (10, 0) to armchair (10, 10), are generated as shown in Figure 1 and 12. The total
number of atoms, diameters and lengths for each chiral nanotube are presented in Table 1.
10,0zigzag 10,1 10,2 10,3 10,4
10,5 10,6 10,7 10,8 10,9
Figure 12. Schematics of different chiral nanotubes.

Table 1. Total number of atoms utilized in MD simulation for chiral SWNTs
H C Nanotube diameter Nanotube length

atoms atoms Å Å
(10,0) SWNT zigzag 20 400 7.83 42.60
(10,1) SWNT 26 400 8.25 40.43
(10,2) SWNT 28 400 8.72 38.26
(10,3) SWNT 32 400 9.23 36.13
(10,4) SWNT 32 400 9.78 34.11
(10,5) SWNT 30 400 10.36 32.20
(10,6) SWNT 40 400 10.96 30.43
(10,7) SWNT 36 400 11.59 28.79
(10,8) SWNT 36 400 12.23 27.27
(10,9) SWNT 38 400 12.89 25.88
(10,10) SWNT armchair 40 400 13.56 24.60
Figure 13 shows the adhesion energy of the composite versus the corresponding chirality
of the nanotube. We can find that the adhesion energy increases slowly when the chirality
becomes higher. However, it is clear that nanotubes with lower chirality indices tend to have
smaller diameter and longer cylindrical axes compared to those with higher chirality, such as
the armchair nanotube (10, 10). The longer the nanotube, the more nanotube surface area to
form bonds between the nanotube and matrix, therefore, the adhesion energy between the
CNT and the polymer strongly depends on the chirality. Thus, our simulations indicated that
the lowest chirality nanotubes are the best nanotube type for reinforcement.
-10
-15
-20
PE
-25
PP
PS
PANI
(10,0)(10,1)(10,2)(10,3)(10,4)(10,5)(10,6)(10,7)(10,8)(10,9)(10,10)
Nanotube chirality
Figure 13. Adhesion energy between different chiral nanotubes and polymer chains.
Filling
The possibility of filling polymers into nanotubes during real-world composite processing
would create the desired structure bridges between nanotubes and polymers. However, the
reality of this filling will be mainly determined by van der Waals interactions between the
polymers and the internal surface of the SWNT. To study the four polymer molecules‘ filling
phenomena, some MD simulations were set up with the polymer chains initially placed near
the opening at one end of the nanotubes along the direction of the nanotube axis.
The simulations show that molecular chains of PE, PP and PS would gradually moved the
entire molecule body into the nanotubes, while PANI molecules can‘t encapsulate into the
nanotube even after a long time of simulation, which maybe caused by the strong polarity of
PANI. The configurations of PE, PP and PS molecules filling into SWNTs, which is observed
at different time steps of the MD simulations, are shown in Figures 14 16.
0ps 20ps 30ps
40ps 55ps 80ps
Figure 14. MD simulation of PE molecule filling into SWNT cavity.

0ps 30ps 70ps
80ps 85ps 95ps
Figure 15. MD simulation of PP molecule filling into SWNT cavity.
0ps 100ps 300ps

400ps 1000ps 2000ps
Figure 16. MD simulation of PS molecule filling into SWNT cavity.
For PE and PP, the molecules were lingering around the opening firstly, then constantly
changing their orientation to facilitate filling into the SWNT opening, and encapsulating into
the nanotube within 100 ps. An equilibrium state of the system was also achieved when the
polymer molecules were entirely in the nanotube. However, as shown in Figures 14 and 15,
we also notice that the PE molecules will stay in the middle of the nanotube, while PP
molecules will stay near the opening instead of going to the middle of the nanotube or the
opposite opening, which maybe because of PP has side groups of methyl and is not as smooth
as PE molecular chains. For PS, however, a very slow filling process is observed compared
with the fast filling phenomenon of PE and PP, which cost about 2000 ps as shown in Figure
16. The PS chain has side groups of aromatic rings, which may be an important factor for the
slow filling.
19200
19150
Potential Energy(Kcal/mol)
19100
19050
19000
(SWNT-PS)
18950 (SWNT-PANI)
18900 (SWNT-PP)
(SWNT-PE)
18850
18800
18750
0 20 40 60 80 100
Time(ps)
Figure 17. Potential energy evolution for SWNT-Polymer composites during 100 ps of filling
simulation.
0
SWNT-PS SWNT-PANI

-5 SWNT-PP SWNT-PE
-10
-15
-20
-25
-30
-35
0 20 40 60 80 100
Time(ps)
Figure 18. Interaction energy evolution for SWNT-Polymer composites during 100 ps of wrapping
simulation.
Figure 17 shows the potential energies and interaction energies during the simulations
and we can see that the potential energies of the four composites are also almost the same
during the simulations. Figure 18 shows the interactions during the filling process for PE, PP,
PS, and PANI. Initially, just as in the wrapping process, for all the polymers the interaction
between SWNTs and polymer chains gradually decreases till they encapsulated into the
nanotube and achieved an equilibrium state. We also notice that the interaction energy will
decreases rapidly for PE, PP and PS once they start filling the nanotube, which reveals that
the filling can improve the load transfer between polymers and SWNTs significantly.
Conclusion I
In this study, some MD simulations are carried out to investigate the interactions of PE,
PP, PS and PANI molecules with SWNTs in a vacuum. The results show that the interaction
between the SWNT and the polymer is strongly influenced by the specific monomer structure
such as aromatic rings, which affect polymers‘ affinities for SWNTs significantly. The results
also show that the attractive interaction between the simulated polymers and the SWNTs
monotonically increases when the SWNT radius is increased while temperature influence is
neglectable for all considered polymers except for PANI. The MD simulations also indicate
that the adhesion energy between the SWNT and the polymer strongly depends on the
chirality and the lowest chirality nanotubes are the best nanotube type for reinforcement.
Lastly, the filling simulations reveal that molecules of PE, PP and PS can fill into a (10, 10)
SWNT cavity due to the attractive van der Waals interactions. The possible extension of
polymers into SWNT cavities can be used to structurally bridge the SWNTs and polymers to
significantly improve the load transfer between them when SWNTs are used to produce
nanocomposites.
INFLUENCE OF CHIRALITY ON THE INTERFACIAL BONDING

CHARACTERISTICS OF CARBON NANOTUBE POLYMER COMPOSITES
In this section, the influence of chirality on the interfacial bonding between the SWNTs
and polymer were investigated using MM and MD simulations. In order to reveal the effect of
the chirality on the interfacial bonding characteristics of the nanotube reinforced composites,
several tubes with different chiralities but similar tube radii and lengths are selected.
Molecular Model
Poly(methyl methacrylate), (PMMA), is a well-known glassy polymer used in a variety

of engineering areas from aircraft glazing to lightweight construction systems. Therefore,
PMMA, with 10 repeating units in each chain, is chosen as a matrix in the study, which is
also due to its simplicity and generic representation feature for polymer materials. Five types
of SWNTs, with different chirality but similar molecular weights, diameters and lengths, are
generated as shown in Figure 19. The total number of atoms, diameters and lengths for each
nanotube are presented in Table 2.
In the simulations, each of the composite systems was composed of a fragment of SWNT
totally embedded inside the amorphous polymer matrix. A model of the composite system
embedded by the pristine SWNT, which consisted of a supercell in the range of 57 Å×57
Å×62 Å, is shown in Figure 20. Each of the configurations was initiated by randomly
generating 112 PMMA molecular chains surrounding the SWNT using an initial density of
1.2 g/cm3. The models were put into an NPT ensemble simulation with a pressure of 10 atm
and a temperature of 300 K for 10 ps with a time step of 1 fs while holding the nanotube
rigid. The purpose of this step was to slowly compress the structure of the matrix polymers to
generate initial amorphous matrix with the correct density and low residual stress. The matrix
polymers were then put into an NVT ensemble simulation and equilibrated for 20 ps with a
time step of 0.2 fs with rigid nanotubes. After that, the composites systems were further
equilibrated for 40 ps at a time step of 2 fs with non-rigid nanotubes to create a zero initial
stress state using NVT ensembles.
Table 2. Total number of atoms utilized in MD simulation for chiral SWNTs
Type of H C Nanotube diameter Nanotube length Chiral angel


SWNTs atoms atoms Å Å
(10, 10) SWNT
40 960 13.56 59.03 30.00
armchair
(12, 8) SWNT 40 960 13.65 58.64 23.43
(14, 5) SWNT 44 960 13.36 59.93 14.71
(16, 2) SWNT 44 960 13.38 59.83 5.82
(17, 0) SWNT
50 960 13.31 60.14 0.00
zigzag
The energy of the composite systems was minimized to achieve the strongest bonding
between the nanotubes and the polymer27, 40. Finally, the interfacial bonding energy and
interfacial shear stress were calculated through pullout simulations.
Figure 19. Schematics of different chiral nanotubes.
a b
Figure 20. Molecular model of The SWNT/PMMA composite system: (a) top view; (b) section view.
The bonding strength between the SWNTs and the polymers can be evaluated by
interfacial energy in the composites. Generally, the interaction energy is estimated from the
difference between the potential energy of the composites system and the potential energy for
the polymer molecules and the corresponding SWNTs as described in equation (18). The total
interaction energy, E , is twice the interfacial bonding energy scaled by the contact area
A [41]:
E
(20)
2A
The pullout simulations were performed to characterize the interfacial shear stress of the
composites. The pullout energy, E pullout , is defined as the energy difference between the full
embedded nanotube and the complete pullout configuration. The pullout energy was divided
into three terms as follows [27]:
E pullout E2 E1 ( E2 ESW NT2 E polymer2 ) ( E1 ESW NT1 E polymer1 )

(21)
( E2 E1 ) ( ESW NT 2 ESW NT1 ) ( E polymer2 E polymer1 )
The pullout energy can be related to the interfacial shear stress, i , by the following
relation:
x L
E pullout 2 r ( L x) i dx r i L2 (22)
x 0
E pullout
(23)
rL2
i
where r and L are the outer radius and length of the SWNT, respectively, and x is the
coordinate along the longitudinal tube axis [24.]
In the first part of the study, the interfacial bonding for armchair SWNT was investigated.
The pullout simulations of a SWNT were performed in order to characterize the interfacial
shear strength of the composites. In order to demonstrate the statistical validity of the results,
the pullout simulations were performed on five molecular models with different initial
configurations and the error was calculated. Figure 21 shows the snapshots of the pullout
simulations. The SWNT was pulled out of the PMMA matrix along the nanotube axis
direction.
The potential energy, interaction energy, and interfacial bonding energy were plotted
against the displacement of the SWNT from the PMMA matrix, as shown in Figure 22.
Figure 22a indicates that the potential energy of the SWNT/PMMA composite system was
increased as the SWNT was pulled out of the PMMA.
Figure 21. Snapshots from the MD simulation of the pullout of the SWNT.
Figure 22. Energy plots during the pullout of the SWNT. (a) potential energy; (b) interaction energy; (c)
interfacial bonding energy.
During the pullout, the interaction energy changed with the displacement linearly and
decreased toward a value of zero, as shown in Figure 22b. This is due to the stable interfacial
binding interaction and the decrease of contact area during the pullout. The interfacial binding
energy kept constant with a value of 0.1 Kcal/mol Å2 during the pullout, as shown in Figure.
22c. After the SWNT was completely pulled out of the PMMA, the potential energy of the
system was level off and the interaction energy then kept zero. The average potential energy
of the composites was 42,122 Kcal/mol at the initial stage of the pull out and increased to
42,501 Kcal/mol after the pullout. From the pullout simulation, the interfacial shear strength
between the pristine SWNT and the PMMA was about 36 MPa.
Next, pullout simulations were performed on the other four kinds of SWNTs and the
influence of chirality on the interfacial bonding has been investigated. In this part of the
simulations, the interaction energy also changed with the displacement nearly linearly and
decreased toward a value of zero during the pullout, as shown in Figure 23. Figure 24 shows
the interaction energy, interfacial bonding energy and shear stress of the composite versus the
corresponding chirality of the nanotube, all of which is the average value during the pullout.
It was shown that the interaction energy, interfacial bonding energy and shear stress attain the
highest value for the armchair system, while the zigzag nanotube composite produces the
least value. Therefore, the binding energy between the SWNTs and the PMMA depends on
the chirality and the armchair SWNT is the best nanotube type for reinforcement.
Chirality dependent conformation of PMMA molecule at nanotube interface has been
investigated through MD simulations by Wei [42]. Local wrapping angle is defined as the
angel between the vector connecting the two ends of a three segment subchain on a polymer
molecule and the nanotube axis. The simulations indicated that, while wrapping around 0
dominate on a small radius armchair SWNT, molecule wrapping shift to larger angels on a
similar radius zigzag tube. The different conformations of polymer molecules at various
SWNT interfaces may cause different interfacial bonding energy and the armchair SWNTs
may have the strongest adhesion with the polymers.
Figure 23. Interaction energy plots during the pullout of the SWNTs from the matrix.
Figure 24. Influence of chirality on the interfacial bonding characteristics of SWNT. (a) interaction
energy; (b) interfacial bonding energy; (c) shear stress.
Conclusion II
In summary, MM and MD simulations have been used to study the influence of chirality
on the interfacial bonding characteristics between the SWNTs and the PMMA matrix. Several
SWNTs, which include (10, 10), (12, 8), (14, 5), (16, 2), and (17, 0), with different chiralities
but similar tube radii and lengths are selected. The simulations indicate that the interfacial
bonding energy between the SWNT and the PMMA depends on the chirality. Substantial
adhesion exists between the nanotube and PMMA when the nanotube has a higher chiral
indices or larger chiral angel. For SWNTs with similar molecular weights, diameters and
lengths, the armchair nanotube may be the best nanotube type for reinforcement. The general
conclusions derived from this work may be of importance in devising advanced nanotube
reinforced composites.
EFFECT OF CHEMISORPTION ON THE INTERFACIAL BONDING

CHARACTERISTICS OF CARBON NANOTUBE POLYMER COMPOSITES
While chemically cross-linking or molecular entanglement method to strengthen the
interface has been conducted [25,41], studies involving the influence of chemical attachment
on interfacial properties of CNTs have not been reported. This section was about finding a
viable way to get SWNT to go into the polymer matrix to act as reinforcement. The
difficulties involved in doing this arise mostly from the fact that CNTs cannot interact well
with other materials. The method pursued here for making the SWNTs capable of effectively
interacting with the polymer was based on the covalent attachment of functional groups to the
surface of SWNTs. By taking advantage of these functional groups, which could act as an
effective interfacial bridge between the SWNTs and the polymeric matrix, an effective load
transfer could be achieved between the SWNTs and the polymer matrix. In this study, the
influence of chemical functionalization on the interfacial bonding between the SWNTs and
polymer was investigated using MM and MD simulations.
Molecular Model
Poly(methyl methacrylate) (PMMA) and polyethylene (PE) are well-known polymers

used in a variety of engineering areas. Therefore, PMMA and PE, with 10 repeating units in
each chain, are chosen as matrixes in the study, which are also due to their simplicity and
generic representation feature for polymer materials. The molecular model of PMMA and PE
are shown in Figure 25. (10, 10) SWNTs, which have diameters of 13.56 Å and lengths of
59.03 Å, are selected for the simulations of the SWNT/PMMA composites except special
conditions. The unsaturated boundary effect was avoided by adding hydrogen atoms at the
ends of the SWNTs.
The literature supplies numerous examples of addition reactions on nanotube surfaces43.
Ying et al. [44] reported the grafting of aromatic rings on the side walls of SWNTs. In the
present work, we used phenyl groups to functionalize the nanotube surface due to its
simplicity and generic representation for the functionalization of CNTs.
As we know, the dispersion degree of the SWNTs in the PMMA matrix can affect the
interfacial bonding characteristics between the SWNTs and the PMMA matrix. However, if
we consider the dispersion degree of the SWNTs, the computational system would be so large
that it would cost too long a time for the simulation.
(a) (b)
Figure 25. Molecular model of PMMA and PE.

(a) with carboxylic groups (－COOH)
(b) with amide groups (－CONH2)
(c) with alkyl groups (－C6H11)
(d) with phenyl groups (－C6H5)
Figure 26. Illustrating of a (10, 10) SWNT with functional groups randomly chemisorbed to 5% of the
carbon atoms.
Also, since the main goal of our research is to investigate the interfacial bonding
characteristics between the SWNTs and the PMMA matrix, the SWNTs were assumed to be
well dispersed in the PMMA matrix and the simulation results would be useful for the
production of SWNTs reinforced polymer composites as well.The composites, reinforced by
pristine SWNT and the SWNTs on which 0.5%, 2.5%, 5%, 7.5%, or 10% of the carbon atoms
had a bonded phenyl group, were simulated by using MM and MD simulations. In the part of
the influence of modification type, MM and MD simulations were performed on composites
reinforced by pristine SWNT and the SWNTs on which 5% of the carbon atoms had bonded
groups (carboxylic group COOH, amide group CONH2, alkyl group C6H11, or phenyl group
C6H5). The functional groups were randomly end-grafted to the surface of the SWNTs.
SWNTs with functional groups randomly chemisorbed to 5% of the carbon atoms are
illustrated in the left panel of Figure 26. The chemical functionalization of the SWNTs has
been performed by attaching functional groups to the surface of the CNTs through chemical
covalent bonding and the functional groups were randomly end-grafted to the surface of the
SWNTs. The associated change in geometry of the atoms on which the phenyl groups are
bonded is illustrated in the right panel of Figure 26, where the bonded nanotube atoms are
―r aised‖ away from the nanotube axes and have hybridizations that change from sp2 to sp3
[45].
In the simulations, each of the composite systems was composed of a fragment of SWNT
totally embedded inside the amorphous polymer matrix. A model of the composite system
embedded by the pristine SWNT, which consisted of a supercell in the range of 57 Å×57
Å×62 Å, is shown in Figure 27. For SWNT-PMMA system, each of the configurations was
initiated by randomly generating 112 PMMA molecular chains surrounding the SWNT using
an initial density of 1.2 g/cm3. For SWNT-PE system, each of the configurations was initiated
by randomly generating 247 PE molecular chains surrounding the SWNT using an initial
density of 0.9 g/cm3.The models were put into an NPT ensemble simulation with a pressure
of 10 atm and a temperature of 300K for 10 ps at a time step of 1 fs while holding the
nanotube rigid.
Figure 27. Cross-section view of SWNT/PMMA system before simulation.

(a) (b)
Figure 28. Illustration of the composite embedded by a (10, 10) SWNT with phenyl groups randomly
chemisorbed to 2.5% of the carbon atoms: (a) top view; (b) side view.
The purpose of this step was to slowly compress the structure of the matrix polymers to
generate initial amorphous matrix with the correct density and low residual stress. The matrix
polymers were then put into an NVT ensemble simulation and equilibrated for 20 ps with a
time step of 0.2 fs with rigid nanotubes.
After that, the composites systems were further equilibrated for 40 ps at a time step of 2
fs with non-rigid nanotubes to create a zero initial stress state using NVT ensembles. The
energy of the composite systems was minimized to achieve the strongest bonding between the
nanotubes and the polymer shown in Figure 28 [24, 40]. Finally, the interfacial bonding
energy and interfacial shear stress were calculated through pullout simulations.
Interfacial Bonding for Pristine SWNT

The pullout simulations of a SWNT were performed in order to characterize the
interfacial shear strength of the composites. The SWNT was pulled out of the polymer matrix
along the nanotube axis direction.
The potential energy, interaction energy, and interfacial bonding energy were plotted
against the displacement of the SWNT from the polymer matrix, as shown in Figure 29 and
30. During the pullout, the interaction energy changed with the displacement linearly and
decreased toward a value of zero, as shown in Figure 29a and 30a.
This is due to the stable interfacial binding interaction and the decrease of contact area
during the pullout. Figure 29b and 30b indicates that the pullout energy of the SWNT/
polymer composite system was increased as the SWNT was pulled out of the polymer.
The interfacial binding energy kept constant with a value of 0.1 Kcal/mol Å2 during the
pullout, as shown in Figure 29c and 30c. After the SWNT was completely pulled out of the
polymer, the potential energy of the system was leveled off and the interaction energy then
kept zero.
Energy (Kcal/molÅ ) Energy (Kcal/mol) Energy (Kcal/mol)

0
-100
-200 (a)
-300
-400
-500
-600
0 10 20 30 40 50 60 70
400
300 (b)
200
100
0
0 10 20 30 40 50 60
-0.06
2
-0.08 (c)
-0.10
-0.12
-0.14
0 10 20 30 40 50
Displacement (Å )
Figure 29. Energy plots during the pullout of the SWNT from the PMMA matrix. (a) interaction
energy; (b) pullout energy; (c) interfacial bonding energy.
0
Energy (Kcal/mol)
-100
-200 (a)
-300
-400
-500
-600
-700
0 10 20 30 40 50 60 70
500
Energy (Kcal/mol)
400
(b)
300
200
100
0
0 10 20 30 40 50 60
Energy (Kcal/molÅ )
2
-0.10
-0.12 (c)
-0.14
-0.16
-0.18
0 10 20 30 40 50
Figure 30. Energy plots during the pullout of the SWNT from the PE matrix. (a) interaction energy; (b)
pullout energy; (c) interfacial bonding energy.
Influence of Chemical Functionalization Degree

The SWNTs and the polymer matrix were not held fixed in the pullout simulation.
Therefore, the pullout energy has been influenced by the deformation of the nanotubes and
polymer during the pullout. Figure 31 and 32 shows the interaction energy changed with the
displacement nearly linearly during the pullout, which is due to the stable interfacial binding
interaction between the SWNTs and the polymer.
0
Energy (Kcal/mol)
-500
-1000
Unfunctionalized
0.5% functionalized
-1500 2.5% functionalized
5.0% functionalized
7.5% functionalized
-2000
10% functionalized
0 10 20 30 40 50 60 70
Displacement (Å)
Figure 31. Interaction energy plots during the pullout of the SWNT from the PMMA matrix.
0
-400
Energy (Kcal/mol)
-800
-1200
-1600
-2000 Unfunctionalized
0.5% functionalized
-2400 2.5% functionalized
5.0% functionalized
-2800
7.5% functionalized
-3200 10% functionalized
-3600
0 10 20 30 40 50 60 70
Displacement (Å)
Figure 32. Interaction energy plots during the pullout of the SWNT from the PE matrix.
However, the interaction energy will keep zero after the nanotubes were completely
pulled out of the polymer because there was no interaction between the nanotubes and the
polymer [25]. Plotted in Figure 33a-b and 34a-b is the calculated interaction energy and
interfacial bonding energy for SWNTs as a function of the degree of functionalization. When
the degree of functionalization is increased, the interaction energy and interfacial bonding
energy between the simulated SWNTs and the polymer monotonically increases toward a
magnitude value, which is about four times the value for pristine SWNT. The interfacial
bonding, which appears to be critically dependent on the nanotube-polymer interface surface
area, will increase linearly with the total interface surface area [10]. When the SWNT is
chemically attached with phenyl groups, the contact area between the nanotube and the
polymer matrix will be drastically increased, which will cause the increase of the interfacial
bonding between the nanotube and the polymer. Figure 33c and 34c shows the growth and
saturation with higher degrees of functionalization. The results show that the shear stress of
nanotube-polymer interface with weak nonbonded interactions can be increased by about
1000% with the introduction of a relatively low density ( 5%) of chemical attachment.
However, the shear stress increases only weakly with the introduction of a relatively high
density (>5%) of chemical attachment. When the functional groups of SWNT were
successfully embedded into the polymer matrix, which may possibly link SWNTs with the
polymer matrix, the shear stress could be effectively increased. The successful embedding
could happen at a low density of functionalization and thus an effective enhancement of the
shear stress could be attained.
Energy (Kcal/mol)
-800
-1200
-1600 (a)
-2000
0 2 4 6 8 10
Shear stress (MPa) Energy (Kcal/molÅ )
-0.1
2
-0.2
-0.3
(b)
-0.4
0 2 4 6 8 10
400
300
200
100 (c)
0 2 4 6 8 10
Degree of functionalization (%)
Figure 33. Influence of chemical functionalization on the interfacial bonding characteristics of SWNT
for SWNT/PMMA system. (a) interaction energy; (b) interfacial bonding energy; (c) shear stress.
Energy (Kcal/mol)
-500
-1000
-1500
-2000
-2500
-3000
-3500
0 2 4 6 8 10
Energy (Kcal/molÅ )
2
-0.2
-0.4
-0.6
0 2 4 6 8 10
600
Shear stress (MPa)
400
200
0
0 2 4 6 8 10
Degree of functionalization (%)
Figure 34. Influence of chemical functionalization on the interfacial bonding characteristics of SWNT
for SWNT/PE system. (a) interaction energy; (b) interfacial bonding energy; (c) shear stress.
However, when the density of functionalization becomes higher, some functional groups
may only contact with the other functional groups, which may lead to a direct result that the
effective contact surface area between the functional groups and the polymer matrix couldn‘t
be strongly increased any more, and thus there is only a weak increase of the shear stress.
Although the covalent attachment of functional groups to the surface of nanotubes can
improve the efficiency of load transfer, these functional groups might introduce defects on the
walls of the perfect structure of the nanotubes, which will lower the strength of the nanotube
filler. Some models predict that the change in mechanical properties of the SWNTs with
lower level ( 10%) of functionalization is negligible [25]. So SWNT with 5% level of
functionalization may be the feasible nanotube type for reinforcement, which could improve
the efficiency of load transfer with negligible influence on the mechanical properties of the
SWNT. It also supports suggestions that chemical attachment of nanotubes during processing
may be in part responsible for the enhanced stress transfer observed in some systems of the
nanotube-polymer composites.
Influence of Chemical Functionalization Type

Figure 35 shows the interaction energy changes with the displacement nearly linearly
during the pullout, which is due to the stable interfacial binding interaction between the
SWNTs and the PE matrix. However, the interaction energy will keep zero after the
nanotubes were completely pulled out of the PE matrix because there was no interaction
between the nanotubes and the PE matrix.
Energy (Kcal/mol) -500
-1000
-1500 Unfunctionalized
With carboxylic groups
-2000 With amide groups
With alkyl groups
With phenyl groups
-2500
0 10 20 30 40 50 60 70
Displacement (Å )
Figure 35. Interaction energy plots during the pullout of the SWNTs from the PE matrix.
Plotted in Figure 36 is the calculated interaction energy (Figure 36a) and interfacial
bonding energy (Figure 36b) of the SWNT-PE system for SWNTs as a function of the type of
functionalization. The figure shows that although the four types of functionalization could
increase the adhesion with the polymer matrix, the special structure of the functional groups
plays a very important role in determine adhesion to the matrix. We can observe that the
SWNT functionalized with -C6H5 directly in the surface has the most strong interaction with
the matrix (about 4 times as compared with that of pristine SWNT), whereas the SWNT with
-COOH, -CONH2, or -C6H11 groups has increased only weakly than pristine SWNT.
The interfacial bonding, which appears to be critically dependent on the nanotube-
polymer interface surface area, will increase linearly with the total interface surface area [10].
When the SWNT is chemically functionalized, the contact area between the nanotube and the
polymer matrix will be drastically increased, which will cause the increase of the interfacial
bonding between the nanotube and the polymer. The aromatic ring (-C6H5) may has more
contact area with the matrix than the other three functional groups for its special ring
structure, which may cause the effective increase of adhesion energy.
Figure 36c shows the interfacial shear stress between SWNTs and PE matrix as a
function of the type of functionalization. The figure shows that the SWNT functionalized with
-C6H11 or -C6H5 has increased the shear stress effectively (-C6H11: about 3 times as compared
with that of pristine SWNT, -C6H5: about 17 times as compared with that of pristine SWNT),
whereas the SWNT with -COOH, or-CONH2 groups has almost the same value as compared
with that of the pristine SWNT. As shown in Figure 26, the backbones of -C6H11 and -C6H5
groups nearly parallel with the radial direction of the SWNT, while the -COOH, or-CONH2
groups nearly parallel with the SWNT axis direction. When the functional groups of SWNT
were successfully embedded into the polymer matrix, which may possibly link SWNTs with
the polymer matrix, the shear stress could be effectively increased. The successful embedding
could be happen when the functional groups parallel with the radial direction of the SWNT,
and thus the shear stress could be effectively increased when the SWNTs are functionalized
with -C6H11 or -C6H5 groups.
Energy (Kcal/mol) -2500

-2000
-1500 Interaction energy
-1000
-500
0
)
-50
-40
Shear Stress (MPa) Energy (Kcal/molÅ
-30 Interfacial bonding energy

-20
-10
0
600
500
400
300
shear stress
200
100
0
Unfunctionalized Carboxyl Amide Alkyl Phenyl
Type of functionalization
Figure 36. Influence of modification on the interfacial bonding characteristics of SWNT-PE system.
The primary objective of this work was to select the appropriate functional groups that
would provide a good interaction mechanism with the composite‘s matrix and act as a load
transfer conduit between the SWNTs and the composite matrix.
Conclusion III
In summary, we have used MM and MD simulations to study the effect of chemical

functionalization on the interfacial bonding characteristics between the SWNTs and the
polymer matrix. The simulations show that functionalization of nanotubes at low densities of
functionalized carbon atoms drastically increase their interfacial bonding and shear stress
between the nanotubes and the polymer matrix. This indicates that increasing the load transfer
between SWNTs and a polymer matrix in a composite via chemisorption may be an effective
way and chemical attachment of nanotubes during processing may be in part responsible for
the enhanced stress transfer observed in some systems of the nanotube-polymer composites.
Furthermore, this suggests the possibility to use functionalized nanotubes to effectively
reinforce other kinds of polymer-based materials as well.
CONCLUSION
In this study, force-field-based MD simulations are performed to study the interaction
between polymers and SWNTs. The ―w rapping‖ of nanotubes by polymer chains was
computed. The influence of temperature, nanotube radius and chirality on polymer adhesion
was investigated. Furthermore, the ―f illing‖ of nanotubes by polymer chains was examined.
The results show that the interaction between the SWNT and the polymer is strongly
influenced by the specific monomer structure such as aromatic rings, which affect polymers‘
affinities for SWNTs significantly. The attractive interaction between the simulated polymers
and the SWNTs monotonically increases when the SWNT radius is increased. The
temperature influence is neglectable for PE and PP but strong for PS and PANI. Also, our
simulations indicate that the adhesion energy between the SWNT and the polymer strongly
depends on the chirality. For SWNTs with similar molecular weights, diameters and lengths,
the armchair nanotube may be the best nanotube type for reinforcement. The simulations of
filling reveal that molecules of PE, PP and PS can fill into a (10, 10) SWNT cavity due to the
attractive van der Waals interactions. The possible extension of polymers into SWNT cavities
can be used to structurally bridge the SWNTs and polymers to significantly improve the load
transfer between them when SWNTs are used to produce nanocomposites.
Also, the influence of chemical functionalization on the interfacial bonding
characteristics of SWNTs reinforced polymer composites was investigated using MM and
MD simulations. The simulations show that functionalization of nanotubes at low densities of
functionalized carbon atoms drastically increase their interfacial bonding and shear stress
between the nanotubes and the polymer matrix. This indicates that increasing the load transfer
between SWNTs and a polymer matrix in a composite via chemisorption may be an effective
way and chemical attachment of nanotubes during processing may be in part responsible for
the enhanced stress transfer observed in some systems of the nanotube-polymer composites.
Furthermore, this suggests the possibility to use functionalized nanotubes to effectively
reinforce other kinds of polymer-based materials as well.
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Lecture Material 7
CARBON NANOTUBES: MOLECULAR

DYNAMICS AND MECHANICAL PROPERTIES
ABSTRACT
Molecular mechanics calculations for the in-place stiffness, shear modulus, and
bending rigidity of both single- and double-walled carbon nanotubes are reported by
calculating the strain energy of carbon nanotubes and graphite sheets subject to various
types of loading. Elastic rod and plate theories are employed to link the material
properties of carbon nanotubes directly to the molecular mechanics calculations. The
length dependence of these material properties is reported and investigated via nonlocal
elasticity theory. In addition, the van der Waals effect on the differences between the
material properties of double- and single-walled carbon nanotubes is also examined. The
diminishment of such differences in large sizes of carbon nanotubes is revealed from the
simulations.
INTRODUCTION
Carbon nanotubes (CNTs), which were discovered by Iijima in 1991 [1], are
macromolecules of carbon in a periodic hexagonal arrangement with a cylindrical shell shape.
As a new type of nano-scale material, CNTs have aroused great interest among researchers
because of their remarkable mechanical, thermal, electrochemical, piezoresistive, and other
physical properties [2-5]. They can be viewed as one (or more) graphite sheet(s) rolled into a
seamless tube. The way this graphite sheet is wrapped is represented by a pair of indices (n,
m) that are called the chirality. When m = 0, the nanotubes are called ―zi
gzag,‖ and when n =
m, they are called ―ar mchair.‖ Research findings have revealed extremely strong mechanical
properties in these materials [6-7]. Of the mechanical properties of CNTs, accurate estimates
of the Young‘s modulus, shear strength, and bending rigidity are critical for their potential
application.
Experimental endeavors have been undertaken to estimate the mechanical properties of
CNTs. Treacy et al. [8] obtained the Young‘s modulus of multi-walled CNTs (MWCNTs) in
a wide range, from 0.4 to 4.15 TPa, with an average of 1.8 TPa, by measuring the amplitude
of their intrinsic thermal vibrations in a transmission electron microscope. Salvetat et al. [9]
used an atomic force microscope to conclude that the elastic and shear modules of a single-
walled CNT (SWCNT) were of the order of 1 TPa and 1 GPa, respectively. Krishman et al.
[10] calculated the Young‘s modulus of SWCNTs to be 0.9 ~ 1.7 TPa by observing their
freestanding room-temperature vibrations in a transmission electron microscope. Wong et al.
[11] experimentally studied the Young‘s modulus of individual, structurally isolated silicon
carbide nanorods and MWCNTs that were pinned at one end to molybdenum disulfide
surfaces and found the values to be 0.7 ~ 1.9 TPa. Poncharal et al. [7] used a transmission
electron microscope to observe the static deformation of an MWCT and indicated that the
Young‘s modulus of the materials was about 1 TPa. The wide variety of experimentally
obtained mechanical properties of CNTs is mainly due to the uncertainty and uncontrollable
environmental effects on the measurements in the experiments. Although a rough estimate of
the order of the mechanical properties of CNTs is available, a more accurate prediction is still
necessary and is, in fact, indispensable for capturing the extensive potential of these materials.
In addition to experimental endeavors, theoretical evaluations of the material properties
of CNTs have also been undertaken. Theoretical modeling is usually classified into two main
categories. The first is atomic modeling, and it includes such techniques as classical
molecular dynamics [12-13], tight-binding molecular dynamics [14], and density functional
theory [15]. Because these atomic methods are limited to systems with a small number of
molecules and atoms, they are restrained to small-scale modeling. Continuum mechanics
modeling, in contrast, is practical for the analysis of CNTs for large-scale systems. Thus, both
continuum modeling and molecular dynamics have been employed to estimate the mechanical
properties of CNTs. Closed-form expressions for the axial elastic properties of chiral CNTs
were recently presented by Chang et al. [16-17]. In their work, a nonlinear stick-spiral model
was developed to investigate the material behavior of SWCNTs, especially the estimates of
the Young‘s modulus and shear modulus of CNTs based on a molecular mechanics concept.
The major conclusion based on this nonlinear model was that the elastic properties are
chirality-independent for SWCNTs with a diameter of less than 2 nm, thus demonstrating an
obvious scale effect. An attempt to estimate the material properties of achiral CNTs was also
made via the concept of a representative volume element of the chemical structure of a
graphite sheet [18-19]. In addition to measurements of the Young‘s modulus and shear
modulus of CNTs, their bending rigidity has also been investigated. Adams et al. [20] applied
quantum molecular dynamics, using both empirical and local density functional methods, to
evaluate the energies of a number of ball-shaped and tubular fullerenes of various sizes and
suggested that the value of bending rigidity was D = 1.62 eV. Lucas et al. [21] estimated
flexural rigidity D to be 1.78 eV by comparing the dispersion relation of the out-of-plane
acoustic phonons in graphite with the flexural vibrational frequency of a continuum plate.
Odegard et al. [18] introduced a representative volume element of the chemical structure of a
graphite sheet and predicted bending rigidity D 1.12eV for armchair nanotubes and
D 1.22eV for zigzag nanotubes, which was supported by the results of Wang [19]. Kudin
et al. [22] reported ab initio calculations of the mechanical properties of two-dimensional
lattices of carbon, boron-nitride, and fluorine-carbon composites. It has been pointed out that
atomic methods are limited to systems with a small number of molecules and atoms and that
continuum mechanics modeling is practical in the analysis of CNTs for large-scale systems.
However, the classical continuum models themselves are not universally applicable, even for
macro-engineering structures. For example, the Euler buckling load is only valid for steel
columns with aspect ratios greater than 30 [23]. Continuum mechanics modeling is also
unable to capture the atomic structures of nano-materials, and hence further verification of its
applicability is necessary. Therefore, it can be concluded that continuum mechanics models

are essential in the study of large-sized CNTs, but that the effectiveness of any proposed
model should be verified through either atomic modeling or ab initio simulations to justify its
applicability. From the aforementioned divergent results on the material properties of CNTs
via different methods, it can be seen that a simple, effective, and efficient modeling method is
necessary to find a reasonable and accurate estimate of the elastic modulus, shear modulus,
and, especially, the bending rigidity of CNTs in a systematic way. Such a simple and explicit
modeling method is expected to employ the classical continuum mechanics theory while
being able to maintain the nature of the atomic structures in the modeling.
The scale effect on CNTs has been an interesting topic in the nano-community. The
modeling of such a size-dependent phenomenon has become an active and interesting subject
for research [24]. Sun and Zhang [25] pointed out the limited applicability of continuum
models in nanotechnology. They indicated the importance of a semi-continuum model to
analyze nanomaterials. In their semi-continuum model for nano-structured materials with
plate-like geometry, results that contrast with those of classical continuum models were
observed. The values of the material properties were found to be completely dependent on the
thickness of the plate structure. Geng and Chang [17] developed a nonlinear stick-spiral
model to investigate the mechanical behavior of SWCNTs. They discussed the elastic
properties, paying special attention to the effects of tube chirality and tube size. However, the
dependence of the material properties of CNTs on their length has not yet been reported in the
available research findings. Nonlocal elasticity was proposed by Eringen [26-27] to account
for the scale effect on elasticity by assuming the stress at a reference point to be a function of
the strain field at every point in the body. In this way, the internal size scale could be
considered in the constitutive equations simply as a material parameter. The application of
nonlocal elasticity in micro- and nano-materials has received a lot of attention from the
nanotechnology community recently. Peddieson et al. [28] investigated the potential for
applying the nonlocal elastic beam theory to micro- and nano-materials by formulating and
applying a nonlocal version of the Euler-Bernoulli beam theory to the study of a cantilever
beam. The small-scale effect on the wave propagation dispersion relation of a CNT was
explicitly revealed [29] for different CNT wavenumbers and diameters via the nonlocal
elastic beam and shell theories. The scale coefficient in nonlocal continuum mechanics was
then roughly estimated for CNTs from the obtained asymptotic frequency. This research
proved effective in predicting the small-scale effect on CNT wave propagation with a
qualitative validation study based on the published experimental reports. Zhang, Liu, and Han
[30] developed a nonlocal multiple-shell model for the elastic buckling analysis of MWCNTs
under uniform external radial pressure in which the effect of small length scale was
incorporated in the formulation of the buckling pressure. Duan and Wang [31] studied the
axisymmetric bending of micro- and nano-scale circular plates and obtained exact nonlocal
solutions under general loading conditions using a variable transformation technique. To the
best of the authors‘ knowledge, the application of nonlocal elasticity to the estimation of the
material properties of CNTs, especially the verification of the theory with molecular
dynamics, has not yet been explored. The verification of this theory with molecular dynamics
simulations is indispensable if it is to be further developed in the analysis of CNTs.
This manuscript is intended to introduce and summarize the progress that the authors
have made in the modeling of the mechanical properties of CNTs. A simple, effective, and
efficient modeling method for the calculation of the in-plane stiffness, shear modulus, and
bending rigidity of CNTs via molecular mechanics simulations and elastic theory is proposed
and discussed. In particular, the strain energy of CNTs and graphite sheets subject to various
types of loading is used to evaluate all of the material properties via elastic rod and plate
theories. A tight link between the molecular mechanics calculations on the strain energy and
the corresponding second derivative and estimate of the mechanical properties of CNTs is
uncovered, particularly for the derivation, and physical interpretations of the results are
provided. In addition, the length-dependent stiffness of SWCNTs is investigated via nonlocal
elasticity by employing an elastic rod that is subject to axial compression to derive the close-
form solution of this material property. Verification of the obtained stiffness from the
nonlocal elastic rod theory is obtained from the molecular simulation results, and a suggestion
for the scale coefficient that is employed in nonlocal elasticity is also proposed. Finally, the
length dependence of the mechanical properties of double-walled carbon nanotubes
(DWCNTs) is explored, and the van der Waals effect on the differences between these
properties in DWCNTs and SWCNTs is investigated.
MECHANICAL PROPERTIES OF SWCNTS

In-plane stiffness, shear modulus, and bending rigidity are investigated using continuum
mechanics and the molecular dynamics principle through calculations of the strain energy for
CNTs and graphite sheets subject to various types of loading. Elastic rod and plate theories
are employed to link the material properties of SWCNTs directly to the molecular mechanics
calculations.
Theoretical Foundation
First of all, t = 0.34 nm, the thickness of a grapheme sheet, is normally assumed for CNT
thickness. However, Yakonson et al. [32] concluded that the effective thickness of CNTs
should be taken to be t = 0.066 nm rather than t = 0.34 nm if the bending rigidity D = 0.85
eV and in-plane stiffness Et 360 J / m 2 are consistent with the classical shell bending
theory. This discussion of the wall thickness of CNT can also be found in Yakobson and
Avouris [33]. The definition of in-plane stiffness is thus defined here as Et (similar to the
definition in solid mechanics) to avoid argument over the value of the effective thickness of
CNTs.
The physical interpretation of the in-plane stiffness of an SWCNT is the rigidity of the
material to the axial loading that is subject to it. Elastic rod theory is employed to link the in-
plane stiffness to the strain energy stored in the structure. From the mechanics of the
materials, an elastic rod that is subject to axial uniform compression or tension can be simply
modeled or represented by a fictitious spring element, with the stiffness given as [34]
k AE / L , (1)
where A is the area of the cross section, E is the Young‘s modulus, and L is the length of
the elastic rod. For an SWCNT, A dt is set, where d and t are the medium diameter of
the SWCNT‘s cross section and wall thickness, respectively. Therefore, the relationship
between the in-plane stiffness of the SWCNT, Et , and the stiffness of the fictitious spring is
obtained by
k (L / d )
Et . (2)
However, it is also known that spring stiffness is normally viewed as the second
derivative of the strain energy restored in the spring with respect to the corresponding
elongation or compression, i.e., U . Hence, the in-plane stiffness of the SWCNT can be
directly obtained as follows in terms of the second derivative of the strain energy stored in the
SWCNT, subject to axial loading [35]:
U (L / d )
Et . (3)
Eq. (3) directly links the in-plane stiffness to the calculations of strain energy and the
corresponding second derivative via molecular dynamics.
Similarly, an elastic rod under torsion can be directly modeled by a rotary spring with the
stiffness k given by [34]
kr GI p / L , (4)
where I p is the polar moment of the inertia of the circular cross section, and G is the shear
modulus. It has been acknowledged that the thickness of CNTs is normally considered to be
very thin compared to their diameters. For example, the diameter of (8,8) armchair CNTs is d
= 1.085 nm, whereas the thickness of the SWCNT is used as 0.076 nm. Hence, the polar
moment of inertia for a thin circular rod can be approximately given by I p d 3t / 4 .
Similar to the spring that is subject to axial loading, the rotary spring stiffness, kr , is
equivalent to the second derivative of the strain energy with respect to the torsion angle in the
spring, or, equivalently, the strain energy stored in the CNTs, with respect to the rotation
angle applied to them, i.e., U r . Hence, the shear modulus of SWCNTs can be easily obtained
from [36]
4U r L / d 3
Gt . (5)
The limited estimates available on bending rigidity are almost all from studies of graphite
sheets [15, 37]. Finally, bending rigidity D is evaluated by applying point loading at the
center of a square graphite sheet with a length of L and the four edges clamped. Based on the
elastic plate theory, the relationship between the loading, P , and the deformation at the
loading position, , is governed by [38]
P D
. (6)
0.0056 L2
Because the work done by the point loading is completely transferred to the strain energy,
the plate is again modeled by a fictitious spring whose stiffness, P / , can be calculated
from the second derivative of strain energy U , as measured from the molecular mechanics
simulations. The expression for D can thus be given as [35]
D 0.0056 L2U . (7)
Eqs. (3), (5), and (7), by virtue of elastic theory, represent the relationship between the
mechanical properties of CNTs and the calculations of the second derivative of strain energy
via molecular dynamics. The measurements of these properties will thus be obtained through
molecular dynamics simulations of CNTs subject to axial loading, torsion, and graphite sheets
by point loading, as shown in the following sections.
Molecular Dynamics
Molecular dynamics is a form of computer simulation wherein atoms and molecules are
allowed to interact for a period of time under the known laws of physics, thus giving a view
of the motion of the atoms [39]. Molecular dynamics simulations are conducted via Materials
Studio, which was developed by Accelrys to study chemicals and materials, including crystal
structure and crystallization processes, polymer properties, catalysis, and structure-activity
relationships. In these simulations, the interatomic interactions are described by the
condensed-phased optimized molecular potential for atomistic simulation studies [40]. This is
the first ab initio force field that was parameterized and validated using condensed-phase
properties, and it has been proved to be applicable in describing the mechanical properties of
CNTs [35-36]. In molecular dynamics, the potential energy of a system can be expressed as
the sum of the valence (or bond), cross-terms, and non-bond interactions:
Etotal = Evalence + Ecrossterm + Enon-bond. (8)
The energy of valence, Evalence, is generally accounted for by terms that include bond
stretching, valence angle bending, dihedral angle torsion, and inversion. The cross terms,
Ecrossterm, account for such factors as the bond or angle distortions caused by nearby atoms to
accurately reproduce the experimental vibrational frequencies and, therefore, the dynamic
properties of molecules. The energy of interactions, Enon-bond, between non-bonded atoms is
primarily accounted for by the van der Waals effect. The molecular simulations are carried
out at a temperature of 1 K to avoid the thermal effect with an adiabatic process. The
molecular dynamics procedure involves the stepwise integration of Newton‘s equations. The
constant volume ensemble, also known as the microcanonical ensemble, is obtained by
solving the standard Newton equation without any temperature or pressure control. Energy is
thus conserved when this (adiabatic) ensemble is generated. The time step in all dynamics
simulations is 1 fs . Once every molecular dynamics process has finished, the configuration
of CNTs or graphite sheets is achieved through a minimizer processor that enables the atoms
in the structures to rotate and move in relation to one another to minimize the potential energy
so that an equilibrium state can be recognized. The simulations are run in parallel for the four
processors on a Sun workstation.
In molecular dynamics simulations of the compression motion of CNTs, the strain energy
0
is collected at every incremental displacement step 0.1 A . Once the strain energy at every
compression step is available, the second derivative of the strain energy with respect to the
compression can easily be obtained through a simple direct finite difference method. Then,
the in-plane stiffness of the SWCNT can be directly determined by Eq. (3). The shear
modulus of CNTs is investigated through a similar procedure. The strain energy at an
incremental step rotation angle, 1 degree, is collected to calculate the corresponding
second derivative and the shear modulus according to Eq. (5). In the estimate of the bending
rigidity of SWCNTs, the boundary of the graphite is updated with hydrogen to cease the
dangling bonds on the edges of the sheets. This is to make the simulation more stable. After
the initial minimization process, the four edges of the sheets are clamped, and the deformation
0
with incremental step 0.02 A at the loading is applied to find the strain energy data of the
plate sheet and the corresponding second derivative of the energy with respect to the
incremental step of the deformation. Then, the bending rigidity can be extrapolated via
Eq.(7).
Simulations
Table 1 lists the in-plane stiffness of achiral (8,0) and (8,8) and chiral (8,4) CNTs. The
diameters and unit cell lengths are 0.626nm and 0.426nm for zigzag (8,0) SWCNTs,
1.085 nm and 0.246 nm for armchair (8,8) SCNTs, and 0.829nm and 1.127 nm for chiral
(8,4) SWCNTs, respectively. Table 1 shows that the in-plane stiffness of (8,0) CNTs
2 2
increases from Et 354.01J / m to Et 375.18 J / m from the shorter
size, L 2.22nm , to the larger size, L 11.99nm . As for the armchair (8,8) CNTs, it can
again be seen that the in-plane stiffness increases with the increase of the tube length from
Et 354.08 J / m 2 for the (8,8) CNT with L 2.66nm to Et 377.58 J / m 2 for the
CNT with L 14.37nm .
Table 1. Molecular dynamics simulation calculations of in-plane stiffness and shear

modulus of SWCNTs
In-Plane stiffness Shear stiffness

(8,0) SWCNT (8,8) SWCNT (8,4) SWCNT (8,8) SWCNT
L (nm) Et (J/m^2) L (nm) Et (J/m^2) L (nm) Et (J/m^2) L (nm) Gt (J/m^2)
2.22 354.01 2.66 354.08 2.20 353.31 2.66 116.75
4.16 363.91 4.35 362.89 4.42 368.65 4.35 118.87
6.10 369.73 6.28 369.99 6.63 373.61 6.28 121.83
8.04 373.31 8.69 375.17 8.84 376.69 8.69 123.31
10.12 374.90 11.57 377.58 10.11 378.96 11.57 123.79
11.99 375.18 14.37 377.58 12.15 379.70
The asymptotic values of the stiffness for the achiral CNTs are all found for tubes larger
than 12 nm from the simulations. An obvious scale effect for the in-plane stiffness is secured
for tubes shorter than 10nm . The asymptotic value of the in-plane stiffness of (8,8) armchair
CNTs is found to be larger than that of (8,0) zigzag CNTs. A similar observation was also
indicated by Geng and Chang [17] via their nonlinear molecular mechanics model. The
measurements of the in-plane stiffness of chiral CNTs are more tedious to obtain than are
those of achiral CNTs, as the repeated units display themselves in a helical direction, rather
than in a straight longitudinal direction as in achiral tubes, thus leading to inaccurate
measurements of the tube lengths. Therefore, a more careful identification of the exact
locations of the repeated units is necessary for accurate measurements of the lengths. The
asymptotic value for the in-plane stiffness of the chiral CNTs is Et 379.70 J / m 2 , and the
lower value is found to be Et 353.31J / m for a tube with L 2.20nm . The noticeable
2
length dependence of the stiffness of the chiral tubes is clearly identified through the
simulations.
Our molecular simulations reveal that the length-dependent in-plane stiffness of CNTs is
in the range of 352 380 J / m 2 , which is close to the estimate of Yakobson et al. [32], 360
J / m 2 , based on data provided by Robertson et al. [41]. Gupta et al. [42] used a similar
method to evaluate the Young‘s modulus and found the in-plane stiffness to be about 420
J / m 2 , which is fairly close to our prediction. In addition, other estimates of the Young‘s
modulus based on experimental results [7,9-10] have also confirmed the range of this
mechanical property when the thickness of CNTs, t 0.34nm , is employed in the
calculations. Such good agreement in the predictions of in-plane stiffness also demonstrates
the effectiveness of the present method.
The in-plane shear modulus is investigated by applying torsion motions to the CNTs. The
last two columns of Table 1 show the variation of the stiffness versus the length of (8,8)
armchair CNTs. The shear modulus increases from Gt 116.75 J / m 2 for a CNT with
L 2.66nm to the asymptotic value Gt 123.79 J / m 2 for CNTs longer than 12nm . The
length-dependent shear modulus is also found in the simulations. Our results are in good
agreement with some of the existing predictions, such as those from lattice dynamics by
Popov et al. [43].
A square graphite sheet subject to point loading at its center is employed to measure the
bending rigidity of SWCNTs [35]. This value was found to be largely length-dependent for
the sheets less than 6.5 nm that we tested. The simulations [35] for bending rigidity are 1.47,
1.30, and 0.99 eV for square sheets with lengths of 6.42, 5.08, and 3.64 nm , respectively.
However, very consistent results are obtained for bending rigidity with a length of 8.48 nm ,
and the average of the values is 1.778 eV . A further measurement attempt was made on a
square plate with a length of 9.4 nm . The calculated bending rigidity is 1.783 eV , which is
very close to the estimate for the previous sheet. The minor difference in the estimates of the
two sheets of different sizes indicates the converged value for the bending rigidity of graphite
sheets larger than 8.48 nm in size. A value of 1.78 eV is thus recommended for the bending
rigidity of SWCNTs and graphite sheets. Such a prediction is in excellent agreement with the
estimate of Lucas et al. [21] and also quite close to that of Adams et al. [20], in which
quantum molecular dynamics methods were applied. The novelty of the method developed
here, however, is that it enables a quick and effective calculation of bending rigidity by
employing the simplicity of continuum mechanics while maintaining the accuracy of
molecular dynamics modeling.
Application of Nonlocal Theory
The length-dependence of in-plane stiffness was revealed in the previous section based
on molecular dynamics simulations. Nonlocal elasticity that employs an elastic rod subject to
axial compression is here introduced to investigate this length-dependent property [44].
According to the theory of nonlocal elasticity [27], the stress at a reference point x is
considered to be a function of the strain field at every point in the body. The basic equations
for a linear, homogeneous, isotropic, nonlocal elastic solid with zero body force are given by
ij , j 0, (9)
ij (x) (x x , )Cijkl kl ( x )dV ( x ) , x V, (10)
1
ij ui , j u j ,i , (11)
2
where C ijkl is the elastic module tensor of classical isotropic elasticity; ij and ij are the
stress and strain tensors, respectively; and ui is the displacement vector. (x x , ) is

the nonlocal modulus or attenuation function that incorporates into the constitutive equations
the nonlocal effects at the reference point x produced by the local strain at source x .
x x is the Euclidean distance, and e0 a / l is defined, where l is the external
characteristic length (e.g., crack length or wavelength). Parameter a describes the internal
characteristic length. For example, the length of a C-C bond is chosen for the analysis of
CNTs [30]. The parameter e0 was given as 0.39 by Eringen [27]. Wang [29] estimated the
scale coefficient as e0 a 2.1nm based on a vibration analysis via the nonlocal Timoshenko
beam theory. Wang and Hu [45] estimated the value to be around 0.28 using the strain
gradient method.
To obtain the length-dependent in-plane stiffness of an SWCNT subject to axial loading,
a nonlocal elastic rod theory is here established based on nonlocal elasticity [44]. Hooke‘s
law for a uni-axial stress state by nonlocal elasticity was proposed in reference [27] and is
given as
2 d2 ( x)
(x) e0 a E (x), (12)
dx 2
where x is the coordinate with its origin at the left end of the rod structure. Thus, the
nonlocal elastic rod theory can be derived as follows, considering the kinematics relation
du ( x)
( x) and the equilibrium equation q (x) 0.
dx x
d 2u ( x ) 2 d 2 q ( x)
E q( x) e0 a 0, (13)
dx 2 dx 2
where u (x) is the compression displacement of the elastic rod under compression, and
q(x) is the distributed axial force applied to the rod. At the limit where the effects of the
strains at points other than x are neglected, or e0 a 0 , the local or classical theory of
elasticity is obtained from the nonlocal elasticity. In our molecular simulations, the CNTs
were subject to compression displacement L on one clamped end. The general equation
(13) can then be re-written as follows, involving the Dirac Delta function and the Heaviside
function to model the molecular simulation process of a CNT with its left end clamped and its
right end subject to ―point‖ loading P AE L / L and based on the local elastic rod theory,
covering the domain from L x L.
d 2u ( x ) P P 2
E H x L H ( x L) e0 a x L ( x L) 0, (14)
dx 2 AL AL
where L is assumed to be very close to L to model a very small portion of the point
1, x x0
loading on the right end of the CNT. H ( x x0 ) and
0, otherwise
,x x0
( x x0 ) H ( x x0 ) are the Heaviside function and the Dirac Delta
0, otherwise
function, respectively, and the prime indicates the derivation of the function with respect to
x . Solving these mechanics problems based on the boundary conditions u (0) 0 and
u ( L) L leads to the displacement at the edge of the enforced domain x L as
2
PL e0 a
u 1 , (15)
AE L
which shows the equivalent size-dependent Young‘s modulus in the form of

2
e0 a
E E/ 1 . Figures 1-3 show the ratios of the in-plane stiffness of the above-
L
studied SWCNTs (see also Table 1) at every specific length to the corresponding asymptotic
in-plane stiffness, thus demonstrating this length-dependent mechanical property.
Figure 1 displays the ratio variation of the zigzag (8,0) SWCNTs from the molecular
simulations based on the data in Table 1 as a solid line by employing the asymptotic value
Et 375.18 J / m 2 . This asymptotic value is independent of CNT size and hence can be
viewed as the in-plane stiffness of the structure based on local elastic rod theory. The stiffness
ratio versus the length of the SWCNTs is also plotted with the different markers shown in the
figure at scale coefficients of e0 a 0.35nm , e0 a 0.65nm , and e0 a 0.95nm ,
respectively. The ratio is shown to be less than unit for shorter CNTs, but to approach unit at
larger sizes, thus indicating the obvious scale effect of length on the measurements of
stiffness. The molecular simulations show that the ratio at a smaller scale coefficient always
provides a higher value. Of the three scenarios, the variation of the ratio at e0 a 0.65nm
fits the molecular simulation results with the least difference. Overall, the comparison of the
ratio between the nonlocal theory and the molecular simulation results verifies the
applicability of the nonlocal elastic rod theory to the estimation of length-dependent stiffness.
A comparison of the measurements of stiffness based on the nonlocal theory with those
via molecular dynamics, which are shown by solid lines, for the armchair (8,8) SWCNTs is
illustrated in Figure 2. Similarly, the stiffness ratio is calculated by the ratio of the in-plane
2
stiffness of CNTs at every specific length to the asymptotic value, Et 377.58 J / m , as
shown in Table 1. The ratio via the nonlocal elastic rod theory is displayed by various
markers at e0 a 0.35nm , e0 a 0.75nm , and e0 a 1.05nm , respectively. The variation
of this ratio can again be observed from the figure, thus indicating a lower level of stiffness
for shorter SWCNTs and an asymptotic stiffness for longer CNTs. In addition, the result
obtained via the nonlocal elastic rod theory at e0 a 0.75nm is found to be best fitted to that
obtained via molecular simulations for the armchair SWCNTs.
The applicability of the nonlocal elastic rod theory is further verified by the
measurements of the in-plane stiffness of the chiral (8,4) SWCNTs. From the molecular
simulations, the asymptotic value for the in-plane stiffness of the chiral CNTs is
Et 379.70 J / m 2 .
0.95 Molecular
Stiffness ratio
simulations
e0a=0.35 nm
0.9
e0a=0.65 nm
0.85 e0a=0.95 nm
0.8
1 3 5 7 9 11 13
Tube length (nm)
Figure 1. In-plane stiffness ratio of zigzag (8,0) SWCNTs.
Molecular
0.95 simulations
Stiffness ratio
e0a=0.35 nm
0.9 e0a=0.75 nm
e0a=1.05 nm
0.85
0.8
1 3 5 7 9 11 13
Tube length (nm)
Figure 2. In-plane stiffness ratio of armchair (8,8) SWCNTs.
0.95 Molecular
Stiffness ratio
simulations
e0a=0.35 nm
0.9 e0a=0.70 nm
e0a=0.95 nm
0.85
0.8
1 3 5 7 9 11 13
Tube length (nm)
Figure 3. In-plane stiffness ratio of chiral (8,4) SWCNTs.

From the comparison between the stiffness ratio via the nonlocal elasticity and that via
the molecular simulations, the length-dependent stiffness for shorter CNTs can again be
examined. In addition, the scale coefficient e0 a 0.7nm is found to be a more adequate
value for the application of nonlocal elasticity in the estimation of the stiffness of chiral
CNTs.
MECHANICAL PROPERTIES OF DWCNTS

In this section, the calculations of the in-plane stiffness and shear modulus of DWCNTs
via the elastic rod theory and molecular simulations, which are similar to the calculations of
SWCNTs presented above, are briefly introduced [46]. The length-dependence of the
mechanical properties is explored, as is the van der Waals effect on the differences between
the material properties of DWCNTs and SWCNTs.
For a DWCNT, the cross section A di do t is set, where t is the wall thickness
of the DWCNT, and di and d o are the medium diameters of the cross section of the inner
and outer CNTs, respectively. According to Eq. (3), the in-plane stiffness of the
DWCNT, Et , is obtained by
UL
Et . (16)
di do
Similarly, the shear modulus of DWCNTs can easily be obtained as follows, based on Eq.
(5).
4U r L
Gt . (17)
d i3 d o3
In building a DWCNT, we choose the inner and outer constituent nanotubes and pick
zigzag (8,0) @ (17,0) and armchair (8,8) @ (13,13) DWCNTs with various lengths. The outer
tubes of the two zigzag and armchair DWCNTs are 1.331 nm and 1.763 nm, respectively.
Five armchair DWCNTs, with lengths of 2.657 nm, 4.351 nm, 6.286 nm, 8.697 nm, and
11.353 nm, respectively, are simulated. The CPU used for the largest calculation for the initial
minimization process of the (8,8) @ (13,13) DWCNT with a length of 11.353 nm and 4116
atoms is 1237.55 seconds.
The in-plane stiffness versus the length of the zigzag (8,0) @ (17,0) DWCNTs is plotted
in Figure 4 by the curve marked with triangle symbols. The stiffness increases from an initial
2 2
value of Et 344.92 J / m to an asymptotic value of Et 375.43 J / m for DWCNTs of
a shorter size, 2.092 nm, and those of a larger size, 11.717 nm. Figure 5 depicts the in-plane
stiffness versus the length of the armchair (8,8) @ (13,13) DWCNTs, showing an increased
variation in the stiffness from an initial value of Et 347.38 J / m 2 to an asymptotic value
2
of Et 376.72 J / m for DWCNTs of a shorter size, 2.657 nm, and those of a larger size,
11.353 nm.
375
In-plane stiffness (J/m^2)
360 (8,0) SWNTs
(8,0)@(17,0)
DWNTs
345
330
0 3.5 7 10.5 14
Length (nm)
Figure 4. Comparison of in-plane stiffness between zigzag DWCNTs and SWCNTs [46].
380
In-plane stiffness (J/m^2)
365 (8,8) SWNTs
(8,8)@(13,13)
DWNTs
350
335
1 4 7 10 13
Length (nm)
Figure 5. Comparison of in-plane stiffness between armchair DWCNTs and SWCNTs [46].
In both scenarios, an obvious scale effect on the in-plane stiffness is secured for tubes
shorter than 12 nm, which is similar to the calculations for SWCNTs. For comparison
purposes, the results of the stiffness of zigzag (8,0) and armchair (8,8) SWCNTs are also
shown by the curves marked with square symbols in Figs. 4-5. By comparing the stiffness of
the DWCNTs with that of the SWCNTs, it can clearly be seen that the stiffness of the former
is obviously less than that of the latter at shorter sizes. Such a difference is diminished only in
longer CNTs. An interpretation of this observation was made based on the van der Waals
effect that occurs between the two walls of the DWCNTs [46]. The interaction of the atoms
between these two different walls is the non-bond van der Waals effect based on the
molecular dynamics principle. The role of this effect is briefly illustrated here by the axial
compression process of a DWCNT. When a DWCNT is subject to compression, the
expansion of its outer wall is greater than that of its inner wall. This is because a larger
diameter leads to a larger gap between the two walls of a DWCNT, and this gap increases
during compression. Because of the widened gap between the two walls, attraction is initiated
due to the van der Walls effect. The force of this attraction means the outer wall is prone to
expand in the length direction, whereas the inner wall contracts further in the longitudinal
direction. With a smaller diameter, the magnitude of the reduction of the inner wall in the
longitudinal direction is greater than that of the extension of the outer wall, thus shortening
the DWCNT further as a whole. This decreasing trend during compression obviously
weakens the resistance of a DWCNT structure that is subject to compression, and hence leads
to the lower in-plane stiffness of the DWCNT. For a longer DWCNT placed under the same
compression as a shorter one, the change in the gap between the two walls becomes less
because of the smaller amount of strain in the radial direction. This results in less attraction
between the two walls due to the van der Walls effect. Therefore, the difference between the
material properties of DWCNTs and SWCNTs is negligible for larger sizes, which can be
seen from the convergence of the two curves in Figs. 4-5. The difference in the stiffness
diminishes for zigzag DWCNTs larger than 12 nm, although only for armchair DWCNTs
larger than 5 nm. A lower level of stiffness was also reported for MWCNTs from
experimental observations [7], and attributed to the occurrence of wavelike ripples.
Figure 6 shows the in-plane shear moduli of armchair (8,8) @ (13,13) DWCNTs, varying
from a value of Gt 115.11J / m 2 for those with a length of 2.657 nm to an asymptotic value
of Gt 123.83 J / m 2 for those with a length of 11.353 nm. It should again be noted that the
van der Waals effect leads to the smaller shear modulus of DWCNTs, as compared with
SWCNTs, and that the difference in this shear modulus diminishes for DWCNTs of larger
sizes. For armchair DWCNTs, the size effect on the difference between the shear modulus of
DWCNTs and SWCNTs becomes negligible when the length of the CNTs is greater than 6
nm.
124
Shear stiffness (J/m^2)
(8,8) SWNTs
120
(8,8)@(13,13)
DWNTs
116
112
1 4 7 10 13
Length (nm)
Figure 6. Comparison of shear modulus between armchair DWCNTs and SWCNTs [46].
CONCLUSION
A comprehensive continuum mechanics and molecular dynamics method for the
measurement of the mechanical properties of both SWCNTs and DWCNTs has been
introduced based on the research findings of the authors‘ group. This tight link between the
principles of the two methods enables accurate and efficient measurement because of the
simplicity of maintaining atomic structures in the continuum mechanics modeling and the
ability to do so via the molecular dynamics principle. The length-dependence of the
mechanical properties of both SWCNTs and DWCNTs is reported and investigated via the
nonlocal elasticity theory. The van der Waals effect on the difference between the material
properties of DWCNTs and SWCNTs is also examined, with the diminishment of that
difference at large sizes of CNTs revealed in the simulations.
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Lecture Material 8
ENGINEERED ELECTRICAL AND MECHANICAL

PROPERTIES OF CARBON NANOTUBE ADDED SI3N4
NANOCOMPOSITES
ABSTRACT
This research explores the use of a variety of nanoparticles to impart conductivity to
ceramic matrices. We have chosen some highly promising families of carbon materials:
multiwall carbon nanotubes (MWCNTs), singlewall carbon nanotubes (SWCNTs),
carbon black nanograins and graphite micrograins for use as fillers. In this book chapter,
we report the results about two types of carbon nanotubes. The MWCNTs and SWCNTs
were dispersed in silicon nitride matrix in different percentages high as 1-10wt%. A high
efficient attritor mill has also been applied for proper dispersion of MWCNTs in the
matrix. In order to get the full use of the benefits provided by carbon nanotubes (CNT) it
is crucial to retain CNT un-attacked in the composites and to optimize the interfacial
bonding between CNT and matrix. By conventional sintering techniques, which are
characterized by the requirement of extended holding at very high temperatures, the
destruction of CNT has been reported. In the present work the development of sintering
processes have been performed to consolidate and tailor the microstructure of MWCNTs
reinforced silicon nitride-based ceramic composites. The silicon nitride nanocomposites
systems retained the mechanical robustness of the original systems. Bending strength
high as 700 MPa was maintained and an electrical conductivity of 10 S/m was achieved
in the case of 3 wt% MWCNT addition. Electrically conductive silicon nitride ceramics
have also been realized by carbon black (in order of 1000 S/m) and graphite additions in
comparison. Examples of these systems, methods of fabrications, electrical percolation,
mechanical properties and potential uses will be discussed.
INTRODUCTION
Carbon nanotubes (CNT) are the most popular reinforcing materials in building
composite structures because they show advantageous mechanical, electrical, and thermal
properties [1-4]. Therefore, the application of CNTs was the main line of building up of the
ceramic composites. From electrical point of view ceramic materials are usually applied
typically as insulator products. During our research a Si3N4 ceramic was developed that had
extreme mechanical and thermal properties [5-7], but from electrical point of view it was
insulator. Our aim was to develop a hi-tech ceramic that has excellent electrically conductive
property as well. To reach this aim, small amount of electrically conductive additives were
mixed into insulator silicon nitride ceramic.
Retaining the CNTs un-attacked in the composites and to optimizing the interfacial
bonding between CNTs and matrix are the further requirements. In this way the toughening
effects characteristic to nano-scale fibre composites could be explored: crack bridging by
CNTs, CNT pullout on the fracture surfaces and crack deflection at the CNT/matrix interface
[8]. This study is focusing on the preparation processes that allow the tailoring of the
microstructure of carbon nanotube reinforced silicon nitride-based ceramic composites.
Experimental procedure has been conducted to effectively disperse the CNTs in the matrix.
Importance was given to temperature-pressure-holding time relation to preserve the carbon
nanotubes in composites and to avoid damaging during high temperature processing.
In the case of appropriate mixing of CNTs the fibrillated second phase interlaces the
material so the electrical current can flow through of it freely in a percolative way. For
comparison effect of the other carbon additives was also examined. To ensure the appropriate
construction two types of sintering method were tried out. Furthermore, a matrix base
material that contains electrically conductive part (aluminum nitride) was applied to improve
electrical conductivity. Carbon-ceramic composites with electrical conductivity in a wide
range have been successfully produced.
PRODUCTION OF CARBON NANOTUBE – SI3N4 NANOCOMPOSITES

The nanocomposites were prepared from the starting powder mixtures consisted of
90wt% α-Si3N4 (Ube, SN-ESP), 4wt% Al2O3 (Alcoa, A16) and 6wt% Y2O3 (H. C. Starck,
grade C) mixing ratios. Different amount of CNTs (multiwall carbon nanotubes MWCNTs,
produced as described elsewhere [9], single wall carbon nanotubes - SWCNTs) were added to
batches (1, 2 and 3 wt%).
The process of carbon nanotube - Si3N4 based nanocomposites is shown in Figure 1.
Figure 1. Schematic view of carbon nanotube - Si3N4 based nanocomposites preparation.
The powder mixtures together with the added CNTs were milled in distilled water in a
highly efficient attritor mill (Union Process Inc.) for five hours. As resulted from weight
measurements each batch contained approximately 1-3 wt% zirconia as contamination from
balls and discs. After milling surfactants (polyethylenglycol, PEG) were added to the powder
mixture. The batches were dried and sieved. Green samples were obtained by dry pressing at
220 MPa. Before sintering an oxidation was carried out at very low heating rates up to 400°C,
to eliminate the PEG from samples. Hot isostatic pressing was performed at 1700°C in high
purity nitrogen by a two-step sinter-HIP method using BN embedding powder. The heating
rate was not exceeding 25°C/min. The gas pressure (2 and 20 MPa) and holding time (0 to 3
hours) were also varied. The dimensions of the as-sintered specimens were 3.5 x 5 x 50 mm.
After sintering, the weight change of the samples was determined. All surfaces of the samples
were finely ground on a diamond wheel, and the edges were chamfered.
STRUCTURE OF CARBON NANOTUBE – SI3N4 NANOCOMPOSITES

The density of the sintered materials was measured by the Archimedes method. Phase
compositions were determined by Philips PW 1050 diffractometer and by EDS analysis
attached on Philips CM-20 transmission electron microscope (TEM). Morphology and
structure of the nanocomposites were studied by field emission scanning electron microscope
LEO 1540 XB, TEM Philips CM-20 and high resolution transmission electron microscope
(HREM) JEOL 3010. Results about the morphology of powder mixture and SWCNT samples
processed with attritor milling are presented in Figure 2.
Figure 2. SEM image of Si3N4 based nanocomposite with SWCNTs addition.
As it is shown in Figure 2 at higher magnification single carbon nanotubes are efficiently

dispersed in α-Si3N4, and sintering additives Al2O3 and Y2O3. In Figure 3a the scanning
electron image of MWCNTs sample is shown. Starting powder mixtures consisted of
crystalline Si3N4, Al2O3, Y2O3 grains and 3 wt% CNTs addition investigated by TEM is
shown in Figure 3b.
The CNTs are located mainly in the inter-granular places and they are well attached to
the silicon nitride grains. The proper separation and dispersion of carbon nanotubes proved to
be a difficult task of the composite preparation. For a better homogeneity, we applied the high
efficient attritor milling at 4000 rpm rotation speed and long time (5h). By applying this
technique advances have been made, but the general tendency of nanotubes (derived from the
high specific surface area), the strong adherence and linking behavior to each other could not
be totally suppressed. As can be observed, the CNTs in most of the cases are in groups, they
can be found as nano- or micrometer sized islands in the matrix after sintering (Figure 4a).
Figure 3. Structural investigation of a) MWCNTs addition, SEM image and b) Si 3N4 based
nanocomposite with MWCNTs addition, plan view TEM image.
Figure 4. a) Cross section TEM image of sintered carbon nanotube – Si3N4 Nan composites, b) HREM
image of Si3N4 grain and carbon nanotubes.
After milling by high efficient attritor mill, as revealed by TEM analysis (Figure 3b),
ceramic powder comprised of ~ 200 nm crystallites and dispersed CNTs. The EDS
measurements approved a primary composition (not shown). The nanotubes are dispersed in
the ceramix matrix. The nanotube radius is ~ 25 nm. The structure of sintered silicon nitride
based ceramic at 1700°C and at 20 MPa is shown in Figure 4. The sample consisted of ~ 300
÷ 400 nm nanocrystalline grains. The porosities were occured between some grains. The
CNTs were located in these porosities. The HREM investigation of β-Si3N4 / CNTs interface
showed that the nanotubes are oriented quite uniformly (Figure 4b).
X-ray diffractograms of sintered samples are presented in Figures 5. The main lines of -
Si3N4 (JCPDS-PDF 41-360), -Si3N4 (JCPDS-PDF 33-160) and ZrO1.96 (JCPDS-PDF 81-
1546) lines can be recognized in the case of samples sintered at 2 MPa.
The -Si3N4 to -Si3N4 phase transformation was completed at 20 MPa (Figure 5). The
X-ray diffractions of sintered samples are showing the main lines of -Si3N4 and two zirconia
phases (Figure 5).
Figure 5. XRD image of sintered Si3N4 based nanocomposite at 2MPa and 20 MPa.
MECHANICAL AND THERMOPHYSICAL PROPERTIES CARBON

NANOTUBE – SI3N4 NANOCOMPOSITES
Mechanical properties, the elastic modulus and four point bending strength for sintered
samples were determined by a bending test with spans of 40 and 20 mm.
Thermophysical properties of samples were tested with the LFA 457 from room
temperature to 900°C. The measured samples were disks with a diameter of approx. 10 mm
and thickness between 1.3 and 2.0 mm. The samples were coated with graphite on the front
and back surfaces in order to increase the absorption of flash light on the samples‘ front
surface and to increase the emissivity on the samples‘ back surface. The samples were
measured five times at each temperature. The specific heat was measured using the
comparative method. For this the system was calibrated with a ceramic standard (Pyroceram).
The mechanical observations are presented in Figure 6 and Figure 7. By increasing the
gas pressure to 20 MPa the similar level of densification and higher strength values can be
achieved for composites with MWCNT.
Figure 6. Modulus of elasticity as a function of the apparent density. a) 2MPa, b) 20MPa sintering
pressure.
For measurement of the thermal diffusivity, specific heat and bulk density at room
temperature were used to compute the thermal conductivity ( l ) by the following equation:
l = r × cp × a (1)
where r is the bulk density, cp is the specific heat and a is the thermal diffusivity of
nanocomposite.
Figure 7. Four point bending strength of composites as a function of the apparent density. a) 2MPa, b)
20MPa sintering pressure.
Figure 8 and Table 1 depict the thermophysical properties of nanocomposite sample with
3 wt% MWCNT (with 3,239g/cm3 density, 233GPa modulus of elasticity and 678MPa four
point bending strength) and a reference sample without carbon nanotube addition (with
3,392g/cm3 density, 254GPa modulus of elasticity and 732MPa four point bending strength).
The unit used in this work was equipped with a high temperature furnace capable of
operation from room temperature to 1100°C. Using th e sample changer three samples can be
measured at the same time. The sample holder for large samples allows sample diameters up
to 25.4 mm. The sample chamber is isolated from the heating element by a protective tube
allowing samples to be tested under vacuum or in an oxidizing, reducing or inert atmosphere.
The temperature rise on the back face of the sample is measured using an InSb detector.
Table 1. Summarized data of thermophysical properties. a) Carbon nanotubes – Si3N4

nanocomposite and b) Carbon nanotubes – Si3N4 without carbon nanotubes
Carbon nanotubes – Si3N4 nanocomposite Si3N4 nanocomposite without CNTs

Density 3.239 g/cm³, Thickness 1.490 mm Density 3.392 g/cm³, Thickness 1.350 mm
Thermal Specific Heat Thermal Thermal Specific Heat Thermal
T(°C) Diffusivity J/(g*K) Conductivity Diffusivity J/(g*K) Conductivity
mm²/s W/(m*K) mm²/s W/(m*K)
26 9.142 0.650 19.247 8.605 0.632 18.465
100 7.588 0.804 19.766 7.154 0.772 18.749
200 6.289 0.899 18.308 5.978 0.868 17.611
300 5.443 0.949 16.725 5.233 0.922 16.376
400 4.881 0.975 15.417 4.671 0.953 15.104
500 4.434 0.998 14.339 4.265 0.980 14.176
600 4.082 1.019 13.474 3.936 1.001 13.363
700 3.796 1.037 12.752 3.654 1.014 12.566
800 3.547 1.050 12.067 3.421 1.025 11.894
900 3.337 1.060 11.457 3.230 1.026 11.246
Figure 8. Thermophysical properties of silicon nitride based nanocomposites. a) carbon nanotubes –

Si3N4 nanocomposite and b) reference sample (silicon nitride nanocomposite without carbon
nanotubes).
Data acquisition and evaluation are accomplished using a comprehensive PC software

package. The data evaluation software allows 2- and 3-layer calculations as well as the
evaluation of the contact resistance. The data can also be corrected for finite pulse and heat
loss effects using any number of models. The instrument can be operated in the fully-
automatic or manual mode.
The specific heat increased with temperature as expected from the Debye theory. The
thermal diffusivity decreased over the entire temperature range. Typical for phonon
conductors is a maximum value in the thermal conductivity nearly at room temperature. This
trend can clearly be seen at both of samples. The thermal conductivity values in the case of
the sample with 3 wt% MWCNT are slightly higher than the values of the reference sample.
The standard deviation of 5 shots at each temperature is less than 1 %.
TRIBOLOGICAL PROPERTIES CARBON NANOTUBE – SI3N4

NANOCOMPOSITES
The tribological test has been performed on each sample using the CSM Tribometer
(TRB), which principle and specifications are shown in Figure 9. The test was prepared in air
atmosphere at 23°C and the humidity was 30%. The linear test mode with 6 mm amplitude
and 5 N normal load, 10 000 laps stop conditions, 5 Hz acquisition rate parameters were used
for all samples.
The TRB is suited to study the friction and wear behaviour of almost every solid state
material combination, with varying time, contact pressure, velocity, lubrication, temperature.
A flat or a sphere shaped static partner is loaded on to the test sample with a precisely known
force. The static partner, (a pin or a ball), is mounted on a stiff lever, designed as a frictionless
force transducer. As the disk is rotating, resulting frictional forces acting between the pin and
the disk are measured by very small deflections of the lever using an LVDT sensor.
Figure 9. Schematic view of CSM Tribometer.

Wear coefficients for both the pin and sample are calculated from the volume of material
lost during a specific friction run. The results of wear study is shown in Table 2. It was
observed that the Si3N4 ball was more damaged with MWCNTs addition nanocomposite than
with pure Si3N4 ceramic. As was observed from tribological measurements, nanocomposite
Si3N4 without carbon nanotubes shows a higher friction coefficient than carbon nanotube -
Si3N4 sample (Figure 10-12).The very big difference that could be shown in this test concerns
the sample wear rate: there was a much higher wear for carbon nanotube - Si3N4 than for
Si3N4 without MWCNTs; factor 10 between the both.
Table 2. Summarized data of the wear study for a) Si3N4 and b) carbon nanotube - Si3N4
nanocomposite
Sample Worn cap Worn track Ball wear rate Sample wear rate
diameter (μm) section (μm2) (mm3/N/m) (mm3/N/m)
Si3N4 726 302 7.61E-06 3.02E-06
CNTs - Si3N4 899 3345 1.79E-05 3.34E-05
Figure 10. Friction coefficient measurements of Si3N4 and carbon nanotube - Si3N4 nanocomposites.
Figure 11. Friction wear test images of reference Si3N4. a) low resolution image, b) high resolution
image.
Figure 12. Friction wear test images of carbon nanotube - Si3N4 nanocomposite. a) low resolution
image, b) high resolution image.
Figure 13 shows the calculation of the worn track section, which was done with a
profilometer Taylor Hobson.
Figure 13. Calculation of the worn track section by profilometer. a) Si3N4, b) carbon nanotube - Si3N4
nanocomposite.
ELECTRICAL PROPERTIES CARBON NANOTUBE – SI3N4

NANOCOMPOSITES
For the electrical measurements electrical contacts (4 on each sample) had to be created
on the surface of the composites. Multilayer contact was developed that consists of 4 layers
and can be seen in Figure 14. The first thin gold layer was vapored on the appropriate part of
surface of the sample. The gold layer was covered by conductive glue as second layer that
contains silver grains. After drying the current feeds (third layer) were fixed on this layer by
ordinary soldering technique (fourth layer). The created contacts should have low resistance
in order to not to overload the measurement system and have good adhesion for fix the
current feeders during the tests.
Figure 14. Four layer multilayer contacts for electrical experiments.
Figure 15. Schematic four wire measurements technique to determine the specific conductivity of the
composites.
After the contacting the DC resistivity measurements was started. To determine the pure
resistivity of the ceramic composites during the test four wires resistance measurement had to
be applied (Figure 15.). Our samples with current feeders have many resistivity but with four
wire technique the pure resistivity of the ceramic between the two inner contacts can be
determined without the resistivity of the contacts.
The basis of the 4 wire resistance measurement is that composite was excited through the
two outside contacts by stable DC current. During the exciting the voltage drop between the
two inner contacts was detected. The resistivity of the section between the two inner contacts
was calculated on the basis of Ohm's law from the DC current and voltage drop and the
device (Agilent 34411A) was done automatically. For choosing the measurement range of the
equipment resistance of the contacts has to be considered. This resistance was determined on
the basis of a simply two points method. Specific conductivity of the samples was calculated
from the geometry of the samples (the part of the samples between the two inner contacts)
and the resistivity.
In the case of some composites impedance spectroscopy measurements were fulfilled
(see Figure 16).
Figure 16. Schematic draw of the lock in technique for AC impedance measurements.
DC CONDUCTIVITY OF CARBON NANOTUBE – SI3N4

NANOCOMPOSITES
During the impedance spectroscopy the absolute impedance of the composites was
determined in a scanned frequency range. For this phase sensitive Lock in measurement
system was adopted. As answer to voltage bias of the signal generator current signal was
detected separated in two components as in-phase and quadrature (respect to excitation
phase). After measuring the two components of the current the sample complex impedance
was evaluated according to Ohm‘s law. The measuring frequency range was from 100 Hz to
100 kHz limited by the Lock-in detector upper limit.
The examined composites can be grouped into insulator and conductor types as resulted
from four points DC resistance measurements. Samples that overloaded our measurement
system (10MΩ measurement limit) can be considered as insulator because their resistance
was not measurable. These samples were: basic ceramics without any type of additives,
samples that contains 1wt% and 5 wt% MWCNTs (Figure 17).
Figure 17. SEM images of insulator ceramics (a) base ceramic and (b) Si 3N4 with 5wt% MWCNT.
The other group of composites is considering as conductor (Figure 18.) because of the
contact and grouping of the additives. As the electrical conduction takes place through the
paths that are made by linked conductive phases these materials behave like percolative
conductors. The percolation threshold is observed in composites with 3wt% for MWCNT.
Figure 18. Specific conductivity of the composites as a function of MWCNT addition and sintering
techniques.
MWCNT addition a larger conductivity range and higher conductivity were detected in
the case of 3wt% addition. In the case of GPS sintering higher conductivity was obtained than
in the case of HIP. The reason of the differences (in electrical aspects) between the two types
of sintering technique is the different grain structure after the heat treatment process. In case
of HIP technique the higher sintering pressure and the holding time performed the β-Si3N4
grains (Figure 19.) opposite of GPS structure that contains α-Si3N4 grains (Figure 20.).
Figure 19. X-ray diffractogram of well conductive composites (Si3N4 with 5wt% MWCNT) and
insulator base material (reference) prepared by HIP sintering.
Figure 20. X-ray diffractogram of well conductive composites and insulator base material prepared by
GPS sintering.
The bigger grain sized β-Si3N4 decreases the spaces for the percolation network (Figure
21). Furthermore, it can generate the crumbling and decomposing of the conductive clusters,
so further decreasing can take place in the conductivity.
Figure 21. SEM images of 5 wt% MWCNT conductor composites produced by (a) GPS sintering, (b)
HIP sintering techniques.
AC IMPEDANCE OF CARBON NANOTUBE – SI3N4 NANOCOMPOSITES

On the insulator sample – that overloaded the DC measurements system – AC impedance
spectroscopy measurements were done. This sample is the nanocomposite with 1 wt%
MWCNT additions. In this nanocomposite, the conductor particles could not form percolation
tracks for current paths. The AC impedance plots of high DC resistance samples are displayed
in Figure 22. It shows the imaginary parts of complex impedance versus real parts. The
imaginary part that represents capacitive character is more dominant over the frequency
range. It means that the conductor parts that do not have connecting segments form capacitor
and this results that the sample works as dielectric insulator material.
Figure 22. Impedance curves of different carbon added Si 3N4 ceramics up to 100 kHz frequency.
In Figure 23 absolute impedance versus frequency is displayed. Except 5 wt% graphite

sample all the curves run with -1 slope in a log-log plot. The impedance has 1 / ω dependence
on frequency. This is a typical capacitive impedance feature (Rc = - i / C ω).
Figure 23. Absolute impedance versus frequency in case of composites that are under the percolation
network.
Figure 24. Specific conductivity range of conductive nanocomposites.

The insulator samples changed their high DC impedance due to frequency variation of
capacitive impedance (Rc) which is characteristic to dielectric materials. Increasing of the
MWCNT additives does not mean higher electrical conductivity because of the nodulation of
MWCNT (Figure 24). Furthermore increasing the MWCNTs additives the well-grown β-
Si3N4 derogates the connection between the MWCNT nodules so the conductivity of the main
composite material decreases. The reason of it is that β-Si3N4 grain grows during the
sintering.
Adding the conductor AlN did not cause the improvement of percolation limit in the
examined compositions, but increased the electrical conductivity of the originally conductor
composites forming more electrical connections between the conductive parts.
INFRARED EXAMINATION OF CARBON NANOTUBE – SI3N4

NANOCOMPOSITES
To do the infrared measurements voltage was connected (Figure 25) on the composite
and the electrical current could flow through the material by percolation way. The material
was heated with the Joule heat of the current. 1 kHz frequency sine signal was used for the
excitation of the composites. The signal was amplified step by step for the sake of the heat
starting up in the sample increases step by step. The stepping of the voltage was continued till
the temperature on the sample became ideal 100 °C for the infrared camera till the maximum
excitation voltage 80 V was reached. During heating of the samples the rate of the applied
excitation was different on the samples because of their different electrical conductivity. The
CEDIP infrared measurement system that can be found in the University of Leoben, Institute
of Structure- and Function Ceramic was used.
Figure 25. Shematic draw of infrared measurement system.
EMISSIVITY OF CARBON NANOTUBE – SI3N4 NANOCOMPOSITES

During the infrared investigations at first the emissivity of the surface of the samples had
to be determined. The emissivity of the surface is needed to know the real temperature of the
surface that can be evaluated during the calibration on each measurement. The emissivity of
the composites was determined by a comparing method. Half of one side of the brick-shaped
sample was covered with a layer that had known emissivity in the examination temperature
range. This emissivity of the layer was 0.95. The result of the exiting samples were heated
and the surface with and without layer showed the same temperature on its equal area (Figure
26). Through the comparison temperatures of the two areas were approximated to each other
and the CEDIP program determined the emissivity of the real surface of the composite.
The emissivity of the surface was determined on more surface points (at the same points
of the lengthwise temperature gradient). The calculated emissivities can be seen in the Table
3. According to the results the Si3N4 carbon-ceramic composites have approximately similar
emissivity in the examination temperature (0.95 – 0.99).
Figure 26. Methods of the emissivity determination, area 1.) covered with layer, area 2.) real surface.
The differences of the carbon additions could not be detected. As the determination of the
emissivity in case of all the samples happened with the same temperature as the temperature
profiles were detected the temperature dependence of the emissivity was not considered in the
following.
Table 3. Surface emissivities of the composites that contain different carbon additions
Sample Carbon addition Specific conductivity Exciting voltage [V] Emissivitiy

sintering [wt%] [S/m]
Si3N4 (GPS) 3 MWCNT 26.6 40 0.96
Si3N4 (HIP) 3 MWCNT 15.9 40 0.97
Si3N4 (HIP) 5 MWCNT 3.56 80 0.98
HEAT CONDUCTIVITY OF CARBON NANOTUBE – SI3N4

NANOCOMPOSITES
A heat conductivity model was adopted for the determination of the heat conductivity of
the composites. This model is a one-dimension heat conductivity model with boundary
condition of two parallel infinitesimal planes with heat source inside [10]. For the evaluations
the temperature gradients of the surface that were detected by the infrared method were used.
Figure 27. One-dimension heat conductivity model by plane walls with heat source.
In the volume of the plane walls the heat source is distributed steadily. The dimensions of
directions deviated from the x direction are so large or the heat gradient is so small in this
direction that he heat conductivity can be considered as one-dimension. During the
examination it was considered that the heat conductivity does not change in the small
temperature range that can be observed on the detected heat profile. This model can be
applied well in the case of electrical conductive materials when some current flow through
producing Joule heat heats the material. The heat conductivity of the model is described by
the following differential equation on the geometry is drafted in the Figure 27:
d2T / dx2 + q / k = 0 (2)

where T is temperature, q is heat flux, k is heat conductivity. For the evaluation of the heat
conductivity the resolution of differential equation of the reviewed model Eq. (2) was applied.
In our case:
ΔT = q · L2 / 2 · k (3)
where L is the length of the examined profile. The heat flux was generated by the Joule
heat emitted in the sample can be determined by Eq. 4 and Eq. 5 calculation:
q = U 2 / ρ · L2 (4)
q = I2 · ρ / A2 (5)
where ρ is specific impedance, U is voltage, I is current, A iscross-section of the sample. The

dimensions and the specific impedance of the samples were given from former measurements
and the generated voltage and later the current through were measured. The model was solved
on a symmetrically arranged parabolic heat profile. The heat profile of our samples did not
symmetrical because of the current leads had different contact resistance and they insured
different heat transfer (Figure 28). In stationer state the border conditions are stable, on the
temperature profile of the section using for the evaluation can be chosen anywhere on our
asymmetrical profile. On this section two segments were found with same temperature (TW)
on the two sides of the maximum (T0).
During the determination of the heat conductivities it was observed that in case of using
the voltage generator excitation the heat conductivity that was determined with the
consideration of the sample resistance in certain cases differs from the exceptions according
to the literacy [11]. The reason of this is that our calculation does not consider the contact
resistance between the sample and the sample holder.
Figure 28. Temperature profile along the sample.

In the case of good insulator samples (graphite or MWCNT additives), where the contact
resistance could be higher than the impedance of the sample the evaluation of the
measurement is not possible (Table 4).
Table 4. Heat conductivity determined by infrared thermography
Excition Carbon phase Specific Conductivity Voltage Current Heat conductivity

[wt%] [S/m] [V] [A] [W/m·K]
Voltage 10 graphite 0.41 80 - 32.5
Voltage 10 graphite 0.52 80 - 37.85
Current 5 carbon black 139 - 0.27 59.68
In the case of current generator drive of small impedance sample the contact resistance is
not in the Eq. 5, its effect has no part in the evaluation so that can be applied on any kind of
electrically conductive sample. For checking a measurement was done using current generator
excitation to determine the heat conductivity of a low resistance sample that can not be
evaluated in the case of voltage generator excitation.
Considering the literacy data [11] the heat conductivity of our material moves between
wide borders. It will be even wider because different porosity material structure become
during the sintering. On the basis of the calculation the obtained heat conductivity values are
in the range of the literacy data.
CONCLUSIONS
Optimisation of the manufacturing processes has been performed in order to thoroughly
disperse the carbon nanotube in matrix, to assure a good nanotube-silicon nitride contact and
to keep intact the nanotubes during high temperature processing.
The grouping of the strength and modulus values as a function of apparent density was
observed in the case of samples, sintered at higher pressure, as compared to the lower
pressure sintered samples. At 20MPa, the highest densification grade, modulus and strength
values were found.
The addition of carbon parts in the ceramics drastically changes the electrical properties
of composites. Using different type and concentration of carbon additives excellent
conductive materials can be produced from the insulators.
In our composites the electrical conduction intervenes in percolative way.
MWCNT additives electrical conduction appeared at 3wt%. Increasing of the MWCNT
additives does not mean higher electrical conductivity because of the nodulation of MWCNT
fibers. Furthermore increasing the MWCNTs additives the well-grown β-Si3N4 derogates the
connection between the MWCNT nodules so the conductivity of the main composite material
decreases. The reason of it is that β-Si3N4 grain grows during the sintering.
Composites that are prepared on low pressure by GPS sintering have better conductivity
than the high pressured HIP sintering because of the more β-Si3N4 formation. The grain size
of β-Si3N4 is effectively bigger than the grain size of α-Si3N4 therefore blocks the conductive
network formation.
Adding the conductor AlN did not cause the improvement of percolation limit in the
examined compositions, but increased the electrical conductivity of the originally conductor
composites forming more electrical connections between the conductive parts.
The results of the infrared analysis showed approximately same emissivity values in the
range of examination temperatures. The emissivities did not depend the mixing ratio and the
type of carbon additions and the sintering methods.
During the processing of the infra images an evaluation method that can be traced back to
a one-dimension heat conductivity problem was worked out. With this method the heat
conductivity of the stationer state samples was determined.
The heat conductivity data are in the same range that can be seen in the literacy data. It
can also be observed that the heat conductivity changes according to the quality and the type
of the carbon additives just as the electrical conductivity. The growth rate is different from the
case of electrical conductivity while the electrical conductivity changes with order of
magnitudes because of the percolation mechanism during the insulator-conductor transition,
the heat conductivity changes according to a mixing rule because the percolation of the
additives is unnecessary here and the additives give its properties a composite phase.
ACKNOWLEDGEMENTS
Author would like to thank the Nanobiotechnology, Florida State University and
Nanoinnovations Inc. Florida, for their help with the infrared measurements.
Authors would like to thank Dr Riaz Khan for helpful discussions for help in sample
preparation and for SEM.
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Lecture Material 9
FLUORINATED CARBON NANOTUBES:

STATE OF THE ART, TRENDS AND ADVANCED
CONCEPTS
ABSTRACT
The present contribution details, in an as exhaustive as possible way, the
fundamental knowledge acquired on fluorinated carbon nanotubes (CNT), also termed
fluorotubes. Since the discrete pioneering articles published in the field a bit more than
10 years ago, around 70 additional references have now appeared which bear witness
tothe dynamism of this emergent part of the chemistry of nanotubes. As a matter of fact,
fluorination stands as the starting point for a great part of the modifications performed on
CNT, rendering fluorotubes fundamental intermediates for the integration of CNT in the
nanotechnology processes. Many synthesis routes use fluorotubes as precursors in view
of the subsequent chemical derivatization of CNT, for instance. In parallel, several
attempts of practical developments based on fluorotubes have also lately appeared
throughout the literature, covering the tribology, nanocomposites, or electrochemistry
sectors, which outline the potential interest of such fluorinated nanostructures. The first
part of this chapter compiles the main knowledge reported to date on the subject
throughout a still reasonable but increasingly abundant body of literature. The
physicochemical properties and special characteristics of the C-F chemical bond in
fluorotubes are critically analyzed in a second part, and the last section is devoted to
some recent tentative applications and future concepts relating to the fluorination of
carbon nanotubes. The present review is essentially depicted from the chemistry of
materials standpoint. Some of the concepts illustrated throughout the text are enriched by
a few works of unpublished data from our group.
FLUOROTUBES - GENERAL ASPECTS

Introduction
The covalent addition derivatives of carbon nanotubes are much less diversified than
those issuing from the parent fullerenes molecules. Owing to the macromolecular topology of
the carbon edifice, a ―rough‖ chemistry consisting of the random fixation of functionalities to
the carbon substrate, in a more or less efficient and controlled way and often inducing defects
in the continuity of the carbon frame, has succeeded to the fine methodologies employed to
functionalize fullerenes. Nanotubes functionalization, most of the time, results from an
oxidation of the carbon substrate in harsh conditions, from methods tested and validated
decades ago on graphite, carbon fibers, or amorphous to semi-crystalline forms of carbon.

Hence, the creation of the basic carbonyl, carboxyl, lactone or hydroxyl functions can be
obtained upon treatment of carbon nanotubes with hot nitric acid or with hot aqueous
solutions containing the permanganate, peroxodisulfate or perchlorate anions, or still with
hydrogen peroxide. If oxygenation turns out to be rather easy, the halogenation of carbon
nanotubes is more difficult to obtain, in spite of the strongly oxidizing character of halogen
atoms, and only fluorine is known to directly and stably fix to such a carbon matrix. The
fluorinated derivatives of carbon nanotubes, the so-called ―f luorotubes‖, have since proven to
be of fundamental importance from both the chemistry of nanotubes and materials
standpoints.
The present synthetic approach describes and analyzes the widely diversified classes of
materials that arise upon fluorine addition to the different tubular carbon allotropes. The first
part reviews the main results reported hitherto concerning the synthesis, structural aspects and
physico-chemical properties of both single-walled and multiwalled fluorotubes. The
characteristics of the C-F chemical bond in fluorotubes will be analyzed in a second part and
the last section of this chapter will be devoted to a prospective overview of the potential
valorization of fluorotubes. Further research in the present field would preliminary require a
perfect knowledge of the unexpectedly complex nature of the fluorinated derivatives of
carbon nanotubes, which is far from being acquired hitherto, as will be shown from a critical
analysis of some previously published data.
Synthesis of Fluorotubes
Single wall nanotubes: The fluorination of single wall carbon nanotubes (SWNTs) is
usually carried out by direct reaction with an atmosphere of gaseous fluorine, over
temperatures ranging from 50 to around 300 °C [1-12]. The latter value is often recognized as
the critical threshold before significant degradation of the tubes begins to manifest. Thus,
beyond this limit, the introduction of surface defects [1,3] and even combustion may become
important, in which case gaseous fluorocarbons will form. The whole set of results reported
converge in the sense that, at constant reaction time, the F/C stoichiometric ratio increases in
parallel with the synthesis temperature, and the maximum functionalization rate seems to be
around 50 to 60%, i.e. C≈2F.
Extremely efficient plasma fluorination techniques have been recently developed [13-16],
allowing access within a few minutes to important fluorination levels. The most common
fluorinating agents are CF4 or SF6. Hence, such processes remain soft, limiting destruction of
the tubular macromolecular edifices, and avoid the use of molecular fluorine, which is
expensive and dangerous.
Multiwall nanotubes: Morphological differences between SWNTs and multiwall carbon
nanotubes (MWNTs) are at the origin of a specific reactivity of each one in regard to fluorine.
Hence, MWNTs appear more resistant than SWNTs regarding fluorination at high
temperature, owing to their higher outer diameters which reduce the strain associated with the
local curvature of the network. Their complete fluorination with an F/C ratio close to 1 has
been obtained near 500 °C [17-19], at the expense of the creation of many defects and of a
more or less pronounced structural disorder. Note that, in the present case, an insufficient
fluorination time will yield surface fluorinated tubes only [19-21]. The use of powerful
fluorinating agents such as XeF2 has been attempted [22], but similarly results in a partial
fluorination, restricted to the outer surface. In parallel, the fluorination of MWNTs can also
be performed at low temperature [17,18], following the catalytic process developed several
decades ago for graphite [23] and based on the use of gaseous HF. Experimental details in the
matter remain scarce, however, since only a few papers on the subject have been published so
far [17-22].
Structure and Fluorination Mechanism
Single wall nanotubes: SWNTs represent the macromolecular form of fullerenes

molecules. In spite of this direct filiation, the fluorination at high temperature of SWNTs
yields covalent addition derivatives resulting from independent and nonhomogeneous
addition patterns, in contrast to fullerenes molecules in which case regioselectivity is
excellent [24]. The direct observation by scanning tunneling microscopy (STM) [2] of the
surface of single-walled fluorotubes (F-SWNTs) indeed showed alternating fluorinated and
non-fluorinated sections along the tube axis. The nanotexture is thus seemingly made of
individual circular ribbons of F addends wrapping the circumference of each tube.
XPS is a major technique for surface analysis and consequently happens to be suited
perfectly to the study of the surface composition of functionalized SWNTs, providing fine
details about fluorine addition to SWNTs. Thus, whatever the final stoichiometry of the
product and synthesis route are used, the C1s XPS spectrum corresponding to F-SWNTs
[6,7,10,11,15] typically consists of a first feature centred near 284.5 eV (see Figure 1), similar
to the one present in the C1s spectra of raw SWNTs and is therefore, related to bare carbon
atoms. It is common knowledge that the C1s core level energy undergoes a substantial primary
shift in the presence of a strong electron-withdrawing element. Accordingly, with reference to
standard tables, the C1s signal related to carbon atoms attached to one fluorine atom usually
appears in the 287-289 eV region, in which a corresponding feature is observed for
fluorotubes. It is also well established that the existence of carbon atoms not directly linked to
a fluorine atom but having fluorinated nearest neighbors gives birth to a secondary shift of the
C1s energy, which, in the case of fluorotubes, splits the first ―gr aphitic‖ signal into a second
component lying at a slightly higher binding energy (≈ 286 eV). Complementarily, weak
shoulders over the 291-293 eV energy range are always present and arise from >CF2 and –
CF3 groups that forms on edges or at local defect sites. At last, XPS almost systematically
detects a few atomic percentages of oxygen in both pristine and fluorinated carbon materials,
respectively.
Figure 1. Typical C1s XPS spectrum of F-SWNTs (here: CF0.5, obtained after 2 hours under a molecular
fluorine gas flow at 300 °C).
The invariable nature of the XPS spectra of F-SWNTs also reflects their texture at the
nanometer scale and provides a compelling evidence of an inhomogeneous fluorine addition
pattern, besides the direct view of the phenomenon by scanning tunnelling microscopy
previously mentioned. Indeed, a uniform halogenation pattern with an F/C ratio close to ½
would constrain each carbon atom to have at least one fluorinated neighbor in its immediate
surrounding, suppressing the photoelectron peak assigned to bare carbon atoms. The
systematic persistence of the latter then necessarily implies a non homogeneous distribution
of fluorine atoms along the longitudinal axis of a tube and the presence of fluorine depleted
zones alternating with fluorinated sections.
The former experimental STM observation has seemed to provide support for a
circumferential exo-addition pattern as being the most probable architecture. Different regular
exo-addition pathways may allow to reach the apparently optimal experimental C2F
composition, but the conclusions of theoretical approaches on the subject [2,25-29],
performed most of the time on virtual fragments of fluorotubes, exhibit too much divergence
to shed light on the point here addressed, the results being highly dependent on the spatial
extension of the system, its initial helicity/diameter parameters, the fluorination rate
introduced and on the level of approximation. Somehow, at constant parametrization except
fluorine location, the modest variations in energy from one model to another should partly
reveal the origin of the overall random addition to SWNTs. Some experimental evidence in
favor of 1-2 addition instead of a more exotic 1-4 pattern have been seemingly obtained
throughout measurement of the 13C NMR chemical shift of F-SWNTs [30]. From the crystal
chemistry standpoint, a high temperature fluorination process does not necessarily imply
destruction of the bundle configuration, provided the latter was initially present within the
pristine tubes sample, as shown on the following micrograph. The bi-dimensional cell
parameter characterizing the compact triangular arrangement of the tubes becomes then
notably expanded upon fluorine addition [7], independently confirming the location of some
fluorine addends on the outer side of each tube.
Figure 2. TEM observation of SWNTs bundles fluorinated at 300 °C for 2h.
Double-walled carbon nanotubes consist of two nested cylinders. They represent the first
step toward MWNTs. Their fluorination has been successfully achieved at room temperature
via the use of a fluorinating agent [31,32], which represents much milder conditions than
heating under molecular fluorine, but some authors reported that direct fluorination at 200 °C
does not break the double-layered morphology [33]. Overall, their apparent fluorination
pattern somewhat recalls the one of SWNTs, consisting of surface addends grafted to the
outermost sidewall, but interestingly the core carbon shell remains preserved. Indeed, the
radial breathing modes characterizing the Raman signatures of the inner shell are kept
unchanged upon fluorination, indicating that addition is effective at the level of the outermost
framework only. The evolution of optical absorption spectra upon fluorination reinforces the
former conclusion. These findings offer a soft transition toward the MWNTs case.
Multiwall nanotubes: In comparison to SWNTs, the reduced curvature within MWNTs
diminishes reactivity, and the conditions required for fluorinating the latter approach those
required to fluorinate their flat homologue graphite. Hence, the formation of covalent fluoro-
derivatives of MWNTs (F-MWNTs) necessitates significant thermal activation and fluorine
atoms tend to graft to the sidewalls of multiwalled nanotubes from 300 °C, near the surface
first. Higher temperatures (500-550 °C) can lead to the bulk fluorination of MWNTs [17-19].
The resulting fluorocarbon compounds still exhibit wiry imprints after undergoing such harsh
conditions. Their overall structure consists of a superposition of more or less fragmented
fluorinated shells, some examples of which are displayed on figures 3 and 4. XRD usually
shows still reasonably resolved (00l)-type Bragg peaks (see example, Figure 3) which reflect
some degree of coherence in the stacking of the fluorinated shells. Their average separation
becomes close to that characterizing the interlayer distance in the graphite fluoride (CF)n,
obtained at 600 °C. Note that the final morphology turns out to strongly depend on the nature
of the precursor MWNTs (Figure 3 and 4): the higher the rate of sidewall discontinuities in
the pristine material is, the higher the disorder in the final fluorinated compound is.
Accordingly, a well-ordered succession of fluorinated domains along both axial and radial
directions can be obtained only by starting from a well organized MWNT. This has been
tentatively explained through the ― zipper‖ model [19], in which the fluorination mechanism is
chronologically decomposed according to the following steps: i) Fluorination of the
outermost sidewall, accompanied by radial inflation of the latter following change in the C/C
bond order ii) Diffusion of F in the increased interlayer space with the underlying sidewall
made possible, resulting in the attack of the latter by fluorine, on edge first iii) Radial
inflation of the second outermost sidewall upon fluorination, inducing progressive cracking of
the above fluorinated shell… and so on. Some authors consequently mentioned [21] that
delamination of the outer fluorinated shells could occur in the course of progression of the
fluorinated front.
Globally, essential divergences arise when comparing the high temperature fluorination
of MWNTs and SWNTs. The maximum stoichiometry that is usually reported for F-SWNTs
is close to F/C ≈ 0.5, whereas fluorination to saturation of the carbon shells occurs in the case
of F-MWNTs, without degradation of the substrate. In the latter case, according to XRD, the
microstructure locally looks like that of the graphite-derived phase poly-monocarbon
monofluoride (CF)n, indicating that more or less ordered rolled perfluorographene sheets
form. This consequently implies that both sides of the carbon shells must be fluorinated
according to a regularly alternating inner/outer addition pattern. Note that partial fluorination
rates, i.e. x<1 in the CFx formula, can also be obtained upon fluorination of MWNTs, but
must be interpreted as the superposition of a set of completely fluorinated layers over a set of
non-fluorinated carbon layers [19,20] (see an example Figure 4). Saturation with fluorine at
the level of a layer is rigorously confirmed by XPS measurements performed on such phases
[21], which do not exhibit the photoelectron peak typical of bare carbon atoms having
fluorinated first neighbors (see peak assignment in Figure 1). In sharp contrast with MWNTs,
the fluorination of SWNTs seems to take place on one side only, according to the current
opinion, though this point needs to be addressed in more details in the following.
In parallel, and adding further difference between the respective fluorination processes of
SWNTs and MWNTs, multiwalled fluorinated products differing from the former covalent
addition derivatives have also been observed.
Two independent studies indeed showed [17,18] that the catalytically assisted low
temperature fluorination of MWNTs gives birth to a surface fluorine intercalation
phenomenon. It is characterized by an increase of the average intershell distance, while the
initial stacking sequence seems preserved, as directly observed by TEM.
This specific behavior is made possible when the topology of the carbon network is
changed from single to multi-walled, creating an interlayer Van der Waals gap that affords
accommodation of guest-species. Such a scenario requires some wall disjunctions to be
initially present or to be created in the course of the fluorination process, however, so that to
allow local inflation of the structure upon intercalation of fluorine atoms [34]. Though the
intercalation phenomenon remains limited to the most external shells in the present case, the
formation of stages was even inferred by Nakajima et al. [17]. Indeed, the related X-ray
signatures recorded presented a good analogy with those of the homologous phases obtained
from graphite. Beyond 60 °C, the latter authors gave evidence for a quantitative diminution of
the intercalation phenomenon, first evidenced by an increase of the stage number, and by
further limitation of insertion to areas closer and closer to the outer surface above 100 °C.
MWNTs - 3h fluorination
MWNTs - Raw
8 10 12 14 16 18 20 22 24 26 28 30 32 34 36 38
Figure 3. CVD-grown MWNTs fluorinated to saturation at 520 °C for 3h (F/C = 1.1) - (Left) SEM
observation: dispersion by spin coating from an ethanol suspension - (Middle) TEM micrograph: lattice
fringes provide evidence for some residual partial order in the stacking of the successive fluorinated
layers - (Right) X-ray diffraction pattern (K Cu): the average intershell distance expands from 0.34 to
about 0.65 nm upon fluorination.
Figure 4. TEM micrograph showing details of a partially fluorinated MWNT - (Left) The black and
white framed zones indicate fluorinated and non-fluorinated regions, respectively. The central channel
of the tube can be easily distinguished - (Right) The ―zipper‖ model: a layer by layer fluorination
mechanism likely accounting for the formation of such organized F-MWNTs. The successive and
progressive rips recall the opening of zippers. Reprinted from ref. [19], with permission from Elsevier
© 2008.
4. Physicochemical Properties
a) Individual Fluorotubes
Stability: From the thermochemical standpoint, fluorinated nanotubes constitute unstable
entities in regard to their transformation into perfluoroalkanes. The expression of their overall
unstable character is activated upon simple annealing. Thus, F-SWNTs simply loose part or
all of their fluorine upon moderate heating in an inert atmosphere [8,11,12] but start to
frankly decompose beyond about 300 °C under the form of gaseous fluorocarbons (mainly
CF4) and COF2 [9,35] (nanotubes always typically contain around 3% atomic oxygen before
and after fluorination), turning back to damaged SWNTs. High temperature (>500 °C)
pyrolysis results in short fragments of SWNTs [9], following gasification of the surface
fluorinated sections.
In compliance with their synthesis temperature higher than that of F-SWNTs, F-MWNTs
appear more resistant to thermal decomposition and show almost no degradation when
annealed at reasonably high temperatures under inert atmosphere (see Figure 5). TEM
observations on SWNTs fluorinated at 300 °C showed the starting coexistence of some multi-
walled fluorinated phases [36], seemingly arising from the transformation of the single-walled
phase, that further confirms that the multilayered state constitutes a better intermediate level
of stability. Alternatively, F-SWNTs have even been observed to evolve toward multi-walled
structures under simple extensive electron-beam irradiation in a TEM [36].
Electronic structure: Contrarily to fullerene molecules, the random addition that takes
place on carbon nanotubes renders the concept of an ―el ectronic structure‖ somewhat
meaningless. Indeed, anticipating any individual behavior implies to consider a unique
residual conjugation scheme at the surface of an ―adde nded‖ tube, which requires a regular
isomeric structure of the carbon substrate plus a periodical distribution of motifs at its surface.
Some theoretical investigations [38-40] have shown that in such conditions, the fluorination
of SWNTs can open or close gaps in the band structure of the final material. This kind of
approach suffers less from the level of approximation used than from its unrealistic
confrontation with actual experimental results, since the former requirements can not be
gathered. So far, the only practically useful data lies in the prediction of an overall lowering
of the Fermi level [39] induced by the presence of fluorine. Hence, each fluorinated carbon
nanotube individually constitutes a potentially oxidizing species.
Figure 5. TGA curves (constant temperatures) illustrating the stability of the C-F chemical bond within
a sample of F- MWNTs, bulk-fluorinated at high temperature (520 °C, here).
This draws a direct link with the electronic properties of the parent fluorofullerene
molecules, well-know to be able to yield stable fluorofulleride anions [24].
Given the structural divergences previously evoked between F-SWNTs and F-MWNTs,
differences in their respective electronic structures emerge. Multiwalled fluorotubes should
not exhibit electronic and macroscopic properties too different from those characterizing the
analogous graphite-derived phase (CF)n.
Chemical bonding: At this stage, we have to first point out that the forthcoming approach
considers an average C-F bond, convenient for the purpose of generalization, but that, in
practice, non-energetically equivalent C-F moieties should sometimes be distinguished, as
detailed in a further section. Globally speaking, a weakening of the C-F bond strength occurs
in curved fluorocarbon networks in comparison to conventional fluorochemicals, but the
amplitude of variation will strongly depend on the characteristics of the carbon substrate.
Thus, if the stability of different fluorinated carbon structures is compared, one essential
factor strongly influencing the C-F bond strength intrinsically lies in the respective topologies
of the carbon lattices considered. For instance, as clearly illustrated from figure 6, the
evolution of the C-F stretching frequency in the IR region shows a concomitant decrease in
the averaged bond strength with the radius of curvature of the carbon network. Admitting that
bond strength also reflects absolute bond energy, the former phenomenon can be basically
explained by a softened covalence between carbon and fluorine. Indeed, bending, when
present, imposes valence angles that prevents the realization of a purely sp3 state at the level
of ―addende d‖ carbon atoms in the final fluorinated structure, which would be associated with
too important local strain. Some sp2 character is then partly retained in such a case,
necessarily implying that the overlap between the hybridized lobes pointing outward the
carbon frame and the fluorine atomic orbitals becomes less efficient.
Figure 6. Experimental IR spectra showing a progressive shift toward lower wave numbers of the
approximate intermediate position of the C-F stretching band, following an increase in the carbon
substrate curvature a) Bulk-fluorinated MWNTs (520 °C, composition CF1) b) SWNTs fluorinated at
300 °C (composition CF0.5) c) 80% fluorinated [70]-fullerene.
If curvature presently plays a significant role in diminishing the thermochemical stability

of fluorotubes, some additional factors may potentially contribute to further weaken the C-F
bond. Thus, an accumulation of adjacent fluorine atoms on one side of a carbon sidewall
should create steric hindrance. Yet, this can not hold for F-MWNTs, in which case nearest
neighbors fluorine alternatively stand on both sides of a carbon sheet, as seen before.
According to theoretical calculations [27], when a given exo- fluorine addition pattern is
envisaged, steric repulsion would tend to stabilize those F-SWNTs endowed with the smallest
diameters (i.e. the most curved). A pronounced curvature effect indeed lengthens the
separation between nearest neighboring addends. At last, one may also legitimately wonder
whether the initial helicity of the tubular substrate can play a role on the thermochemical
stability of the C-F moiety within fluorotubes. Computational investigations [27,37] showed
that the latter factor has a minor influence compared to that exerted by diameter.
Alternatively, it has often been inferred that bond weakening within fluorotubes arises
from a polarization of the C-F bond, an opinion issued from the several decades long earlier
background dedicated to fluorographites. However, in spite of the ability of fullerenic carbon
frames to undergo charge withdrawal [34], such a bonding concept happens definitely
obsolete, as discussed later on.
To close this paragraph, we have to point out that a more reasoned overview on chemical
bonding in fluorotubes must be provided but appears so unexpectedly complex that a whole
detailed part will be devoted to it in the following. The present short section aims only at
being a useful preliminary guide to account for some of the properties described below.
Reactivity: Standard fluorocarbons are known for their chemical inertness, a property
inherent to the generally extreme intrinsic robustness of their C-F bonds. One may then
expect the less energetically stable C-F bond present in fluorotubes to confer the latest an
enhanced chemical reactivity. This naïve picture is at the origin of a frequently committed
mistake in the interpretation of the physicochemical properties of fluorotubes, described
below.
The notion of reactivity will be essentially restricted to single-walled fluorocarbon
matrices only, since as seen previously, F-MWNTs consist of rather stable
perfluorographene-like shells, resembling their chemically inert flat homologue graphite
fluoride (CF)n. In contrast, F-SWNTs can undergo defluorination upon simple prolonged
annealing [8,11,12] at moderate temperature or upon chemical reduction with hydrazine
[1,10,43,47] or with some solid halides and sulfides [43], turning back to their initial SWNTs
state. More importantly, the latter fluorocarbon matrices are prone to nucleophilic attack, as
well. Important pioneering works [41,42] had similarly led to the grafting of functionalities to
the [60]-fullerene cage by starting from a fluorinated precursor, whereas some of these
reactions may otherwise be more difficult to obtain from the C60 molecule, especially when a
high functionalization rate is desired. The interested reader may refer to wider review articles
on the subject [24]. Originally initiated within the framework of the study of the chemical
reactivity of fluorofullerenes, investigations on this latter aspect have since been intensively
extended to fluorine atoms grafted to the surface of a SWNT and have shown the ability of
the halogen atom to be readily displaced. Hence, the attachment of alkyl chains and various
oxygen or nitrogen-based functional groups [1,43-47] to the sidewall of SWNTs has been
reported, by starting from F-SWNTs. So far, simple substitution of mobile fluorine species,
which are initially easy to fix to the carbon substrate, turns out to be the best means to
enhance the rates of derivatization of fullerenic carbon networks (= fullerenes and tubes).
One should be careful to a common confusion when interpreting the above properties.
Indeed, the ability of some fluorotubes to undergo defluorination does not arise from the
previously illustrated diminution of the absolute average C-F bond energy, but instead likely
possesses a kinetic origin [48]. Indeed, absolute bond energies stand with respect to the free
atoms and provide information on thermochemical stability, once the energy offset
corresponding to the constitutive elements in their more conventional thermodynamic
standard state is introduced. Thus, in comparison with ―nor mal‖ bonds in common
fluorochemicals, especially poly-monocarbon fluoride (CF)n, C-F moieties in fluorotubes
happen to be destabilized, i.e. the creation of C-F bonds according to Cgraphite + x/2 F2 → CFx
is less energetic when the final CFx network is bent. But interpreting the formerly described
physicochemical properties of fluorotubes from an energetic standpoint implies to rather
consider the bond dissociation energy (BDE), which itself contains an enthalpy contribution
relating to the formation of the native carbon network. It turns out that the latter term exerts
the major influence on the BDE value, as shown from the following example. In the absence
of direct thermochemical measurements on fluorotubes, a parent fluorofullerenic framework
is compared to the fluorinated derivative of graphite. Hence, direct defluorination of a
gaseous fluoro[60]fullerene molecule according to CFx → C + x F∙ necessitates on good
average 291 kJ/mol C-F [49,50], while the complete defluorination of poly-monocarbon
fluoride (CF)n necessitates only 254 kJ/mol C-F [51] (cohesive energies in the fluorinated and
non-fluorinated condensed states of graphite are of nearly the same order, due to only weak
van der Waals interaction between layers, so that the corrective terms involved in the
transition to isolated layers can be safely neglected). Overall, the C-F bond cleavage happens
then to be +37 kJ/mol more endothermic for an isolated C60Fx molecule than for a
perfluorographene sheet. This energy amount compares very favorably with the +42 kJ/mol C
representing the enthalpy of formation of a gaseous C60 molecule from graphite.
Consequently, it can be extrapolated that fluctuations in the average BDE of any fluorocarbon
lattice turn out to be almost entirely contained in the sole enthalpy of formation of the pristine
carbon frame. It follows that the average BDE of a fluorocarbon matrix will increase
concomitantly with the radius of curvature of the naked carbon precursor.
Consequently, great care should then be taken when evoking poorly bound fluorine
atoms, while referring to the physicochemical properties of fluorotubes previously recalled. In
fact, regarding the above mentioned energetic considerations, fluorine atoms within
fluorotubes turn out to be tightly bound to their carbon substrate since defluorination of a
tubular network will require more energy than that of graphite fluoride. However, it turns out
that this energetic statement appears in strong contradiction with all experimentally
established trends, since F-SWNTs can readily release fluorine throughout different
processes, as seen before, whereas graphite fluoride (CF)n is an extremely stable compound
until around 500 °C under inert atmosphere, decomposes beyond into gaseous fluorocarbons
instead of undergoing defluorination, and is chemically inert. Such a paradox, opposing
thermodynamics, clearly means that the easy fluorine departure from F-SWNTs compounds
has a pure kinetic origin, so that a terminology like ―l abile fluorine‖ should be rigorously
preferred. The underlying kinetics can be reasonably understood by referring to hybridization
states in conjunction with activation energies. In pristine SWNTs, the sp2 character of the
carbon atom is limited by the pyramidalization angle imposed by curvature, while the sp3
character of the carbon atom in the fluorinated derivatives is in turn limited for similar
valence angle reasons, so that in the course of either a fluorination or defluorination process,
part of the hybrid character initially present is retained. Hence, this should lower the energy
barrier over which to pass to reach the transition state involved in the transformation. This
makes an initially bent carbon skeleton easier to fluorinate and in parallel, its fluorinated
homologue also easier to defluorinate. In contrast, the fluorination/defluorination of graphite
is a total sp2↔sp3 transformation involving complete conversion of the hybridization state of
carbon. This introduces kinetic limitation which can be overcome only under harsher
experimental conditions.
Though fundamentally correct and laying the foundation stone of the physico-chemistry
of F-SWNTs, we have to here again emphasize that the present approach implicitly stands for
an averaging of the chemical bonding properties and that, in practice, partial distinction
should be introduced between C-F moieties.
b) Solid State
Solubility/dispersion: Bulk samples of F-SWNTs prepared at 250 °C exhibit excellent
dispersion properties upon sonication in alcohols [47], whereas perfluorinated solvents
happen surprisingly quite inefficient in this purpose, in spite of the ―l ike dissolves like‖
tendency. This was tentatively attributed to an enhanced dipolar moment of the C-F chemical
bond that may facilitate the formation of hydrogen bonds from the partly charged fluorine
atoms.
Conductivity: It can be readily foreseen that the functionalization of carbon nanotubes
should generate insulating materials, upon random breaking of conjugation. Thus, not
surprisingly, the resistivity of an initially conducting film of SWNTs was found to increase by
six orders of magnitude after fluorination [1]. In another study [5], the resistivity was also
found to increase in parallel with the level of fluorination. Let‘s note that such phenomena,
though intimately correlated with the evolution of the ―ov erall‖ electronic structure, reflect a
sum of individual behaviors within a sample rather than a true collective behavior.
In regard to the resemblance that F-MWNTs bear to poly-monocarbon fluoride, it can be
taken for granted that the latter similarly exhibit strong insulating properties. Nevertheless,
when not bulk-fluorinated, it is expected that they can also exhibit a mixed behavior
combining the insulating properties of their outer fluorinated shells with those of their
conducting to semi-conducting inner pure carbon core.
II. CHARACTERISTICS OF THE C-F CHEMICAL BOND IN

FLUOROTUBES
1. A Weakened Plus Versatile C-F Bond
As for summary of the essential points established in the former part I, one may first
basically recall that the high affinity of carbon for fluorine usually makes this combination
one of the most energetic simple bonds. Thus, in thermodynamic equilibrium conditions, the
simple pairing of one fluorine atom and one carbon atom yields a stable.:CF(g) radical
fragment, called fluoromethylidyne, whose bond dissociation energy is 540 kJ/mol [52,53].
We then evoked, independently of any local structural aspect, how the global stability of the
latter atomic association in the solid state is made dependent on the carbon frame. The present
part II aims at providing a final description of the fluorine addition pattern to carbon
nanotubes. Focus will be made on the variability of the C-F binding energy that can appear in
fluorotubes, which arises from its ability to be frozen in local minima.
MW Fluorotubes: The tendency toward metastability of the C-F bond in fluorotubes is
clearly exemplified throughout the low and high temperature forms of multiwall fluorotubes,
which clearly denote the opportunity of formation of kinetic products. We have depicted in
the first part how the multi-walled tubular carbon allotrope mimics by several ways the
chemistry of graphite, especially from the point of view of its reactivity with fluorine. Thus,
the high temperature fluorination of MWNTs yields products that compare well with the
graphite fluoride form (CF)n prepared at high temperature, whereas low temperature catalytic
conditions yields a surface phase analogous to the so-called intercalation compounds of
graphite with fluorine. It can be intuitively guessed that the varied addition modes of fluorine
to a MWNT, under the form of either intercalation or addition final compounds, are
intrinsically associated with changes in the nature of the C-F chemical bond. The
experimentally checked quenching of the intercalation process upon increase of the synthesis
temperature is the signature of rapid thermal decomposition, providing evidence for the
instability of the C-F bond involved in it. The increase in the synthesis temperature alleviates
kinetic limitations and then turns the fluorination process toward the formation of a true
addition compound involving the thermodynamically stable covalent C-F moiety.
SWNTs: Though manifesting in a different way, the versatile character of the C-F bond in
F-SWNTs has also asserted itself and several authors have explicitly mentioned the
coexistence of mixed bonding properties in their samples of single-walled fluorotubes
[5,9,12-14,61]. These have been evidenced most of the time by an apparent splitting of the
associated F1s XPS signal [5,13,14], or through the wide wave number range associated to the
stretching of the C-F bond [9,12,61]. Simple gravimetric techniques exploiting controlled
thermolysis have recently provided a quantitative determination of the different bonding
modes present in F-SWNTs [12]. As illustrated below, three different modes of fluorine
chemisorption can be unambiguously evidenced within single-walled fluorotubes prepared at
100 °C, for instance. The TGA curves of such samples exhibit distinct steps of fluorine
desorption, tentatively interpreted as the successive rupture of weak to stronger C-F bonds
over the respective ranges 50 °C<T<150 °C, 150 °C<T<250 °C and T>250 °C. The respective
amounts of each C-F bond type can be quantified when selective thermal desorption is used, a
sequential cleavage of the three different types of C-F bonds progressively taking place. Thus,
figure 7shows that the thermolysis of such fluorotubes stabilizes after one day at 150 °C,
providing evidence of the complete removal of a first category of F atoms, involved in the
weakest bond type. A further and again stabilized weight loss can be achieved at 250 °C, after
a similar duration, signing for the complete removal of another kind of bonds. Note that about
24h are required to overcome kinetic limitations each time. By weight difference with respect
to the initial total fluorine content of the sample, the existence of a third bonding mode of
fluorine is deduced, supposedly representing the strongest and therefore, thermodynamically
stable, interaction.
A first tentative explanation of such a gradual classification might be that the C-F bond
dissociation energy in F-SWNTs correlates to the diameter and/or helicity of a tube, as seen
previously. Since samples of SWNTs consist of mixtures of individual entities with different
intrinsic characteristics, their fluorination might indeed be expected to result in fluctuating
bond strengths inside a same sample. However, variation in the tube diameter seems to be the
parameter having the greatest influence on bond strength and the size distribution in SWNTs
samples being usually narrow, it seems that the present hypothesis can not contribute
significantly to the drastic variations observed.
Figure 7. (Left) Thermogravimetric analysis curve of a sample of SW fluorotubes, prepared at 100 °C

for 4h (composition CF0.34), showing successive fluorine desorption steps. (Right) Relative amounts of
the different C-F bond types in the same sample, as revealed by sequential thermal desorption.
Reprinted from ref. [12], with permission from Elsevier © 2007.
According to computational investigations [25], isomeric addition configurations usually

exhibit too small energy differences and can again not account for so large fluctuations in the
C-F binding energies. At this stage, the ultimate hypothesis of metastability becomes fully
justified. From the experimental standpoint, the increased amount of the most stable bonding
mode with the increase of the synthesis temperature seems to confirm an influence of kinetic
factors. It is then highly likely that owing to a high activation energy regarding the formation
of the most thermodynamically stable bond and/or difficulty in getting ordered at the surface
of a tube, some fluorine atoms can be trapped into local energy minima. Accordingly, out of
equilibrium C/F subsystems should provide a satisfying explanation for the non-
homogeneous bonding scheme experimentally revealed in F-SWNTs. Their origin lies in the
existence, dictated by the topology of the carbon substrate, of secondary binding sites that can
readily allow fluorine storage. Thus, beside the outer surface of a tube, different regions can
contribute to fluorine fixation, including both the interstitial channels between contiguous
tubes belonging to a same bundle and the inner hollow core of a tube.
In preliminary conclusion, F-SWNTs exhibit exaggeratedly important deviations in their
bonding properties for a regular bonding scheme to be inferred and further characterization is
required to really find out about the different ways fluorine can bind to the tubular carbon
substrate. Alternatively, it has sometimes been argued [5,9,13,14] that the former
differentiation in the C-F bond strength within single-walled fluorotubes has to be related to
the emergence of an ionic character at the level of some bonds. Such arguments are
manifestly wrong, as shown in the next section.
2. C-F Bond Strength Versus C-F Polarity: Revision Required?
The C-F bond within a fluorinated inorganic carbon network containing graphene-like
structural units usually possesses constant characteristics within a same sample, but can be
variable from sample to sample according to the synthesis conditions. Hence, it has long been
claimed that the C/F interaction in this case is able to evolve from ionic to covalent, including
a somewhat exotic intermediate state set between ionic and covalent. Though rather
improperly, the terminology ―s emi-ionic‖ or ―s emi-covalent‖ has been of common use to
describe the latter intermediate state, instead of ―i
onocovalent‖.
Accordingly, the most part of interpretations regarding chemical bonding within
fluorofullerenic networks (the term stands for fluorofullerenes + fluorotubes) have been and
still keep on being performed in a state of mind imposed by the several decades long earlier
background on fluorographites, that is, bond softening has become a synonym of bond
polarity. For instance, the presence of notably polar C-F groups was claimed [54-56] in
fluorofullerenes, following shifts observed in their IR and XPS features. The versatility of the
C-F bond in fluorotubes has also often been explained via this model and depending on the
fluorination conditions, bonds qualified by the authors as covalent or ―s emi-covalent‖ can
form and seemingly even coexist [5,9,13,14]. While the temperature threshold ensuring the
formation of as covalent as possible bonds seems to be sample dependent, a general
consensus was reached in that the higher the synthesis temperature is, the more developed the
covalent character of the final C-F bond in the sample is.
If the role played by the curvature of the carbon network in regard to the softening of the
C-F bond within fluorotubes has not to be demonstrated anymore, may one complementarily
see in the latter weakening effect the signature of a polarization of the C-F moiety ? In this
context, it may be interesting to have a closer look at the extent to which some of the ideas
that have prevailed for decades on fluorographites should be reconsidered in the light of some
late investigations, unhappily never taken into account. It follows that many of the
interpretations published so far on the nature of chemical bonding in fluorotubes and
fluorofullerenes are widely open to criticism and should be properly revisited. Indeed, while
the either ionic or covalent characters of the C/F interaction in graphite fluorides seem well
established and have obtained general acknowledgement among researchers involved in the
physico-chemistry of inorganic fluorocarbons, the pertinence of the ionocovalent
(―semicovalent‖) bonding mode happens obsolete in the light of some experimental and
theoretical data.
Thus, as early as 1999, Panich [23] noted that the covalent (CF)n and supposedly
ionocovalent CFx forms of graphite fluoride exhibit similar 13C NMR chemical shifts. The
shielding effect being partly function of the electron cloud symmetry around the nucleus, no
real change in the hybridization state of the carbon atom must consequently occur from one
form to the other, which would otherwise induce sensitive difference in their respective 13C
values. In other words, no significant modification in the bond character must occur from one
form to the other. He also noted that the respective positions of the 19F lines are reversed in
regard to what would be expected from an increased partial charge on the fluorine atom,
therefore sharply contrasting with the idea of a variation in bond polarity between the two
fluorographitic forms. He then concluded that variation in the ionicity of the bond should not
stand as a correct explanation of the bond strength weakening in the intermediate form of
graphite fluoride, hitherto thought as ionocovalent. Further NMR studies [57] have since
confirmed that the so-called ―s emicovalent‖ fluorographite form consists of a mixture of C-
Csp2-C and Csp3-F carbons, implying that the C-F bond has a true covalent character. Recent
neutron diffraction results by Sato et al. [58] have provided further highlight in favor of the
non-existence of an ionocovalent bonding mode in the latter compound. Their analysis of the
carbon environment has led to an estimated carbon-fluorine separation of 0.140 nm. This
represents only a slight elongation with respect to the 0.136±0.003 nm bond length present in
any common fluorochemical, while one may intuitively expect interatomic distances to
strongly vary in case of charge separation. A structural model was proposed in which the
carbon planes are buckled, contrarily to all previous models, indicating that orbital overlap
between fluorine and sp3 carbon, i.e. a normal covalence state, prevails. An independent
computational study [59] has also confirmed that the energetically favorable configuration
necessitates the realization of a sp3 hybridization state for those carbon atoms directly linked
to a fluorine atom. In other words, the C-F bond in the intermediate form of graphite fluoride
appears as a conventional, but softened, covalent bond. The reason for bond weakening was
shortly discussed by Sato et al. [58] who proposed that the latter effect results from the
participation of the valence shell of the fluorine atom in a hyper-conjugation phenomenon,
involving the subsisting conjugated -scheme present in each partially ―addende d‖ carbon
layer.
Given the obsoleteness of the ionocovalent model in the case of fluorographites, it seems
clear that the same must hold true for fluorofullerenes and fluorotubes and that weakened, but
nevertheless fully covalent bonds do form in these compounds. Consequently, both the
overall softening effect and the mixed character of the C-F bond in fluorotubes are
independent of any variation in the polarity of the bond, as sometimes inferred earlier. The
respective contributions to the overall softening of the bond strength remain curvature and
potential steric hindrance, to which we will now add the metastable location of some fluorine.
These naturally go in parallel with a soften IR frequency associated to the main vibration
mode, while a higher core molecular level also results for fluorine, which accounts for the
upshifting effect sometimes recorded from the F1s XPS signal. The present conclusion that
partial ionicity can not stand as a correct explanation for bond weakening in fluorofullerenic
compounds is further reinforced by an independent computational study [27], who established
that the Mulliken charges on the fluorine atoms in PTFE, CH2F2 and a C2F fluorotube isomer
are similar, rendering the existence of an exotic charge state in the latest highly doubtful.
Anticipating the True Bonding Scheme of F-SWNTs
The set idea considering exo- fluorine chemisorption on the surface of SWNTs has
intuitively prevailed hitherto, guided by the initial STM view [2] on the outer morphology of
single-walled fluorotubes and probably also by antecedent findings regarding the parent, exo-
functionalized, fluorofullerene molecules [24]. Yet, attention should be drawn on the facts
that STM does not allow observation of inner regions, which might well bias our empirical
apprehension of such systems, and that fullerene molecules are closed cages which only
permit external addending, while this might not be the case for tubes, usually open on tips.
Consequently, a more rigorous state of mind on the question is necessary and in parallel,
seems also dictated by the most recent experimental characterizations.
Hence, the simplest structural model that forces itself upon us and that would account for
versatile C-F bonding modes through multiple locations of F atoms is depicted in figure 8.
The present description comprises a first group of F atoms involved in (CF)n-like sections
which should represent the thermodynamically stable addition configuration. A second group
is made of fluorine atoms involved in a fluorofullerene-like addition pattern, of intermediate
stability. A third and last group consists of endo- fluorine atoms, likely destabilized.
The pertinence of this multi-site model finds at least partial justification throughout
different experimental and theoretical facts. First, the structure of F-MWNTs was claimed to
be made of rolled (CF)n-like sheets [17-20], so that at least partial filiation is highly likely to
occur with F-SWNTs. In parallel, the scarce computational investigations [39,40] that
considered an a priori exotic succession of exo/endo fluorine addends on the sidewall of an F-
SWNT, concluded in the enhanced or competing thermodynamic stability of the latter in
regard to an equivalent but purely exo- functionalization pattern. Finally and more
importantly, the 19F NMR spectrum of high temperature prepared F-SWNTs is made of a
broad hump centered on -175 ppm [47], which is very close to the -180 ppm chemical shift
recorded for the (CF)n compound [23]. This suggests almost unambiguously that (CF)n-like
sections should constitute a notable part of the fluorine addition pattern of F-SWNTs. IR
spectroscopy strictly confirms this assumption. Indeed, the IR spectrum displayed in figure 6
spreads over a wide wave number range but distinctly shows a first tooth at 1205 cm-1,
approaching the 1220 cm-1 feature that characterizes the (CF)n compound. The small
frequency downshift simply occurs as a manifestation of the curvature effect on bond

strength, as seen before. In the second place, IR spectroscopy seems to also confirm the
existence of fluorine atoms belonging to the second group envisaged. Indeed, a broad but
distinct feature is also apparent over the 1100-1150 cm-1 zone in figure 6, which is the typical
absorption range of C-F moieties in fluorofullerene molecules (see Figure 6). The presence of
a fluorofullerene-like subsystem in the overall addition pattern of F-SWNTs is therefore
highly likely, as well. The existence of the third group happens to be more difficult to
evidence since no equivalent reference compound is known. However, some authors reported
[7] that, at constant final F/C ratio, the fluorination of open and closed-end SWNTs generates
a lesser intertubular expansion in the case of open tubes. This might be an indirect indication
that part of the fluorine can be located on the inner side of tubes. Our own most recent work
[60] has also confirmed the possible endo-functionalization of SWNTs.
Figure 8. Provisional and schematic overall fluorine addition pattern to SWNTs.
At last, the influence of synthesis conditions on the final constitution of the mixed
bonding scheme in F-SWNTs may be discussed. Experimental observations indeed indicated
that the synthesis temperature is a key parameter affecting the respective proportions of each
bonding mode [12] in the final fluorotubes sample, but the fluorination method retained
(gaseous fluorine vs fluorinating agents, plasma techniques…), the reaction time and some
morphological characteristics (average diameter size, bundled or individual tubes…) at the
level of the precursor SWNTs batch may also more or less significantly influence the final
result. The dependence of the final global addition pattern on the experimental conditions
undoubtedly explains why characterization of F-SWNTs has often led to different results
throughout the literature, depending on the chemisorption mode that prevailed within each
sample studied.
From the energetic standpoint, we can intuitively postulate stability in the respective
order: group I> II> III, so that II and III appear as metastable subsystems at the origin of
softened covalent bonds. Indeed, fluorine atoms of group I abide by the architecture of the
stable (CF)n compound, while group II addends undergo higher local strain at the level of the
carbon frame plus probable steric hindrance between nearest neighbors in eclipsed position, if
1-2 addition is respected. The group III endo-addition pattern possesses a still more reduced
stability, owing to the concave deformation generated at the surface of a convex carbon
surface plus also probable steric hindrance. Therefore, the maintenance of group II and III
with respect to the more energetically favorable group I arise from kinetic limitation that is
progressively overcome by increasing the synthesis temperature and/or reaction time,
favoring evolution of the global addition pattern toward the most stable of the three
subsystems.
The first hypothesis inferred to interpret the origin of kinetic effects is that the formation
of bonds necessitates an increased activation energy in the order: group III< II< I. It can
indeed be anticipated that the final p character of an ―addend ed‖ carbon atom regularly
diminishes from group I to III, meaning that the final hybridization state of carbon atoms
linked to fluorine is all the more close to their initial hybridization state in the precursor
carbon substrate that the group number is high. Consequently, the pathway to the transition
state might be energetically facilitated in the order III easier than II easier than I. The second
and more reliable assumption is that the group I pattern likely derives from group II and III
motives, after ―backs ide completion‖ of the latter, which additionally implies potential
rearrangement if 1-2 addition initially prevails. Indeed, in terms of probability, the direct
formation of group I motives seems difficult because simultaneous attack on opposite sides
by two fluorine molecules is required. Therefore, the creation in two steps of addends of the
first group may slow down their apparition, so that again, high temperature and time will be
required to observe their proliferation on the surface.
At last, some complementary morphology-related mechanistic grounds responsible for
endo-fluorination can be reasonably anticipated, as well. Indeed, an arrangement under the
form of bundles sometimes prevails in SWNTs sample. Owing to the compactness of such
nano-ropes, diffusion of fluorine molecules into the carbon lattice is hindered from the lateral
sides. Fluorine admission must then rather occur from the bundles tips, where different
apertures pre-exist. Consequently, fluorine species diffuse into the compact carbon lattice
either via migration in the intertubular channels or directly through the hollow core of a tube.
The cross section of a tube being about ten times that of an interstitial channel, the contact of
fluorine molecules with the inner side of a tube rather than with the outer side should be
clearly favored, facilitating endo-addition. In such conditions, a high temperature route will
be necessary to initially provide sufficient kinetic energy to the fluorine molecules and carbon
lattice, so that to force ―de-cohesion‖ within the ropes and to allow the parallel formation of
the other subsystems.
III. APPLICATIONS AND PROSPECTS: ADVANTAGES OF A WEAK AND

VERSATILE ENERGETIC BONDING SCHEME
Developments and Future Progress
In this last section, it will be dealt with the potency of fluorinated nanotubes in regard to
advanced technological developments. So far, attempts of valorization of research performed
on fluorotubes have been mostly driven by some antecedent industrial exploitation of
fluorochemicals. Graphite fluorides or some fluorinated polymers are indeed currently used
under the form of lubricants, cathode materials, or fibers entering into composite materials or
textiles. In comparison to the standard fluorocarbon materials previously cited, the C-F bond
weakening in fluorotubes is at the origin of an enhanced mobility of the fluorine atoms which
may allow optimization of some performances.
In parallel, it is expected that fluorotubes may benefit to new kinds of applications.
Indeed, fluorination stands so far as the starting point for a great part of the modifications
performed on carbon nanotubes, rendering fluorotubes fundamental intermediates for the
integration of nanotubes in nanotechnology processes.
Functionalization: We will first briefly recall that in sharp contrast with the very bad
nucleofugal properties of the fluorine atom in classical fluoroalkanes, fluorine atoms in
fluorotubes can be readily displaced, rendering fluorotubes important synthons in the context
of nanotubes chemistry. Thus, many synthesis routes use F-SWNTs as precursors [1,43-47] in
view of the subsequent chemical derivatization of SWNTs. The basic aspects of fluorine
displacement by some functions have been formerly depicted and we will then restrict this
paragraph to an example of the new synthesis developments recently achieved [61]. It turns
out that the differentiated bonding scheme of fluorine atoms within F-SWNTs can be turned
to good account in some advanced synthesis patterns yielding to the controlled bi-
functionalization of the SWNTs surface by heteroelements.
Hence, it has been observed [61] that in an acidic aqueous medium, partial pre-
fluorination stimulates the subsequent grafting of hydroxyl terminations to the sidewall of
SWNTs via the classical substitution of F by O, resulting in an oxyfluorinated compound. A
significant upshift in the position of the F1s peak (Figure 9) in the course of the XPS analysis
of the products indicated an evolution in the strength of the C-F bonds before and after acid
treatment.
C-OH
C-F
C-O
3600 3200 2800 2400 2000 1600 1200 800

-1
Wave number (cm )
Oxyfluorinated SWNTs
Fluorinated SWNTs
678 680 682 684 686 688 690 692 694 696 698
BE (eV)
Figure 9. (Left) IR spectrum of oxyfluorotubes obtained upon acid treatment of F-SWNTs. (Right) F1s
XPS spectra of F-SWNTs before and after treatment in acidic conditions, showing an evolution in the
strength of the C-F bonds. Reprinted from ref. [61], with permission from Elsevier © 2008.
This resulted in the conclusion that a selective substitution of fluorine by oxygen can be
achieved through the displacement of the sole most weakly bonded, and therefore
nucleofugal, F atoms, since only the strongest C-F bonds subsist after substitution. Such a
tunable reactivity of the F atoms in a fluorotube paves the way toward a quantitative
functionalization process of the SWCNT surface by hetero-elements. In practice, the
advantage of the concept remains limited, yet, the selectivity in the exchange power of the F
atoms being quenched in the presence of too strong nucleophilic reagents [61,62].
Electrochemistry: The covalent perfluoro-derivative of graphite (CF)n has been
commercialized as a primary lithium battery component for several decades now. Cathode
materials are probably the field par excellence where the weakening of the C-F bond can
benefit to optimized performances. Indeed, the thermodynamics of a CFx/Li battery is
governed by the Gibbs energy of decomposition of the fluorocarbon phase into carbon, which
acts as an unfavorable factor, compensated by the free enthalpy of formation of LiF upon
discharge. The higher the bond energy is, the higher the increase in the absolute value of the
Gibbs energy of the cell reaction is, so that any loss in the C-F bond strength is interestingly
recovered under the form of power for the battery. As expected, fluorotubes show open circuit
voltages superior to the one characterizing perfluorographite, when used as cathodes materials
in lithium batteries, but preliminary tests [63-65] also globally displayed modest
performances owing to a significant potential drop in the course of discharge and to a limited
Faradaic capacity. Future developments are required to achieve a more competitive system, in
which partially fluorinated MWNTs could play a role. As seen in a former section, the latter
indeed consist of fluorinated shells superposed to a pure carbon core, i.e. consist of an outer
electrochemically active layer wrapping an electron conducting central part that might limit
Ohmic drop in the cell.
Supercapacities are the second main electrochemical energy storage device. Samples of
fluorotubes do not offer good capacitances but fluorination followed by defluorination at high
temperature seems to constitute an efficient means to develop the porosity within a sample of
raw carbon nanotubes, and as a corollary, to increase its specific capacitance [66].
Mechanical properties: Some theoretical calculations predicted [67] that the exceptional
intrinsic mechanical properties of carbon nanotubes are preserved upon addition of fluorine.
Additionally, the presence of surface C-F groups offers a way to favorably tune the strength
of interfacial interaction or adhesion between SWNTs and another external medium. Such
fundamentals clearly render fluorotubes potential reinforcing agents for composite materials.
The discovery of the homogeneous dispersion properties of fluorotubes represents one
concrete step in view of such a practical development. Let‘s here recall that fluorinated tubes
form stable suspensions in alcohols over a long time and that they also seem to present a good
affinity for polymers, satisfying one more key requirement for their processing under the form
of tentative fillers for polymer matrices. Thus, a growing number of studies now report the
use of F-SWNTs [68-70] or F-MWNTs [71-73] as reinforcing agents in nanocomposites
made from different kinds of polymers. A significant increase in the mechanical properties is
claimed most of the time, in parallel with the fluorination level and/or load percentage of
fluorotubes. Fluorination happens to strongly improve interfacial adhesion since fluorine
atoms can be removed and subsequently substituted by direct covalent bridges with the matrix
[69,70].
In the short term, new synthesis routes allowing a controlled chemical modification of
carbon nanotubes could play a key role in the expansion of nanotube-based composites. The
previous example of oxyfluorotubes offers a nice overview on one essential realization that
might be achieved in this field. One may indeed imagine that the relative proportions of each
type of C-F bond in a fluorotube may be modulated according to its preparation conditions,
and that so may be the selective exchange of fluorine by oxygen (or by some other hetero-
element) and the consequent final O/F/C stoichiometric ratio. The practical interest of such
hetero-functionalized derivatives as reinforcing materials is obvious, since the filler-matrix
interactions might be tuned through the ratio of the hetero-elements, opening a window on
materials with desired mechanical performances. At last, one may also try to anticipate to
which extent partially fluorinated MWNTs as those displayed in figure 4 may efficiently
ensure the mechanical reinforcement of composites. Their use would indeed involve a
complex combination of effects, including that of a fluorinated outer surface improving
dispersion and/or interactions at the fibre-matrix interface and that of a sliding and highly
resilient inner carbon core.
Tribology: The exceptional friction properties of C-F covered surfaces are well-known
and commercially exploited under the form of Teflon® or poly-monocarbon fluoride based
lubricants. This fully justifies the initial suspicion that similar or even better properties might
be obtained from fluorotubes. However, the lubrication durability of F-SWNTs was shown
[74] to be limited by cracking, owing to the weakening of the frame induced by fluorination.
In this context, F-MWNTs might here again appear as a possible alternative, at first
glance. Indeed, the partial fragmentation induced by fluorination results in a relaxation of the
internal strain which improves flexibility and hence, might help in the preservation, under a
mechanical constraint, of tubular-shaped nanoparticles playing the role of nano-bearings.
Nanoelectronics: One of the most promising and exciting future development, that has
boosted expansion of research on carbon nanotubes, is the integration of the latter as
nanometer scaled components into a new generation of electronic circuits. At first glance,
functionalization does not appear as adding value at this level, because its random character
makes difficult the control of the electronic properties of tubes by this means. Yet, surface
fluorinated MWNTs couple insulating outer properties with conducting inner properties that
make these fluorocarbon architectures look like nano-sized electric wires. One might then
expect the latter to be used to carry electricity inside nanodevices, so that to ensure
connections between nanocomponents.
Processing: Finally, we will shortly emphasize the potential impact of fluorination as a
―pr ocessing method‖ for carbon nanotubes. Due to the reversibility of fluorination,
fluorotubes might well be seen as simple intermediates in the course of an overall
transformation scheme of raw carbon nanotubes. Thus, an etching effect on the carbon
skeleton following high temperature pyrolysis of fluorotubes was reported [9], but sounds
rather anecdotic since a similar effect can be obtained upon prolonged sonication of a
suspension of raw tubes.
The key point slowing down the development of carbon nanotubes-based devices is the
control of the intrinsic characteristic of the tubes during their synthesis, which, so far, results
in a mixture of conducting and semi-conducting entities. Trends in this field have lately
turned toward post-synthesis chemical sorting methods. Though reported several times from
different kinds of oxidizing reagents [34], the selective ―s orting effect‖ accompanying the
reaction of raw SWNTs with fluorine seems worth being noticed. As a matter of fact, the
selective quenching of Raman and optical absorption features observed during a [room
temperature fluorination/ high temperature defluorination] cycle showed that a preferential
destruction of small diameter metallic tubes occurs in this case [75], providing a basis to an
enrichment process. The latter results have been seemingly confirmed by an ab initio study
[76] stipulating that metallic tubes interact more favorably with fluorine than semi-conducting
tubes, beyond a critical diameter threshold.
Prospects
As for conclusion to this chapter, we will detail below a few concepts, remaining
speculative at this stage, but that may stimulate future research in the field. Some are inspired
with commercially exploited applications of other perfluorocarbons, a fact reflecting in itself

a promising potency in regard to future valorization.
Gas storage: From kinetic grounds, we have previously justified why F-SWNTs contain
rather labile fluorine atoms. If synthesis conditions favoring metastable addition are
employed, the labile character of the fluorine atom happens to be highly reinforced. In such a
case, simple heating at moderate temperatures can rapidly permit the removal of important
amounts of fluorine (see Figure 10). The precursor carbon matrix could then be
advantageously envisaged for fluorine storage. An obvious underlying interest arises,
regarding some potential energy storage devices, given the highly exothermic reactions to
which elemental fluorine, once liberated, can give birth.
Figure 10. TGA curve of a SWNTs sample fluorinated at room temperature, showing how some
fluorine can be rapidly given off.
Soft fluorinating agents: Alternatively, transforming an initially weak C-F bond into a
stronger one, by transferring the fluorine atom to another compound in which the C-F energy
level would be deeper, appears straightforward from the thermochemical standpoint.
Therefore, due to their ability to release fluorine, some fluorotubes are good candidates likely
to act as donors and could probably be employed as soft fluorinating synthesis agents. The
fluorine exchange process may be activated upon heating a mixture of the respectively donor
and acceptor solid phases, for instance, and might be efficient regarding H substitution in –
CHx groups or for the halogenation of pure carbonaceous materials.
Figure 11. Annual publication trend, estimated on the basis of the whole bibliography herein
referenced, and reflecting evolution in the research effort dedicated to fluorinated carbon nanotubes
over the past 12 years.
Smart textiles: The notoriety of some fluorine containing synthetic fibers has entered our
everyday life, the most famous being probably Gore-Tex®, which is nothing else than thermo-
mechanically expanded PTFE. Like their non-fluorinated precursor, fluorotubes can likely be
processed under the form of fibers, paving the way toward their integration into tissues and
canvas. In this framework, the combination of the robustness of their carbon frame with the
specific properties of the C-F bond may provide a bunch of interesting characteristics. The
well known hydrophobicity of the C-F group intuitively suggests fluorotube-made anti-soil
and/or water proof textiles, or highly resistant and water repellent surface coverage tissues,
for instance. Furthermore, the stabilization of the C-F bond in F-MWNTs may render them
compatible with biological tissues, opening the door toward novel implant materials or
growth templates, which require chemical inertness.
…and much more: Nanosized particles are known to modify the barrier properties of
polymers. In parallel, the introduction of -CFn groups in a polymer chain is also known to
have a similar influence. Fluorotubes are a perfect example of a nanoload that would combine
both effects and that may, for instance, benefit the tuning of the separation properties of
membranes. Alternatively, the reasonable resistance to thermolysis of F-MWNTs (see Figure
5) might also render the latter useful agents for improving the fireproof performances of some
polymers.
CONCLUSION
Still at an exploratory stage only a few years ago, the development of research on
fluorinated carbon nanotubes is now becoming routine. Its evolution is well reflected by a
growing number of contributions to the subject year after year, a picture of which is given
below. Among all possible future work directions, one is clearly emerging. Indeed, some key
knowledge concerning some characteristics of the C-F chemical bond in fluorotubes still
remains to be ascertained before application attempts can confidently build on a solid
fundamental basis. In parallel, one may regret that most studies have been performed on
fluorinated SWNTs, so far, leaving fluorinated MWNTs on the fringe of research in this field.
The latter really possess an appealing applicative potential that I have tried my best to
emphasize throughout this chapter.
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Lecture Material 10
CARBON NANOTUBES: APPLICATIONS IN THE

DEVELOPMENT OF ANALYTICAL METHODS
OBJECTIVE
Carbon nanotubes also have unique electronic properties, different from graphite, and
present an exciting potential for nanoscale electronic devices. The predominant synthesis
methods for carbon nanotubes produce two distinct kinds of nanotube materials: single-wall
carbon nanotubes (SWCNTs) and multi-wall carbon nanotubes (MWCNTs). In a typical
single-wall synthesis, the nanotubes vary in diameter from 1.0 nm to 1.4 nm, and can bond to
one another in a regular parallel array on a triangular lattice. Such composite structures are
sometimes referred to as ropes or bundles. A multi-wall nanotube, on the other hand, is a
composite structure comprising many single-layer nanotubes, all nested concentrically. The
spacing between the layers is close to the natural Van der Waals layerlayer spacing for
graphite, 3.4 Å. The combination of size, structure and topology gives nanotubes important
mechanical and surface properties. The electrical properties of CNT depend sensitively on
their diameter and chirality. In this lecture, we will present a brief review of the application of
CNTs in the analytical sciences. Finally, we will demonstrate new analytical data (a case
study) obtained in our research groups involving the development of a solid phase extraction
system for cobalt determination using CNT as well as a new potentiommetry stripping
method for antimony determination using CNT as the electrode.
INTRODUCTION
The importance of nanomaterial chemistry is nowadays uncontested and new and
sophisticated materials with surprising properties have influenced both the chemical and the
materials sciences. Considering the materials classified as nanomaterials, carbon nanotubes
(CNTs) are the most representative. This material was synthesized for first time by Iijima
(1991), using the graphite pyrolysis process in plasma under a helium atmosphere. Despite
this discovery, it is important to observe that in 1889 a patent related that filaments of carbon
can be formed from hydrocarbons in metal crucibles at high temperatures (Herbst et al. 2004).
It should be emphasized that the discovery of CNTs by Iijima as well as the discovery of
fullerenes in 1985, created new boundaries for the chemistry and physic of carbon. Carbon
nanotubes can be conceptually considered as originating from single layered sheets of
graphite, rolled up into perfectly seamless tubes with nanometer dimensions (Rotkin and
Subramoney 2005). The sp2 carbon-carbon covalent bond in graphite is among the strongest
known bonds in chemistry, and carbon nanotubes are therefore mechanically robust. The
bonds between layers of graphite have relatively weaker Van der Waals interactions that give
graphite its natural lubricating properties. According to their structural parameters, SWCNTs
can behave as a metal, a semiconductor or a small-gap semiconductor. Since the topological
defects in nanotubes result in local perturbations of their electronic structure, the pentagonal
defect of caps make them more metallic than cylinders (Filho and Fagan 2007). Due to their
interesting properties, such as high surface area, high chemical and physical resistance and
electrical properties, great attention has been paid in recent years to the development of new
analytical methods covering all fields of analytical science. The use of carbon nanotubes as an
analytical tool in filters and membranes, as solid sorbent material, in electrochemical systems
involving the building of sensors, biosensors or carbon nanotube paste electrodes for stripping
techniques and in separation methods including chromatography and electrophoresis can be
found in the literature.
ATTRACTIVE PHYSICAL PROPERTIES OF CARBON NANOTUBES FOR

APPLICATION IN ANALYTICAL SCIENCES
In the last two decades no other chemical structure has aroused so much attention as have
carbon nanotubes. The confluent ability of carbon nanotubes to gather together physics,
chemistry and molecular biotechnology is one of the main reasons for the great interest in
these unidimensional nanostructures (Dai 2002, Yan et al. 2007, Katz and Willner 2004).
Research results from the basic single structure to more complex network arrays have
motivated scientists of different areas, yielding in the first quarter of 2004 more than 3500
publications dealing with carbon nanotubes (Wildgoose et al. 2006). The specific interest of
CNTs to analytical sciences starts from the very basic modification of their surface, and
making them functionalized areas that can be used, for instance, in a large range of biosensors
based on protein modified surfaces. Since, the physical properties of nanotubes are totally
dependent on surface modifications, understanding the first principles of the molecular
structure of CNTs may help researchers to better comprehend and predict the effects observed
in their experiments.
The Structure of CNTs: Wrapping a Graphene Sheet into a Carbon Tube
Carbon nanotubes are derived from a planar graphene sheet (Figure 1), which, alone, is
composed of sp2 hybridized carbon atoms arranged with 6 point group symmetry.
The overlap of the unhybridized pz orbitals yields a complex both above and below the
plane containing the atoms, which is related the high electron mobility and high electrical
conductivity of graphene. In that manner, a single-walled carbon nanotube can be described
as a rolled up graphene sheet that is closed at each end with half of a fullerene. A nanotube is
usually characterised by its diameter dt and the chiral angle (0 30 ). The chiral
vector Ch is defined with two integers (n,m) and the basis vectors of the graphene sheet
(Dresselhaus et al. 1995):
Ch = na1 + ma2 ≡ (n,m) (1)
The chiral angle is the angle between the chiral vector Ch and the so-called ―zi gzag‖
direction (n,0). The integers (n,m) determine dt and . Since there are different ways to wrap
carbon nanotubes, two types of achiral nanotubes can be achieved. These special classes of
nanotubes are the so-called ―armchair‖ nanotubes (n,n) and the ― zigzag‖ nanotubes (n,0). All
chiral‖nanotubes (n,m) with n m and m 0 (Figure 1). The rectangle that is
the others are ―
formed with the translation vector T and the chiral vector Ch define the unit cell of a nanotube
that can be translated in only one direction. Whether a nanotube is a conductor or a
semiconductor is determined by its chirality. If n-m = 3p, where p is an integer, then the
nanotube is a metal and otherwise it is a semiconductor.
Figure 1. Definition of the unit cell of a carbon nanotube. OO‘ defines the chiral vector C h = na1 + ma2.
Translator vector, T, is along the nanotube axis and perpendicular to C h. a1 and a2 represent the two
lattice translation vectors.
SWCNTs are often found in bundles that are formed by a triangular arrangement of
individual SWCNTs. The nanotubes are held together by weak van der Waals forces. On the
other hand, MWCNTs are nanotubes with more than one graphene cylinder nested one into
another.
Morphological Variations of Carbon Nanotubes

While a SWCNT can be described as a single tube of graphite, MWCNTs are made of
more complex structures and present several morphological variations (Wildgoose et al.
2006). Among these, the most commonly reported are the ―hol low tube‖, ―her ringbone‖ and
―bam boo-like‖ forms, which are consecutively depicted in Figure 2(a)-(c).
Banks et al. 2006 have suggested and presented experimental evidence that, similar to a
graphite electrode, the end of the tubes can be described as ―ed ge-plane-like‖ while the tube
walls themselves are ―bas al-plane-like‖ (Figure 2(d)). Actually, herringbone and bamboo-like
forms are expected to possess a high density of these edge-plane-like defects because, in both
cases, the plane of the graphite sheet is at an angle to the axis of the tube (Banks et al. 2006).
Most of the reports dealing with electrochemical sensors based on carbon nanotubes
compare the electrochemical activity of the nanotube-modified electrode with glassy carbon
(GC) electrodes, which do not present edge-plane-like defects, thus, leading to

misinterpretations about the original activity of the carbon nanotubes.
Figure 2. Illustration of morphological variation for forms of carbon nanotubes, (a) hollow-tube, (b)
herringbone, and (c) bamboo-like. (d) Representation of edge-plane-like defects and the basal plane in a
single MWCNT.
Intrinsic Chemical and Physical Properties
Because of the non-polar nature of their bonds, carbon nanotubes are insoluble in water.
This hydrophobicity stems both from the more positive enthalpy of forming weak SWCNT-
water hydrogen bonds, as compared to strong water-water hydrogen bonds, and from the
decreased entropy of water molecules at the non-polar SWCNT surface. SWCNTs can be
made to form stable suspensions in certain organic solvents like toluene, dimethyl formamide
(DMF) and tetrahydrofuran (THF), but they are generally insoluble in any medium without
chemical modification or coordination with surfactants.
Nanotubes are subject to the rules of carbon chemistry, which means that they can be
covalently functionalized. While they are not especially reactive, SWCNT respond well to
strong acids and other chemical oxidizers that are believed to attach functional groups (e.g.,
hydroxyl and carboxyl groups) to the side walls. Direct fluorinations of side walls have also
been achieved. Subsequent substitution reactions with alkyl-lithium or Grignard reagents, for
example, provide a route to a more complex derivatization. As might be expected, the more
highly strained caps at the ends of SWCNT contain the most reactive carbon atoms, and
smaller diameter SWCNTs are more reactive than larger ones (Abiman et al. 2008, Leventis
et al. 2005, Bahr et al. 2001).
Important to its derivatization is the fact that each carbon atom on a nanotube is
accessible to both the interior and exterior chemical environments. In this way, a SWCNT has
an extremely good surface area to volume ratio (where the volume of the lumen must be
excluded because it contains no atoms belonging to the molecule). When coupled with its
excellent van der Waals physisorption properties, this attribute makes nanotubes a natural
candidate for gas filtration, sensing, and energy storage applications (Peng et al. 2001, Li et
al. 1996, Zhang et al. 1999, Heller 2004).
SWCNTs exhibit excellent thermal stabilities in inert atmospheres. They are routinely
vacuum annealed at temperatures up to 1200 oC and can survive temperatures in excess of
1500 oC, although in the latter case the coalescence of several small tubes into larger diameter
SWCNTs has been reported. Like graphite, nanotubes burn when heat-treated in air or a
similar oxidizing environment. Small diameter tubes burn at lower temperatures than larger
diameter tubes due to the difference in strain energy (Smith and Luzzi 2004, Thien-Nga et al.
2002).
Intrinsic Electrical Properties of Carbon Nanotubes
Electric transport in carbon nanotubes is still controversial. The scattering properties of

the metallic sub-bands, i.e., the central two sub-bands of metallic tubes, and the
semiconducting sub-bands (those bands that do not cross the Fermi level of undoped metallic
tubes) are very different. While superconductivity should not be expected in low dimensional
quasi 1D system due to the low density of states (DOS), it has been observed in different
types of carbon nanotubes (Sasaki et al. 2007). On the other hand, ballistic transport in single-
walled carbon nanotubes with a diameter of 1.4 nm was theoretically estimated and has been
reported in the literature (Berger et al. 2002, Bachtold et al. 1999).
Unlike large diameter multi-walled carbon nanotubes, which are essentially two-
dimensional conductors, single-walled carbon nanotubes are considered as ideal one-
dimensional systems. In addition, it is strongly believed that when the Fermi level crosses the
semiconducting sub-bands in doped metallic SWCNTs, the two metallic sub-bands provide
the main contribution to the electrical transport. In MWCNTs the situation becomes more
complicated due to the chiralities of the different shells constituting the nanotube. It means
that, for a MWCNT presenting several shells with the same chiral angle, the periodicity along
the nanotube axis enables one to commpensate them. However, experiments have
contradicted this assumption showing that only the outer layer in MWCNTs conduct (Berger
et al. 2002).
Why Do Their Characteristics Make Carbon Nanotubes Attractives to

Electroanalysis?
The electrochemical sensitivity of any electrode is mainly affected by the noise due to the
capacitive current at the electrode/electrolyte interface and thus is proportional to the surface
area of the electrode and the specific capacitance at the interface. In voltammetry
experiments, for example, the peak of the current (ipeak) of the redox signal is the sum of two
terms: a linear diffusion and a nonlinear radial diffusion, according to:
Do Do
i peak nFACo * nFACo * (2)
t r
where the terms of the equation have their usual meanings and the first term is proportional to
the electrode surface area and decays to zero over time while the second term is proportional
to the inverse of the electrode radius and represents the steady state current due to a constant
flux of material to the surface (Bard and Faulkner 2001, Li and Mayyappan 2006). As it can
be clearly seen, signal to noise may be improved by 1000 times if the radius is reduced from
20 m to 20 nm and, in this case, the second term of equation 2 predominates over the first
one.
At macroelectrodes, the electron transfer rate, s, is calculated using the Nicholson
method. In this way, charging current effects often distort the cyclic voltammetric curve
leading to erroneous values of s. Since mass transport increases with decreasing electrode
size, the electron transfer in nanoelectrodes can take place faster than in the case of
heterogeneous charge transfer. This means that the reaction is kinetically controlled and the
electron transfer rate can be determined from the potential shift of the curves without any
interference from charging current (Oldham and Zoski 1988, Oldhan et al. 1989).
Ultrafast electrode processes with time constants as short as 1 ns might also be reached,
because the response time ( ) is also a function of the electrode dimension, according to
equation 3, where the term refers to the conductivity of the electrolyte.
rC 0 / 4 (3)
This suggests that the rates of electrode reactions could be investigated up to the diffusion
jump limit (Engstrom et al. 1986). Some general characteristics of carbon nanotubes as
nanoelectrodes are shown in Table 1.
Table 1. General characteristics and benefits of CNTs as nanoelectrodes
Electrochemistry with Nanoelectrodes Observed Characteristics

Steady-state current in a Faradaic process is
Limiting current
reached rapidly.
Small ohmic drop, which enables electrochemistry
Electrode/solution interface
in highly resistive media.
Due to the small size very small volumes are
Sample volume
needed.
Signal/noise ratio Faradaic-to-charge current ratio is very high.
Mass transport in nanoelectrodes is higher than
Mass transport
that in macroelectrodes.
Fast electron transfer processes Time constant can be as short as 1 ns.
Detection limit Lower due to the higher signal/noise ratio.
APPLICATION OF CARBON NANOTUBES IN ELECTROANALYTICAL

METHODS
Today, CNT have received significant attention for the preparation of electrochemical
sensors owing to their intrinsic properties, such as high surface area and high chemical and
electrical properties. Various kinds of CNT have been used to build novel electrodes with
interest for electroanalytical applications. Gas sensors and biosensors with immobilized
biomolecules are examples of these applications of CNT. Their application in voltammetric
methods is especially favorable, but they are also employed for sorption of different analytes
and in electrochemical stripping methods. In the field of gas sensor preparation the CNTs
have been successfully applied for determination of nitrogen dioxide, amnonia, hydrogen and
inorganic vapors general (Sayago et al. 2007, Nguyen et al. 2007, Valentini et al. 2004).Their
applications are based on changes in the electrical properties of CNTs, as a result of their
interactions with analytes. Owing to the properties of CNTs as sorbents of gases, some papers
have developed systems for removal of nitrogen oxides from the burning of fossil fuel (Herbst
et al. 2004).
Recently, Wang et al. 2007 synthesized composites of MWCNTs grafted with polymers
for sensing organic vapors of methanol, chloroform and tetrahydrofuran. Santhosh et al. 2007
developed an amperometric sensor based on multi-wall carbon nanotube (MWCNT) grafted
polydiphenylamine (PDPA), MWCNT-g-PDPA, for the determination of carbon monoxide
(CO). The MWCNT-g-PDPA-ME exhibits high sensitivity for the oxidation of CO in a
0.5 mol L-1 HClO4 solution. The dependence of the response current on CO concentration was
explored under optimal conditions and an excellent linear concentration range between 10 and
200 ppm (correlation coefficient r = 0.9941) with a significantly low detection limit of
0.01 ppm was obtained. A novel highly selective gas sensor was recently designed by
chemical modification of multi-walled carbon nanotubes containing carboxyl groups
(MWCNT-COOH) with poly(ethylene glycol) (PEG) in the presence of N,N-
dicyclohexylcarbodiimide (DCC) (Niu et al. 2007). The resistance responsiveness of the film
samples against various organic vapors, such as chloroform and acetic acid was investigated.
The use of carbon nanotubes for the design and fabrication of sensor chips also has
gained attention. Chavali et al. 2008 fabricated an active wireless UHF (433 MHz) RF-
powered sensing system for monitoring volatile organic compounds. The sensor chip
developed consisted of a thin film of gas-responsive composite material, based on modified
multiwalled carbon nanotubes (m-MWCNTs) and polypyrrole (Ppy), coated over two comb-
like interdigitated gold electrodes. The feasibility of the sensor was evaluated for detecting
the volatile anesthetic agent, fluoromethyl 2,2,2,-trifluoro-1-(trifluoromethyl) ethyl ether
(sevoflurane). Another approach of using carbon nanotubes as sensing sensor for gases relates
to the development of novel hybrid materials. Ionescu et. al. 2008 prepared a hybrid material
by means of metal-decorated multiwall carbon nanotubes (MWCNTs) dispersed on nano-
particle metal oxides. The sensor was successfully applied in the detection of different
hazardous species, specifically NO2, CO, C6H6 and NH3, at low operating temperature.
Regarding biosensors these devices are promising applications of CNTs, because this
material can enhance the electrochemical reactivity of important biomolecules and can
promote the electron-transfer reactions of proteins at low overpotentials (including those
where the redox center is embedded deep within the glycoprotein shell) (Shapter et al. 2003,
Yu et al. 2003). In addition to enhanced electrochemical reactivity, CNT-modified electrodes
have been shown useful to accumulate important biomolecules (Wang et al. 2003) and to
alleviate surface fouling effects (Musameh et al. 2002). The remarkable sensitivity of CNT
conductivity to surface adsorbates permits the use of CNT as highly sensitive nanoscale
sensors. There are over 200 dehydrogenases and 100 oxidases. Many of these enzymes
specifically catalyze the reactions of clinically important analytes (e.g., glucose, lactate,
cholesterol, amino acids, urate, pyruvate, glutamate, alcohol, hydroxybutyrate) to generate the
electrochemically detectable products NADH and hydrogen peroxide. Similar sensitivity and
stability improvements have been illustrated for electrochemical biosensors based on other
enzymes, including tyrosinase, peroxidase, organophosphorous hydrolase or alkaline
phosphatase. There are different ways for confining CNT onto electrochemical transducers.
Most commonly this is accomplished using CNT-coated electrodes (Musameh et al. 2002,
Wang et al. 2003, Luong et al. 2004) or using CNT/binder composite electrodes (Wang and
Musameh 2003, Rubianes and Rivas 2003). In this later approach, which is the simplest for
building biosensors, a variety of binders, like mineral oil, Teflon or epoxy resins can be
employed to produce CNT pastes or composites, with rigid epoxy based CNT composites
being less exploited.
CNT-based biocomposite electrodes are attractive avenues for preparing CNT-based
amperometric enzyme electrodes involves CNT/insulator/enzyme biocomposites (Wang and
Musameh 2003, Rubianes and Rivas 2003). Conventional carbon-paste composites have been
widely used for the design of renewable amperometric enzyme electrodes (Gorton 1995). A
composite electrode based on mixing MWCNTs and bromoform as a binder was prepared by
Britto et al. 1996. Carbon nanotube paste enzyme electrodes were prepared by mixing CNT
with mineral oil (Rubianes and Rivas 2003). Such composite electrodes combine the ability of
carbon nanotubes to promote electron-transfer reactions with the attractive advantages of
paste electrode materials. The preparation of a binderless biocomposite based on mixing an
enzyme (GOx) with CNT has been reported (Wang and Musameh 2003). The resulting
biocomposite was packed within a needle and was used as a microsensor for glucose. Another
simple avenue for preparing effective CNT-based electrochemical biosensors involves the use
of CNT/Teflon composite materials (Wang and Musameh 2003). These biocomposite devices
rely on the use of CNT as the sole conductive component of the transducer rather than
utilizing it as the modifier in connection to another electrode surface. The bulk of the
resultingCNT/Teflon electrodes serve as a ―r eservoir‖ of the enzyme.
Biosensors for monitoring glucose are one of the most reported. A glucose biosensor
based on the same rigid and renewable carbon nanotubes (CNT) was recently reported
(Merkoçi 2006). The biosensor was based on the immobilization of glucose oxidase (GOx)
within the CNT epoxy-composite matrix prepared by dispersion of multi-wall CNT inside the
epoxy resin. The use of CNT, as the conductive part of the composite, ensures better
incorporation of enzyme into the epoxy matrix and faster electron transfer rates between the
enzyme and the transducer. Experimental results showed that the CNT epoxy composite
biosensor (GOx-CNTEC) offers an excellent sensitivity, reliable calibration profile and stable
electrochemical properties together with significantly lower detection potential (+ 0.55 V)
than GOx-graphite epoxy composites (+0.90V; difference ΔE = 0.35 V). The results obtained
favorably compare to those of a glucose biosensor based on a graphite epoxy composite
(GOx-GEC).
Timur et. al. 2007 showed the used of Pseudomonas putida DSM 50026 cells as the
biological component with measurement based on the respiratory activity of the cells
estimated from electrochemical measurements. The cells were immobilized on carbon
nanotube (CNT) modified carbon paste electrodes (CPE) by means of a redox osmium
polymer, viz. poly(1-vinylimidazole)12-[Os-(4,4′-dimethyl-2,2′-dipyridyl)2Cl2]2+/+. A
microbial biosensor was also prepared by using phenol adapted bacteria and then calibrated
for phenol. After that, it was applied for phenol detection in an artificial waste water sample.
Pereira et al. 2007 evaluated the performance of MWCNTs as transducer, stabilizer and
immobilization matrix for the construction of amperometric biosensors based on lactate
dehydrogenase (LDH) and Meldola's Blue (MB). The enzyme was immobilized onto the
MWCNT adsorbed with MB by cross-linking with glutaraldehyde. The sensor was applied
directly in measurements of lactate in blood samples.
Santos et al. 2006 in similar work developed a amperometric biosensor based on
immobilization of alcohol dehydrogenase (ADH) and Meldola!s Blue (MB) in MWCNTs for
the determination of ethanol in a great variety of alcoholic beverages. A biosensor based on
carbon nanotube modified glassy carbon (CNT/GC) was used for enhancing the sensitivity of
electrochemical measurements of enzymatically generated thiocholine (Liu et al. 2005). The
highly sensitive electrochemical detection of enzymatically generated thiocholine with this
CNT sensing platform holds great promise for preparing an acetylcholinesterase biosensor for
monitoring organophosphate pesticides and nerve agents.
Another alternative for confining biomolecules on the surface of electrodes modified with
CNTs can also be by covalent immobilization. Kumar et. al. 2008 investigated the covalent
immobilization of SWCNTs on chemically functionalized glass surfaces. According to the
authors, the covalent immobilization of SWCNTs on glass has an important role in fabrication
of carbon nanotubes-based electrodes for biosensor and bionanotechnology applications.
The detection of NADH via electrocatalytic oxidation at single-walled carbon nanotubes
modified with Variamine blue has been proposed by Radoi et. al. 2008. The authors prepared
screen-printed electrodes (SPEs) modified with Variamine blue (VB), covalently attached to
the oxidized single-walled carbon nanotubes (SWCNTs-COOH). The Variamine blue redox
mediator was covalently linked to the SWCNTs-COOH by N,N′-dicyclohexylcarbodiimide
(DCC) and N-hydroxysuccinimide (NHS) chemistry. This chemical sensor was used for the
detection of the reduced nicotinamide adenine dinucleotide (NADH). The preparation of
chitosan film containing SWCNT (CHIT–SWCNT) has been proposed by Tkac et. al. 2007.
The stabilization of the film was done by chemical crosslinking with glutaraldehyde and the
free aldehyde groups produced a substrate used for covalent immobilization of galactose
oxidase (GalOD). This new galactose biosensor offers highly reliable detection of galactose
with RSD well below 2% and it has been successfully applied for assaying galactose in blood
samples with recovery indices between 101.2 and 102.7%.
As regards CNT-coated electrodes, the drawback for developing of these devices is the
insolubility of CNT in most solvents. Earlier reported CNT-modified electrodes have thus
commonly relied on casting a CNT/sulfuric acid solution onto a glassy carbon surface
(Musameh et al. 2002). Nafion films have been used extensively for the construction of
amperometric biosensors owing to their unique ion-exchange, discriminative and
biocompatibility properties. The ability of Nafion to solubilize CNT provides a useful avenue
for preparing CNT-based electrode transducers for a wide range of sensing application (Wang
et al. 2003). The resulting biosensors greatly benefit from the coupling of the
antifouling/discriminative properties of Nafion films with the efficient electrocatalytic action
of CNT towards hydrogen peroxide. Further improvements in the electroactivity of hydrogen
peroxide (with detection limits down to 25 nmol L-1) can be obtained by dispersing platinum
nanoparticles in the Nafion/CNT coating (Hrapovic et al. 2004).
The application of CNTs in some methods using stripping voltammetry has also been
reported, based on the strong sorption properties of these materials. In stripping
measurements of several organic analytes, the first step of determination (sorption of
analytes) has been carried out by an open circuit on electrodes modified with CNTs. In the
next step, the electrochemical oxidation or reduction of accumulated analyte is carried out.
These procedures have been developed for anodic determination of xanthine (Zhao et al.
2002), 6-benzylpurine (Zhao et al. 2003) and fluphenazine (Zeng and Huang 2004), as well as
for determination of 4-nitrophenol based on a very sensitive, well-defined reduction peak at
the SWCNT-modified GCE (Yang 2004). In the first stage of voltammetric stripping
determination of transition metal cations with electrodes modified with MWCNTs, cations
have been adsorbed from solutions containing iodide, then reduced at -0.6 V (Hg2+) or -1.2 V
(Cd2+ and Pb2+), and, in the last step, anodic stripping has been employed (Yi 2003, Wu et al.
2003). Such methods are alternatives to conventional anodic stripping voltammetry using
thin-film mercury electrodes. The stripping voltammetry of Cd(II) was also reported using a
GCE modified with an MWCNT/Nafion composite film (Tsai et al. 2004). Conventional
anodic stripping voltammetry of Cd(II) with a GC disk electrode has been used to detect
DNA hybridization based on CNTs loaded with CdS tags (Wang et al. 2003).
APPLICATION OF CARBON NANOTUBES IN SOLID PHASE

EXTRACTION
Considering the interesting properties of CNTs and the compatibility of these properties
with the fields of separation sciences, CNTs are used as stationary phases both in solid phase
extraction and chromatography. The following paragraphs demonstrate some techniques and
different applications of CNTs that have been reported in recent years on this area.
Solid Phase Extraction (SPE) and Chromatography
Solid phase extraction (SPE), sometimes called accumulation columns, has been an
important tool for the analytical sciences since 1970 (Fritz 1999) and many practical
applications are described in the literature, such as for environmental, pharmaceutical,
chemical and biological analyses. Considerable attention has been paid to SPE as a way to
isolate and concentrate desired components from a sample matrix. In addition to being fast,
efficient and easily automated, SPE is a clean analytical procedure (Fritz et al. 1995). This
technique has a simple principle regarding the selective retention of the desired analyte from a
liquid phase by a solid stationary phase and the control of these retention properties based in
an equilibrium (K) concept, where:
amount in the solid phase

K
amount in the liquid phase
A high value of K indicates strong retention and a lower value of K suggests low
retention.
Different from SPE, chromatography is performed to separate different molecules
contained in the same sample. Snyder and Kirkland 1979 defined the differential migration of
various compounds (solutes) in the original sample, and a spreading along the column of
molecules of each solute as two features of chromatographic separation. The Figure 3 (a)
shows a hypothetical separation in a column of three types of analyte and their respective
chromatogram (b).
Nowadays, chromatographic separations with both liquid and gaseous mobile phase are
widely used as analytical tools for applications such forensic, petroleum, pharmaceutical and
chemical determinations. In other words, the demand is large and great attempts have been
focused for the development of new techniques to obtain considerable advances in this
separation field.
Concerning SPE and chromatographic separation techniques, we will emphasize the use
of CNTs as a stationary phase in both cases.
Carbon Nanotubes Used in Solid Phase Extraction (SPE)
SPE offers a number of important benefits in comparison with laborious classical liquid–
liquid extraction, such as reduced solvent usage and exposure, low disposal costs and short
extraction times for sample preparation (Pyrzyńska 2007). Several types of solid compounds
have been used as stationary phases for this technique, with silica-based and polymer-based
phases being the most important, used in either the normal or the reversed phase mode. The
two most practical significant applications of SPE are clean-up/purification and concentration
(Cruz-Vera et al. 2008).
In the first case, when the matrix effect is considerable, the use of solid phase extraction
is the first choice when compared with liquid-liquid extraction, due to its versatility based on
specificity, recovery of the analyte, absence of emulsion, large interfacial area of the particles
(about 200-800 m2/g) (Fritz 1999) and others.
Figure 3. The hypothetical separation of three distinct molecules in a liquid chromatographic colums (a)
and the chromatogram obtained after their detection.
The use of SPE as a preconcentration method has been increasing since 1970, and is one
of the most studied applications of CNT. It consist basically on passing a large quantity of
sample through a column containing a solid phase, which, due to the very low concentration
of the analyte in the raw sample, can not be analyzed by simple techniques without this
preconcentration step. This technique has been successfully applied for various metal ions in
different kinds of samples (Pyrzyńska 2007). Moreover, sorbent extraction can be used for
selective retention of some particular chemical forms of a metal, thereby enabling speciation
(Campanella et al. 1996, Pyrzyńska et al.1999). Environmental samples and trace analyses
samples are good examples of the application of this resource. To illustrate, some applications
will be described.
Another advantage of CNT is their ease of handling to prepare the column in the
laboratory. In general the column is made using two conical tips with quartz wool placed at
each end to keep the CNT inside or by using polypropylene syringe tubes with polyethylene
frits to hold the adsorbent.
The functionalization of the carbon nanotubes nowadays contributes so much to increase
its possible applications of separation techniques. The literature describes different types of
functionalizations for distinct functions.
Metal Ions
CNT have been exploited for their interactions with metal ions and the possibilities
generated for separation techniques. This has resulted in an enormous quantity of applications
demonstrating by metal ion preconcentration.
Gil et al. 2007 recently demonstrated the ability of MWCNTs as a substrate for the on-
line preconcentration and speciation of vanadium both: V(IV) and V(V) prior to
determination by electrothermal atomic absorption spectrometry (ETAAS). In this paper, the
authors used flow injection analysis (FIA) coupled with ETAAS and found good recovery
levels of both vanadium species in drinking water at the ng L-1 level. Moreover, with the
speciation technique using the masking reagent CDTA (1,2 cyclohexanediaminetetraacetic
acid) for V(IV), it was possible to quantify selectively both species. This method is based on
the non-retention of the complex V(IV)-CDTA by the MWCNTs. The optimum concentration
of CDTA that inhibits retention of V(IV) was 1.0 x 10-4 mol L-1 at pH 4.0. This pH was
determined by experiments, in which sample solutions were adjusted over the range 1.0 – 9.0
and between pH 3.5 – 5.0 adsorption close to 100 % was achieved.
The interferences on vanadium adsorption by MWCNTs by common coexisting ions
(cations and anions) in waters was studied and it was demonstrated that in the most of cases
the tolerance level was higher than the concentrations in which these ions commonly occur in
the sample to be analyzed. It was concluded that these ions had no influence on the retention
properties of MWCNTs in this study.
The role of pH on extraction is very important, as described by Liang et al. 2005, due to
the effective control of the adsorption by the MWCNTs. It can be justified by the
functionalization that has occurred on the surface of the carbon structures producing oxygen-
containing groups when submitted to oxidation. This oxidation leads to a reduction of the
isoeletric point (IEP) of the MWCNTs to lower pH values. Under these conditions the sample
pH needs to be higher than the IEP of the oxidized MWCNT. This is justified by the negative
charge on the surface that provides electrostatic attraction that is favorable for adsorbing
cations. In other papers, if the sample has a lower pH, the negative charge is neutralized and
consequently the retention of cations for the MWCNTs decreases quickly. Nitric acid is
widely used as the reagent responsible for MWCNT oxidation. Data has demonstrated this for
preconcentration of different ions (El-Sheikh 2008). The need of oxidation of the column
prior to the extraction process was shown because non-oxidized structures show an
accentuated decrease on sensitivity, consequently increasing the detection limits and reducing
the linear ranges. The pH influence was also discussed and another theory of retention was
proposed concerning the interaction of metal hydroxides, which are formed in basic pH, with
functional groups on the adsorbent surface (carboxyl, phenolic, lactonic) present in their
deprotonated forms.
Stafiej and Pyrzyńska 2007 carried out a study that corroborates these theories about pH
and surface charges by establishing a relationship among sorption level with ion
electronegativity. The affinity order of metal ions toward MWCNTs at pH in the range 7.0 –
9.0 was Cu(II)>Pb(II)>Zn(II)>Co(II)>Ni(II)>Cd(II)>Mn(II). In other words, the relation
between metal ion electronegativity and sorption level is directly proportional. However, this
relation must be accepted only in cases when the cation is sorbed in its ionic form, not in a
complexed form.
In this context a work of Tuzen et al. 2008 may exemplify the alteration observed in the
sorption behavior of MWCNTs. When the cation is chelated with ammoniumpyrrolidine
dithiocarbamate (APDC) before extraction, some of main parameters such as the pH of the
sample that was found to give good recoveries was in the 2.0 – 6.0 range and decreased after
pH 7.0. This can be explained by the APCD pKa value (3.29) that supports better results in an
acidic medium. According to work performed by Tuzen et al. 2008, excellent quantitative
recoveries were found in environmental samples for the determination of Cu(II), Cd(II),
Pb(II), Zn(II), Ni(II) and Co(II) on MWCNTs.
Tuzen and Soylak 2007 used a similar procedure of speciation with APDC to concentrate
and quantitatively determine chromium (VI) and total Cr in environmental samples (river
water). They proved experimentally that Cr retention by MWCNTs must be mediated by
APDC complexation and it is concentration-dependent. Good results were found in this study
for detection limit (0.9 µg L-1) with a preconcentration factor of 100-fold. So, they suggested
also that this method is superior to other recent studies about Cr speciation due to its
selectivity, detection limit, applicable pH range, capacity of adsorption (9.5 mg g-1) and,
additionally, no consumption of organic solvents.
Normally much attention of this research is devoted to comparison of MWCNTs with
other kinds of sorbent, in which the MWCNT show better features than conventional
sorbents. However, it is possible to observe an increase in the number of studies that compare
carbon nanotubes with themselves. Recently, Lu and Chiu 2008 compared the efficiency of
MWCNTs on Zn2+ adsorption after distinct oxidization process, where they submitted the
carbon nanotubes to oxidation separately with HNO3/H2SO4, HNO3, KMnO4 and NaClO.
Improvements of the physicochemical properties of MWCNTs were observed, mainly on its
surface where a negative charge was generated increasing the adsorption of Zn2+. Among the
oxidation agents tested, NaClO was the most effective. However, it important to point out that
this subject needs to be explored case by case, because each one has its particularities,
depending of the element to be concentrated and on the characteristics of sample. In another
work, Xiao et al. 2007 suggested KMnO4 as the oxidizing agent.
As is well known, carbon nanotubes possess different dimensions when purchased and
these dimensions exert an important role on nanotube efficiencies. Taking into account this
factor, El-Sheikh et al. 2007 studied this effect on MWCNTs enrichment efficiency for metal
ions. Briefly, they suggested that long MWCNTs (5-15 µm) are better than short MWCNTs
to concentrate metals. Regarding external diameter, the best enrichment efficiency was found
within 10-30 nm.
Based on the principle of silica-bonded stationary phase, some authors have been
applying similar procedure to use the MWCNTs as a support for different ligands. In a recent
work, Amais et al. 2007 synthesized a nanocomposite based on alumina (Al2O3) supported on
MWCNTs to be applied as a sorbent for nickel preconcentration prior to flame atomic
absorption spectrometry (FAAS). Significant results mainly on the recovery studies (higher
than 95%), stability of the column for more than 200 cycles without loss of sorption capacity
and a high analytical frequency due to the high flow used in the preconcentration step were
achieved.
Some procedures based on biosorption can be found in the literature. Baytak and Turker
2005 used this resource to determine Fe (III), Co (II) and Cr (III) in water samples by using a
preconcentration system with Saccharomyces carlsbergensis immobilized on amberlite XAD-
4 and in a similar work (Baytak and Turker 2005) they got to preconcentrate Fe (III), Co (II),
Mn (II) and Cr (III) using as a solid-phase extractor an structure composed by Agrobacterium
tumefacients immobilized on Amberlite XAD-4. Tuzen et al. 2008 describes the use of the
well known pathogenic bacteria Pseudomonas aeruginosa immobilized on the MWCNT
surface as a biosorbent for the determination of Co (II), Cd (II), Pb (II), Mn (II), Cr (III) and
Ni (II) in distinct kinds of samples, such as tomato leaves, bovine liver, boiled wheat, canned
fish, black tea, lichen and natural water samples.
Organic Compounds
Applications of carbon nanotubes as SPE for organic compounds have been widely
reported for several types of molecular classes like mycotoxins, phenols, herbicides,
fungicides, pollutants and pharmaceuticals, among others.
Yu and Lai 2007, associating MWCNTs with another interesting analytical technique
called molecularly imprinted polymer (MIP), designed a micro solid-phase extraction (µSPE)
system to determine ochratoxin A, a type of mycotoxin, in red wine by multiple pulsed

elutions coupled to high performance liquid chromatography (HPLC) with fluorescence
detection (FLD). Different from the majority of the SPE techniques that use a mini-column
packed with the solid phase, this work consisted in synthesizing inside a stainless steel frit
with a defined pore size of 0.5 µm a film of combined molecularly imprinted polypyrrole
(MIPPy) and MWCNTs to be used as a stationary phase. The metallic composition of the frit
had an important role on the synthesis of MIPPy-MWCNT film because the polymerization
process was inducted by an electrochemical method in which a constant potential of +0.85 V
was applied on the frit by a potentiostat.
Several applications of CNT in the analysis of pesticides like fungicides and herbicides
have been reported, especially in that cases where the analyte has a phosphoric group, that
shows a good affinity for the MWCNT surface. In a recent paper Du et al. 2008 demonstrated
a different system of determination based on SPE for nitroaromatic organophosphates
(NAOP) by using MWCNTs as a sorbent phase to retain and subsequently determine methyl
parathion (MP), a kind of NAOP widely used around the world and which is responsible for a
lot of cases of severe human intoxication. Many SPE processes using CNT for some classes
of pesticides have already achieved excellent results concerning figures of merit, like limits of
detection and quantification, precision and linear range. Moreover, good rates of enrichment
and recovery were showed by carbon nanotubes. The determination of cyanazine,
chlorotoluron, and chlorbenzuron, belonging, respectively, to the triazine, phenylurea and
benzylacylurea pesticide groups was described by Zhou et al. 2007-a. The same authors, in
other work (Zhou et al. 2007-b), describe a sensitive and simultaneous determination at trace
levels of four fungicides (metalaxyl, diethofencarb, myclobutanil and tebuconazole) and
prometryn, another triazine herbicide. Pyrzyńska et al. 2007 evaluated the sorption behavior
of acidic herbicides (3,6-dichloro-2-methoxybenzoic acid and 2,4,5-trichlorophenoxyacetic
acid) by CNT and confirmed a greater herbicide adsorption capability than that shown by
graphitized carbon black (GCB) and bonded C18 silica. Comparing MWCNTs and C18 silica
for enrichment power Zhou et al. 2007 found interesting results that demonstrated a higher
efficiency of MWCNTs in comparison with C18 silica in environmental water samples, thus
indicating the effectiveness of MWCNTs in clean-up process of complex matrices.
In the pharmaceutical field some works apply CNT as a tool, such as that developed by
Wang et al. 2006, in which four benzodiazepine residues (diazepam, estazolam, alprazolam
and triazolam) were determined in pork, and another food of animal origin food, after clean-
up by SPE with a CNT packed cartridge.
DEVELOPMENT OF NEW ANALYTICAL METHODS:

SOME CASE STUDIES
As application examples, the potential of using carbon nanotubes as an advantageous
material for the development of analytical methods, two methods will be described. The first
comprises the development of a sorbent flow preconcentration system for cobalt
determination coupled to UV/Vis spectrophotometry while the second emphasizes the
potentialities of using carbon nanotubes as an electrode in potentiometric stripping analysis
for antimony determination.
Spectrophotometric Flow Injection System Using Mwcnts as Solid

Concentrator for Cobalt Determination
Trace metal determination currently has been performed in association with solid-phase
preconcentration/separation techniques, basically aiming at the enrichment of metallic species
and/or matrix elimination. These techniques have gained special importance in analyses of
complex matrices owing to some advantageous features, such as higher preconcentration
factor, simplicity, easy coupling to flow injection analysis (FIA), better repeatability, high
sample throughput and easy regeneration of the solid-phase (Gama et al. 2006). Basically, a
solid sorbent can be employed as a chelate forming functional group or as a support and the
properties of both components determine the features and the applications of the respective
materials. Hence, the choice of an effective sorbent regarding selectivity and efficiency of the
preconcentration step in the analytical method is made by taking into account the nature of
the functional groups and the physico-chemical properties of the sorbent, such as mechanical
and chemical stability, surface area, pore volume and kinetic characteristics (Bilda et al.
1998). The literature presents a variety of sorbents and the most prominent among them are
C18 silica, activated carbon and polymeric resins (Liu et al. 2004). Nevertheless, most of them
have shown some drawbacks, such as the limited breakthrough volumes and the narrow pH
stability range for C18 silica and the poor selectivity of some polymeric resins (Liu et al.
2004). Therefore, currently there is interest in the development of new solid phase extractors
with high sorption capacity, selectivity and preconcentration factors. There are numerous
attempts in this direction and recently nanomaterials have shown advantages over traditional
sorbents, mainly owing to high chemical activity since their surface atoms are unsaturated
and, therefore, can bind strongly with other atoms; large surface area and high chemical
stability. As examples, nanoparticles of titanium dioxide and alumina have been used as solid
sorbents for the enrichment of metal ions, followed by their determination by FAAS or
inductively coupled plasma optical emission spectrophotometry (ICP OES), respectively
(Liang et al. 2004, Hang et al. 2007).A literature survey showed that the MWCNTs were first
used as sorbent in off-line preconcentration mode for cadmium, manganese and nickel metals,
using ICP OES as the quantification technique (Liang et al. 2004). Recently, we published
two papers that are based on sorbent flow preconcentration using MWCNTs for cadmium and
lead, with, respectively, thermospray flame furnace atomic absorption spectrometry (TS-FF-
AAS) and FAAS for metal determination (Tarley et al. 2006, Barbosa et al. 2007). These
works have emphasized the powerful performance of MWCNT when associated to an
element-selective technique. However, there are no investigations regarding the selective
performance of MWCNTs for metal ions using flow injection preconcentration coupled to
UV-vis molecular spectrophotometry, a technique that is less selective and less expensive
than atomic absorption spectrometry. Herein, a reliable flow injection preconcentration
system coupled to UV-vis molecular spectrophotometry for cobalt determination using
MWCNTs as sorbent is described. Cobalt determination was accomplished after
preconcentration followed by elution and complexation with PAN, 1-(2-piridylazo)-2-
naphtol), a complexant more broadly applicable to heavy metals.
Apparatus and Reagents Used for the Development of the Analytical Method
A Femto UV-vis spectrophotometer (São Paulo, Brazil) model 482 equipped with a flow
cell of 1 cm optical length was used for FIA measurements. Data acquisition was carried out
from an interfaced (Advantech) PCL 711S mode and a computational program was developed
on an EXCEL® spreadsheet, using macros of Visual Basic®. An Ismatec Model IPC peristaltic
pump (Ismatec IPC-08, Glattzbrugg, Switzerland) furnished with Tygon® tubes was used to
propel all sample and reagent solutions. The preconcentration/elution steps were selected by
using a home-made injector-commutator made of Teflon® (PTFE, polytetrafluoroethylene).
Adjustment of sample pH was done using a Handylab 1 Schott pHmeter (Stafford, UK).
Multi-wall carbon nanotubes (MWCNTs) were supplied by CNT Co., Ltd. (Yeonsu-Gu,
Incheon, Korea) with > 93% purity, diameters between 10-40 nm and lengths of 5-20 m.
Prior to use, MWCNTs were submitted to acid treatment to create carbonyl and carboxyl
groups on the MWCNT surfaces that are potentially involved in metal sorption. More details
are found in the articles by Barbosa et al. 2007 and Tarley et al. 2006. All solutions were
prepared with water obtained from a Milli-Q purification system (Millipore, Bedford, MA,
USA) having a resistivity of 18.2 MΩ cm. Analytical grade chemical reagents were used
throughout.
Sorbent Flow Preconcentration System

A diagram of the flow preconcentration system is shown in Figure 4. At the
preconcentration step position, the sample, buffered at pH 5.2, is percolated through a
minicolumn of MWCNTs (50 mg) at 4.3 mL min-1 flow rate for 2 min and 33 s. After this
stage, by switching the central part of the injector-commutator, a stream of 0.50 mol L-1
HNO3 displaces the cobalt ions at 2.4 mL min-1 flow rate. Afterwards, the eluted cobalt ions
react with a 150 µmol L-1 PAN solution at the 2.4 mL min-1 flow rate. The Co(PAN)2
complex is driven to the spectrophotometer where the absorbance measurements are
continuously made at 575 nm. All absorbance signals were taken as peak height.
Optimization of Chemical and Flow Variables

The influence of the sample pH on the adsorption of cobalt onto MWCNTs was studied
within the experimental domain from 4.2 to 10.2. As can be seen in Figure 5, the best pH
values for cobalt adsorption were found to be from 4.2 to 8.0; the analytical signal decreases
gradually above pH 8.0. These results indicate the robustness of the preconcentration method
over a large sample pH range. A sample pH of 5.2 was chosen for further experiments, since
an acetate buffer at this pH presents good buffering capacity.
Figure 4. Schematic diagram in the loading position of the in line flow preconcentration system for
cobalt ions onto MWCNT with spectrophotometric determination. S = sample, P = peristaltic pump, W
= waste, RC = reaction coil. For more details, see text.
Figure 5. Effect of sample pH on the adsorption of cobalt ions onto MWCNTs. Sample volume: 11 mL,
cobalt concentration: 100 g L-1, sample concentration flow rate: 5.5 ml min-1, sample buffer
concentration: 0.01 mol L-1, Pan concentration: 100 mol L-1 in 1.25 mol L-1 (pH 9.82) ammoniacal
buffer; surfactant concentration (SDS) = 0.005 mol L-1, eluent and PAN flow rates = 2.4 ml min-1,
reaction coil (RC) = 120 cm.
In order to evaluate the effect of sample buffer concentration, assays were carried out
with CH3COOH/CH3COO- buffer concentrations from 0.0051 to 0.05 mol L-1. Figure 6
shows the increase in buffer concentration decreases the adsorption of cobalt onto MWCNTs,
probably due to competition of buffer for the adsorbent sites. A buffer concentration of 0.005
mol L-1 was selected as the best condition.
Figure 6. Effect of sample buffer concentration on the adsorption of cobalt ions onto MWCNTs.
Sample volume: 11 mL, cobalt concentration: 100 g L-1, sample pH: 5.2, sample preconcentration
flow rate: 5.5 ml min-1, surfactant concentration (SDS) = 0.005 mol L-1, eluent and PAN flow rates =
2.4 ml min-1, reaction coil = 120 cm.
The effect of the sample preconcentration flow rate was examined in the range 1.5-8.0 ml
-1
min . As shown in Figure 7, the analytical signal decreases with increasing flow rate. This
behavior was expected, considering the solid phase extraction process and allows concluding
that the adsorption kinetics of cobalt is slow. By employing a lower preconcentration flow
rate (1.5 ml min-1), the required time for concentrating 11 ml of sample would be more than 7
min, an unusual and unsatisfactory condition in an flow system. Therefore, in order to
maintain a comprise between sensitivity and sample throughout, the preconcentration flow
rate was set at 4.3 mL min-1, being carried out during 2 min and 33 seconds.
It is well known that a surfactant provides better solubilization of PAN in aqueous media.
Moreover, the molar absorptivity of the complex Co(PAN)2 is high in the presence of
micellar aggregates. Thus, the influence of the nature of the surfactant was investigated using
the following surfactants at 0.005 mol L-1 concentration: CTAB (cetyltrimethyl ammonium
bromide), Triton X-100, SDS (sodium dodecyl sulphate) and Triton X-114 (Figure 8). SDS
gave the best results based on peak height. Hence, the SDS was chosen for subsequent
studies.
After establishing the importance of surfactant to the performance of the system, the
influence of its concentration was investigated from 0.005 to 0.035 mol L-1. The results
shown in Figure 9 show a slight increase of analytical signal from 0.005 to 0.015 mol L-1, as a
result of the high efficiency of solubilization of Co(PAN)2 . For surfactant concentrations
above 0.018 mol L-1, the sensitivity remained essentially unchanged, hence this value was
selected.
Figure 7. Effect of sample preconcentration flow rate on the adsorption of cobalt ions onto MWCNTs.
Sample volume: 11 mL, cobalt concentration: 100 g L-1, sample pH: 5.2, sample buffer concentration:
0.005 mol L-1, surfactant concentration (SDS) = 0.005 mol L-1, eluent and PAN flow rates=2.4 ml min-1,
reaction coil = 120 cm.
Figure 8. Effect of kind of surfactant on analytical signal. Sample volume: 11 mL, cobalt concentration:
100 g L-1, sample pH: 5.2, sample preconcentration flow rate: 4.3 ml min -1, sample buffer
concentration: 0.005 mol L-1, eluent and PAN flow rates = 2.4 ml min-1, reaction coil = 120 cm.
Figure 9. Effect of surfactant concentration on analytical signal. Sample volume: 11 mL, cobalt
concentration: 100 g L-1, sample pH: 5.2, sample buffer concentration: 0.005 mol L-1, sample
preconcentration flow rate: 4.3 ml min-1, eluent and PAN flow rates = 2.4 ml min-1, reaction coil = 120
cm.
The buffer concentration of the PAN solution is an important factor because it

demonstrates the capacity of the medium to maintain a favorable pH for the formation of the
Co(PAN)2 complex. In this process, the eluent solution pH should increase quickly to favor
Co(PAN)2 complex formation, because the acid eluent solution containing the cation merges
with the reagent solution at a T. In this work, NH4+/NH3 buffer solutions were used.
According to Figure 10, the analytical signal increased significantly with the increase in PAN
buffer concentration from 0 to 0.25 mol L-1, with only a slight variation in the analytical
signal above 0.25 mol L-1 buffer concentration being observed. Even though this value
showed a high absorbance, its buffering capacity is not favorable in the presence of the 0.5
mol L-1 nitric acid used as eluent from the preconcentration step. This result probably reveals
that the complexation reaction between Co(PAN)2 is faster than the neutralization reaction.
Despite the low difference in absorbances over 0.25 to 1.25 mol L-1 range, the best signal was
observed with a 0.75 mol L-1 PAN buffer concentration, this value being adopted throughout
the remainder of this study.
The effectiveness of Co(PAN)2 complex formation depends on the PAN solution pH. The
effect of this parameter on the analytical signal is shown in Figure 11. For this study the pH
range was varied from 8.3 to 10.20 using a 0.75 mol L-1 NH4+/NH3 buffer. The profile of the
curve obtained in Figure 11 indicates that high pH values favored complex formation in
agreement with the literature, in which the complexation reaction of cobalt ions with PAN is
favored at high pH owing to the pKa values of the ligand. Therefore, a pH 10.2 was chosen as
the best value for this study.
Figure 10. Effect of PAN buffer concentration on analytical signal. Sample volume: 11 mL, cobalt
preconcentration flow rate: 4.3 ml min-1, surfactant concentration (SDS) = 0.005 mol L-1, eluent and
PAN flow rates = 2.4 ml min-1, reaction coil = 120 cm.
Figure 11. Effect of PAN solution pH on analytical signal. Sample volume: 11 mL, cobalt
PAN solution concentration plays an important role in the analytical signal. Its
concentration must be sufficiently high to guarantee the total complexation of cobalt. The
range studied was from 4.0 to 200 mol L-1. For all values studied there is an excess of PAN,
with respect to cobalt ions. However, above 80 mol L-1 PAN the analytical signal remains
constant (Figure 12). Considering that in the optimization studies the cobalt concentration
was set at 100 g L-1, it was decided adopt a 150 mol L-1 PAN concentration for further
experiments, having in mind the construction of the analytical curve.
Figure 12. Effect of PAN concentration on analytical signal. Sample volume: 11 mL, cobalt
Characteristics of the Analytical Method

The analytical curve (duplicate measurements at each point) gave a linear range from 2.5
to at least 150 g L-1, with the use of a 2 min and 33 s preconcentration time (Figure 13). The
precision of the method, expressed as the RSD, for the determination of 15 and 30 µg L-1 Co
(II), were 1.75 and 3.33%, respectively (n =10). The enrichment factor (EF), defined as the
ratio of the slopes of the linear sections of the analytical curves before and after the
preconcentration, was 12.6. CE defines the preconcentration factor attained by a sorbent flow
concentration system during 1 min of preconcentration. As the time of the preconcentration
step was 2 min and 33 s, the CE was found to be 4.96 min−1. The consumptive index, defined
as the volume of sample in milliliters consumed to achieve a unit EF, was 0.87 ml. The
detection limit, defined as the preconcentration of analyte giving a signal equivalent to three
times the S.D. of the blank signal plus the net blank intensity (Long and Winefordner 1983),
was measured to be 0.75 g L-1. The determination rate was 20 samples per hour under the
optimal conditions.
Figure 13. Analytical curve obtained for the on-line flow preconcentration system under optimized
conditions. For more details, see text.
In order to investigate the selective separation and determination of cobalt ions from
water containing different metal ions, aliquots of aqueous solutions (11 mL) containing 1.1
µg of cobalt ions and increasing amounts of possible interferents ions were taken and
submitted to preconcentration procedure. The ratios analyte:concomitant employed were 1:1,
1:10 and 1:100 for all metals evaluated. The results were compared with cobalt
preconcentration in the absence of concomitant. When the analytical signal was changed,
positively or negatively above of 15 % the presence of interference in the procedure was
considered. The cations Cu2+ (1000 µg L-1), Zn2+ (1000 µg L-1), Mn2+ (10 mg L-1), Mg2+ (10
mg L-1), Cr3+ (100 µg L-1), Cd2+ (100 µg L-1) and Pb2+ (1000 µg L-1) interfere in cobalt
preconcentration at the levels specified. Despite the interference, it is important to stress that
the amounts of these metals are usually lower in drinking waters and natural waters. Recovery
studies were performed in mineral water and tap water by spiking different amounts of cobalt
ions. Recovery percentages varying from 93.4 to 102.0% demonstrate that the presence of
foreign ions in this sample have no effect on the recovery of cobalt ions.
Antimony Determination by Potentiometric Stripping Analysis Using

Carbon Nanotubes Paste Electrode
Electrochemical stripping techniques, including anodic stripping voltammetry (ASV) and

cathodic stripping voltammetry (CSV) have been mostly used for trace metal determination.
In stripping voltammetry a preconcentration step is combined with a stripping step, thereby
enhancing sensitivity and selectivity. During the preconcentration step, the trace metal of
interest is collected onto or in a working electrode and during the stripping step the collected
metal is oxidized or reduced back into solution, providing the measurement. Another
modality of stripping analysis is potentimetric stripping analysis (PSA) described by Jagner
and Graneli 1976. The elements that can be determined by PSA are those that can also be
determined by voltammetric stripping analysis. The deposition step in PSA is the same as in
stripping voltammetry, but the stripping step (oxidation) is different as the elements are
chemically oxidized, usually by Hg (II) ions in excess, and not by a potential sweep as in
ASV. As a consequence, PSA does away with the necessity for deoxygenation of the sample
and is little affected by the presence of surface active agents and electroactive organic matter.
The majority of data reported using potentiometric stripping procedures require metal
accumulation onto a hanging mercury drop electrode. However, these techniques require
tedious experimental precautions regarding the stability and recovery of the mercury drop
after each experiment. The potential dangers associated with mercury has led to the
development of various electrodes, such as the glassy carbon electrode coated with mercury
film modified with Nafion (Merkoai et al. 2000, Vidal et al. 1992), cellulose acetate (Wang
and Hutchins-Kumar 1986), a naphthol derivative (Esnafi and Naeeni), cysteine (Bai et al.
1998) etc. However, to the best of our knowledge, the assessment of carbon nanotube paste as
an electrode using PSA for metal determination has not been reported in the literature. The
data presented here offer an innovative methodology for antimony determination by PSA as
well as emphasizing the great potentialities of carbon nanotubes in the field of stripping
analysis.
Apparatus and Reagents Used for the Development of the Analytical Method
All electrochemical experiments were performed using a potentiostat/galvanostat
Autolab® PGSTAT-12 (Eco Chemie B.V.; The Netherlands). Experiments were performed in
a conventional single-compartment three-electrode cell. A carbon nanotube paste electrode
was employed as the working electrode. A platinum wire was employed as the auxiliary
electrode. All potentials were recorded in relation to an Ag/AgCl reference electrode. All
metal ion solutions were prepared daily by appropriate dilution of 1000 mg L-1 stock
solutions from Merck (Darmstadt, Germany). Acetate buffer solution was prepared without
further purification from acetic acid and its respective sodium salt, purchased from Merck.
Aqueous solutions were prepared with deionized water ( > 18.2 M cm, Millipore Milli-Q
system). The MWCNTs used for electrode preparation were supplied by CNT Co., Ltd.
(Yeonsu-Gu, Incheon, Korea) with > 93% purity, diameters between 10-40 nm and lengths of
5-20 m.
Potentiometric Stripping Analysis for Antimony Determination

Prior to the electrochemical measurements, the carbon nanotube paste electrode was
submitted to chemical and electrochemical activation. Chemical activation was carried out by
immersing the electrode in a 2.0 mol L-1 HNO3 solution for 2 min. At the end of this step,
electrochemical activation was carried out by cyclic voltammetry, cycling the potential
between -1.0 and 1.0 V (18 cycles) in a 0.3 mol L-1 acetate buffer solution, pH 3.6.
The potentiometric stripping analysis procedure was performed using an electrochemical
cell with 15 mL capacity, in which the carbon nanotube paste electrode, Ag/AgCl reference
electrode and the platinum auxiliary electrode were immersed 10 mL of stirred 0.3 mol L-1
acetate buffer solution (pH 3.6) containing Hg2+ ions at 10 mg L-1. An aliquot of a stock
solution of antimony was added to the solution and a constant potential of -0.70 V (vs.
Ag/AgCl) was applied for during 180 s. Afterwards, the rest period of 20 s was waited and
the chemical stripping step was performed from -0.70 V to -0.15 V in an unstirred solution.
Stripping potentiograms were recorded as dt/dE vs E (V). All experiments were carried out
without removal of oxygen.
Composition of the Carbon Nanotube Paste Electrode

As previously related, the literature has demonstrated the useful incorporation of CNT in
composite matrices using different binders, such as Teflon (Wang and Musameh 2003),
bromoform (Britto et al. 1996) and mineral oil (Rubianes and Rivas 2003). Indeed, the
composite of CNTs with mineral oil presents several advantages for electrode building, such
as simplicity, low cost and feasibility for several electroanalytical techniques. However, the
content of mineral oil is an important aspect to consider when preparing carbon nanotube
paste electrodes. Therefore, the first study carried out was establishes the best composition of
carbon nanotube paste. The assays were performed in a medium containing 0.3 mol L-1 of
acetate buffer solution (pH 3.6) in the presence of 10 mg L-1 Hg (II) ions. The concentration
of antimony solution was 500 g L-1. This solution was submitted to the application of a
constant potential of -0.5 V for 180 s. The equilibrium time was 60 s and the chemical
stripping step was carried out from -0.5 V to -0.15 V. The potentiograms obtained are shown
in Figure 14.
Figure 14. Effect of paste composition on the analytical signal for antimony. The experiment was
performed in a medium containing 0.3 mol L-1 acetate buffer (pH 3.6) in the presence of 10 mg L-1 Hg
(II) ions. The concentration of the antimony solution was 500 g L-1 and the deposition potential
applied was -0.5 V for 180 s.
In spite of the facility to handle the paste containing 77.0% w/w oil, it proved to be the
worst composition regarding the analytical signal. This result was expected due to the reduced
conductivity of the paste. When pastes were used containing only 20.0 % w/w oil, a high
analytical signal was observed, but the background was somewhat high. Moreover, this
composition was difficult to handle. In order to circumvent this drawback, graphite powder
was inserted in the paste. The composition CNT: mineral oil: graphite (70/20/10 w/w)
showed a reduced background as well as a high analytical signal, similar to that observed in
paste containing 20.0 % w/w oil. Another advantage that can be pointed out is related to the
surface of electrode, which was smoother than those of the other electrodes tested.
Effect of Deposition Potential and Time Deposition

The profile of the analytical signal as function of the influence of the deposition potential
is shown in Figure 15. The potential range was varied from -0.4 to -1.0 V and, as expected,
the best results are obtained at the more negative deposition potential. However, when a
deposition potential more negative than -0.8 V was employed a significant decrease in the
antimony peak was observed, probably due to reduction of interferents present in the acetate
buffer solution. Thus, -0.7 V was chosen to avoid possible interferences. A study for
evaluating the influence of the deposition time on the sensitivity of the antimony peak was
performed from 60-180 s. The response for the analyte peak increased with increasing
deposition time, reaching a saturation condition at 180 s. This result is lower than previously
published results (Rise et al. 1997).
Figure 15. Effect of deposition potential on the analytical signal for antimony. The experiment was
performed in a medium containing 0.3 mol L-1 acetate buffer (pH 3.6) in the presence of 10 mg L-1 Hg
(II) ions. The concentration of the antimony solution was 500 g L-1.
Effect of pH and Ionic Strength

Potentiometric stripping analysis, similar to other electroanalytical techniques, requires
the use of supporting electrolytes. In antimony determination, acidic electrolytes are
commonly more used so that the hydrolysis of antimony could be avoided. In this study, a
hydrochloric acid solution at pH 1.0 as supporting electrolyte was first studied for antimony
determination, but no antimony peak was observed. This result indicates that more than one
mechanism can be involved in the retention of antimony onto the electrode surface. The first
comprises the deposition of antimony on the mercury film formed on the carbon nanotube
surface. The second is related to antimony adsorption directly onto carbon nanotube surface.
This later mechanism can be used to explain the high affinity of antimony for those sites of
carbon nanotubes at pH higher than 3, due to presence of carboxylic and hydroxyl groups
with pKa values ranging from 3 to 5. This consideration was confirmed by studying a large
pH range, in which the best antimony peak was observed at pH 3.6 in acetate buffer solution.
After choosing the sample pH the influence of ionic strength was investigated. Figure 16
shows the behavior of the analytical signal over a range varying from 0.05-0.7 mol L-1. From
the results, the analytical signal increased as the ionic strength increased until 0.3 mol L-1. At
high ionic strength the analytical signal decreased as a consequence of the reduction of
diffusion coefficient of the antimony ions towards the electrode surface. Therefore, a 0.3 mol
L-1 acetate buffer concentration was established as the best value in this study.
Figure 16. Effect of ionic strength on the analytical signal for antimony. The experiment was in the
presence of 10 mg L-1 Hg (II) ions. The concentration of the antimony solution was 500 g L-1 and the
deposition potential was -0.7 V.
Effect of Mercury Concentration

As is well known, the stripping step in potentiometric stripping analysis can be performed
by using chemical oxidation or by constant current. In this work, when constant current (25
A) was used, the analytical signal of antimony was very small. On the other hand, when
mercury ions were used as chemical oxidant, the signals were significantly increased. This
result makes it possible to emphasize that mercury ions play a more important role in the
stripping analysis than constant current. In addition, as observed from the pH studies, during
the deposition step the antimony can be reduced both in the mercury film and on the carbon
nanotube surface. Therefore, the effect of mercury concentration on the analytical signal was
investigated from 2-16 mg L-1. As observed in Figure 17, the best mercury concentration was
10 mg L-1, this value being chosen for the remainder of the study. At low mercury
concentrations there are not enough mercury ions to create the film as well as to chemically
strip off antimony ions. Concentrations of mercury ions higher than 10 mg L-1 provide a
decrease of analytical signal due to competition between antimony and mercury ions on the
carbon nanotubes surface.
Figure 17. Effect of mercury concentration on the analytical signal for antimony. The experiment was
performed in a medium containing 0.3 mol L-1 acetate buffer (pH 3.6). The concentration of the
antimony solution was 500 g L-1 and the deposition potential was -0.7 V.
Characteristics of the Analytical Method

Under the optimal conditions described above, the analytic curve and the detection and
quantification limits were determined. Peak height was linearly dependent on concentration in
the range 0.4 to 2.40 µmol L-1 with correlation coefficients of 0.990 (Figure 18). Limits of
detection and quantification were calculated as concentrate ions corresponding to three and
ten times, respectively, according to IUPAC (Long and Winefordner 1983). They were found
to be 0.085 and 0.286 µmol L-1, respectively.
Figure 18. Insert: Analytical curve for antimony determination with the optimized potentiometric
stripping method. For more details, see text.
Table 2 presents the results obtained from determination of antimony in the presence of
different concentrations of several interfering ions. A given species was considered to
interfere if it resulted in ±5% variation of the antimony peak. For this study different amounts
of the species were added to a 250 µg L-1 solution of antimony. The ratios of
antimony/interferent investigated was 1:1, 1:10 and 1:100. As observed, the foreign ions Pb
2+
, Ni2+, Co2+ and Cr3+ decreased the analytical signal of antimony only at high amounts of
interferent; however, it is important to stress that this condition is not usually found in real
samples, such as natural waters.
FINAL REMARKS
We have shown that in recent years there has been considerable interest for applying
CNTs directly or indirectly in the field analytical chemistry. The wide range of possible
applications of CNTs, including solid phase extraction procedures and biosensor and
electrochemical sensor construction indicate that these materials are promising candidates to
enhance the performance of analytical methods, mainly in situations where the exploitation a
new on different physics or chemistry is still necessary.
Table 2. Results obtained for several foreign ions in the potentiometric stripping
analysis for antimony determination
Interferent Ratio antimony/interferent Recovery of antimony

analytical signal (%)a
3+
Al 1:1 96.5
1:10 103.5
1:100 102.8
2+
Mg 1:1 98.5
1:10 98.5
1:100 99.1
Fe3+ 1:1 105.6
1:10 99.6
1:100 102.2
2+
Cd 1:1 97.0
1:10 97.4
1:100 102.9
Pb2+ 1:1 99.5
1:10 84.5
1:100 -
2+
Zn 1:1 101.3
1:10 103
1:100 98.8
Ni2+ 1:1 105.2
1:10 95.2
1:100 71.04
2+
Co 1:1 78.9
1:10 13.5
1:100 -
Cr3+ 1:1 90.7
1:10 33.6
1:100 -
a Recovery percentage for the antimony analytical signal using an antimony solution at 250 µg L-1.
In this way, the chemical and electrochemical properties of CNTs can be particularly
considered as suitable for the design of a variety of sensors. The electrochemical activities of
CNT-based electrochemical sensors, including potential shifts compared to the corresponding
non-modified sensors have been reported. To some extent this is explained by the presence of
oxidants, such as strong acids, that can open the ends of CNTs or introduce defects in their
sidewalls. Moreover, the good compatibility of carbon nanotubes with biological tissues and
their nanometric size are properties suitable for the assembling of miniaturized biocompatible
transducers and biosensors to be used for in vivo measurements.
In the field of sorption process CNTs have been proven to possess potential as superior
sorbents for preconcentration or removing divalent metal ions from aqueous solution, owing
to their high chemical resistance and high surface area. However, it is important to note that
the relatively high unit cost restricts their practical application for treatment of high volumes
of effluent. Hence, the practical use of CNTs as sorbents is more indicated for
preconcentration systems with the aim of obtain highly sensitive analytical methods. Within
this subject, we have reported a trace enrichment method based on the application of
MWCNTs as a SPE sorbent for cobalt determination by spectrophotometry using a flow
injection system. Besides the good analytical performance of this flow system, including a
high preconcentration factor, simplicity, low detection limit and low cost of implementation,
the MWCNT minicolumn has also shown excellent stability since the same packed
minicolumn has been used more than 800 times without losses of sorption capacity.
Besides the adsorptive properties of CNTs, the exploration of these materials for the
development of electrochemical stripping methods is still in its beginnings. Thus, another
successful application of CNTs described in this chapter is the study of a potentiometric
stripping method utilizing a carbon nanotube paste electrode for the determination of
antimony dissolved in aqueous solutions. Our results show that the use of carbon nanotubes
brings several advantages. Compared to other electrodes, mainly glassy carbon electrodes
coated with mercury film, carbon nanotube paste electrodes are easier to prepare, do not
require significant time for mercury accumulation onto the CNTs and are much less
expensive. Analysis based on the results obtained, it can be suggested that CNTs will surely
find application for the development of new voltammetric or potentiommetric stripping
methods for metal ion determinations as well as to make speciation studies.
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Lecture Material 11
CARBON NANOTUBES:
DISPERSION AND FIELD EMISSION PROPERTIES
ABSTRACT
Since carbon nanotubes (CNTs) show strong tendency to form aggregates due to
their high van der Waals interactions, their dispersion is a prerequisite for practical
applications. In order to enhance the dispersion of multi-walled carbon nanotubes
(MWNTs) in texanol, optimum type of dispersant and its concentration for six
commercial dispersants were evaluated based on the rheological results. Moreover, the
cutting and dispersion efficiencies of MWNTs were compared using conventional ball
milling and high energy milling, whereby the latter was found to be more effective. High
energy milling for two hours produced a large portion of MWNT agglomerates smaller
than 150 nm, showing a drastic increase in slurry viscosity due to the dispersion into
individual CNTs. On the other hand, 120-hour ball milling was required to achieve the
agglomerate size of 300 nm with less viscosity increase upon milling. Decrease in the
degree of MWNT crystallinity was observed by both milling, even though 2-hour high
energy milling showed slightly less damage than 120-hour ball milling based on XRD
and Raman spectroscopy results. The field emission properties of MWNTs were
examined using a screen-printed thick film prepared by 3-roll milling of MWNTs and
UV-sensitive binder solution, with a diode-type configuration in a vacuum for field
emission display (FED) application. The effects of MWNT milling, and various types of
ceramic and metal fillers on the emission current density and turn-on field were evaluated
by design of experiment (DOE), whereby the emission properties were shown to be
dependent significantly on both factors. Considerably enhanced emission properties were
obtained with the paste containing 5 – 10 wt. % of either ITO or the glass frit compared
with those without a filler. The paste containing 10 wt. % ITO represented an emission
current density of 176.4 μA/cm2 at 5V/μA, a turn-on field of 1.87 V/μA for an emission
current density of 1 μA/cm2 and a field enhancement factor of 7,580. The paste
formulation was also found to be suitable for fine patterning using UV-lithography
techniques. A long-term stability test for 110 hours of a paste containing 10 wt. % ITO
revealed a half-life of approximately 30,000 hours, which is appropriate for commercial
applications.
INTRODUCTION
Since their discovery by Iijima in 1991 [1], carbon nanotubes (CNTs) have been
extensively studied for application to field emission display (FED). Figure 1 shows a
schematic of CNT-FED structure with the CNT emitters on a cathode substrate and with red-
green-blue phosphor dots on the anode side. CNTs are acclaimed as one of the best cold
cathode emitters for FED applications due to their large aspect ratio, high mechanical strength
and electrical conductivity, and possible large-area application via thick film processing.
Although very similar to a cathode ray tube which is equipped with a single electron gun,
FED uses many electron emitters with a flat panel structure and features the advantages of
high screen quality, low power consumption, fast response time, wide viewing angle of about
170o, and low cost compared to the modern plasma display panel (PDP) or liquid crystal
display (LCD) [2].
Proposals for the fabrication of FEDs using CNTs have included direct growth [3],
electrophoresis [4] and the printing method [5]. The screen printing method is considered
particularly appropriate as the best candidate for the fabrication of large-area FED due to its
simple process, readiness for mass production and low cost. The limitation of this process in
achieving the level of micro-patterning which is required to increase the resolution has been
resolved by using a photosensitive binder and photolithography technique [6]. Even though
Toshiba and Canon tried to commercialize the surface-conduction electron-emitter (SED)
display based on similar technology in 2007 [7], there are no consumer production CNT-FED
models available yet. The challenges for the commercialization of CNT-FED are in ensuring
long-term stability and uniform electron emission [2].
Depending on the number of graphene layers, CNTs can be divided into single-walled
carbon nanotubes (SWNTs) and multi-walled carbon nanotubes (MWNTs). SWNTs generally
show a shorter lifetime, in spite of their higher degree of structural perfection and emission
current density, than MWNTs do [8].
Figure 1. Schematic diagram of CNT-FED.
The long-term stability which ensures a uniform electron emission for more than 30,000
hours is the most important challenge for the commercialization of CNT-FED [2]. Therefore,
current efforts are mainly being focused on MWNTs with a diameter of several nanometers
for actual applications [9].
Since CNTs tend to form aggregates due to their high van der Waals interactions, their
dispersion into each thread is a prerequisite to achieve uniform emission sites for FED
application. From this reason, researchers put their efforts to enhance the dispersion of CNTs
by surface treatments or cutting. Many kinds of surface treatment using plasma [10] or
UV/ozone [11], and functionalization techniques with polymeric agents have been introduced
[12-14]. Various physical or chemical CNT cutting methods include the utilization of
ultrasound [15], ball mill [16], vibration mill [17] or two-roller mill [18], oxidation in acidic
condition [19], fluorination and pyrolysis [20] and gamma radiation [21].
MWNT agglomerates are usually broken up and dispersed into smaller units through
mechanical milling for initial dispersion. The conventional ball mills and attrition mills that
have been used for this purpose, requiring more than 100 hours of milling to get an average
length of 200 – 300 nm [19,22]. However, modern high-energy mills are known to be very
efficient in milling due to their high-speed rotor rotating at up to several thousand rpm and
use of small grinding media with diameters of 0.05 – 0.8 mm [23]. Figure 2 presents a
schematic of (a) high energy mill system (MiniCer, Netzsch, Germany) and (b) the grinding
chamber of this mill. As high-energy mills have recently been introduced, however, there are
few reports thus far regarding its efficiency in MWNTs dispersion. In the subsequent
stabilization process, dispersion is achieved by electrostatic and steric mechanisms, both of
which prevent the close approach of MWNTs by using electrostatic repulsive force and
hindrance via the adsorption of polymeric molecules on the particle surface, respectively [23].
The paste for screen printing requires numerous ingredients such as CNT, binder resin,
solvent, dispersant, metal filler, photo initiator, monomer and glass frit, all of which should be
optimized in order to obtain a favorable emission property. Design of experiments (DOE) is a
powerful tool for establishing relationships among many experimental factors and output
responses for such a complex system. It utilizes a structured DOE table in which the input
factors are varied in a planned manner in order to identify efficiently the factors that most
influence the results as well as those that do not [24]. In spite of the numerous types of
available fillers, only few materials such as Ag and Al2O3 have been used as a filler for
MWNT paste without any systematic approach yet [25]. The filler is one of the important
ingredients, as it confers the adhesion strength of the paste and may also change the emission
properties at the same time.
In this overall perspective, we tried to cut and disperse CNTs using a high energy mill for
the first time and systematically compared the efficiency of this method with that of
conventional ball milling after adding a commercial dispersant into CNT slurry. Since CNTs
are known to be damaged by milling process [17], the degree of damage associated with two
different milling methods was characterized using high resolution transmission electron
microscopy (HRTEM), Raman spectroscopy and X-ray diffraction (XRD). Moreover, a full
factorial DOE was performed in order to verify the effect of each factor and to determine the
optimum MWNTs paste condition for FED application using three experimental input factors:
the treatment of high-energy milling for MWNTs, Ag filler type, and polymer/MWNTs ratio.
The emission properties such as turn-on field, current density and field enhancement factor
for eight kinds of paste were analyzed as output responses.
Figure 2. Schematic of (a) high-energy mill and (b) the milling chamber of the high energy mill [26]. 1:
Rotor with discs, 2: inlet, 3: milling media, 4: cooling jacket, 5 and 6: separation system.
Statistical analysis software, MINITAB, was used to generate the DOE table and to
analyze the results. Finally, this work examines the effects of four kinds of fillers on the
electron emission properties of MWNT paste, and the results are compared with those of the
paste without filler for FED application.
EXPERIMENTAL
Materials and the Determination of the Best Dispersant
MWNTs grown by catalytic CVD method (CMP-310F, Iljin Nanotech, Korea) with a
mean diameter of 3-5 nm, a length of 10-20 μm, and a specific surface area of 797 m2/g were
used for this experiment. In order to compare the dispersion efficiency of commercial
dispersants, six types of dispersant were chosen according to vendors‘ suggestion, as shown
in Table 1. After 10 minutes of ultrasonication (HD 2070, Bandelin using 20 kHz and 40 W,
Germany) for the slurries containing 2 wt. % of MWNTs in texanol (C12H24O3: 2,2,4-
Trimethyl-1,3-pentanediol monoisobutyrate), the rheological behavior of slurries with
different dispersants was investigated at 20oC using a computer-controlled cone-type
viscometer (LVDV-II+, Brookfield, MA, USA) with CPE-52 spindle at various shear rates.
Each dispersant was added at a ratio of 20 wt. % with respect to the CNT content.
Texanol was used as a solvent because it is widely used in the synthesis of CNT paste for
FED application [25]. Since BYK P-104 gave the most favorable results among the six types
of dispersant, the optimum amount of this dispersant in MWNT slurry was further decided by
varying the addition amount to 20, 50 and 100 wt. % with respect to MWNTs.
Table 1. Details of the six dispersants used in this study
Dispersant Abbreviation Supplier Suppliers‘ description on the functional group of

dispersant
Rhodafac RE-610 RE-610 Rhodia Nonylphenol ethoxylate based phosphate esters
Disperbyk-2001 BYK-2001 BYK Modified acrylate block copolymer
Disperbyk-111 BYK-111 BYK Copolymer with acidic groups
Disperbyk-103 BYK-103 BYK Copolymer with pigment affinic groups
BYK-9076 BYK-9076 BYK Alkylammonium salt of a high molecular weight

copolymer
BYK P-104 BYK P-104 BYK Low molecular weight unsaturated polycarboxylic
acid copolymer
Ball Milling and High Energy Milling Process
In order to compare the milling efficiency between ball milling and high energy milling,
the following experiment was performed. For ball milling, slurry was prepared by adding 1.4
grams of BYK P-104 and 7 grams of MWNTs (i.e., 20 wt. % of dispersant with respect to the
MWNT content) into 350 grams of texanol to form a slurry of 2 wt. % MWNTs with respect
to the solvent content. This slurry was ball-milled in a 500 cc polypropylene bottle for 120
hours with 2 mm spherical ZrO2 balls. Figure 2 shows the schematic of high energy mill
system. This mill had a ZrO2 lining with an 180cc of milling chamber which was filled with
80 vol. % of 0.8 mm ZrO2 balls. High energy milling was performed at 2,000 rpm for the
same composition of MWNT slurry for 2 hours by adopting a continuous circulation method,
where a small amount of sample was taken at a planned time for the characterization.
The following characterization was performed for the samples collected occasionally
during this experiment. Optical microscopy (ICS 305B, Sometech, Korea), field emission
scanning electron microscopy (FESEM: S-4100, Hitachi) and HRTEM (H-7600, Hitachi
using 200 kV) were utilized for the morphological observation after the ultrasonication and
during both the milling operations. In order to evaluate the degree of MWNT damage upon
milling, XRD (RINT 2000, Rigaku using Cu Kα line for 2θ = 20 – 80o) and FT-Raman
spectroscopy (FRA 106/S, Bruker Optics, Germany using Ar excitation (λ=514.5 nm)) were
performed using the dried MWNT samples before and after milling. Zeta potential/particle
size analyzer (PSA: Zetasizer Nano ZS, Malvern) and viscometer were also used to evaluate
the degree of MWNT dispersion with the slurry samples during both of milling.
Paste Synthesis and the Design of Experiment (DOE)

Eight different types of paste were prepared for DOE. The high-energy milling for
MWNTs, Ag filler type, and polymer/MWNTs ratio were chosen as the three controlled
factors with two levels for each factor, as shown in Table 2. In order to obtain the adequate
rheology for screen printing, the amount of nano-sized Ag filler was adjusted to 10 wt. %
with respect to the total paste weight, while that with micro-size is fixed at 20 wt. %.
Table 2. Eight types of MWNT pastes for experimental design
No High-energy milling Silver filler Polymer/CNT ratio

1 No Nano 10 wt.% 4/1
2 Yes Nano 10 wt.% 4/1
3 No Micro 20 wt.% 4/1
4 Yes Micro 20 wt.% 4/1
5 No Nano 10 wt.% 6/1
6 Yes Nano 10 wt.% 6/1
7 No Micro 20 wt.% 6/1
8 Yes Micro 20 wt.% 6/1
The eight paste compositions were pre-mixed using a stirrer and subjected to the 3-roll
milling. The MWNTs pastes were screen-printed onto an indium-tin oxide (ITO) coated soda
lime glass to a thickness of approximately 2 μm and dried at 90oC for 10 minute in air. Since
the residue of organic materials in the pastes causes problems such as out-gassing and arching
during the field emission measurement, binder burnout was performed at 450oC in a nitrogen
atmosphere, followed by an activation process comprising the vertical alignment of MWNTs
using a sticky tape. The field emission characteristics of the pastes were measured in a
vacuum chamber with a parallel diode-type configuration at a pressure of 10-6 Torr using a
pulse generator with 1/500 duty. The gap between anode and cathode was 200 μm, and each
sample had an area of 1.0 ×1.0 cm2. The surface resistivity of the screen-printed thick film
was measured using the four-point probe method.
The Effects of Fillers
Coarse (average particle size of 705 nm) indium tin oxide (ITO) powder was milled into
a fine particle size (67 nm) using an high energy mill (MiniCer, Netzsch, Germany) at a rotor
speed of 3,000 rpm with 0.4 mm ZrO2 beads for 3 hours, and added to the paste at 1, 5, 10
and 20 wt. % as filler. A glass powder (78 nm) with a composition of 72 SnO-23 P2O5-5
SiO2/Al2O3 in wt. % was also high energy-milled at the same milling condition and used as a
filler by adding 1, 5 and 10 wt. %. An additional two paste formulations were prepared as
references: one containing 5 wt. % Ag powder (392 nm), and the other containing no filler.
Figure 3 shows SEM images of the fillers used in this study. The amounts of MWNT,
initiator and acrylic binder resin were fixed to 1.6, 3.0 and 23.0 wt. %, respectively, in all
pastes for comparison. Polycarboxylic acid polymer (BYK-P 104, BYK Chemie, USA) at a
ratio of 100 wt. % with respect to the CNT weight was used as the dispersant.
The nine paste formulations, including that without a filler, were pre-mixed using a stirrer
and subjected to 3-roll milling. Three-roll mill employs the shear force created by three
horizontally positioned rolls rotating at opposite directions with different speeds relative to
each other in order to homogenize the paste by decreasing the gaps between the rolls.
Figure 3. SEM morphology of the fillers used in this study: (a) ITO, (b) glass frit and (c) Ag powder.
Average particle size is shown.
The field emission characteristics of the pastes were measured with the same condition
that mentioned above. In order to check the UV-development properties of these pastes for
practical application, 25 μm–diameter dots at a spacing of 400 μm were deposited
simultaneously by UV-lithography. The MWNTs and printed morphologies were examined
by field emission scanning electron microscopy (FESEM: S-4100, Hitachi) and high
resolution transmission electron microscopy (HRTEM: H-7600, Hitachi).
RESULTS AND DISCUSSIONS

Materials and the Determination of the Best Dispersant
As-received MWNTs have a highly entangled structure as shown in Figure 4, which

needs to be dispersed, preferably up to single nanotube level, for practical applications. Figure
5 shows the rheological behavior of slurries containing 2 wt. % of MWNTs in texanol with
six different dispersants after 10 minutes of ultrasonication. All show pseudo-plastic
behavior, as indicated by the viscosity decrease with increasing shear rate. Even though all six
slurries with dispersant showed lower viscosity than that without dispersant (85 mPa.s at a
fixed shear rate of 60 s-1), the slurry with BYK P-104 showed the lowest viscosity (38 mPa.s)
among the six types of dispersants tested (range of other five types: 54 – 67 mPa.s).
The purpose of this measurement was in the determination of optimum dispersant by
measuring the overall slurry viscosity instead of more detailed and complicate measurement
such as isotherm adsorption. Since ultrasonication for 10 minutes alone could not confer the
dispersion into each CNT thread, however, we may regard the CNT agglomerates as close-
packed bundles at this stage. Even though there might be some portion of disentangled CNTs,
many researchers considered them before further treatment as entangled-bundles [27-30].
Even if CNTs behave like bundles, the slurries containing different dispersants would still
show difference in viscosity depending on the properties of each dispersant such as amount of
absorption, configuration and different functional groups. When CNTs are considered as
bundles, the CNT slurry with more dispersed bundles will show lower viscosity, showing a
pseudo-plastic behavior as slurries containing ceramic particles [31]. Therefore, BYK P-104
could be considered as the most efficient dispersant with the lowest slurry viscosity.
This effect of different dispersants on the rheological behavior of slurries with MWNT
agglomerates was further confirmed using the optical microscopic images shown in Figure 6.
Figure 4. SEM image of as-received CNTs with highly entangled structure.
Figure 5. Rheological behavior of slurries containing 2 wt. % of MWNTs in texanol without dispersant
and with six different types of dispersant: (a) Viscosity as a function of shear rate and (b) viscosity at a
fixed shear rate of 60 s-1.
Since better-dispersed slurry with MWNT bundles displays lower viscosity due to the
particle mobility offered by the fluid interparticulate layer [31,32], the dispersant with high
slurry viscosity generally tends to show more agglomerated MWNT morphology than the
slurry with lower viscosity. According to Figure 6, slurries containing BYK-2001, BYK-103
and BYK P-104 showed lower viscosity and more dispersive morphologies than those with
BYK-9076, BYK-111 and RE-610, which is consistent with the viscosity data shown in
Figure 5.
Figure 6. Optical microscopic images of MWNT slurries containing no dispersant or 20 wt. % of the
six dispersants after 10 minutes of ultrasonication.
The slurry dispersion depends on the sign and magnitude of the total energy of interaction
between particles [33]. Stabilization is usually achieved by electrostatic and/or steric
mechanisms, both of which prevent particles from approaching close due to the electrostatic
repulsive force and hindrance to the adsorption of polymeric molecules on the particle
surface, respectively [33]. The difference in slurry viscosity and morphology in this case
seems to be more attributed to the steric hindrance caused by the different functional groups
of the dispersants than to the electrostatic mechanism because of the low dielectric constant of
texanol (εr≈2.0, based on our measurement using an SI 1260 instrument with 12964A sample
holder, Solartron, UK). Zeta potential, the magnitude of electrostatic repulsion, of the above
slurries in texanol could not be measured using a PSA. On the other hand, the zeta potential
of the above slurries in ethanol (εr=24.3) containing the same amount of dispersants was less
than 10 mV, which would be smaller in texanol due to its low dielectric constant.
Figure 7 shows the rheological behavior of MWNTs slurry as a function of BYK P-104
content. The viscosity continuously decreased with increasing BYK P-104 content: 85, 38,
28, and 24 mPa.s at a shear rate of 60 s-1 for 0, 20, 50, and 100 wt. % with respect to MWNT
weight, respectively. A well-dispersed slurry displays lower viscosity, while slurry with
insufficient dispersant shows higher viscosity due to the incomplete adsorption on the particle
surface. Therefore, 100 wt. % of BYK P-104 with respect to MWNT weight can be
considered as the optimum content because of the lowest slurry viscosity.
However, a milling experiment was further performed with 20 wt. % of BYK P-104
dispersant because the formulated binder phase contained approximately 80 wt. % of
additional dispersant in case of paste synthesis for our CNT-FED application. The slurry
viscosity decreased significantly by adding 20 wt. % of BYK P-104 compared to the slurry
without dispersant based on Figure 7. Since the dispersant is a simple facilitator which
ultimately needs to be removed, the minimum amount able to confer slurry dispersion is
desirable.
Figure 7. Rheological behaviors of slurries containing 2 wt. % of MWNTs in texanol with various
amounts of BYK P-104 dispersant.
Ball Milling and High Energy Milling Process
Figure 8 compares the morphology of the MWNTs using optical microscopy after various
treatments: (a) 10 minutes of ultrasonication, (b) 120 hours of ball milling, and (c) 2 hours of
high energy milling. Figure 8 (a) shows much portion of MWNT agglomerates even though it
looks like well-dispersed slurry under low magnification as shown in Figure 6 (g).
Figure 8. Optical microscopic images of MWNTs: (a) ultrasonicated for 10 minutes, (b) 120-hour ball-
milled and (c) 2-hour high energy-milled.
Compared to the sample only with ultrasonication in Figure 8 (a), the milled samples in
Figures 8 (b) and (c) showed more dispersed images due to larger portion of individual CNTs.
According to the evolution of average particle size on milling time for the two kinds of
milling method shown in Figure 9, 2-hour high energy milling resulted in finer MWNT size
than that with 120-hour of ball milling. High energy milling generated a much greater portion
of MWNTs smaller than 150 nm after 2-hour milling, even though it showed bimodal
distribution, while it was difficult to obtain such a fine size with ball milling even after 120-
hour milling. Figure 10 compares the morphology of as-received and 2 hour high energy-
milled MWNTs using SEM and HRTEM images. A comparison of the SEM images in Figure
10 (a) and (b) indicates that the MWNTs were not only dispersed but also cut by high energy
milling. The as-received MWNT sample in Figure 10 (c) shows four graphene layers with a
diameter of approximately 4.8 nm and with some impurities attached on the straight MWNTs.
Since energy dispersive X-ray analysis (EDX) results (data not shown here) only showed a
carbon peak, these impurities must have been amorphous carbon, fullerenes or
nanocrystalline graphite. The high energy-milled MWNTs in Figure 10 (d) shows somewhat
curved morphologies, indicating damage from the milling, while the overall tube-type shapes
are still maintained after 2-hour high energy milling.
Figure 9. Evolution of particle size distribution for two different milling methods: (a) ball milling and
(b) high energy milling.
Figure 10. Comparison of the morphologies of as-received and 2-hour high energy-milled MWNTs in
texanol: (a) and (b) SEM, and (c) and (d) HRTEM images.
The behavior of slurry viscosity with milling time, represented in Figure 11, showed
somewhat different characteristics between the two kinds of milling. The viscosity of the
milled slurries drastically increased, especially at the initial milling stage, i.e., up to 10 hours
for ball milling and 10 minutes for high energy milling. However, the viscosity of the high
energy-milled slurries was much higher than that of the ball-milled ones, in spite of the
shorter milling time. For example, the viscosity increased from an initial 85 mPa.s to 9,735
mPa.s for the high energy-milled slurry after 10 minutes of milling, while that of the ball-
milled slurry increased to only 3,169 mPa.s after 10 hours of milling. The viscosity of a slurry
with solid particles generally decreases with increasing dispersion due to the particle mobility
offered by the fluidity between interparticulate layers, so long as a new surface is not created
during the milling process. However, when dispersion means the disentanglement of
spaghetti-liked MWNTs into individual CNTs by milling as in this case, viscosity will be
increased by dispersion due to the enhanced resistance to flow resulting from internal friction
between the nanotubes and solvent. Therefore, the degree of viscosity increase on milling
indicates the dispersion efficiency of the system. Based on these observations, we may
conclude that the dispersion efficiency of high energy milling is much higher than that of ball
milling.
Figure 11. Changes in slurry viscosity for two different milling methods: ball milling and high energy
milling.
Paste Synthesis and the Design of Experiment (DOE)
Figure 12 (a) and (b) compares the side-view images of the activated screen-printed
pastes containing MWNTs of (a) without and (b) with 2-hr, high-energy milling. Since the
high-energy milling enhanced the dispersion as well as the cutting of MWNTs, the sample
shown in Figure 12 (b) has a much greater density of surface-exposed MWNTs than that in
Figure 12 (a). Figure 12 (c) and (d) shows the top surface morphology of the pastes
containing (c) nano-sized and (d) micro-sized Ag. Based on the agglomerated Ag particles in
Figure 12 (c), it seems that 3-roll milling alone is insufficient to disperse the nano-sized Ag
particles.
Figure 13 shows (a) the electric field vs. emission current density and (b) Fowler-
Nordheim (F-N) plots for four selected samples: numbers 2, 6, 7 and 8 among the 8 samples
described in Table 2. In order to compare the MWNT emitters, Table 3 presents a comparison
of the eight samples in terms of the turn-on electric field for an emission current density of 10
μA/cm2, film surface resistivity, emission current density at 5 V/μm, and the calculated field
enhancement factor (β factor), assuming an MWNT work function of 5 eV, which is the same
as that of graphite. All values were different, confirming the importance of parameter
optimization. Sample No. 8, synthesized from 20 wt. % micro-sized Ag with a polymer to
high-energy milled MWNT ratio of 6/1, showed the smallest turn-on field of 1.95 V/μm for
10 μA/cm2 emission density with the highest emission current density of 101.0 μA/cm2 under
5 V/μm and the highest β factor of 24,255.
Figure 12. MWNT pastes screen-printed on ITO glass: side-view of samples (a) No. 1 and (b) No. 2;
top-view of samples (c) No. 2 and (d) No. 4.
Figure 13. (a) I-V characteristic and (b) F-N plot with field enhancement factor ( ) for four selected
MWNT emitters.
Table 3. Values of turn-on field, surface resistivity, current density and -factor for
eight samples
No Turn-on field for 10 Surface resistivity Current density at 5 -factor

A/cm2 (V/ m) (kΩ/square) V/ m ( A/cm2)
1 2.75 184.97 84.7 11,180
2 3.35 1.72 63.8 7,151
3 2.25 26.81 93.0 10,816
4 2.65 1.26 90.1 9,960
5 2.90 219.67 77.0 10,475
6 2.90 31.99 76.5 10,979
7 2.65 1.47 93.3 10,000
8 1.95 1.97 101.0 24,255
The F-N model expresses the relationships among the emission current density, local
electrical field, and work function (Φ) of the emitter, where ln(J/E2) vs. 1/E shows a linear
relationship based on the following equation [34]:
3
2
J a 0.95b 2
ln( 2 ) ln( ) (1)
E E
where J is the emission current density (A/m2), E is the applied electric field (V/m), a and b
are constants (a=1.54×10-6 A·eV/V2, b=6.83×109 eV3/2), is the field enhancement factor,
and Φ is the work function of CNT. The slope of ln(J/E2) vs. 1/E from equation (1) is given
by:
3
d (ln( J / E 2 )) 0.95b 2
Slope (2)
d (1 / E )
Therefore, can be calculated from equation (2) since the other parameters are known.
The factors of the 8 samples ranged from 7,151 – 24,255, which is relatively larger than that
of theoretical or individual CNT values of few thousand [35]. Based on their experimental
results, Berdinsky et al. [36] proposed that this 10 – 50 times higher factor for CNT paste
than that for an individual CNT was due to the addition of emission currents from the CNTs
in an array. Many of other researchers also reported such a high factor for a CNT array
[5,37-40].
The main effects plot for emission current density at 5 V/μm, which can verify the effect
of each input factor, is shown in Figure 14. A dashed reference line is drawn at the average of
the eight samples (=84.9 μA/cm2), and the circle symbols in the graph correspond to the
emission current density at each level of the corresponding input factor. The straight line
connecting between two circle symbols does not mean a real linear output response for this 2-
level factorial design. The most significant factor was the Ag filler type in the paste, with a
larger emission density being shown with the 20 wt. % micro-sized Ag compared to the 10
wt. % nano-sized sample. Due to the different content, however, the size effect of the Ag
filler in the emission current density could not be distinguished. CNT paste containing fillers
such as Ag and Al2O3 is known to show higher emission density and more adequate
rheological properties than the paste without filler [25]. Based on the higher density of the
surface-exposed MWNTs in Figure 12, the paste containing high-energy milled MWNTs was
expected to have a much larger emission density than that of the intact sample, but this was
contradicted by the result shown in Figure 14.
Figure 14. Main effects plot for emission current density at an electric field of 5V/μm.
Figure 15. Main effects plot for surface resistivity of MWNT thick film.
According to our previous report with the same type of unmilled MWNTs [9], the higher
surface-exposed density resulted in the larger emission current density along with a smaller
surface resistivity. Although the surface resistivity of the screen-printed film was decreased
drastically with the high-energy milled MWNTs due to their high surface-exposed density
compared to the paste with intact sample in this study, as shown in Figure 15, the paste
containing high-energy milled MWNTs did not show such a high emission current density
relative to the surface-exposed density. As shown in Figure 16 (a), there was no correlation
between the emission current density of the paste with high-energy milled MWNTs, while
current density increased with decreasing surface resistivity for the pastes with intact
MWNTs, which showed a similar trend to the results of our previous study shown in Figure
16 (b). Even though the larger MWNT content in the paste increased the emission density of
the intact MWNTs, the content was fixed at 2 wt. % in this study due to the excessively high
paste viscosity for screen printing and the difficulty in dispersion for higher content.
In order to explain the lower average emission current density of the paste with high-
energy milled MWNTs over the surface-exposed density, XRD patterns and Raman spectra
were compared between the as-received and high-energy milled MWNTs, as shown in Figure
17. Based on the XRD patterns of MWNTs shown in Figure 17 (a), the peak at 2θ of 25.60o
(P1), which corresponded to the {003} plane of synthetic carbon/graphite [JCPDS Card #: 26-
1079], showed an especially wide diffraction angle, indicating the existence of amorphous-
like carbonaceous impurities. Miller indices for each peak were not available because JCPDS
cards contain more than eighty kinds of carbonaceous materials which have different
diffraction peaks. The peak at 2θ of 42.80o (P2), which corresponded to {101} plane of the
synthetic carbon/graphite material, decreased and broadened significantly after the milling.
The relative intensities of this peak compared to that at 2θ of 25.60o (=P1/P2) were 1.078 and
0.315 for the as-received and 2-hr, high-energy milled samples, respectively. The small peaks
at 2θ of 62.34 and 78.66o, both corresponded to {105} and {110} plane of the above material,
respectively, with the as-received sample had almost disappeared after the milling. The
decrease in intensity and peak broadening suggested a degraded MWNT crystallinity due to
milling damage. The Raman spectra, an alternative method for checking the degree of
damage, of MWNTs using Ar excitation (λ=514.5 nm) is shown in Figure 17 (b). The G-band
near 1587 cm-1 that originated from the vibration of sp2-bonded atoms in two-dimensional
hexagonal lattices, which is referred to as a highly ordered or crystalline carbon [41], and the
D-band near 1279 cm-1 was indicative of some disruption, disorder or defects in the graphitic
layers and/or carbonaceous particles [6].
Figure 16. Relationship between emission current density and film surface resistivity for (a) the present
study results with 2-hr, high-energy milled MWNTs and (b) the previous study results with unmilled
MWNTs.
Figure 17. (a) XRD patterns and (b) Raman spectra of MWNTs before and after 2-hr, high-energy
milling.
The relative band height ratio of G- to D-band was 7.03 for as-received MWNTs, but was
nearly halved to 3.63 after 2-hr, high-energy milling. These XRD and Raman results indicated
that the MWNT amorphism was increased by milling, and that such damage is inevitable due
to the collision and shear forces exerted on the MWNTs during the milling process. Even
though there has been no report of decreased electron emission with MWNTs after milling,
we considered that the structural damage associated with the milling decreased in the
emission current density despite the high density of surface-exposed MWNTs.
The Effects of Fillers
Figure 18 shows a typical SEM side-view image of a paste, which was screen-printed
onto ITO glass, indicating the high surface-exposed MWNT density after the activation
process. Since electrons are emitted from the MWNT tips, a higher surface-exposed density is
desirable for achieving a high emission current density.
Figure 18. SEM side-view image of the screen-printed MWNT paste containing the glass frit.
The filler in the paste mainly plays two roles: enhancing the adhesion strength of
MWNTs to the substrate, and offering an electron-conducting path from the cathode to
MWNTs. In this respect, the glass frit and Ag filler are closely related to the former and latter,
respectively, while ITO might act for both purposes. At the same time, the addition of filler
should not deteriorate the overall paste properties including the screen printability and
processing capability for micro-patterning. The MWNT dot patterns, which have a diameter
of 25μm and were formed by UV-lithography after screen printing of 4 different paste
compositions, show a clear boundary, as shown in Figure 19. This suggests that the binder
system and filler selection are suitable for screen printing and fine patterning.
Figures 20 (a) and (b) show the current-voltage (I-V) characteristics and the
corresponding Fowler-Nordheim (F-N) plots for selected samples, containing different
amounts of ITO and glass frit, respectively. The I-V curves for the pastes containing 5 wt. %
Ag and no filler are also included for comparison. The overall emission current densities of
the ITO added pastes are much higher than that without the filler and 5 wt. % Ag added paste.
Moreover, the emission current density showed a continuous increase up to a ITO content of
10 wt. %. The paste with 20 wt. % of ITO showed a slightly lower emission current density
than that with 10 wt. %, indicating the maximum emission properties at this composition.
Similar emission properties were observed with the glass frit added pastes, as shown in Figure
20 (b), while the maximum current density is shown at 5 wt. % addition. The F-N plots
(insets in Figs. 20(a) and (b)) show an almost linear slope, suggesting the field emission of
electrons from MWNTs.
Figure 19. SEM images of the MWNT cathode dot patterns formed by UV-lithography containing (a)
no filler, 5 wt. % of (b) ITO, (c) glass frit and (d) Ag powder.

Figure 20. I-V characteristics and Fowler-Nordheim (F-N) plots (inset) for selected samples of the
pastes containing (a) ITO and (b) glass frit. I-V curves for the pastes without filler and with 5 wt. % of
Ag are added for comparison.
Figures 21 (a) and (b) show the electron emission current density in an electric field of 5
V/μA and a turn-on field for an emission current density of 1 μA/cm2 for the ITO and glass
frit added pastes, respectively. Compared with the emission current density of the paste
containing no filler (28.9 μA/cm2) and with 5 wt. % Ag (40.0 μA/cm2) at 5V/μA, the 10 wt.
% ITO and 5 wt. % glass frit added pastes show very high values of 176.4 and 170.2 μA/cm2,
respectively. At the same time, the turn-on field for an emission current density of 1 μA/cm2
was decreased from 2.60 V/μA for the paste without a filler to 1.87 and 1.80 V/μA for the 10
wt. % ITO and 5 wt. % glass frit added samples, respectively. The sample with the higher
emission current density tended to show a lower turn-on field, as shown in Figure 21. A low
turn-on field is preferred by the panel display industry for rapid switching. This considerable
increase in emission current and decrease in turn-on field in these samples might be due to the
optimum electron-conducting path and adhesion strength at these compositions. Further
increases in the amount of filler can enhance the adhesion strength but hinder electron
conduction due to the low electric conductivity of the filler.
The factors calculated from the slope of equation (2) for the samples with 10 wt. % ITO
and 5 wt. % glass frit were 7,580 and 7,750, respectively, assuming that the work function of
MWNTs is 5 eV, which is the same as that of graphite. The effective electric field at the
MWNT tips is not simply the applied voltage divided by the gap between two electrodes
(=V/d), but is increased by (= E), which reflects the enhancement of the electric field at the
CNT tips for electron emission. Based on these results, the type of filler and its amount are
important parameters for obtaining optimal emission properties.
The long-term emission stability of the CNT emitters was checked for 110 hours at a
constant electric field of 5V/μm. Figure 22 shows the change in current density for the sample
containing 10 wt. % ITO. The figure shows stable emission properties without any significant
fluctuation and degradation during the measurements. An extrapolation of this data indicated
this sample to have a half-life of approximately 30,000 hours, which is suitable for
commercial application.
Figure 21. Behavior of the emission current density and turn-on field as a function of the amount of
filler for (a) ITO and (b) glass frit.
Figure 22. Long-term emission current density stability of the MWNT emitter containing 10 wt. % of
ITO filler.
CONCLUSION
The results presented in this study enable the following conclusions to be drawn:
1. Six types of commercial dispersant showed different degrees of MWNT dispersion in

texanol, which was explained mainly by the steric hindrance effect due to the low
zeta potential value of the slurry. Based on the comparison of MWNT milling
between conventional ball and high energy milling, the latter was much superior to
the former in terms of milling efficiency with less damage to the MWNTs. High
energy milled MWNTs can be applied for many practical applications including
paste for CNT-FED, where higher current density is expected due to the larger
surface-exposed MWNTs density at low loading rate.
2. High-energy milling significantly enhanced the MWNT dispersion and the surface-
exposed density in a paste form. Based on the field emission properties of the 8
samples investigated using DOE technique, however, the overall emission current
density of pastes containing 2 wt. % high-energy milled MWNTs was lower than that
of the paste containing the same amount of intact sample, despite their higher density
of surface-exposed MWNTs. Further characterization using XRD and Raman
spectroscopy indicated that the degree of crystallinity with high-energy milled
MWNTs was lower than that of the intact samples due to the collision and shear
forces experienced during the milling process, which explained the lower emission
current density of high-energy milled MWNTs. The paste synthesized from 20 wt. %
macro-sized Ag with a polymer to high-energy milled MWNT ratio of 6/1

demonstrated the optimum condition among the 8 samples, with an emission current
density of 101 μA/cm2 under 5V/μA.
3. The effects of ITO and glass frit fillers on the field emission properties of MWNT
paste were examined by varying their amounts for 9 types of samples. The electron
emission properties, such as the emission current density and turn-on field, were
found to be dependent on the type and amount of filler. The optimum filler results
were obtained with the paste containing either 10 wt. % ITO or 5 wt. % glass frit. At
these concentrations, the field enhancement factor for the pastes containing 10 wt. %
ITO and 5 wt. % glass frit was 7,580 and 7,750, respectively. The emission current
densities of the pastes for optimum conditions were approximately 6 times higher
than that without the filler. At the same time, the turn-on field for these samples was
decreased significantly from the paste without the filler, which highlights the
importance of a filler in a paste formulation. Moreover, the samples showed good
long-term emission properties for 110 hours, indicating a half-life of 30,000 hours
based on data extrapolation, which is suitable for commercial CNT-FED application.
These results also suggest that the UV-sensitive binder system can be used for fine
patterning.
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Chapter 12
CARBON NANOTUBES:
COAGULATION–FRAGMENTATION EQUATIONS
ABSTRACT
The existence of single-wall carbon nanotubes (SWNTs) in organic solvents in the
form of clusters is discussed. A theory is developed based on a bundlet model for clusters
describing the distribution function of clusters by size. The phenomena have a unified
explanation in the bundlet model of a cluster, in accordance with which the free energy of
an SWNT involved in a cluster is combined from two components: a volume one
proportional to the number of molecules n in a cluster, and a surface one proportional to
n1/2. A comparative study of the droplet and bundlet models is carried out. The model
yields an activation barrier and predicts that pores with a radius below a certain critical
value are unstable, while those above this radius will grow indefinitely until the
membrane ruptures. During the latter stage of the fusion process, the dynamics were
governed by the displacement of the volume of liquid around the fusion site. Based on a
simple kinetic model, micellization of rod-like aggregates occurs in three separated
stages. A convenient relation is obtained for <n> at transient stage; at equilibrium another
relation determines binding energy . A relation with surface dilatational viscosity is
obtained. The model predicts that pores with a radius below a certain critical value are
unstable.
Keywords: nanostructure, diffusion, phase equilibrium, thermodynamic property,

transport property.
INTRODUCTION
Among the unusual properties of fullerene solutions, it should be mentioned the
nonmonotonic temperature dependence of solubility [1] and the nonlinear concentration
dependence of the third-order nonlinear optical susceptibility [2]. The solvatochromic effect
is exhibited in a sharp alteration in the spectrum of the optical absorption of C70, dissolved in
a mixture of organic solvents, of a result of a slight change in the solvent content [3, 4]. The
behaviour of fullerene solutions is attributable to cluster formation [5, 6]. It was examined the
decrease in pyridine-soluble material observed after soaking coals in solvents, because of an
increase in cross-linking associated with the formation of ionic domains [7]. It is not possible
to extract C60–70 from a solution in toluene to water and from a dispersion in water to toluene
[8]. In water C60 spontaneously forms a stable aggregate (C60)n with nanoscale dimensions
[9]. Water might form a donor–acceptor complex with C60 leading to a weakly charged
colloid [10–12]. C60, dissolved in water via complexation with cyclodextrin8, was extracted to
toluene [13, 14]. In C60 incorporated into artificial lipid membranes, it was not extracted to
toluene, but extraction became possible once the vesicle was destructed by adding KCl [15].
Addition of KCl was required to extract poly(vinylpyrrolidone)-solubilized C60–70 to toluene
[16–20].
In earlier publications, periodic tables of single-wall carbon nanotubes (SWNTs) were
discussed [21, 22]. A molecular modelling comparative study of SWNT solvents and
cosolvents provided a classification in best, good and bad solvents [23–26]. A program based
on the AQUAFAC model was applied to calculate the aqueous coefficients of SWNTs [27].
The bundlet model for clusters of SWNTs was presented [28–32]. The aim of the present
report is to perform a comparative study of the properties of fullerenes (droplet model) and
SWNTs (bundlet). A wide class of phenomena accompanying the behaviour of SWNT
solutions is analyzed from a unique point of view, taking into account the tendency of
SWNTs to cluster formation. Based on the droplet model of C60–70 the bundlet model of
SWNTs is proposed. The next section presents the computational method. Following that the
asymptotic coagulation–fragmentation equations are described. Next the calculation results
are discussed. The last section summarizes the perspectives.
COMPUTATIONAL METHOD
A solubility mechanism is based on SWNT cluster formation in solution. Aggregation
changes SWNT thermodynamic parameters, which displays phase equilibrium and changes
solubility. The bundlet model is valid when the characteristic number of SWNTs in the
cluster n >> 1. In saturated SWNT solution, the chemical potentials per SWNT for dissolved
substance and for a crystal are equal. The equality is valid for SWNT clusters. The free
energy of a cluster is made up of two parts: the volume part proportional to the number of
SWNTs n in the cluster, and the surface one proportional to n1/2 [33–43]. The model assumes
that clusters consisting of n >> 1 particles have a cylindrical bundlet shape and permits the
Gibbs energy Gn for a cluster of size n to be
12
Gn G1n G2n (1)
where G1–2 are responsible for contribution to Gibbs energy of molecules placed inside the
volume and on surface of a cluster. Chemical potential n of a cluster of size n is
n Gn T ln Cn (2)
where T is temperature. With (1) this results
12
n G1n G2n T ln Cn (3)
where G1–2 are in temperature units. In a saturated SWNT solution, cluster-size distribution
function is determined via equilibrium condition linking the clusters of a specified size with a
solid phase, which corresponds to the equality between the chemical potentials for molecules
incorporated into clusters and into crystal, resulting in the cluster-size distribution function in
saturated solution
An Bn1 2
f n gn exp
T (4)
where A is the equilibrium difference between interaction energies of an SWNT with its
surroundings in solid phase and in cluster volume, B, the similar difference for SWNTs
located on the cluster surface, gn, the statistical weight of a cluster of size n. We shall neglect
gn(n,T) dependences in comparison with exponential (4). Normalization for distribution
function (4)
f nn C
n 1 (5)
requires A > 0. Here C is solubility in relative units. As n >> 1 normalization (5) results
An Bn 1 2
C gn n exp dn
n 1 T (6)
where gn is the statistical weight of a cluster averaged over the range of n that makes the
major contribution to integral (6), and C0, SWNT molar fraction. The A, B and C0 have been
taken equal to those for C60 in hexane, toluene and CS2: A = 320K, B = 970K, C0 = 5·10–8
(molar fraction) for T > 260K. A correction takes into account the different packing
efficiencies between C60 and SWNTs
cyl cyl
A A B B
sph and sph (7)
where cyl = /2(3)1/2 is the packing efficiency of cylinders, and sph = /3(2)1/2 the one of
spheres (face-centred cubic, FCC). Software is available from the authors.
DESCRIPTION OF THE ASYMPTOTIC COAGULATION–

FRAGMENTATION EQUATIONS
Finding a manageable approximation to the behaviour of the coagulation–fragmentation
equations is challenging. The approximation is presented by asymptotic analysis. Results
have been checked against numerical solutions to the equations dealing with the Becker–
Döring equations. Typical models for the binding energy of a n cluster follow. For rod-like
aggregates
n n 1 k BT (8)
where kBT is the monomer–monomer binding energy [44–46]. In the Becker–Döring model,
reactions only between monomers and other clusters are taken into account. Expression (8) is
suitable for aggregates of certain kinds of lipids, when these form rod-like clusters. The lipid
molecules typically have a hydrophilic head and a hydrophobic tail so, in aqueous solution,
they spontaneously arrange themselves so that tails are away from the surrounding water, and
heads in contact with it. Depending on the shape of the particular molecule, they can form
spherical aggregates with tails pointing inwards and heads pointing outwards, or form lipid
bilayers, where lipid molecules form a double layer with heads on the external surface and
tails on the inside. Clusters formed by lipids in aqueous solution are called micelles, and the
process by which they form is called micellization. To determine the time scale, one needs a
measure of the kinetic coefficient of the d decay reaction, which was set equal to one. A
convenient relation could be an equation, which in dimensional units is <n> ≈ (d t)1/2. In case
the self-similar size distribution is not reached during the intermediate phase, another way to
determine d is to study the equilibration era and compare the experimental size distribution
with the numerical solution of the model. By combining early ~ s/ with <n> ≈ (d t)1/2 it is
obtained <n> ≈ (d s/ )1/2. The original software used in the investigation is available from
the authors.
CALCULATION RESULTS AND DISCUSSION

The line graphs in Figure 1 depict <k>2rk as a function of x = k/<k> for the times
= 0.5 105, 105 and 1.5 105. They are nearly superimposed on top of each other. The heavy
dots correspond to the plateau time = 20, so the change in the distribution shape over the
whole time span 20 < < 1.5 105 is not rather great.
Figure 1. Approximate self-similar behaviour of the size distribution at times = 50 000, 100 000 and
150 000 (solid lines). Notice that <k>2rk is approximately the same function of k/<k> at different times.
The dots correspond to = 20.
Figure 2 illustrates the evolution of the size distribution for the rescaled binding energy
= e– / = 4.54 10–4 (corresponding to = 10 and = 0.1), where the binding energy of the
k cluster k = (k–1) kBT, and is the ratio number of particles/number of spaces. It records
the time-dependent behaviour of the average cluster size <k>. It is a log-log plot of <k>/e vs.
. It reveals an initial rapid growth of <k> to a plateau value close to e, roughly located in the
interval 10 < < 100. In the subsequent growth after the plateau large clusters with k >> 1
eventually appear. Figure 2 indicates that by time = 5 104, k clusters having <k> ≈ 10 are
prevalent. In the time interval 2 104 < < 5 105, the log-log plot of <k>/e vs. is close to a
straight line of slope 1/2. This strongly supports the existence of a self-similar stage of the
kinetics. Notice that the average cluster size <k> corresponding to the intermediate transient
(Figure 2, dotted line) approaches the asymptotic value (straight line of slope 1/2).
Figure 2. Evolution of the average cluster size <k>/e vs. the scaled time (thick solid line). The dotted
line corresponds to the intermediate transient with an initial condition corresponding to the dot. The
straight line of slope 1/2 corresponds to the asymptotic self-similar continuum size distribution.
1000
C60
SWNT
Energy (K)
0 SWNT -correction
SWNT n
SWNT -correction n
-1000
0 20 40
Number of molecules in cluster
Figure 3. SWNT interaction energy with its surroundings in cluster volume or surface.
0.0008
C60
Solubility, molar fraction
0.0006
SWNT
SWNT -correction
0.0004
SWNT n
0.0002 SWNT -correction n
0
200 300 400
Temp erature (K)
Figure 4. Temperature dependence of solubility of C 60 (droplet) and SWNT (bundlet).
Figure 3 illustrates the equilibrium difference between the Gibbs free energies of
interaction of an SWNT with its surroundings, in the solid phase and in the cluster volume, or
on the cluster surface. On going from C60 (droplet model) to SWNT (bundlet) the minimum is
less marked (55% of droplet), which causes a lesser number of units in SWNT (nminimum ≈ 2)
than in C60 clusters (≈8). Moreover the abscissa is also shorter in SWNT (≈9) than in C60
clusters (≈28). In turn when the packing-efficiency correction (7) is included, the C60–SWNT
shortening decreases (68% of droplet) while keeping nminimum ≈ 2 and nabscissa ≈ 9.
The temperature dependence of SWNT solubility S (cf. Figure 4) shows that S decreases
with temperature, because of cluster formation. At T ≈ 260K, the C60 crystal presents an
orientation disorder phase transition from FCC to simple cubic. The reduction is less marked
for SWNT, in agreement with the lesser number of units in SWNT clusters. In particular at
T = 260K on going from C60 (droplet) to SWNT (bundlet), S drops to 1.6% of droplet. In turn
when the packing-efficiency correction is included (7), the shortening decreases (2.6% of
droplet).
The concentration C dependence of the heat of solution H in toluene, benzene and CS2,
calculated at solvent temperature T = 298.15K (cf. Figure 5), shows that for C60 (droplet), on
going from C < 0.1% of saturated (<n> ≈ 1) to C = 15% (<n> ≈ 7), H decreases by 73%. In
turn for SWNT (bundlet) H increases by 98% in the same interval. However, when the
packing-efficiency correction (7) is included, the increment in H is reduced to 54%. The
discrepancy between various experimental H data of fullerenes and SWNTs may be ascribed
to the sharp C dependence of H.
The results for the dependence of diffusion coefficient D on C in toluene, at T = 298.15K
(cf. Figure 6), show that the cluster formation in a solution close to saturation decreases D by
58%, 73% and 69% for C60, SWNT and SWNT with packing-efficiency correction,
respectively, as compared with D0 for an SWNT. For SWNT (bundlet) D decreases by 35%
with regard to droplet. In turn when the packing-efficiency correction (7) is included, the
decrease is reduced to 27%. The discrepancy between experimental data, on fullerene–SWNT
Ds, may be because of the sharp concentration dependence of D for the systems. The results
for SWNT (bundlet) with packing-efficiency correction and extrapolation n ∞ are
superposed to SWNT (bundlet, n ∞).
C60
Heat of solution (kJ/mol)
0
SWNT
SWNT -correction
-5 SWNT n
SWNT -correction n
-10
0 0.5 1
C/C sat
Figure 5. Heat of solution vs. concentration in toluene, benzene and CS2 at 298.15K.
8E-10
C60
Diffusion coefficient (m 2 /s)
SWNT
6E-10
SWNT -correction
SWNT n
4E-10 SWNT -correction n
0 0.5 1
C/C sat
Figure 6. Diffusion coefficient vs. concentration of C60/SWNT in toluene at 298.15K.
From the discussion of the present results the following inferences can be drawn.
1. Based on a simple kinetic model, micellization of rod-like aggregates occurs in three

separated stages: (a) many clusters of small size are produced while the number of
monomers decreases sharply; (b) aggregates are increasing steadily in size, and their
distribution approaches a self-similar solution of the diffusion equation; (c) a simple
mean-field Fokker–Planck equation describes the third era until the equilibrium
distribution is reached. In order to validate the theory by an experiment, it would be
important to measure the average cluster size as a function of time. To determine the
time scale, one needs a measure of the cluster diffusion coefficient d that was set
equal to 1. A convenient relation in dimensional units is <k> ≈ (d t)1/2. In case the
self-size similar distribution is not reached during the intermediate phase, another
way to determine d is to study the equilibration era and compare the experimentally
obtained size distribution with the numerical solution of the model. At equilibrium
<k>2 ≈ e , and this relation determines the dimensionless binding energy .
2. Fullerene–SWNT cluster formation suggests that the cluster sheath is filled with
pores. The membranous character of growth process in clusters explains
experimental data dispersion. The model yields an activation barrier and predicts that
pores with a radius below a certain critical value are unstable, while those above this
radius will grow indefinitely until the membrane ruptures. During the latter stage of
fusion the site expansion velocity slowed down by two orders of magnitude.
Dynamics were governed by the displacement of the volume of liquid around the
fusion site.
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__________________________________________________________________
Topic page
Lecture 13: Artificial fossilization process: A shortcut to nanostructured materials

from natural substances 296
Lecture 14: Biomimetic mineralization and mesocrystals 312
Lecture 15: Nano-fabricated structures and biomolecular analysis 342
Lecture 16: Bionic Superhydrophobic Surfaces Based on Colloidal Crystal Technique 360
Lecture 17: Nanomaterials:Green synthesis 397
Lecture 18: Targeted nanoparticles in cancer therapeutics 446
Lecture 19: Lithographically-Structured,gripping devices 466
Lecture Material 13
ARTIFICIAL FOSSILIZATION PROCESS:

A SHORTCUT TO NANOSTRUCTURED MATERIALS
FROM NATURAL SUBSTANCES
ABSTRACT
The combination of various synthetic chemical processes and biological assemblies
provides a promising strategy for the design and preparation of functional materials with
tailored structures and properties. Precise replication of natural substances with inorganic
matrices results in artificial materials possessing the initial biological structures and
morphologies. An so-called ―a rtificial fossilization process‖ was developed to achieve
inorganic replicas of the biological species which possess the corresponding finest
structure details and morphological hierarchies all the way down to nanometer scale. And
it was successfully applied to natural cellulosic substances such as filter paper, cotton and
cloth to obtain the corresponding metal oxide replicas. The resultant man-made materials
are hierarchical ceramics composed of metal oxide nanotubes, which are precise hollow
replicas of the initial cellulose nanofibers. This approach has been employed to prepare
various nanostructured metal oxide materials as well as metal oxide nanotube-metal
nanoparticle hybrid material.
INTRODUCTION
Replication of the morphologies and structures of natural substances with inorganic
matrix, that is, synthesis of the inorganic analogues of biological species, is called artificial
biomineralization; and it is believed that this specific replication can introduce some of the
superb properties of biological assemblies into man-made materials [1, 2]. Biological
organisms are produced from self-assembly of highly ordered functional units and are
inherently complex and hierarchical, possessing micro-to-nanoscale features. Employing
biological substances as templates for the formation of inorganic functional materials is a
facile, low-cost and environmentally benign short-cut to functional materials with unique
multilevel structures and morphologies.
A large variety of biological substances, such as shell, skeleton, wood, eggshell
membrane, diatom, bacterium, virus and living cell, as well as synthesized organic assemblies
such as sugar-based organogels[3, 4] and biomacromolecules such as DNA[5] and
polysaccharide [6], have been utilized with different chemical processes for the syntheses of
various inorganic materials like silica, titania, silicon and zeolite in the forms of
negative/positive or true copies of the initial biotemplates. Chemical processes like chemical
vapor deposition (CVD) [7], atomic layer deposition (ALD) [8], gas solid displacement
reactions [9], as well as wet-chemistry techniques such as sol gel polycondensation [4 6, 10]
were employed for the replication. While morphological replication in these instances have
been achieved only in the micrometer scale and the nanoscopic details could not be
reproduced. The structural organization of natural templates cannot be then finely controlled
in the resultant inorganic analogues, hence the unique morphological features of natural
substances are not maintained.
Therefore, it still remains a challenge to achieve inorganic replicas of the biological
species which possess the corresponding finest structure details and morphological
hierarchies all the way down to nanometer scale. The formation of fossils such as siliceous
woods is a short-cut route to reproduce morphological hierarchies of the original plants by
replacing the wood components with silica in intricate details. Such fossilization processes
would be realized artificially if morphologically complex surfaces of the biological structure
are faithfully lined with ultrathin inorganic layers accompanied by subsequent removal of the
organic template. To this end, an artificial fossilization process was developed by taking
advantage of the surface sol gel process to faithfully replicate the morphological hierarchies
of biological substances from macroscopic to nanometer scales, and it was successfully
applied to natural cellulosic substances such as filter paper, cotton and cloth to obtain their
metal oxide replicas.
The surface sol gel process is a facile methodology to deposit nanometer-thick metal
oxide films on hydroxyl- or carboxyl-terminated surfaces, which is based on chemical
adsorption of metal alkoxide from solution onto hydroxylated substrate surface to form
covalently-bound monolayer, followed by hydrolysis to give a new hydroxylated gel layer for
successive film deposition. Subnanometer thickness can be attained for an individual metal
oxide layer under carefully controlled experimental conditions [11].
The replication procedure employing the artificial fossilization process is schematically
illustrated in Scheme 1. Natural cellulose fibers possess abundant surface hydroxyl groups,
and provide a suitable substrate for metal oxide deposition via the surface sol gel process. In
this artificial fossilization process, ultrathin metal oxide gel films are deposited on
morphologically complex surfaces of the cellulosic substances with nanometer precision.
Their hierarchical morphologies are retained in metal oxide films to give macroscopic fossils
after removal of the organic substances by calcination. The resultant fossils are hierarchical
ceramic materials, in which the structures of the original substance are faithfully replicated.
The ceramics are composed of metal oxide nanotubes, as precise hollow replicas of the initial
cellulose nanofibers. This approach has been employed to prepare nanostructured titania,[12]
zirconia [12], tin oxide [13], and ITO [14] with tens to hundred nanometer outer diameters
and controllable wall thicknesses ca. ten nanometers, as well as hierarchical titania nanotube-
gold nanoparticle hybrid material [15]. And this process was applied to deposit ultrathin tin
oxide coatings on other kinds of biological templates like silica diatom frustules [16].
Scheme 1. Representative illustration of the precise replication of natural cellulose fibers by metal oxides.
NANO-PRECISION REPLICATION OF NATURAL CELLULOSIC

SUBSTANCES BY TITANIA
In order to make a ―titania fossil‖ of natural paper, titania gel films were deposited on the
morphologically complex surface of paper, and the resultant paper/titania composite was
calcined to remove the original filter paper. In a typical procedure, a piece of commercial
filter paper was placed in a suction filtering unit, and was washed by suction filtration of
ethanol, followed by drying with air flow. Ten ml of titanium n-butoxide solution (Ti(OnBu)4,
100 mM in 1:1/v:v toluene/ethanol) was then passed through the filter paper slowly within 2
minutes. Two 20-ml portions of ethanol was immediately filtered to remove the unreacted
metal alkoxide, and 20 ml of water was allowed to pass in order to promote hydrolysis and
condensation. Finally, the filter paper was dried with air flow. By repeating this
filtration/deposition cycle, thin titania gel layers covered the surface of the cellulose fibers.
The resultant paper/titania composite was calcined in air at 723 K for 6 hours (heating rate 1
K/min ) to remove the original filter paper.
The resultant titania fossil possessed morphological characteristics of the original filter
paper except for a little shrinkage in size due to calcination, which is commonly observed
after calcination of a sol–gel material (Figures 1a and 1b). The resulted titania sheet is self-
supporting and highly porous with thickness of ~0.22 mm and mass of ~1.5 mg. The sheet
size and thickness depend on the original filter paper used.
The original morphology of the filter paper was found to be faithfully replicated by
titania films, and the cellulose fibers were precisely copied as irregular titania nanotubes as
clearly recognized in SEM and TEM images (Figures 1c and 1d). The ―t itania paper‖ records
the morphological information of the original paper at the nanometer scale. The outer
diameter of the tube varies from 30 nm to 100 nm, and the thickness of the tube is uniform
along its length with wall thickness of ca. 10 nm. The wall thickness can be controlled by
changing the number of deposition cycles of titania layers. The titania nanotube assembly
manifests the original morphology of interwoven cellulose fibers, and the nano-branched
structure of the initial fibers can be clearly seen (Figure 1c). The SAED pattern from
agglomerated titania tubes shows diffraction rings typical of the anatase crystal, and it is
revealed by TEM observation that the titania nanotubes are composed of anatase fine particles
with sizes of around 10 nm.
Fihure 1. Nanoprecise titania replica of filter paper. (a) Photograph of a piece of filter paper after deposition
of 10-nm thick titania layer. (b) Photograph of pure titania sheet obtained by calcination of the filter paper
sample shown in (a). (c) FE-SEM image of the sample displayed in (b), showing titania nanotube assemblies.
(d) TEM image of an individual titania nanotube isolated from the assembly.
Similar replication processes are readily applied to other natural cellulosic substances like
cloth and cotton, resulting in ―t itania cloth‖ and ―t
itania cotton‖ (Figure 2). The hierarchical
morphologies of these natural substances are retained in titania films to give macroscopic
fossils, in which the structures of the original substance are again faithfully replicated from
macroscopic to nanometer scales. The fine titania thread shown in Figure 2a is a copy of an
individual fiber that makes up strands in the original cloth; and the fine titania hair displayed
in Figure 2b shows the spiral twist of natural cotton lint. The corresponding high
magnification SEM images demonstrate that both of them are composed of arrays of tortuous
titania nanotubes (insets of Figures 2a and 2b), as precise replicas of cellulose fiber
assemblies. The topographic differences among the titania fossils (as shown in Figures 1c, 2a
and 2b) mirrors the structural differences of the initial paper, cloth and cotton, although they
are all natural cellulosic substances.
Among various oxidic nanotubes, titania nanotube is particularly attractive due to its
unique electronic, photonic and catalytic properties. The current approach presents a practical
and environment friendly approach to produce titania nanotubes. Structural design of the
nanotube is achieved by proper selection of template materials. These features are not readily
attained by the reported chemical methods such as sol gel template syntheses using porous
membranes [17] and polymer fibers [18], or alkali treatment on titania powders [19].
The multi-helical morphology of the titania nanotubes shown in Figure 2b is worthy of
mention. Helical inorganic fibers are unique class of advanced functional materials, important
and challenging. As clearly shown here, replication of natural helical structures with
inorganic matrices can be a shortcut route to helical inorganic materials. It is known that each
natural cotton hair is a thin flattened tubular cell with a pronounced spiral twist when it is
fully mature and dry, and its length is several centimeters. Precise duplication of cotton hairs
with titania via the current petrifaction process gives ―t itania cotton‖ composed of multi-
helical titania nanotubes.
Fihure 2. Titania replicas of natural cellulosic substances. (a) SEM image of ―

titania cloth‖. (b) SEM image
of ―
titania cotton‖.
Fihure 3. Electron micrographs of ―zirconia paper‖. Deposition of zirconia thin films was repeated 20 times
for this sample. (a) FE-SEM image, showing zirconia nanotube assemblies. (b) TEM image of an individual
zirconia nanotube isolated from the assembly.
OTHER NANOTUBULAR METAL OXIDE MATERIALS DERIVED FROM

CELLULOSIC SUBSTANCES
Since the discovery of carbon nanotubes, nanotubular materials have been attracting great
attention in both fundamental and industrial studies due to their peculiar properties superior to
the corresponding bulk materials and isotropic nanoparticles [20]. In contrast, general and
efficient synthetic approaches have not been available for oxidic nanotubes [20a]. As
described above, the current ― artificial fossilization‖ process provides a shortcut to produce
metal oxide nanotubular materials. Metal oxides other than titania can be similarly employed
in the surface sol gel process, and are suitable as replicating matrices. For instance, Figure 3
shows artificial zirconia fossil derived from natural paper by using zirconium n-butoxide
(Zr(OnBu)4) as the precursor compound. The initial fiber assembly in paper leads to well
aligned zirconia nanotube arrays (Figure 3a), and the zirconia nanotubes are uniform with an
ultrathin wall thickness of ca. 10 nm (Figures 3b) and an extremely high aspect ratio (length
vs. diameter).
Tin oxide nanotubular materials were also prepared by using a natural cellulosic
substance (filter paper) as template [13]. Cellulose fibers of filter paper were firstly coated
with SnO2 gel layers by the surface sol gel process using Sn(OiPr)4 as precursor compound,
followed by calcination in air to give SnO2 nanotubular materials as hollow replicas of natural
cellulose fibers. The final resulted material was self-supporting ceramic sheet as being a
macroscopic fossil of the template filter paper. For an as-deposited sample prepared by
repeating the surface sol gel deposition cycle for twelve times, the SnO2 nanotubes obtained
by calcination at 450 ºC were amorphous-like and composed of fine particles with sizes
smaller than ca. 5 nm; the outer diameters are tens to two hundred nanometers and wall
thicknesses are 10 15 nm. While calcination at 1100 ºC yielded tube-like polycrystalline
SnO2 nanocages (outer diameter, 100 200 nm), which were composed of rutile-phase SnO2
nanocrystallites with sizes of 10 20 nm. In the previous examples given by other research
groups, cotton fibers were employed as template for the preparation of SnO2 microtubes by
chemical deposition technique through olation and heterogeneous nucleation of tin difluoride
[21]. The initial template morphology was replicated only on the micrometer scale by SnO2
matrix, resulting in SnO2 tubules with diameters and wall thicknesses in the micrometer
regime. Only by taking advantage of the surface sol gel process, the natural cellulosic
substance can be replicated at all levels of morphological hierarchies from nanometer to
centimeter regimes.
The conditions of thermal treatment strongly affect the structure of the SnO2 nanotubes
formed. Thermogravimetric analysis (TGA) showed that during calcination the as-deposited
sample experienced two-step weight losses at 71 °C and 319 °C, which indicated evaporation
of solvents (isopropanol and methanol) and combustion of the organic moiety, respectively.
The weight loss was 92 wt% up to 500 °C and then slightly decreased to reach 94 wt% at
1300 °C. It is clear that calcination temperatures above 500 °C are needed to obtain pure
SnO2 from the as-deposited composite sheet. Figure 4 shows XRD patterns of such SnO2
powders obtained at different calcination temperatures from 300 to 900 °C. All diffraction
peaks agree with the tetragonal structure (rutile type) in JCPDS 41-1445 (cassiterite), which
indicate that the specimens consist of SnO2. The intensity of each peak increased with
increasing calcination temperatures. In particular, the sample calcined at a temperature above

500 °C showed typical diffraction peaks of cassiterite, suggesting that the sample was
composed of pure SnO2 without any impurity. The crystallite size increased along with
calcination temperature gradually up to 700 °C, but more quickly after that temperature. The
crystallite size of the powder calcined at 300 °C was only about 2.0 nm. This is smaller than
that of SnO2 derived from SnCl4 (about 4 nm obtained at 300 °C) [22]. Even at the calcination
temperature of 900 °C, the crystallite size was suppressed to below 10 nm. TEM observation
indicated that sizes of SnO2 nanoparticles are ca. 5 nm and 10 20 nm at calcination
temperatures of 450 °C and 1100 °C, respectively. These data are in fair agreement with
crystallize sizes estimated for each calcinations temperature from the XRD measurements,
although the two sets of data for 1100 °C cannot be directly compared. In any case, it is clear
that crystal growth is suppressed even at high calcination temperatures. The mechanism of
such suppressed crystal growth of SnO2 in the current case is not clear. It may arise from the
unique preparation process of the SnO2 sheet. SnO2 nanoparticles are formed separately on
the cellulose fiber matrix and probably possess limited numbers of contacts with the
neighboring particles in the sheet.
Being a stable wide-bandgap n-type semiconductor, SnO2 is a promising key functional
material for a wide range of practical applications particularly for gas sensing. The unique
nano-cage morphology of the SnO2 nanotubes obtained herein could offer advantages in
fabricating novel gas sensors. The high surface area and the fine grain size could enhance the
interaction between SnO2 surface and gas molecules to be detected, and the nano-cage
structure would facilitate fast and full gas access to SnO2 nanocrystals. In the case of a thin-
film SnO2 sensor, concentration gradients of sensing gases may arise between the outermost
layer of the film and the bottom layer directly attached to the electrode substrate. Open
nanotubes will not have this problem. The tubular SnO2 nano-cages are hence expected to
provide better gas sensitivity and sensing reversibility compared with SnO2 films or
nanobelts. Figure 5 shows response transients of a SnO2 nanotube sheet sensor to 100 ppm
H2, 100 ppm CO and 20 ppm ethylene oxide in the working temperature range of 350 500
°C. The resistance (Ra) of the sensor in dry air was 1 2 106 Ω at tested temperatures and
increased slightly with decreasing temperatures. This Ra level is close to that of SnO2
crystallite [23]. Among the gases tested, the response transient to the smallest hydrogen gas
was larger than those to the other two gases, although a low concentration range was
employed for ethylene oxide. It is known that the sensitivity to target gas strongly depends on
the ease of diffusion of gas molecules inside the sensor. It is clear that hydrogen molecule is
diffused most easily inside the deeper region of the sensor and reacts with oxygen adsorbed
on SnO2 surface. Figure 6 shows dependence of sensitivity, S, of the test gases on the working
temperatures. The sensitivity to hydrogen gas gives a concave shape with a peak at 450 °C (S
16.5, which is comparable to the conventional SnO2 sensor), while S to the other two gases
show downward trends with temperatures. Unfortunately, these S values at 450 °C seems not
to be so high, compared with other reported results (S values are 500 and 1150 to 800 ppm H2
and 800 ppm CO respectively, at 350 °C) [23]. The grain size effect is well known for SnO2
gas sensor and the gas sensitivity increases drastically with crystallite size of less than 6 nm
[24]. The known relationship of crystallite size and sensitivity predicts that the gas sensitivity
of the current SnO2 nanotube could be much larger than the observed value. The present SnO2
sheet is composed of highly entangled nanotubes. Therefore, the access of gas molecules to
SnO2 nanotubes may not be favored, compared with that toward isolated nanotubes and to
bundles of a few nanotubes. It is also essential that nanotubes either have porous wall or tube
ends are open, in order to secure efficient access of gaseous reactants. Such favorable
situations are apparently not attained in the present case, and its sensitivity, S, is not superior
to that of the thin-film SnO2 sensors. For our current cellulose fiber templated SnO2
nanotubular materials, isolated and/or better aligned nanotubes could lead to enhanced gas
access.
Tin-doped indium oxide (indium tin oxide, ITO) is a promising degenerate wide band gap
n-type semiconductor, and has become the best-known transparent conductive oxide (TCO)
material in optoelectronics due to its relatively low resistivity, high optical transmittance in
the visible and near-infrared regions and high reflectance in the infrared region. Nanotubular
ITO materials, if they possess nanotube topography and electronic conductivity, could offer
unique opportunities in the development of functional devices and sensors. However, existing
investigation is extremely sparse. By employing the artificial fossilization process developed
herein, free-standing nanotubular ITO sheets with different In/Sn ratios were fabricated using
commercial filter paper as template. The resulting materials have a hierarchical structure
originating from the morphology of cellulosic paper, and the ITO nanotubes are composed of
interconnected layers of ITO nanocrystals of a few nanometers. As shown in Figures 7a and
7b, the resulted pale-yellow ITO sheet (inset of Figure 7a) is self-supporting, and its
morphological characteristics from macroscopic to microscopic sizes are similar to the
template material. It is composed of irregular nanotube networks (Figure 7a) originating from
the cellulose fiber network of the filter paper. The initial cellulose fiber assembly leads to
aligned ITO nanotube arrays, and individual nanotubes can be clearly identified. The
nanotubes possess very high aspect ratios, with outer diameters of a few tens of nanometers to
ca. two hundred nanometers. High-resolution FE-SEM images of the individual ITO
nanotube (Figure 7b) clearly show that the tube is composed of nanoparticles. This cage-like
nanotube morphology was confirmed by transmission electron microscopy (TEM), and the
ITO nanoparticle size is seen to be ca. 10 nm. The selected-area electron diffraction (SAED)
pattern indicates the polycrystalline nature of the ITO nanotube.
ITO gel layers were deposited on individual cellulose nanofibers by using precursor
solutions of indium methoxyethoxide and tetraisopropoxytin with a total concentration of 12
mM with different In/Sn molar ratios. The precursor mixtures of In/Sn ratio of 10/0, 9/1, 2/1,
2/8 and 0/10 were used and named as In10, In9Sn1, In2Sn1, In2Sn8 and Sn10 hereafter,
respectively, from the In/Sn ratio in the precursor solution. The practical In/Sn ratio in the
nanotube sheets was determined by electron probe micro analysis (EPMA) (Table 1). The
observed In/Sn ratio is always smaller than that of the precursor solution. This difference may
be caused by the greater reactivity of the indium alkoxide relative to that of the tin alkoxide
[11]. The average thickness of ITO sheet, measured by optical microscopy, is in the range of
90-220 m (Table 1). The apparent density of ITO sheet was determined from the nominal
volume and weight of 10 10 mm square pieces of the replica. It was very low, being in the
range of 1.8 4.7% of the ideal density calculated from the bulk density of In2O3 and SnO2
and the observed In/Sn ratio (Table 1). This indicates that the replica sheet is highly porous.
Fihure 4. XRD patterns of the SnO2 powder obtained by calcination of the as-deposited SnO2 sheet for 3 h at
various temperatures.
Fihure 5. Response transients of SnO2 nanotubes to 100 ppm H2, 100 ppm CO and 20 ppm C2H4O at varied
temperatures.
Fihure 6. Temperature dependence of sensitivities of SnO2 nano-tube sensor to 100 ppm H2, 100 ppm CO and
20 ppm C2H4O.
Figure 7. ITO (In2Sn1) nanotubes obtained by the artificial fossil approach using commercial filter paper as
template, deposition of ITO thin films was repeated 12 times for this sample. (a) Low-magnification FE-SEM
micrograph of the ITO nanotube sheet, showing nanotube networks; inset is a macroscopic photograph of the
sheet, which was obtained by calcination of a half of an as-deposited ITO gel/filter paper composite sheet. (b)
FE-SEM image of one individual ITO nanotube isolated from the assembly, inset shows high-magnification
image of the boxed area. (c) Arrhenius plots of the electrical conductivity of nanotubular ITO sheet. (d) I-V
curves of In9Sn1 at 20 C.
Table 1. Compositions and characteristics of ITO nanotubular sheets
†
Precursor In/Sn ratio Thickness / Density / Fractional 298 / Ea /
mixture m g cm-3 Density / % S cm -1
kJ
mol
In10 98 5 0.33 4.7 5.89 10-3 16
In9Sn1 93.5/6.5 218 12 0.23 3.3 0.533 1.2
(14.4/1)
In2Sn1 80.3/19.7 95 11 0.13 1.9 7.58 10-3 4.0
(4.1/1)
In2Sn8 30.4/69.6 172 26 0.16 2.2 4.27 10-3 20
(1/2.3)
Sn10 165 19 0.13 1.8 1.03 10-3 13
†
at 298 K.
The powder XRD study revealed that the crystal phases of the ITO sheets varied with
changing metal compositions. In10 sample consists of the cubic In2O3 phase and a small
amount of the rhombohedral In2O3 phase. In9Sn1 has a rhombohedral In2O3-type phase. The
preferential formation of a rhombohedral phase may be due to the small particle-size, since
the current sample consists of ITO nanoparticles less than 10 nm in diameter. Sn10 consists
of a single tertragonal SnO2 phase. In2Sn8 sample also shows the single pattern of the
tetragonal SnO2-type phase, but the peak width is broader than that of Sn10. In2Sn1 is a
mixture of rhombohedral In4Sn3O12 and rhombohedral In2O3-type phases, although the
relative intensity of peaks is much weaker than those of others. These XRD results suggested
that the nanoparticles in the ITO nanotube are crystalline and do not reveal phase separation
between indium and tin oxide.
The electrical conductivity parallel to the sheet surface was measured for a free-standing
ITO sheet by four-probing van der Pauw method. Arrhenius plots of electrical conductivity,
, of ITO sheets indicate that all the sample are semiconducting with increasing with
1
temperature (Figure 7c). In9Sn1 reveals the highest , 0.53 S cm , at ambient temperature
and the smallest activation energy, Ea, (1.2 kJ mol 1) of all the ITO sheets (Table 1). In2Sn1
exhibits an Ea value similar to that of In9Sn1, but its is lower than that of In9Sn1 by a factor
of 102 over the measured temperature range. Other three samples show apparent temperature
dependence of with large Ea compared to those of In9Sn1 and In2Sn1. The values of In10
and In2Sn8 are in the order of 10 3 S cm-1 at 20 C, but they increase to about 0.3 0.4 S cm 1
with temperature rise to 400 C. The conventional ITO materials, such as dense film, single
crystal and sintered pellet reveal a metallic behavior at 300 700 K, the conductivity decreases
with temperature increasing. The current ITO sheets exhibit the opposite temperature
dependence, indicating that its scattering mechanism is different. The I-V curve of In9Sn1 at
room temperature is not linear in nature, and shows an upward deviation from the Ohm‘s law
at the voltage region higher than 5 V (Figure 7d). Other ITO sheets also show a similar
character in I-V curves. This feature of the I-V curve can be attributed to the effect of charge
trapping at the grain boundary. The grain boundary of polycrystalline semiconductors
contains a large amount of point defects which induce trapping of the carriers. This charge
trapping produces the depletion layer with a potential barrier at the grain boundary, impeding
the carrier motion from one crystallite to another. The influence of such grain boundary
scattering must be significant for our samples, because they are composed of the particle of a
few nm that is commensurate with the mean-free path (~ 3 nm) in nanocrystalline ITO films.
The electrical transport property of the free-standing ITO sheet is determined mainly by the
nature of the grain boundary. The other possible scattering mechanism is related to
crystallinity, the effect of low crystallinity of our sample to conductivity is rather small. The
value of In9Sn1 at 20 C (0.53 S cm 1) is lower than that of the commercial ITO (5 103 S
cm 1) by a factor of 104. However, when the conductivity was corrected for the apparent
density, the effective conductivity of solid In9Sn1 was calculated to be 160 S cm 1. This
value is only an order of magnitude lower than that of the commercial ITO film. And the
effective electrical conductivity of the present free-standing ITO sheet is apparently higher
than those of other nanostructured ITO materials, if effective conductivities are compared.
The high conductivity of the current nanotube may originate from its structural feature.
The uniform ITO gel layer is formed on the template cellulose fiber with nanometer
precision, and this gel layer is converted upon calcination to nanotubes composed of
interconnected nanocrystals. Since macroscopic morphology and the free-standing nature of

the template filter paper are maintained even after calcination, the interconnection is extended
from nanometer to macroscopic scales. This means that all the nanocrystals are covalently
connected up to the macroscopic scale (Figures 7a and 7b) in spite of extremely small space
occupation of the specimen. Such a structural property of the sample may be efficient for the
electron percolation than that of the pelletized sample of mesoporous ITO powder, where the
electrical contact between the particles is achieved by physical contacts. The covalent
connection of nanocrystals may also lower the grain boundary scattering, since the chemical
bonding at grain boundaries can decrease the number of charge trapping sites such as
dangling bonds. The ITO sheet with In/Sn ratio of 93.5/6.5 exhibited the highest effective-
electrical-conductivity of 160 S cm-1, and this value is higher than those of the nanostructured
ITO prepared by other template-synthesis process as well as than that of the single-crystalline
nanowhisker prepared by vapor-liquid-solid (VLS) technique. The morphological and
electrical characteristics in the current specimen arise from the covalent interconnection of
ITO nanocrystals from nanometer scale up to macroscopic scale. The morphological
adaptability of the current ITO system is superior to those of other conductive transition-
metal oxides such as CuO, NiO and MnO2. The latter metal oxides would not give flexible,
nano-precision morphologies, even if proper template structures are used. Therefore, the
nano-structured ITO, for example, single ITO nanotube, may find unique applications in
nano/micro-sized electronic devices. On the macroscopic scale, combination of high electrical
conductivity and high surface area, as exemplified by the nanotubular ITO sheet, should be
advantageous as electrodes in many applications such as electrochemical battery,
electrochemical capacitor, electrolytic wastewater treatment, and light-electricity conversion
system. It is anticipated that the nanostructured ITO sheet affords a novel strategy to design
electronic micro-devices and electrode materials.
HIGHLY STABILIZED HIERARCHICAL HYBRID OF TITANIA

NANOTUBE AND GOLD NANOPARTICLE
The artificial fossilization process developed herein makes it possible to use natural
substances as scaffold and platform for the fabrication of functional composite nanostructured
materials. Here gives an example of nanoparticle/nanotube hybrid. [15] Nanoparticle/nanotube
hybrid materials in which certain nanoparticles are attached onto the wall of host nanotubes
combine unique structural features of nanotubes and outstanding functionality of
nanoparticles, and promise wide applications such as heterogeneous catalysis and molecular
sensors. The titania nanotube and gold nanoparticle hybrid nanomaterial synthesized herein is
composed of gold nanoparticles and titania nanotubes with the morphological hierarchy of the
template cellulosic substance (filter paper). The inset of Figure 8a shows the resulted bulk
material. For this sample, 15 layers of titania films were firstly deposited on the cellulose
nanofibers, followed by adsorption of a layer of gold nanoparticles (positively-charged), and
5 more layers of titania film were additionally deposited. The as-deposited sample was then
subjected to calcination to remove the original filter paper as well as the organic ligand on
gold nanoparticles, giving a dark-brown gold/titania hybrid sheet. This gold/titania composite
sheet is self-supporting and weighs ~2.4 mg. It contains as much as ~40% Au by weight.
An FE-SEM image displayed in Figure 8a shows that the gold/titania hybrid sheet is
composed of hierarchically templated assemblies of gold nanoparticles and titania nanotubes.
The individual hybrid nanotube is uniform with an extremely high aspect ratio, and possesses
tube walls of ca. 10 nm thickness. The individual gold nanoparticles were attached to titania
nanotube uniformly as monolayer. The TEM image displayed in Figure 8b visualize high
coverage of the titania nanotube with gold nanoparticles. Each gold nanoparticle is covered
by an ultrathin titania shell (Figure 8c). The interparticle distance of ca. 3 nm is apparently
determined by presence of the long chain organic ligand on the gold nanoparticle employed.
Figure 8. A hierarchical hybrid material of titania nanotubes and gold nanoparticles, [(TiO2)15/Au-
nanoparticle/(TiO2)5], derived from filter paper. (a) FE-SEM image of the hybrid. The inset shows a
macroscopic photograph of the material. (b) TEM image of an individual titania nanotube that is fully coated
with gold nanoparticles. (c) Schematic illustration of the gold nanoparticle/titania nanotube hybrid (not to
scale).
The titania thin layer is additionally deposited on the gold nanoparticle, so that the
individual nanoparticles are wholly covered by the titania layer. Gold nanoparticles are
known to undergo melting at relatively low temperatures [25], and this facilitates fusion of
the unprotected nanoparticle. In the case of gold nanoparticles (sizes, 6 1 nm) on carbon
nanotube, the fusion was observed after heating for 30 s at 300 C [26]. The titania layers that
surround individual gold particles should suppress fusion of adjacent gold particles even at
higher temperatures. The gold nanoparticles are protected by coating with 5 titania layers
(thickness ~2.5 nm) in the case of sample [(TiO2)15/Au-nanoparticle/(TiO2)5]; and their
average size and the standard deviation are 4.9 and 1.4 nm, respectively. In fact, the original
size distribution (5 1 nm) is not altered after a long period of calcination (6 h at 450 C). In
contrast, the particle fusion was observed when the nanoparticle was not protected by the
titania layer. For the hybrid material of titania nanotube and gold nanoparticle, it can be
ensured with large surface areas, high and uniform metal loading, as well as enhanced particle
stability in the hierarchical morphology. The one-pot fabrication of such complex loading
matrices is rendered feasible by appropriate design of hierarchical templates, and should be
extremely beneficial from the practical standpoint. Combining the rich varieties of
nanoparticles and ceramic nanotubes, the present approach can produce versatile nano-
precision systems with unique physical and chemical functions
SUMMARY
In summary, a general chemical procedure, artificial fossilization process, was developed
for nanoscale to macroscale duplication of the complex hierarchical morphology of natural
cellulosic substances with metal oxide matrices. It extends the range of replication techniques
that already exist, by allowing hollow replication on both the micron and the nanometer scale
simultaneously. This new nano-copying methodology provides replicas (both positive and
negative) of targeted objects in nanometer precision. As pointed out by R. A. Caruso [12b],
this synthetic procedure provides a pathway to probe structures of biosystems at nanometer
scales as well, and obviously is a practical, low-cost and environmentally friendly route to
produce nanotubular ceramic materials with unique structural features. And Moreover, this
method was extended to fabricate nanostructured conjugated polymer material [27] and
functional bioactive material [28] using natural cellulosic substances as scaffolds.
The biotemplate-derived functional materials have shown promising potentials for
various practical applications due to their unique structures and properties like high inner
surface area. The artificial biomineralization science and technology are at the crossing point
of biology, chemistry, physics, and materials science; and is a bridge to connect the new-age
nanotechnology and classic biological science. Precise replication of the natural three-
dimensional biological structures at the nanometer spatial scales and further at single
molecular level with a certain guest material is still a fairly unexplored field. New
methodologies such as nanocopy technique [29] are now in great demand.
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Lecture Material 14
BIOMIMETIC MINERALIZATION
AND MESOCRYSTALS
ABSTRACT
Self-assembly into highly ordered superstructures and control over the shape and the
size of inorganic materials is an important character of natural growth phenomena. In
biological systems, the results of long-term evolutionary optimization processes are
intimately related to specific functions. Bio-inspired materials synthesis is a powerful
strategy for the synthesis of advanced materials with complex shape, hierarchical
organization and controlled size, structure and polymorph in aqueous environments under
ambient conditions. Increased understanding of biomineralization mechanisms has
greatly enhanced the possibilities of biomimetic mineralization and template synthesis
approaches. Bio-inspired materials with complex structures and advanced functions
always attract attention because of their unique properties, which have paved the way to
many potential applications. Organic templates such as biopolymers and synthetic
amphiphilic polymers can be employed to understand the interaction of the organic
matrix with the developing inorganic crystals at a molecular level and to address the
question which factors lead to the remarkable crystallographic orientation of the
crystalline phase, crystal growth and nanoparticle assembly, which is often observed in
biomineralization. Clear evidence has shown that crystallization does not necessarily
proceed along the classical crystallization process, which is the attachment of
ions/molecules to a primary nanoparticle forming a single crystal, instead, crystallization
can also proceed along a particle mediated self-assembly pathway. Mesocrystal is a
quasi-single crystal consisting of ordered assemblies of small, anisotropic, and vectorially
aligned nanoparticles, thus forming an entirely new class of porous metamaterials
through mesoscopic transformations and nanoparticle precursors. Such composite
crystals are of considerable interest to a broad range of disciplines including materials
chemistry and life science as well as crystallization-related fields in general. It has been
found that nanoclustered crystal growth, mediated by organic templates, is a basic
characteristic of biomineralization that enables the formation of composite materials with
elaborate morphologies and structures. During the past decade, exploration as well as
application of these bio-inspired synthesis strategies has led to novel materials with
specific size, shape, orientation, composition, hierarchical organization and assembled
superstructures. This overview tries to capture the concepts and recent progress in this
rapidly developing field, and prospects for the future in this field has been discussed.
INTRODUCTION
Learning from nature is a constant principle for there are numerous mysterious features in
nature, which have developed over millions of years of evolution and will inspire us to
develop new functional materials. Bio-inspired and biomimetic concepts borrowed from
Nature have been developed for synthesizing novel functional materials such as bio-inorganic
materials, bio-inspired, multiscale structured materials, bio-nanomaterials, hybrid
organic/inorganic implant materials, and smart biomaterials [1]. These bio-inspired, smart
materials are attracting much interest because of their unique properties, which have paved
the way to many potential applications. Interfaces between biomolecules and inorganic
materials have been the focus of research in various fields such as biochemistry, materials
chemistry, biomedicine and bionanomaterials.
Materials with complex shapes and interface functions always attract attention and
fascination. In view of the huge time frame Nature had to optimize and perfect functional
materials, it is obvious that scientists are highly interested to develop synthetic strategies that
mimic these natural processes. Especially promising materials in this respect are biominerals,
which combine complex morphology over different length scales with superior materials
properties and environmentally friendly synthesis and biocompatibility. This makes them
very attractive archetypes for materials chemists. To mimic the synthesis of these materials,
the main purpose is not to simply emulate a particular biological architecture or system, but to
abstract the guiding principles and ideas and use such knowledge for the preparation of new
synthetic materials and devices. Based on these concepts a rapidly developing research field
has evolved, which can be summarized as bioinspired or biomimetic materials chemistry [2].
The creation of superstructures resembling naturally existing biominerals with their unusual
shapes and complexity, is meanwhile an important branch in the broad area of biomimetics
[3]. During the past decade, exploration as well as application of these bio-inspired synthesis
strategies has resulted in materials with specific size, shape, orientation, composition, and
hierarchical organization [4]. This chapter will summarize these developments and give some
examples what can already be achieved by applying natures‘ strategies for biomineral
synthesis. This review can not be comprehensive, therefore we have selected some topics,
those are closely mimicking biomineralization strategies. We consider biomimetic
mineralization as mineralization in aqueous solutions under ambient or nearly ambient
conditions borrowing strategies from biomineralization processes.
This chapter is organized into five sections. We have structured the paper in a way that
first the main concepts underlying biomineralization are introduced and then, the specific
examples for biomimetic mineralization are given in the following. The main contents of this
review involve strategies for biomineralization, biomimetic mineralization and mechanisms,
Non classical crystallization and mesocrystals, and bio-inspired functional nanomaterials.
Finally, summary and outlook is discussed.
BIOMINERALIZATION
Biomineralization is the process by which living organisms secrete inorganic minerals in
form of skeletons, shells, bone, teeth, magnetic iron minerals in bacteria, etc. (Figure 1) [5]. It
is already a rather old process in the development of life, which was adapted by living beings
probably at the end of the Precambrium more than 500 million years ago [6]. Materials found
in nature combine many inspiring properties such as sophistication, miniaturization,
hierarchical organizations, hybridation, resistance and adaptability [7]. Elucidating the basic
components and building principles selected by evolution to propose more reliable, efficient
and environment respecting materials requires a multidisciplinary research.
Biominerals are highly organized from the molecular to the nano- and macroscales, often
in a hierarchical manner, with intricate nanostructures those ultimately make up a lot of
different functional soft and hard tissues (Figure 1). Under genetic control, biological tissues
are produced in aqueous environments under mild physiological conditions by using
biomolecules, primarily proteins but also carbohydrates and lipids. Biomolecules both collect
and transport raw materials, and consistently and uniformly self- and co-assemble subunits
into short- and long-range-ordered nuclei and substrates [8]. For example, magnetotactic
bacteria, are able to form nano-sized, membrane-bound magnetic iron minerals, magnetite
(Fe3O4) or greigite (Fe3S4), by a mineralization process with precise biological control over
iron accumulation and mineral deposition [9]. The unexpected and unusual features of these
biogenic magnetite crystals are not only a narrow size distribution, but above all, a diameter
range of 40±120 nm, which thus allocates them the highest magnetic moment. This diameter
range corresponds to magnetite crystals with a single magnetic domain [10] (Figure 1f). More
examples of biominerals can be addressed as the caption in Figure 1. Whether in controlling
biomineral formation, biological functions or physical performance, bimolecules are an
indispensable part of biological structures and systems. A simple conclusion is that next-
generation biomimetic systems should include biomolecules in synthesis, assembly or
function [11].
a b c
d e f
Figure 1. Representative examples of biologically synthesized complex biominerals. (a) scanning electron
microscope (SEM) image of mouse enamel. It is a hard, wear-resistant material with highly ordered
micro/nano superstructure comprised of hydroxyapatite crystallites that assemble into woven rod architecture
(inset: schematic cross-section of a human tooth) [12]. (b) SEM image of a growth edge of abalone (Haliotis
rufescens) displaying aragonite platelets (blue) separated by organic film (in orange) that finally becomes
nacre (mother-of-pearl). This is a layered, tough, and high-strength biocomposite (inset: transmission electron
microscope (TEM) image [13]. (c) sponge spicule (with a cross-shaped apex shown in inset) of Rosella has
layered silica with excellent optical and mechanical properties, a biological optical fiber (SEM image) [14].
(d) SEM image of siliceous skeletal structures in diatomaceous earth. Actinopoda and diatoms, single-celled
organisms, create amorphous siliceous units that are resting spores with highly intricate and symmetrical
geometrical shapes [15]. (e) SEM of the peripheral layer of a dorsal arm plate (DAP) from Ophiocoma
wendtii with the microlenses structures in brittlestars. Skeletal elements of echinoderms are each composed
of a single crystal of oriented calcite shaped into a unique, three-dimensional mesh. Ophiocoma wendtii is a
highly photosensitive species, and it changes color markedly, from homogeneous dark brown during the day
to banded grey and black at night [16]. (f) magnetite nanoparticles formed by magnetotactic bacterium
(Aquaspirillum magnetotacticum, inset: TEM image) are single-crystalline, single-domained and
crystallographically aligned [9].
It is commonly assumed in the biomineralization field, that the remarkable biomaterials

morphologies are fabricated under total control of specific biomolecules so that
biomineralization is eventually a genetically controlled process, which transforms the
genetically engineered organic scaffolds into soft and hard matter. On the other hand, the
recent nacre retrosynthesis example [17] has already indicated that the natural biopolymers
can be replaced by synthetic polymer analogues so that some of the biomineralization
mechanisms are much related to physicochemical principles such as nucleation inhibition
rather than specific biopolymer structures and functions. Although a protein may exhibit a
complex structure, its actual function may be simple, for example, serving as a polyelectrolyte
in a biomineralization process. Therefore, it makes sense to investigate, on how far
physicochemical principles play a role in biomineralization. An excellent example for this is
the pattern formation in diatoms, which can be explained by a phase separation model of
amphiphilic polyamines (Figure 2) [18].
Figure 2. Schematic drawing of the templating mechanism by the phase-separation model (a–d) and
comparison with the stages of the developing cell wall of C. wailesii (e–h). (a) The monolayer of polyamine-
containing droplets in close-packed arrangement within the silica-deposition vesicle guides silica deposition.
(b, c) Consecutive segregations of smaller (about 300 nm) droplets open new routes for silica precipitation.
(d) Dispersion of 300 nm droplets into 50 nm droplets guides the final stage of silica deposition. Silica
precipitation only occurs within the water phase (white areas). The repeated phase separations produce a
hierarchy of self-similar patterns. (e–h) SEM images of valves in statue nascendi at the corresponding stages
of development. Reprinted from Ref. [18] with permission of the American Association for the Advancement
of Science.
In this model, amphiphilic polyamines phase separate and form an emulsion of closely
packed microdroplets in a hexagonally arranged monolayer within the flat silica deposition
vesicle (SDV, Figure 2a). Phase separation is induced by coordination of positively charged
polyamines with phosphate, and also incorporated in the polyamine itself in form of
phosphoserins [19], which acts as a cross linking agent [20].
The silica precursors in form of a polyamine stabilized sol are located at the aqueous
interfaces between the microdroplets. As a consequence, upon silica formation, a honeycomb-
like hexagonal framework is produced (Figure 2e). After partial consumption of the
polyamines by inclusion into silica, a further segregation of the initial microdroplets into
smaller droplets is assumed (Figure 2b). The newly created interfaces again serve as the
template for silica deposition (Figure 2f) and a further fraction of the polyamines is
consumed. This leads to a further phase separation of the polyamines into 300 nm sized
droplets (Figure 2c) and silica deposition at the aqueous interface between the droplets
(Figure 2g). A final phase separation of the nanodroplets into only 50 nm sized droplets
(Figure 2d) with subsequent mineralization of the aqueous interface between the droplets
results in the observed hierarchically hexagonal self-similar patterns. This pattern formation
mechanism, fully based on a series of consecutive phase separation steps, results in a pattern,
which matches that of the diatom valve of Coscinodiscus [18].
Above result indicates that such physicochemical principles can be transferred to
biomimetic mineralization for the generation of advanced materials as summarized for the
polyamine case in Ref. [21]. It will be interesting to reveal further physicochemical
mechanisms in bio- and biomimetic mineralization such as the minimization of interface
energies by the formation of an amorphous surface layer and growth inhibition by foreign
additives/impurities [22]. These principles will make us to a deeper understanding of
biomineralization processes and thus extend the toolbox of biomimetic mineralization by a
transfer of biomineralization principles to the synthetic materials field.
BIOMIMETIC MINERALIZATION
Biomimetic mineralization based on the biomineralization principle of templating of
inorganic structures by soft organic templates has already been transferred to materials
synthetic science. Organic templates can be employed to understand the interaction of the
organic matrix with the developing inorganic crystals at a molecular level and to address the
question which factor result in the remarkable crystallographic orientation of the crystalline
phase, which is often found in biomineralization. The templates are thus used as mimics of an
oriented structural matrix in biomineralization. Importance in this respect is Langmuir
monolayers as a template, because they are available with different head groups and can be
compressed so that a range of regular template structures can be adjusted. There have been
numerous reports on the selective nucleation of certain crystal faces under Langmuir
monolayers as summarized in Refs [23], and initially, epitaxy or stereochemical resp.
geometrical match between the arrangement of the charged groups of the monolayer and the
ion arrangement on the nucleated crystal surface was discussed. More recently, Cavalli et al.
demonstrated that flexible self-organizing -sheet lipopeptide monolayers led to a new
growth habit of calcite and thus the formation of indented calcite crystals. This study
confirmed the importance of flexibility of the template in crystal oriented nucleation [24]. In
addition to Langmuir monolayers, self assembled monolayers (SAM‘s) can also be used to
investigate the influence of the functional groups and other parameters on the mineral
deposition and orientation. The advantage is that the SAM‘s are chemically fixed to the
substrate and that they can be patterned by PDMS stamping or surface lithography.
The most remarkable result with patterned SAM‘s was reported by Aizenberg et al. for the
direct fabrication of large micropatterned calcite single crystals [25], which can be considered
to be a rough mimic of the oriented single crystal calcite microlens arrays in brittlestars [16].
First, photoresist micropatterns were formed on a glass surface by photolithography. Then,
the surface was coated with Au or Ag. As a very important step, a localized nanoregion of a
polar alkanethiol was deposited on the surface with an AFM tip serving as a single nucleation
center for calcite with a known orientation. Then, the remainder of the metal surface was
coated with alkanethiols with varying length and functionality, which created a disordered
surface and therefore favored the formation of amorphous calcium carbonate (ACC). First, a
mesh of metastable ACC filled the interstices of the framework followed by site specific
nucleation of a calcite single crystal at the deposited nucleation spot. This single crystal grew
by transformation of the surrounding ACC finally leading to a micropatterned single crystal
as imaged by polarization microscopy. The micropatterned surface was found to serve for the
release of stresses, water and impurities during the formation of the final crystal.
As a soft template used in biomimetic synthesis, double hydrophilic block copolymers
(DHBCs) are a new class of amphiphilic macromolecules of rapidly increasing interest. They
are water-soluble polymers in which amphiphilicity can be induced through the presence of a
substrate or by temperature and pH changes. Their chemical structure can be tuned for a wide
range of applications such as colloid stabilization, crystal growth modification, induced
micelle formation. In particular, mineralization processes can be controlled by using DHBCs
as inspired by biology, which have a molecular head group reacting with the metal ions and a
central non-reactive part similar to proteins containing hydrophilic and mineralophilic sites
[26]. Such polymers help to control the size, mineral forms, structure and assemblies of
inorganic crystals. Indeed, original superstructures have been prepared, as well as aligned
hydroxyapatite whiskers or mineral crystals having complex morphologies [2, 26].
More recently, a DHBC polymer with high molecular weight has been applied for the
crystallization process of BaCO3 crystals [27]. At the critical point between aggregation
towards long fibres and spheres, the short nanofibers at starting pH 5.5 self-organized towards
most striking dynamic ring structure patterns on the large scale. Figure 3 shows typical SEM
images of the obtained quasiperiodic wave patterns grown in solution for 1 day. Light
microscopy images indicate that those patterns were already formed in the aqueous solution.
This periodic wave pattern has multiple centres, from which concentric rings with even
spacing radiate outwards (Figure 3a), reminiscent of the target (concentric) waves in the
spatially extended Belousov-Zhabotinsky (BZ) reaction. A set of coupled chemical reactions
necessary for the establishment of a reaction-diffusion system could be formulated including
an autocatalytic formation of a Ba-polymer complex. On the substrate, many groups of
concentric rings grow at the same time and stop when merging with each other. The enlarged
SEM image shows that each ring (band) is composed of short nanorods standing on the
substrate instead of lying, and tending to form bundles on the substrate, which are organized
into a circular pattern around the center (Figure 3b, c). It is noteworthy that the experimental
window is narrow for the formation of this concentric circle pattern [27].
The periodic pattern formation in this reaction system belongs to a self-organization
process, in which competition between autocatalytic particle growth and educt diffusion
occurs. This concentric circle pattern is a vivid and ubiquitous phenomenon in the reaction-
diffusion system. In the meantime, numerical simulations using a modified Oregonator model
for the reaction-diffusion equations qualitatively agree with the experimental observations.
Figure 4d shows the simulation result for the BZ reaction, similar to the experimental ring
patterns. It is important to note that similar patterns can also be found in natural minerals;
spiral patterns of nacre (aragonite CaCO3) have been found on the growing inner surface of
nacre [28], and in that case, screw dislocations were believed to be responsible for spiral
growth of nacre.
Calcite crystals with exposed (001) faces have recently been obtained in the presence of
poly(sodium 4-styrene sulfonate-co-N-isopropylacrylamide) (PSS-co-PNIPAAM) (Figure 4a,
b) [29]. Usually, calcite is not able to expose the (001) faces because these faces are
composed of only CO32 or Ca2+ ions in a hexagonal orientation, respectively, and therefore
are highly charged faces. Such highly charged faces would exhibit high surface energies and
cannot exist in the absence of growth modifiers. The fact that this face now becomes
dominant can be ascribed to multiple Coulomb binding of the negatively charged polymer
molecules to the positively charged (001) plane. This leads to surface stabilization and
inhibition of growth along this direction. The oriented self-assembly of subunits toward
larger, single-crystalline superstructures results in mesocrystal formation based on non-
classical crystallization processes [30]. A Cerius2-colour model of the double truncated
trigonal calcite structure viewed along the 001 face is shown in Figure 4c, in agreement with
our experimental observations.
This assignment of faces and orientation can be well revealed by polarized optical
microscopy images of calcite mesocrystals which show that the exposed faces are not
birefringent, confirming that the new exposed face is (001). The (001) direction, due to its
symmetry, is the only axis in calcite which is not birefringent. In addition, the mesocrystals
are not symmetrical along the [001] axis. While one side of the mesocrystals exhibits the
truncated (001) face as shown in Figure 4b, the opposite side of the mesocrystal has the shape
of a pyramid tip. This anisotropic shape supports the model of mesocrystal assembly from
nanoparticles by dipole fields suggested in the previous work [31], and leading to an
assembly with opposite charge of the opposite sides along the [001] axis with the truncated
(001) face being the positive face, the pyramid tip negatively charged.
Figure 3. SEM images (a-c) of the obtained concentric circle pattern of BaCO3 crystals grown for 1 day.
[polymer] = 1 g L 1, [BaCl2] = 10 mM, starting pH = 5.5. (d) showing a simulated pattern using a modified
Oregonator model for the reaction-diffusion equations. Reproduced with permission from Angew. Chem. Int.
Ed., 2006, 45, 4451. Copyright © 2006, Wiley-VCH.
Figure 4. SEM images (a, b) of the obtained calcite superstructure with (001) faces exposed grown for 6 days,
polymer: 1 g L−1, [CaCl2] = 10 mM. (c) Cerius2-colour model of double truncated trigonal calcite structure
viewed upon the 001 face (yellow), red = (104) faces. (d) SEM image of the obtained vaterite hexagonal
plates grown for 6 days at a middle polymer and CaCl2 concentration. Polymer: 0.5 g L 1, [CaCl2] = 5 mM.
(e) SEM image of the aragonite superstructure obtained at the lower polymer and CaCl2 concentrations.
Polymer, 0.1 g L 1, [CaCl2] = 1.25 mM, 6 days. Reproduced with permission from Wiley-VCH.
When polymer and CaCl2 with an intermediate concentration (polymer: 0.5 g L 1, [CaCl2]
= 5 mM) was used for crystallization, the vaterite phase was obtained after the same reaction
time (6 d). Figure 4d presents a typical SEM image of the obtained sample showing
predominant vaterite hexagonal plates, slightly contaminated with minor traces of aragonite.
Obviously, these particles with the diameter ranging from about 15 to 20 m are uniform.
Moreover, a high resolution SEM image (Figure 4d) shows that an individual hexagonal plate
is composed of hundreds of primary hexagonal small plates. Thin primary platelet-like
crystals are aligned to a multilayered stack, and build up the complete mesocrystal. Although
on the nanometer scale the superstructure looks random, the whole crystal is a quite regular
hexagon with sharp facets and edges. It has to be noted that the results clearly indicate that
kinetic rather than thermodynamic factors control the formation of the vaterite phase. The
interactions between anionic moieties in the polymer and calcium cations in solution or at
mineral surfaces are believed to be responsible for initiating and stabilizing non-equilibrium
crystal polymorphs and superstructures.
The aragonite phase was selectively produced when crystallization was carried out at the
lower polymer and Ca2+ concentrations. The morphology and structure of as-synthesized
sample produced at a polymer concentration of 0.1 g L 1 and 1.25 mM CaCl2 (reaction time:
6 days) is shown in Figure 4e, displaying “sheaf bundle” crystals as the predominant
morphology. This structure was also observed in a previous study using surface adsorbed
triblock copolymer microgels as additive [32]. In this work, successful realization of
polymorph switching of calcium carbonate could prove useful for a deeper understanding of
the related biomineralization processes of CaCO3. In addition, polymorph switching in
general is of great technical importance.
Different kinds of polymers can be employed for mineral crystallization. The presence of
polymers clearly improves the quality of oriented assembly, as the nanoparticle surfaces are
obviously ‗code‘ by the selective polymer adsorption for subsequent oriented attachment. An
impressive example is the formation of BaSO4 or BaCrO4 fibers of about 30 nm in diameter,
which are, however, defect free up to hundreds of micrometers in length. These primary
fibers further assemble to hierarchical fiber bundles and cones (Figure 5).
The sodium salt of polyacrylic acid serves as a very simple structure-directing agent for
the room temperature synthesis of highly ordered cone-like crystals [33] or very long,
extended nanofibers of BaCrO4 or BaSO4 with hierarchical and repetitive growth patterns
[34]. Temperature and concentration variation allow the control the finer details of the
architecture, namely length, axial ratio, opening angle, and mutual packing [35]. The
observed [210] growth axis implies that the polyanion adsorbs onto all parallel faces to this
axis on the nucleated nanoparticles, just leaving the negatively charged (210) faces free for
direct interaction. This makes them the highest energy faces, which fuse together by oriented
attachment to form the fibers.
Figure 5. Complex forms of BaSO4 bundles and superstructures produced in the presence of 0.11 mM sodium
polyacrylate (Mn = 5100), at room temperature, [BaSO4] = 2 mM, pH = 5.3, 4 days. (a) highly ordered
funnel-like superstructure with multiple cones aligned orderly; (b) zoomed image of the detailed
superstructures with repetitive patterns; (c) zoomed image of the aligned bundles; (d) magnified surface
structures of the nanofiber bundles. Reprinted with permission from Nano Lett., 2003, 3, 379. Copyright ©
2003, American Chemical Society.
At least three different formation mechanisms of the superstructures shown in Figure 5

can be identified, which are all based on oriented attachment of nanparticles. The chosen
crystallization path was determined by the availability of nanoparticles in solution available
for further oriented attachment and polymer complexes were identified as the earliest species,
subsequently transforming to amorphous nanoparticles, which themselves are the precursors
of the crystalline nanoparticles for oriented attachment. The complexity of this biomimetic
crystallization system is remarkable and shows the significant analytical challenges associated
with an already seemingly simple two components biomimetic mineralization system.
These results make it very clear that the nanoparticles alignment has many similarities to a
controlled ‗polymerization‘ process, where the defined nanoparticles take the role of the
organic monomers. Controlled nucleation or initiation in a short period of time sets the basis
of a process that is terminated by the depletion of a material or external stimuli, such as
electric fields, or curvature or stress fields. In this way, monodisperse aggregates and
superstructures can be created. This similarity between controlled assembly and controlled
polymerization has also been demonstrated by other reports.
Specific biopolymers can exert a strong influence on both crystal nucleation and growth
rates in addition to temperature, pH, ionic strength and composition. Recently, investigation
of biomineralization of inorganic nano- and microstructures and their relevance to biogenic
templates has emerged as an active research field between biomimetics and nanotechnology.
Biopolymers, existing in living organisms due to their role in biomineralization, are often a
natural soluble additive choice for morphogenesis of complex superstructures. It has been
found that chiral copolymers of phosphorylated serine and aspartic acid with molar masses
between 15000 – 20000 g/mol were very efficient additives for the generation of helical
calcite superstructures consisting of elongated 70 nm wide, uniform and highly aligned calcite
nanoparticles where the helix turn corresponding to the copolymer enantiomer [36]. The
helical structures formed when a high degree of phosphorylated Ser (75 mol%) and 25 mol%
Asp in the copolymer were adopted in combination with the ten-fold Ca2+ concentration with
respect to the monomer, which is similar to the conditions where a shellfish forms a shell. The
formation mechanism of the chiral crystalline superstructure is still unclear at present and
needs to be investigated in the future.
NON CLASSICAL CRYSTALLIZATION

AND MESOCRYSTALS
Besides the classical crystallization in templates or confined reaction environments,

biomimetic mineralization can also follow non classical particle mediated assembly
pathways. Some evidences were found that crystallization does not necessarily proceed along
the classical crystallization process, which is the attachment of ions/molecules to a primary
particle forming a single crystal. Instead, crystallization can also proceed along particle based
reaction routes [37]. Particle mediated crystallization pathways were identified, which
produces crystals in the process of a so-called mesoscopic transformation including self-
assembly or transformation of metastable or amorphous precursor particles [38]. Such
crystallization pathways especially apply to systems far from equilibrium, for which the
classical thermodynamic considerations are not valid anymore to predict the morphology or
size of the crystals. But even for systems, which were so far considered to crystallize via the
classical pathway, indications were found that nanoparticles are involved in the crystallization
process. This nanoparticle mediated crystallization pathway involving mesoscopic
transformation is called ―N on-Classical Crystallization‖ and involves multiple nucleation
events of nanoparticles, which form a nanoparticle superstructure in contrast to a single
nucleation event to form a single crystal. Non classical crystallization involves self-
organization of pre-formed nanoparticles to an ordered superstructure, which then can fuse to
a single crystal. Overall crystallization pathways are depicted in Figure 6 and discussed below
in more detail under the respective headings.
d)
Amorphous particles
Nucleation
clusters or ?
Liquid droplets
Crystal growth
Temporary Crystal with
Mesoscale complex shape
Primary Stabilization
assembly
nanoparticles
> 3 nm
a) b) c) Mesocrystal
Amplification Mesoscale
assembly
Single crystal Oriented

attachment
Fusion Fusion
Figure 6. Schematic representation of classical and non-classical crystallization process. Pathway a)

represents the classical crystallization pathway where nucleation clusters form and grow until they reach the
size of the critical crystal nucleus growing to a primary nanoparticle, which is amplified to a single crystal
(path a). The primary nanoparticles can also arrange to form an iso-oriented crystal, where the
nanocrystalline building units can crystallographically lock in and fuse to form a single crystal (Oriented
Attachment, path b). If the primary nanoparticles get covered by a polymer or other additive before they
undergo a mesoscale assembly, they can form a mesocrystal (path c). Note: Mesocrystals can even form from
pure nanoparticles. There is also the possibility that amorphous particles are formed, which can transform
before or after their assembly to complicated morphologies (path d). Reproduced with permission from
Wiley-VCH.
One of the so far most investigated synthetic mesocrystals is the hexagonal prismatic seed
crystal of fluoroapatite, formed in a gelatin gel, which further grows to spherical particles via
dumbbell intermediates. In addition, structural defects attributed to a collagen triple helix
strand as well as self-similar nano-subunits nucleated by gelatin were detected, which agree
with the mesocrystal scheme presented in Fig. 6c but for hexagonal building units. Although
the above system is very well investigated, the formation process of the mesocrystal is still
not fully revealed. Precursor nanoparticles were already experimentally found and dipole
fields suggested being responsible for their almost perfect alignment, but the system proves to
be highly complex on several hierarchy levels. The full growth mechanism was so far only
reported for two mesocrystal examples. One of them is a copper oxalate mesocrystal. Here,
nanoparticles were found to arrange almost perfectly to a mesocrystal, which could be
influenced in terms of morphology by hydroxymethylpropylcellulose. The polymer
influences nucleation, nanocrystal growth and aggregation by face selective interaction. Upon
aggregation of the nanocrystals, a mesocrystal is formed as intermediate but is apparently not
stable due to the low repulsive electrostatic and steric forces. Later, the “brick by brick” self-
assembly mechanism could be experimentally revealed in a time dependent study.
The face selective PAA adsorption onto orthorhombic K2SO4 crystals led to the formation
of tilted unit crystals, which were assembled in a diffusion limited condition resulting in
various complex morphologies such as helices or zig-zag assembly of twinned crystals. A
conclusive explanation of the various possibilities of particle growth in an anisotropic
diffusion field was proposed. It is remarkable that the K2SO4-PAA system has six hierarchical
levels from the nm scale to that of several hundreds of microns, which is a typical feature of
biominerals and was so far only rarely reported for a synthetic material [39]. The
superstructure designed at each level was controlled by changing the polymer concentration
and the observed hierarchy was attributed to the interaction between crystals and polymers
and the diffusion-controlled conditions. A similar hierarchical system has recently been
reported for potassium hydrogen phthalate and PAA [40]. Again, plate-like units are
composed of aligned crystalline nanocrystals, therefore, the well facetted plates on the micron
scale can be considered as mesocrystals although the authors of the original paper also
discuss mineral bridges between the subunits as a possibility for the explanation of their
mutual crystallographic alignment. As the concept of particle assembly in diffusion fields
coupled with face selective polymer adsorption was demonstrated for both – inorganic and
organic crystals, it seems to be much more versatile than known so far.
Uniform NH4TiOF3 mesocrystals comprising orientationally ordered primary crystallites
have been recently prepared by a simple, room-temperature surfactant-mediated route [41].
Figure 7a-c show typical SEM images of the obtained particles, which have regular square
morphology. High-magnification cross-sectional SEM image of a particle (Figure 7c) shows
that this particle consists of small nanoparticles. TEM measurements provide more details of
the structure of the as-prepared NH4TiOF3 mesocrystals (Figure 7d-f). The regions of low
contrast between individual crystallites indicate interstices with diameters of about 5–10 nm
exist within these particles (Figure 7e). The interstitial walls are formed by nanocrystals
whose lengths can extend to 25 nm or more, which is consistent with the 26 nm in the
[100]/[010] directions from the XRD results. SAED of an area ca. 250 nm in diameter
(Figure 7d) shows single-crystal diffraction with minor distortions, indicating that the whole
assembly of particles behaves as a single crystal. The distortions come from the mismatch
between boundaries of the small particles, which are typical for a mesocrystal. Importantly,
when the sample was moved in the microscope in diffraction mode, the pattern remained the
same, except for some minor brightness changes, as clearly shown in Figure 7f.
CaCO3 (vaterite) mesocrystals with a hexagonal morphology and uniform size were
successfully prepared in the presence of an N-trimethylammonium derivative of hydroxyethyl
cellulose via aggregation-mediated crystallization using a simple gas-diffusion method
(Figure 8) [42]. Uniform hexagonal particles show sharp facets and edges, and assembled by
the aggregation of spherical nanoparticle subunits (Figure 8b, [polymer]: 1 g L 1, [CaCl2] =
10 mM) or hexagonal nanosheet subunits (Figure 8d, [polymer]: 0.5 g L 1, [CaCl2] = 10 mM),
depending on the adapted polymer concentrations. ED pattern and HRTEM data confirmed
that these hexagonal micrometer-sized plates display 3D highly oriented superstructures. The
selective adsorption of polymer molecules on specific faces of crystals plays a key role in this
mesoscale transformation. This mesoscale transformation involving cooperative
reorganization of coupled inorganic and organic components can be relevant for the model of
matrix-mediated nucleation in biomineralization. An understanding of the 3D-oriented
aggregation will be helpful in controlling the aggregation-driven formation of complex,
higher-order structured materials, and further provide new insights into biomineralization
mechanisms.
a d f
Figure 7. SEM (a-c) and TEM (d-f) images of the obtained uniform hexagonal NH4TiOF3 mesocrystals in the
presence of 23.1 wt% Brij 58 at 35 oC for 20 h. (a) top view of an as-obtained particle, (b) cross-sectional
view of a particle, (c) high-magnification cross-sectional image of a particle. (d-f) TEM, HRTEM and ED
images of an as-prepared particle. Reproduced with permission from Royal Society of Chemistry.
Figure 8. SEM images of the obtained uniform hexagonal plates of vaterite mesocrystals grown in the
presence of polymer for 1 day, a full view of vaterite CaCO3 particles (a), and high resolution SEM image (b)
showing each hexagonal plate consists of nanoparticles. [polymer]: 1 g L 1. [CaCl2] = 10 mM. (c) SEM
image and (d) high resolution SEM image of vaterite mesocrystals under [polymer]: 0.5 g L 1. [CaCl2] = 10
mM. The edges of primary hexagonal discs are perfectly parallel to each other, indicating each large particle
is a single crystalline aggregate with a preferred c-axis orientation. Reproduced from Wiley-VCH with
permission.
A typical example for a non-classical crystallization pathway of mesocrystal formation

and morphology evolution of calcite CaCO3 crystals in the presence of a polystyrenesulfonate
(PSS) was also demonstrated [43]. Variation of the concentration of calcium chloride and
PSS solutions by a CO2 gas diffusion technique can result in the formation of unusual CaCO3
superstructures, which transformed from the typical calcite rhombohedra, to rounded edges,
to truncated triangles, and finally to concavely bended lens-like superstructures (Figure 9).
The strong binding effect of PSS to free calcium ions will shift the mechanism from
traditional ionic growth to mesoscale assembly. In addition, PSS can also bind selectively to
the otherwise non exposed (001) calcite face, resulting in mesocrystals composed of truncated
triangular units instead of the typical rhombohedra. Usually, calcite is not able to expose the
(001) faces because these are composed of only CO32 or Ca2+ ions in a hexagonal
orientation, respectively. The fact that this face becomes dominant can be ascribed to multiple
Coulomb binding of the negatively charged polymer molecules to the positively charged
[001] plane, leading to surface stabilization and inhibition of growth along this direction. The
oriented self-assembly of subunits toward larger, single-crystalline superstructures is an
example for mesocrystal formation, which is currently identified to be relevant in a wide
range of crystallization processes. Those crystals are porous and composed of almost
perfectly 3D-aligned calcite nanocrystals. The results suggest that the inner field effects
within the nanocrystal building units cannot be neglected, which play a key role in the
polymer-controlled crystallization processes.
Figure 9. SEM images of calcite mesocrystals obtained on glass slips by gas diffusion reaction after 1 day
with different concentrations of Ca2+ and PSS. (a) [CaCl2] = 1.25 mM, [PSS] = 0.1 g L 1. (b) [CaCl2] = 1.25
mM, [PSS] = 1.0 g L 1. (c) [CaCl2] = 5 mM, [PSS] = 0.1 g L 1. (d) [CaCl2] = 5 mM, [PSS] = 1.0 g L 1.
Copyright © 2005, American Chemical Society.
On the other hand, self-similar growth of well-defined polyhedral building units can also
lead to mesocrystals; this is a growth process, which relies on the shape of the primary
building units and some elementary assembly rules. For instance, the construction of a
mesocrystal can be provided by geometric packing of the building units, which are aligned by
edge-to-edge or face-to-face connections. The driving force for the geometrical orientation is
the minimization of the interfacial mismatch energy, through forming a coherent interface and
reducing the exposed surface area. Self-similar growth was found for the spontaneous self-
assembly of ca. 5 m sized octahedral silica polyhedrons [44] on glass substrates. These
crystals are formed by the {111} faces of the cubic phase and can self-assemble in an oriented
manner due to the capability of edge sharing (Figure 10). As the primary building blocks are
well-defined in size, as well as the contact edges to the next particles, a self-similar growth
process is induced, leading to a mutual crystallographic orientation of the primary particles
and thus to the formation of a mesocrystal. A large variety of arrangements are possible, as
the initially formed mesocrystal units can themselves assemble into higher-order structures
(Figure 10e, f), because of the octahedral structure of the primary building units, these
mesocrystals have porosity due to the structural voids. Larger pores are formed by packing
defects and larger cage formation (Figure 10b). The above principle of octahedral edge
sharing is learnt from some crystals built by atoms [45] and also by nanoparticles [46], it
indicates that the same building principles can be applied regardless of the size of the building
units, which is the basis for self-similarity even within one structure.
Figure 10. Schematic illustrations of edge-sharing stacking: a) primary octahedral units, face-on
configurations, b) quartet-octahedron model for the secondary structure, c) tertiary structure with filled
corners, d) tertiary structure with unfilled corners, e) a high-order structure from primary octahedra, f) a high-
order structure from tertiary units. Reproduced with permission from Wiley-VCH.
Besides the self-similar assembly strategy for mesocrystals, the crystallographic

orientation of the nanoparticles building up the mesocrystal can also be achieved by spatial
constraints in an already existing mesocrystal. However, the distinction between spatial
constraints and self-similar assembly is not easy to make on the basis of the final
mesocrystals, and only a time resolved investigation is able to reveal how the mesocrystals
are formed.
A very recent example of the self-similar assembly of obtained calcite mesocrystals is
shown in Figure 11 [47]. In this case, calcite nanoparticles with a characteristic shape aligned
to a mesocrystal with a shape resembling that of the primary nanoparticles, with triangular tip
structures.
The sample consists of a large number of elongated calcite particles with well defined
faces and edges (Figure 11a, b, e). Elongation direction of each microparticle is along the c-
axis, as marked in Figure 11e. The high-magnification SEM images show that the
microparticles appear to be self-similar mesoporous superstructures themselves (Figure 11d),
formed by aggregation of the primary nanoparticles. The size of those nanoparticles
determined from SEM images is ca 23-30 nm, in agreement with the XRD measurement.
There are totally eight faces for each microparticle. Two basal faces of each elongated
microparticle have a three-fold symmetry (the equilateral triangular crystal planes (Figure 11b
& c) have three angles of 60o, that is, the crystallographic planes of each end surface belong
to the {001} family), corresponding to the (001) and (00-1) face [48].
g)
10-1
011
001 1-11 -11-1 00-1
0-1-1
-101
Figure 11. SEM images of the obtained trigonal calcite mesocrystals with triangular capped building blocks
in the presence of PS-MA. (a) overall product morphology; (b), (c) and (d) High magnification SEM images
showing the basal faces of elongated microparticles; (e) and (f) SEM images showing the lateral faces along c
axis; (g) a modelled calcite morphology of a combination of {001} and {011} forms constructed by the
Cerius2 software. Grey lines and face indices are those at the back. Polymer: 0.1 g L 1, [CaCl2] = 1.25 mM.
Inset in (c) is Sierpinski triangles. (Image reproduced from [47] with permission of Wiley-VCH.)
a b c d
30 nm 100 nm 300 - 600 nm 40 m
Figure 12. Proposed formation mechanism of hierarchical self-similar calcite mesocrystals made of triangular
calcite subunits. (a) nearly spherical CaCO3 nanoparticles formed in the initial reaction stage. (b)
crystallization of calcite nanoparticles exposing {011} & {001} faces and their aggregation. (c) further
aggregation of the nanoparticles into an aggregate with the shape of its subunits presumably along the {011}
faces; (d) formation of large calcite 3D mesocrystals consisting of triangular calcite building blocks via
mesoscale transformation. (Image reproduced from [45] with permission of Wiley-VCH.)
By comparing the measured interfacial angles with the theoretical angles of six lateral
faces, it can be concluded that these six lateral faces are (-101) (10-1), (1-1) (-11-1) and (011)
(0-1-1) (Figure 1e, g). For example, the isosceles triangular crystal planes (Figure 11e) have
angles of 76 1o and 28 1o, that is, the crystallographic planes of this surface and the
opposite surface parallel to this surface are ascribed to the {011} family (Figure 11e, g). All
eight faces are clearly displayed in Figure 11g, which is a modelled calcite morphology
constructed by the Cerius2 software (Accelrys). In fact, the modelling morphology (Figure
11g) for a set of {001} and {011} faces is quite similar to the calcite morphology grown
(Figure 11a, e).
The side faces from the {011} family are neutral faces, which align according to their
surface ion pattern. The oriented self-assembly of subunits toward larger, single crystalline
superstructures (Figure 11) can be seen in the framework of self-similar growth of a
mesocrystal from particulate subunits. This observation indicates that an overall
crystallographic relationship exists among the nanocrystallites that constitute the whole three-
dimensional mesocrystals. The SEM images depict the formation of a hierarchical structure.
This structural hierarchy indicates that both the primary units and the pre-assemlbed
intermediates can undergo further oriented attachment, with the larger structures also being
able to support larger pores while packing towards the superstructure. The scattering behavior
is of a single crystal because the triangle-capped subunits are arranged with the same
crystallographic orientation (three edges of each triangle are parallel to those of other
triangles). This is schematically visualized in Figure 12.
The above example gives clear evidence that mesocrystals can self assemble even from
complex shaped subunits via edge or face energy minimization.
More recently, crystallization of calcium carbonate in 5 mM CaCl2 solution with PSS-MS
(PSS-co-poly(maleic acid)) copolymer as additive at a concentration of 0.1 0.25 g L 1 led to
the formation of octahedral calcite mesocrystals, as shown in Figure 13 [49]. Figure 13a
presents a typical scanning electron microscopy (SEM) image of the obtained product, clearly
displaying octahedral morphology, which is unexpected for calcite crystals grown in a dilute
solution. These Platonic octahedral particles are uniform, with well defined faces and edges,
and a size of several tens of micrometers. High resolution SEM images (Figure 13c, d) show
that each octahedron consists of a large number of smaller subunits, thus displaying
mesocrystal features. Comparable observations were reported for fluoroapatite systems of
self-similarity [31] and truncated triangular calcite mesocrystals [29], indicating a mesoscale
transformation process occurs for these octahedral calcite aggregates.
Figure 13e shows an intermediate of dodecahedra derived from time-dependent
experiments performed with different Ca2+ and polymer concentrations. Strikingly, all SEM
images of these intermediates suggest a rhombohedral P-surface morphology [50] made up by
nanoparticles. Figure 13f shows a unit cell of P-surface. Typically, minimal surfaces observed
on the micro- and macroscale occur only for liquid phases with surface tension, such as lipid-
water or surfactant-water mixtures as well as copolymers and liquid crystalline mesophases of
amphiphiles in water, while on the scale of atoms to few crystalline unit cells, minimal
surfaces were also reported for zeolites and equipotential lines in crystal grids [51].
Looking more carefully onto those structures, it is found that there are six smooth and six
coarsened facets per dodecahedron. We ascribe the former to the {104} family, thus being the
usual exposed faces of calcite, whereas the latter do not look like real faces at all, as they
seem to have curvature and blips with a typical nanogranular surface structure. A non-
classical crystallization mechanism is employed for the morphogenesis with elements of
liquid and solid behavior resulting in the first observation of a minimal rhombohedral
primitive surface in a synthetic crystallization reaction.
a b e
c d f
Figure 13. SEM images of octahedral calcite mesocrystals. (a-d): [PSS-co-MA] = 0.1 0.25 g L 1, [CaCl2] =
5 mM, 2 weeks; (e): [PSS-co-MA] = 0.0025 g L 1, [CaCl2] = 1.25 mM, 3 days; (f) a unit cell of P-surface
with minimal surfaces. (Image reproduced from [47] with permission of Wiley-VCH.)
BIO-INSPIRED FUNCTIONAL
NANOMATERIALS AND ASSEMBLY
Biomaterials naturally have highly organized structures at the nanoscale with a variety of
functions. Various bioassemblies are shown to template complex, multidimensional inorganic
architectures that are typically not available by our synthetic methods. In addition to the
various naturally occurring templates, the powerful techniques developed by life sciences are
an interesting tool for engineering strategies for materials science [52]. Besides a structural
and morphological control during synthesis, biotemplating approaches may add another
dimension to inorganic materials such as biofunctionality. This section is focused on recent
advances in the preparation of inorganic materials through use of biomolecule assemblies. We
try to elucidate chemical methodologies and presents examples of templates based on protein,
lipid and peptide building blocks that were successfully exploited to synthesize inorganic
nanostructures.
In relation to the surfactant-templated growth of nanostructured materials, the recent use
of microorganisms to control inorganic crystal formation has been promoted as genetically
engineered polypeptides binding to selected inorganics (GEPIs), such as Au [53] and silica
[54]. GEPIs are based on three fundamental principles: molecular recognition, self-assembly
and DNA manipulation, and they promise numerous successes in bio-inspired strategies.
Although we have not clearly known about the mechanisms of in vivo crystallization
processes at a molecular level, proteins play a key role in the formation of inorganic
materials. However, this in vivo synthesis is just limited to certain materials such as calcium
carbonate, silica, or magnetite. Therefore, the knowledge of biological concepts, functions,
and design characteristics has been implemented into approaches for the synthesis of new
technologically important materials which have no isomorphous complement in nature.
Proteins have many specific recognition capabilities to drive assembly into defined
superstructures but are less programmable than the DNA templates. They are, however, based
on a very broad platform of chemical diversity. Recent progresses in combinatorial biology
allow the identification of amino acid sequences with specific affinity for inorganic crystals,
ranging from metal oxides and semiconductors to metals, providing a route to create novel
interfaces between biomolecules and inorganic crystals. Besides specific recognition
capabilities (e.g., antibody–antigen, biotin–avidin), proteins display various functionalities
such as catalytic (e.g., enzymes) and motility functions (e.g., motor proteins) for potential
biofunctional inorganic materials. What‘s more, it has been confirmed that nanoparticles may
also influence the structure and function of the conjugated biological structures [52]. Some
common used biotemplates are schematically summarized in Figure 14 [52].
Biotemplates with well-defined chemical and structural heterogeneity have recently been
exploited for the precise control of the size and shape of the formed nanostructures including
metal nanostructures of Ag [58], Ni, and Co nanowires [59], Au [60], complex two- or three-
dimensional assemblies of Au nanoparticles [61] and semiconductor CdS, PbS, ZnS
nanostructures [62, 63] A DNA membrane complex as a simplified prototype system can be
used as a nanoreactor to template the growth of CdS nanorods [64]. The strong electrostatic
interactions within such complexes align the CdS (002) polar planes parallel to the negatively
charged sugar-phosphate DNA backbone, which suggests that molecular details of the DNA
molecule have been replicated onto the inorganic crystal structure.
Sastry and co-workers reported a reaction of the extract of the lemongrass plant with
aqueous chloroaurate ions, which can produce high yields of thin, flat, single-crystalline gold
nanotriangles in a one-step reaction at room temperature [65]. By using a similar method,
pure metallic silver and gold nanoparticles and bimetallic Au/Ag nanoparticles can also be
obtained through reaction between metallic ions with the extract of Azadirachta indica leaf
[66].
Processes based on biotemplates take advantage of the characteristic nanoscale
dimensions of the biological specimen and replicate their morphology using inorganic
components. Biomolecular components exhibit dimensions from lower nano- to micro-sized
ranges with interesting surface features and functionalities. The replication process generates
a positive (hollow), negative or exact copy of the template.
Figure 14. Schematic illustration showing the structure and dimensions of several protein assemblies used as
biotemplates for materials synthesis. Reproduced from ref. [55] with permission of the American Chemical
Society, from ref. [56] with permission of Wiley-Blackwell, and ref. [57] with permission of Wiley-VCH.
Figure 15. Casting of silver nanowires using peptide nanotubes. (A) The nanowires are formed by the
reduction of silver ions within the tubes, followed by enzymatic degradation of the peptide mold. (B) TEM
analysis (without staining) of peptide tubes filled with silver nanowires. (C, D) TEM images of silver
nanowires that were obtained after proteolyticlysis of the peptide mold. Reprinted with permission from ref
[67]. Copyright 2003 American Association for the Advancement of Science.
A very short peptide, the Alzheimer‘s -amyloid diphenylalanine structural motif can be
self assembled into discrete and stiff nanotubes. The peptide nanotubes were used to serve as
molds for casting metal Ag nanowires (Figure 15a) [67]. The tubes were added to boiling
ionic silver solution, and the silver was reduced with citric acid to ensure a more uniform
assembly of the silver nanowires. TEM measurements indicate the formation of silver
assemblies within the majority of the tubes (Figure 15b). Proteolytic lysis of the peptide
mold, by the addition of a proteinase K enzyme to the silver-filled nanotubes solution,
resulted in the attainment of individual silver nanowires 20 nm in diameter, as seen by TEM
(Figure 15c, d). The diameter of the nanowires is smaller than that of the tubes, which further
suggests that casting was done inside the tubular structure.
Bio-inspired strategies that use viruses and genetically engineered bacteriophages have
been employed to prepare nanometer-sized structures. Some groups have used combinatorial
cell surface display or phage display methods to identify peptides that bind strongly to, and in
some cases induce the precipitation of, synthetic inorganic materials [68]. Ahmad et al.
recently demonstrate that peptides (BT1 and BT2 peptides) identified by phage display
biopanning are capable of inducing the rapid, room-temperature formation of tetragonal
barium metatitanate, ferroelectric BaTiO3, from an aqueous precursor solution at near neutral
pH [69]. Obviously, this bio-inspired synthetic route shows more advantages over
conventional synthetic methods.
Key to the successful application of nanotechnology on an industrial scale, however, is
the ability to manipulate these nano-objects into a spatially ordered pattern. In recent years,
interest in using biomolecules, such as crystalline S-layer proteins and ferritin protein cages,
as templates to scaffold inorganic nanostructures has arisen [52]. Of the various
biomolecules, phospholipids, owing to the strong amphiphilicity stemming from possessing
both polar heads and aliphatic tails, are able to spontaneously arrange themselves along a
phase boundary or external surface. This property permits the generation of a multitude of
microscopic structures, that is, micelle, vesicle, bilayer, microtubules, and nanotubes, and
would also render the phospholipid an ideal material to drive the assembly of metal
nanoparticles. Lipid nanotubes and solid-supported lipid multilayers were used to induce the
self-assembly of the metal particles [70].
More recently, Yoon et al. have reported a novel method of using a solid-supported liquid
crystalline lipid membrane as a template to synthesize nanometer-sized particles as well as to
force the encapsulation of the resulting particles with lipid molecules (Figure 16) [71]. In the
multilayer formed by lipids in the liquid crystalline state, the lipid membrane becomes
flexible as a result of chain melting. Capitalizing on the flexibility of the liquid-crystalline
lipid membrane, when metal film was deposited onto the liquid-crystalline lipid multilayer,
the flexible lipid membrane allows penetration of the metal and subsequent formation of
metal nuclei within the multilayer membrane. Subsequent spontaneous encapsulation of metal
particles by the lipid molecules would limit the further growth of metal particles beyond an
equilibrium size by confining the volume and shape of the metal particles. The unique aspect
of our approach lies in the fact that the encapsulating lipid molecules are highly mobile and
thus able to induce redistribution of metal to produce monodisperse nanoparticles. The
mobility of the lipid molecules also expedites the formation of highly ordered nanoparticle
superlattices (Figure 16). Dissolving the Ag-embedded lipid membrane in a polar solvent,
chloroform, also resulted in the formation of ordered superlattices, but with increased density
of mis-registered Ag particles.
(a)
Figure 16. (a) TEM images of Ag-embedded membrane of lipid DOPC. (b) TEM image of 2D self-assembled
Ag nanoparticles formed after dissolving in iso-octane. (c) TEM image of 3D self-assembled Ag
nanoparticles formed after dissolving in iso-octane. d) Magnified image of (c). (e) Low- and (f) high-
magnification TEM images of the ‗‗honeycomb‘‘ structure formed by dissolving the Ag-embedded
membrane in chloroform. (Image reproduced from [71] with permission from Wiley-VCH.)
Figure 17. Spontaneous formation of protein–nanoparticle chains by binding of Ni-NTA functionalized Au

nanoparticles to a histidine tag modified, stress-related protein oligomer (6His-SP1). Reproduced from ref.
[72] with permission of the American Chemical Society.
Besides directly depositing the inorganic material on the biological specimen,

biomolecules have been applied to assemble preformed inorganic building blocks into
superstructures after binding to biomolecules with specific recognition capabilities. Medalsy
et al. have reported the formation of ordered arrays and Au nanoparticle/protein hybrid
superstructures using a stress-related stable protein 1 (SP1) [72]. SP1 is a boiling-stable ring
protein complex, isolated from aspen plants (populus tremula) expressed during drought, 11
nm in diameter, which self-assembles from 12 identical monomers. SP1 can be utilized to
form large ordered arrays; it can be easily modified by genetic engineering to produce various
mutants; it is also capable of binding gold nanoparticles (GNPs) and thus forming protein-
GNP chains made of alternating SP1s and GNPs. The protein is extremely stable towards
high temperature and detergents, e.g., it reveals a melting temperature of 107 oC and
resistance to detergents such as sodium dodecyl sulfate (SDS). Large, two dimensionally
ordered arrays with a periodicity of 11 nm were assembled by applying the native SP1
oligomer and a phospholipids mixture. The N-termini of the SP1 could be modified by six
histidine tags and the mutant could be expressed in E. coli. Histidine tags strongly bind to Ni
ions and provide anchor points for Ni–NTA conjugates. 1.8 nm sized Au nanoparticles
covered by Ni–NTA ligands could be bound to the inner pore of the histidine tag modified
variant. Upon incubation of the Au nanoparticle/6His-SP1 hybrids in buffer solution protein–
nanoparticle chains are spontaneously formed (Figure 17). Such particle 6His-SP1 hybrids
may serve as potential building blocks for various other nanostructures, e.g., two-dimensional
arrays.
DNA possesses remarkable molecular recognition properties and structural features,
which make it one of the most promising templates to create patterned materials with
nanoscale precision. The emerging field of DNA nanotechnology strips this molecule from
any preconceived biological role and employs its simple code to produce addressable
nanostructures in one, two, and three dimensions. These structures have been used to
precisely position proteins, nanoparticles, transition metals, and other functional components
into deliberately designed patterns [73]. A strategy based on DNA hybridization to link
colloids has been demonstrated [74] and was successfully applied in biological sensing [75].
The employment of DNA for materials synthesis and the use of genetically engineered
proteins and organisms for inorganic growth and self-assembly opens up new avenues for the
design of original nanostructures. During the past few years, two-dimensional (2D) arrays
with amazing regularity have been produced using DNA [76]. However, the assembly of 3D
dimensional structures has proven difficult. More recently, two groups have independently
reported on the DNA-mediated assembly of nanoparticle crystals [71, 72].
Gang et al. have reported the formation of three-dimensional crystalline assemblies of
gold nanoparticles mediated by interactions between complementary DNA molecules
attached to the nanoparticles’ surface (Figure 18) [77]. It is found that the nanoparticle
crystals form reversibly during heating and cooling cycles. Moreover, the body-centered-
cubic (bcc) lattice structure is temperature-tunable and structurally open, with particles
occupying only 4% of the unit cell volume. It can be expected that this DNA guided
crystallization strategy, and the insight into DNA design requirements it has provided, will
facilitate both the creation of new classes of ordered multicomponent metamaterials and the
exploration of the phase behavior of hybrid systems with addressable interactions. Similar
study on DNA-mediated 3D Au nanoparticle assembly was also reported by Mirkin et al.
[78].
Figure 18. TEM and SEM images of Au nanoparticles before and after assembly at room temperature. SAXS
patterns and corresponding structure factors S(q) for as-assembled systems at room temperature (blue curve)
and annealed at Tpm (yellow curve). An illustration of the b.c.c lattice is shown, where the proposed CsCl-
type particle arrangement is coded with blue and red colors representing particles with complementary DNA
cappings. Reproduced with permission from Nature Publishing Group.
CONCLUSION
Since organisms have spent millions of years optimizing structural biomaterials for
performance, durability, and appearance, it is reasonable that scientists are highly interested
in designing functional materials and are developing a curiosity about how Nature has solved
problems that are often encountered in materials science and technology. Bio-
inspired/biomimetic chemistry has now become a separate branch of materials chemistry,
wherein lessons learnt from biological systems are implemented into in vitro syntheses. The
current chapter shows that a much progress has been made in recent years to transfer
biomineralization principles to synthetic materials chemistry. This is triggered by an
increased understanding of biomineralization mechanisms as well as an increased
understanding of self-assembly and bottom up materials synthesis approaches. Bio-inspired
mineralization can now be used for the room temperature synthesis of a large variety of
inorganic minerals, metals and organic crystals, producing materials with new and exciting
properties. Biomimetic materials and systems such as adaptive materials, nanomaterials,
hierarchically structured metamaterials, mesocrystals, materials compatible with ecological
requirements, should become a major preoccupation in advanced technologies. It can be
expected that this research field will still gain further relevance in the next years because of
the importance of hierarchically structured composite materials, one dimensional materials,
morphosynthesis approaches, polymorph and size control of crystals and self-organization in
materials synthesis. Nature teaches excellent lessons how to achieve these elusive goals and it
can be expected that many new and exciting materials can be produced in the future by
mimicking biomineralization processes. Bio-inspired selective multifunctional materials with
associated properties such as separation, adsorption, catalysis, sensing, biosensing, imaging,
multitherapy, will appear in the near coming years.
The current overview reveals that much progress has been made in recent years to transfer
biomineralization principles into synthetic materials chemistry. Despite the efforts made this
past decade to elaborate bio-inspired materials, characterize their structural and physico-
chemical properties, understand their structure–function relationships and particle mediated
assembly of mesocrystals, many unknown mechanisms still need to be investigated in the
future study. Prospects for the future include the development of novel transcription
methodologies, as well as the creation of new transcriptive templates which present a greater
degree of structural complexity or can give rise to extended, complex architectures. One of
the principal challenges remaining consists of finding novel methodologies for obtaining
transcribed structures with a high degree of order at the multiscale level.
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Lecture material 15
NANO-FABRICATED STRUCTURES FOR

BIOMOLECULE ANALYSIS
ABSTRACT
In this chapter, the recent development of biomolecule analysis, especially
biomolecule separation using nano-fabricated structures was reviewed. Fundamental
fabrication techniques for micro- and nano-structures on silicon or glass substrates,
various approaches for biomolecule separation based on different separation mechanisms,
and typical applications such as DNA separation will be included, and practical
applications such as DNA separation are described from the aspect of ―n anomaterials and
nanotechnology for bio-analytical chemistry‖.
INTRODUCTION
Since the concepts of uTAS was demonstrated in 1990 by Manz [1], micro- and
nanofabrication technique have been extensively developed in the semiconductor and MEMS
industries, and now, nanofabricated structures are available for many researchers as a novel
analytical tool in uTAS. uTAS contains several elements for the acquisition, pretreatment,
separation, post-treatment, and detection of samples. In this multi elements system,
microfluidics plays a central role to the development of uTAS because these elements must be
able to handle liquid or gas samples and be miniaturized and incorporated onto a single chip.
To construct the system, various types of components such as valves, pumps, mixers, filters,
and interconnects, are still required to transport, mix, and separate samples. Nanofabricated
structures have potentials to inspire the current microfluidics and provide extra functions on
the components, and explore understanding of a new field, nanofluidics.
The early generations of uTAS performed the functions of large analytical devices in
small, often disposable, units. The potential benefits of uTAS include reduced consumption of
samples and reagents, shorter analysis times, greater sensitivity, portability that allows in situ
and real-time analysis, and disposability. As a consequence of these potential benefits, there
has been considerable interest in the development of uTAS. In the current nanostructures-
equipped uTAS, many researches focus on fundamental understanding of nanofluidics as well
as their practical applications. A typical example of practical applications of nanostructures-
equipped uTAS is periodical nanoslits for DNA separation. The periodical nanoslits consist of
periodical deep and shallow channel to fractionate large DNA molecules which have larger
gyration radius than the shallow channel. This entropic barrier generates different duration
time for large and small DNA molecules and lead to successful separation. This periodical
nanoslits offered a new type of sieving matrix for DNA molecules, in which large DNA
molecules migrated faster than small DNA molecules under electrophoresis. This type of
separation could not be achieved in the past gel or polymer based separations.
In this chapter, the recent development of biomolecule analysis especially biomolecule
separation using nano-fabricated structures will be reviewed. Fundamental fabrication
techniques for micro- and nanostructures on silicon and glass substrates, various approaches
for biomolecule separation based on different separation mechanisms, and practical
applications such as DNA separation are described from the aspect of ―na nomaterials and
nanotechnology for bio-analytical chemistry‖.
FABRICATION PROCESSES OF MICRO- AND NANOSTRUCTURES FOR

BIOMOLECULE ANALYSIS
In this chapter, typical fabrication processes for micro- and nanofluidic devices are
described according to the substrate materials.
Silicon Fabrication
Since silicon is an indispensable component for the semiconductor and MEMS industries,
extensive efforts to develop fabrication techniques have been made. These well-developed
techniques could directly apply to fabrication of micro- and nanofluidic devices for chemical
and biological applications. Two typical fabrication processes, standard photolithographic and
sacrificial layer process, are introduced in this section.
Microfluidic channels and micron-sized structure could be fabricated on a silicon wafer
using standard photolithography and reactive ion etching (RIE) technique as shown in Figure
1. First, photoresist is spin coated on a silicon wafer with a thickness of several tens of
micrometers. The required patterns are transferred by UV irradiation through pre-patterned
photomask. After the post exposure bake is provided if required, the exposed patterns are
developed and hard bake is performed to harden the cross-linking structures in the remaining
photoresist. The patterned resist was used as an etch mask in the successive process, RIE,
which is most widely used as an anisotropic dry etching process. Through-wafer fluid access
holes to the microfluidic channels are then fabricated in precise locations by laser
micromachining, sandblaster, and potassium hydroxide (KOH) etching. After this process, in
order to provide an electrical insulation layer, the wafers are oxidized by a thermal oxidation
process to grow a silicon dioxide layer approximately 100 to 500 nm. This insulation layer is
absolutely necessary especially for electrophoresis experiments. The wafers are then bonded
to a transparent glass slide to ensure optical observation by anodic bonding. Some research
groups use an adhesive silicone, polydimethylsiloxane (PDMS), which was coated on a glass
coverslip about 25- [2, 3]. Polypropylene and PDMS tubes
are glued above the access holes for liquid operations.
Figure 1. Typical fabrication process of silicon microfluidic channel by photolithography and reactive ion
etching. Oxidation process is for growing insulation layer for the following electrophoresis experiments.
Pyrex glass is widely used to seal the microfluidic channel ensuring optical path. (Reproduced from Cabodi,
M. et al., Electrophoresis 2002, 23, 3496. With permission)
The other fabrication method is the use of sacrificial layer process which could integrate
the floor and ceiling of the microchannels and nano- and micron-sized structures in a single
monolithic design [4, 5]. This fabrication method has advantages over earlier methods, in
which the ceiling of the microchannels was created by sealing a glass coverslip to opened
microchannel fabricated by the above mentioned standard photolithographic technique.
Superior uniformity and precise control of the microchannel height is achieved because this
dimension is determined by the thickness of the sacrificial layer. The method is also more
tolerant of particulate contamination because the structure is disrupted only in the immediate
vicinity of a particle. Since the ceiling layer consists of silicon nitride, optical observations
are available through the transparent and the negligible autofluorescent ceiling layer. The
process steps are outlined schematically in Figure 2. The silicon wafer is thermally oxidized
and the layer of silicon dioxide is grown up to 1.0 m. After the oxidation, silicon nitride is
deposited by low-pressure chemical vapor deposition (LPCVD) and form 190 nm of silicon
nitride layer. Polysilicon is subsequently deposited at 500 nm and a 100 nm hard-mask layer
is thermally grown in the polysilicon layer. Over this oxide hard-mask layer, 40 nm of
aluminum was thermally evaporated to assist in the following pattern transfer and provide a
conductive substrate for electron beam lithography (EBL). Polymethylmethacrylate (PMMA)
is spin coated about 200 nm over the aluminum as an electron beam resist. EBL is carried out
to pattern nano-scaled features with an optimal electron dose. The patterned electron beam
resist is developed and transferred to the lower aluminum layer by RIE with chlorine (Cl2),
boron trichloride (BCl3), and methane (CH4) gas. The patterned aluminum layer is then used
as a mask to etch the next SiO2 hard-mask layer by RIE using CF4 gas. As a final etching
step, the pattern of the SiO2 hard-mask layer is transferred to the polysilicon sacrificial layer
with a three-step RIE with Cl2, BCl3, and H2 gas. At this point, patterning of the sacrificial
layer is completed and the required nano- and micron-scale structures such as nanopillar array
and microchannel structures are fully constructed. The open space in the sacrificial layer will
become isolated nano- and micron-scale structures in the gap between the top ceiling and
floor layer after the top layer deposition and the sacrificial layer removal. Silicon nitride was
then deposited by low-stress LPCVD by 320 nm thick and the buried silicon nitride in the
sacrificial layer will later form nanopillar array structures inside a microchannel. To remove
the sacrificial layer, fluid access holes are fabricated by a photolithography and a CF4 RIE.
The sacrificial layer was removed by dipping the wafer in a 5% tetramethylammonium
hydroxide (TMAH) solution over 30 min. Very low temperature oxide (VLTO) silicon
dioxide layer is grown about 2.5 m over the silicon nitride top layer to reseal the fluid access
holes for the sacrificial layer removal. The other fluid access holes for the subsequent
separation experiments were fabricated by a photolithography and a CHF3 RIE.
Quarts Fabrication
Quartz is one of the most suitable materials for biomolecule analysis because it has
excellent optical properties and chemical stability especially at low pH range. Standard
silicon process has been basically applied to quartz fabrication so far. However, due to the
slow etching rate in dry etching process, some modification methods are required especially
for etching and bonding process. Three different fabrication processes, standard
photolithography, nanoimprint, and focused ion beam (FIB) milling are introduced in this
section.
Figure 2. Fabrication process of a sealed monolithic obstacle array using sacrificial layer. (Reproduced from
Chou, C. et al., Proc. Natl. Acad. Sci. USA 1996, 96, 13762. With permission)
When the required structures are over approximately 1.0 m, standard photolithography
and RIE are applicable to quartz substrate as well as silicon. For example, an array of
micropillars oriented in a hexagonal lattice whose width was 2 m and height was 2 m was
fabricated using standard photolithography and RIE [6, 7]. The micropillar arrays were sealed
using glass cover slips with a spin coated thin-layer silicone elastomer. To make the silicone
elastomer hydrophilic and adhesive to the quartz, the silicone elastomer surface was treated
for 1 min in an oxygen plasma. This sealing method through silicone elastomer could be
applied for mechanically strong micron-sized pillars but not for more fragile nano-sized
pillars. Fluid access holes for buffer solutions and DNA samples were drilled mechanically.
When nano-scale and high-aspect-ratio pillar structures are desired to use for separation
experiments, more complicated fabrication processes are required including EBL and deep
RIE. For example, nanopillar structures with high aspect ratio of 20 were achieved by the
following fabrication process [8, 9]; (Figure 3) Thin Cr and Pt layer (~10 nm) were sputtered
on the quartz substrate, and then, positive type EB-resist was spin coated about 1.2 m thick on
the Cr/Pt layer. The nanopillar pattern was delineated by EBL. To produce high aspect ratio
nanopillars, Ni was electroplated into the pattern of nanopillars as a mask for the subsequent
SiO2 dry etching using the Cr/Pt layer as cathode in the appropriate nickel sulfate solution.
For the microchannel patterning, standard photolithography was employed. The nanopillar
and microchannel patterns were etched by deep RIE with a planar-type neutral loop discharge
(NLD) plasma of CF4. The surfaces of a quartz cover plate and the patterned plate were
treated with a mixed solution of ammonia and hydrogen peroxide and left overnight in contact
with each other under pressure at room temperature. After that, they were directly bonded at
1100˚C for 3 hrs without applying pressure.
As well as the nanopillar array structures, the nanochannels that could confine a DNA
molecule offer novel bio-analytical techniques. The nanochannles that were 100 nm in width
with a depth of 200 nm were fabricated on quartz substrate for DNA dynamics study [10] by
using the imprinting technique developed by Chou and coworkers [11]. The high-density
arrays of nanochannels were fabricated using nanoimprint lithography (NIL). The NIL mold
was generated by interferometric lithography (IL) and has 200 nm period gratings over a 100-
mm-diamter wafer. By using this method, millions of enclosed nanochannels with dimensions
smaller than 10 nm have been fabricated. The nanochannels were then bonded with quartz
cover plates by a combination of the surface-cleaning protocol(RCA), room-temperature
bonding, and annealing at 1000˚C.
FIB milling is one of the more recently developed method for a nanochannel fabrication.
Nanochannels were prepared by FIB milling after first coating with a 5-nm Au layer on
quartz substrate. The gold layer was removed by aqua regia, and then, the device was sealed
with a quartz coverslip. After the microchannel fabrication by photolithography and RIE,
reservoirs were affixed over the fluid access holes at the ends of the microchannels [12, 13].
Figure 3. (A) Fabrication process of quartz nanopillar structures. (B-E) Fabricated nanopillar array structures
on a quartz substrate before sealing by a cover slip. The scale bars are all 500 nm. (Reproduced from Kaji, N.
et al., Anal. Chem. 76, 15, 2004. With permission)
Plastics Fabrication
Most microfluidic devices had been fabricated in glass, quartz, or silicon. The
microchannels were fabricated in these substrates using photolithography and various etching
processes, and then, enclosed by flat substrates using anodic or fusion bonding. Although
these materials and fabrication methods have been already developed, there are some
limitations especially for the rapid development and testing of new concepts in microfluidic
devices. The fabrication processes are slow and requires expensive clean-room facilities. The
materials are fragile and too expensive for disposable use. In these materials, silicon is
relatively easy materials for fabrication but optical and electrical properties are not suitable
for certain types of bio-analytical experiments. To compensate these potential defects, plastics
are trying to use as an emerging materials for microfluidic devices. Various plastics such as
polydimethylsiloxane (PDMS), polymethylmethacrylate (PMMA), and cyclic olefin
copolymer (COC), have been applied for microfluidic devices. Here, from the viewpoint of
ease of fabrication and disposability, only PDMS device fabrication is described.
Negative-type thick photoresist (SU-8) was spin coated onto silicon wafers to create
master mold. A microfluidic channel design was printed on a transparent sheet by a high-
resolution printer and used as a mask in the following photolithography step. After
development and post-exposure bake processes, the master molds were placed in a desiccator
under vacuum for 2 hrs with a vial containing a few drops of tridecafluoro-1,1,2,2,-
tetrahydrooctyl-1-trichlorosilane for the surface silanization. This chemical treatment
facilitates the removal of the PDMS replica from the master mold. A 10:1 mixture of PDMS
prepolymer and curing agent (Sylgard 184) was stirred thoroughly and then degassed under
vacuum. The prepolymer mixture was poured onto the master wafer and cured for 1 hr at
65˚C or overnight at room temperature. After curing, the PDMS replica was peeled from the
master wafer. For making fluid access holes on the PDMS replica, glass posts were placed on
the master wafer before pouring the prepolymer mixture or the cured PDMS replica was
punched. To seal the microchannels irreversibly, a PDMS flat plate or a glass plate were
contacted immediately after the brief treatment of plasma oxidization. These processes do not
require clean room operation except the master molds fabrication, rapid and mass productions
are possible. Although there is a size limitation of fabricating structures in this PDMS process
(typically 20 m), transparent and insulating PDMS microchannels are useful for many bio-
analytical applications. Novel nanomaterials, which are described in the following section,
could be easily packed in the PDMS microchannel and applied for separation experiments.
Nanomaterials
Molecular sieving matrices are indispensable for biomolecule separations such as DNA
and protein separations. Generally these sieving matrices show high viscosity and it becomes
increasingly difficult to load the sieving matrices inside microchannels. To resolve the
problem and expand application filed, a new type of nanoparticles called nanoballs was
developed [14]. A core-shell type of globular nanoparticles, which was prepared by the
multimolecular micellization and subsequent core polymerization of block copolymer of
polyethylene glycol(PEG) with polylactic acid(PLA) possessing a methacryloyl group at the
PLA end (PEGm-β-PLAn-MA1; Mw(PEG/PLA) = 6,100/4,000, m ≈ 100, n ≈ 40, l ≈ 70) in
aqueous medium, was developed. The hydrophobic PLA segments form a spherical core,
which is covered by tethered, flexible PEG chains at a fairly high density. The methacryloyl
groups located in the particle core were polymerized to form stable core-shell type
nanospheres having a diameter of 30 nm. The nanospheres have no surface charge, and they
have a narrow size distribution and low viscosity in aqueous media (0.94 cP at 1.0%)
compared with conventional polymers (e.g., methylcellulose: 8.8 cP at 0.5% and 104 cP at
1.0%), owing to their globular structure.
To avoid loading problem of sieving matrices, nanostructures consist of nanomaterials
were constructed inside microchannel based on self-assembled mechanism. Self-assembled
colloidal arrays [15, 16] and molecular sized cavities interconnected by nanopores [17, 18]
have been demonstrated to work as a DNA sieving matrix. In self-assembled colloidal arrays,
the colloidal suspension containing monodispersed plain polystyrene (PS) microspheres and
silica nanospheres was injected into the microchannel reservoirs. The aqueous solution fills
the channels spontaneously, and then, recede of the liquid meniscus accompanying
evaporation induced colloidal growth within the microchannels. Before separation
experiments, the water in the reservoirs was substituted with the running buffer and
equilibrate over 20 min. In molecular sized cavities interconnected by nanopores, a clean
glass slip was immersed in an aqueous suspension of negatively charged polystyrene beads.
The colloid was deposited on the substrate forming a two-dimensional, hexagonally packed
monolayer by slow evaporation under ambient conditions. A 30% acrylamide monomer
solution containing 6% bisacrylamide, ammonium persulfate, and
tetramethylethylenediamine (TEMED) was introduced into the crystalline lattice and
polymerized to form a dense and transparent hydrogel film. PS beads were dissociated in
toluene overnight, and finally, a closepacked array of cavities and interconnections of its six
nearest neighbors were left behind. The remained gel as shown in Figure 4 was equilibrated in
the running buffer and provided for DNA electrophoresis.
PRACTICAL APPLICATIONS OF
MICRO- AND NANOSTRUCTURES
In this chapter, practical applications of micro- and nanostructures which were fabricated
by above mentioned technologies are described.
Micron-sized Pillars
The pioneering work using the device fabricated by microfabrication technique for
practical applications was demonstrated by Austin et al [19]. They fabricated the micropillars
which are 0.15 m high, 1.0 m in diameter, and 1.0 m spacing on a standard 3 inch diameter
silicon wafer. A 0.5 m SiO2 layer was grown into the surface of the micropillar array for
optical observation and electrical insulation. One set of arrays were made with the
micropillars on a square lattice, and another set also on a square lattice but with the lattice
rotated 45˚. Although the effective pore size of 1.0 m corresponds to a physically unstable
0.05% agarose gel, they indicated that the micropillar array is capable of length fractionation
up to a length of ~100 kbp in a DC field through direct observation of DNA migration in the
micropillar array.
Figure 4. (A) Fabrication process of self-assembled colloidal arrays in a microfluidic channel. (D,E) SEM
images of a constructed matrix of 330-
(F) SEM image of a hexagonally closed packed 2-
(Reproduced from Zeng, Y. et al., Anal. Chem. 2007, 79, 2289. With permission)
The micropillar array was applied to pulsed-field electrophoresis for long DNA
separations [6, 7]. The past pulsed-field electrophoresis in an agarose gel was time-
consuming, typically requiring over 12 hrs, and consumed running buffers more than 1 L.
Therefore the micropillar array system was strong candidate to substitute the conventional
pulse-field electrophoresis systems. Bakajin et al. demonstrated that T4 (168.9 kbp) and (48.5
kbp) DNAs could be resolved into two clearly separated bands within 10 s by the micropillar
array system [7]. This result corresponds to a mass resolution of 6% in 11 min in a 1-cm-long
array. In this system, they used entropic focusing method to concentrate and form a thin band
of DNA samples for DNA injection at the entrance of the micropillar array. On the other
hand, continuous DNA sample injection and separation by pulsed-field electrophoresis were
achieved in the DNA prism by Huang et al [6]. In this prism, microfluidic channels surround
the micropillar array and connect it to fluid access reservoirs where electric fields are applied.
These microfluidic channels provide continuous sample loading and collection ports and
create uniform electric fields across the entire micropillar array, whereas conventional pulsed-
field electrophoresis systems create uniform electric fields by surrounding the slab gel with
many electrodes. The prism separated 61-209 kbp DNA molecules in 15 s with 13%
resolution.
Figure 5. The trajectories of different sized DNA fragments, 15 and 33.5 kbp, in the Brownian ratchet array.
(Reproduced from Chou, C. et al., Proc. Natl. Acad. Sci. USA 1999, 96, 13762. With permission)
Another important demonstration of micron-sized pillars is Brownian ratchet array which

consists of asymmetric micron-sized obstacles. The basic concept is that, by using a regular
lattice of asymmetric obstacles to rectify the lateral Brownian motion of the molecules, they
follow different trajectories through the device based on the molecular sizes [20]. Compared
with conventional gel electrophoresis, one of the advantages of this technique is that sample
loading and separation could be achieved continuously as in the DNA prism. In the initial
design of the Brownian ratchet array, samples consists of different components were injected
in a line stream at the top corner of the device, separated through the array and collected
different positions at the bottom edge [4] as shown in Figure 5. This Brownian ratchet
achieved a nominal 6% resolution by length of DNA molecules in the size range 15-30 kbp.
To produce a fine stream at sample loading, a novel our-of-plane injection scheme was
proposed by Cabodi et al [21]. They successfully separated a mixture of T2(164 kbp) and
T7(37.9 kbp) coliphage DNA. However, the separation of large DNA molecules in these
microfabricated Brownian ratchet array was slow because it totally relies on a diffusion
process. Tilted Brownian ratchet array improved the separation resolution and speed by
factors of 3 and 10, respectively [22, 23]. Whereas previous Brownian ratchet arrays with no
tilt required about 140 min of running time to resolve 48.5 from 164 kbp DNA molecules
with resolution 1.4, the same resolution could be achieved in only 14 min at a flow tilt angle
of 7.2˚. In this tilted Brownian ratchet array, great reduction of the amount of diffusion
required for ratcheting led to faster separation without any loss of resolution.
As mentioned above, the array of the asymmetric micron-sized obstacles was designed
that inherently rely on diffusion for separation. DNA molecules that could be regarded as
hydrodynamically equivalent particles have many different migration paths based on different
diffusion lengths. Even if the same size of DNA molecule was loaded, the DNA molecule
was eluted as a broaden zone by multipath effect such as size-exclusion chromatography. To
eliminate this multipath zone broadening, Huang et al. demonstrated a separation process that
creates equivalent migration paths for each particle in a mixture [3]. Their separation process
uses laminar flow through a periodic array of micron-scale obstacles. The mixture of 0.8-,
0.9-, and 1.0-
chromosomes (BAC) of 61 and 158 kbp were separated in 10 min with a resolution of 12%.
In this device, the separation is based on a physical displacement, not on a random process
such as diffusion. Therefore a much sharper transition occurred when the flow velocity was
increased to minimize diffusion effects. In the case of large DNA molecules, higher fields
resulted in lower resolution, possibly because of random deformation and stretching of DNA
by collisions with the obstacles. Having said that, this separation technique is very useful for
relatively rigid spherical particles to achieve faster separation than Brownian ratchet. Using
this technique, Davis et al. demonstrated the fractionation of whole blood components and
isolation of blood plasma with no dilution [2]. Whole blood components were separated based
on their hydrodynamic size, but not their mass, in this device. They successfully separated
white blood cells, red blood cells, and platelets form blood plasma at flow velocities of 1000
m/s and volume rates up to 1 l/min.
Figure 6. Separation of (48.5 kbp) and T4 (165.6 kbp) DNA by the nanopillar chip under DC electric field.
(Reproduced from Kaji, N. et al., Anal. Chem. 2004, 76, 15. With permission)
Nano-sized Pillars
An array of nano-sized pillars, nanopillars, was applied for DNA separation by Turner et
al. [5] and Kaji et al. [9]. Turner et al. demonstrated that the devices could function as a
molecular sieve since they observed a significant mobility difference between two different
types of DNA molecules [5]. In the experiments of Kaji et al., the size of pillars and the
spacing between pillars are designed as a DNA sieving matrix for optimal analysis of large
DNA fragments over a few kbp. DNA fragments ranging from 1 to 38 kbp were separated as
clear bands, and furthermore, the mixture of (48.5 kbp) and T4(165.6 kbp) DNAs were
successfully separated by a 380-m-long nanopillar channel within only 10 s even under a DC
electric field [9] as shown in Figure 6.
Periodical Nanoslits
The microchannel which consists of narrow constrictions and wider regions causes size-
dependent trapping of DNA at the onset of a constriction. This process creates electrophoretic
mobility differences and enables efficient DNA separation without use of a gel matrix.
Surprisingly, longer DNA molecules migrated faster than shorter DNA molecules in this
system. This mechanism is explained from the viewpoint of the energy barrier across the
narrow constrictions, nanoslits. DNA molecules were entropically trapped at the constriction
and escaped with a characteristic lifetime. The difference of mobility comes not from the
escaping activation energy barrier, but from the fact that the surface area of a DNA molecule
facing the thin slit is different because of the difference in their size. Longer DNA molecules
escape faster simply because more monomers are facing the thin slit, and are able to form a
beachhead for escape [24-26]. The selectivity of the DNA separation was shown to be
dependent on the depth of deep and shallow channel regions, applied electric field, and
number of entropic barriers [27]. Recently, Fu et al. expand the application field of the
entropic trapping array to shorter DNA and protein separations [28-30]. They showed
experimental evidence of the crossover from Ogston-like sieving to entropic trapping,
depending on the ratio between nanofilter constriction size and DNA size [29]. The crossover
from Ogston sieving to entropic trapping was measured by mobility of DNA of a size ranging
from 0.5-8 kbp which corresponds to the gyration radius of 40-220 nm in a 73 nm nanofilter
array. Nanofilter arrays with a gap size of 40-180 nm were fabricated and applied for SDS-
protein complexes [30]. Separations of SDS-protein complexes and non-denatured proteins
are also demonstrated in a microfabricated anisotropic sieving structure consisting of a two-
dimensional periodic nanofluidic filter array [28] as shown in Figure 7. This device
successfully demonstrated high-resolution continuous-flow separation of a wide range of
DNA molecules from 50 bp to 23 kbp and proteins from 11 kDa to 400 kDa in just a few
minutes.
Figure 7. Whole design of a microfabricated anisotropic sieving structure consisting of a two-dimensional

periodic nanofluidic filter array for DNA and protein separations. (Reproduced from Fu, J. et al., Nat.
Nanotech. 2007, 2, 121. With permission)
Nanochannels
When the polymer that is freely coiled in solution is confined in a nanochannel, a

confinement elongation is observed. In the nanochannel, self-avoidance increases the scaling
exponent for the contour length because the polymer is prevented from back-folding.
Tegenfeldt et al. extended genomic-length molecules sized over 1 Mbp in arrays of imprinted
nanochannels and measured the length of the extended DNA molecules directly at a single
molecule level [10] as shown in Figure 8. They demonstrated that nanochannel-based
measurements of DNA length have several advantages over current electrophoresis-based
techniques. Using the nanochannel array with diameters of 100-200 nm, restriction mapping
of DNA molecules by restriction endonucleases was demonstrated by Riehn et al. [13].
Combining DNA introduction into the nanochannels by electrophoresis and the formation of
concentration gradient of magnesium ion (Mg2+) and EDTA in the nanochannels by diffusion,
the restriction reactions could be observed at desired locations in the nanochannels. The
measurement of the positions of restriction sites were achieved with a precision of 1.5 kbp in
1 min. Wang et al. demonstrated the direct imaging of GFP-LacI repressor proteins bound to
bacteriophage
DNA molecules in nanochannels [12]. The number of bound protein molecules was counted
by using an integrated photon molecular counting method, and then, the locations of the
bound protein were determined.
Figure 8. Imprinted nanochannels and the extended DNA molecules in the nanochannels at a single molecule
level. (Reproduced from Tegenfeldt, J. et al., Proc. Natl. Acad. Sci. USA 2004, 101, 10979. With permission)
Although the above mentioned nanochannels were fabricated by nanoimprint lithography

and FIB milling, soft lithography based on elastomer replicas was used for fabrication of
relatively large-scale nanochannels, ~1 m. This relatively large nanochannel was successfully
applied for DNA stretching by using low-ionic-strength buffers. Enzymatic labeling of
specific sequences on elongated DNA molecules inside the nanochannels was imaged via
fluorescence resonance energy transfer by Jo et al. [31].
Nanomaterials
A microchannel filled with novel nanomaterials, which were fabricated by ―bot tom-up
approach‖ based on organic chemistry and polymer synthesis, could be also regarded as a
new type of nano-fabricated structures in microchannels. A typical example of nanomaterial
for DNA separation is so-called nanoball, core-shell type nanospheres, in microchip
electrophoresis14. DNA fragments up to 15 kbp were successfully separated within 100 s
without observing any saturation in migration rates. DNA fragments migrate in the medium
while maintaining their characteristic molecular structure. Although this medium requires
little complicate procedures combining pressure and electric field for sample injection, low-
viscosity and high-resolution medium could be widely applicable to simple microchannels.
Molecular sized cavities interconnected by nanopores were used to investigate
confinement effects on long DNA molecules [17, 18]. In this study, they indicated the
possibility of DNA separation by the cavities. Self-assembled colloidal arrays, which have
contradictory structures of the above mentioned cavities, demonstrated their separation ability
as a sieving matrix. The flexibility of pore size enabled by this methodology provided
separation of biomolecules with a wide size distribution, ranging from proteins (20-200 kDa)
to dsDNA (0.05-50 kbp) [16]. Under moderate electric fields, complete separation can be
finished in minutes, with separation efficiency comparable to gel/polymer-filled or micro-
/nanofabricated microsystems. To realize continuous separation and collection, a two
dimensional colloidal self assembly bed surrounded by multiple microchannels was prepared.
High-throughput separation of 2-50 kbp DNA was achieved by this 2D microsystem.
CONCLUSION
As seen in this chapter, nanofabricated structures allows biomolecule separations based
on novel principles such as entropic trapping and pillar arrays that is difficult to achieve using
the past gel or polymer based separation. Precise control of nanospaces, which are produced
by nanofabricated structures, explores fundamental understanding of nanofluidics and
polymer dynamics in confined space. Single molecule approaches such as the nanochannel
array approach might be an alternative to achieve high-throughput analytical systems beyond
the current TAS. For construction of integrated systems from cell lysis to biomolecule
analysis on a single chip, combining microfluidics and nanofluidics depending on analytes‘
sizes and features are essential. The continued successful development of TAS including
nano-fabricated structures is a promising effort to disclose a new insight of chemistry and
biology.
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Lecture Material 16
BIONIC SUPERHYDROPHOBIC SURFACES BASED ON

COLLOIDAL CRYSTAL TECHNIQUE
ABSTRACT
Biomimetic research reveals that superhydrophobicity with a self-cleaning effect of a
lotus leaf is ascribed to the combination of both a hierarchical micro-/nanostructure on
the surface and a low surface-energy material covering the surface. The syntheses of
colloidal crystals and micro/nano structured arrays based on the colloidal crystals have
been well developed, a lot of ordered micro- or nanostructured arrays can be prepared
using the colloidal monolayer templates. These ordered arrays and the colloidal crystals
are rough on the surfaces in the micro- or nanoscale, which gives a good chance to create
the superhydrophobicity on the sample surfaces. In this chapter, we review the
superhydrophobic surfaces based on the various colloidal crystal techniques. As we
know, the micro- or nanostructured arrays by colloidal monolayer templates have
important applications in photonics, photoelectronic devices etc. This suggests that
nanodevices built from these nanostructured arrays could be waterproof and self-cleaning
in addition to their special device functions after possessing the superhydrophobicity.
INTRODUCTION
Biomimetic research has recently revealed many interesting phenomena of natural
organisms, such as self-cleaning effect of a lotus leaf (so-called lotus effect) that removes
contamination on its surface and striking adhesive force of a gecko‘s foot [1]. These unique
functionalities are attributed to the combination of hierarchical micro- and nanostructures on
the surfaces of the natural organisms. Specially, the lotus effect is related to the
superhydrophobicity with a water contact angle (CA) larger than 150° and a sliding angle
(SA) less than 10°, which is caused by the combined effect of both a hierarchical micro-/
nanostructure on the surface and a low surface-energy material covering the surface. This
superhydrophobic property can be widely used for preventing the adhesion of water or snow
to windows, antioxidation coating, self-cleaning utensils, and microfluidic devices [2].
Inspired by the lotus effect, various techniques to synthesize bionic superhydrophobic
surfaces have been recently developed [3]. Additionally, theoretic investigation about
wettability on rough surfaces was also explored and two famous models were established to
explain the wettability phenomena on the rough surface. Generally, when a water droplet dips
into the pores or grooves of a rough surface, Wenzel gave a quantitative description of the
surface wettability as follows [4]:
cos r r cos (1)
where r is the roughness factor and is defined as the ratio of the total surface area to the
projected area on the horizontal plane. r and are the CAs for the film with rough and
smooth surface, respectively. For this Wenzel‘s type surface, obviously, increased roughness
can enhance the hydrophobicity and/or hydrophilicity of hydrophobic and/or hydrophilic
surfaces. When a liquid droplet contacts with a rough surface and is completely lifted up by
the roughness features, or it cannot dip into pores or grooves on the rough surfaces, another
model was presented by Cassie as the following equation [5]:
cos θr = f1 cos θ - f2 , (2)
where f1 and f2 (= 1 - f1) are the area fractions of a water droplet in contact with the solid and
air on the rough surface, respectively. Obviously, increasing f2 can lead to larger r . It means
that the area fraction on pores or grooves in the surface is important to the hydrophobicity for
Cassie‘s type surface. According to these two models, one can find that a superhydrophobic
surface should be of two features, the enough roughness and low surface free energy which
can produce a hydrophobic property on the native flat surface. Generally, the low surface free
energy can be obtained by chemical modification using the low surface free energy materials,
such as coating with fluoroalkylsilanes or thiol [3].
Recently, the syntheses of colloidal crystals and micro/nano structured arrays based on
the colloidal crystals have been well developed. Plentiful order micro- or nanostructured
array films could be synthesized using the colloidal monolayer as a template, including pore
arrays, pillar arrays, hierarchical micro/nano structured arrays etc [6]. The surfaces of these
ordered arrays and the colloidal crystals are rough at the microscale or nanoscale. It is
expected that such ordered structured arrays could show superhydrophobicity. This means
that nanodevices built from these nanostructured arrays could be waterproof and self-cleaning
in addition to their special device functions if they possess the superhydrophobicity.
In this chapter, we review the recent developments of the bionic superhydrophobic
surface based on the colloidal crystal, including synthesis and corresponding theoretic
analyses, in the following sections.
SUPERHYDROPHOBIC SURFACES OF
PLASMA-ETCHED COLLOIDAL MONOLAYERS
As we know, the typical colloidal monolayer exhibits a hexagonal-close-packed (hcp)
arrangement and can supply the well ordered nanoscaled surface roughness. Therefore, it may
give a possibility to induce the superhydrophobicity. However, Chen et al. found that the
polystyrene (PS) colloidal monolayer with hcp arrangement and PS-sphere size of 440 nm
just exhibited hydrophobicity with a water CA of 132°, after coating a gold layer and
modification with octadecanethiol, instead of superhydrophobicity as expected [7]. In order to
achieve the superhydrophobicity, they reduced the PS sphere size by the oxygen plasma
etching, but the periodicity of the PS colloidal monolayer was kept unchanged. By this route,
the hcp colloidal monolayer could be changed to the one with a hexagonal-non-close-packed
(hncp) arrangement, which is very helpful to induce the superhydrophobicity after chemical
modification. For instance, the PS sphere sizes in the colloidal monolayer can be reduced and
well controlled from 400 nm to 190 nm by oxygen plasma etching, as shown in Figure 1 [7].
These samples displayed the increasing hydrophobicity with decrease of the PS phere size
after modification with thoil. When the sphere size was less than 330 nm, the array would
demonstrate the superhydrophobicity. The Cassie`s model can give a good explanation for the
water dewetting behavior on these nanostructured films. With reduction of the sphere size, the
liquid-solid contact area will decrease if we assume that the water contact line lies on the
upper part of the spheres, the decreasing liquid-solid contact area will result in the
enhancement of hydrophobicity according to Cassie`s equation. Additionally, they found that
the water CA on the double layer PS sphere arrays had larger value than those on the
monolayer, which can be attributed to the less defects of the double layer than the monolayer.
a b
c d
Figure 1. SEM images of size-reduced PS sphere by reactive ion etching and corresponding water droplet
shapes on the sample surfaces. The sizes of PS spheres and water CAs are (a) 400 nm, 135°, (b) 360 nm,
m. Reprinted with permission from Ref. 7,
Copyright 2004 America Chemical Society.
Figure 2. Fabrication procedure of binary assemblies by consecutively dip-coating CaCO3-PNIPAM particles

and silica or PS colloidal spheres in aqueous solution. Reprinted with permission from Ref. 8, Copyright
2005 America Chemical Society.
a b
c d
Figure 3. (a) SEM image of a 2 dimensional CaCO3-PNIPAM particle non-close-packing array produced by
first-step dip-coating a 0.1 wt% aqueous particle aqueous suspension. (b) SEM image of binary colloidal
assemblies by second-step dip-coating a 1.0 wt % 296 nm silica spheres on the CaCO3-PNIPAM particle
non-close-packing arrays. Low (c) and high (d) magnification SEM images of binary colloidal assemblies by
second-step dip-coating a 2.0 wt% 296 nm silica sphere aqueous suspension on a CaCO3-PNIPAM particle
non-close-packing array. Reprinted with permission from Ref. 8, Copyright 2005 America Chemical Society.
SUPERHYDROPHOBIC SURFACES FROM

BINARY COLLOIDAL ASSEMBLY
In order to obtain the superhydrophobic surfaces, enough roughness on the surface is very
important. Wang et al. prepared binary structures with hierarchical roughness by two-step
consecutive dip-coating (Figure 2). They first synthesized the CaCO3-loaded poly (N-
isopropylacrylamide) spheres (denoted as CaCO3-PNIPAM in the following) by in situ
mineralization of CaCO3 within PNIPAM gel spheres. These CaCO3-PNIPAM microspheres
have uniform sizes and can be easily self-assembled into colloidal monolayers with a non-
close-packed arrangement by dip-coating and subsequently drying with effect of the
shrinkage of these hydrogel spheres. The silica or PS spheres were fabricated on the substrate
with CaCO3-PNIPAM sphere arrays by second-step dip-coating and the binary packed
structures would be achieved, as shown in Figure 3. Additionally, these binary colloidal
assemblies demonstrated the enhanced mechanical stability after heating treatment, producing
a good durability in applications, for example, in self-cleaning surfaces.
After modification with low surface free energy molecules, the as-prepared film with
binary colloidal structures displayed an enhanced hydrophobicity. The silicon wafer with
CaCO3-PNIPAM microsphere non-close-packed arrays took on hydrophobicity with a water
CA of 99o, however, the binary colloidal assembly obtained by second-step dip-coating of 1.0
wt % aqueous suspension of 296 nm silica spheres showed the improved hydrophobicity with
water CA of 130o and the one by higher concentration (2.0 wt % silica sphere with the same
size) led to the superhydrophobicity with water CA of 160o. The enhanced hydrophobicity of
binary colloidal assembly was ascribed to the increasing roughness compared with CaCO3-
PNIPAM microsphere non-close-packed arrays. With increase of concentration of silica
sphere aqueous suspension, the roughness on the binary structures will increase in some
contents, and finally results in the superhydrophobicity on the surface.
ORDERED POROUS SEMICONDUCTOR ARRAY FILMS

Tunable Wettability Caused by the Precursor Concentration
As we reported [6], ordered pore array films can be prepared by colloidal monolayer
templates and their morphologies are closely dependent on the experimental conditions, for
example, precursor concentrations. Recently, we have found that the morphologies of ZnO
ordered pore arrays can be well controlled by increasing the precursor concentrations. The
surface roughness increases with increase of precursor concentration. Therefore, it is expected
to use this phenomenon to control the surface wettability. Figure 4 shows ZnO ordered pore
arrays with different morphologies prepared by solution dipping method with different
precursor concentrations [6f, 9]. These three kinds of surface microstructures correspond to
different precursor (zinc acetate) concentrations. At a low precursor concentration (0.3 M,
Figure 4a), the pores in the film demonstrate truncated hollow spheres, and the pore sizes are
smaller than the diameter of the colloidal sphere of the template. The depth is also smaller
than the radius of the template. With increase of precursor concentration to 0.5 M, each pore
looks like a hollow hemisphere and the pore size increases to about the PS diameter (Figure
4b). If the concentration is further increased (1.0 M), the pores show a noncircular shape from
the top view. The surface morphology exhibits a hierarchical structure, which is composed of
close-packed rough wreaths and some small particles with 30 nm in size, as shown in Figure
4c and d.
The wettability of the as prepared ZnO porous arrays with different surface morphologies
was by measured the water CA by carefully dropping water droplets upon their surface in a
dark chamber. Figure 5 (a-c) gives the photographs of the water droplets on different films
and the corresponding water CAs are 125o, 131o, and 143o, respectively. These indicate that
such ordered porous structures can effectively increase the hydrophobicity in comparison
with relatively flat ZnO films (CA, 109o). Moreover, the wettability shows clearly
dependence on surface microstructures, which is determined by the precursor concentration.
Moreover, after chemical modification with low surface energy materials, fluoroalkylsilanes,
the above-mentioned three samples demonstrate superhydrophobicity and the corresponding
water CAs increase to 152o, 156o, and 165o, respectively, as shown in Figure 5 (d-f), where
the water droplets display rather spherical shapes.
a b
c d
Figure 4. SEM images of ZnO ordered pore array fims prepared by solution dipping method using different
precursor concentration: (a) 0.3 M, (b) 0.5 M, (c d) 1.0 M. Reprinted with permission from Ref. 9, Copyright
2005 Elsevier.
Figure 5. Water droplet shapes on ZnO pore arrays and the corresponding static CAs on the as-prepared
samples (a–c), corresponding to Figure 4a–c; (d–f) show the water droplets and CAs on the same samples
after chemical modifications, corresponding to (a–c), respectively. Reprinted with permission from Ref. 9,
Copyright 2005 Elsevier.
Interestingly, the water droplets on the surfaces of the modified samples shown in Figure
4 a and b did not slide even when the surfaces are almost tilted vertically. But, slightly tilted
(less than 5o), the water droplets on the modified surface of Figure 4 c rolled off quickly. This
phenomena can be described more accurately by the dynamic CAs, advancing CA ( A ) and
receding CA ( R ). The dynamic CAs of the modified ordered pore array films fabricated by
1.0 M precursor are too small to be measured (sliding angle, <5o). Nevertheless, its CA
hysteresis H (= A - R ) is the smallest due to its largest water CA and smallest sliding
angle. Our results indicate that, with rise of precursor concentration, the CA hysteresis of
such as-prepared and modified nanostructured films both decrease. Moreover, the as-prepared
samples have a relatively larger CA hysteresis (more than 21°) while the modified films have
a smaller one (less than 9°).
As we know, dewetting behavior of the rough surface that can be described with Wenzel
model (Wenzel surface) show larger H than the surfaces that can be described with Cassie
model (Cassie surface) due to the bigger adhesive force between the water and film. In this
case, the as-prepared sample has a large H (more than 21°). Consequently, this dewetting
behavior principally prefers the Wenzel model. From Figure 4, it can be found that the
roughness of the porous array film increases with increase of precursor concentration.
Correspondingly, according to Wenzel model, r should increase, which agrees well with
these results. After modification with fluoroalkylsilane, these ordered pore array films exhibit
superhydrophobicity (CA>150°) with a small H (less than 9°), which corresponds to Cassie
surface. After modification with lower surface free energy materials, air can be trapped into
such ordered porous film. For the modified samples, since the parameter f1 decreases with
the precursor concentration, corresponding water CA also increases.
With increase of precursor concentration, CA hysteresis decreases for both the as-
prepared and modified samples, possibly due to the gradual reduction of the adhesive forces
between water droplets and films induced by the concentration-dependent roughness. When
the precursor concentration is not high (say, 0.3 M or 0.5 M), although the modified samples
are of superhydrophobicity, water droplets on their surface do not roll off when the films are
tilted. This is mainly attributed to the continuous, stable three-phase (air-liquid-solid) contact
line for such net-like ordered pore array films. However, when the precursor concentration is
high enough (say, 1.0 M), the surface of the ordered porous film is much rougher and shows
hierarchical structure (Figure 4 c-d), which is similar to that of the well-known lotus leaves,
leading to superhydrophobicity with large water CA (165°) and small sliding angle (less than
5°). This film could be expected to show self-cleaning effect.
Controlled Superhydrophobicity Based on Structural Periodicities
Ordered indium oxide pore array films with different morphologies were prepared by sol-
dipping method using the PS colloidal monolayers as templates. These porous films took on
hydrophilicity. However, after modification with fluoroalkylsilanes, all of these pore array
films displayed superhydrophobicity due to rough surface and low surface free energy
materials on their surfaces. Interestingly, with increase of the pore size in the films, the
superhydrophobicity could be controlled and was gradually enhanced due to the
corresponding increase of roughness caused by nanogaps produced by the thermal stress in
the annealing process with increase of film thickness. The ordered pore array films could be
fabricated on the substrate after removal of the PSs and annealing at 400 oC in air for 1 hour.
Figure 6 shows the SEM images of macropore array films prepared using the colloidal
monolayer with different PS sphere sizes ((a),
exhibit the orderly hexagonal arrangement, which corresponds well to the colloidal
monolayer template.
When the PS sphere size template, the honeycomb structured,
ordered indium oxide macropore array films with hexagonal alignment are fabricated by
above-mentioned method, as shown in Figure 6 a and b. If the size of PS sphere increases to
the pore array film also can be formed and pore shapes are close to hollow hemispheres
(Figure 6c and d). However, most walls between the two neighboring pores cracked and many
nanogaps were produced, as clearly shown in the magnified image (Figure 6d). Further
ed
and changed from hollow hemisphere shape to irregular shape. Additionally, many nanogaps
with big size also were produced on the walls between the two neighboring pores, as shown
in Figures 6 e and f.
In the annealing process of film materials, some thermal stress will be generally produced
between materials and their supporting substrates. With increase of the film thickness, the
influence of thermal stress on the film morphology is becoming much serious, and even the
rupture. In this work, with increase of PS sphere size in the colloidal monolayer template, the
thickness of macropore array film fabricated using such template will correspondingly
increase. When the PS sphere size is relatively small (1
array film is about half of sphere size and the thermal stress has nearly no influence on pore
array films in the annealing process at 400 oC for 1 hour, and so morphology of the pore array
film corresponds perfectly to that of the colloidal monolayer template and exhibits
hexagonally arranged regular pores. However, when the PS sphere diameter is of 2
which leads to the thicker pore array film, the thermal stress begins to play an important role
in formation of pore film and produce many nanogaps on the pore walls. If the thickness of
pore array film is further increased by the colloidal template w
influence of thermal stress become more serious, resulting in the deformed pore shapes and
nanogaps on the pore walls. In addition, for the pore array film, it belongs to a kind of rough
surface in essence. According to these results, one can clearly see that roughness of pore array
film increases with increase of the PS sphere size in the colloidal monolayer templates due to
the effect of thermal stress.
Before modification with fluoroalkylsilane, all samples took on the hydrophilicity (water
CA<90o) and the water CA decreased with increase of pore size, as shown in curve b in
Figure 7. After modification, the morphologies of such ordered pore arrays were nearly
unchanged. However, the wettability was changed from the original hydrophilicity to
superhydrophobicity. Moreover, with increase of the pore size from 1
o
, 158o and 163o, respectively, as shown
in curve a of Figure 7. Additionally, the water droplet on them was gradually close to
spherical shape with increase of pore size in the films, as shown in insets in Figures 6a, 6c,
6e, respectively, reflecting that the superhydrophobicity enhanced with increase of pore size.
Our results indicate that the superhydrophobicity can be well controlled by change the
periodicity of colloidal templates.
Figure 6. SEM images of the as prepared indium oxide pore array films. The diameter the PS sphere in the
colloidal monolayer (a)- - -
(c) and (e), respectively. The precursor concentration: (a) 0.2 M, (c) 0.4 M, (e) 05 M. The insets in (a), (c)
and (e) are photographs of the water droplets on the corresponding pore arrays after chemical modifications
with fluoroalkylsilane, and the corresponding water CAs are 155o, 158o and 163o, respectively. Reprinted
with permission from Ref. 10, Copyright 2007 Elsevier.
a
b
Figure 7. The water CAs as a function of pore size of the indium oxide pore array film before (b) and after (a)
modification with fluoroalkylsilane. Reprinted with permission from Ref. 10, Copyright 2007 Elsevier.
The Wenzel model can well explain why such pore array films exhibit hydrophilicity and
the hydrophilicity enhances with increase of pore size. For a Wenzel type surface, obviously,
high roughness can enhance both hydrophobicity of hydrophobic surface and hydrophilicity
of hydrophilic surface. Theθ value of a relative flat indium oxide film, prepared by dip-
coating method without using colloidal monolayer template, was 85o, indicating that the
wettability of a flat film is hydrophilic. In this work, these indium oxide pore array films are
much rougher than the relative flat surface, and the roughness of pore array films increases
with increase of pore size due to many nanogaps produced on the pore walls as mentioned
above. The relative flat indium oxide film displayed the hydrophobicity with water CA of
115o after chemical modification. According to Wenzel model, for the same reason, the
indium oxide pore films are so rough that the hydrophobicity is enhanced to water super-
repellence and all such films displayed the superhydrophobicity. The superhydrophobicity
increases with increase of pore size due to corresponding increase of roughness.
Reversible Wettability
Very interestingly, we find that ultraviolet (UV) light irradiation leads to transition of the
wettability from hydrophobicity to hydrophilicity for the as-prepared ZnO pore array films.
Figure 8 demonstrates the results for the as-prepared sample kept in the dark chamber for 7
days before and after irradiation for 2 h by UV light from a 500 W Hg lamp with a 400 nm
filter. The differences between hydrophobic and hydrophilic CA increase from 89o o
with rise in precursor concentration. If we alternate keeping the samples in dark for 7 days
and then exposing them to UV light, this wettability transition is reversible.
As we know, the electron-hole pairs generated by UV irradiation will move to the surface
and these holes will react with the lattice oxygen to form the surface oxygen vacancies.
Meanwhile, water and oxygen may compete to adsorb on them. The defective sites are
kinetically more favorable for hydroxyl adsorption than oxygen adsorption. Such reason and
the rough surface can lead to hydrophilicity. Due to the instability of the hydroxyl absorption
and the thermodynamical favorite of the oxygen adsorption, oxygen atoms will replace the
hydroxyl groups adsorbed on the defective sites gradually when the UV irradiated ordered
porous films were placed in the dark chamber. So when the original state of the surface is
recovered, the wettability is reconverted from hydrophilicity to hydrophobicity. According to
Wenzel model, the surface roughness enhances both hydrophobicity of hydrophobic surfaces
and hydrophilicity of hydrophilic ones. The increase of the porous film roughness factor
induced by increasing precursor concentration can enhance both the hydrophobicity and
hydrophilicity of the surface with two contrary states, leading to ever-increasing CA
differences of reversible wettability transition with rise in precursor concentration, shown in
Figure 8. Such a surface is very important in application of microfluidic devices.
Figure 8. Transition between hydrophobicity and hydrophilicity induced by UV irradiation for the as-
prepared ZnO porous films. Reprinted with permission from Ref. 9, Copyright 2005 Elsevier.
SUPERHYDROPHOBIC SILVER HIERARCHICAL

RING-LIKE ARRAYS
Using a colloidal monolayer with PS sphere of 5 µm as the template and silver acetate
(AgAc) as precursor after heating at 360oC for 3 h, silver orderly structured array was
fabricated on the substrate by solution dipping method, as shown in Figure 9. This ordered
array with the periodicity of 5 µm exhibits a hexagonal alignment. Each unit is of a ring-like
structure (Figure 9 a, b) in the array and the whole array has rough inner walls composed of
nanoparticles with average size of ca. 135 nm, as shown in Figure 9c. Most silver
nanoparticles are sintered with each other because of the surface melt during the heating
process. The heating process leads to the formation of highly durable micro/nano structured
arrays: these structures were not destroyed and the whole hierarchical arrays were not
detached from a substrate even when the substrate was ultrasonically washed in water for 30
min.
When the water droplet with a small volume was added on the as-prepared silver
hierarchical ordered structured array, it spread out rapidly and the surface exhibited the
hydrophilicity with water CA of 23o. However, after modification with thiol, the shape of
water droplet was nearly spherical and the sample showed the superhydrophobicity with
water CA of 169o, as shown in Figure 10. Additionally, the sliding angle for water droplet
was only 3o, indicating a self-cleaning effect.
In order to reveal the origin of the superhydrophobic property of the as prepared
micro/nano-structured film, a relatively flat silver film (Figure 11a), a silver nanoparticle film
(Figure 11b) and a pore array film with smooth inner walls (Figure 11c) were prepared. The
water CAs of these films were systematically investigated before and after the modification
with thiol. The flat silver film was of water CA of 69o and was transformed to be hydrophobic
with a water CA of 108o after surface modification (Figure 11a). For silver nanoparticle film,
the water CA was 35o and 134o before and after modification, respectively. These
experimental data indicate that the silver flat film and nanoparticle film were unable to induce
superhydrophobicity. The silver pore array film had a water CA of 33o, which was smaller
than the flat silver film, but 151o after the surface modification (Figure 11b). In addition, the
water droplet of 3 mg cannot roll off on its surface when the film was tilted to any angle, even
upside down, which is mainly due to the stable, continuous three phase contact line (air-
liquid-solid) formed on such netlike pore array structure when the water drop is added on the
surface. Although the silver pore array exhibited the superhydrophobicity after modification,
it was far away from the requirement of the self-cleaning effect. Therefore, the results
demonstrated that the self-cleaning effect of the as prepared ordered ring-like array film
resulted from the special combination of micro and nano structures of hierarchical array, like
lotus leaves.
The Wenzel model can well explain why these surfaces are more hydrophilic and more
hydrophobic before and after modification. In above cases, the hierarchical ring-like array has
the smallest water CA and the biggest CA before and after modification, respectively, which
indicates that this hierarchical structure has the largest roughness. When a water droplet is
added on the chemically-modified ordered ring-like array film with the hierarchical structure,
air can be trapped in interstices or corrugations that are produced between the microstructure
and the nanostructure. In this case, according to the Cassie equation, f2 for the as prepared
surface can be calculated to be 0.98. For the pore array film shown in Figure 11c, however,
the corresponding f2 is only 0.90. This analysis reveals that the hierarchical micro/nano-
structured surface produces large amount of air traps between the microstructure and the
nanostructure, and that the strong superhydrophobicity of the bionic surface is mainly caused
by the unique hierarchical micro/nano structures and the subsequent surface chemical
treatment.
Figure 9. SEM images of the as prepared silver hierarchical ring-like array film. (a) low magnification. (b) an
image of a feature ring-like unit. (c) morphology within a ring-like unit with larger magnification. Reprinted
with permission from Ref. 11, Copyright 2007 America Chemical Society.
a b
Figure 10. Water droplets on the silver ring-like structured array film shown in Figure 9. (a) before
modification with thiol, CA: 23o, (b) after modification, CA: 169o. Reprinted with permission from Ref. 11,
400
b nm
5
c m
Figure 11. Morphology and wettability for the Ag surfaces with different structures. (a) a relatively flat silver
film on the glass substrate obtained by the thermal evaporation deposition. (b) a silver particle film on the
glass substrate synthesized by the decomposition of AgAc coating at 360 oC for 3h. (c) a silver pore array
film with smooth pore walls synthesized by the electrodeposition using colloidal monolayer with sphere size
thiol, respectively. Reprinted with permission from Ref. 11, Copyright 2007 America Chemical Society.
WETTABILITY OF SILICA MICRO/

NANOSTRUCTURED ARRAYS
The SiO2 ordered pore array film can be fabricated by sol-dipping method using colloidal
monolayer as template, as we previous reported [12]. The as-prepared 2D SiO2 ordered pore
array is shown in Figure 12. Figure 13 shows the water droplet shapes and corresponding
CAs on such silica macropore array film and flat film before and after modification by
(heptadecafluoro-1,1,2,2-tetrahydrodecyl) triethoxysilane. When a water droplet was added
on the silica macropore array films, it spread out rapidly on surface of the sample, which
shows superhydrophilicity with a CA of about 5° (Figure 13 a). After modification with low
free energy materials, however, such porous film presented superhydrophobicity. The shape
of the water droplet on such film was inclined to be spherical and the corresponding CA was
154°, as displayed in Figure 13b and Figure 14. For comparison, the wettability of the flat
silica film (about 1
monolayer template, was also investigated, the water CA of the film was about 10° (Figure
13c), which is mainly caused by rich hydroxyl with good affinity to water vapor on such high
specific area surface materials. After modification, the water CA was only 114° (Figure 13d).
These results indicate that, before and after modification, the porous silica film can
effectively improve its wettability to superhydrophilicity and superhydrophobicity,
respectively, compared with the sample without macropores. According to Wenzel model,
high roughness can enhance the hydrophilicity of hydrophilic native surface. In this case,
such porous film is much rougher than relatively flat surface obtained without using template.
So this model can explain why as-synthesized films show superhydrophilicity with a CA
about 5°.
Figure 12. Morphology of SiO2 ordered pore array film by the sol-dipping method using the colloidal
monolayer as a template. Reprinted with permission from Ref. 12, Copyright 2006 Institute of Physics.
Figure 13. Water droplet shapes and water CAs on different silica film surfaces. (a) and (b): macropore array
silica film before and after surface modification by (heptadecafluoro-1,1,2,2-tetrahydrodecyl) triethoxysilane,
respectively. (c) and (d): flat silica film without macropores before and after modification, respectively.
Reprinted with permission from Ref. 12, Copyright 2006 Institute of Physics.
Figure 14. A photograph of a water droplet on the modified ordered macropore array film shown in Figure
13b. Reprinted with permission from Ref. 12, Copyright 2006 Institute of Physics.
After modification with a lower surface free energy material, the air can be trapped in
such ordered pore arrays. The measurements of water CAs show that the surface takes on
superhydrophobicity with water CA of 154°. In this case, Cassie equation is available. Based
on a simplified model, as schematically illustrated in Figure 15, the relationship between f 2
(the area fraction of a water droplet in contact with air in macropores) and radii of the
macropores at film surface (r) and PS (R) can be described by f 2 3 r 2 . Here, r=470
6 R2
nm, R=500 nm. We can thus obtain the values f 2 =0.8013. The value, for the flat silica
surface without macropores, after modification, is experimentally about 114°. So the CA for
our modified macroporous sample can be estimated to be about 152.5° by Cassie‘s equation,
which is in good agreement with experimental result. Although the modified samples are of
superhydrophobicity, a small water droplet (5 mg) on their surface cannot roll off when the
films are tilted to any angle, even upside down. It still firmly pinned on such surface.
Generally, the three-phase (air-liquid-solid) contact line plays an important role for sliding
behavior of water droplets. The proper design of this contact line can lower the energy barrier
for droplet motion and improve its sliding on the surface. According to the morphology of
such netlike ordered pore arrays, it is easy to form the continuous, stable three-phase contact
line (Figure 16), which leads to larger the energy barrier for water droplet motion and make it
difficult to slide on surface. Such special properties have great significance on liquid
microtransport without loss in microfluidic devices. However, it is not suitable to use in
application of self-cleaning surfaces due to unable sliding behavior of water droplet on its
surface.
Figure 15. A schematic illustration of unit with a macropore. R and r are the radii of PS sphere in the
template and the macropores at film surface, respectively. Reprinted with permission from Ref. 12, Copyright
2006 Institute of Physics.
Most recently, we have found that the ordered structures with a quite high surface
roughness could be prepared based on heat-deformed colloid crystal as template [12-14]. For
example, ordered silica network with triangular prism shaped pillars on almost each node can
be fabricated by deformed colloidal monolayer which sintered at 120 C for 10min (see
Figure 17a). Before modification with fluoroalkylsilane, such film also takes on
superhydrophilicity (CA<5°). After modification, however, its water CA was increased to
165°. From Figure 17a, we know that the height of regular triangular section of the prism is
about 170nm. According to the geometric relationship, f 2 , f1 can be easily calculated with
the values of 0.9615, 0.0385, respectively. By Cassie‘s equation, the water CA of such
nanostructured film is evaluated to be 167.7°, which accords with our results. Corresponding
result of the sliding angle, 36°, further indicates such silica film with nanopillar array shows
obviously enhanced superhydrophobicity in comparison with ordered pore array film obtained
by template without sintering, which suggests that we can control the superhydrophobicity
using the heat-deformed template with different extent. Additionally, the existence of 36°
sliding angle indicates that the continuous, stable three-phase (air-liquid-solid) contact line for
the ordered pore film (Figure 16) can be broken and the energy barrier for water droplet
motion can be lowered by the such pillar array on it (Figure 17b) [15, 16], which is very
useful for improving the water droplet motion and enhancing superhydrophobicity.
Figure 16. 2D (X-Y) presentation of the silica macropore array surface and probable three-phase contact line
(dark line). Reprinted with permission from Ref. 12, Copyright 2006 Institute of Physics.
a
Figure 17. (a) SEM image of silica porous film with nanopillar array. Inset on left top in (a) is monolayer
colloid crystal sintered 120℃ for 10min, the bar is 1µm. Inset on right top in (a) is the water droplet shape
and corresponding CA (165°). (b), probable three-phase contact line (dark line) on the nanopillar array film.
Reprinted with permission from Ref. 12, Copyright 2006 Institute of Physics.
HIERARCHICAL MICRO/NANO COMPOSITE ARRAYS

Recently, fabrication techniques of monolayer colloid crystals have been well developed
because of the promising applications in optical gratings, optical filters, and antireflective
surface coatings. The monolayer colloidal crystals with large area could be easily fabricated
by self-assembling process, for example, spin coating, dip coating etc. Because colloidal
crystals that consist of hexagonally close-packed microspheres provide surfaces with a
regularly ordered and well-defined roughness, they may lead to an enhancement of the
surface hydrophobicity. Unfortunately, such surface roughness is not sufficient to induce
superhydrophobicity. Therefore, to achieve superhydrophobicity using monolayer colloidal
crystal, a much rougher surface texture should be provided on the colloidal crystals. For
instance, if we decorate zero dimensional (0D) structures (i.e. nanoparticles) or one
dimensional (1D) ones (i.e. nanotubes or nanowires) on the colloidal monolayer with
microsized PS spheres, the hierarchical micro/nanostructures like lotus leaves will be easily
obtained, which gives us a big opportunity to get a surface with self-cleaning.
Figure 18. Scheme for the fabricating process for the bionic surfaces with the hierarchical microsphere/
nanoparticle composite arrays. Reprinted with permission from Ref. 17, Copyright 2007 America Chemical
Society.
0D Nanostructures on Microsized PS Spheres [17]
Recently, we have found that silver nanoparticles can be formed homogeneously on the
substrate by thermal decomposition of silver acetate (AgAc) at a low temperature. So we tried
to prepare the silver nanoparticles on the colloidal monolayer in order to get bionic
hierarchical micro/nanostructured arrays by following experiments. The fabrication process of
the bionic surfaces is illustrated in Figure 18. The monolayer PS colloidal crystal with the
area of about 2 cm2 was prepared on a glass substrate by spin coating. Subsequently, AgAc
aqueous solution was dripped onto the colloidal crystals, forming a thin AgAc coating on the
PS colloidal crystal. The colloidal crystal with the AgAc coating was heated in an oven,
leading to the formation of comparatively uniform decoration of silver nanoparticles on the
surfaces of the PS spheres. Consequently, hierarchical structures consisting of ordered PS
microspheres and silver nanoparticles were created.
(a) (b)
Figure 19. SEM images of bionic surface fabricated using the colloidal monolayer (the diameter of PS
o
C for 3 h. (a) SEM image of the surface at a low
magnification. The insets on left and right top are the photos of water droplets before and after modification,
respectively. The water CAs were 29o (left) and 168o (right), respectively. (b) the local magnification of (a) .
Reprinted with permission from Ref. 17, Copyright 2007 America Chemical Society.
Figure 19 presents SEM images of the synthesized hierarchical micro/nano structures.

The monolayer colloidal crystal has the periodicity of 5 m and the nanoparticles on the
colloidal crystal have an average size of 180 nm. The hierarchical structure was fabricated
with a precursor solution of 0.5 M and at a heating temperature of 200 oC for 3 h, which was
the optimized experimental condition for the fabrication. The synthesized hierarchical
structure well mimicked the surface of a lotus leaf. The water CA of the as-prepared bionic
structure was measured to be 29o, showing hydrophilicity (the inset at left top in Figure 19a).
However, the wettability of the film was changed into superhydrophobicity after the chemical
modification of the surface with 1-Dodecanethiol, a kind of low surface-energy material. The
water CA was dramatically increased to 168o (the inset at right top in Figure 19a). In addition,
the modified surface exhibited a small sliding angle of about 2o. These results indicated that
the fabricated surface with the hierarchical micro/nano structure had a typical self-cleaning
property. In the process of CA measurement, we found it difficult to add a water droplet on
the fabricated bionic surface, demonstrating that the surface has very low adhesive force and
very small CA hysteresis. The fact of superhydrophobicity with a very low sliding angle and
the difficulty in dropping water on the surface provide strong evidence of the lotus effect for
the synthesized bionic surface with the hierarchical micro/nano structure.
In order to identify the origin of the superhydrophobic properties of the synthesized
structures, we prepared a uniform silver films, with 30 nm in thickness, by plasma sputtering
(Figure 20a), a silver nanoparticles‘ film on a flat substrate by the thermal decomposition of
AgAc (Figure 20 b). The flat silver film is of the water CA 68o, and shows hydrophobicity
with a water CA of 110o after surface modification (the insets of Figure 20a) by 1-
dodecanethiol. The silver nanoparticle film had a water CA of 41o, which was lower than the
flat silver film, but was enhanced to 135o after the surface modification (Figure 20b). The
silver-coated monolayer PS colloidal crystal showed the water dewetting behavior similar to
the silver nanoparticle film: the CA was 43o and increased to 129o after the surface
modification (Figure 20c). These results show that both the silver nanoparticle film and the
PS colloidal crystal do not induce superhydrophobicity and suggest that the strong
superhydrophobicity of the synthesized bionic surface originates from the hierarchically
combined micro/nano structure of the surface.
In the presented method, the distribution density of nanoparticles can be controlled by
changing the concentration of AgAc precursor. The number density of the nanoparticles
decreased with decreasing the precursor concentration. If the precursor concentration was low
(say, 0.3 M), silver nanoparticles were not completely coated on the PS spheres (Figure 21),
and some PS spheres were melted during heating because of the incomplete coating of AgAc
on the PS spheres (Figure 21b). These resulted in smaller surface roughness and
corresponding lower water CAs compared to the optimized precursor concentration. The
experimental results showed that the superhydrophobicity was obtained at the precursor
concentration of 0.35 - 0.65 M. Moreover, a well-controlled heating temperature was also
crucial for the successful synthesis of the superhydrophobic bionic surfaces. Heating
temperature below 180 oC was insufficient to convert AgAc precursor on the PS spheres to
silver nanocrystals. At a heating temperature above 220 oC, PS spheres were gradually
decomposed and the bionic structures collapsed (glass transition temperature Tg of PS is
nearly 100 oC), leading to the decrease in the CA. The experimental results showed that the
temperature in the range of 195 – 215 oC was optimum value for the formation of the bionic
surfaces with superhydrophobicity. Heating treatment has two important roles: firstly,
decomposition of AgAc coating into silver nanoparticles and secondly, enhancing the
mechanical stability of bionic surfaces. A bare monolayer colloidal crystal can be easily
peeled off from the substrate. However, the PS spheres can be slightly melted during heating
at a temperature higher than Tg of PS, leading to the PS spheres sintered tightly to the
substrate and that the silver nanoparticles embedded into PS spheres. The embedment of
silver nanoparticles in PS microspheres is clearly seen in Figure 21b. Consequently, the
heating process induces the highly durable hierarchical micro/nano bionic films: the
organized films were not detached from a substrate even when the bionic surface was
ultrasonically washed in water for 1 h.
The periodicity of the microstructures can be tuned by changing the PS sphere size in
colloidal monolayer. For example, Figure 22 is the FESEM image of the bionic structure
synthesized from the PS colloidal monolayer of 1.3 m sphere size. After the modification
with thiol, this structure also induced superhydrophobicity with a water CA of 163o and a
sliding angle of about 6o.
We have presented a facile synthetic route to bionic surfaces with remarkable
superhydrophobic and self-cleaning properties by preparing the 0D nanostructures on the PS
sphere surfaces. The bionic surfaces were synthesized by decorating silver nanocrystals
generated from thermally-decomposed AgAc on PS colloidal crystals and the subsequent
surface modification. Since the bionic surfaces consist of regularly-ordered rough structures,
one of features is that the films are of superhydrophobicity on the whole surface.
Additionally, the wettability of the bionic surface can be easily tuned by the AgAc precursor
concentration, the treatment temperature, and the PS sphere size, which is very helpful in the
microfluidic devices.
(a)
(b)
(c)
Figure 20. FESEM images and water droplets on (a) a flat silver film surface, (b) a silver nanoparticle film,
and (c) a colloid monolayer with a thin silver coating. Insets at left (right) top of the figures are the water
droplets before (after) the chemical modification by 1-dodecanethiol. The water CAs before and after the
modification were (a) 68o and 110o, (b) 41o and 135o, (c) 43o and 129o, respectively.
melted PS
(a) (b)
Figure 21. FESEM image of a bionic surface fabricated on PS monolayer after dipping the AgAc solution of
0.3 M and heating at temperature of 200 oC for 3h. (b) A local magnification of (a). Reprinted with
permission from Ref. 17, Copyright 2007 America Chemical Society.
Figure 22. FESEM image of a bionic surface fabricated using the colloidal monolayer with the PS diameter
and 0.5 M AgAc after heating at 200 oC for 3h. Reprinted with permission from Ref. 17,
1D Nanostructures on Microsized PS Spheres
Besides the hierarchical micro/nano structured arrays by decorating 0D nanostructures on

the microsized PS spheres, we also presented a facile and alternative method to create
superhydrophobic bionic surfaces with hierarchically micro/nano combined structures by
loading 1D nanostructures (carbon nanotubes) on the microsized PS spheres. The
microstructure was prepared by PS colloidal monolayer on a glass or silicon substrate and the
nanostructure was supplied by single-walled carbon nanotubes (SWCNTs) decorated on the
micro-structures by wet chemical self-assembly [18-20]. The morphology and the number
density of the nanostructure can be easily controlled by the concentration of SWCNT
solution. The presented route well exhibits the concept of bionic fabrication. The morphology
of the resultant product bears more resemblance to the natural lotus leaf, and consequently
shows strong superhydrophobicity.
The fabrication process is illustrated in Figure 23. The monolayer PS colloidal crystals
with cm2 size were prepared on glass substrates by spin coating using PS colloidal
microsphere suspension. The colloidal crystals were then heated at a temperature of 130 oC
(above than the glass transition (105 oC) of the microspheres) for 40 min, which strongly
increases the adherence of the PS crystals to the substrate [21, 22]. Subsequently, a gold layer
of 30 nm thickness was coated on the microsphere surface by plasma sputtering. The sample
was then dipped into 0.1 mol L-1 aqueous solution of mercaptoethylamine for the
funtionalization of amino group on the gold surface. SWCNTs with carboxylic acid
functionality (–COOH) solution was prepared by ultrasonating the raw CNTs (Iljin Nanotech
Co., Ltd., Korea) in the mixture of concentrated sulfuric and nitric acids with a volume ratio
of 3:1 for 6 h, followed by redispersing the SWCNTs into acetone after filtration by
sonication. When the prepared SWCNTs solution was dropped on the microsphere surface,
the SWCNTs can be self-assembled on the PS shperes due to the condensation reaction
between the –COOH and –NH2 as well as the electrostatic attraction and Van der Waals
interactions between the carbon nanotubes [23]. The SWCNTs were then decorated randomly
on the surfaces of PS microspheres with almost uniform distribution density. As a result, the
bionic surface comprising PS microspheres and SWCNT was created.
Figures 24 a-c are the SEM images of the bionic structure fabricated by 2.0 mg L-1
SWCNTs solution and the colloidal monolayer with periodicity of 5.0 m. The images
clearly show that dense SWCNTs were assembled on the microspheres surface by the wet
chemical self-assembly method described above. SWCNTs on the microspheres take on
interlaced ‗net‘ structures and they tightly adhere to the microsphere surfaces. The whole
hierarchical microsphere/ SWCNTs composite arrays exhibit hexagonally close-packed
arrangement. The SEM image of a natural lotus leaf surface was also shown for the
comparison of the morphology (Figure 24 d) [24]. The synthesized hierarchical structure well
mimicked the surface of a lotus leaf.
The wettability of the synthesized hierarchical structures was investigated by both static
water CAs and sliding angles (SAs). The water CA of the as-prepared samples with the
hierarchical structures was measured to be 33o (Figure 25a), exhibiting hydrophilicity. In
order to reduce the surface energy on the structures, the surface of the as-prepared samples
were chemically modified with fluoroalkylsilane: the samples were immersed in a hexane
solution of 20 mM 1H, 1H, 2H, 2H–perfluorodecyltrichlorosilane (Alfa Aesar) for 30 min and
subsequently dried in an oven at 50 oC for 30 min. After the chemical modification, the CA of
the sample was dramatically increased to 165o and the water droplet was nearly spherical
(Figure 25b, c). In addition, the surface exhibited a small SA of about 5o. The presence of
superhydrophobicity with a very low SA provides strong evidence of the lotus effect for the
synthesized hierarchical micro/nano structures.
Figure 23. Schematic fabrication process of bionic surface with micro/nano structures. Reprinted with
(a)
100 m
(b)
(c) (d)
1 m 1 m
Figure 24. SEM images of the bionic surface with microsphere/SWCNTs composition array (the
concentration of SWCNTs was 2.0 mg L-1). (a): low magnification. (b) and (c): the local magnification of (a).
(d): the surface microstructure of a natural lotus leaf [2]. Reprinted with permission from Ref. 18, Copyright
2007 America Chemical Society.
We systematically investigated the CAs of a monolayer PS colloidal crystal coated with

gold film and a SWCNTs film on flat substrate (Figure 26). A SWCNT film was prepared on
gold coated silicon substrate by a wet chemical self-assembly as described above. The CA of
the gold-coated colloidal monolayer was 94o and increased to 138o after the surface
modification (Figure 26a). Moreover, the SWCNT film on flat substrate exhibited
hydrophilicity with a water CA of 63o and showed hydrophobicity with the CA of 132o after
the surface modification (Figure 26b). These results indicate that the strong
superhydrophobicity of the fabricated bionic surface originates from its unique hierarchical
structure combined by the nano-scaled net structure of SWCNTs and the micro-scaled PS
spheres.
Before the surface modification, the SWCNT film on the flat surface was hydrophilic
(CA: 63o), which is attributed from –COOH groups on the CNT surfaces. Evidently,
SWCNTs on the colloidal monolayers make the surface roughness increase. According to the
Wenzel model [17a], a rough surface is more hydrophilic than a flat surface and accordingly
the hydrophilicity increases with increasing the roughness [17b], which can explain why the
water CA (33o) of the SWCNTs on the colloid monolayer is smaller than the CA (63o) of the
SWCNT film on the flat substrate. When the bionic surfaces are modified with the low
surface-energy material, the air can be trapped in grooves or interstices on the bionic surfaces.
In this case, Cassie can be used to explain our phenomenon. Since the given water CAs of the
flat graphite surface [25] and the bionic surface modified with fluoroalkylsilane are 108o and
the 165o, respectively, f2 is calculated to be 0.95. This indicates that such strong
superhydrophobicity of the bionic surface is mainly due to the air trapped in the rough surface
by the combining microstructure of the ordered PS sphere arrays and the nanostructure of the
SWCNTs adsorbed on their surface.
In this route, the SWCNT distribution can be controlled on the microsphere surfaces by
changing the concentration of SWCNT solution and corresponding wettability can also be
tuned due to the morphology change. For a typical example, by decreasing the SWCNT
concentration from 2.0 mg L-1 to 1.0 mg L-1, the corresponding number density of SWCNT
was reduced and the microsphere surfaces were not completely coated by SWCNTs, as
displayed in Figure 27. This reflects that, at lower SWCNT concentrations, a smaller quantity
of SWCNTs was assembled during wet chemical self-assembly process on the PS spheres.
Consequently, the surface roughness becomes less, resulting in lower CA of 156o after the
surface modification. The experimental results showed that the CAs decreased further with
decreasing the SWCNT concentration and that the super-hydrophobic bionic surfaces with the
CA higher than 150o were obtained at the concentration range of 2.5- 0.7 mg L-1.
(a) (b)
(c)
Figure 25. Water droplets on the fabricated bionic surface (a) before and (b) after the surface modifcation.
The corresponding CAs are 33o and 165o, respectively. (c): a photo of a water droplet on the as-prepared
bionic surface. Reprinted with permission from Ref. 18, Copyright 2007 America Chemical Society.
Figure 26. SEM images of PS sphere array with gold coating (a) and SWCNT film on the substrate (b). The
insets on the left top in (a) and (b) are the water droplets on the surface before modification (the CAs are 94o
and 63o, respectively). The insets on right in (a) and (b) are water droplets after the surface modification (the
CAs are 138o and 132o, respectively). Reprinted with permission from Ref. 18, Copyright 2007 America
Chemical Society.
Figure 27. A SEM image of the PS microsphere/SWCNTs composition array corresponding to the SWCNTs
concentration of 1.0 mg L-1. The left inset is the local magnification and the right inset shows the water
droplet on the surface. Reprinted with permission from Ref. 18, Copyright 2007 America Chemical Society.
As we know, SWCNTs are more expensive than multi-WCNTs. If we can use the
MWCNTs to replace the SWCNTs to fabricate the bionic surfaces, the costs will be largely
reduced. Therefore, we also tried to fabricate the MWCNTs on the microsized PS spheres
using the same route as that of SWCNTs. Figure 28 shows the typical morphology of the
hierarchical microsphere/MWCNT composite arrays and the water droplet shapes on the
surface of the arrays. MWCNTs combined with the monolayer colloidal crystals also
exhibited superhydrophobicity. The hierarchical structure was obtained from the MWCNT
solution with concentration of 2.0 mg/L and the PS spheres of in diameter. After
surface treatment, the surface also displayed the superhydrophobicity with a CA of 166o and a
SA of 5o.
It should be noted that, compared to lotus leaves with randomly-distributed
microstructures of non-uniform sizes, the prepared bionic surfaces consist of regularly
ordered microsphere/CNT composite arrays. Therefore, the synthezied bionic films had a
very uniform wettability on the whole surface (the deviation of the measured water CA was
around 1o on the whole fabricated surface).
This work has following advantages: (1) the synthesized bionic surfaces were very
similar to the natural lotus leaves, that consist of rugged, hierarchical micro/nano structures,
and thus exhibited strong superhydrophobicity with a low SA after the surface treatment, like
lotus leaves. (2) The wettability of the bionic surface can be well controlled by the
distribution density of CNTs on the colloidal monolayers as well as the periodicity of
colloidal monolayers, which makes them have many potential applications in the fields of
microfluidic devices, bioseparation devices, and liquid transportation without loss. (3) CNTs
with a continuous and homogeneous distribution on PS spheres have unique electrical and
electrochemical properties, and such rough surface with hierarchical microsphere/CNTs
structures has a large specific area. Thus, the fabricated bionic structures can also be used for
other devices such as gas sensors with good selectivity and high sensitivity.
Figure 28. SEM image of the microsphere/MWCNT composite array and a water droplet on the surface.
Reprinted with permission from Ref. 18, Copyright 2007 America Chemical Society.
SUPERHYDROPHOBIC SURFACES
ON THE CURVED SUBSTRATES
Recently, people have found that a water strider can float and move quickly on water
surface. Biomimetic researches reveal that this interesting feature mainly resulted from the
superhydrophobicity of the water strider‘s legs [1]. A water strider‘s leg has special
hierarchical structure consisting of microsetae with nanogrooves, leading to the
superhydrophobicity. A leg of a water strider gives a supporting force corresponding to 15
times the water strider‘s weight against water surface. This phenomenon inspires the
scientists to fabricate the similar objects, which has important applications in the miniaturized
aquatic devices operating on water or under water and lossless liquid transportation channels.
Although various techniques to synthesize superhydrophobic surfaces have been developed,
most of these techniques are restricted on flat substrates. However, in order to allow the
striking water-supporting force to an object with curved surface, all the outer surface of the
object should be enclosed with superhydrophobic materials. So far, only few methods to
fabricate superhydrophobic surfaces that can mimic the legs of water striders have been
investigated such as superhydrophobic surfaces on the metal wires by the electrochemical
deposition utilizing the property of physical chemistry of metals, which can well mimic the
legs of water strider [26]. It has been found that polymer colloidal monolayer on a substrate
can be transferred onto another substrate, while retaining its integrality [27]. Inspired by this,
we fabricated bionic superhydrophobic surfaces on a curved surface. The fabricated
superhydrophobic coating on a convex tube exhibited a strong water-repellent property and
supplied a high supporting force when it was floated on water surface.
The fabrication process of the superhydrophobic coating on a curved substrate is
illustrated in Figure 29. In previous studies, we found that polymer colloidal monolayer on a
substrate can be transferred onto another substrate [27]. We applied this technique for the
fabrication of the superhydrophobic coating on a curved surface using a precursor solution as
a medium. At first, a monolayer PS colloidal crystal is prepared on a glass substrate by spin
coating. Subsequently, the colloidal monolayer on the substrate is gradually dipped into the
silver acetate (AgAc) solution with an inclination angle of about 30o. As a result, the colloidal
monolayer is peeled off from the glass substrate and floated on the AgAc solution surface
while retaining its integrality. Then, the colloidal monolayer is slowly picked up with a glass
tube and accordingly the colloidal monolayer coated with AgAc covers the outer surface of
the tube, followed by drying at room temperature. Finally, the samples were heated at 200 oC
for 3 h in an oven. Glass tubes with two different outer diameters of 1.4 mm and 4.87mm
were used in the experiments. After the heat-treatment, AgAc is transformed into silver
nanoparticles and consequently hierarchical structures that consist of PS microsphere arrays
coated with silver nanoparticles are created. The photographs to show each process described
above are displayed in Figure 30.
Figure 29. Scheme for fabricating a superhydrophobic coating on a curved surface. (a) A PS colloidal
monolayer is fabricated on a flat substrate by self-assembling. (b) The colloidal monolayer on the substrate is
gradually dipped into the AgAc solution and then the monolayer is peeled off from the substrate. (c) The
colloid monolayer floats on the solution surface. (d) The monolayer is picked by a glass tube with curved
surface. (e) The PS colloidal monolayer with AgAc coating is formed on the curved surface. (f) After heating,
the hierarchical structure consisting of micrometer-sized PS spheres and silver nanoparticles is prepared on
the curved surface. Reprinted with permission from Ref. 17, Copyright 2007 America Chemical Society.
Figure 31a presents a typical FESEM image of the fabricated hierarchical micro/nano
structure on a convex glass substrate. Close-packed arrays of monolayer PS spheres with 5
m diameter completely covered the glass substrates and nanoparticles with an average size
of 180 nm uniformly decorated on the PS spheres. Using the same strategy, such a
hierarchical micro/nano structure was also fabricated on a concave surface of the inner wall of
a glass tube (Figure 31b). These hierarchical micro/nano structured arrays exhibited a strong
superhydrophobicity with water CA of 168o and a small SA of about 2o after modification
with low free energy materials. The fact of superhydrophobicity with a very low SA and the
difficulty in dropping water on the surface provide strong evidence of the lotus effect for the
synthesized surface with the hierarchical micro/nano structure.
In order to demonstrate the water-repellent property of the superhydrophobic coating, we
prepared such a coating on a glass tube and measured the supporting force of the tube (Figure
32). Both ends of the tube were sealed to prevent water from permeating into the inside of the
tube. The outer diameter, the inner diameter, and the length of the tube were 4.87 mm, 3.05
mm, and 13.15 mm, respectively. The weight of the tube was 332 dynes, which is about 300
times heavier than a water strider (an adult Gerris remigis ) [28].
(a) (b)
(d) (c)
Figure 30. Photographs corresponding to the manipulation process described in Figure 29. Reprinted with
2 m
Figure 31. SEM images of the hierarchical structure on (a) an outer surface of a glass tube (outer diameter:
1.4mm) and (b) inner surface of glass tube (inner diameter: 3.0mm ). The insets are the locally magnified
images. Reprinted with permission from Ref. 17, Copyright 2007 America Chemical Society.
a b
c d
Side view
Figure 32. (a) Photographs of a bare glass tube (transparent one) and a glass tube with the superhydrophobic
coating, respectively. (b) The behavior of the two tubes when slowly put on water suarface. (c) The glass tube
with the superhydrophobic coating on water surface. Deformed meniscus is clearly seen on the water surface
near the tube. (d) The glass tube with the superhydrophobic coating weighted with plastic beads. Reprinted
with permission from Ref. 17, Copyright 2007 America Chemical Society.
Surprisingly, when a glass tube with the superhydrophobic coating was dipped into water
and then taken out, no water droplets were found on the tube, exhibiting that the coating has
an excellent water-repellent superhydrophobic property. As displayed in Figure 32b, a bare
glass tube sank in the water and finally on the bottom of vessel. However, the glass tube
covered with the hierarchical structure can easily float on the water surface. Moreover,
deformed meniscus is clearly seen on the water surface near the tube (Figure 32c), reflecting
that strong supporting force was produced by the superhydrophobic coating on the tube. For
measuring the maximum supporting force, the glass tube was weighted with plastic beads
through a thin plastic wire that was fixed at the middle of glass tube until the glass tube sank
to the bottom of vessel (Figure 32 d).
The maximum supporting force is obtained when mg = Fb + Fc. Here, m is the total mass
of the glass tube, the plastic beads, and the thin wire that make the weighted tube sink to the
water bottom. g is the gravitation acceleration speed. The total weight mg is supported by
combination of two forces, the buoyancy force (Fb) and the curvature force (Fc) [26, 28]. Fb
can be determined by integrating the hydrostatic pressure over body area in contact with
water. Fc is associated with the surface tension and thus this is more important factor
characterizing the non-wetting property of the superhydrophobic coating than Fb. The total
weight mg was measured to be 1420 dynes, which is around 4.3 times the weight of the glass
tube. In addition, the maximal curvature force was calculated to be 386 dynes, which is 4.5
times larger than that of a leg of water strider (an adult Gerris remigis ) reported by Lu and
his co-workers [28]. The results show that a very high supporting force was produced by the
fabricated superhydrophobic coating, which can effectively capture the air in the interstices
among the micros/nano hierarchical structure.
CONCLUSION
Biomimetic research reveals that a self-cleaning effect of a lotus leaf is ascribed to the
combination of both a hierarchical micro-/nanostructure on the surface and a low surface-
energy material covering the surface. According to this phenomenon, one can find that it is
necessary to meet two conditions to create the superhydrophobic surfaces. One is large
enough roughness factor on the surface and the other is a coating of low surface free energy
on the surface. Inspired by the lotus effect, a lot of techniques to prepared bionic
superhydrophobic surfaces have been developed. Due to well development of the synthesis of
colloidal crystals and micro/nano structured arrays using on the colloidal crystals as
templates, a lot of order micro- or nanostructured arrays could be prepared using the
colloidal monolayer templates. These ordered arrays and the colloidal crystals are rough on
the surfaces in the micro- or nanoscale, which gives a good chance to create the
superhydrophobicity on the sample surfaces. In this chapter, we review the superhydrophobic
surfaces based on the colloidal crystal techniques. From these results, one can clearly see that
hierarchical micro/nano structures are very important in realizing the superhydrophobicity
with self-cleaning effect. This can be well proven by the examples of silver hierarchical ring-
like arrays, hierarchical PS microsphere/silver nanoparticle composite arrays and hierarchical
PS microsphere/CNT composite arrays, which exhibited the very large water CAs and very
small SAs. Importantly, the technique of fabricating superhydrophobic surfaces on the curved
substrates has important applications in the miniaturized aquatic devices operating on water
or under water and lossless liquid transportation channels. As we know, the micro- or
nanostructured arrays by colloidal monolayer templates have important applications in
photonics, photoelectronic devices etc. This suggests that nanodevices built from these
nanostructured arrays could be waterproof and self-cleaning in addition to their special device
functions after possessing the superhydrophobicity. However, the as prepared
superhydrophobic surfaces with self-cleaning effect still can not be used in the practical
applications (for example, the coating on the building) due to the high costs, easily being
contaminated. Besides these facts, another big problem is that the surfaces are easily damaged
because of the low mechanical stability with increase of surface roughness, which also
prevents from the normal applications. Therefore, the design and preparation of bionic self-
cleaning superhydrophobic surfaces with low costs and high mechanical stability will be an
important and challenging task based on the colloidal crystal techniques. We also believe that
more attention will be paid in these problems and they would be resolved in the near future.
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Lecture Material 17
NANOMATERIALS:
BIOPOLYMER-ASSISTED GREEN SYNTHESIS
ABSTRACT
Nanomaterial science and technology have generated great enthusiasm in recent
years because these novel technologies are guaranteed to have an impact on the energy,
chemical, electronic, and aerospace industries. Researchers in various fields of chemistry
have been studying novel methods through which the morphology and the dimensions of
nanomaterials can be controlled at the micro- or even the nanoscopic level. As far as the
synthesis of nanoparticles is concerned, there is an ever-growing need to develop clean,
non-toxic, and environmentally friendly (― green chemistry‖) synthetic procedures in the
pursuit of nanotechnology, especially for nanoproducts targeted at bioapplications. The
selection of an environmentally acceptable solvent system, an eco-friendly reducing
agent and a nonhazardous capping agent for the stabilization of the nanoparticles are
three criteria for a totally green nanoparticle synthesis. Thus, during the last few years,
―g reen‖ has become a common term to designate those nanomaterials with the aim of
replacing nondegradable and toxic regents, thereby reducing the environmental pollution.
Another reason that is propelling the green chemistry is the great profusion and
availability of environmentally friendly biopolymers in Nature. These (together with
purely academic curiosity) have led to the development of bioinspired approaches for the
growth of advanced materials. Parallelly, when considering synthetic routes towards the
growth of inorganic nanocrystals and subsequent organization into hierarchical and
functional systems, one approach that is proving successful in addressing these problems
is the use of biopolymers as templates, scaffolds, and interconnects. The structure-
directing effect of such organic materials and the presence of functional groups provide
site-specific crystal growth, thus rendering crystallite dimensions on the nanometer-scale,
porosity, and three-dimensional ordering to the inorganic products. Therefore, this is not
surprising given that some of biopolymers act as fascinating templates for creating
inorganic materials. Herein, we focus our attention on the biopolymer-assisted green
synthesis of nanomaterials in this chapter, which covers our recent interesting results and
mainly includes (1) nanomaterials obtained by biopolymer-assisted green method and (2)
their properties.
INTRODUCTION
Background of Green Synthesis
The field of nanotechnology spans the synthesis of nanoscale matters,

understanding/utilizing their exotic physicochemical and optoelectronic properties, and
organization of nanoscale structures into predefined superstructures. It has witnessed
impressive advances in various aspects and promises to play an increasingly important role in
many key technologies of the new millennium [1]. Over the past decade, researchers in
various fields of chemistry have been studying novel methods through which the morphology
and the dimensions of inorganic materials can be controlled at the micro- or even the
nanoscopic level. As far as the synthesis of nanoparticles is concerned, there is an ever-
growing need to develop clean, non-toxic, and environmentally friendly (― green chemistry‖)
synthetic procedures in the pursuit of nanotechnology, especially for nanoproducts targeted at
bioapplications [2]. The selection of an environmentally acceptable solvent system, an eco-
friendly reducing agent and a nonhazardous capping agent for the stabilization of the
nanoparticles are three criteria for a totally green nanoparticle synthesis. Thus, during the last
few years, ―gr een‖ has become a common term to designate those nanomaterials with the aim
of replacing nondegradable and toxic regents, thereby reducing the environmental pollution
[3]. Another reason that is propelling the green chemistry is the great profusion and
availability of environmentally friendly biopolymers in Nature [4].
Nature is the source of a wide number of biomacromolecules that can be involved in the
preparation of nanomaterials [5]. There is no dearth of examples in both the plant and animal
kingdom that produce inorganic materials [6]. Some of the more well-known examples
include magnetotactic bacteria (which synthesize magnetite nanoparticles) [7], diatoms
(which synthesize siliceous materials) [8], and S-layer bacteria (which produce gypsum and
calcium carbonate layers) [9]. Thus, the use of naturally occurring biomolecules in the
synthesis of nanoparticles can meet the above criteria for ―gr een‖ nanoparticle synthesis
because it will help to reduce the amount of waste products and to diminish environmental
pollution, leading to sustainable development [10]. This (together with purely academic
curiosity) has led to the development of bioinspired approaches for the growth of advanced
materials [11]. For example, with the use of DNA, polylysine, or cytochrome c3, several
nanowires have been obtained [12], and due to its cylindrical and double helical structure,
DNA also has been confirmed to be useful in the assembly of nanoparticles to two-
dimensional (2D) or 3D [13] and in the alignment of discrete 1D nanomaterials [14]. By
following this strategy, different research teams have also developed biomaterials that mimic
the exceptional features of natural nanocomposites [15]. For example, Deville et al. have
replicated the nacre structure by mimicking a process that takes place in nature, giving rise to
tough and ultra-lightweight materials [16].
Advantages of Biopolymer
Nanomaterial science and technology have generated great enthusiasm in recent years
because these novel technologies are guaranteed to have an impact on the energy, chemical,
electronic, and aerospace industries [17]. Just as addressed above, there is a growing need to
develop environmentally benign nanoparticle synthesis processes that do not use toxic
chemicals in the synthesis protocol. Nature is adept at greenly producing remarkable
structures, optimised in form and property for their designated function [18]. As a result,
researchers in the field of nanoparticle synthesis and assembly have turned to nature, in the
respect that nature has ingeniously succeeded in giving rise to impressive varieties of
biological species with particular functional structures. Parallelly, when considering synthetic
routes towards the growth of inorganic nanocrystals and subsequent organization into
hierarchical and functional systems, one approach that is proving successful in addressing
these problems is the use of biopolymers as templates, scaffolds, and interconnects [13b, 19].
The structure-directing effect of such organic materials and the presence of functional groups
provide site-specific crystal growth, thus rendering crystallite dimensions on the nanometer-
scale, porosity, and 3D ordering to the inorganic products. Therefore, this is not surprising
given that some of biopolymers act as fascinating templates for creating inorganic materials
[20]. For example, starch, one of the most abundant polysaccharides stored in plants, meets
all of these requirements. It has also been confirmed that the structure of starch is chain-
shaped with many hydroxyl groups (–OH) on the surface and can be adopted as morphology-
directing agent to the nanomaterials and nanocomposites syntheses [21]. In addition, in
aqueous solution at high temperature, agarose (a polysaccharide) behaves as a semi-flexible
polymer. On cooling it forms double helices that bundle together giving rise to fibrils,
resulting in a water filled gel. This network structure and the ability to control the pore sizes
within the gel by altering the agarose concentration have made it widely applicable as
chromatographic and electrophoretic media [22], and as stabilizers and thickeners in food
preparations and pharmaceutical products [23]. It was also shown that agarose could be used
as a template for the fabrication of porous inorganic structures [24]. The in situ reduction of
gold in a gelling agent has been reported, where the urea derivatised amphiphiles acted both
as reductant and gelling agent in aqueous tetrachloroauric acid solutions [25]. Wang et al.
found that a mixture of tetrachloroauric acid solution and guanine with a molar ratio of 50:1
could produce submicrometre scale gold plates with nanometre thickness [26]. The similar
result was obtained by reacting an aqueous solution of tetrachloroauric acid with serum
albumin protein (BSA) at the physiological temperature, where BSA provided the dual
function of Au(III) reduction and directing the anisotropic growth of Au(0) into plate-like
structures [27]. Epoxide–iron gels have been used for the production of iron oxide magnetic
mesoporous architectures (aerogels and xerogels) endowed with the monolithic characteristics
of the gels [28]. Cellulose acetate membranes have been utilized for the production of porous
titanium and zirconium dioxide [29]. Commercial [30] and natural [31] cellulose fibers
resulted in the biomimetic synthesis of magnetite and maghemite fibers, and 3D pollen grains
for the bioreplication of silica, calcite, brushite, and monetite [32]. Aqueous solutions of
sodium carboxymethyl cellulose (NaCMC) were used for the morphosynthesis of
biopolymeric gels with spherical (capsules) or wirelike architectures after the ionotropic
gelation with ferric cations [33]. This method has been extended recently to prepare evenly-
distributed Au/TiO2 nano-hybrid [34].
In all, natural biopolymers are selected as the attractive candidates for the green synthesis
of nanomaterials among the biomolecules because of their following outstanding features.
Firstly, their benign water-solubility makes synthesis processing in water medium. This is
very crucial for ―gr een chemistry‖. Secondly, biopolymers belong to a class of materials that
can swell largely in water other than organic solvents and maintain their 3D network structure
on a nanometer length scale in the swollen state. This makes them have many potential
applications including high-surface-area supports for catalysts, separation media, and
templates for the synthesis of other nanoscopic materials, which undoubtedly motivates the
preparation of nanostructural materials inside biopolymers [35]. In addition, the extensive
number of functional groups, such as carboxyl groups and hydroxyl groups can anchor to the
surfaces of the materials and stabilize them. The third concern is the reducing properties of
biopolymers. Reducing agents, such as hydrazine, sodium borohydride (NaBH4), and
dimethyl formamide (DMF) that are used in most systems are highly reactive chemicals and
pose potential environmental and biological risks [2]. Most of biopolymers contain abundant
reductive hydroxyl groups, which resemble the behaviors of glucose[2, 36] and are exploited
for in situ reduction of metal ions and nucleation of metallic clusters. Kept to the three main
steps in the preparation of materials evaluated from a green chemistry perspective, which are
the choices of the solvent medium used for the synthesis, an environmentally benign reducing
agent, and a nontoxic stabilizing agent [2], the present synthetic procedure tallies well with
green chemistry. Fourthly, the most important feature of biopolymers is that their topologies
may experience distinct change under experiment conditions. The reduction reaction of the
starting materials and the assembly process of inorganic nuclei in and by the biopolymer sols
may be expected to be accompanied by the formation of nanoscopic morphologies other than
those are accessible by classical routes. Finally, the use of biopolyers doesn‘t require stringent
conditions, so could be readily extended to various systems. So, the selection of biopolymers
for the fabrication of nanomaterials should have significant benefits over existing
technologies because the method is facile, environmentally benign, and amenable to
processing.
Thus, biological processes have recently been considered as possible methods for the
synthesis of nanoparticles, especially the development of ―gr een‖ synthetic approaches.
Important requirements for the green applications of biopolymers are that they should possess
the ability to guide the oriented growth of organic or inorganic substances, and should be
thermally and chemically stable, easy to obtain, cheap, and, most importantly,
environmentally friendly. Combined with traditional chemical techniques, this biopolymer-
assisted synthesis method would have proven promising in the generation of a large variety of
inorganic structures that are currently unattainable through any other methods [19e]. Herein,
we focus our attention on the biopolymer-assisted green synthesis of nanomaterials in this
chapter, which covers our recent interesting results [37] and mainly includes (1)
nanomaterials obtained by biopolymer-assisted green method and (2) their properties.
Other significant examples using biopolymers as templates involve the formation of low-
dimensional inorganic nanostructures [13b], and helical arrays of nanoparticles on lipid
nanotubes [38] and nanoribbons [39].
NANOMATERIALS OBTAINED BY
BIOPOLYMER-ASSISTED GREEN METHOD
Noble Metal Nanomaterials
3D Noble Metal Sponges

Noble metals-containing porous frameworks have been used extensively in catalysis,
electrochemistry, heat dissipation, biofiltration, biosensor technology, as transport media, and
as porous electrodes [40]. Thus, different techniques for the production of various porous
materials have been developed under different conditions [41]. Generally speaking, porous
metals can be prepared following three procedures. First is dealloying process, which
involves the selective dissolution of a specific metal from a metal alloy [41a, 42]. For
example, porous gold can be prepared by dealloying a silver–gold alloy—the silver phase is
dissolved using nitric acid, leaving the gold phase intact [43]. Second is electrochemical
process in the presence of templates (soft or hard). Ordered macroporous gold and platinum
films, for example, have been produced by electrochemical reduction of gold or platinum
complexes dissolved in aqueous solution within the interstitial spaces of a polystyrene
colloidal array [44]. Using this method, mesoporous metals including Pt, Sn, Se, Co, Pd, Ni,
Te, Rh, Pt–Pd, Pt–Ru, and Cd–Te have also been successfully fabricated [45]. Third approach
for porous materials is by (thermal reduction of metal-ion-impregnated porous supports and
simultaneous) template-sacrificed route. This route involves first soaking a porous template in
a colloidal metal sol or metal salt solution to load the template with the metal or its soluble
precursor, porous metal structures can then be obtained by calcinating the organic phase or,
particularly in the case of inorganic templates, by dissolving away the original porous
materials. For example, echinoid (sea urchin) skeletal plates were immersed in gold paint and
a continuous coating of gold was deposited over the whole surface area [46]. Dissolution of
the original calcium carbonate support in acid solution produced a porous structure with 15
mm channels. Mann and coworkers have demonstrated the fabrications of macroporous
frameworks of silver, gold and copper oxide, as well as composites of silver/copper oxide or
silver/titania by heating metal-salt-containing pastes of the polysaccharide, dextran, to
temperatures between 500 and 900 °C [47]. Recently, Zhang and Cooper modified this
approach, by using emulsion-templated polymers as scaffolds, for the production of
macroporous materials from nanoparticulate building blocks [48]. In addition, Yamada and
coworkers recently prepared nanoporous films with different particle sizes and agglomerated
states by two-step strategy, i. e. preparing colloidal solutions and subsequent salting-out the
colloidal solutions with salts [49]. Although these various synthetic approaches have been
successfully used to fabricate many porous metals, the preparations of precursors for the
dealloying process and the prerequisite interstitial spaces for the electrochemical process
make these two methods restricted. Comparatively, the third route is more advisable and
practical. Herein we modified the third route by introducing biopolymer, HAPS (hyaluronic
acid potassium salt), for the production of macroporous frameworks of gold [37a]. FESEM
image (Figures 1A and 1B) revealed that the obtained materials consisted of an
interconnected framework of gold filaments, which were approximately 0.6 μm in width and
composed of fused micrometre-sized particles that enclosed pores 1-4 μm in size. The above
method can also be extended to the fabrication of silver sponges with open framework
structures by transferring an HAPS / AgNO3 aqueous solution into a stainless steel autoclave
with a Teflon liner and heating in an oven. Figures 1D and 1E showed that metallic silver
sponges were composed of fused crystallites, 200-400 nm in diameter that enclosed pores 0.4-
2 μm in size.
A B
C D
Figure 1. FESEM image (A) and a magnification version (B) of gold sponges on ITO substrate with the
initial molar ratio 1:1 of HAPS to Au(III) for a reaction time of 6 h at 1800C. (C) and (D) are the FESEM
micrograph and a magnification version of silver sponges with the initial molar ratio 1:1 of HAPS to Ag(I)
for a reaction time of 24 h at 1800C. Reprinted with permission from Ref. 37a, Copyright 2005 Institute of
Physics.
2D Single Crystalline Gold Disks

Gold nanostructures have long been the focus of much scientific research due to their
potential use in photocrystals, plasmonic waveguides, chemical or biological sensors, optical
filters, and drug delivery, and therefore, considerable attention from both fundamental and
applied research has been paid to the synthesis and characterization of gold nanomaterials
[50]. On the other hand, it has been well demonstrated that the physical and chemical
properties of the nanometer-sized materials is closely related to their size and shape [51]. As a
result, there has been an increasing interest in exploiting new methods for fabricating shape-
controlled nanoparticles, and many reviews are now available [52]. Among the various
morphologies of Au nanostructures, the planar gold nanostructures may hold promise for
scanning tunneling microscopy (STM) substrates [53] and can be processed to be
nanostructural building blocks such as nanogears. Although the exciting development in
fabricating planar Au nanostructures, it is still a challenge for materials scientists to
synthesize planar gold nanostructures with a preferential growth direction along the (111)
plane. In order to solve this problem, we developed a facile hydrothermal synthetic strategy
for the fabrication of single-crystalline Au disks growing preferentially along the (111) plane
by combining the stabilizing and reducing properties of biopolymer, sodium alginate, and the
advantages of hydrothermal process [37d].
The overall morphology of the sample is shown in Figure 2A, which indicates that the
sample is composed of a large quantity of hexagonal nanodisks. This is also verified by the
magnification image (Figure 2B). The interesting feature is that the hexagonal nanodisks form
2D arrays on the ITO substrate. This is possibly ascribed to two reasons. One is the
interactive force between the hydrophilic surface of the ITO substrate and the hydrophilically
passivated gold nanocrystals. The other is the lowest energy and most stable state resulting
from the lying state. The XRD pattern of the product is compiled in Figure 2C. The reflection
peaks can be indexed to face-centered cubic (fcc) structure of metallic gold with space group
Fm-3m (JPCDS file, card no. 65-2870) and substantiate the formation of crystalline gold [54].
The outstanding aspect features the extremely strong reflection peaks of (111) and (222)
planes and other negligible peaks. Moreover, the ratio of the intensity between the (200)
(shown in inset for clarity) and (111) diffraction peaks is far lower than that given in the
standard file (JCPDS; 0.47 versus 1). These observations also confirm that the nanodisks are
primarily dominated by (111) facet and have high purity in crystalline phase.
The three admitted experimental facts are as follows: (1) Adding NaBH4 to the colorless
supernatant after the termination of reaction gives no gold particles, which indicates that
HAuCl4 in the mixture is completely depleted by sodium alginate. (2) The reduction of gold
by NaBH4 at atmospheric pressure instead of hydrothermal condition produced gold
nanoparticles instead of nanodisks under otherwise identical conditions. (3) The products also
show nonplanar morphology when the same reaction system was protected by N2 gas. These
three pieces of evidence accompanying with the extremely strong reflection peaks of (111)
and (222) planes in XRD patterns indicate that this biopolymer-assisted hydrothermal process
is a high-yield approach for the preparation of large gold nanodisks with a preferential growth
direction along the Au (111) plane.
C
140000 1800
200
111
Intensity / Counts
1600
Intensity / Counts
1400
120000
220
1200
1000
311
100000 800
600
400
80000 200
40 50 60 70 80
60000 Two Theta / degree
40000
222
20000
0
40 60 80 100 120 140
Two Theta / degree
Figure 2. FESEM image (A), a magnification version (B), and XRD pattern (C) of gold nanodisks. Reprinted
with permission from Ref. 37d, Copyright 2008 Bentham Science.
A Sodium Alginate Nucleation New Nucleation

+ HAuCl4 Nanodisks
Reduction Reduction
Nucleation, Reduction
Nanodisks Nanodisks
+Nanoparticles Rearrange
Scheme 1. Simple representation of disk growth inside sodium alginate sol. (A) a simplified course. (B)
cross-linking of sodium alginate via Au(III). (C) nucleation resulting from the reduction of sodium alginate
and subsequent appearance of elongated particle shapes because of steric hindrance in the cross-linking
system. (D) reorganizing of the cross-linking structure and forming of classical multiply twinned particles, i.
e. hexagonal nanodisks, when the constraint exerted by particles onto the cross-linking system is too high.
The sodium alginate chains are omitted. Reprinted with permission from Ref. 37d, Copyright 2008 Bentham
Science.
What mechanism is operative here? Generally speaking, it is believed that the growth of
gold disks should be completed by two steps, i.e. nucleation and growth, respectively, which
is illustrated in Scheme 1. According to the colloidal particle nucleation and growth model
proposed by La Mer and Dinegar, nucleation occurs when ion concentration reaches super-
saturation concentration [55]. This condition is likely to be met when gold precursor is
directly mixed with the aqueous solution of sodium alginate. All metallic ions are in sodium
alginate sol and thus available for nucleation to occur. Hydrothermal treatment makes the
initial nucleation processes very quickly and causes that ―t ransversal‖ particle size is larger
than that of the cross-linking domains in the sol. Thus the growth is slowed by the cross-
linking systems and the transversal Au(III) flux is limited, which results in elongated particle
shapes. When the constraint exerted by particles onto the cross-linking structure in the sol is
too high, restructuring of sodium alginate sol occurs. Consequently, classical multiply
twinned particles such as hexagonal shape grow. The remnant Au(III) ions form new
nucleation site and follow the same course to form hexagonal gold nanodisks. These
observations in turn exhibit the effect of reaction time on the product morphology.
It is well-known that the formation of anisotropic nanostructures in the solution phase is
normally related to the following two factors: One is surfactant-based soft templates, which
can induce the preferential growth direction of the nanocrystal [56]. The other is selective
adsorption of small molecules or polymers on certain crystal planes, which can control or
confine the growth rate of the nanocrystals along a certain direction [51b, 57]. As we know, it
is impossible for sodium alginate to form a disklike soft template. Thus, we believe that the
formation of disklike Au structures is associated with the selective adsorption of the sodium
alginate on the originally preformed ultrafine Au nanocrystals.
No external reductant was needed for Au(0) production in the present method, suggesting
that sodium alginate itself is responsible for the reduction of Au(III). The hydroxyl groups in
the repeating units of sodium alginate is hypothesized to be oxidized into carboxyl groups
when Au(III) are reduced to Au(0), which can be referred to the literature [58]. Esumi et al.
also proposed that the reduction of Au(III) ions occurs through oxidation of alcohol groups
into carbonyl ones [59]. In fact, the spontaneous reduction of Au(III) to Au(0), that is, without
addition of an external reducing agent, has also been reported in other media [55, 60].
Semiconductor Nanomaterials
ZnO-based Hollow Microspheres

Zinc oxide (ZnO), an important II-VI semiconductor with a direct wide band gap (3.37
eV) and a large exciton binding energy (60 meV), has become one of the key technological
materials and of considerable interest due to its wide applications ranging from surface
acoustic wave filters, photovoltaic and optoelectronic devices, sensors, varistors,
(photo)catalysis, light-emitting diodes, photodetectors, optical modulator waveguides, and
solar cells, together with its biosafety and biocompatibility [61]. Therefore, we have
witnessed growing efforts toward synthesizing various ZnO nanostructures [62] such as
nanobelts, nanowires, nanotubes, nanoribbons, nanopins, nanocables, nanorods, and
nanoneedles over the past decades. The aesthetic morphologies, wide band gap, and strong
exciton binding energy have triggered great interest in ZnO-based nanoscience and
nanotechnology, thus making great progress in developing various routes (including vapor-
liquid-solid growth, thermal evaporation, thermal decomposition, electrochemical deposition,
and solution-phase processes) to tailor the morphology and size for the purpose of obtaining
better properties or for applying them to practical use [61e, 62a, 63]. Of these methods, the
facile solution procedures may be the most simple and effective way to prepare well-
crystallized materials at a relatively low temperature. Moreover, the advantages of solution-
based methods have also involved the remarkable influence of organic additives on the size
and morphology of the final products [64].
Many of the previously reported synthetic methods are limited to the formation of the
preferred 1D ZnO nanostructure due to its highly anisotropic growth rate along the c axis.
Their self-assembly into 2D and 3D ordered superstructures are urgently needed to meet the
demand for exploring the potentials of ZnO [64]. In fact, different 1D, 2D, and 3D
nanostructures have been fabricated by several self-assembly processes based on different
driving mechanisms [62j, 64, 65]. However, compared with this great success in the spatial
orientation of nanocrystals, little attention has been devoted to the controlled organization of
primary building units into 3D curved nanostructures. Furthermore, fine control of curved
nanostructures is relatively difficult due to the lack of understanding of the formation
mechanism, which hinders the development of secondary organization of subunits. From this
point of view, more investigation of the fabrication methods and systematic research on the
formation mechanism are highly desired for controlling the growth of complex
nanostructures. Based upon the situation mentioned above and the significance of green
synthesis, we developed a low-temperature and environmentally benign solution-phase
approach to fabricate 3D ZnO-based hollow microspheres in the presence of water-soluble
biopolymer, sodium alginate [37b].
50 m
1 m
1 m
Figure 3. FESEM
3. FESEM
Figure images ofimages
the ZnOofmicrospheres:
the ZnO microspheres: (A) low-magnification
(A) low-magnification image, (B) high-
image, (B) high-magnification
image and (C) the central
magnification imagepartand
of the microspheres.
(C) the centralReprinted withmicrospheres.
part of the permission fromReprinted
Ref. 37b, Copyright 2006
with permission
American
fromChemical
Ref. 37b,Society.
Copyright 2006 American Chemical Society.
Low-magnification FESEM observations show that the panoramic morphology is mainly
composed of uniform, spheric architectures ranging from 3 μm to 5 μm in diameter (Figure
3A). Closer observations (Figure 3B) show that the surface of the architecture is not smooth.
Furthermore, there might be some pores presented on such surface. The examination of an
intermediate (Figure 3C) vividly reveal that the structure of these spheric architectures is built
from a single layer of radially oriented nanorods with average diameter of ca.100 nm and
length of 1-1.5 μm, self-wrapping to form hollow interiors with 3 μm to 5 μm in outer
diameter. It is worth noting that all the constituent nanorods are radially aligned with their
growth axes perpendicular to the surface of the microspheres without any substrate support.
B
A
20 μm 5μm
C D
1 μm 2μm
E
10 m
Figure 4. FESEM images of samples obtained from control experiments: (A) in the
Figure 4. FESEM images of samples obtained from control experiments: (A) in the absence of SA under
absence of SA under other identical conditions; (B) 5 times the dosage of sodium alginate
other identical conditions; (B) 5 times the dosage of sodium alginate under other identical conditions; (C)
under other identical conditions; (C) decreasing the pH value from 10.38 to 9.96;
decreasing the pH value from 10.38 to 9.96; increasing the pH value from 10.38 to 10.48 (D) and 10.54 (E).
increasing the pH value from 10.38 to 10.48 (D) and 10.54 (E). Reprinted with permission
Reprinted with permission from Ref. 37b, Copyright 2006 American Chemical Society.
from Ref. 37b, Copyright 2006 American Chemical Society.
As a polar crystal with hexagonal phase, ZnO is highly anisotropic and tends to grow
along the c axis [66]. Under certain kinetic conditions, metastable 1D ZnO nanostructures can
self-assemble into hierarchical structures in liquid media. The kinetic conditions could be
reaction temperature, pH value of liquid media, and organic additives. We selected sodium
alginate as organic additives due to large amount of carboxylic groups within its skeletal
framework (the pKa values of SA are 3.5 for guluronic acid and 4.0 for mannuronic acid)
under alkaline conditions after adding ammonia into the reaction system. For comparison, we
also carried out hydrothermal experiment without using sodium alginate as additives. Figure
4A shows the FESEM image of the products obtained without sodium alginate. The inset
clearly shows that ZnO nanorods are rooted in one center and radially extended toward
outside. As a result, there is no interior hole. On the other hand, when 5 times dosage of
sodium alginate was used, another kind of hierarchical hemispheric superstructure self-
assembly from ZnO nanorods was observed (Figure 4B and the inset). These control
experiments demonstrate that the presence of an appropriate amount of water-soluble sodium
alginate is crucial for the formation of hollow microspheres.
Scheme 2. Schematic illustration of the proposed formation mechanism for the as-obtained hollow
microsphere superstructures under typical synthetic conditions. (A) the electrostatic attraction between the
positively charged Zn2+-water complex and the carboxylic groups of the negatively charged sodium alginate;
(B) sodium alginate stabilized ZnO colloidal nanoclusters; (C) formation of secondary spherical particles; (D)
coagulation of secondary spherical particles into large particles; (E) formation of nanorod-based shells at the
cost of the small nanoclusters in the system. Reprinted with permission from Ref. 37a, Copyright 2006
American Chemical Society.
In classical colloid chemistry, self-assembly of colloidal particle into secondary blocks

with particular micrometer size and special structure and corresponding mechanism are well
known, which is based on particle aggregation [62j, 65, 67]. In this case, sodium alginate is
dispersed within liquid medium mainly as random coil sol bearing abundant negatively
charged carboxylic groups. These random coils behave as organic matrixes binding many
zinc cations. In Scheme 2, a possible mechanism is proposed. There is electrostatic attraction
between the positively charged Zn2+-water complex and the negatively charged carboxylic
groups of sodium alginate. The introduction of ammonia changed the pH value and caused
Zn2+ ions to hydrolyze to form precursor, which was still attached to sodium alginate. Upon
hydrothermal treatment, the precursor decomposed and ZnO could be formed. With the
process of reaction, more ZnO would be produced and may still attach to sodium alginate
chains and aggregate into colloidal particles in order to minimize their surface energy. The
spherical aggregates may further coagulate to form large particles [68]. Driven by the natural
preference of polar crystal growth, ZnO tends to grow towards the exterior of the large
particles since randomly oriented growth is physically limited. This is accompanied by the
energetically favorable self-assembly of tiny rods and Ostwald ripening with intermediary
phases transfer from smaller nanocluster particles within the reaction system. As a result,
ordered nanorod-based mesoporous shells are constructed [69].
Hierarchical CuO Hollow Micro/Nanostructures

Over the past few decades, the synthesis of hierarchical micro/nanostructures with well-
defined morphologies has progressively attracted considerable attention from both
fundamental research and technological aspects [70]. The physicochemical nature of their
organic and inorganic components and the synergy between these components endow them
with higher functionality and performance [71]. Thus, design and construction of hierarchical
structures facilitate tuning materials properties through tailoring the kinds and accessibility of
functional components, curvature of interfaces, and style and degree of the internal
organizations. None of these parameters can be manipulated by traditional material
engineering in a single length scale [72]. As a result, the ever-increasing research interest has
been generated in developing a variety of approaches to construct hierarchical structures for
the development of new functional materials [72, 73]. Up to now, many methods have been
used to prepare complex hierarchical micro/nanostructures, such as hydrothermal methods
[74], thermal reduction and oxidation process [75], oriented aggregation or self-assembly of
building blocks [76], and template-assisted synthesis [77].
As an important p-type transition-metal-oxide semiconductor with a narrow band gap (Eg
= 1.2 eV), copper oxide (CuO) has been widely exploited for a versatile range of applications
such as superconductor [78], gas sensing [79], heterogeneous catalysis [80], magnetic storage
media [81], field-emission sources [82], solar cell devices [83], lithium ion electrode materials
[84], construction of a variety of organic-inorganic nanocomposites with unique
characteristics [85], and so on. Among all these potential applications, the use of CuO as
electrode materials for next generation rechargeable lithium-ion batteries have been
intensively studied because of their high theoretical capacity, high safety, environmental
benignity, low cost, etc [84c]. One of the bottlenecks restricting it from application in lithium-
ion batteries is the large volume variation during the lithium uptake/release process, which
leads to severe mechanical strains and very rapid capacity decay. In this context, a great effort
has been made to use nanoarchitectured electrodes for overcoming such drawback and
improving the electrochemical performance [86]. Hitherto, various methods, such as
hydrothermal method [87], sol-gel technique [88], gas-phase oxidation [75], micro-emulsion
[89], thermal decomposition, and template techniques, have been developed and improved
for the organization of CuO micro/nanostructures with different morphologies such as
nanoparticles, nanoellipsoids, nanorods, nanoneedles, nanoribbons, nanoshuttles, nanoleaves,
nanotubes, 3D peanutlike patterns [90], pricky/layered microspheres [83a], dandelion-like

CuO hollow microspheres [91], and flower-shaped structure [72, 92]. All these techniques
have indeed been demonstrated as fabrication tools for formulation of CuO nanoparticles into
the manipulated position with controlled structures. But the required toxic raw materials, the
removal of the template and the contamination of the byproducts limit their exploitation at the
application level. Therefore, it is required to develop a clean and friendly method to
synthesized complex CuO nanostructures in large-quantity under mild conditions. Herein, we
demonstrated biomolecule-assisted green method for synthesizing hierarchical hollow CuO
micro/nanostructures simply by using CuSO4, NH3·H2O, and tyrosine instead of any toxic and
dangerous reagents [37h].
Low-magnification FESEM observations show that the panoramic morphology of the as-
obtained CuO product is in large quality and mainly composed of uniform, spheric
architectures ranging from 1.5 to 3 m in diameter (Figure 5A). The clear view (Figure 5B
and 5C) displays that the surface of the architecture is not smooth and consists of many
nanosheets with average diameter of ca. 250 nm. The examination of a crashed microsphere
(Figure 5D) vividly reveals that the structure of these spheric architectures is assembled from
nanosheets, self-wrapping to form hollow interiors with 1.5-3 m in outer diameter. It is
worth noting that all the constituent nanosheets are aligned without any substrate support, and
the driving force originates from the biomolecule [37b, 71a, 72]. In this case, tyrosine is
dispersed within liquid medium mainly as a random coil sol bearing abundant negatively
charged carboxylic groups. These random coils behave as organic matrixes, binding many
copper cations. The introduction of ammonia changed the pH value and caused Cu2+ ions to
hydrolyze to form intermediate, which was still attached to tyrosine. Upon hydrothermal
treatment, the intermediate decomposed and CuO could be formed. The gradual attachment of
reactive constituents onto the nucleation centers leads to formation of relatively flat plates.
With an increased growth rate, a highly curved structure may form due to the lattice tension
or surface interaction [37b, 71a, 72]. As the reactant is consumed, the subsequent growth of
the crystals takes an energetically favorable self-assembly of tiny sheets and is confined by
the curved structure. As a result, ordered nanosheet-based mesoporous shells are constructed.
After sonication treatment of the sample for 10 min, the morphology can be well kept (Figure
5E), which shows the highly mechanical stability. In the absence of the tyrosine, the dominant
morphology is mat composed of interconnected nanosheets as shown in Figure 5F. All these
observations verify the formation of mechanically stable and hierarchical CuO
micro/nanostructures with the help of biomolecule, tyrosine.
A B
m m
C D
nm m
E F
m m
Figure 5 FESEM images of the sample: (A) low-magnification image, (B) a single
Figuremicrospehere,
5. FESEM images of the
(C) the sample:
surface (A)microsphere,
of the low-magnification image,CuO
(D) crashed (B) micro/nanostructure,
a single microspehere,and(C) the
surface
(E)ofafter
the microsphere, (D) crashed
sonication treatment. (F) CuO
is themicro/nanostructure,
FESEM image of theand (E) after
product sonication
in the absencetreatment. (F) is
of tyrosine.
the FESEM image
Reprinted of permission
with the productfrom
in theRef.
absence
37h, of tyrosine.2008
Copyright Reprinted with permission
American from Ref. 37h,
Chemical Society.
Eu3+-doped ZnO Urchins

In response to ever-increasing demands for displays, liquid crystal display, back light,
and alternatives to general lighting (incandescent light bulb), a tremendous emphasis is being
placed on white light as a more efficient replacement for conventional lighting sources [93].
Besides the difficulties associated with the employment of ultraviolet-light-emitting diode
(UV-LED) with triple-wavelength red-green-blue (RGB) phosphors [94], single host lattice is
not able to produce white light with satisfactory CIE coordinates. These have generated great
interest in developing white light emitters via chemical approaches.
It is well known that ZnO exhibits wide visible emission from UV near-band-edge
emission to a visible deep-level one [95]. But, it is difficult to get white light of special CIE
coordinates with pure ZnO. Fortunately, visible PL from ZnO could be tuned in a wide range
from blue to green and orange through chemical doping [96]. As host materials, ZnO
theoretically enables full color (blue, green, and red) via energy transfer (ET) to longer
wavelength emitters. Therefore, ZnO is a potential candidate for fabricating white light-
emitting materials. Meanwhile, Eu3+ ions are ideal red emitter due to its efficient red emission
from 5D0→7FJ. In an earlier effort, Ishizumi et al. tried to dope ZnO nanorods with Eu3+ ions
and the characteristic Eu3+-related PL could be observed when excited by the characteristic
absorption of Eu3+ ions, 466 nm, but not the case for excited by 325 nm, which means that
white lighting could not be attained by single wavelength light excitation [97]. To achieve
such objective, it is desirable to control the doping level of the red emitter and tune the ET
process from host to guest in the ZnO system simultaneously. In order to achieve this goal,
we obtain uniform Eu-doped ZnO urchins with white light by the hydrothermal thermolysis
of the Zn(Ac)2 in the presence of water-soluble biopolymer, sodium alginate [37c]. Figure 6A
(FESEM images) shows that the morphology of the product is uniform micrometer urchins. A
closer observation (Figure 6B) displays that such urchin-like product are secondary structure
consisting of orderly aggregates of nanorods. The SAED (shown in the inset in Figure 6B)
exhibits strong and ordered single-crystallite diffraction spots, demonstrating crystalline
nature of this product. The powder XRD (Figure 6C) displays that all the diffraction peaks
could be indexed to phase-pure würtzite-type ZnO (JCPDS 80-0075). No characteristic peaks
could be observed other than ZnO. Comparable experiment, without using sodium alginate as
additive, yielded mixture of Eu2O3 and ZnO, which was accompanied with dramatic change
in terms of morphologies. These results verify the unique function of such biopolymer for the
fabrication of doped ZnO nanomaterials with well-defined morphology.
To make full understanding of the Eu doping and the defect state in the urchins, Raman
spectrum was measured. The inset in Figure 6C shows the Raman spectrum of the sample
recorded at room temperature. The mode assignment at ambient conditions is well established
in the literature [98]. The sharp and strong Raman peak at 437 cm-1 is attributed to the ZnO
nonpolar optical phonons (E2) mode [99], which is one of the characteristic peaks of würtzite
ZnO and consistent with XRD result [100]. The bands at 172, 327, and 400 cm-1 are assigned
to A1, the overtone of A1, and E1(TO), respectively [101]. The absence of peak at 581 cm-1,
which is related to the E1 mode of oxygen deficiency [102], in our product implies that the
sample is of excellent crystallinity. Meanwhile, there is no signal corresponding to any
europium species in the Raman spectrum. All these demonstrated that Eu-doped ZnO urchins
can be created by a facile one-step low-temperature hydrothermal route in the presence of
water-soluble biopolymer, sodium alginate.
16000
14000 C
Intensity (a.u.)
12000
10000
Intensity
8000 200 400 600 800 1000

-1
Wavenumber (cm )
6000
4000
2000
0
20 40 60 80 100
2 (deg.)
Figure 6. A and B are low-magnification image and a closer observation of the product. C and the inset in it
are the XRD pattern and the room temperature Raman spectrum of the sample. Reprinted with permission
from Ref. 37c, Copyright 2006 American Institute of Physics.
Ag/ZnO Nanocomposites
ZnO, especially on nanometer-scale, has become more and more attractive because of
their different physical and chemical properties from bulk materials [103]. However,
nanometer-scaled materials, such as nanoparticles and nanorods, with a high surface-to-
volume ratio tend to aggregate during the preparation and photocatalysis process, which
results in the reduction of the photocatalytic efficiency. An available way to prevent the
nanoparticles from aggregation and maintain the high photocatalytic efficiency is to organize
these nanoscale materials into hierarchical structures [104]. On the other hand, the strong
antibacterial activities of both metallic Ag and Ag+ ion have been known for a long time
[105]. The difference between them is that the antimicrobial activities of Ag nanoparticles are
significantly influenced by the dimensions of the particles: the smaller the particles, the
greater the antimicrobial effect [106]. However, with the decrease of particle size, Ag
nanoparticles can easily be aggregated, which causes deterioration of their chemical
properties and decreases their antibacterial properties. Moreover, in order to make use of
silver economically, there is also a need to find cheaper ways of using silver in potential
applications without jeopardizing its functionalities. So, Ag nanoparticles have been
supported on SiO2 [107], zeolites [108], and carbon fiber [109]. However, these supports are
inert of antibacterial activity. Considering the fact that ZnO is another inorganic antibacterial
agent and relatively cheaper, it can be expected that ZnO will be an excellent support for Ag
nanoparticles. In addition, this Ag/ZnO metal-semiconductor nanocomposite may inhibit the
bacteria synergistically due to the strong interaction between them. Recently, we succeeded in
modifying 3D ZnO hollow microspheres with Ag with the assistance of sodium alginate
through a facile one-step hydrothermal method [37f].
The representative FESEM patterns of the sample with a Ag content of 1.62 at.% are
shown in Figure 7A. This low-magnification observation indicates that the panoramic
morphology of the as-prepared sample is sphere-like with a diameter ranging from 3-5 μm.
The high magnified FESEM image of a fragment of the broken sphere (Figure 7B) reveals
that the architecture of the hollow microsphere is built from a single layer of oriented
nanorods. The TEM/HRTEM images in Figure 8 corroborate the morphologies observed in
the FESEM images. Figure 8A (a fragment of the broken microsphere) confirms that the wall
of the hollow microsphere is composed of oriented nanorods, which were aligned with their
growth axes perpendicular to the surface of the microsphere. A typical medium-magnified
TEM image of an individual Ag/ZnO nanorod is shown in Figure 8B. The 2D lattice fringes
in the HRTEM image in Figure 8C signify the crystallinity of the ZnO nanorod. The distance
between two fringes is about 0.528 nm, which is close to the d spacing of the (0001) plane,
indicating that the <0001> direction (c axis) is the preferential growth direction of ZnO
nanorods. And in Figure 8D, lattice fringes with interplanar spacing of 0.236 nm,
corresponding to the (111) plane of polycrystalline Ag nanoparticles, are also observed.
Instead of sodium alginate by tyrosine, we obtained rod-like Ag/ZnO nanocomposite (Figure
9) [37g].
A B
Figure
Figure 7. FESEM
7. FESEM images
images of the as-synthesized
of the as-synthesized 1.62 at.%
1.62 at.% Ag/ZnO Ag/ZnO
hollow hollow low-magnification
microspheres: microspheres:
low-magnification panoramic view (A) and a fragment of one broken microsphere (B).
panoramic view (A) and a fragment of one broken microsphere (B). Reprinted with permission from Ref. 37f,
Reprinted
Copyright with
2008 permission
American from
Chemical Ref. 37f, Copyright 2008 American Chemical Society.
Society.
A B
C
D
Figure 8. TEM images of the as-synthesized 1.62 at.% Ag/ZnO hollow microspheres: (A) low-magnified
image of a fragment of a broken microsphere, (B) high-magnified image of an individual nanorod, (C)
HRTEM image of the ZnO (the inset is the corresponding FFT pattern) and (D) HRTEM image of the
Ag/ZnO (the inset is the corresponding FFT pattern of Ag). Reprinted with permission from Ref. 37f,
Figure 9. Low-magnification (left) and high-magnification (right) SEM images of the as-prepared samples
with different initial materials: (a) pure ZnO prepared in the absence of tyrosine, (b) pure ZnO prepared in the
presence of tyrosine, (c) 1.2 at.% Ag/ZnO composite using tyrosine as shape-conductor and reductant.
Reprinted with permission from Ref. 37g, Copyright 2008 Institute of Physics.
Magnetic Nanomaterials
As described in multiple publications, the controlled synthesis of magnetic nanoparticles

is of high scientific and technological interest [110]. Magnetite (Fe3O4) is a common ferrite
having a cubic inverse spinel structure. The compound exhibits unique electric and magnetic
properties based on the transfer of electrons between Fe2+ and Fe3+ in the octahedral sites.
Interest in the magnetite has centered on applications [111] such as multi-terabit magnetic
storage devices, ferrofluids, sensors, spintronics, separation processes, MRI contrast
enhancement agents, and especially biomedical fields.
Magnetite nanoparticles are usually synthesized by the co-precipitation of ferrous (Fe2+)
and ferric (Fe3+) ions by a base [112]. Other synthetic methods include the thermal
decomposition of an alkaline solution of an Fe3+ chelate in the presence of hydrazine, and the
sonochemical decomposition of an Fe2+ salt followed by thermal treatment [113]. Uniformly
sized magnetite nanoparticles were synthesized by the high-temperature reaction of Fe(acac)3
in octyl ether and oleic acid or lauric acid, or a mixture of four solvents and ligands, namely
phenyl ether, 1,2-hexadecanediol, oleic acid, and oleylamine [114]. Even though they produce
highly crystalline and uniformly sized magnetic nanoparticles, these synthetic procedures are
not exempt of drawbacks, because they require expensive and often toxic reagents,
complicated synthetic steps, and high reaction temperatures. To understand the environmental
implications of these nanoparticles, and to facilitate their potential applications, it is important
to develop a simple, green, and generic method for the preparation of Fe3O4 nanoparticles.
Herein we extended such one-pot green chemistry reaction method for the formation of high-
purity magnetite nanocrystals from sodium alginate and FeCl3 in alkaline solution [37e].
FESEM observations show that the panoramic morphology of the as-obtained product is
mainly uniform and spheric architectures (Figures 10A and 10B). TEM image (Figure 10C)
further verified the spheric structure of the products. An XRD pattern (Figure 11A) of the
prepared product can be clearly seen and indexed to the face-centered cubic spinel structure
of pure Fe3O4 with a lattice parameter of a= 8.393 Å, which is very close to the reported
value (JCPDS 65-3107) and expected for this green synthesis route; i.e., the reduction of Fe3+
by biopolymer leads to magnetite as the final product. The broadening of these diffraction
peaks indicated that the sample was composed of nanosized particles. In this biopolymer-
assisted route, the addition of sodium alginate and its amount are the key factors to the green
synthesis of Fe3O4 nanoparticles. There are two functional groups (COO- and OH) on the
sodium alginate molecules, which are hydrophilic groups and can provide coordination sites.
When Fe3+ ions enter into aqueous solution, the sites provide the necessary heterogeneous
nucleation sites, and Fe3+ forms complexes with the hydrophilic functional groups. Herein,
when the temperature is increased, these OH groups on the sodium alginate molecules will
reduce Fe3+ ions to be Fe2+ ions, urea will decompose and release OH-, which with Fe3+ and
Fe2+ ions forms the Fe3O4 crystalline nucleus and grow up on the coordination sites.
Without the addition of sodium alginate, the final products are pure Fe2O3, even when
other experimental conditions are kept the same (Figure 11D). With increasing the amount of
sodium alginate from 0 mL to 5 mL and 15 mL, the products are mixtures of Fe2O3 and Fe3O4
(Figure 11C and Figure 11B, respectively). Until the volume of sodium alginate is increased
to 25 mL or more, the product is pure Fe3O4 (Figure 11A). No external reductant was needed
for Fe3+ reduction in the present method, further suggesting that sodium alginate itself is
responsible for this reduction reaction. The hydroxyl groups in the repeating units of sodium
alginate is hypothesized to be oxidized into carboxyl groups when Fe(III) are reduced to
Fe(II), which can be referred to the literature [37d]. These results show that magnetite
nanoparticles can be fabricated using a biopolymer (sodium alginate)-assisted green route via
redox-based hydrothermal method.
Figure 10. A typical FESEM image (A), high-magnification image (B), and a TEM image (C) of Fe3O4
nanoparticles. Reprinted with permission from Ref. 37e, Copyright 2008 American Chemical Society.
# # Fe3O4
* Fe2O3
Intensity (a.u.)
#
#
# #
# # ## #
a *#
b
* *
c *
d * ** * * * * **
30 40 50 60 * 70 80
Two Theta / degree
Figure 11. XRD patterns of (a) sample obtained in the presence of 25 mL of 25.4 mM sodium alginate
aqueous solution, (b) sample obtained by decreasing the amount of 25.4 mM sodium alginate aqueous
solution from 25 to 15 mL, (c) sample obtained by decreasing the amount of sodium alginate from 25 to 5
mL, and (d) sample obtained by decreasing the amount of sodium alginate from 25 to 0 mL under other
identical conditions. Reprinted with permission from Ref. 37e, Copyright 2008 American Chemical Society.
PROPERTIES OF THE OBTAINED NANOMATERIALS

Some properties, such as the surface-enhanced Raman scattering (SERS), photocatalytic
and magnetic properties, and antitibacterial activity have been investigated on the basis of
these nanostructured materials. These results supply useful theoretic foundations for their
further applications.
SERS-active Substrates
Since the discovery of SERS from pyridine absorbed on an electrochemically roughened

silver electrode reported by Fleischmann in 1974 [115], many researchers have been working
in this field in order to explore its basic nature and to apply this new technique to surface
science, trace analysis, and sensing [116]. Recent studies have shown that intentional
aggregation and the formation of nanoparticles can significantly increase the observed SERS
enhancement [50f, 117]. From the morphology of the obtianed gold sponges (having
abundant particle junctions, Figure 1), it is reasonable that these gold sponges should show
evident SERS activity. To study their SERS effect, 4-mercaptobenzoic acid (MBA) was
employed to be the probe molecule [37a]. A has several binding sites, i. e. the benzene δ-
electron system, carboxyl groups, and thiol groups. Curve a shown in Figure 12 is the
representative SERS spectrum of MBA on gold sponges substrate, which is dominated by the
strong band at about 1080 cm-1 and is assigned to out-of-plane vibration of the benzene ring
[118]. In contrast to this characteristic peak, no Raman signal was observed in the sample
prepared by dropping 20 μL of ethanol solution of 1.0×10-4 mol·L-1 MBA onto a glass slice
(Curve b in Figure 12). Thus the spectrum recorded in Curve a was believed to result from the
surface enhancement effect of gold sponges. The application potential of the prepared silver
sponges was also tested in SERS from R6G. The Raman shifts (Figure 13) and the
corresponding assignments were compared well with previously reported data [50f]. This
result revealed that the silver sponges were also shown to be an excellent SERS substrate
[37a].
Raman Intensity (a.u.)
1080
500 750 1000 1250 1500 1750 2000

-1
Raman Shift (cm )
Figure 12. SERS spectra of gold sponges (a) and no substrate (b) using 10-4 mol·L-1 MBA as analyte.
Reprinted with permission from Ref. 37a, Copyright 2005 Institute of Physics.
From many literatured reports on SERS, it is widely accepted that there are two
enhancement mechanisms [50f, 119]. One is the long-range EM effect and the other is the
short-range chemical (CHEM) effect, which are simultaneously operative. Hereby, the SERS
enhancement of sponges must also come from both EM and CHEM effects. Firstly, to the EM
effects, the as-prepared gold sponges, different from spherical nanoparticle SERS substrates
that rely on inter-particle coupling or aggregations, have great number of particle junctions
with distinct edges and corners, which can act as ―h ot sites‖ for surface plasma and cause the
―rough surface‖ EM enhancement. So the unconventional architecture present is considered
as independent substrates to enhance local scattering field. Secondly, the CHEM effect is also
important because the unique surface properties of the gold (silver) sponges and sulfur
(nitrogen) species from the analyte molecules facilitate efficient adsorption or binding of the
analyte molecules onto the gold (silver) sponges‘ surface. And then the fluorescence energy
of the analyte molecules can transfer to the metal surface, which results in the decrease of
fluorescence intensity and amplification of the Raman spectrum [119]. Of course, theoretical
studies as well as further experiments are required in order to get a complete understanding of
CHEM mechanism. Instructive discussion of the SERS mechanism can refer to the many
excellent review articles on this subject [120].
30000
25000
Raman Intensity
1649
20000
1508
1362
1574
15000
10000
5000
0
1200 1400 1600 1800
-1
Raman Shift (cm )
Figure 13. SERS spectrum of silver sponges using 10-9 mol·L-1 R6G as analyte. Reprinted with permission
from Ref. 37a, Copyright 2005 Institute of Physics.
Photoluminescence (PL) Properties
ZnO is an important II-VI semiconductor with a direct wide band gap and a large exciton
binding energy (60 meV). Its PL properties are strongly dependent on the morphology. Figure
14 displayed the room temperature PL measurement of the 3D ZnO-based hollow
microspheres (Figure 3) [37b]. The emission spectrum shows wide band emission covering
the blue and green regions. The calculated CIE coordinates are 0.24 and 0.31, which fall at
the edge of the white region (the 1931 CIE diagram) [121]. It is generally accepted that the
green emission results from the recombination of a photo-generated hole with a singly ionized
charge state of the specific defect [122]. Thus, it is reasonable to predicate that there exist a
few crystal defects in the hollow ZnO microspheres. Blue emission of ZnO nanostructures,
including ZnO nanotubes [123], ZnO clusters inside mesoporous silica [124], and ZnO
nanorods [63h], has been observed in previous reports. However, the origin of this emission
still remains unclear, although it was speculated to be related to the radiative defects at the
interface of the components of ZnO microspheres, or the existence of interstitial zinc in ZnO
lattices [124]. The emission spectrum obtained here is dramatically different from those of the
products synthesized under different conditions [63h, 125], verifying that the optical
properties of ZnO crystals are very sensitive to the preparation conditions. What‘s more
important is that the wide band emission of the hollow ZnO microspheres suggests that such
superstructure might serve as a potential host for white-light emitting materials. Under this
background, the Eu-doped ZnO urchins (Figure 6) were synthesized and their PL properties
(Figure 15) were measured, which shows that the emission covers nearly the whole visible
spectral region [37c]. The calculated CIE coordinates are 0.28 and 0.35, which fall within the
white region [126]. The emission bands at green and blue region are attributed to ZnO-related
ones. The green emission at 537 nm is assigned to the transition between the antisite oxygen
and donor-acceptor complexes [127]. The blue emission of the sample is likely linked to the
radiative defects at the interface of the components of ZnO urchins [128], or the existence of
interstitial zinc in ZnO lattices [124]. In addition to the ZnO-related emission, two sharp
peaks at 593 and 613 nm are attributed to the 4f-4f intrashell transitions of 5D0→7F1 and
5
D0→7F2 of Eu3+ ions, respectively. Since Ishizumi et al. only observed emission of ZnO with
similar Eu3+ doping level in nanorods [97], a plausible reason for this might be ascribed to ET
from ZnO to Eu3+ ions, which is a suitable energy facilitating the characteristic red light
emission of Eu3+ ions. The ET during such process might be due to the presence of
biopolymer, which means sodium alginate might act as an ―ant enna‖ for Eu3+ ions or enhance
3+
the ET process between ZnO and Eu ions and therefore mediates the ET process. The
results suggest that this biopolymer-assisted hydrothermal method is a feasible route for the
fabrication of ZnO materials doped with optically active impurities.
Intensity (a.u.)
400 450 500 550 600 650

Wavelength (nm)
Figure Figure
14. Room 14. Roomemission
temperature temperature
spectrumemission spectrum
of hollow ZnO of hollow
microspheres. ZnO
Reprinted withmicrospheres.
permission
from Ref. 37b, Copyright 2006 American Chemical Society.
Reprinted with permission from Ref. 37b, Copyright 2006 American Chemical
Society.
Intensity (a.u.)
400 500 600 700

Wavelength (nm)
Figure 15. Room 15.

Figure Room temperature
temperature emission
emission spectrum spectrum
of Eu-doped ZnOof Eu-doped
urchins. ZnOwith
Reprinted urchins.
permission from
Ref. 37c,Reprinted with American
Copyright 2006 permission fromofRef.
Institute 37c, Copyright 2006 American Institute
Physics.
of Physics.
3.5
800
Specific capacity(mA h g )
-1
700
3.0 charge 600
500
400
2.5 300
Voltage (V vs. Li /Li)
200
+
2.0 100
0
0 5 10 15 20
1.5 Cycle number
1.0
0.5 discharge
0.0
0 200 400 600 800 1000 1200 1400

-1
Specific capacity(mA h g )
Figure First
16First
Figure 16. discharge/charge
discharge/charge voltage
voltage profile of the profile of the present
present hierarchical hierarchical CuO
CuO micro/nanostructures at a
micro/nanostructures at a rate
rate of C/10. The inset is variation of C/10.
in charge capacityThe inset
versus cycleisnumber
variation
for thein charge sample
as-prepared capacity versus
at a rate
of C/10.
cycle Reprinted
number with
for thepermission from Ref.
as-prepared 37h, Copyright
sample 2008of
at a rate American
C/10. Chemical
Reprinted Society.
with permission
from Ref. 37h, Copyright 2008 American Chemical Society.
Performance as Electrode Materials for Lithium-ion Batteries

To demonstrate the potential application of the nanomaterials with high electronic

conductivity, we carried out a preliminary investigation into their electrochemical
performance of the present hierarchical CuO micro/nanostructures toward Li uptake-release
(Figure 16). It was found that the first discharge/charge voltage profile displayed three
pseudoplateaus (2.5-2.0, 1.35-1.25, and 1.0-0.02 V vs Li+/Li, respectively) for the Li reaction
with CuO, corresponding to the multistep electrochemical Li reaction process or additional
sites for Li uptake, whereas there are no obvious potential plateaus for Li release from the
crystal lattice of CuO [84b, 129]. It is a typical reaction of transitional metal oxide with Li.
Below 0.7 V, the potential tends to decrease gradually as the discharge depth increases. The
behavior is similar to that described in the literatures [84b, 84c, 129]. The first charge
capacity of hierarchical CuO micro/nanostructures is 560 mA h g-1, which is larger than that
of reported CuO nanomicrospheres [84c]. This result indicates that the construction of
hierarchical micro/nanostructures is an effective way to improve the electrochemical
performance. The improvement may be attributed to the following two factors. On one hand,
the 3D configuration of hierarchical hollow micro/nanostructures can be considered as a 3D
current collector network, which provides negligible diffusion times (short diffusion length),
and enhanced electronic conductivity, and hence is the key to the good power performance.
On the other hand, in view of the large volume expansion of CuO during Li+ uptake (CuO
converts to Cu and Li2O, about a 174% volume expansion), the 3D hierarchical hollow
micro/nanostructures may also be considered as an elastic buffer to relieve the strain
associated with the volume variations during Li uptake-release, suppress particle
pulverization, maintain electronic contact and guarantees good power performance and
capacity retention [16a]. Unfortunately, it is reported that the initial discharge capacity of
CuO is approximately 1240 mA h g-1, higher than the theoretical one (670 mA h g-1) based on
a maximum uptake of 2 Li per CuO [84b,129] and that of the as-prepared hierarchical CuO
hollow micro/nanostructures (560 mA h g-1). Usually, the first discharge capacity of CuO
considerably exceeds the nominal capacity and is ascribed to the electrolyte being reduced to
form a solid electrolyte interphase (SEI) layer and organic conductive polymer [130], the
reduction of the adsorbed impurities on CuO surfaces, the initial formation of lithium oxide
due to the presence of some residual OH- groups in the surface of active CuO, and possibly
interfacial lithium storage [131]. Either electrolyte reduction decomposition or impurities will
consume additional energy irreversibly during the charge/discharge process. Therefore, the 1st
charge capacity is merely 560 mA h g-1, corresponding to 1.67 Li release. Owing to the
hierarchical micro/nanostructures, favourable cycle calendar life and reversible capacity can
be achieved as shown in the inset of Figure 16. It is suggested that practical electrode has
outstanding chemical/mechanical robustness. Therefore, the use of this low-cost and high-
performance hierarchical CuO micro/nanostructures for lithium ion batteries is feasible and
promising.
Photocatalytic Performance
Orange G is selected as a representative organic pollutant to evaluate the photocatalytic

performance of the Ag/ZnO hollow microspheres with various Ag contents (shown in Figure
7) [37f]. As an example, the absorbance spectra of the dye solution were shown in Figure 17
in the presence of Ag/ZnO microspheres with a Ag content of 1.62 at.%. It can be seen that
the absorbance peaks at 248, 331 and 472 nm were reduced significantly, indicating the
degradation of the dye molecules. The photocatalytic experimental data can be converted to a
linear pattern using pseudo-first kinetics model, and the results were shown in Figure 18. A
control test without photocatalyst showed that the photoinduced self-sensitized photolysis can
be neglected compared with the photocatalysis. Also, as a photocatalytic reference,
commercial TiO2 (Degussa P-25) was used to evaluate the activity of Ag/ZnO samples
qualitatively. It can be seen from Figure 18 that the photocatalytic performance of ZnO
microspheres can be significantly improved by depositing an appropriate amount of Ag
nanoparticles. The positive effect of Ag deposits is commonly due to the fact that Ag
nanoparticles on the semiconductor surface behave like electron sinks, which provide sites for
the accumulation of photogenerated electrons, and then improve the separation of
photogenerated electrons and holes. This can be understood based on the proposed charged
separation of Ag/ZnO under UV irradiation shown in Scheme 3. Because the bottom energy
level of the CB of ZnO is higher than the new equilibrium Fermi energy level (Ef) of Ag/ZnO,
the photoexcited electrons on the CB under UV irradiations could transfer from ZnO to the
Ag nanoparticles. It has been proposed that the charge separation is the outcome of a Schottky
barrier formed at the metal-semiconductor interface [106]. Therefore, Ag nanoparticles,
acting as electrons sinks, reduce the recombination of photoinduced electrons and holes, and
prolong the lifetime of the electron pairs. Subsequently, the electrons can be captured by the
adsorbed O2 and the holes can be trapped by the surface hydroxyl, both resulting in the
formation of hydroxyl radical species (·OH) [133]. It is accepted that hydroxyl radical species
show little selectivity for attacking dye molecules and are able to oxidize the pollutants due to
their high oxidative capacity (reduction potential of ·OH is 2.8 V). Thus the possible
mechanistic pathway of Ag/ZnO microspheres for degradation of Orange G can be proposed
as follows [134]:
ZnO + (1)
ZnO hh (UV)
(UV) ZnO
ZnO (e
(ecbcb hhvb+vb)) (6)
(1)
(6)
+ (2)
(2)
Ag
Ag Ag
Ag + ee (7)
(7)
H ++ O2 H
ee O
O22 (ads)
(ads) O
O22 H O
O22H
H O2 H O
O22 HH22OO22
e (3)
(3)
e O
O22 OH
OH OH
OH (8)
(8)
Ag + (4)
(4)
Ag + ee Ag
Ag (9)
(9)
h +vb OH OH (5)
(10)
h +vb OH OH (5)
(10)
OH Orange G Degradation Products (6)
(11)
OH Orange G Degradation Products (6)
(11)
0.20
0.18
0.16 0 min
10min
0.14 20min
30min
Absorbance
0.12
40min
0.10 50min
0.08
0.06
0.04
0.02
0.00
200 250 300 350 400 450 500 550 600 650
Wavelength (nm)
Figure 17. The time-dependent UV-vis absorbance spectra of Orange G in the presence of Ag/ZnO sample
with a Ag content of 1.62 at.%. Reprinted with permission from Ref. 37f, Copyright 2008 American
Chemical Society.
1.62 at.% Ag/ZnO

4
3.30 at.% Ag/ZnO
0.83 at.% Ag/ZnO
6.45 at.% Ag/ZnO
Degussa P25
3
Pure ZnO
Without Catalyst
ln( C0/C)
0 10 20 30 40 50 60
Time (min)
Figure 18. The ln(C0/C) versus time curve of photodegradation of Orang G. C0

and C are the initial concentration after the adsorption equilibrium and the
reaction concentration of Orange G, respectively. The experimental data are fitted
using the persudo-first-order kinetic equation: ln(C0/C) = kt. Reprinted with permission
from Ref. 37f, Copyright 2008 American Chemical Society.
However, it is found from Figure 18 that as the Ag content increases, the photocatalytic
performance of the Ag/ZnO has not been enhanced monotonously. When the Ag content is
relatively lower (< 1.62 at.%), the photocatalytic activity of the Ag/ZnO increases with the
increase of Ag content (ZnO < 0.83 at.% Ag/ZnO < 1.62 at.% Ag/ZnO). On the other hand,
when the Ag content is relatively higher (>1.62 at.%), the photocatalytic activity of the
Ag/ZnO decreases with the increase of Ag content (1.62 at.% Ag/ZnO > 3.30 at.% Ag/ZnO >
6.45 at.% Ag/ZnO). Thus the optimal Ag content is approximately 1.62 at.%. Several groups
have suggested that, at higher metal content than the optimized, the over accumulations of
electron on metal deposits could attract the photogenerated holes to the metal sites. This may
encourage the recombination of charge carriers and the metal deposits reversely behave as
recombinant centers [135]. In addition, higher surface loadings may decrease the catalytic
efficiency of the semiconductor due to the reductive availability of semiconductor surface for
light absorption and pollutant adsorption [136]. In this case, this may result from the variation
of the surface hydroxyl content of Ag/ZnO with different Ag content.
Scheme 3. Proposed charge separation process and photocatalytic mechanism of as-prepared Ag/ZnO
samples under UV irradiation. Due to that the energy level of CB for ZnO is higher than the Fermi energy
level of Ag, the photoinduced electrons are transferred to the metallic Ag. Then the electrons in the Ag sinks
can be trapped by the chemisorbed O2 and the hole can be captured by the surface hydroxyl. Reprinted with
permission from Ref. 37f, Copyright 2008 American Chemical Society.
In the process of photocatalysis, after the photogenerated electrons and holes are
separated from recombination, they can be trapped generally by the oxygen and surface
hydroxyl, respectively, to produce ultimately the primary oxidizing species of the hydroxyl
radicals (·OH). So, the OH groups play a significant role in the photocatalytic oxidation
process [137]. Figure 19 shows the variation of persudo-first-order rate constants and the
surface hydroxyl contents of Ag/ZnO with the different Ag content. It can be seen that the
photocatalytic performance of Ag/ZnO microspheres is in good line with the surface hydroxyl
content, i.e., the more the surface hydroxyl content is, the more efficient the photocatalyst is.
From this point, we can reasonably conclude that the difference in photocatalytic activity is
related to various contents of surface hydroxyl on the Ag/ZnO microspheres caused by
different Ag contents. Therefore, the 1.62 at.% Ag/ZnO sample, which has the highest surface
hydroxyl content, exhibits the highest photocatalyst performance in our work.
Antibacterial Activity
For more environmental application, the antibacterial activity of the Ag/ZnO

nanocomposites (shown in Figure 9) with the Ag content of 1.20 at.% was checked. For
comparison, the antibacterial activity of pure Ag and ZnO are also tested [37g]. The
antibacterial activities against microorganisms considered were qualitatively and
quantitatively assessed by determining the presence of inhibition zones and the minimum
inhibitory concentration (MIC) values, respectively. Antibacterial effect in the form of

inhibition zones, evaluated by the disk diffusion assay, is shown in Figure 20. The
bactericidal activity against E. coli and S. aureus showed clear zone of inhibition within and
around the disk impregnated with ZnO and Ag/ZnO, respectively. These results indicate that
Ag/ZnO nanocomposites have efficient antibacterial capability for both Gram-negative (G-)
and Gram-positive (G+) bacteria.
To further quantitatively investigate the antibacterial activity, the MIC value of Ag, ZnO
and Ag/ZnO is determined (Figure 21). The results are shown in Table 1.
0.070
70
0.065
0.060
60
0.055
Rate constant ( min )
-1
OH percent (%)
0.050
50
0.045
0.040
40
0.035
0.030 30
0.025 Rate constant vs. Ag content

OH percent vs. Ag content
0.020 20
0 2 4 6
Ag content (%)
Figure 19. Persudo-first-order rate constant k and the surface hydroxyl content of Ag/ZnO as a function of Ag
content. The value of rate constant k is equal to the corresponding slope value of the fitting line shown in
Figure 18. Reprinted with permission from Ref. 40f, Copyright 2008 American Chemical Society.
Table 1. Minimum inhibition concentrations (MIC) assay values for Ag, pure ZnO and
Ag/ZnO nanocomposites
Minimum inhibition concentrations (μg/mL)

Bacteria
Ag ZnO Ag/ZnO (1.6 wt.%)a
-
Escherichia coli (G ) 15 3500 600
Staphylococcus aureus (G+) 25 1000 400
a
The weight percent of 1.20 at.% Ag/ZnO is 1.6 wt.%.
Figure 20. The qualitative paper disk diffusion assay results: (a) E.coli; (b) S.aureus. Reprinted with
permission from Ref. 37g, Copyright 2008 Institute of Physics.
It can be seen that the MIC values of Ag/ZnO nanocomposites were 600 μg mL-1 for E.
coli and 400 μg ml-1 for S. Aureus, respectively. Considering the fact that the mass percentage
of Ag in Ag/ZnO nanocomposites was about 1.6 wt.%, the MIC values of Ag in Ag/ZnO
nanocomposites were about 9.6 μg mL-1 and 6.4 μg mL-1, respectively. It can also be seen that
the MIC values of pure Ag nanoparticle were 15 μg mL-1 for E. coli and 25 μg mL-1 for S.
aureus, and those of pure ZnO were 3500 μg mL-1 for E. coli and 1000 μg mL-1 for S. aureus,
respectively. From the practical and economical point of view, the Ag/ZnO nanocomposites
were more attractive compared to pure Ag and ZnO. Also, we find that though both Ag and
ZnO show antibacterial activities, their antibacterial efficiency is different for G- and G+
bacteria. That is, Ag shows better antibacterial activity for G- bacteria than G+ bacteria, while
ZnO shows better antibacterial activity for the G+ bacteria than G- bacteria. This phenomenon
was also reported in the literature for Ag [105g, 105h] and for ZnO [138]. So when Ag and
ZnO are coupled together to form Ag/ZnO nanocomposites, they showed enhanced
antibacterial capacity for both G- and G+ cells.
Moreover, it was found that the efficiency of Ag/ZnO nanocomposites was not simply the
sum of the efficiency of pure Ag and pure ZnO. It can be seen that in Ag/ZnO
nanocomposites, the components of Ag and ZnO may undergo antibacterial activity
synergistically. One reason is that ZnO matrix decreases the aggregation of Ag nanoparticles.
Then the Ag nanoparticles can have more opportunities to attach to the cell membranes and
interact with sulfur- and phosphorus-containing compounds in them. It has been reported that
this interaction between the Ag nanoparticles and the cell membrane will disturb the cell‘s
power function such as permeability and respiration, leading to cell death [105i, 106b].
Figure
Figure 21 Growth
21. Growth inhibition
inhibition against coli
against Escherichia coli and
and Staphylococcus
Escherichia of (a) Ag nanoparticles,
Staphylococcus
aureus aureus (b)
ZnO nanorod and (c) 1.2 at. Ag/ZnO nanocomposite. Each point represents
of (a) Ag nanoparticles, (b) ZnO nanorod and (c) 1.2 at. Ag/ZnO nanocomposite.the mean ± 1 standard error. C 0:
the concentration of initially incubated bacteria. **: significantly different from C0 (P < 0.01). Reprinted
Each point represents the mean ± 1 standard error. C0: the concentration of
with permission from Ref. 37g, Copyright 2008 Institute of Physics.
initially incubated bacteria. **: significantly different from C0 (P < 0.01).
60
30
M (emu/g)
0
-30
-60
-5.0 -2.5 0.0 2.5 5.0

H (KOe)
Figure 22. Magnetization-hysteresis (M-H) loops of Fe3O4 nanoparticles measured at room temperature.
Reprinted with permission from Ref. 37e, Copyright 2008 American Chemical Society.
Magnetic Properties
Figure 22 presents hysteresis loop for Fe3O4 nanomaterials (shown in Figure 10)
measured at room temperature [37e]. The hysteresis loop demonstrates a ferromagnetic
behaviour with the saturation magnetization (Ms) value about 62.1 emu / g, which is different
from those reported for Fe3O4 nanomaterials [139]. The remanent magnetic induction (Mr)
and coercivity (Hc) values are about 8.9 emu / g and 93.6 Oe, both of which are lower than
those of bulk Fe3O4 [140]. As we know that the effects of size, structure, and morphologies
are concerned with the magnetic properties of the nanomaterials. Typical reasons for this
include the reaction or complexation of the surface atoms of magnetic nanoparticles with
surfactant, which may create a magnetically dead layer [141]. With a significant fraction of
surface atoms, any crystalline disorder within the surface layer may also lead to a significant
decrease in the nanoparticle saturation magnetization. The underlying factor affecting the
magnetic properties in our case needs further investigation.
CONCLUSION
An interdisciplinary culture has evolved in nanotechnology that requires the collaboration
between physicists, chemists, biologists and engineers. The corresponding disciplines have
made remarkable progress in the synthesis, visualization, manipulation, modification and
control of materials on the nano- and micrometer scale. As a promising strategy, bioinspired
synthesis has proven to be a completely new technological approach and environmentally
friendly route to construct inorganic materials with confined crystalline units and assembly
hierarchy and tailored correlative properties [142]. Thus, the exploration of bioinspired
morphosynthesis strategies, using self-assembled organic superstructures, organic additives,
and/or templates with complex functionalization patterns, to construct inorganic materials
with controlled morphologies, has drawn a lot of attention. The development of reliable, eco-
friendly, biologically inspired experimental processes for the synthesis of nanomaterials
becomes more and more important aspect of nanotechnology today.
Based upon this background, we developed such biopolymer-assisted green synthesis to
fabricate metal, semiconductor, and magnetic nanomaterials. By taking advantage of the
biopolymers, unique metal sponges, 3D ZnO hollow microspheres, hierarchical CuO hollow
micro/nanostructures, doped ZnO with hierarchical structures, and spherical Fe3O4
nanoparticles can be created by one-step hydrothermal technique. The drawbacks (the use of
toxic reagents in the synthesis of magnetic nanomaterials) can be overcome by using
biopolymers as reducing and stabilizing agents. These preliminary results verify the
feasibility of such biopolymer-assisted green synthesis for the fabrication of nanomaterials. In
addition, the biopolymer are safe reagents compared with those chemical routes utilizing
special and poisonous chemicals. The use of environmentally benign and renewable materials
and the combination of nanoparticles and biological molecules are very attractive and have
gained tremendous attention from academics and industry, because such a strategy offers
numerous benefits ranging from environmental safety to ready integration of these
nanomaterials to pharmaceutically and biologically relevant systems. This aspect makes the
present method possible for use in further green nanoparticle syntheses. Although replication
of such natural structures in terms of form and function remains an enduring and challenging
goal, the future development of novel bionanocomposites with improved properties and
multifunctionality can be envisaged as an emerging, open field of research, with plenty of
possibilities because of the great abundance and diversity of biopolymers in Nature, as well as
the advantage of their synergistic combination with inorganic nanosized solids.
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Lecture Material 18
BIOLOGICALLY TARGETED NANOPARTICLES

AS CANCER THERAPEUTICS
ABSTRACT
The concept of biological targeting has led to the development of a new
generation of therapeutic agents. These molecularly targeted agents have achieved
great clinical results not previously seen with conventional agents. Their success, in
turn, has inspired the development of biologically targeted nanoparticles for
therapeutic applications, especially for the treatment of cancers. Biologically targeted
nanoparticles are engineered by functionalizing the nanoparticles‘ surface with
targeting ligands, such as antibodies, peptides, small molecules and oligonucleotides.
Targeting ligands can improve the differential accumulation of nanoparticles at
specific sites in the diseased tissue. Preclinical data have demonstrated that targeted
nanoparticles are more efficacious and less toxic when compared to non-targeted
nanoparticles. In this chapter, we will review the rationale for molecularly targeting
nanoparticles, overview the various classes of targeting ligands, discuss the
formulation of targeted nanoparticles, and highlight interesting examples from the
preclinical data on targeted nanoparticle therapeutics.
INTRODUCTION
Advances in genetics, biochemistry, cell and molecular biology over the last several
decades have dramatically improved our understanding of human cancers. We have
uncovered important mechanisms of cancer pathogenesis as well as identified numerous
biomarkers of cancer tissue. Based on this new knowledge, a new generation of therapeutics,
targeted specifically against the cancer specific pathways and biomarkers have been
developed. These targeted agents have the ability to selectively treat diseased tissue without
causing significant side effects on normal tissue. Over the last two decades, targeted cancer
therapeutics have achieved great clinical results that have eluded conventional therapeutics
[1-5]. Examples include small molecules such as imatinib for the treatment of chronic
myelogenous leukemia (CML) and monoclonal antibodies such as trastuzumab for the
treatment of human epidermal growth factor receptor type 2 (HER2) positive breast cancer.
Small molecule targeted agents treat cancer by inhibiting important oncogenic pathways.
As an example, imatinib (Gleevec, Novartis) is a relatively selective tyrosine inhibitor of the
kinase ABL and a first line therapy for the treatment of CML [6, 7]. In CML, more than 90%
of the cases are caused by the Philadelphia chromosome (t(9;22) (q34;q11)), which results in
the BCR-ABL fusion with constitutively active ABL kinase activity [8, 9]. Despite the fact
that many normal tissues express the ABL kinase and that imatinib does not discriminate
between normal and diseased tissue, the targeted inhibition of ABL by imatinib showed
striking efficacy and little toxicity in the treatment of CML [5, 10-12]. In contrast,
monoclonal antibody-based therapeutics target both disease specific biomarkers as well as
oncogenic pathways. These antibodies are generally selected against cancer specific
biomarkers that have important signaling properties and are important to cancer pathogenesis.
Examples include trastuzumab against HER2 in breast cancer, bevacizumab against vascular
endothelial growth factor (VEGF), cetuximab against epidermal growth factor receptor
(EGFR), and rituximab against CD20. Some of the best clinical data come from trastuzumab,
a monoclonal antibody targeted against the extracellular juxtamembrane domain of HER2.
HER2 is a transmembrane tyrosine kinase receptor that normally regulates cell growth and
survival. It is over-expressed in 20 to 30% of breast cancers and has been associated with
poor treatment outcome and decreased overall survival [13-15]. Trastuzumab binds to HER2
and inhibits its signaling by preventing the activation of the tyrosine kinase [16]. Since HER2
is also a surface biomarker, trastuzumab is physically targeted against breast cancer as well as
molecularly targeted against breast cancer cells. In clinical trials, trastuzumab achieved
remarkable results: improved both disease-free survival and overall survival [1, 17, 18].
In additional to biologically targeted agents, another major advance in cancer therapeutics
that is equally as significant is the development of nanoparticle cancer therapeutics.
Conventional chemotherapeutics have several significant challenges that affect their
therapeutic index. These include poor solubility, short circulating half-life, and uniform
biodistribution between diseased and non-disease tissue. The development of nanoparticles
has provided a solution to these challenges. In particular, precise engineering of nanoparticles
allows unprecedented freedom to modify fundamental properties such as solubility,
diffusivity, biodistribution, release characteristics and immunogenicity. Nanoparticle
therapeutics have been shown to have longer circulation half-lives, superior bioavailability
and lower toxicity when compared to their small molecule counterparts [19, 20]. Nanoparticle
agents quickly translated to clinical practice. Today, there are twenty-four nanoparticle
therapeutics in clinical use [21]. In addition, a 2006 global survey conducted by the European
Science and Technology Observatory (ESTO) revealed that more than 150 companies are
developing nanoscale therapeutics, highlighting the high enthusiasm for nanoparticle-based
therapeutics [22].
Another important characteristic of nanoparticle-based chemotherapeutics is that they are
passively targeted to the cancers due to the enhanced permeability and retention effect (EPR)
[23]. EPR is the property by which macromolecules, such as nanoparticles, tend to
accumulate in tumor tissue much more than they do in normal tissues [24]. This phenomenon
is due to the characteristics of tumor microvasculature: defective architecture, impaired
lymphatic drainage, and permeability to macromolecules [25, 26]. Nanoparticle extravasation
is dependent on the size of open interendothelial gaps and transendothelial channels. The pore
cutoff size of these transport pathways has been estimated between 400 and 600 nm [27]. In
general, smaller particles are more effective for extravasating the tumor vasculature.
The success of both biological targeted agents and nanoparticle therapeutics has led to
increasing interest in combining molecular targeting and nanoparticle delivery. Biologically
targeted nanoparticles have been formulated by functionalizing the nanoparticles‘ surface
with targeting ligands. By combining biological targeting with nanoparticles, it can lead to
higher intratumoral drug concentration and lower systemic toxicity. Furthermore, the binding
of targeting ligand to tumor cells often induces intracellular uptake of the nanoparticles,
leading to higher intracellular drug concentration. Therefore, actively targeted nanoparticles
have several important advantages: the ability to partition more of the nanoparticles within
target tissue, increased uptake into target cells, higher therapeutic efficacy and lower toxicity.
Targeting ligands include monoclonal antibodies, aptamers, peptides, antibody fragments and
small molecules, all of which have been conjugated to nanoparticles. Preclinical data obtained
from these targeted nanoparticles thus far have supported the advantages of targeted
nanoparticles [28-35]. Although there is no clinically approved targeted nanoparticle
therapeutics to date, many are in preclinical and clinical development.
TARGETING LIGANDS
Our improved knowledge of disease specific processes/biomarkers enables the
identification of targeting ligands against these targets. The major targeting ligand classes
include monoclonal antibodies, antibody fragments, peptides, aptamers and small molecules.
These targeting ligands differ in their size, process of production/synthesis, selection process,
immunogenicity, and stability. Each of the targeting ligand classes will be reviewed below.
Monoclonal Antibodies
Antibodies are proteins produced by the B cells of the immune system to target foreign
agents. They can bind molecules ranging from small organic compounds to large proteins
with high affinity and specificity, making them excellent targeting ligands. Antibodies are
glycoproteins around 150kDa in size [36]. The basic functional unit of each antibody is an
immunoglobulin (Ig) monomer [37]. The Ig monomer is a "Y"-shaped molecule that consists
of four polypeptide chains; two identical heavy chains and two identical light chains. Each
heavy chain has two regions, the constant region and the variable region. The variable region
of each heavy chain is approximately 110 amino acids long. A light chain also has one
constant domain and one variable domain. Each antibody contains two light chains that are
always identical. The variable domains of the light chains and heavy chains make up the tips
of the ―Y‖ shaped antibody. This region of the antibody is called the Fab (fragment, antigen
binding) region and is responsible for the binding of antigen [38]. The base of the ―Y ‖ is
called the Fc (Fragment, crystallizable) region. It is composed of constant regions of two
heavy chains. Its primary function is to modulate the immune response for a given antigen
[39].
Today, the applications of antibodies include laboratory agents, diagnostic tests and
therapeutic agents. The key challenge of utilizing antibodies for biomedical applications has
been the production of monospecific antibodies (monoclonal antibodies). Monoclonal
antibody (mAb) production was first described by Kohler et al. in 1975 [40]. In this
technique, myeloma cells are fused with normal B-cells from a mouse that has been
immunized with the desired antigen to form hybrid cells (hybridomas). The hybridomas can
grow indefinitely and are able to produce the desired monoclonal antibodies in large
quantities. However, such mAbs are immunogenic in humans and can lead to allergic
reactions as well as human anti-mouse antibodies (HAMA). Various approaches have been
developed in the last several decades to address this problem. In one approach, recombinant
DNA technology is utilized to generate half-mouse and half-human antibodies. These are
called chimeric antibodies or humanized antibodies depending on the proportion of the human
component [41-43]. Another approach involves transgenic mice expressing human antibody
gene sequences in order to produce more human-like antibodies [44]. One technique for
screening and selecting mAbs is the display (phage, ribosome and yeast) technology. It
involves the engineering of recombinant antibodies through the use of viruses or yeast.
Immunoglobulin gene segments are cloned and expressed to create libraries of antibodies
from which antibodies with desired specificities are selected [45, 46]. The display technology
can enhance the mAb specificity and their stability.
As targeting ligands for nanoparticles, mAbs‘ advantages include high binding specificity
and affinity, a relatively easy selection process for a particular antigen, and well-established
in vivo toxicity profiles. However, they have their share of limitations. Monoclonal antibodies
are large, complex molecules that penetrate tumors poorly and require significant engineering
at the molecular level to be effective [47, 48]. They are expensive to manufacture and there
exists variation from batch to batch, limiting their efficiency as targeting molecules.
Antibodies are sensitive to high temperatures and changes of pH. They typically expire in
weeks unless stored near freezing temperatures. The Fc domain can lead to non-specific
interactions and immunologic reactions. Lastly, despite the engineering efforts to make mAbs
more humanized, they still contain non-human components, which can lead to allergic
reactions and HAMA.
Antibody Fragments
Advances in molecular biology and recombinant DNA technology have lead to the
development of antibody fragment molecules. Antibody fragments contain antigen binding
domains of the antibody but are without other structural domains such as the Fc region [49].
They are smaller in size, have lower immunogenicity and lower toxicity when compared to
full antibodies. Antibody fragments can be categorized as either monovalent or multivalent
molecules. Fab, single chain variable fragments (scFv) and nanobody are monovalent
antibody fragments. Multivalent molecules include Fab2, Fab3, minibody, diabody, triabody,
tetrabody, and small antibody mimetics. These molecules‘ size and composition summarized
in Table 1.
Table 1.
Size Composition
Fab [38, 49] ~55 kDa One constant and one variable domain from each heavy and
light chain
scFv [50] ~28 kDa The variable regions of the heavy (VH) and light chain (VL)
Nanobody ~15 kDa Heavy chains only, both the variable and constant domains
[51]
Small 3 kDa Fusion of two complementarity-determining regions that
antibody retained the antigen recognition of their parent molecules.
mimetics [52]
Fab2 ~110 kDa Covalently linked dimers of Fab
Fab3 ~165 kDa Covalently linked trimers of Fab
Minibody ~75 kDa Fusion between scFV and a CH-3 domain that self-assembles
[53] into a bivalent dimer
Diabody [54] ~50 kDa Covalently linked dimers or non-covalent dimers of scFVs
Triabody ~75 kDa Covalently linked trimers of scFVs
Tetrabody ~75 kDa Covalently tetramers of scFVs
Single-domain antibody fragments, despite their small size, generally retain the specific
antigen-binding affinity of the parent antibody [50]. On the other hand, because of their
smaller size, these molecules have better tissue penetration than full antibodies. They are also
able to target cryptic epitopes that may be poorly accessible to full antibodies [55]. The small
size of single-domain antibody fragments is also advantageous for molecular targeting of
nanoparticles. Smaller size of targeting ligand generally means smaller targeted nanoparticles,
less immunogenicity, and longer circulation half-lives. Cheng et al. compared whole
monoclonal antibody, Fab fragments and scFv as targeting ligands for liposomal doxorubicin
[56]. In this study, a anti-CD19 monoclonal antibody, its Fab fragment, and its scFv fragment
were conjugated to immunoliposomes. It showed Fab targeted liposomes had longer
circulation half-life than whole antibody targeted liposomes or scFv targeted liposomes, and
had the best treatment results. The shorter-half life of scFv targeted liposomes was thought to
be due to the particular scFv construct. A number of Fab molecules have been FDA approved
for clinical application. These include Fab against gpIIb/gpIIa for cardiovascular diseases;
Fab against snake venom as rattlesnake antidote; Fab against CEA for colorectal cancer
imaging.
Nanobodies, also called single domain antibodies or VHH antibodies, are composed of
heavy chains only. They were first discovered by Hamers-Casterman et al. who observed that
half of the antibodies produced by camelids (bactrian camels, dromedaries, and llamas) lack
the light chain [51]. Despite the lack of light chains and ten times smaller, nanobodies‘
binding affinity and specificity are similar to that of full antibodies. In addition, nanobodies
are more resistant to heat and pH, and may retain their activity as they pass through the
gastrointestinal tract [57]. Therefore, nanobodies are highly suited for applications that
require a high stability, which includes biological targeting of nanoparticles. Nanobodies have
been engineered to target the EGF receptor and CEA for cancer targeting, though they still
remain in a preclinical investigational stage of development [58, 59].
Small antibody mimetics are the smallest of all antibody fragment molecules. They were
first reported by Qiu et al. who fused two complementarity-determining regions (CDRs),
VHCDR1 and VLCDR3 through a cognate framework region VHFR2 [52]. Small antibody
mimetics are approximately 3 kDa in size, 50 times smaller than a full antibody. Qiu et al.
demonstrated that the small antibody mimetics retained the binding properties of the parent
antibody. In vivo imaging studies showed the small antibody mimetics had enhanced tumor
localization and penetration ability when compared to the parent antibody. When linked to a
bacterial toxin, the antibody mimic-toxin conjugate had greater efficacy than parent antibody-
toxin conjugate.
One disadvantage of single-domain antibody fragments as targeting ligand is their fast
dissociation rates and short retention times on the target antigen in vivo [49]. To improve the
functional affinity and retention time of antibody fragments, multivalent antibody fragments
have been engineered from monovalent antibody fragments. Furthermore, multivalent binding
of cell-surface receptors can result in the activation or inhibition of transmembrane signaling
pathways, which may have therapeutic effects of its own. For example, the binding of CD20
or epidermal growth factor receptor by multivalent molecules can lead to tumor cell death
[60, 61]. Fab and scFV have been engineered into dimeric, trimeric or tetrameric conjugates
which are called Fab2, Fab3, diabodies, triabodies, tetrabodies, using either chemical
reactions or recombinant DNA technology [62-65]. Compared to the parent antibodies, these
multivalent antibody fragments have better retention and internalization properties.
The advantages of antibody fragments as targeting ligands for nanoparticles include
smaller size, higher stability, easier production process, and less batch to batch variation.
However, the production of antibody fragments, like antibodies, still relies on in vivo
production systems, such as bacteria and hybdridomas. Therefore, antibody fragments have
higher cost, more batch to batch variation, and higher immunogenicity when compared to
targeting ligands such as peptides or small molecules.
Peptides
Peptides are short polymers formed from L-amino acids. It has been shown that certain
peptides can bind to their targets with high specificity and affinity, similar to antibody
fragments. For example, cilengitide is a cyclic peptide that binds to integrins, which is
currently in phase II clinical trials for the treatment of non-small cell lung cancer and
pancreatic cancer [66]. The reduction of size from antibody to antibody fragment has resulted
in lower immunogenicity, higher stability and increased tissue penetration. Similarly, peptides
have the lowest immunogenicity, highest stability and highest tissue penetration when
compared to the other amino acids-based targeting ligands: antibodies and antibody
fragments.
The selection process of peptides against a particular target utilizes the display
technologies, which allows the screening of more than 1011 different peptide sequences
against a target [67]. There are two major types of display technologies: biological display
systems that employ biological hosts/biological reactions, and nonbiological display systems
that use chemical and engineering techniques. Regardless of the type of display, each display
library consists of three components: the displayed entity, linker, and a corresponding code.
These components are different for each type of the display techniques. Over the past decade,
development of peptide phage libraries, bacterial peptide display library, and plasmid peptide
library for the selection of peptides for biomedical applications have significantly contributed
to the popularity of peptides as targeting ligands [68].
There are several peptides that are distinct in their characteristics. These include A-
domain proteins, AdNectins and affibodies. AdNectins were initially derived from the 10FN3
sheets that are responsible for structural diversity [69]. AdNectins ars thermostable and
resistant to various proteases. A large library of AdNectins can be created by introducing
diversity into the three loops. Using the library, AdNectins capable of binding to specific
molecular targets can be isolated using enrichment strategies. Recently, an AdNectin for
human vascular endothelial growth factor receptor 2 (VEGFR2), Angiocept, has been isolated
by AdNexux Pharmaceuticals. It entered Phase I clinical trials for treating advanced solid
tumors and non-Hodgkin's lymphoma in 2006 [70].
A-domain proteins are naturally occurring, cell-surface proteins that bind their target
through multiple points of attachment [71]. The first A-domain protein was found in the low-
density lipoprotein receptor (LDLR) by Tschopp and Mollnes [72]. Today, there are
approximately 200 A-domain proteins that have been discovered. They are generally 40
amino acids in size and can bind to a diverse set of targets. Several structural features are
conserved in A- -sheet,
and a high-affinity calcium-binding site. The N- and C-terminal portions of the molecule are
folded into loops. The C-terminal loop contains the calcium-binding site. Because of their
very small size (4–5 kDa), A-domains are frequently linked together to create higher binding
avidity. A large number of residues can be randomly mutated to generate libraries of A-
domain proteins, which can be screened against a particular target [71].
Affibodies are small (6kDa) affinity proteins that were first described by Nord et al. in
1997 [73]. The first affibody was derived from an antibody-binding domain (Z domain) of
staphylococcal protein A (SPA) [74]. The three-helix Z domain is a single peptide that is
capable of rapid and independent folding, and has a native capability of high binding affinity
with target proteins [75]. Affibody were inspired from Z domains. Using combinatorial
protein engineering and display technologies, libraries of affibodies can be constructed and
screened against target molecules. Since their conception, many affibodies have been selected
against clinical significant targets. For example, a 6kDa affibody with subnanomolar affinity
to HER2 receptor has been isolated [76].
Peptides possess some of the most favorable characteristics of targeting ligands: small
size, high stability, high-throughput screening process, and most importantly, ease of
production. Compared to antibody and antibody fragments, peptides are the smallest and have
the highest stability. Display technologies allow high-throughput screening, which translates
into fast and efficient selection process. Because peptides are small and generally fold
spontaneously, their productions are through chemical synthesis rather than relying on in vivo
systems. Therefore, peptides do not exhibit batch-to-batch variation and have lower
immunogenicity.
Aptamers
Aptamers are single-stand DNA or RNA oligonucleotides that can bind to target
molecules such as proteins, sugars and phospholipids, with high sensitivity and specificity
[77]. These single-stand oligonucleotides can fold by intra-molecular interaction into unique
3D conformations, allowing them to bind to target molecules. Aptamers range in size from 20
to 80 bases (~6 to 26 kDa) and have binding affinities similar to that of antibodies. The most
unique aspect of aptamers are the selection process. In 1990, the Szostak and Gold groups
independently described a method for isolating aptamers against specific targets [78, 79]. This
method, called ―i n vitro selection‖ or ―s ystematic evolution of ligands by exponential
enrichment‖ (SELEX), took the core concepts of natural evolution-diversification, selection
and replication. SELEX starts with an initial random pool of (>1010) aptamers of 30-60
nucleotides in length. The pool is incubated with the target molecule and the bound
oligonucleotides are collected and amplified by PCR. The new collection of aptamers
becomes the new pool and process is repeated. In this process, the aptamers with higher
binding affinity and specificity are preferentially enriched. Eventually, one or several
aptamers with the highest binding affinities and specificities will be selected. For biomedical
applications, aptamers are generally modified to prevent nuclease degradation. After
modifications, aptamers are relatively stable over a wide range of buffer conditions and are
resistant to degradation. However, they are still somewhat susceptible to nuclease degradation
and hydrolysis at teh 2‘ OH position [80].
Aptamers have several important advantages as targeting ligands: high binding affinity
and specificity to their targets, versatile selection process, production through chemical
synthesis, low immunogenicity, resistant to physical or chemical degradation, and small
physical size [81, 82]. The production process through chemical synthesis means production
can be scaled up with ease without batch-to-batch variations [83, 84]. This is a significant
advantage over antibody-based targeting ligands. Since the first description of SELEX, more
than 200 aptamers have been isolated [85, 86]. Some examples include RNA aptamers against
the Vascular Endothelial Growth Factor (VEGF)165 isoform and the prostate membrane
specific antigen (PSMA) [87-89]. It has also resulted in FDA approved targeted therapeutic.
Pegaptanib, an aptamer targeted against the VEGF165, was approved by the FDA in
December of 2004 for the treatment of neovascular macular degeneration, underscoring the
rapid progress of aptamers from its original conception to clinical application. Aptamers do
have unfavorable characteristics which include their low serum stability, the need for
chemical modifications, and high production cost.
Small Molecules
Small organic molecules are the smallest targeting ligands. Many naturally occurring
small molecules, such as folic acid, are involved in receptor binding for normal cell functions.
These molecules can be utilized for tumor targeting when cancer cells over express their
receptors. Folic acid is the best example for small molecules targeting tumors. The high
affinity vitamin folate is a commonly used ligand for cancer targeting because folate receptors
(FRs) are frequently over-expressed on tumor cells [90]. Folate specifically binds to FRs
with a high affinity (KD = ~10-9 M) and mediates the process of endocytosis. When
conjugated to drug delivery vehicles such as liposomes, the binding of folic acid to FR
mediates the intrcacellular uptake of the conjugate. Non-naturally occurring small molecules
can also be selected for the targeting of tumor markers. Urea-based molecules have been
shown to bind to prostate specific membrane antigen (PSMA) [91, 92]. PSMA is a well-
established marker for prostate cancer cells. Its expression is primarily prostate specific, with
very low levels seen in the brain, salivary glands and small intestine [93]. The percentage of
prostate cancer cells that express PSMA approaches 100% with highest expression in
androgen-independent prostate cancer cells [94, 95]. The urea-based molecules can not only
bind to PSMA with high avidity and specificity, they can also mediate endocytosis similar to
folic acid. Small size, high stability and production through chemical synthesis are favorable
characteristics of small molecules as targeting ligands. The main challenge for small molecule
targeting ligands is the selection process. For a given cellular target, there is a lack of efficient
and high-throughput methods to identify a small molecule ligand.
NANOPARTICLE PLATFORMS
Liposomes, dendrimers and polymeric micelles have been used for the formulation of
targeted nanoparticle therapeutics. Each nanoparticle platform has unique advantages and
disadvantages as drug delivery vehicles. Their properties are discussed below.
Liposomes
Liposomes have been widely used as pharmaceutical carriers in the past decade, with
eleven formulations approved for clinical use and many more in clinical development. Some
of the commonly used therapeutics include: liposomal amphotericin (Ambisome, Gilead),
liposomal doxorubicin (Doxil, Ortho Biotech), and liposomal daunorubicin (DaunoXome
Gilead). Liposomes are spherical vesicles that contain a single or multiple bilayered
membrane structure composed of natural or synthetic amphiphilic lipid molecules[96, 97].
Their biocompatible and biodegradable composition, as well as their unique ability to
encapsulate hydrophilic agents in their aqueous core and hydrophobic agents within their
lamellae makes liposomes excellent therapeutic carriers. Liposomes can also be coated with
biocompatible and antibiofouling polymers, such as polyethylene glycol (PEG), to prolong
their circulation half-life [97]. Such surface modifications are crucial in generating
biofunctionalized liposomes because the polar heads of the lipids themselves would be a
relatively poor site for conjugation to a targeting ligand. The polymer coating of the
liposomes, on the other hand, can be engineered to carry a functional group which can be
used for targeting ligand conjugation.
A liposomal drug formulation will typically alter the pharmacokinetics and
biodistribution of a drug in a way that may be favorable for a clinical problem. For example,
Doxil greatly reduces the volume of distribution of doxorubicin (from approximately 1000
L/sq m in the free drug form to 2.8 L/sq m) by restricting the distribution within the plasma
only [98, 99]. At higher plasma concentrations, the drug can diffuse into sites of cancer
(through the EPR effect) at much higher concentrations than that of free drug form. On the
other hand, liposomal doxorubicin reduces drug concentration in normal tissues, such as the
heart, resulting in lower toxicity. Doxil also increases the residence time of the drug in the
body (i.e. higher AUC relative to free doxorubicin) by reducing the clearance rate, which
leads to higher therapeutic efficacy [99].
Liposomes have been the popular delivery vehicle for chemotherapeutic agents
principally because of the extensive body of knowledge that exists on their properties (both in
vitro and in vivo), formulation procedures, and chemical modifications. Recent research
interest has been focused on the development of targeted liposomes. Liposomes have been
modified with various targeting ligands, including antibodies, peptides, and small molecules
[100-102]. Other innovations in creating targeted drug delivery using liposomes include the
engineering of pH responsive liposomes [103]. The major shortcomings of liposomes are
their relative instability and their larger size when compared to the other drug delivery
platforms.
Dendrimers
Dendrimers are well defined, regularly branched macromolecules that are 2.5-10 nm in
size [104]. They are synthesized from either synthetic or natural elements such as amino
acids, sugars, and nucleotides. The core of a dendrimer is denoted generation zero and each
additional level of branching is called a generation. Dendrimers‘ favorable characteristics as
therapeutics carriers include nanoscale spherical architecture, narrow polydispersity,
multifunctional surface and large surface area. Over 100 dendrimer families with different
surface modifications have been reported [105, 106]. Among the families, the
polyamidoamine (PAMAM) and poly(propylenemine) (PPI) have been most widely used for
biomedical applications. The specific molecular structure of dendrimers enables them to carry
various drugs through their multivalent surfaces by covalent conjugation or electrostatic
adsorption. Alternatively, dendrimers can be loaded with drugs using the cavities in their
cores through hydrophobic interaction, hydrogen bond or chemical linkage. Their surface can
be engineered to provide precise spacing of surface molecules and to conjugate targeting
molecules.
The preclinical development of dendrimers has been focused largely on forming
dendrimer-drug conjugates [107], synthetic techniques [108, 109], gene delivery [110-114],
and in identifying newer, less toxic materials for the dendrimer scaffold [35, 107, 115].
Relatively early in their development, dendrimers were found to be excellent in transfecting
cells with DNA in vitro [116]. It has lead to the commercialization of a dendrimer-based in
vitro transfection agent, SuperFect (Qiagen). However, dendrimers applied in gene delivery
have been unable to advance towards clinical application largely due to the inherent toxicity
of the cationic dendrimers that were used to complex with DNA. It is believed that the size
and positive charge of the dendrimer act to destabilize the cell membrane [31], leading to
leakage of ions into and out of the cell, and triggers both necrosis and apoptosis. Indeed, use
of any dendrimer-based transfection agent requires optimization to balance efficacy against
toxicity. Reductions in the toxicity of dendrimers are possible by modifying the surface with
PEG or other chemically inert structures [32, 117].
Self-assembling nanoparticles have been made from dendrimer-like materials that are less
toxic than typical dendrimer formulations. These nanoparticles are considerably larger than a
typical dendrimer when complexed with DNA, with a typical diameter being approximately
150 nm. This nanoparticle delivery platform has been shown to deliver plasmid DNA more
effectively than some commercially available transfection reagents [30].
Dendrimers have been developed for a wide variety of indications in medicine, although
none have yet been approved for use as a drug delivery vehicle by the FDA. Dendrimers
themselves have been used as drugs by Starpharma (Melbourne, Australia) in their VivaGel
formulation (a vaginal microbicidal gel), which has advanced through phase I clinical trials in
Australia. Various formulations are currently being explored as drug delivery vehicles; some
of the front runners in commercializing this technology are projecting to be in the clinic by
2009.
Polymeric Nanoparticles
Biodegradable polymer nanoparticles have been extensively investigated as therapeutic

carriers [28]. They are generally formed by the self-assembly of block-copolymers consisting
of two or more polymer chains with different hydrophobicity. These copolymers
spontaneously assemble into a core-shell structure in an aqueous environment. Specifically,
the hydrophobic blocks form the core to minimize their exposure to aqueous surroundings
while the hydrophilic blocks form the corona-like shell to stabilize the core [34]. This
structure provides an ideal drug delivery nanocarrier. The hydrophobic core is capable of
carrying therapeutics with high loading capacity (5-25 % weight). The hydrophilic shell not
only provides a steric protection for the micelle, but also provides functional groups for
further particle surface modifications. Polymeric nanoparticles have been formulated to
encapsulate either hydrophilic or hydrophobic small drug molecules, and macromolecules
such as proteins and nucleic acids [117]. The release of encapsulated drugs occurs at a
controlled rate in a time or condition dependent manner. Furthermore, the rate of drug release
can be controlled by modification of the polymer side chain, development of novel polymers,
or synthesis of copolymers [29]. In general, these biodegradable polymer systems can provide
drug levels at an optimum range over a longer period of time than other drug delivery
methods, thus increasing the efficacy of the drug and maximizing patient compliance, while
enhancing the ability to use highly toxic, poorly soluble, or relatively unstable drugs.
Poly(D,L-lactic acid), poly(D,L-glycolic acid), poly( -caprolactone), and their copolymers at
various molar ratios diblocked or multiblocked with poly(ethylene glycol) are the most
commonly used biodegradable polymers, while PEG is the most commonly polymer used to
engineer the polymeric micelle surface [30].
Biodegradable polymeric nanoparticle micelles have been investigated extensively for the
delivery of hydrophobic drugs. Notably, poly(lactic-co-glycolic acid) (PLGA) has been used
as a material because of its low toxicity and favorable degradation characteristics, which can
provide controlled release formulations for drugs. Recently, PLGA-block-PEG (PLGA-PEG)
has been used as a material for targeting docetaxel, a poorly water soluble drug, to prostate-
specific membrane antigen (PSMA) expressing prostate cancer cells using the A10 anti-
PSMA aptamer as a targeting ligand. The preclinical data showed that intra-tumoral injection
of aptamer-functionalized PLGA-PEG nanoparticles effectively reduced local tumor burden
with improved side effect profiles in mice [118]. In addition, it was found that precisely
engineering the nanoparticle surface to balance steric repulsion effects (due to the PEG) with
cell binding elements (such as the A10 aptamer in this example) is required to produce
maximal accumulation in tumors [33]. In other words, it is important to find the optimal
amount of targeting ligand on the surface of a nanocarrier, since too little does not produce
enough effect and too much probably increases the interaction of the nanocarrier with the
immune system. PLGA-PEG has also been blended with poly-L-histidine-block-PEG, a
polypeptide-PEG conjugate whose side chain has a pKa of 6, which results in acid-sensitive
accelerated nanoparticle degradation [34]. While PLGA microparticle formulations have been
used in humans extensively, biodegradable polymeric micelle nanoparticle formulations are
just beginning to make their way to the clinic. A poly(lactic acid)-block-PEG polymer
nanoparticle formulation of paclitaxel, Genexol-PM (Samyang) is currently being evaluated
in a phase II trial for locally advanced or metastatic pancreatic cancer.
Compared to dendrimers and liposomes, polymeric micelles appear to have good
potential in succeeding in in vivo targeted delivery. Factors contributing to polymeric
micelles‘ potential include their small size, high stability, ease of conjugation to targeting
ligands, the ability to engineer nanoparticles with precise control over targeting ligand
density, and high drug payload. The limitations of polymeric micelles include their relatively
unknown toxicity profile in humans and that it is difficult to encapsulate small hydrophilic
drugs.
CONJUGATION STRATEGIES
The conjugation of targeting ligands to nanoparticles can be accomplished through
covalent or noncovalent reactions. Certain conjugation strategies maybe preferred for a
particular targeting ligand class, such as maleimide-thiol chemistry for peptide ligands. In
general, covalent conjugation strategies are preferred, because of their higher stability in
physiologic salt concentrations. We will discuss the some of the most common conjugation
strategies in this section. For a detailed review of all bioconjugation strategies, please refer to
―Bionc onjugate Techniques‖ by Hermanson [119].
Maleimide-thiol Coupling Chemistry
Maleimide-thiol coupling chemistry is the most common strategy utilized for the
conjugation of amino acid-based targeting ligands, including antibodies, antibody fragments
and peptides. Maleimides (maleic acid imides) are derived from maleic anhydride and
ammonia. They readily react with sulfhydryl groups in pH range of 6.5 to 7.5. One of the
carbons in the carbon-carbon double bond in maleimide undergoes nucleophilic attack by the
thiolate anion, resulting in a stable thioether bond (Figure 1). Although maleimide can cross-
react with amines at higher pH, the reaction of maleimide with sulfhydryl is very specific at
pH around 7. The rate of maleimide-thiol reaction is more than 1000 times that of maleimide-
amine reaction at pH 7.
In general, maleimides are engineered onto the nanoparticles‘ surface through the
incorporation of maleimide modified polymers or lipids into the nanoparticles. Sulfhydryl
groups can be engineered into the targeting ligands by the incorporation of cysteine, which
has a thiol side chain. Aptamers can also be modified with thiol groups at either the 3‘ or the
5‘ end. Because the maleimide-thiol reaction is spontaneous and requires no catalysts, the
reactants (nanoparticles and targeting ligands) can simply be incubated together in a neutral
buffer. During the conjugation reaction, reducing agents such as dithiothreitol (DTT) and
tris(2-carboxyethyl) phosphine (TCEP) should be added to prevent disulfide formation
between the targeting ligands.
O O
+H+
Ligand S N Ligand S N
pH 6.5 - 7.5
O O
Figure 1. Maleimide-Thiol Conjugation Reaction.
Succinimidyl Ester-amine Chemistry
Succinimidyl esters-amine chemistry has been commonly used for the conjugation of
carboxylate groups to amine groups. Carboxylate containing molecules can be converted to
active esters by reacting with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and N-
hydroxysuccinimide (NHS) or sulfo-NHS. The activated succinimidyl ester can then react
with a primary amine to form a stable amide bond (Figure 2). NHS esters are relatively water
insoluble where sulfo-NHS esters are water soluble and hydrolyze slowly in water. NHS
esters have a half-life on the order of minutes to hours under physiological pH conditions.
The rate of hydrolysis increases with increasing pH. In general, to maximize the conjugation
reaction and minimize the effects of hydrolysis, high concentrations of reactants are used for
the conjugation.
For targeting ligand-nanoparticle conjugation, a primary amine or carboxylate is
engineered into the targeting ligand where the other group is engineered into the nanoparticle
surface. The succinimidyl ester-amine chemistry is generally preferred for aptamers and small
molecules such as folate. Aptamers can be easily modified with amines at the 5‘ and/or 3‘ end
where small molecules can contain either primary amines or carboxlates. Folate, for example,
contains a carboxyl group and can be converted to folate-NHS ester.
Avidin-biotin Chemistry
Avidin is a glycoprotein found in egg. It contains four binding sites for biotin (vitamin
H). The avidin-biotin interaction is one of the strongest noncovalent interactions. The avidin-
biotin complex is very resistant to break-down, even at high salt concentrations and extreme
pH conditions. One major disadvantage of avidin is its tendency for non-specific binding to
negatively charged molecules. Streptavidin is a bacterial analogue of avidin, derived from
Streptomyces avidinii. It is less prone to nonspecific binding compared to avidin. The
attachment of streptoavidin and biotin to nanoparticles and targeting ligands requires
conjugation chemistries such the maleimide-thiol coupling chemistry or succinimidyl ester-
amine chemistry mentioned above. The major advantage of using avidin-biotin chemistry is
the high specific and affinity for avidin and biotinylated molecules, even in extreme reaction
conditions.
-H+
+H+
N H
O
H2N Ligand Ligand N
O O
O
Figure 2. Succinimidyl ester-amine Conjugation Reaction.
PRECLINICAL RESULTS
Increased Intratumoral Concentration
Functionalizing nanoparticles‘ surface with targeting ligands can lead to preferential

accumulation of the nanoparticles in the tumors. This has been demonstrated by a number of
preclinical studies. Kukowska-Latallo et al. has shown that folate targeted, fluorescently
labeled PAMAM dendritic polymers concentrated more in KB tumor xenografts in severe
combined immunodeficiency (SCID) mice than that of non-targeted nanoparticles. Our own
experience also showed higher concentration of targeted nanoparticles in tumors when
compared to that of non-targeted nanoparticles. We studied the biodistribution of aptamer
targeted polymeric nanoparticles and non-targeted nanoparticles. We used the A10 aptamer
which is against prostate specific membrane antigen (PSMA), and prostate cancer xenograft
models for our study. We demonstrated that the tumor accumulation for targeted
nanoparticles was 2.5 times higher than that of non-targeted nanoparticles. The tumor
accumulation of targeted nanoparticles is highly dependent on the characteristics of the
targeting ligands and the nanoparticles. As more in vivo studies on targeted nanoparticles
become reported, we will obtain more information on the factors determining tumor
localizations.
Increased Intracellular Uptake
The most significant effect of targeted nanoparticles is increased intracellular uptake by

the target cells. In one study, Kim et al. showed that folate targeted polymeric nanoparticles
had more than 6.7 times cell uptake than non-targeted nanoparticles. Our own data showed
that the A10 aptamer targeted nanoparticles had 77-fold increase in intracellular uptake study
by PSMA expressing prostate cancer cells in vitro when compared to non-targeted
nanoparticles. Oyewumi et al. studied folate targeted polymeric nanoparticles‘ uptake into
KB cells. Folate-coated nanoparticles showed higher uptake in comparison to PEG-coated
nanoparticles. At a nanoparticle concentration of 180 mg/ml, KB cell uptake of folate coated
nanoparticles was 20-fold higher than non-targeted nanoparticles. Kirpotin et al. formulated
anti-HER2 MAb-liposome conjugates to study tumor targeting. In this study, targeted
liposomes had 6-fold higher intracellular uptake when compared to non-targeted
nanoparticles.
Increased Efficacy
Targeted delivery of therapeutics has also been shown to achieve greater efficacy. Park et
al. studied anti-HER2 immunoliposomes encapsulating doxorubicin in tumor xenograft
models. They demonstrated that in four different xenograft models, immunoliposome-dox
was significantly superior to free dox, liposomal dox, and anti-HER2 monoclonal antibody.
Bartlett et al. showed that transferrin targeted siRNA nanoparticle is more effective than non-
targeted siRNA nanoparticles despite similar biodistribution and tumor accumulation of the
two nanoparticles. Using mouse xenograft tumors expressing luciferase and siRNA against
luciferase, they showed that transferring targeted nanoparticles reduced luciferase activity to
50% of compared to non-targeted nanoparticles. Increased efficacy was also seen in the
Kukowska-Latallo study. Folate targeted methotrexate (MTX) lead to statistically slower
tumor growth compared to non-targeted MTX. In our own experience, we have developed
aptamer targeted nanoparticles (NP-Apt) that target the prostate specific membrane antigen
(PSMA) on prostate cancers. Using NP-Apts encapsulating docetaxel and a murine xenograft
model of prostate cancer, the targeted nanoparticles effectively decreased tumor size
following a single intra-tumor injection while non-targeted nanoparticles did not.
CONCLUSION
Advances in biological targeting inspired the development of biofunctionalized targeted
nanoparticles. In the coming years, many biofunctionalized targeted nanoparticles including
targeted polymeric nanoparticles will be entering clinical trials. These nanoparticles will
likely have higher intracellular uptake, higher target tissue concentration, improved efficacy
and lower toxicity compared to non-targeted nanoparticles, making them better delivery
vehicles for therapeutic agents. By optimizing the nanoparticle surface and size, as well as the
targeting ligand, more and better targeted nanoparticle systems will be discovered as well. We
believe that targeted nanoparticles will be become an integral component of cancer treatment
in the near future.
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Lecture Material 19
LITHOGRAPHICALLY-STRUCTURED, BIOLOGICALLY-
INSPIRED, GRIPPING DEVICES
ABSTRACT
Nature is the best engineer on the nanoscale; biology is full of nanoscale functional
machines and large structures that are exquisitely structured on the nanoscale. Some
examples include viruses, organelles, butterfly wings, claws, and sea shells. In each case,
precise nanoscale engineering is used to facilitate remarkable mechanical, electrical or
optical properties that are unparalleled in human engineering. Moreover, human beings
have always been fascinated by the construction of machines that look and behave like
biological ones. In this chapter, we review strategies developed in our laboratory to build
functional structures such as mobile claws and containers that are triggered with
nanoscale thin films. Specifically we describe functional structures in the form of mobile
claws or cages that can be used to capture very small objects including live ones. The
self-closing claws and cages do not need any electrical power or tethers, can be moved
from afar in any arbitrary trajectory and close spontaneously based on thermal or
chemical cues. The ability to trigger machines based on chemical cues is a challenge in
human engineering but is widely seen in nature. Examples include the opening and
closing of flowers, the capture of a pathogen by a macrophage. Hence, we believe the
ability to trigger the functional responses e.g. closing or capture based on a chemical cue
is a first step towards fabricating adaptive machines and ones that respond only to highly
specific chemical cues. The self-closing claws and cages also demonstrate the utility of
self-assembly and biomimicry as an attractive paradigm for enabling remote and
autonomous control of miniaturized machine-based function in human engineered
systems.
INTRODUCTION
From assemblies of molecules too small to see with the naked eye, to the complex tissues
and organs within the human body, nature is without equal, the greatest engineer. Nature has
found a way to create extraordinarily complex structures using simple building blocks.
Additionally, structures fabricated by nature span a range of length scales from the meter to
the submicron, and the processes used to construct them are robust and defect tolerant [1].
Throughout history, scientists and engineers have attempted to mimic these natural
fabrication and design paradigms to engineer more advanced materials, structures and
devices.
In this chapter, we will discuss some technologies that have been developed in an effort
to mimic one of nature‘s simplest machines: a grasping appendage which opens or closes on-
demand. Grasping appendages can be quite varied in form, some more familiar than others
(hands, feet, paws, or pincers), but we will generalize them as any structure that can close
around an object and hold it firmly. We will first review some examples of such devices in
nature, including the Venus flytrap, the aquatic mussel, and human macrophage. We will then
discuss some human-engineered gripping devices fabricated at the sub-mm scale. It should be
noted that while it is relatively easy to construct grippers in the macroscale (we define macro
as larger than 1 mm), it is extremely difficult to construct them at the sub-mm scale. Though
machining of joints, gears, and motors can be achieved down to a millimeter scale with a high
precision multi-axes lathes or machine presses, as we move into to the sub-mm scales,
lithographic fabrication needs to be utilized. However, traditional lithographic
microfabrication methods such as Multi-User MEMS (Microelectromechanical systems)
Processes (MUMPS), bulk micromachining, and LIGA (Lithographie, Galvanoformung,
Abformung) are based on an inherently two-dimensional (2D) lithographic process [2, 3], and
it is extremely challenging to construct truly three-dimensional, let alone patterned, structures
at sub-mm size scales. Assembling such small-scale components into functional devices
precisely is a bigger challenge, and moreover, it is difficult to power sub-mm scale devices.
There are several metrics that can be used to judge the effectiveness of a gripper, or any
device for that matter, at small size scales. The first is specificity – the ability of a device to be
triggered only by a particular cue and perform the designed function. A device which has a
very high specificity could be described as being able to discriminate between target objects
and the surrounding medium. The importance of this trait can be highlighted by the following
example: suppose a device is constructed which can travel through the blood stream and
remove plaque buildup from the walls of the blood vessels. If this machine removes plaque
incredibly well, yet also removes portions of the blood vessel wall in the process, then its
unfortunate lack of specificity would render it unusable in medicine.
The second condition of effectiveness is reversibility – the ability for a device to
repeatedly move between two or more states. This includes both the ability to reverse an
action as well as to repeat it several times before losing functionality. In nature, some
grasping structures are more reversible than others. For example, the human macrophage can
consume over 100 different pathogens before dying [4], whereas the Venus flytrap is only
capable of around 10 closures before it loses its ability for motion [5]. This condition should
not be used as the primary judge of a device‘s effectiveness, but instead as more of an
additional advantage used for judging overall effectiveness. It is also important for an
engineered device, since it is a determining factor in both production costs, as well as patient
usage.
The third condition, and perhaps the most difficult to replicate/fabricate in a laboratory, is
autonomy – the degree to which the device can make decisions, move, and perform its
function without guidance from an external source. This concept, well-developed by nature, is
still far from being fully-implemented in current human-engineered technologies. Most
endoscopic devices currently used in a clinical setting require some sort of tube or tether with
which to power the camera and record images. Though technologies have been developed
which allow devices such as cameras to enter the body without tethers, these devices have no
means yet of self-propulsion and digestive tract probes rely on peristalsis [6]. If an
autonomous device could be developed which housed its own power supply and possessed a
system to maneuver within the human body, the incidence for minimally invasive surgery
needed for most endoscopes would be greatly reduced. These technologies could even
eventually result in the phasing out of more invasive surgical procedures from medicine.
The three criteria of specificity, reversibility, and autonomy are very important from an
intellectual perspective as well as in the commercial/economical aspect of engineering, which
is often ignored when discussing new scientific technologies. Let us take for example, a
hypothetical device which is highly specific and almost fully autonomous, yet lacks any
reversibility and must be discarded after a single use; if the cost of mass-producing each unit
is too great, then the new technology may experience slow acceptance, if any at all, as doctors
and patients are resistant to investing time and money into unproven technologies.
In order to better understand how the three criteria can be used to judge emerging
technologies, we will first use them to analyze the functionality and effectiveness of
structures fabricated by nature. By starting at the large-scale with the Venus flytrap and the
aquatic mussel, then moving down to the microscopic macrophage, we will see how
characteristics and functionality change as we move down the size-scale.
MACROSCOPIC CLAW-LIKE MACHINES

CONSTRUCTED BY NATURE
The Venus Flytrap
Dionaea muscipula, more commonly known as the Venus flytrap, is one of the few plants
with the capability for rapid response. The ―t rap‖ portion of the plant is actually composed of
the terminal lobes of the leaf. These lobes are hinged along the midrib to create two grasping
structures that clamp down and capture prey for digestion. In light of the criteria previously
discussed, we will discuss the merits and disadvantages of the Venus flytrap as a gripper.
The lobes of the Venus flytrap are stimulated to close by a series of projections inside the
trap opening called trigger hairs [7]. Each hair-like trichome is a hinged structure that, when
touched under certain conditions, will cause the trap to quickly close (around 100
milliseconds) [8]. The trap is specific in that the Venus flytrap must distinguish between
insects (prey) and rain/debris (non-prey) which may fall into the opening of the trap. In order
to increase specificity, the Venus flytrap has evolved so that the trap will not begin to close
unless two trigger hairs are simultaneously stimulated, or a single trigger hair is stimulated
more than once in rapid succession [7, 9]. If two trigger hairs happen to be touched by falling
debris, the trapping mechanism will begin to close, but not fully to its sealed state. The trap
requires continuous stimuli (provided by a trapped insect attempting to escape) to the trigger
hairs in order to close completely. Though not perfect, this system allows the plant to
discriminate between live prey and other debris, and it is fairly specific in its function.
An exact explanation of the mechanisms involved in the trapping of prey is not fully
known, but it is generally accepted that the geometry of the leaf is the main trait enabling
rapid closure of the Venus flytrap [8, 10-12]. While in the open state, the leaf lobes of the
Venus flytrap are curved outwards, exposing the maximum amount of inner surface area and
allows insects a greater chance of stimulating the trigger hairs. It is believed that during this
open state, a great amount of stress is placed on the leaves due to the curvature and cell wall
strain. This results in a rapid conformation change when the leaves are triggered, analogous to
when a spring-loaded mouse trap snaps shut when the triggering bar is knocked loose. This
relatively simple concept could be used by researchers to create microdevices that close with
great speed, yet only use energy mechanically stored in spring-like structures.
Figure 1. Optical image of Venus flytrap leaves. Each open trap is held at high tension, such that when
triggered, the geometry of the leaf causes the two halves to close in approximately 100 milliseconds.
In terms of specificity, the Venus flytrap performs its task adequately. Although lacking a
centralized brain or processing unit, it is able to take stimuli from the trigger hairs and
transfer that information to the leaves, which in turn close around and trap the prey. This is an
example of how a device can be built that is not reliant on a processing microchip to perform
a rather complicated function. Simplicity in this case, results in a system which performs
effectively.
The Venus flytrap is able to reuse its trapping mechanisms over a long period of time, but
they do wear out. Also, the trap needs time to reestablish cell wall tension. Partial closures
caused by falling debris require around 12 hours before returning to the fully open state [5],
which means that there is a relatively long period of time associated with a failed trapping
attempt. After about ten to twelve closures, whether partial or complete, the trap will cease to
function as a prey catching mechanism and revert to photosynthesis [5] for the plant‘s energy
needs. Although the traps do eventually lose their mechanical proficiency and have their
efficiency reduced by a long reset time, the Venus flytrap is capable of reversibility.
Regarding autonomy, while the trapping mechanism of the Venus flytrap is nourished
and supported by the plant body and root system, the actual mechanics and trigger systems
are self-contained within the trap. Since the plant has no nervous system, the trap can be
considered a single autonomous unit, able to function without the explicit control of another
source.
Bivalve Mollusks
Bivalves, which include clams, mussels, oysters, and scallops, are a very large and
diverse portion of the mollusk phylum, but are classified based on the presence of two
symmetric valves on either side of the hinged shell line. These aquatic animals are composed
of two shells which can be opened and closed for feeding or defense, and they can range in
size from the meter long Giant Clam (Tridacna giga) [13] to the sub-centimeter Pea Clam
(Pisidium nitidum)[14]. Across this wide range of sizes, mollusks share common mechanisms
to open and close their shells. For the purposes of this chapter, we will focus on the relatively
common freshwater mussel (Subclass Palaeoheterodonta) as an example. This organism has
become both a staple of human seafood consumption, as well as a nuisance to parts of the
Laurentian Great Lakes of North America, where Zebra mussels (Dreissena polymorpha)
have caused problems with sewage and shipping around the area [15, 16].
Unlike the Venus flytrap where the trapping component was only an element of the entire
plant, the mussel‘s entire structure is encased within its shell. This means that complexities
that are present in the larger versions of the organism are still present in the small, such as the
muscles, sensory organs, and nervous system. Mussels have sensory organs that are present
around the margins of the mantle, capable of responding to chemical and pressure [17, 18].
Although the nervous system is considered primitive in that there is no brain, it uses this
simple system in conjunction with its sensory organs to respond to stimuli, such as closing its
shell when the tide goes out. [Figure 2]
Figure 2. Optical image of freshwater mussels. The two shell halves are held closed with great force using
only a few muscle groups. When inundated with water, the mussels will open and release filters with which
they feed.
The way in which the mussel shells are designed allows for the animal to open and close
its shells with such force that many predators cannot penetrate it. This design has high
reversibility, since it relies on only a few groups of muscles to actuate the shells together [19].
By minimizing the number of moving parts, the two shells can be opened and closed almost
indefinitely as long as the animal is alive. The placement of the muscle groups near the ends
of the shell provides a large amount of torque which requires an incredible amount of force to
pry open. However, the mussel trades off opening size and angle for this strength, since the
two shell halves are designed for filter feeding, not for wide openings.
Considering that the entire organism is encased inside its two shells, this system would be
perfectly autonomous if replicated in human-engineered form. It would be a great
achievement if engineers could recreate everything a bivalve contained within its shell, from
the chemoreceptors to the muscular foot it uses for propulsion. The mussel‘s foot is a
particularly interesting device to study, mainly because of its ability to serve multiple
functions while remaining simple in concept. As a propulsion device, the muscular foot is
able to move the animal through various types of media present on the sea floor, such as sand
or gravel. To do this, the mussel projects the foot forward, then uses it to pull the rest of the
body along, very much like how an earthworm moves through soil [19]. Secondly, the foot
acts like an anchor for the mussel once it has reached a satisfactory feeding location. By
creating a vacuum with the foot and then excreting byssus, a protein adhesive, into the
vacuumed space under the foot, the mussel is able to create an incredible bond with the
surface[19, 20].
If biotechnology could develop a device inspired by the mussel and its foot, small enough
to enter the human body, then the unit would able to reach areas of the body which would
normally only be accessible through surgery. Imagine a microdevice that can travel through
the large intestine and attach to the intestinal wall using some kind of organic adhesive. Once
there, this device is able to extend small probes into the internal fluid to scan for particulates,
much like the mussel extends filters to collect food particles. This device could collect
information from its sensors and transmit them to an outer unit using radio waves, providing a
constant data stream for as long as needed. Or perhaps if this device were sufficiently
miniaturized, the mussel-inspired machine could attach to the wall of a blood vessel, using its
sensors to constantly monitor the blood sugar levels of diabetes patients. This method would
provide constant and immediate information, with the ability to detect blood sugar drops
within seconds, along with the benefit of reducing or removing the need for blood sampling
tests.
Macrophages
As we move down the size scale to the cellular level, function analogous to grasping can
be carried out in completely different ways. There are situations on the cellular level that
require the ability to capture or encapsulate objects to isolate or destroy them. At the
macroscale, as we have already discussed, this is often done using gripping appendages such
as hands, traps, or shells. At the microscale, however, the most common method of capturing
objects is through phagocytosis [4]. Macrophages encapsulate objects with their cell
membrane into internal vacuoles, and are thus able to perform the same actions which we
have used to define a gripper, but at the cellular level [21]. [Figure 3]
One of the main functions of macrophages is to ingest and destroy pathogens which enter
the body [22]. In the human immune response system, the macrophage responds to antigens
which are present on the surface of pathogens. The macrophage destroys the pathogens it
engulfs by fusing a lysosome filled with digestive chemicals with the vacuole containing the
pathogen. Once broken down, the contents of the vacuole are discarded by the macrophage by
fusing the vacuole with its cell membrane. In order to facilitate a better downstream seek and
destroy process of these pathogens, however, the macrophage requires help from antibodies
and helper T-cells. When a certain pathogen is digested by the macrophage, the antigen which
was located on the pathogen surface is presented to the helper T-cell. In turn, the helper T-cell
produces antibodies which attach to the antigens on that type of pathogen‘s surface, enabling
a quicker response for future macrophage phagocytosis [23]. This system of positive feedback
gives the macrophage very high specificity, since it is able to further target and digest specific
cells once antibodies have been released.
A macrophage, like any living organism, cannot function indefinitely. The lifespan of a
macrophage is often decided by the number and type of pathogens that it ingests. The typical
macrophage is capable of ingesting more than 100 bacteria before succumbing to its own
digestive enzymes [22]. This translates to high reversibility, as any humanly engineered
device which was capable of functioning 100 times at such a small scale would be considered
quite reversible [24].
Figure 3. Plate-based phagocytosis assay. Phagocytosis of apoptotic human neutrophils (arrowheads; stained
brown) by adherent human monocyte-derived macrophages (unstained) [37].
Autonomy is something we always strive to emulate in engineering, especially on the

cellular level. Although the microphage requires help from other cells and proteins to help the
immune system perform optimally, it is still able to judge pathogens on an individual basis
initially [25]. Later in this chapter, we will discuss human engineered devices that, while not
as specific as biological systems, can also use chemical cues to trigger functional changes. By
taking inspiration from these organisms, cells, and systems, researchers hope to improve upon
technologies to better suit medical and scientific problems.
BIOLOGICALLY-INSPIRED DEVICES
We have discussed organisms and organic structures which have different ways of
performing the same general function. An approach to developing better human-engineered
structures that mimic a natural function is to first understand how it happens in nature. It is
important to keep in mind that there are many different ways to approaching a problem such
as this, as it is impossible to predict the optimal method.
Currently, medical techniques are often constrained by the size of the instrument.
Biopsies for example, are a medical procedure requiring doctors to enter the body with the
intent of retrieving a specific cluster of cells. With current technologies, the easiest way to do
this is to insert endoscopes into the body and take a small amount of tissue using an apparatus
at the end of the device. Although less invasive than most surgeries, it still has many
drawbacks. Commercial biopsy forceps that are used for endoscopic tissue sampling have a
usual diameter of 5 to 10 mm[26], which is still not small enough to get into some of the
more intricate organs and tissues deep within the human body. Though minimally-invasive,
biopsies currently still can involve minor surgery and are not immune to some of the same
hazards faced in surgery, such as infection. Some organs, such as the pancreas, can be
reached by endoscopes, but the area is too convoluted to allow a tethered device to be pushed
into the organ without causing damage. Also, endoscopies often rely on an existing body
cavity through which to thread the endoscope, which means portions of the body that are not
in the vicinity of major blood vessels or the intestinal track are often not accessible with
traditional endoscopic methods.
Capsule endoscopy is a newly-developed technology which tries to remedy some of these
problems [6]. By fitting a small camera in a pill-sized capsule, doctors are able to image the
entire digestive track by having their patients swallow the device. Though this does facilitate
imaging of the gastrointestinal tract better than a traditional endoscope, it does not help the
patient who is in need of a biopsy. In order to remedy this, a gripping device of sufficient
specificity to locate the specific tissue of interest and autonomy to perform its job without the
need of tethers must be developed [27]. If a device small enough were to be created with
these traits, many procedures which currently require minimally-invasive surgery could be
performed with by less invasive methods.
HUMAN-ENGINEERED DEVICES
There have been several devices developed by researchers and scientists around the world
which attempt to emulate many of nature‘s devices and have the ability to open and close
around an object. The desire is often to make them smaller and precise enough to hold and
manipulate single cells. Additionally, creating autonomous devices that can be manipulated
remotely without tethers would allow for greater freedom of movement. With small enough
devices, these traits could allow for applications inside the body and may reduce the number
of surgical procedures by removing those which can now be solved through other methods.
Several new or developing technologies are presented here, but the list is by no means
inclusive of all the ideas being pursued at this time. These example devices are discussed to
describe a sampling of the different paths which various researchers have taken to enable
microscale manipulation. It should be noted that many of these devices have been fabricated
for in vitro use.
Manipulation of single cells has always been difficult, due to the small size of cells, their
fragility, as well as the relatively large tools which are available. Traditional systems use a
suction pipette to hold the cell in place, while a needle from a secondary system is used to
conduct the procedure, whether that is DNA injection or nuclear extraction [28]. This system
has several shortcomings, mainly stemming from the fact that it is incredibly difficult to
accurately manipulate the cell. Technicians must be formally trained and must practice for
many months before becoming adept. To address this limitation, a device has been developed
by researchers at Osaka University in Japan to enable easier cell manipulation [29].
The researchers were inspired by the basic mechanics of chopsticks. Called the ― Two-
fingered micro-hand,‖ the device is able to grasp, rotate, and move with nanometer precision
by using two structures which are manipulated like chopsticks [30]. This system utilizes a
system of actuators which all manipulate a single plate. This plate is called a parallel
mechanism, has high accuracy and precision because of the stiffness of support given by the
three actuators. The use of three actuators also allows for movement with 3 degrees of
freedom (DOF). Two of these parallel actuator plates make up each micro-hand, one for each
finger, allowing for a total of 6 DOF. When two micro-hands are used in conjunction, the
same tasks which were difficult to do with the pipette and needle method are now much
simpler. This device presents several advantages over the current systems available. Because
of its intuitive nature, there is much less training time required for technicians to become
fluent in manipulating cells, improving productivity and cost effectiveness. Also, the
precision of the device is much better than previous systems [30], since it allows for
manipulation on the nanoscale. The device‘s selectivity is improved compared to current
devices, such as the pipette and tweezers system, since it can resolve objects and manipulate
them at the nanoscale[30]. Its reversibility is also very high, since the ―chops tick‖ mechanism
is something easily manipulated around a multitude of objects and can be done so repeatedly
without wear on the device. Its autonomy, however, is something which limits its uses in the
medical setting. Since this is a device meant for in vitro applications, it does not solve the
problem of how to create a device that can help reduce the need for minimally-invasive
surgery in certain medical circumstances. Since the device is not built for in vivo applications,
there is a need for a device which has the same small-scale capabilities as this device, but is
able to enter the body without the need for tethers.
Sometimes the most innovative solutions are not the ones that create entirely new
materials, but rather use existing materials in new ways. Researchers at the UCLA
Department of Bioengineering followed this train of thought by harnessing the power of
human cardiac muscle cells in order to create a functional micromachine [31]. Millions of
years of evolution, they reasoned, had produced a product which was fine-tuned for motor
contraction, so it was an obvious choice to use these cells in their research. Since there was no
defined way to produce freely-moving structures which used cardiac muscle cells at the
time[31-33], the researchers created a new approach: producing a gold structure which could
bend and flex with the contractions of muscle cells. To do this, they first created a structural
cantilever and support from which to build the device out of silicon dioxide and a silicon
substrate. This structure was then coated in poly-N-isopropylacrylamide (PNIPAAm) and a
thin layer of gold (Au) was deposited. Since cardiac muscle cells bind to Au much better than
to PNIPAAm, the functional bundle of muscle cells grew above the Au layer[33] [Figure 4].
Once the cells matured and the supporting structure was dissolved and removed, the device
was released. The resulting free-moving structure opened and closed as the muscle fibers
were stimulated. However, there are several things that still need to be accomplished in order
to design a device that satisfactorily meets all of the criteria set forth at the beginning of this
chapter. First, although these structures are not tethered and can be triggered remotely, there
is no way to direct their motion within a defined space. In order to be useful in medicine,
many hundreds of these devices would have to be introduced into the body in order to
increase the probability of the random encounters with the target. Also, since this device
requires the use of a biological component, there are compatibility issues regarding in vitro
use; there is the possibility that the device can create an allergic reaction in the patient, or that
the patient‘s immune system will destroy the device before it has performed its function.
Finally, though this device can grasp using muscle contractions, there is no functional way to
hold a contracted state, since the cardiac muscle fibers are only designed for quick pulses of
force. Holding a closed state with this system may cause tearing or other damage to the
muscles that would render the device unusable. Remedying this problem requires thoughtful
analysis and experimentation of different ideas, such as using different types of muscles to
power the device, or using two groups of muscles together, contracting and relaxing much
like muscle groups work in the human body.
Figure 4. Microscopic images of a single half-circle muscle bundle with the plane of the gold film oriented
perpendicularly to the underlying substrate with a thickness of 29.1 ± 2.7 μm. The circled area of the
muscle/gold is also redrawn to better show the interface between the muscle and the underlying gold film
[31].
Figure 5. Scanning electron microscope (SEM) image of a tetherless microgripper. The microgrippers are
triggered en masse using relatively benign cues, such as small temperature changes or chemical cues.
TETHERLESS, THERMO-CHEMICALLY
ACTUATED MICROGRIPPERS
This section describes recent work performed in our own laboratory [34-35]. In order to
mimic as closely as possible the features of many small-scale natural grippers, we developed
a process to mass-produce tetherless microgrippers that could be remotely triggered en masse
by temperature, as well as chemical cues, under biological conditions. The microgrippers
utilized a self-contained actuation response, removing the need for external tethers in
operation, while still retaining the ability of en masse actuation even when spatially separated.
The microgrippers were demonstrated to perform diverse functions, such as picking up beads
on a substrate and removing cells from tissue embedded at the end of a capillary (an in vitro
biopsy). Our work was inspired by the observation that autonomous microscale function in
nature is often achieved by organisms and cellular components that are triggered en masse by
relatively benign cues, such as small temperature changes and biochemicals. The use of
chemical actuation is quite different from any other demonstrated work with gripping devices
and could enable the high specificity and selectivity observed in biological machinery.
The microgrippers were remotely actuated when exposed to temperatures above 40°C or
selected chemicals. The temperature trigger is in the range experienced by the human body at
the onset of a moderate to high fever, and the chemical triggers include biologically benign
reagents, such as glucose and cell media. The microgripper digits were inspired by the
dicondylic joints of arthropods [36], designed as a series of rigid nickel (Ni) and Au
phalanges interconnected by hybrid organic-inorganic flexible trilayer joints. To remain
tetherless, the devices utilized an actuation mechanism that utilized joints composed of a
polymer trigger and a stressed bimetallic thin film patterned between rigid phalanges. The
polymer layer enabled triggered control over the closing of the gripper. Another favorable
trait of the device was the use of ferromagnetic Au-coated Ni which enabled remote
manipulation through the use of magnetic fields. This remote manipulation capability
allowed the devices to be guided through tortuous channels and to targets of interest, such as
a bead or cluster of cells. The devices could then be remotely-triggered to close around their
target and subsequently retrieved magnetically. To explore the utility of their gripper in
microsurgical applications, an in vitro biopsy was performed on a tissue sample from a
bovine bladder loaded into a glass capillary tube. The presence of connective tissue in the
bladder necessitated using the manipulation magnet to also rotate the gripper such that the
claw phalanges could cut through the connective tissue and extricate the cells.
Since this microgripper was capable of being positioned using magnetic fields as well as
triggered by temperature and chemical signals, the microgripper was able to be directed to
and obtain cells from very specific locations. This specificity, however, was reliant on how
the device was positioned, as the gripper could indiscriminately close once triggered. With
this system, care must also be taken to prevent unintentional triggering of the microgripper by
a chemical cue which is similar to the one generally used to trigger the gripper. Also, in its
current form, the device unfortunately has no reversibility. Once closed, the contents of the
grippers can only be biocompatibly-retrieved by mechanical disruption. After retrieving
microgrippers with captured contents, the grippers need to be mechanically agitated using a
variety of methods (including force applied via Pasteur pipette tip, prying open with 22G
syringe tips, and overtaxing) to release the cells. However, under nonbiological conditions, a
series of chemicals could be used to close and open the grippers for one cycle [35]. Thus, the
grippers are limited to use in single-function applications. However, since the grippers are
fabricated en masse and are inexpensive, this lack of reusability and reversibility is not a big
concern. Still, we are continuing to explore methods to add a greater degree of reversibility,
especially under biologically-compatible conditions.
While not strictly self-autonomous in the sense that the method of propulsion is not
internalized and requires the use of an external, actively-manipulated magnetic field actively,
the microgripper is very adept in moving through body cavities without having the need for
tethers to control the triggering or movement. The use of magnetic propulsion solves many of
the problems that come with internalized power supplies, the most obvious being weight and
size constraints.
CONCLUSION
In this chapter, we have discussed how gripping devices in the biological world have
served as inspiration and as a starting point for the engineering technologies being developed
today. The point we wish to make with the analysis of these devices is that seemingly trivial
tools present in both nature and technology are no longer trivial at smaller size scales; tasks
easily performed with mechanical tweezers on the macroscale suddenly become quite
challenging at the microscale. Determining, recreating, and successfully implementing the
ways nature has been able to overcome these challenges is something that we, as engineers,
continue to strive to do.
One promising direction that needs to be pursued is to construct hybrid systems i.e. those
that utilize both human engineered and biological components. In our own laboratory we are
investigating tethering molecular motors such as the F1 subunit of Adenosine-5'-triphosphate
(AT -shaft can spin counter-clockwise
(when viewed from above) at 150Hz at saturating ATP concentration. There is also the
possibility of tethering microorganisms such as bacteria to aid in autonomous propulsion.
These ideas are novel and considerable challenges need to be overcome before they can be
implemented. Even if it is possible to construct such devices, several important questions
need to be answered before they can be implemented in vivo. How will the devices be
retrieved? How does one prevent an immune response? What about side effects? All these
questions are important and form the basis of a new and rapidly developing field of
―nanom edicine,‖ and there may come a day when we might be able to swallow or inject
miniaturized surgical tools with grasping appendages into the body.
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Topic page
Lecture 20: Protein engineering tools for interfacing proteins and solid supports with exquisite
chemical control 481
Lecture 21: Bioinspired colloidal systems 503
Lecture 22: Bacilli, Green Algae, Diatoms and Red Blood Cells
How nanobiotechnological research inspires architecture? 519
Lecture 23: Biopolyelectrolyte multilayer microshells: Assembly,property and application 554
Lecture 24: Synthesis and Electron Field Emission from Different Morphology Carbon
Nanofibers 574
Lecture 25: Carbon Nanotubes: A New Alternative for Electrochemical Sensors 635
Concluding Remark 681
Lecture Material 20
PROTEIN ENGINEERING TOOLS FOR INTERFACING

PROTEINS AND
SOLID SUPPORTS WITH EXQUISITE
CHEMICAL CONTROL
ABSTRACT
Immobilization of proteins onto surfaces is of great importance in numerous
applications, including protein analysis, drug screening, and medical diagnostics, among
others. The success of all these technologies relies on the immobilization technique
employed to attach a protein to the corresponding surface. Non-specific physical
adsorption or chemical cross-linking with appropriate surfaces results in the
immobilization of the protein in random orientations. Site-specific covalent attachment,
on the other hand, leads to molecules being arranged in a definite, orderly fashion and
allows the use of spacers and linkers to help minimize steric hindrances between the
protein and the surface. The present paper reviews the recent development of new
chemical and biological technologies for the site-specific immobilization of proteins onto
inorganic materials and their potential applications to the fields of micro and
nanotechnology.
Keywords: Protein immobilization, Native Chemical Ligation, Protein Splicing, Protein

Chips, Staudinger Ligation, Click Chemistry, capture-ligand immobilization
INTRODUCTION
Nano-biotechnology is an exciting emerging field that lies at the interface of recent
advancements in nanoscale science/technology and biotechnology [1]. The biological,
biomedical, and medical applications of nanotechnology are some of the most promising,
most exciting, and potentially most rewarding. Sensing and therapeutics using tools from
micro and nanotechnology [2], the use of nanoscale drug delivery devices for targeted therapy
[3], development of devices using micro-nano fabrication and scaffolding techniques [4], and
miniature drug screening and discovery [5] are only some of the rapidly emerging
possibilities.
The key for the success of all these applications relies, however, on interfacing the
typically ―s oft‖ biomolecules found in biological systems (i.e., mainly proteins and DNA)
with the generally ―har d‖ inorganic materials found in micro and nanotechnology, ensuring
that the interfaced biomolecules retain their remarkable biological properties. Although there
has been an enormous amount of progress made in interfacing/immobilizing DNA
biomolecules onto different inorganic materials [6], the immobilization of proteins has been a
challenging task. This is mainly due to the heterogeneous chemical nature of protein surfaces
and the marginal stability of the native active structure over the denatured inactive random
coil structure.
Most of the available methods for the immobilization of proteins onto inorganic materials
have relied on the use of non-specific adsorption of proteins or on the reaction of naturally
occurring chemical groups within proteins (mostly amines and carboxylic acids) with
complementary reactive groups chemically introduced onto the corresponding inorganic
material [4, 7, 8]. In both cases, the corresponding proteins are attached onto the surface in
random orientations, which may cause the loss of the protein‘s biological activity [9]. The use
of recombinant affinity tags addresses the orientation issue. However, in most cases, the
interactions of the tags are reversible and unstable over time [10-14].
Site-specific covalent immobilization (Figure 1), on the other hand, allows the proteins to
be arranged in a definite, controlled fashion. The reaction between these two groups should be
highly chemoselective, thus behaving like a molecular ‗Velcro’ [15, 16]. Finally, the use of
hydrophilic spacers and linkers may also help to minimize the potentially detrimental
interaction between the protein and the inorganic support.
The scope of this work is to review some of the most commonly employed methods as
well as the latest developments for the chemoselective immobilization of biologically active
proteins onto inorganic supports.
Figure 1. General concept of a chemoselective reaction between a protein and an appropriately modified
surface.
IMMOBILIZATION OF PROTEINS ONTO

INORGANIC SUBSTRATES
Most methods used for chemoselective immobilization of proteins are based on ligation
methods originally developed for the synthesis, semi-synthesis, and selective derivatization of
proteins by chemical means (see Table 1) [16]. All these methods involve the derivatization
of a protein with a unique chemical group at a defined position, which will later react
chemoselectively with a complementary group previously introduced into the inorganic
surface.
Surface Modification
Silicon, metals (mainly Au and Ag) and semiconductor (i.e., Ag2S, CdS, CdSe, and TiO2)
based substrates are among the most common materials used to immobilize proteins in micro-
and nano-biotechnology. Si-based substrates are usually modified using derivatized
trialkoxysilanes such as (3–aminopropyl)-trialkoxysilane (APS) or (3-mercaptopropyl)-
trialkoxysilane, which are able to introduce an amino or thiol groups, respectively. These
functionalities can be further chemically modified to introduce appropriate linkers where the
proteins can be covalently attached in a chemoselective fashion.
Table 1. Summary of different chemoselective methods commonly used for the site-
specific immobilization of proteins onto inorganic substrates
See reference [16] for a more detailed review.
Sulfur- and selenium-containing compounds can also be used for the modification of
substrates based on several transition metals (Au, Ag and Pt) [17, 18] or semiconductor
materials (e.g.., Ag2S, CdS and CdSe) [7]. The most studied system, however, is the use of
alkanethiols on gold surfaces. Chemisorption of alkanethiols as well as alkyl disulfides on
clean gold gives rise to similar levels of surface coverage, although thiols react faster than
disulfides [17, 19]. Alkylaminothiols are the most common species used for the preparation of
functionalized gold surfaces. Among them, cysteamine (HS–CH2–CH2–NH2) is probably the
most used [20-23], in part due to its availability. Mercaptoalkyl carboxylic acids (HS–(CH2)n–
CO2H) can also be used to introduce a reactive group onto the gold substrate [7, 23].
In our group, we have developed a new efficient synthetic solid-phase scheme for the
rapid generation of modified alkanethiols (Figure 2) [24, 25]. We have used this approach for
the chemical synthesis of different modified alkane thiols that were successfully used to
immobilize different biological functional proteins onto Si-based and Au surfaces [24-26].
Figure 2. Synthetic scheme for the rapid and efficient preparation of chemically modified thioalkanes [24-26,
82]. The solid ellipsoid represents the different molecular functionalities that can be incorporated in the
thioalkane for surface modification.
IMMOBILIZATION OF THIOL-CONTAINING PROTEINS

Cysteine is the only naturally occurring amino acid containing a thiol group in its side-
chain, and its relative abundance in the average protein is rather small (< 2%). Thiols have a
pKa of around 8.5, and they are nucleophilic at pH 7. Under these conditions, they can be
reacted with high selectivity with some chemical groups like -haloacetyl and maleimide-
containing compounds to form a stable thioether bond. There are a great variety of
commercially available reagents that can be used for introducing maleimide as well as
iodoacetyl groups on amine-derivatized inorganic supports [7, 24-29]. This provides a unique
window of reactivity for the chemoselective attachment of proteins to surfaces through the
thiol group of the amino acid Cys. The only requirement to have control over the orientation
during the protein immobilization process is the presence of a unique and reactive Cys residue
on the protein. Reactive Cys residues should not be involved in any structural element and
should be exposed in a solvent accessible region of the protein. In the absence of an
endogenous Cys residue, it will be necessary to introduce a Cys residue through mutation of a
native residue. The effect of this Cys mutation on the structure and function of the protein can
be easily minimized by following a few simple rules:
(1) If the tertiary structure of the protein is known, choose a region remote from the
active site of the protein, preferably in a flexible surface loop.
(2) Choose the mutation to be as conservative as possible, e.g., Ser → Cys or Ala →
Cys.
(3) Avoid mutation of residues that are conserved across a gene family.
(4) Take advantage of any known mutational data on the system since the effect on
structure/function of a specific residue mutation may already be known.
In our laboratory, we used a genetically modified Cow Pea Mosaic Virus (CPMV) to
create assembled viral nanostructures on maleimide-containing surface templates [24]. In this
work, the CPMV was genetically engineered to present Cys residues at geometrically
equivalent positions in the solvent-exposed E- F loop of the viral capsomer [30]. The
maleimide containing surface templates were created by microprinting techniques and
scanning probe nanolithography on a gold-coated mica surface using a PEGylated amino thiol
linker. The amino group was then reacted with N-(maleimido-propionyloxy)-succinimide
ester (MPS) to yield the corresponding maleimide-containing surface template.
As shown in Figure 3, the Cys-mutated CPMV was chemoselectively attached only on
those areas containing the maleimide function. Mirkin and co-workers [31] have also used a
similar scheme to template viral deposition on chemically modified surfaces.
The use of the Cys thiol group is probably the most common method used by the
scientific community for achieving chemoselective immobilization of proteins and viruses
onto inorganic supports (reviewed in detail in [16]). It should be noted, however, that is not
general method and can only be applied when there is only one reactive Cys residue, either
artificially introduced or naturally occurring, on the protein to be attached. When the protein
contains multiple Cys residues, it is better to use alternative chemoselective methods.
Figure 3. (A) Chemoselective attachment of genetically modified CPMV virus with unique cysteine residues
(Cys-CPMV). (B) Atomic force microscope (AFM) height image of Cys-CPMV virus assembled on a
micron-sized template containing the maleimide function. (C) AFM height image of a monolayer-thick of
virions assembled on a parallel line pattern created by nanografting with a maleimide-containing linker [24].
PROTEIN IMMOBILIZATION USING

EXPRESSED PROTEIN LIGATION
One of the most efficient ways to site-specifically immobilize biologically active proteins
onto solid supports is by using Expressed Protein Ligation (EPL) [32-35]. Key to this
approach is the use of protein -thioesters (see Table 1), which can be efficiently attached to
surfaces containing N-terminal Cys residues through Native Chemical Ligation (NCL) as
shown in Figure 4 [36, 37]. In this reaction, two fully unprotected polypeptides, one
containing a C-terminal α-thioester group and the other an N-terminal Cys residue, react
chemoselectively under neutral aqueous conditions with the formation of a native peptide
bond at the ligation site.
Figure 4. Principle of Native Chemical Ligation [36, 37].
We recently immobilized several biologically active proteins onto modified glass

surfaces through their C-termini using this approach [25]. In this work, two fluorescent
proteins (EGFP and DsRed) and a SH3 domain protein C-terminal -thioesters were readily
expressed in E. coli, using an intein expression system [38]. The -thioester proteins were
then immobilized onto a N-terminal Cys-containing glass slide. The chemical modification of
the glass slide was accomplished first by silanization with (3-acryloxypropyl)-
trimethoxysilane and then reacting with a mixture of PEGylated thiol linkers 1 and 2, shown
in Figure 5, in a molar ratio of 1:5, respectively. Linker 1 contained a protected N-terminal
Cys residue for the selective attachment of the -thioester proteins. Linker 2 was used as
diluent to control the number of reactive sites on the surface. Linker 1 contains a longer PEG
moiety than linker 2 to ensure that the reactive Cys groups were readily available to react with
the corresponding protein -thioester in solution. When the derivatization was complete, the
corresponding protecting groups of the Cys residue from linker 1 were removed by a brief
treatment with trifluoroacetic acid. The surface was rinsed, neutralized, and quickly used for
micro-spotting (Figure 6). The ligation reaction was kept in the dark at room temperature for
36 h, and the protein-modified slide was then extensively washed. As shown in Figure 6, only
specific attachment between the -thioester proteins and the Cys-containing glass surface was
observed. No fluorescence signal was observed where the control EGFP protein lacking an -
thioester was spotted. It is interesting to note that the immobilized DsRed protein, which only
has red fluorescence as a tetramer, retained its red fluorescence thereby indicating that its
quaternary architecture was unaffected by the attachment to the PEGylated glass surface.
Figure 5. (A) Site-specific attachment of a protein -thioester through its C-terminus. (B) PEGylated thiol
linkers used to attach C-terminal -thioester proteins onto acryloxy-modified glass surfaces [25].
Figure 6. Selective attachment of EGFP (green) and DsRed (red) -thioesters onto a Cys-containing glass
slide [25]. Epifluorescence image of the glass slide after the protein micro-spotting (top) and after PBS
(phosphate buffer solution) washes (bottom). EGFP with no -thioester was used as control. Spotting was
carried out using 100 µM protein solutions. Dot size is 100 µm.
Yao and co-workers have also used NCL and EPL, for the selective immobilization of N-
terminally Cys-containing polypeptide [39] and proteins [40] onto -thioester coated glass
slides. In this case, the polypeptide/proteins are site-specifically immobilized through their N-
termini, which may be convenient in cases where the C-terminal immobilization, described
earlier, affects the activity of the protein.

THE STAUDINGER LIGATION REACTION
Azido-containing proteins can be chemoselectively immobilized onto solid supports

modified with a suitable phosphine via a modified version of the Staudinger ligation reaction
[41-44]. This reaction allows the formation of an amide bond between an arylphosphine
moiety and azide group (Figure 7). This reaction is highly chemoselective and works with
better yields when Z is –CH2- and X is sulfur (i.e., a thioester function). The introduction of
the arylphosphine derivative in a carboxylic-containing surface can be conveniently carried
out by using an activated carboxylic surface and the diphenylphosphinomethanethiol, the
synthesis of which was recently reported by Raines and co-workers [10] for the
immobilization of small azide-containing synthetic polypeptides onto glass slides.
The azido function is not present in any naturally occurring protein. Tirrell, Bertozzi, and
co-workers, however, have reported a novel method for incorporation of azido groups into
recombinant proteins.[45, 46] Unnatural azido-containing aminoacids were incorporated in
recombinantly expressed proteins using engineered methionyl-tRNA synthetases in
combination with auxotroph E. coli strains. This approach, however, is not site specific for
proteins containing more than one methionine residue.
Figure 7. Chemoselective attachment of proteins to surfaces using a modified version of the Staudinger
reaction. (A) Proposed mechanism for the Staudinger reaction. (B) Site-specific immobilization of an azide-
containing protein onto a solid support using a traceless version of the Staudinger reaction [41-44].
An elegant way to overcome this limitation was recently developed by Waldmann and
co-workers [47] using in vitro EPL for the site-specific introduction of an azido group at the
C-terminus of a protein. This approach also allows the incorporation of other functional tags,
labels, or reporter groups. More recently, Raines and co-workers [48] have shown that the
thioester linkage between a target protein and intein can also be efficiently cleaved by
bifunctional hrydrazides bearing an azido group. This procedure appends an azido group to
the target protein in a single step without the need to use strong reducing conditions.

“CLICK” CHEMISTRY
The Cu(I)-catalized Huisgen 1,3-dipolar azide-alkyne cycloaddition, also known as
―click‖ chemistry,[49] has also been successfully used to immobilize azido- or alkyne-
containing proteins onto alkyne- or azido-coated surfaces, respectively.[50-52] (see Figure 8)
This cycloaddition reaction appears to be very forgiving and does not require any special
precautions. In the presence of Cu(I) as catalyst, the reaction proceeds to completion in 6 to
36 h at ambient temperature in aqueous buffers at pH 7-8. Under these conditions, the
cycloaddition is highly regioslective producing the corresponding 1,4-disubstituted tetrazole
as the only product (Figure 8A). It has been found that meanwhile a number of Cu(I) sources
can be used directly, and the catalyst is better prepared in situ by reduction of Cu(II) salts,
such as CuSO4•5H2O. Among the different reducing agents that can be used in this
cycloaddition, TCEP is one of the most competent reagents for the in situ reduction of Cu(II)
and has been shown to react only very slowly with aliphatic azides[53].
The site-specific incorporation of an alkyne group at the C-terminus of proteins has also
been accomplished using in vitro EPL [50] or nucleophilic cleavage of intein fusion proteins
with derivatized hydrazines [48]. More recently, Poulter and Distefano have independently
reported the use of protein farnesyltransferases (PFTase) for the selective alkylation of C-
terminal Cys residues of proteins with farnesyl analogues containing the alkyne function[51,
52]. PFTases catalize the alkylation of the thiol function in the Cys located in C-terminal
CaaX motifs, where X = Ala, Ser by farnesyl diphosphate. This reaction is general for any
soluble protein bearing a C-terminal CaaX motif and works very well for the
chemoenzymatic incorporation of alkyne and azido groups in the C-terminus of proteins.
Although this cycloaddition reaction can in principle be used for the chemoselective
immobilization of alkyne- or azide-modified proteins onto azide- or alkyne-coated surfaces,
respectively, Lin and co-workers[50] have found that alkyne-modified proteins react more
efficiently with azide-coated surfaces. This effect has been attributed to the fact that Cu(I)
coordinates with alkynes in solution more rapidly and with higher affinity than with the azide
group, thereby enhancing the rate of the cycloaddition reaction with the surface azido group.
Using these conditions, the minimal concentration of protein required for acceptable level of
immobilization was found to be in the low µM range.[50, 51]
Figure 8. Site-specific immobilization of proteins onto surfaces using the Cu(I) catalyzed Huisgen 1,3-dipolar
cycloaddition or ― click‖ chemistry. (A) Mechanism and regiospecificity of the Huisgen 1,3-dipolar
cycloaddition. In the absence of Cu(I), the reaction requires high temperatures and usually results in a
mixture of the 1,4- and 1,5-disubstituted tetrazoles. The addition of Cu(I) as catalyst produces only the 1,5-
regiosomer in very mild conditions.[49] (B) Immobilization of azide- and alkyne-containing proteins using
catalyzed Huisgen cycloaddition [50-52].
CHEMOENZYMATIC METHODS FOR

THE SITE-SPECIFIC IMMOBILIZATION OF PROTEINS
All the methods described so far rely on pure chemoselective reactions with little or no
activation at all. That means that the efficiency of these reactions depends on the
concentration of the reagents (i.e., on the concentration of the protein to be attached to the
corresponding surface) to bring both reactants close enough to allow them to react in an
efficient way.
One way to overcome this intrinsic barrier and make ligation reactions more efficient is
to introduce complementary moieties in the protein and the surface, which can form a stable
and specific intermolecular complex. Once formed, this complex brings both reactive groups
in close proximity, thus increasing the local effective concentration of both reactants (Figure
9).
Figure 9. Attachment of a protein to a surface by using a capture-ligand approach reaction [16].
PROTEIN IMMOBILIZATION USING ACTIVE

SITE-DIRECTED CAPTURE LIGANDS
The idea of using reactive ligands to capture proteins has been used by Meares and co-
workers [54] for creating antibodies with infinite affinity. In this interesting work, the authors
create an antibody against a metal-complex ligand, which contained a reactive electrophile
close to the binding site. When the antibody and the ligand are apart, their complementary
groups do not react, mainly due to the dilution effect. However, when the antibody
specifically binds the ligand, the effective concentration of their complementary groups is
greatly increased, leading to the irreversible formation of a covalent bond.
Figure 10. Site-specific immobilization of C-terminal mutant O6-alkylguanine-DNA alkyltransferase (AGT)-

fusion proteins [60]. In this enzymatic alkylation approach, the target protein is fused to an AGT of ≈ 200
residues. Selective immobilization can be accomplished by using O 6-benzylguanine-coated solid supports.
Mrksich and co-workers [55] have used this same principle for the selective attachment
of protein onto surfaces with total control over the orientation. They use the protein
calmodulin fused with the enzyme cutinase as a capture protein. Cutinase is a 22 kDa serine
esterase that is able to form a site-specific covalent adduct with chlorophosphonate ligands
[56]. The chlorophosphonate group mimics the tetrahedral transition state of an ester
hydrolysis. When it binds specifically to the active site of the enzyme, the hydroxyl group of
the catalytic serine residue reacts covalently with the chlorophosphonate to yield a stable
covalent adduct that is resistant to hydrolysis.
In this case, the authors use a gold surface to immobilize the cutinase inhibitor. The
attachment is extremely selective and can be carried with the whole crude E. coli periplasmic
lysate containing the cutinase fusion protein [55].This approach has also been used to prepare
antibody arrays on self-assembled monolayers presenting a phosphonate capture ligand [57].
Walsh and co-workers [13] have also recently reported a very elegant scheme for the
chemoenzymatic site-specific modification of proteins. In their approach, the target proteins
are expressed as fusions to a peptide-carrier protein (PCP) excised from a nonribosomal
peptide synthetase (NRPS). NRPS PCPs are relatively small (8-10 kDa), autonomously
folded, compact, and stable domains. These domains contain one specific Ser residue that can
be catalytically phosphorylated by the phosphopantetheinyl (Ppant) transferase SFP using
CoA (Coenzyme A) as a substrate. Using the Ppant transferase SFP from B. subtilis, the
authors were able to specifically label proteins with Ppant-biotin using biotin-CoA as
substrate. These biotin-labeled proteins were used to produce protein microarrays onto an
avidin-coated glass slide. Johnsson and co-workers [58] have used a similar approach
involving the transfer of phosphopantetheine derivatives to a peptide-acyl carrier protein
fused to the protein of interest. In a similar way, this approach could be used for the site-
specific immobilization of PCP-fusion proteins onto surfaces derivatized by CoA.
Johnsson and co-workers [59] have developed a novel approach for the site-specific
labeling of recombinant proteins using a mutant of human O6-alkylguanine-DNA
alkyltransferase (AGT). This modified enzyme can efficiently transfer a benzyl group to itself
when presented with O6-benzylguanine (BG) derivatives. The mutant enzyme is promiscuous
with respect to the substituents appended to the benzyl group, enabling a range of probes to
be used for site-specific labeling. The same group has recently reported the use of this active
site-directed capture approach for the selective immobilization of different AGT-fusion
proteins onto O6-benzylguanine-coated slides (Figure 10 [60] The fact that the AGTs from E.
coli and yeast do not react with BG derivatives allows direct immobilization without
purification of AGT-fusion proteins expressed in the above microorganisms. It should be
noted, however, that for AGT-fusion proteins expressed in mammalian cells, the use of
specific inhibitors is required to prevent the unwanted attachment of cognate AGTs. These
inhibitors have been designed to inhibit endogenous AGTs without affecting the engineered
AGT used for the site-specific attachment [61].
Los and Wood have also recently reported the use of an engineered haloalkane
dehalogenase, the HaloTag protein (HTP) for site-specific derivatization of proteins (Figure
11) [62, 63]. The native HTP is a monomeric protein (MW ≈ 33 KDa) that cleaves carbon
halogen bonds in aliphatic halogenated compounds] Upon nucleophilic attack by the
chloroalkane to Asp106 in the enzyme, an ester bond is formed between the HaloTag ligand
and the protein (Figure 11). HTP contains a critical mutation in the catalytic triad (His272 to
Phe) so that the ester bond formed between HTP and HaloTag ligand cannot be further
hydrolyzed. HaloTag ligands labeled with small organic dyes or immobilized onto surfaces
can be used for the site specific modification of proteins [64] or production of protein
microarrays [65], respectively.
H O
N O O O
H2N O O Cl HO Asp106 H2 N Asp106
O
O
HTL
HTP HTP-HTL
HTP-fusion protein
O
HO
O O
O
Cl O
Figure 11. Schematic of the specific immobilization of proteins to solid supports mediated by the HaloTag
protein (HTP) and its ligand (HTL=HaloTag ligand) [62, 65].
PROTEIN IMMOBILIZATION BY
PROTEIN TRANS-SPLICING
One of the main limitations of site-specific capture methods for site-specific
immobilization of proteins is that the linker between the protein of interest and the surface is
always another protein or protein domain. In some cases, the presence of such a large linker
could give rise to problems, especially in those applications where the immobilized proteins
will be involved in studying protein–protein interactions with complex protein mixtures [66,
67].
To address this problem, our group has developed a new traceless capture ligand
approach for the selective immobilization of proteins to surfaces based on the protein trans-
splicing process [26] (Figure 12). This process is similar to protein splicing[68, 69] with the
only difference being the intein self-processing domain is split in two fragments (called N-
intein and C-intein, respectively).[70, 71]
In our approach, the C-intein fragment is covalently immobilized onto a glass surface
through a PEGylated-peptide linker while the N-intein fragment is fused to the C-terminus of
the protein to be attached to the surface. When both intein fragments interact, they form an
active intein domain, which ligates the protein of interest to the surface at the same time the
split intein is spliced out into solution (see Figure 13).
Figure 12. Proposed mechanism for protein splicing in cis (left) and in trans (right).[38, 69, 89]
Figure 13. (A) Site-specific immobilization of proteins onto a solid support through split-intein mediated
protein trans-splicing [26]. (B) Structures of linkers used for the derivatization of a glass substrate.
Key to our approach is the use of the naturally split DnaE intein from Synechocystis sp.
PCC6803 [72]. The C- and N-intein fragments of the DnaE intein are able to self-assemble
spontaneously (Kd = 0.1-0.2 µM), not requiring any refolding step[26, 73]. The DnaE intein-
mediated trans-splicing reaction is also very efficient under physiological-like conditions ( 1/2
≈ 4 h and trans-splicing yields ranging from 85% to almost quantitative) [26].
Using this split intein, we have successfully immobilized Maltose Binding Protein (MBP)
and Enhanced Green Fluorescent Protein (EGFP) to chemically modified SiO2-based
substrates [26]. Both proteins were modified at the DNA level to append the DnaE N-intein
fragment (IN, residues 1-123 of the DnaE intein) at their C-termini. The two fusion proteins
(MBP-IN and EGFP-IN) were readily expressed in E. coli and purified by affinity
chromatography.
In order to enable the site-specific attachment of the IN fusion proteins onto a glass
support, an amine-functionalized glass slide was first treated with maleimidopropionic acid
N-hydroxysuccinimide ester (MPS) and then reacted with a mixture of PEGylated thiol
linkers 3 and 4, in a molar ratio of 3:97, respectively (Figure 13B). Linker 3 contains the
DnaE C-intein fragment (IC, residues 124-159 of the DnaE intein) followed by the
corresponding C-extein sequence CFNK. The Cys residue in the C-extein sequence was
protected with a S-tBu protecting group. This was required to ensure that the IC-containing
linker was selectively immobilized through its PEGylated terminal thiol onto the maleimido-
coated glass surface.
Linker 4 was used as diluent to control the density of reactive sites on the modified glass
surface. As shown in Figure 14A, different concentrations of pure EGFP-IN and MBP-IN
fusion proteins were spotted onto IC-coated glass slides. As a control, a solution of EGFP with
no IN fragment was spotted in the same slide (Figure 14A). In addition, a solution of MBP-IN
was spotted onto a glass slide derivatized with only the non-functional linker 4 (Figure 14B).
The trans-splicing reaction was incubated in a humidified chamber at 37°C for 16 h. Only
specific immobilization of the proteins containing the IN polypeptide to the IC-containing
glass surfaces was observed. No fluorescent signal was detected from the control protein
EGFP, lacking the IN polypeptide, after washing. In addition, the MBP immobilization was
minimal when the trans-splicing active MBP-IN fusion protein was spotted onto a control
glass slide coated with the non-functional linker 3 (Figure 14B). The trans-splicing mediated
attachment of EGFP and MBP in both cases was efficient in a range of concentrations. It is
interesting to note that the attached EGFP retained its characteristic green fluorescence,
indicating that its tertiary structure was unaffected by the attachment to the PEGylated glass
surface.
We have also explored the ability to selectively immobilize IN-containing fusion proteins
from complex mixtures through protein trans-splicing. We have shown that cellular fractions
of E. coli overexpressing MBP-IN can be selectively immobilized onto IC-coated glass slides
with minimal background, as described above (Figure 15A). The use of MBP-IN expressed
using an E. coli-based in vitro transcription/translation (IVT) systems also gave similar results
(Figure 15B).
Figure 14. Selective attachment of EGFP-IN (green) and MBP-IN (red) onto derivatized glass slides [26]. (A)
Epifluorescence image of an IC-coated glass slide after the spotting of different concentrations of EGFP-IN
and EGFP (top) and after buffer washes (bottom). (B) Different concentrations of MBP-IN were spotted onto
a PEG- (top) and IC-coated (bottom) glass slides. After washing, the immobilized MBP was detected by
immunofluorescence. Dot size is ≈ 100 µm.
Protein immobilization using protein trans-splicing is highly specific and efficient. It

allows the use of protein mixtures and eliminates the need for the purification and/or
reconcentration of the proteins prior to the immobilization step. The required minimum
protein concentration for efficient immobilization was estimated to be sub-micromolar [26].
More importantly, once the protein is immobilized to the surface, both intein fragments are
spliced out into solution, providing a completely traceless method of attachment. All these
features allow this methodology to be easily interfaced with cell-free protein expression
systems with rapid access to the high-throughput production of protein chips and other types
of biosensors.
Figure 15. Selective immobilization of MBP-IN from complex mixtures [26]. (A) Soluble cellular fraction of
E. coli cells overexpressing MBP-IN. (B) MBP-IN expressed in vitro using a cell-free system. Protein
concentrations in the cell lysate and IVT crude reaction were estimated by Western Blotting. In both cases,
MBP was detected by immunofluorescence after washing. The level of non-specific background can be seen
at the bottom of the slide when MBP with no DnaE N-intein was spotted.
CONCLUSION
We have reviewed some of the latest developments in site-specific immobilization of
active proteins onto solid supports. The ability to interface active biomolecules such as
proteins with solid supports is of great importance for the development of new technologies in
biotechnology and biophysics. For example, functional protein microarrays are starting to
become a key research tool in proteomics research.[9, 25, 26, 74-76] Like DNA microarrays,
protein microarrays allow for high-throughput analysis of thousand of proteins
simultaneously for rapid characterization of new protein–protein, protein–nucleic acid, and
protein–small molecule interactions as well as enzymatic activity and post-translational
modifications [75, 77, 78].
Other potential applications for site-specific immobilization of protein onto surfaces
involve the creation of optimized biosensors [79, 80]. An ordered protein film will have a
higher activity density that a random protein film, where a significant percentage of the
protein molecules may be bound to the surface in potentially inactive conformations. With
ordered proteins, biosensors can be miniaturized without losing sensitivity. Also, the
combination of recent advances in nano-lithography technique [81] combined with the ability
to bind proteins in an extremely ordered fashion allows for the creation of molecular
nanopatterns that can be used to better understand the processes involved in macromolecular
assembly [24, 82].
In summary, proteins represent an extremely fertile territory in micro- and particularly
nano-biotechnology because they possess a variety of properties ideal for engineering. They
have sophisticated architectures at nanoscale dimensions, complex chemistry composition,
and a wide variety of enzymatic activities. Proteins are capable of carrying out complex tasks
in living cells. The flagellar motors of bacteria, the linear motors of muscle and cytoskeleton,
voltage-gated ion channels, DNA replication complexes, or the photosynthetic reaction
centers are just a few examples. The development of new chemical/biochemical technologies
for the engineering of proteins with absolute control will shape the future of nanotechnology,
allowing us to harness the full power of proteins to create new components for materials and
devices.
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Lecture Material 21
PREPARATION AND APPLICATION OF BIO-INSPIRED

COLLOIDAL SYSTEMS
ABSTRACT
Nanostructured core-shell particles and their counterpart hollow capsules are two
classes of colloidal systems that can be designed to mimic biocolloids. A variety of
procedures have been employed for the manufacture of core-shell particles. One of the
simplest and most versatile approaches is the layer-by-layer self-assembly (LbL)
technique, which enables coating of various colloids with diverse composition and
controllable thickness. This chapter provides an overview of our recent work on the
preparation of bio-inspired colloidal systems via biofunctionalization of colloidal
particles with nano-structured shell composition via the LbL self-assembly technique.
The emphasis is on biocompatible colloidal systems that have applications in ultra-
sensitive immunoassay and biomolecule encapsulation with controlled release.
INTRODUCTION
Nanostructured colloidal systems (core-shell particles and their counterpart hollow
capsules) have attracted great interest due to their great potential for use in a wide variety of
applications. A number of procedures have been employed for the manufacture of core-shell
particles in the nanometre to micrometre size range [1-3]. Among them, learning from and
mimicking the supramolecular architecture of structures found in nature has provided a
promising way to design and fabricate complex colloidal systems with sophisticated
functions.
In the last decade, the LbL self-assembly method [4-6], which is based on the stepwise
electrostatic self-assembly of oppositely charged species, has emerged as a promising and
versatile approach to the fabrication of functional core-shell particles with well-defined shell
structures. The great advantages offered by the LbL technique are its ability to coat colloidal
particles with uniform layers of varying composition (e.g., synthetic and natural polymers,
biological macromolecules, dyes and nanoparticles) and the achievement of controllable
thickness at a molecular scale. These colloidal particles have sizes varying from nanometres
to many micrometres, and composition spanning a range from inorganic and polymer
particles to those composed of biomacromolecules or low molecular weight species. In
addition, the subsequent removal of the template core has led to a viable approach for
preparing hollow capsules with controlled composition and wall thickness [7-9]. The diameter
of the capsule is dependent on the size of the template and the shell thickness is controlled by
the number of deposited layers. The capsule surface can be further modified to alter the
functionality and/or improve the colloidal stability of the capsules, and various materials can
be sequestered into the capsule interior for drug delivery, sensing or catalysis applications.
In this chapter, we highlight our recent work on the preparation of biocompatible
colloidal systems via biofunctionalization of mesoporous silica particles with nano-structured
shell composition via the LbL self-assembly technique, and their applications in ultra-
sensitive immunoassay and biomolecule encapsulation. In the Section 3 we introduce a new
method to prepare a novel class of fluorescent biolabels by templating hollow periodic
mesoporous organosilica (H-PMO) particles. The biolabels are formed by loading dye
molecules into H-PMO particles, followed by biofunctionalization with polyelectrolyte and
antibody via LbL self-assembly. Their performance as fluorescent immunoassay biolabels is
also discussed. In Section 4 we discuss the preparation of biocompatible polyelectrolyte
capsules by combining colloidal templates and the LbL technique. We then introduce a facile
and universal strategy to encapsulate biomolecules with high loading and retention of
bioactivity by using mesoporous silica particles as both capsule and biomolecule loading
templates.
DYE-LOADED MESOPOROUS SILICA PARTICLES AS FLUORESCENT

BIOLABELS FOR IMMUNOASSAYS
Among the currently available analytical techniques, fluorescent immunoassay (FIA) is
one of the most promising strategies for achieving bioassays with high sensitivity and high
specificity, and it has been widely used in early disease diagnosis. As the sensitivity of FIA is
mainly determined by the number of light quanta emitted per analyte molecule, increasing the
fluorescent dye to biomolecule ratio (i.e., the F/P ratio) is crucial for achieving signal
amplification (and hence sensitivity). However, labeling antibodies with large numbers of
fluorophores usually leads to reduced specificity and binding affinity as well as a reduced
quantum yield due to dye self-quenching effects [10]. Thus the optimal dye/protein ratio is
normally kept below ~4 [11], which greatly limits the sensitivity of FIA.
Recently, we reported a novel approach for the preparation of a novel class of fluorescent
biolabels with high F/P ratio based on dye-loaded hollow periodic mesoporous organosilica
(H-PMO) particles [12]. As illustrated in Figure 1, the biolabels were prepared by a stepwise
process beginning with the loading of organic dye fluorescein diacetate (FDA) molecules
onto H-PMO spheres. Ultrathin polyelectrolyte multilayers were then deposited on the surface
of the particles using the LbL technique to provide a suitable ―i nterface‖ for further
attachment of antibodies. Compared with previous strategies, this method offers a widely-
applicable method for preparing label systems with high F/P ratio. In addition, it allows
control of the size and shape of the label systems by choosing suitable templates, and
provides a promising way to prepare multi-dye label systems for particular applications.
Figure 1. Schematic illustration of the preparation of the antibody coated H-PMO-FDA-PEM biolabels: (a)
FDA loading into H-PMO particles; (b) LbL assembly of oppositely charged polyelectrolytes onto H-PMO-
FDA particles; (c) Further coating of IgG resulting in the formation of H-PMO biolabels. (Adapted from [12]
with permission of the American Chemical Society.)
Loading Dye Molecules into Hollow Periodic Mesoporous Organosilica (H-

PMO) Particles
Periodic mesoporous organosilicas (PMOs), with organic functional groups

homogeneously distributed throughout the mesoporous silica walls, have attracted much
attention recently [13-15]. Compared with other mesoporous materials, these materials have
highly ordered mesoporous structures, as well as better hydrothermal and mechanical
stability. More interestingly, their surface composition and properties (e.g., charge, polarity
and hydrophobicity) can be tuned by changing the organic component, which makes them
very attractive in applications such as catalysis, sensors, drug delivery and separation.
As shown in Figure 2, the H-PMO particles used in this work have well-defined
mesostructures with pore size from 3.6 to 3.8 nm. The diameters of the particles are in the
range from 300 to 400 nm with an average diameter of 375 nm (Figure 2d) and a shell
thickness of ~50 nm. This unique structure provides the H-PMO particles with a high surface
area of 844 m2.g─1 and pore volume of 0.587 cm3.g─1, which makes them very attractive for
material loading purposes.
Figure 2. Low (a) and high (b, c) magnification TEM images of H-PMO particles. (d) Size distribution of as-
synthesized H-PMO particles. (Adapted from [12] with permission of the American Chemical Society.)
FDA loading was carried out by dispersing the dry H-PMO particles into the DMSO
solution of FDA under sonication. The driving forces for adsorbing FDA onto the particle
surface are mainly physical interactions. The FDA loading amount can be estimated by
thermo-gravimetric analysis (TGA) and was calculated to be ~504 mg FDA/g H-PMO. This
value is much higher than the loading amount of FDA on conventional mesoporous silica
material MCM-41 (~ 140 mg FDA/g MCM-41) [16]. The higher loading ability of H-PMO is
probably due to its hollow structure and the hydrophobic nature of the organosilica that is
favourable for the loading of organic dye molecules.
Biofunctionalization of Dye-loaded Particles via LbL Technique
After washing off the loosely adsorbed FDA, the FDA loaded H-PMO particles (H-PMO-
FDA) were coated with 2 bilayers of poly(diallyldimethylammonium chloride)
(PDDA)/poly(sodium 4-styrenesulfonate) (PSS) by the LbL technique (denoted as H-PMO-
FDA-PEM). Due to the loss of some FDA molecules during the coating process, the final
FDA loading amount was reduced to be ~443 mg FDA/g H-PMO. In addition, although the
thickness of the four layer polyelectrolyte coating is only about 3-4 nm [17], the
polyelectrolyte network can effectively prevent further desorption of FDA from the particle
and more importantly, the outmost PSS layer provides the FDA-loaded particle a suitable
surface for further antibody attachment (see below).
Goat anti-mouse IgG (Gt α-M IgG) was then immobilized onto (PDDA/PSS)2 multilayer
coated H-PMO-FDA particles by physical adsorption. The approximate IgG surface coverage
of 0.89, 2.3 and 5.1 mg.m─2 were obtained when the same mass of particles were incubated in
50, 100 and 200 μg.mL─1 IgG solutions, while the theoretically calculated surface coverage
values for a closely packed Gt α-M IgG monolayer are in the range of 2.0-5.5 mg.m─2,
depending on the orientation of the IgG molecules [18]. By using the FDA loading amount of
443 mg FDA/g H-PMO and average H-PMO particle size of 375 nm, the number of FDA
molecules per particle is calculated to be 9.1 X 106. From this value and the protein surface
coverage amounts, the F/P ratios can be calculated to be within a range of 1000~6000,
depending on the IgG concentration used for adsorption. These values are much higher than
the conventional covalently coupled fluorescent immunolabel, e.g., an IgG–FITC conjugate
(F/P ratio 4~8). As F/P ratio is the key parameter for improving the sensitivity of fluorescent
immunoassays, these novel biolabels have great potential to create highly amplified
bioassays.
Solid-phase Sandwich Fluorescence Immunoassays
The utility of these biolabel systems for immunoassays was examined by using a model
sandwich immunoassay for mouse IgG (M IgG) detection. Figure 3 illustrates the scheme for
performing the immunoassays. The capture antibody (Gt α-M IgG, Fc Specific) was pre-
adsorbed on the micro-titer plate. The analyte M IgG is added and immobilized by the capture
antibody. After exposing to the biolabels and carefully washing off the unadsorbed biolabels,
FDA in captured biolabels was released and converted into fluorescein molecules by adding
40% NaOH and sonicating for 1h. The addition of NaOH is to dissolve the silica particles and
free the loaded FDA molecules. It also acts as a hydrolysis catalyst and pH adjustor to turn
the diacetate form of fluorescein into its di-anionic form, which was fluorescent and soluble
in water and can be quantitatively measured [10]. The fluorescence intensity of the system
was then measured by a fluorescence reader.
The performance of three kinds of biolabels with different Gt α-M IgG surface coverage
was tested and the fluorescence intensity in response to different analyte concentrations is
shown in Figure 4. It was observed that for all three biolabels, the fluorescence intensity
increases with the analyte concentration and finally reaches a plateau. When compared with
FITC labeled commercial antibody system (F/P ratio 4.2), a 10 ~ 52-fold higher sensitivity is
achieved by using the particle biolabels, depending on the analyte concentration. In addition,
no increase of background signal was observed when using 500 ng.mL─1 Goat IgG as a
negative control, indicating that this method also has good selectivity.
Figure 3. Scheme of a solid sandwich immunoassay using H-PMO biolabels. (Adapted from [12] with
permission of the American Chemical Society.)
Figure 4. Sandwich fluorescence immunoassay of M IgG using three biolabels with different antibody
surfafce coverage: 2.3 (a), 5.1 (b) and 0.89 (c) mg.m─2. Error bars correspond to standard deviations of n = 6.
(Adapted from [12] with permission of the American Chemical Society.)
BIOCOMPATIBLE POLYELECTROLYTE CAPSULES FOR BIOMOLECULE

ENCAPSULATION
Preparation of Biocompatible Polyelectrolyte Capsules
LbL multilayer capsules are formed by the consecutive deposition of positively and
negatively charged polymers onto colloidal particles via electrostatic interaction, followed by
removal of the sacrificial colloidal template. The typical preparation process is illustrated in
Figure 5 [19]. The initial steps (a-d) involve stepwise film formation by repeated exposure of
the colloidal particles to polyelectrolytes of alternating charge. The excess polyelectrolyte is
removed by cycles of centrifugation and washing before the next layer is deposited. After the
desired number of polyelectrolyte layers is deposited, the coated particles are removed (e) to
leave hollow polyelectrolyte capsules (f). A comprehensive review on colloidal templates,
wall materials and the procedure used for preparing PE capsules was given by Peyratout and
Dähne in 2004 [9].
Recently, there has been increasing interest in using biocompatible polyelectrolytes,

including natural polymers and polypeptides (e.g.,) as building blocks to prepare multilayer
capsules with biocompatible or biofunctionalized surfaces aimed at biomedical applications.
For example, Berth et. al. reported the preparation of micrometer-sized hollow shells of
chitosan and chitosan sulfate by means of the LbL technique [20]. Sukhorukov‘s group
demonstrated the preparation of dextran sulfate/protamine capsules by sequential adsorption
of dextran sulfate (Mw 500 000) and protamine (Mw 5 000 with a high content of arginine)
on melamine formaldehyde (MF with diameter of 5.12 μm) particles followed by dissolving
MF core [21]. The same group also prepared sodium alginate (SA)/chitosan capsules by
templating biodegradable poly-DL-lactic acid (PLA) particles and studied the permeability of
the resultant capsules [22]. Another important biocompatible multilayer films are poly(L-
lysine)/poly(glutamic acid) (PLL/PGA) films, which have been widely studied on flat
substrates [23-26]. Yu and Caruso et. al. were the first to assemble PLL/PGA multilayer films
onto mesoporous silica (MS) particles to prepare enzyme encapsulated hollow capsules (see
following section) [27].
A convenient way to characterize the multilayer film build-up is to monitor the ζ-
potential changes of the particle after each deposition step. For example, at pH 5.5, PLL is
positively charged while PGA is negatively charged (PI~3.5). The original MS spheres have a
ζ-potential of ca. 40 mV. The ζ-potential varies after each PLL and PGA adsorption step,
becoming alternately positive (~+40 mV) or negative (~ 35 mV) respectively (Figure 6),
indicating that the multilayer surface is being charge-overcompensated in each adsorption
step, thereby facilitating adsorption of the next oppositely charged polypeptide to form the
multilayer film. According to ζ-potential measurements, PLL/PGA capsules could be formed
in both pH 7 and pH 5 buffer solutions, but the capsules prepared at pH 5 have a much thicker
shell compared with those prepared at pH 7. This is due to fact that PLL/PGA multilayer
films undergo an exponential mass growth with the number of deposited layers and this
nonlinear growth tendency becomes more distinct under low pH conditions.
Figure 5. Schematic illustration of the polyelectrolyte deposition process and of subsequent core
decomposition. (Adapted from [19] with permission of Wiley-VCH.)
60
40
potential, mV
20
-20
-40
0 1 2 3 4 5 6
Layer number
Figure 6. Variation in the ζ-potential for the deposition of PLL/PGA multilayer shells on MS spheres. The ζ-
potential measurements were performed in Milli-Q water at pH 5.8. Layer number 0 corresponds to the bare
MS spheres. Layer numbers 1, 3, 5 correspond to PLL deposition, and layer numbers 2, 4, 6 correspond to
PGA deposition.
Encapsulation of Biomacromolecules into Polyelectrolyte Capsules
Loading biomolecules into preformed LbL capsules. Arguably one of the most important
features of polyelectrolyte capsules is the fact that the properties and structure of the
multilayers of which they are formed are sensitive to conditions of the surrounding media
(e.g., pH, temperature, solvent mixture) and changes may result in altered permeability of the
capsules. For example, PAH/PSS hollow capsules are only permeable to water, small ions
and molecules with MW less than 7 000, but if the polyelectrolyte capsule shell is assembled
from materials that are responsive to salt (e.g., dextran sulfate/PAH) [28] or pH (e.g.,
PSS/PAH) [29], pores within the shell can be opened to allow larger molecules to diffuse into
the capsules.
This technique enables the loading of a range of materials into the preformed capsule
interior as long as the size/molecular weight of the diffusing molecule permits its permeation
through the capsule pores. The limitation of this technique is that the loading amount
achieved is typically low (particularly for large biomacromolecules), as the maximum
concentration of therapeutic inside the capsules is normally limited to the concentration in the
solution. Additionally, the loading reproducibility is often poor, and the conditions for
inducing permeability (i.e., extremes of salt or pH) may not be appropriate for many
biomolecules.
Direct coating enzyme crystals. In instances where the loaded materials form crystals,
encapsulation can be performed by direct assembly of polyelectrolyte multilayers onto the
crystals [30-33]. This method has been employed to encapsulate enzyme crystals of catalase
[30-32] and lysozyme [33]. Although amorphous particles composed of protein aggregates
such as lactate dehydrogenase [34] and chymotrypsin [35], can also be encapsulated within
polyelectrolyte multilayers, the formation of such particles relies on optimization of delicate
solution conditions and the results are often polydisperse and unstable. The advantage of this
technique is that a very high concentration of enzyme can be obtained in a capsule. However,
the obvious disadvantage is that this method is limited to encapsulation of compounds that
form crystals.
Using mesoporous silica (MS) particles as templates. To overcome these problems in the
LbL encapsulation techniques, we recently proposed a versatile process for encapsulating
biomacromolecules in a free state at high loading by using mesoporous silica particles as
sacrificial templates for both enzyme immobilization and PE multilayer capsule formation
[27].
As illustrated in Figure 7, the enzymes are pre-loaded into MS particles by exposing the
MS spheres to enzyme solution. Following several washing cycles to remove loosely
adsorbed enzyme, the enzyme-loaded MS particles are coated with alternating PE multilayer
film via LbL self-assembly. Subsequently the porous silica template cores are removed by
exposure to a hydrofluoric acid (HF)/ammonium fluoride (NH4F) buffer (pH 5), resulting in
enzyme encapsulated polyelectrolyte microcapsules. The encapsulated enzymes can be easily
released by altering the shell permeability.
Figure 7. Schematic representation of the procedure for encapsulating enzyme in polyelectrolyte

microcapsules using MS spheres as templates. (Adapted from [27] with permission of Wiley-VCH.)
The advantages of this approach over other LbL encapsulation strategies are obvious. The
salient feature of this method is that it is applicable to a wide range of materials for
encapsulation. It is not limited to species that undergo crystallization, or dependent upon
adjustments in electrostatic interactions within PE microcapsules to alter shell permeability
characteristics. Any materials that can be either physically adsorbed or chemically bonded to
the silica surface can be encapsulated. The mesoporous silica particles can also be surface-
modified through the well-established siliceous chemistry to suit the special requirements of
particular materials. Another important feature is the high degree of loading that can be
achieved using mesoporous materials as the loading matrix. MS materials are well-known for
their high specific surface areas (up to about 1500 m2.g-1) and high specific pore volumes (up
to about 2 cm3.g-1) which confer a high loading ability for drugs and biological molecules
[36-38].
The MS material used in this work are so-called bimodal MS (BMS) spheres, which have
a bimodal pore s
ranging from 2–4 µm, a surface area of 630 m2 g-1 and a pore volume of 1.72 cm3 g-1 (Figure
8) [39].
Protein adsorption onto MS surfaces is a complex process and is affected by various
factors including the nature of the MS spheres, nature of the enzyme (size, isoelectric point)
and the experimental conditions used for immobilization. Due to the large molecular size of
biomacromolecules, the pore size of the MS material has been found to be a key factor in
achieving high loading [40, 41].
Figure 8. TEM images of BMS spheres at different magnifications. (Adapted from [39] with permission of
Royal Society of Chemistry.)
Figure 9. CLSM images of (PLL/PGA)3 microcapsules loaded with FITC-labeled catalase. The scale bar in
the inset corresponds to 800 nm. (Reproduced from [27] with permission of Wiley-VCH.)
After enzyme loading, a multilayer shell is assembled on the MS sphere surface to

encapsulate the protein in the mesoporous particles via the LbL procedure. Shells with
various compositions and properties have been reported, including PAH/PSS [42],
poly(diallyldimethylammonium chloride) (PDDA)/PSS [39], PGA/PLL [27, 43],
poly(vinylpyrrolidone) (PVON)/poly(methacrylic acid) (PMA) [44], chitosan/dextran
sulphate [45], and PDDA/silica nanoparticles [39]. During the assembly of the multilayer
shells (especially the first polyelectrolyte layer), the loaded protein can be partially released
from the MS particle due to competitive adsorption between the polyelectrolyte and protein at
the silica walls. The amount of enzyme desorbed is dependent on the enzyme size as well as
the strength of the interaction between MS spheres, enzyme molecules and polyelectrolyte. If
the enzyme is small and the interaction between the enzyme and polyelectrolyte is stronger
than the interaction between the enzyme and the MS spheres, the enzyme molecules tend to
desorb from the MS spheres and be replaced by polyelectrolyte. However, the leakage of
protein significantly decreases during subsequent deposition steps, and is negligible after
three bilayers of PE coating [46]. This suggests that a PE multilayer coating of six layers is
sufficient to encapsulate the loaded protein in the MS particles.
A dilute HF solution or a buffer solution of 2M HF/8M NH4F (pH 5) has been widely
used to remove the MS template core through the formation of [SiF6]2-, which can easily
diffuse through PE multilayers [47]. The final enzyme amount encapsulated in the capsules
varies greatly for different enzymes and depends on the experimental conditions used for
protein adsorption, multilayer deposition and core dissolution. For example, the maximum
loading (expressed as a concentration) in (PLL/PGA)3 microcapsules for catalase adsorbed
from a 1 mg.mL-1 / pH 7.0 phosphate buffer for 72 h was 40 mg.mL-1 [27]. However, for the
smaller enzyme lysozyme (Mw 14,000, size 3.0 nm x 4.5 nm) adsorbed from 2 mg.mL-1 / pH
7.0 phosphate buffer for 3 h, the concentration within the microcapsules was 185 mg.mL-1
[27]. Compared with other LbL encapsulation techniques, the high enzyme loading is
noteworthy, particularly for a large enzyme such as catalase (~10 nm) which is difficult to
load into pre-formed hollow capsules.
Another important feature of this encapsulation approach is that the encapsulated

enzymes are in a free state. Figure 9 shows a confocal laser scanning microscopy (CLSM)
image of (PLL/PGA)3 microcapsules loaded with fluorescein isothiocyanate (FITC)-labeled
catalase (7 wt% loading). The CLSM image shows that the capsules are well separated in
solution. The homogeneous fluorescence from each capsule indicates that catalase molecules
were uniformly distributed throughout the capsule interior and suggests that most of the
catalase molecules are in a free state, i.e., not forming a complex with the PE shell.
The encapsulated enzymes were shown to maintain their bioactivity. Evidence for this
from our experiments is the fact that urease-loaded PLL/PGA capsules are capable of
catalyzing the hydrolysis of urea and have been shown to induce the exclusive formation of
CaCO3 particles inside the capsules [48]. Urease-containing capsules were injected into a
solution of 0.5 M urea and 1 M CaCl2, which had been degassed by nitrogen bubbling. In the
presence of urease, urea decomposes to form CO32- inside the capsules. The carbonate anions
then react with metal cations from the surrounding solution to precipitate calcium carbonate.
Figure 10 shows typical SEM images of urease-loaded (PLL/PGA)3 multilayer capsules
before (a) and after carbonate precipitation in 0.5 M urea and 1 M CaCl2 solution for 10 (b)
and 20 min (c). It can be seen that bulky, spherical particles with diameters ranging from 2~4
μm are formed, which is the same size as the PE capsule. The fast and uniform CaCO 3
precipitation process is mainly due to the high enzyme loading and notable enzymatic activity
within the PE capsules. It should also be noted that no CaCO3 forms outside the capsules
during the reaction indicating that the PE shell can effectively prevent the leakage of the
enzyme and restrict the CaCO3 precipitation to only occur within the PE capsules.
Figure 10. SEM images of urease-loaded (PLL/PGA)3 capsules before (a) and after 10 min (b) and 20 min (c)
carbonate precipitation in 0.5 M urea and 1 M CaCl2 solution. (Images A and C are reproduced from [43]
with permission of Royal Society of Chemistry.)
Release of Biomacromolecules from Polyelectrolyte Capsules
The encapsulated biomolecules can be released from the capsule interior by changing the
permeability of, or by rupturing the capsule shells through various methods [49]. Traditional
stimuli used to regulate intermolecular interactions in electrostatically assembled systems
include pH and ionic strength [27-29]. Over recent years, significant interest has also emerged
in other environmental triggers such as chemical stimuli [44], enzyme degradation [45] and
light [33, 50-51]. Additionally, recent studies on permeability change of capsules

incorporating temperature responsive polymers [52-53] and magnetic nanoparticles [54]
indicate that temperature and magnetic fields may provide promising new strategies for
stimuli resulting in the release of the encapsulated materials.
As weak polyelectrolytes (e.g., PLL and PGA) undergo conformational changes upon
changes in solution pH and salt concentration, encapsulated enzymes can be released from the
capsule interior by the same mechanism as that used for loading biomolecules into preformed
hollow capsules. For example, catalase in (PLL/PGA)3 microcapsules prepared in pH 5.5
buffer can be released under other pH conditions or by adding salt, as shown in the enzyme
release profiles in Figure 11 [27]. Capsule stability and enzyme retention were observed when
the buffer solution remained at a pH similar to that used for capsule preparation (pH 5.5).
However, by decreasing the pH to 2, almost 80% of the enzyme was released from the
capsule within 30 min. A similar result was observed at the other extreme of pH, a value of
11. In addition, added salt also enhances protein release, as salt induces conformational
changes in the three-dimensional structure of weak polyelectrolytes, increasing capsule
permeability. However, total enzyme release may be prevented due to the association of some
of the encapsulated catalase with the capsule walls.
Because different enzymes show optimum activity at different pH values, it is possible to
select certain release conditions for a given enzyme. For example, the released catalase
molecules keep activity when released at pH 7.0 and 5.5 while activity reduced to ~70% after
release at pH 11. Following release at pH 2, the enzyme showed negligible activity.
Figure 11. Influence of pH and salt on catalase release with time for catalase-loaded (PLL/PGA)3
microcapsules. The layers were deposited from pH 5.5, 0.05 M MES buffer. The lines drawn are to guide the
eye. (Reproduced from [27] with permission of Wiley-VCH.)
CONCLUSION
This chapter has highlighted some of our recent work in preparation of bioinspired
colloidal systems via the LbL self-assembly technique and their applications in biological
fields. We demonstrated a general and effective method to prepare a novel class of
biochemical assay labeling systems based on utilization of mesoporous organosilica particles
as templates. The biolabels could be constructed by first loading dye molecules followed by
biofunctionalizing polyelectrolyte multilayer and IgG molecules via LbL self-assembly
technique. A high F/P ratio (from ~1000 to 6000) of the resulting biolabels could be achieved
owing to the high surface area, hollow structure and the hydrophobic surface nature of H-
PMO. The immunoassay experiments indicated that the biolabels were immuno-active, and
generated an optimal signal that was ~50 times higher than the conventional dye-labelled IgG
system. In the second part of the chapter we introduced an effective and novel method, based
on mesoporous silica particle templates, for the preparation of high content biomolecule-
loaded biocompatible microcapsules that can be switched by pH or salt to alter their
permeability and release active biomolecules. This approach is significant as it is not limited
to materials that crystallize or by the shell characteristics of preformed hollow polyelectrolyte
capsules to effect encapsulation. Further, due to the immobilization capacity of mesoporous
silica for various materials and the versatility of the LbL method, this technique is likely to
find wide application in the encapsulation of other interesting materials, such as drugs,
fragrances and nanoparticles.
The prime advantage of the LbL technique is its versatility and modularity as regards
materials and properties. This has led to rapid development of the technology in widespread
research and application areas in the last 10 years. There is no doubt that LbL deposition has
become a powerful tool to prepare biocolloids with defined structure and functionality.
Currently, we are working on different ways to improve the sensitivity of our fluorescent
biolabels. For example, the synthesis of H-PMO particles with larger mesopore size and
vertical pore direction, should facilitate the infiltration of dye molecules into the hollow part
of the particle thus increasing the dye loading amount and therefore the F/P ratio. We are also
considering utilizing smaller size of mesoporous silica particles in order to reduce the steric
effect during immunoassays, and employing quantum dots instead of fluorescent dyes to
increase the fluorescence intensity and reduce the self-quenching effect. Regarding
polyelectrolyte capsules, efforts are directed towards synthesis of biocompatible polymers
with different functionality; understanding of the structure–property relationship and the
permeability response of the multilayer film upon external physical and chemical change; and
developing ‗smart‘ capsules with biocompatible nature and specific response characteristics.
These fundamental studies are also beneficial in developing new encapsulation and release
strategies for biomolecules, for example those that respond to multiple stimuli and deliver
multiple active molecules in a controlled way. In addition, further functionalization of the
capsule shell with superparamagnetic nanoparticles, integrin receptors, and antibodies to
enable the capsules target to specific areas is very attractive for controlled drug delivery and
release.
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Lecture Material 22
BACILLI, GREEN ALGAE, DIATOMS AND RED

BLOOD CELLS –
HOW NANOBIOTECHNOLOGICAL RESEARCH
INSPIRES ARCHITECTURE
ABSTRACT
Biological processes, structures, functions and materials provide powerful inspiration
for novel approaches in architecture. In this chapter, a variety of biological systems are
introduced: Bacillus subtilis, the green alga Euglena gracilis, diatoms and red blood cells.
Subsequently results of bionanotechnological research performed (by physicists) on these
systems are presented. In the next step, the systems and the results are discussed with an
architect, resulting in a multitude of ideas, possible approaches, experiments and projects.
Such interdisciplinary access corroborates the power of collaboration across established
fields in modern science and technology.
INTRODUCTION
Biological systems with functional units in the micro- and nanometer regime
continuously inspire novel micro- and nanotechnological applications [1]. Synergies between
biology and mechanical engineering are manifold [2] and were the motivation to investigate
synergies between bionanotechnology and architecture. Effective collaboration requires
interdisciplinarity. However, with the huge knowledge in different fields, nowadays it is
impossible for a single person to know and understand more than just a fraction.
Nevertheless, the awareness and understanding of different approaches and concepts is a
paramount prerequisite of interdisciplinary work.
A common language of bionanotechnologists and architects, in which descriptions at

different level of detail are more compatible, is attempted here – the chapter is intended to be
readable by both occupational groups.
General principles that can be applied by architects comprise integration instead of
additive construction, optimization of the whole instead of maximization of a single
component feature, multi-functionality instead of monofunctionality, and energy efficiency
and development via trial-and-error processes. Systematic technology transfer from
bionanotechnology to architecture thereby becomes generally accessible (Figure 1).
Terminology. As in many other fields, for example information technology, the

interpretation of the term „architecture―was extended from the initial building construction
related to the interpretation of a general system and construction principle, adaptable to any
technical discipline.
In this way „nanoarchitecture―can be interpreted as the design of new nanotechological
materials and surfaces, by the controlled assembly of nano-scale „building― bl ocks.
On the other hand nanotechnology increasingly finds its way into our everyday lives, so
into the classic architecture and building construction as well. In order to avoid confusion, we
will refrain from using the term nanoarchitecture for the application of nanotechnology in
architecture.
Aims. The encounter of four selected projects in nanotechnology with current
architectural developments will widen the interpretation and understanding of both disciplines
and identify future research fields.
Figure 1. Conceptual diagram of this bookchapter.
Current Application Fields of Nanotechnology in Architecture
Literature research was carried out for nanotechnological applications in architecture, to

give an overview of the present state of the art. As proposed by Gruber P. in ― Biomimetics in
Architecture‖ in 2008 [3] architecture is interpreted as multiscale discipline, from nanoscale
to regional and urban planning. In general nanotechnology delivers functional optimisation of
specific characteristics of building and construction materials. Comprehensive collections of
nanotechnological applications in architecture are given by S. Leydecker [4] and the
association of German engineers VDI [5]. Nanotechnology and a biomimetic approach have
already delivered new products for architecture and building industry.
By means of further development in chemistry, physics and material science innovation
in the field of nanotechnology in architecture are made possible. In the following, present
research aims are presented:
Production of new nano-objects and nano-systems, for example spheres, crystals,

plates, fibres, layers and branching structures.
Design, control and regulation of new or optimised material characteristics, for
example chemical reactivity, hardness, flow properties, colour and transparency,
protective function, electrical conductivity and magnetism.
Improvement of materials and production processes, also concerning environmental
and economic issues.
Solution of complex problems in an intelligent and economic way.
Improvement of interior climate, comfort of living and safety in buildings.
Improvement of durability of buildings, amongst others elements the facades,
windows, doors, roofs etc.
Reduction of energy consumption.
Improvement of energy efficiency and durability of cement-bound materials.
Improvement of durability of tarmac, a widely used pavement material.
Current application fields are not restricted to surfaces, but this field is vast in contrast to
the fewer developments on the side of actual materials. The difference between surface
structure and material characteristics will be the main criterion used to order the field. This
might be in opposition to materials research, but is a useful differentiation used in architecture
and building.
Surface Coatings
Surface coatings of construction materials and products are very common. They are used
in the inside and outside of buildings to change the characteristics of the elements surface in
providing a new functionality that cannot be achieved with the material as such. The
production of these coatings is usually by bulk, and the application techniques range from
conventional painting to spraying and other methods. The effectiveness of the coatings is due
to the designed surface geometry and catalytic processes.
Functions that can be achieved are described in the following.
Self cleaning and easy-to-clean function

Self cleaning of surfaces is based on the well known Lotus-effect [6], by the
creation of a super hydrophobic surface. The company STO is the market leader in
facade colour and plaster (http://www.sto.de). Similar principles are used for
protection from graffiti on wall painting.
The contrary effect, hydrophilic surfaces, is used for anti-fog function. They
create an invisible thin film of water instead of visible droplets.
Photocatalytic effects work in the combination of air humidity, UV radiation,
oxygen and the nanoscale catalyst TiO2. The surfaces are cleaned with water from
decomposed dirt particles.
In contrast to the Lotus-effect surfaces, easy-to-clean surfaces are smooth,
hydrophobic and oleophobic. Water rolls off due to the hydrophobic surface
characteristics.
UV protection
Inorganic UV protection is a novelty in building industry. The three chemical

compounds, TiO2, ZnO and CeO are used to absorb UV radiation and protect
material from ageing. Particle sizes below 15 nm ensure transparency. Many research
groups and companies are working in this field.
Switchable transparency and darkening
A new development in electrochromic glasses, whose transparency and
darkening is switched with electric voltage, allows the use of adaptive darkening
without continuous use of electrical energy. A memory effect in the functional nano
layer is responsible for the effect. Different grades of transmission are possible.
Photochromic glass changes colour according to environmental light conditions.
Large glass producers meanwhile offer electrochromic products for architectural
application under the title ―s mart glass‖ (for example Saint Gobain, Sage
Electrochromics) [7]. [The basic technology was developed by the Lawrence
Berkeley National Laboratory in the 1990ies [8].
Anti reflectivity
Nano-scale surface structures that are smaller than the wavelength of visible
light, for example SiO2 spheres, deliver efficient anti-reflective solutions. The use of
biomimetic structures is also possible: In 2006 the US company Reflexite developed
an antireflective knobbed surface that yields in the wavelength band between 400 and
700 nanometers a reflectivity of less than one percent [9]. The inspiration for this
product came from work on moth eyes from Vukusic and Sambles that was
published in Nature [10].
Anti fingerprint
Anti fingerprint coatings change the refraction of metal or glass surfaces in a
way that fingerprints are no longer visible, delivering a clean appearance.
Fire protection
SiO2 based gels are used in a millimetre thin layer as functional filling in glass,
and deliver excellent fire protection. Nanoparticles in coatings are applied to glass,
wood, metal, plastics or concrete. In case of fire they produce a ceramic protective
layer. Other nano-additives are used in organic fibre materials reducing
inflammability.
Anti bacterial effect
Silver nanoparticles deliver photocatalytic and anti bacterial effects, and are used
for interior materials, for example floors, textiles, sanitary surfaces etc. In this way
the use of disinfectants can significantly be reduced.
Scratch proof and abrasion proof
Self healing or glasslike scratch proof and abrasion proof surface coatings can be
used for different materials, for example wood, metal and ceramics.
Air cleaning
Catalytic processes are used for air cleaning. The degradation of organic
substances delivers uncritical components.
Microcapsules for fragrances
Microcapsules set fragrances free when they break during pressure or friction.
They are used in interior design, and are activated by the user.
Materials
Nanomaterials are either thin layers or composite materials, which are nano-structured.
They are used for:
Insulation: Vacuum panels and Aerogels

Vacuum panels deliver better thermal insulation values than conventional
materials. The panels consist of a metal or plastic foil, enclosing the evacuated layer
of foam, powder or fibrous material.
Aerogels, one of the few really nano-structured materials, is used in form of a
granulate material for insulation fillings in glass or polycarbonate panels. Aerogel is
an ultra light nanostructured foam with exceptional thermal and acoustic insulation
properties [11].
Temperature control: Phase change materials
Phase change materials deliver latent energy storage by melting and freezing.
Paraffin in nanoscale plastic spheres absorbs energy without temperature increase.
By absorbing peak temperatures, interior spaces stay cool for a longer period of time.
The process works the other way around as well.
High performance lightweight concrete
Nanoparticles improve the bond in the so-called ―N ano-concrete‖, thus
improving material properties as well.
Optimisation of characteristics of cement bound materials in general
Materials for energy harvesting
High temperature fuel cells for electricity and heat production with gas can be
more efficient with the use of nanostructured membranes and catalysts.
Solar cells
Nanotechnology is used to create dye-sensitized solar cells, a promising
development in photovoltaic applications.
LEDS
Light emitting diodes are increasingly used as energy efficient lighting
technology.
Atomic Force Microscopy and Spectroscopy
Bioimaging with the atomic force microscope (AFM) has become an ambitious field of
research in the last two decades. It has increasingly been used to image microbiological
samples at ultrahigh resolution. There are several advantages of using the AFM for biological
investigations. The AFM is not only an instrument for imaging sample surfaces, there is also
the possibility to measure various physical properties such as mechanical properties, surface
charges, molecular interactions, magnetic properties, friction forces and surface
hydrophobicity [12]. The AFM techniques can also be used to manipulate living samples and
surfaces. The main advantage of using the AFM for biological specimens is the ability to
analyze non-conducting surfaces without additional preparation such as metalizing with gold
which would have an influence on the biological properties of the samples. The second
advantage is the non-destructive method of imaging the biological samples by using the AFM
in the so-called dynamic mode.
Yet another AFM imaging mode is called phase imaging [13]. By mapping the phase of
the cantilever oscillation during the dynamic mode scan (the cantilever is one of the main
parts of the AFM with a sharp tip on its end used to scan the sample surface), phase imaging
goes beyond simple topographical mapping to detect variations in composition, adhesion,
friction, viscoelasticity, and possibly other properties. Applications include identification of
contaminants, mapping of different components in composite materials, and differentiating
regions of high and low surface adhesion or hardness.
One interesting topic for medicine, biology, industry and ecology is nanoscopic
investigation of cell surfaces. Cell walls have properties that can provide information on the
interaction of pathogens with tissues and the accumulation on implants [14] in medicine,
show advantages in biotechnology, such as cell immobilization in reactors and water safety
treatment, e.g. Bacillus subtilis for managing drinking water quality safety.
The reason why cell surfaces play such an important role is their ability to interact with
the environment and to protect the cytoplasm from outer dangers. Cell surfaces act as
molecular sieves and control interfacial interactions, such as cell adhesion and aggregation. In
order to understand these functions, the structural and physical properties of the cell surface
have to be investigated. There are several methods [15, 16] for investigating these properties:
X-ray photoelectron spectroscopy, infrared spectroscopy, electrophoretic mobility
measurements, electron microscopy, and many other chemical and technical methods, which
all have the disadvantage of destroying useful information during the preparation process,
because of the required extensive cell preparation procedures. By using the AFM it is possible
to investigate the cell surface properties under physiological conditions and at ultrahigh
resolution. One of the current investigation topics is the change of the cell surface structure
under native conditions and the visualisation of effects induced by external agents. AFM
investigations of the influence of chemicals, enzymes, solvents, ions and antibiotics may
reveal significant changes of the properties of microbiological cells that cannot be detected
with other methods [17]. This advantage provides that cell growth, budding processes (i.e.
forming a new organism by the protrusion of another organism) and the change in cell surface
morphology resulting from treatment with external agents can be investigated in situ.
Molecular interactions play a big role in understanding medical and biological processes and
are essential for the study of human health. By using an AFM equipped with a closed fluid
cell, the living conditions and nutrients can be changed during the measurement in order to
observe changes of cell structures and their morphology in different environments. The
sample fixation, using a fluid cell, is challenging, but in some cases, especially for vegetative
samples, it is indispensable to use a fluid cell.
Another useful AFM application is the cell-probe technique, where the cells are directly
attached to the AFM probe. The cell-probe method can be used for recording force vs.
distance curves. Another field of usage of the AFM is the nanomedical investigation. AFM
cantilever tips functionalized with biomolecules are used to investigate forces between
ligands and receptors. For obtaining representative results of the chemical properties and
intermolecular forces of a biological sample surface, it is important to know the chemical
characteristics of the cantilever. There have been many probes developed with a well defined
tip-chemistry. These probes are usually functionalized with self-assembled monolayers
(SAMs), which are very sensitive to chemicals and have a high spatial resolution [18]. For the
investigation of cells with the AFM it is necessary to work in parallel with an optical
microscope to control the cantilever tip approach to distinct cellular features [19]. The cell
behaviour influenced by growth, gene expression and cell cycle progression is closely related
to changes in their physical properties [20]. The measurement of the changes of elasticity will
provide a better understanding of these processes.
The elasticity of the cell can be measured by using indentation techniques [21]. By
pressing the cantilever with a pre-defined force into the sample surface, force vs. distance
curves are obtained to determine the compressibility of the cell wall. The AFM tip is
approached towards and retraced from the surface, while the force between the tip and the
surface of the sample is recorded. The force vs. distance curves are a function of the z-value
and the deflection signal of the cantilever [22]. The comparison of force vs. distance curves
can give information about elasticity differences in various samples.
Cross et al. report in 2007 in Nature Nanotechnology a change in stiffness in cancer cells
as compared to benign cells [23]: Samples of lung, breast and pancreas cancer were studied
with atomic force microscopy. It was found that the stiffness of metastatic cancer cells is
more than 70 % smaller with a standard deviation over five times narrower than the stiffness
of benign cells that line the body cavity.
Dulinska and co-workers investigated erythrocytes from patients with haemolytic
anemias and patients with anisocytosis and found statistically relevant differences in stiffness:
the Young's modulus of pathological erythrocytes was higher than in normal cells [24].
Observed differences indicate possible changes in the organization of cell cytoskeleton
associated with various diseases.
Atomic force microscopy and spectroscopy studies on two systems, Bacillus subtilis and
red blood cells, will be presented below. The results of these studies, and two more biological
systems of interest, Euglena gracilis and diatoms, are further on discussed with an architect.
Bacilli
Bacillus subtilis (lat. bacillum/bacillus, stick; subtilis, simple) is a rod-shaped, gram-

positive bacterium (gram staining is a method to differentiate bacterial species) with flagellae
providing the mobility and the ability to sporulate (Figure 2, sporulation is a form of
adaptation to starvation).
Like every bacterium of the species Bacillus, B. subtilis grows in aerobe conditions and
produces endospores as a result of sporulation. The endospores allow the organism to resist
extreme environmental conditions concerning e.g. pH, temperature and nutrient shortage [25].
B. subtilis is not harmful to human health and its robust spores may serve as safe model
organisms for pathogenic microorganisms in drinking water (e.g. in testing the efficiency of
water treatment methods).
Morphogenesis
Sporulation is a mechanism of bacteria to adapt to starvation. In contrast to most adaptive
responses in bacteria, sporulation takes many hours and includes major changes in cellular
morphology as well as in biochemistry and physiology. Morphogenesis relies on the
cooperation of two sister cells, which both are starting with the same genome. The first cell is
packaged into a tough resistant coat, while the other cell contributes most of its resources to
this process and then lyses (i.e. the cell dies after destruction of the cell membrane with the
aid of a protein called Lysine). Morphogenesis is also an example for the differentiation of
certain cells and for elaborated feedback mechanism between these two specific cells. The
genetic interactions can be explained by the action of transcription factors. Sporulation is
among the best understood of developmental systems and helps answering basic questions of
biology at the molecular level.
Figure 2. Stages of sporulation in Bacilli.
By using the AFM phase imaging mode several cell properties can be measured. As
mentioned above it is possible to investigate material properties such as adhesion and
elasticity by recording the phase shift as a function of the driving signal. Although this
imaging method is not very easy to perform on living cells, this imaging method is evolving
rapidly [26]. Forces such as lateral and shear forces are reduced by using the dynamic mode,
without damaging the cells. By using phase imaging the obtained image can be explained as a
map of viscoelastic variations on the sample surface, and provide insight to the cells. A small
positive phase shift indicates stiffer, a small negative phase shift softer regions, and might be
displayed by brighter and darker regions within the recorded image, depending on the contrast
settings of the AFM software [27]. One influence of the phase shift signal is the surface
stiffness; the second is the result of the viscoelasticity.
UV-sensitive and UV-resistant Spores

The resistance to UVC and UVB radiation depends on the method inducing the
sporulation process and therefore which type of spore is formed. Bacillus subtilis spores have
been successfully established for the validation of water treatment processes like filtration
techniques and disinfection processes most importantly for the biodosimetry of UV

disinfection systems [28].
Green Algae
The algal flagellate Euglena (Figure 3) has for long been an outstanding subject of study,
its species is one of the most completely studied [29]. These small yet complex unicellular
organisms dispose over a plethora of bionanotechnological machinery in order to live, survive
and procreate. Their long feature list includes amongst others a proteic crystal that acts as
highly efficient light sensor, a flexible outer shell (the pellicle) that can actively change shape,
a mobile thread (the emergent flagellum) enabling them to move with high speed through the
liquid, and chloroplasts converting sunlight photons into storable energy.
The ability to ―f eed‖ mainly on sunlight (although it must take up the vitamin B-12,
which it cannot synthesize itself, from the surrounding) and to survive even with no light
present at all by forcing itself to heterotrophic metabolism has raised some discussion
whether this organism is more ‘plant-like‘ or ‘animal-like‘. Photosynthesis (and its
prerequisites, the chloroplast plastids within the cell) is a strong argument to classify it
amongst plants, yet an Euglena cell has no cellulose cell wall (its protective hull, the pellicle,
is inside the plasmalemma) and is a very active swimmer using its whip-like flagellum, an
ability rather associated with animals. This lets guess the unusual cross-functionality
regarding metabolism, organelles and lifecycle exhibited by Euglena.
Interesting subsystems of Euglena gracilis can be found on every scale [30], from its
entire body pellicle down to the light capturing and transdu ction mechanism where even
single atoms play a major role. Many aspects are still actively researched such as the
mechanics of proteinaceous strips composing the pellicle, the biochemistry of rebuilding lost
chloroplasts, the electrochemistry of aforementioned signal transduction from the stimulated
light receptor, the biophysics of concerted microtubuli contraction and detraction for
euglenoid movement (a second mode of movement described below) and many other
processes sometimes also to be found in other living systems.
Figure 3. Euglena gracilis cells under the optical microscope. The length of one cell is about 50 m. The
rightmost cell can be seen in the process of cell division. This takes about 2-4 hours after which two equal
daughter cells can begin their new lifecycles. In that sense an Euglena gracilis cell never dies, but lives on in
its offspring [31].
Photoreceptor
The word "Euglena" is formed from the two Greek words "eu" and "glene" which mean
"good" and "eyeball" respectively, because of the clearly visible (with optical microscopes)
stigma (Figure 4), also called the eyespot [32]. Originally it was thought that this "eyespot"
was light-sensitive and used by the alga to direct itself towards the light. But we know today
that its role is only to intermittently shade the photoreceptor (attached to a flagellum inside
the reservoir of the cell) as the cell revolves around its long axis. In this sense the eyespot is
only a sub-part of the algal optical system.
The ability to perceive light and adapt to changing light conditions is crucial to
photosyntactic organisms, therefore detecting low light intensities becomes an adaptive
advantage. A photosynthetic organism in dim light can obtain more metabolic energy if it is
able to discriminate and move toward better illuminated areas.
The photoreceptor is connected to the flagellar rod and protected by a surrounding
membrane. It is a highly efficient light detector and is shielded on one side by the stigma (the
eyespot). The stigma consists of tiny carotenoid granules that absorb light in the sensitivity
range of the photoreceptor. This simple but complete visual system allows Euglena to
orientate itself towards a light source [33]. The light-orientated movement of the cell, called
phototaxis, is caused by the teamwork of the stigma and the photoreceptor. During its
movement, the cell permanently rotates and the stigma comes between the light source and
the photoreceptor. Euglena experiences a periodical dip in light intensity and changes its
direction of movement until the detected light is no longer modulated by the stigma. Then the
cell is moving towards the light source.
Figure 4. Apical part of Euglena viridis. Image adapted from [34].
Figure 5. Three images of the apical part of the same cell. The leftmost image shows the cell as seen in phase
contrast, with no UV-light illumination. The image in the middle shows the cell during illumination with
365 nm light - upon absorption of photons of this wavelength the rhodopsin-like proteins present in the
Euglena photoreceptor change into the excited state. The green emission of the photoreceptor is only faint. In
the rightmost image, the cell is irradiated with photons of 436 nm wavelength. At this wavelength the light
sensitive proteins fall back into the ground state, emitting bright green light and completing the photocycle.
Arrowheads indicate the photoreceptor crystal in the images, the arrow indicates the stigma responsible for
intermittently shading the alga when rotating during swimming. The scale bar in each of the images is 20 m.
The photoreceptor is the exceptional light-sensing unit of the alga. It is a small proteic
crystal and enables the alga to detect even very low light intensities (i.e. single photons).
Although the photoreceptor is only 1 m in diameter, it reaches an absorption rate close to
100% of the incident light within its absorption band spectrum. The sensitivity of its
photoreceptor is so high because it is made of a stack of many pigment containing membranes
(around 100 layers) [35].
Embedded within the layered structure of the photoreceptors is its main ingredient, a
rhodopsin-like protein. Rhodopsins are special proteins for intercepting light, universally used
from archebacteria to humans, consisting of a proteic part, the opsin, organized in seven
transmembrane helices, and a light-absorbing group, the retinal (i.e. the chromophore). The
retinal is located inside a pocket of the opsin, approximately in its center. Several properties
make the retinal-opsin complex an excellent light detection unit. It has an intense absorption
band whose maximum can be shifted into the visible region of the spectrum, over the entire
range from 380 nm to 640 nm. Second, light isomerizes the retinal inside the protein very
efficiently and rapidly. The isomerization, i.e. the event initiating the vision reaction cascade,
can be triggered almost exclusively by light. In the dark it occurs only about once in a
thousand years!
The isomerization and possible conformational changes of the protein follow a
photocycle and are therefore repeatable (Figure 5). The photocycle leads through a series of
conformational changes from the initial state to an excited state and back again. Usually also
a number of intermediates can be identified. The photocycle of the chromophore in the
photoreceptor of Euglena shows such a cycle including a ground and an excited state. As
different conformational states possess different fluorescence characteristics, a photoreceptor
whose chromophore proteins are mostly excited differs in emission characteristics from a
photoreceptor containing chromophore protein mostly in the ground state.
The time it takes for the whole photocycle to complete is on the order of microseconds or
less. The isomerization alone is one of the fastest processes ocurring in nature, completing in
about 200 femtoseconds (i.e. 200*10-12 s) [36].
Pellicle
If the cell cannot use its flagellum for locomotion it can move by the so-called ―eugl enoid
movement‖ (similar to that of a worm contracting the back and extending the front part) -
sliding along solid material and being lubricated by its own muciferous lubricant excreted
from pellicle pores. This kind of movement can be induced when only little space is given to
the alga to move, e.g.between two flat glass slides. The pellicle of Euglena has evolved into a
very elastic and refined structure that supports such a viscous change in shape during
euglenoid movement.
The boundaries of the Euglena body are formed by the outer tripartite (three-layered)
plasmalemma membrane surrounding the pellicle composed of interlocking and articulating
flexible strips (Figs. 6 and 7). These ribbon-like strips are arranged in a left-handed spiral and
interwoven with microtubuli.
Figure 6. Twice the same Euglena cell as seen with an optical microscope, using UV-light illumination. The
focal plane in the left image is adjusted such that the part of the pellicle closer to the observer is rendered.
The image one on the right shows the far side of the pellicle. The light-dark pattern follows the alignment of
pellicle strips fusing partly at top and bottom.
The pellicle defines the basic shape of the cell. Its role is vital to the organism as it must
function as protection from the environment, yet cannot be fully impermeable as it must
permit e.g. exchange of information or matter with the exterior as in sensory pathways or
uptake of the vitamin B12. Additionally, euglenoid movement requires the strips of the
pellicle to be highly flexible and articulate against each other. The cells also show excellent
pressure resistance up to 100 bar and beyond.
There is strong evidence that the microtubuli within the strips (aligned in parallel)
together with motor proteins are responsible for the sliding of the strips against each other,
meaning the pellicle changes its shape actively at the command of the cell! The fact that this
protective shielding is self-assembling through means of specific binding sites inside the
plasmalemma membrane and binding proteins adds to this exceptional part of Euglena [38].
Figure 7. On top of a swimming Euglena: The schematic drawing depicts a transverse section of its cell
surface. Details of the articulating S-shaped strips of the membrane skeleton and the infrastructure associated
with strip overlap. The position of the skeleton and the bridges are well suited to mediate the sliding of
adjacent strips occurring during shape changes. The portion of the plasma membrane not subtended by the
cytoskeleton may provide the fluid region, which accommodates sliding as well as a region for the insertion
of new strips during surface replication. The traversing fiber is positioned to maintain the S-shaped
configuration and it may contribute an elastic component to the sliding skeleton. MAB1 and MAB2,
microtubule associated bridges; MIB-A and MIB-B, microtubule independent bridges; PM, plasma
membrane; T, traversing fiber [37].
If the cell is disrupted the pellicle can be seen dissociated along the striations into flat
strips of material which have a thickened edge and a thinner flange. Electron microscopy
sections clearly show how these strips interlock and how they pass helically along the cell.
These strips are intracellular structures lying immediately beneath the plasmalemma, a
continuous tripartite membrane about 0.8 – 1 m thick. The pellicle is thus not equivalent to a
cell wall, since the latter is always laid down outside the plasmalemma (like a cellulose wall
of plant cells). The throughs between adjacent strips start as a whorl at the posterior end of the
cell, bifurcate a few times before passing helically along the length of the cells and then meet
again as they reach the canal opening. The strips of the pellicle curve over and continue into
the canal, where they also fuse. Although variation occurs concerning thickness and shape the
form of construction is the same in all euglenoids. The cross section of a pellicle surface can
be seen in a schematic drawing in Figure 7.
Diatoms
Diatoms are unicellular microalgae with a cell wall consisting of a siliceous skeleton
enveloped by a thin organic case. The cell walls of each diatom form a pillbox-like shell
consisting of two parts that fit within each other. These microorganisms vary greatly in shape,
ranging from box-shaped to cylindrical; they can be symmetrical as well as asymmetrical and
exhibit an amazing diversity of nanostructured frameworks [39, 40] (Figure 8).
Diatoms are found in both freshwater and marine environments, as well as in damp soils
and on moist surfaces. They are either free floating (planktonic forms) or attached to a
substrate (benthic forms) via biogenic adhesives, and some species may form chains of cells
of varying lengths. Individual diatoms range in size from 2 μm up to several millimeters,
although only a few species are larger than 200 μm. Diatoms as a group are very diverse, with
12 000 to 60 000 species reported 41, 42].
Diatoms can serve as model organisms for micro- and nanotribological investigations
[43-45] and as templates for novel three-dimensional microelectromechanical systems
(MEMS) [46, 47]. In ambient conditions, these organisms produce nanostructured amorphous
silica surfaces. Some diatom species have rigid parts that in relative motion act like rubber
bands when elongated [48] and subsequently released, whereas other diatom species have
evolved strong, self-healing underwater adhesives [49]. Diatoms are small, mostly easy to
cultivate, highly reproductive, and, since many of them are transparent, are accessible using
optical microscopy methods.
Already in 1999, Parkinson and Gordon [50] pointed out the potential role of diatoms in
nanotechnology via designing and producing specific morphologies. In the same year, at the
15th North American Diatom Symposium, Gebeshuber and coauthors [51] introduced atomic
force microscopy and spectroscopy to the diatom community as new techniques for in vivo
investigations of diatoms. These scanning probe techniques not only allow for the imaging of
diatom topology, but also for the determination of physical properties like stiffness and
adhesion [52-57]. A representative example of the fruitful exchanges in the area of diatom
nanotechnology can be found elsewhere [58].
Figure 8. a) b) c) SEM images of Solium exsculptum, an Eocene fossil (45 million years old) from a deposit at
Mors, Denmark. b) and c) show the linking structures in more detail. Scale bars: 20 μm, 5 μm, and 5 μm,
respectively. The sample is from the Hustedt Collection in Bremerhaven, Germany, # E1761. (Reproduced
with permission. © F. Hinz and R.M. Crawford.)
Hinges and interlocking devices in diatoms are very stable and can still be seen in fossil
deposits millions of years old (Figure 8) [59]. In 2006, Gebeshuber and Crawford [60]
presented scanning electron microscopy (SEM) images of extinct and recent diatom species
with linking structures with the aim of correlating structure and function. Figure 8 shows four
connections of two Solium exsculptum sibling cells that lived 45 million years ago and are
still in good condition.
Perhaps we might even soon be able to evolve the kind of nanostructures we want and
replicate them in large numbers via the way diatoms naturally replicate – cell division: a
compustat [61, 62] could monitor diatom properties and selectively destroy cells that do not
evolve in the desired direction. In this way, directed evolution would take place. This
conveyor belt-type production could yield nanostructures for use in technological
applications. The amorphous silicate material of the diatom cell wall is not very interesting
for technological applications. However, recently Sandhage and co-workers introduced a
method to replace the diatom silica with materials of technological relevance (such as silicon)
while preserving the shape [63-66]. In this way, tailored diatom nanostructures are even more
usable for devices.
Red Blood Cells
Blood consists of blood plasma and different types of cells, the red blood cells
(erythrocytes), the white blood cells (leukocytes) and the platelets (thrombocytes). The
erythrocytes give the blood its red colour and are responsible for the oxygen transportation
within the human body.
Red Blood Cells

Red blood cells carry oxygen to the periphery of the body and CO2 from there back to the
lung, exchange the gases and start the circle again. This is possible with the help of
haemoglobin. There are about 4.5 - 5 million red blood cells per microliter blood.
Erythrocytes are about 7.5 µm in diameter and 2 µm in thickness. The cells are shaped
biconcavely have a surface area of about 135 µm² and a volume of about 90 fl.
The erythrocytes shape indicates the hydration status of the body. Therefore certain
diseases can be diagnosed by the investigation of the shape of the red blood cells and their
surface properties. For example, hypertone dehydration leads to shrunken and wrinkled
erythrocytes whereas hypotone dehydratation provokes balloon shaped red blood cells.
Erythropoietin
Erythropoietin (EPO) is a hormone that is produced by the kidney. It stimulates the bone
marrow to produce red blood cells. Synthetic EPO is clinically used for treating renally
caused anaemia and kidney diseases. In serious sports recombinant erythropoietin is used for
doping. Recombinant erythropoietin is a glycoprotein that is generally expressed in Chinese
hamster ovary cells that were transfected with DNA encoding for human erythropoietin.
Applying recombinant EPO enhances the number of erythrocytes by about 5-20 %. This leads
to increased oxygen transfer and better performance of the doped athletes. Synthetic EPO is
difficult to detect, because of its natural occurrence and because it is metabolised within 6-
12 hours.
A medical device based on a method similar to AFM stiffness evaluation that is fast and
reliably detects doping with EPO directly on site would find wide applications in serious
sports.
MATERIALS AND METHODS

Bacilli
By inducing adverse environmental conditions to living B. subtilis cells the sporulation

procedure is successfully initiated. Two methods of spores‘ production resulting in different
types of spores are included in the investigation. UV-sensitive and UV-resistant spores are
prepared by standard methods and kindly supplied by the Institute for Hygiene of the Vienna
Medical University [67].
Preparation of spores in aqueous solution. The B. subtilis spore solution is centrifuged in
order to remove unintentional artefacts. About 1 ml of this solution is dropped on poly-L-
lysine coated and uncoated glass slides.
After fifteen minutes drying in air, some of the samples are washed with PBS (phosphate
buffered solution) in order to reduce the spore density on the substrate. Depending on the
inclined position of the slides during the washing and the amount of PBS used more or less B.
subtilis cells remain attached to the slide. Then the sample slides are dried in air for about
forty minutes before the atomic force microscope measurements are started.
Preparation of vegetative B. subtilis cells. Due to the limited lifetime of the vegetative B.
subtilis cells and their ability to move by using their flagella the immobilization is an
important aspect. The use of adhesively coated substrates turned out to be sufficient for
mechanical fixing for AFM imaging.
The preparation is the same as for the B. subtilis spores in aqueous solution described
above, but in this case only substrates coated with adhesive poly-L-lysine for proper
immobilization and PBS buffer solution for diluting instead of distilled water are used. The
distilled water would initiate the bacteria membrane to burst because of the osmotic pressure.
The vegetative B. subtilis are usually dissolved in a nutrient solution. Nutrients can cause
artifacts on the substrate surface during imaging.
To prevent artifacts on the glass-slide, a proper method is to smear off the excess solution
with another slide. Due to the absence of water after drying the sample one has to consider
that the bacteria will possibly start the morphogenesis, which can be intended in some cases
e.g. for real-time imaging of changes in the bacterial membrane. To initiate the
morphogenesis, the drying must occur not too fast because a little amount of water is required
for the conversion from the living bacteria to the spore. Otherwise the majority of the bacteria
would die before any morphological changes can happen.
Atomic force microscopy imaging. Dynamic mode AFM was used to investigate the B.
subtilis samples. In dynamic mode there is only intermediate contact between the tip of the
cantilever and the sample surface because the cantilever is oscillating at or close to its
resonance frequency. This technique is used to avoid damaging the sample by scratching over
it. Amplitude, phase and frequency information is obtained by tip-sample interaction forces.
These modifications provide information about the sample‘s characteristics. In contrast to
contact mode, in dynamic mode the cantilever oscillation amplitude is kept constant.
Therefore the amplitude is permanently measured and a feedback loop adjusts the value of the
cantilever z-value due to the distance between the tip and the sample surface which is defined
by the set-point amplitude. Through this process the topography of the sample surface is
obtained.
Different tip driving frequencies were used for imaging the spores. The best images were
obtained using cantilevers with a resonance frequency of 70 kHz and a spring constant of
1.8 nN/nm (Olympus OMCL-AC240TS). The prepared samples were analyzed at different
positions with an investigation area of 20x20 µm². The parts of interest were then magnified.
Measurement of indentation depth. Force vs. distance curves are used to determine the
indentation depth of the cantilever tip into a sample due to a preset force trigger point. The
penetration depth is a parameter for the stiffness of the cells. It is calculated by the position of
the cantilever when the predefined trigger point is reached, minus the position of the
cantilever when it is in first contact with the sample surface. On each spore a path is created
that consists of 15 predefined points. On these locations force vs. distance curves are
recorded. The preset trigger forces are 3 nN and 12 nN, respectively, and the curves are
acquired with 1 Hz, 2 Hz and 4 Hz (Hz refers to the number of force vs. distance curves
recorded per second, indicating different pulling speeds). Altogether, 2760 indentation data
points on UV-sensitive and UV-resistant spores are obtained.
Red Blood Cells
There are several reasons for using the atomic force microscope in studying blood cells;
the most important is that the AFM is a general purpose instrument for analyzing surfaces at
ultrahigh resolution, in ambient, fluid or vacuum conditions. Compared to other analytic
instruments the AFM and especially the ambient AFM has a variety of advantages. The main
advantage is the ability to analyze non-conducting samples without additional preparation
such as metalizing with gold or similar techniques.
For the experiments the stiffness of blood samples of renal insufficient patients (i.e.
patients with kidney problems), who are medicated with synthetic EPO and blood samples of
a control group (healthy individuals) are compared. The maximum age of donors was 50 a.
After the standard procedure of preparation of the blood samples [68-71], the cells are
imaged with the AFM using dynamic mode. Subsequently AFM force vs. distance curves are
recorded and evaluated for differences in penetration depth.
The images are recorded in dynamic mode to prevent damaging the sample by scratching
over it, with cantilevers with 70 kHz resonance frequency in air and a spring constant of
1.8 nN/nm (Olympus OMCL-AC240TS). The scan frequency is 0.43 Hz (i.e. a little bit less
than two lines per second), the image size is originally 20x20 µm2, regions of interest are
subsequently scanned with smaller scan size.
For the question if there is a difference in the stiffness and plastic deformability of the
control group and EPO medicated patients, the most interesting parameter is the penetration
depth. The penetration depth is the parameter for the deformability of the cells. It is evaluated
by the position of the maximal movement of the piezo sensor in z-direction, which is the
coordinate when the sensor reaches the trigger point of 3 µN, minus the position of the sensor
when the tip is in first contact of the surface of the red blood cell.
Bio-Inspired Nanomaterials and Nanotechnology in Architecture
For the development of further concepts, a mutual biomimetic approach is used. Usually
– as described methodologically - biomimetic translation occurs from either end of the
process: the natural phenomenon or the technical problem. The connotations ―bot tom-up‖ and
―top-down‖ biomimetics (as introduced by Speck and co-workers [72]) are not used since
they might imply hierarchical relation between biology and technology. According to
Gebeshuber and Drack (2008) the methods of biomimetics by analogy and biomimetics by
induction are used simultaneously, in imposing current issues and discussions in a specific
field (architecture) onto a set of bionanotechnological investigations (bacilli, green algae,
diatoms and red blood cells, Figure 1).
RESULTS AND DISCUSSION

Bacilli
This section presents the first high resolution AFM results on various stages of the
sporulation process in B. subtilis. The different stages of the sporulation process are clearly
discernible (Figure 9). Starting from the vegetative B. subtilis (Figure 9a) the bacillus enters
stage I (asymmetric cell division, Figure 9b) and the developing endospore is appearing (stage
II, Figure 9c). The development of the prespore (Figure 9d, the cell is in stage III, IV or V)
leads towards the release of the mature spore by lysis of the mother cell (stage VI, Figure 9e).
Stages III, IV and V are not discriminable with surface methods such as AFM. At the final
stage of the sporulation process, the spores appear (stage VII, Figure 9f).
Furthermore, phase images of UV-sensitive and UV-resistant spores reveal distinct
differences in the structure compound which refers to different elasticity of the spore coats
(Figure 10). The darker regions (i.e. smaller phase signal) in Figure 10 indicate softer spore
surface areas and the brighter regions indicate stiffer spore surface areas. The structure of the
UV-sensitive spore is divided into small softer regions surrounded by a stiffer grid (Figure
10). The UV-resistant spore (Figure 10, right trace) has large regions that are stiffer than the
rest of the spore. These differences in stiffness are representative for UV-resistant and UV-
sensitive spores, and are found in all recorded samples.
2760 indentation data points on UV-sensitive and UV-resistant spores are recorded. The
UV-resistant spores are overall stiffer than the UV-sensitive spores, which correlates with the
obtained phase images. It is found that the indentation depth in UV-sensitive and UV-resistant
spores shows distinct differences when the indentation force exceeds a few nanonewtons:
Considering the mean indentation depth of a preset force of 3 nN there are no significant
differences in the penetration depth and therefore no difference in the stiffness of the spore.
Using a preset force of 12 nN shows distinct differences in the indention depth of the UV-
sensitive and UV-resistant spores (Figure 11).
Figure 9. Stages of the sporulation of B. subtilis imaged with atomic force microscopy. Dynamic mode,
imaging parameter amplitude. The different stages are clearly visible (see text).
Figure 10. Left: Typical phase trace of a UV-sensitive endospore: there are only few stiff regions separating
small softer areas. Scan size 667x667 nm2. Right: Typical phase trace of a UV-resistant endospore: large stiff
areas with small soft regions in between. Scan size 892x892 nm2.
Biomimetic Inspiration by Bacilli for Architecture – Results of the Discussion
Transformation of Principles
The most interesting characteristics of Bacillus subtilis are its sporulation ability, the
adaptive shape and spore UV sensitivity in combination with the skin structure of the spores.
Spores
The spores of the bacilli must sense the environment and check if the environmental
conditions have improved. How this is achieved is not known yet.
Figure 11. Indentation depth in UV-sensitive and UV-resistant B. subtilis spores. Differences are
very distinct for an indentation force of 12 nN (right trace) as compared to an indentation force of
3 nN (left trace). UV-sensitive spores have smaller stiffness.
Sporulation as Escape
The bacillus stops reproducing if there is a shortage of nutrients and retracts into spore
state. If viewed as a retraction phase, the spore phase is very interesting for architecture: if the
outside conditions are adverse, retraction provides protection for the inside. In terms of
architecture, the analogy is shelter, providing protection from attack or other hostile
influence. Extreme environments like polar stations or outer space require permanent
extraordinary protection measures. Temporary measures of retraction, for example the retreat
of inhabitants to warmer core zones of the house in winter, are used as strategies for energy
efficiency in traditional architectures. The zoning of space according to temperature is again
state of the art in sustainable design, but the architecture itself cannot yet be retracted.
Currently rooms that are not needed are only abandoned, but remain existent. A more
dynamic approach to architectural material could provide spore-like states as living
environment as well and might prove to be more energy efficient. Adaptive shapes that react
to daily or seasonal changes would be the solution.
On another scale, migration according to climate already exists in many also modern
human cultures. The so-called snowbirds, retired people in the US who stay during the
summer in the north of the US and move to warmer states in the winter, and the fast
increasing tourism industry are recent examples. Both phenomena use plenty of resources and
energy.
Principle of Mother and Daughter

The development of the one inside the other is also interesting for propagation in adverse
environments. The interior environment is separated from the outside, controlled and provides
the milieu that is needed for replication. Possible scenarios are toxic ambient air, too little
oxygen, dangerous radiation etc. For space applications, an analogue environmental problem
exists, with the hostile space environment. For space missions, the transport weight and
volume are crucial parameters. Everything has to be sent in a space-saving way, so often
folded structures are used. But on the other hand for space habitats sealing is always
challenging, e.g. if a cap has to be added to a foldable structure. The development of the one
inside the other would mean to develop a larger space habitat inside the milieu of the already
sealed space of the transport capsule.
UV Sensitivity
The membranes of B. subtilis UV-sensitive and UV-resistant spores differ structurally
concerning the dispersal and pattern of hard and soft areas. The differentiation may have a
functional relation with UV sensitivity, but it is still unknown. Apart from showing increased
stiffness, the pattern of the UV resistant spore membranes seems to be a highly ordered
system. The change in stiffness areas of the cell membrane in a pattern hints to a more
dynamic kind of adaptability as well – maybe shape change is possible here as well. More
investigation on the structure/function relation would clarify the case.
UV-sensitivity of B. subtilits is exploited as a biosensor. Sense, if and how the
environment changes and react to it. UV absorbtion is important in building industry. The
development of better and cheaper ways of blocking UV radiation would be economically
very interesting.
Red Blood Cells
After imaging the erythrocytes, force spectroscopy with trigger forces of three
micronewtons is performed on each single cell along preset paths in ambient air. About 200
force vs. distance curves are recorded per sample, 26 samples are investigated.
The penetration depth does not reveal statistically relevant differences in healthy and
EPO blood samples. However, the penetration depth of samples 10 and 11 is four times
higher than the penetration depth of the other samples (see Figure 12).
There are also abnormalities of the surface of the blood cells of the donor of samples 10
and 11. The erythrocytes do not have the typical donut shaped form. These blood cells are
oblate and very flat. These two blood donations are from the same donor. The measurements
are repeated in order to ensure that no destruction of the sample during transportation or
preparation took place. The previous result is confirmed, the cells are oblate and flat and the
penetration depth of the cantilever is four times higher than in the other samples, indicating
softer cells. After more detailed medical investigation a rare case of diabetes is diagnosed in
the donor of samples 10 and 11.
Penetration depth
4,50E-07
4,00E-07
Penetration depth [m]
3,50E-07
3,00E-07
2,50E-07 healthy
2,00E-07 EPO
1,50E-07
1,00E-07
5,00E-08
0,00E+00
21 26 25 23 22 24 9 19 20 10 11 2 1 16 15 13 17 4 5 6 8 7 18 14 3
Samplenumber
Figure 12. Penetration depth of all 25 blood samples samples.
The AFM successfully proves as a nanodiagnostic tool. Minamitani and co-workers

measured the deformability and viscoeleasticity of erythrocytes of patients with Diabetes
mellitus, the common type of diabetes, by microchannel flow systems and atomic force
microscopy. The blood cells of the patients with diabetes mellitus are harder than the control
group [73, 74]. The reason for the softening of erythrocytes in the rare case of diabetes
presented here and the hardening of erythrocytes in diabetes mellitus is yet to be determined.
Erythrocytes of people with diabetes also have a shorter live span compared to erythrocytes of
a healthy control group. Currently, diabetes is diagnosed solely via chemical methods.
Biomimetic Inspiration by Red Blood Cells for Architecture – Results of the Discussion
The most interesting characteristics of red blood cells are their shape change possibilities
and the change of properties revealed in the indentation experiments presented above.
Shape Change due to Environmental Influence

Sodium is the most important ion in the extracellular space. In hypertone dehydration one
looses more water than sodium, and the concentration of ions in the extracellular space is
larger than inside the erytrocytes. Water leaves the red blood cells, and the erytrocytes shrink
(Figure 13d). If the sodium concentration in the extracellular space is smaller than inside the
erytrocytes, water moves into the erytrocytes and they bulge. This can happen e.g. if one
drinks too much distilled water – in this case the erytrocytes can even burst.
The most interesting question; is the shape change of the red blood cells also functional?
Does the change of form help to cope with potentially dangerous situations of the organism?
As far as we know, there is no functional reason discovered yet.
Figure 13. a) Regular erythrocytes, b) Stack of erythrocytes, c) bulged erythrocytes d) thorn apple shape.
Image source: Gray's anatomy of the human body, 20th U.S. edition, 1918. Public domain material.
Shape change due to environmental influence is the abstract phenomenon that is

interesting for technical application. In the case of red blood cells the concentration of a
specific substance triggers a radical change of form (Figure 13). In architecture, changes of
environmental conditions, for example temperature difference between inside and outside, or
the presence of toxic substances in the air outside, could be used to initiate a functional form
change – for example by reducing the exposed surface area. More interesting is adaptability
on a materials scale. Thermal insulation is a large and expensive issue in building
construction. Currently we tend to increase thermal insulation to an extent that prevents the
use of the free energy from outside. Adaptability would be a solution for the daily and
seasonal changes of requirements. A shape changing layer in a composite wall could react to
humidity of temperature difference by increasing the layer thickness, thus increasing the air
layer and insulation properties. This layer does not have to be continuous. Single points of
shape changing material would be sufficient. Passive mechanisms would avoid the use of
additional energy for the adaptation control - energy efficiency by passive adaptation to
environmental conditions.
Another materials application is the use of the shape change principle for a sensor. The
microscale reaction to environmental change could bring a macroscopic reaction with it that
could be sensed by humans, for example colour change.
The Change of Properties

The change of properties of the red blood cells is not easy to handle, as the phenomenon
is quite new. The indentation method does not deliver structural information. If we knew
more about the structure we could suggest a way to change surface characteristics of
materials. The adaptation of material properties, in this case hardness and softness, would be
a wonderful thing to have in built environment.
Change of blood pressure is the macroscopic effect of the shape change of the red blood
cells. Change of flow properties by shape changing particles – could find technical
application. If we had particles that could change shape on command, we could use them as
flow control mechanism. A passive version could be: if the flow of a fluid full of particles
becomes too fast and turbulent, the particles lose shape and break the flow, if the flow is slow
enough again, the particles change back into initial shape.
Diatoms and Euglena Gracilis
Biomimetic Inspiration by Diatoms for Architecture – Results of the Discussion

The most interesting characteristics of diatoms are the fact that they biomineralize
amorphous silica, the connections they establish between sibling cells and their resting stages
and resting spores.
Biomineralization
Biomineralization is the process by which living organisms produce minerals, often to
harden or stiffen existing tissues. Examples include silicates in algae, carbonates in diatoms
and invertebrates, and calcium phosphates and carbonates in vertebrates. These minerals often
form structural features such as sea shells and the bone in mammals and birds. Organisms
have been producing mineralized skeletons for the past 550 million years. Other examples
include copper, iron and gold deposits involving bacteria. Organisms biomineralize more than
50 different minerals such as SiO2 and CaCO3 [75]. Linking Structures – Connections. Some
diatoms grow in colonies, the single cells are connected to each other (Figure 14). Why are
there four posts in Figure 8 and not three? If I had two connections I could still bend the
structure. Three would be enough for the reduction of the degrees of freedom. The
connections shown in Figs. 8, 15 and 16 provide a certain distance, yet keep the cells
together. The connections may function as dampers.
Table 1 Overview with organism examples and phenomena interesting for transfer.
Several criteria and aspects important for the specific example are described, and
estimated values for transfer issues like scaling and grade of abstraction are provided
Figure 14. Many diatoms grow in chains. The single cells are connected via junctions. The junctions can be
mucilage pads, fused extensions of the cell walls, interlocking spines of varying complexity and size fitting to
each other like a key to its lock or ball-and-socket arrangements [76] © 2006, Sibirian branch of the Russian
Academy of Sciences.
As can be seen below and in Figs. 15 and 16, different amounts degrees of freedom are
obviously possible for different species of algae. In spite of that, they may fulfill the same
function – generate space between the cells, but hold the cells together, and allow some
movement between the cells.
Single linkage structure connecting siblings – single extension intertwined with
neighboring one, the distance can change. Something that appears to have escaped notice
until recently [77] was that although considerable movement along the connecting axis is
possible, such an arrangement cannot be rotated. This is most easily appreciated by clasping
one´s forearm with both hands and attempting to rotate one arm. Examples for having a single
linking structure comprise species of the genera Rutilaria (Figure 15) and Syndetocystis.
Two linking structures connecting siblings – Maluina in Figure 16 has two extensions
meet in a connection that is very similar to Solium exsculptum. The distance is fixed, only
very little movement is possible, perhaps bending in cross section is still possible. The linking
structures outside look as if they are a system that reacts to compression – like a bumper. The
inside looks as if they react to tension, with interdigitating ―f ingers‖, altogether hints to a
semi-rigid connection, exposed to pressure and tension. On the outside of the connection area,
the shell is perforated in a regular way. The function of these openings is unclear – as the
material itself is not flexible, the perforated area can not deliver damping characteristics, but
is still less strong than the connecting bumpers and fingers. The openness hints to exchange
of something, maybe soft cell material.
Species of the genera Maluina, Hemiaulus, Climacodium, Keratophora and Briggera are
examples for having two linking structures.
Three linking structures connecting siblings. Trinacria regina is an example for having
three linking structures.
Figure 15. Single linkage structure in Rutilaria. Scale bar 50 m. © R.M. Crawford and P.A. Simsi [[1].
Four linking structures connecting sibling. Solium exsculptum in Figure 8 has four
connections at a fixed distance and position between neighboring cells. Only straight large
scale colonies seem possible. Solium exsculptum (Figure 8) and the Trinacria species
presented in Crawford and Sims, 2008 [78] (this species has tapering linking structures at
each of the four elevations that may pull apart without damage) are examples.
Five linking structures connecting sibling. Solium pentagona.
Many linking structures. Most of the genera showing linking structures between sibling
cells have many linking structures, attaching the cells firmly or not so firmly, holding them at
a distance, some are easily separable, some not, here a lot of analysis has to be performed.
However, in most of the cases the spines allow no movement of the two sibling valves vis à
vis one another. The genera Skeletonema, Stephanopyxis, Lamyloseira, Aulacoseira,
Cymatoseira, Strangulonema, Plagiogrammopsis, Fragilarimforma, Fragilaria,
Staurosirella, Staurosira, Preudostaurosira and Punetastriata exhibit such multiple linking
structures.
Many linking structures, ball-and-socket like. Kisseleviella.
Q: Interesting would be: is there evolutionary sequence of this functional solution?
A: In general, the connections become more complicated the more recent in the fossil
record one looks, but that there are examples of ―pr imitive‖ linking features in species that
appeared later and even among living diatoms [78].
Figure 16. Two linkage structures in Maluina. Scale bar 20 m. © R.M. Crawford and P.A. Sims [78].
Structure of Solium Exsculptum

The most obvious characteristic about the structure of Solium exsculptum, its rectangular
shape, can not be explained yet.
Between the main ribs the silica structure is extremely thin and interspersed with pores.
In these fragile plate-like structures, secondary stiffening by small, undirected ribs can be
observed. These reinforcement structures prevent the buckling that flat shells elements are
prone to. There are no reinforcement ribs on the top and bottom part of S. exsculptum - the
cytoplasm might provide enough mechanical support. The flange structure around the rim,
which obviously belongs to the primary structure, is interesting – perhaps it helps in the
attachment of the valves or serves as an attachment structure for the cell membrane (Figure
8). Altogether the structural differentiation is very material efficient.
The primary structure of S. exsculptum consists of the dark areas that can be
distinguished in Figure 8 (for sketch see Figure 17). These are reinforcement ribs, delivering
the main force transmission areas. In the primary structures there are no holes in the
amorphous silica (Figure 17). But there are reinforcement ribs. The whole structure is made
from thin material. Plates or shells without ribs would be prone to buckling. The silica
structure might be so thin since many pores are needed; the mechanical stiffness comes from
the reinforcements ribs. Less material is needed if thin material and reinforcements ribs are
used. There are no reinforcement ribs on the top and bottom part of the Solium image – only
the holes. Inside this top and bottom parts resides the cytoplasm, it might provide enough
mechanical support to prevent mechanical damage.
Figure 17. Left: Primary structure of Solium exsculptum, as far as discernable. Dark regions interpreted as
reinforced main structural elements. Dashed lines indicate secondary ribs, reinforcing the thin plates. Right:
Scheme of approximate extension of thin plates, enabling exchange by interspersed holes.
Spore Formation
Some diatoms, especially those whose natural habitats are soil and rock, can survive
desiccation for decades. Dormant vegetative cells, resting stages and resting spores ensure
survival [79]. Diatom resting spores (hypnospores) are heavily silicified stages in the life
cycles of marine centric diatoms and a few freshwater and pennate diatoms [80-83]. In some
genera, resting spores superficially resemble the parent vegetative cell, whereas in others,
spores and vegetative cells are morphologically quite different.
Biomimetic Inspiration by Euglena Gracilis for Architecture – Results of the

Discussion
The most interesting characteristics of the Euglena are its sensing and orientation system,
together with its movement and skin structure.
Integrated Orientation and Locomotion System

Locomotion and sensing are combined functions within one complex ―or gan‖. The
rotational movement initiated by the flagellum is used for periodic shielding of the sensing
device in order to deliver a signal difference that is interpreted directly and defines the overall
direction of movement.
The same principle works with other than light signals.
Orientation in space is essential for all living organisms. Human orientation depends on
signal difference between pairs of sensing organs, and locomotion is as important for
orientation. In contrast to common knowledge which accredits orientation to sight, sound is
very important for spatial orientation.
Technical devices for orientation in space already exist, for example the GPS system. It
depends on signal contact to a range of satellites. The easy availability of those systems for
private use has already influenced our perception of built environment and our behaviour.
Spatial orientation is not sufficient for human survival in complex environment.
Orientation in architecture and in built environment in general, is not always easy. Large
buildings, for example hospitals, traffic infrastructure, etc. need signage to transfer
information to the users, which has to be applied or inserted into the built environment.
Another system, providing selective information when needed would be helpful. The
visibility of pathways taken by other people, following the principle of chemical tracing of
ants, could be helpful for finding one‘s way. I could also use state transitions to activate
something, e.g. a path to an exhibition gets bright if many people walk on it. This would yield
reactive illumination in buildings, on floors, on streets and new concepts for illuminating
streets, parks, halls, etc.
Storage Medium
The crystals inside the sensing organ of Euglena provide a switch triggered by light. This
change in state of rhodopsin could be exploited, or transferred to another nanoscale system,
which delivers a similar effect.
This could be implemented into surface materials to detect the presence of people,
initiated for example by touch or acoustics. Another example would be reactive shirts for
soccer players that indicate fouls by being pulled by a change of colour (the more pull the
more intense the color).
The principle could as well be transferred to a larger scale system, monitoring something
happening in a ―sensing material‖ that also delivers information to the environment.
Skin Structure of Euglena
Skin Structure Built from Inside

A protein layer builds up the skin of Euglena from the inside. This method of growth is
interesting as it could be transferred to a light and soft system forming space in synthesizing
material and structure from inside out. The delivery of material works easily in liquid
solution, but would have to be solved in a different way in air. The interior system would
have to be stable enough to maintain itself and integrate a transport system for the material.
Inside out technologies could be interesting for rapid prototyping or production technologies.
Skin Structured in Stripes

The outer skin of Euglena is structured in stripes. One of the biggest challenges that are
currently explored in architecture is the construction of complex shapes with (mostly) two-
dimensional flat material. The conservative building industry is still working based on two
dimensions, but new production technologies allow three-dimensional material processing.
With two-dimensional stripes of material one can generate open geometries (cylinders),
but for having more complex, three-dimensional curved shapes the stripes would need to
deform, change width and taper off to zero – all this to achieve a static form. Change of cross-
sectional geometry of the stripes themselves would be another way of solving this.
Flexibility of the material itself enables a specific amount of change already, but other
mechanisms exist in Euglena that allowing not only for adaptability but also active
locomotion.
Flexibility of Connection
The connection of the skin-stripes is very interesting from a functional/geometric point of
view. A model would help to understand the functionality of and the options provided by the
connection. It is perhaps useful for the development of a new connection system of cladding
material, or even useful as a model for the connections of monocoque elements. [1] The first
association of a biologists to transfer the principle to a technical membrane must fail - „soft―
membranes are pre-stressed to maintain stability under different loading conditions. A rigid
frame would have to be used where a soft membrane could be attached to.
Movement Mechanism
The stripes of the shell move along each other. There are decentralised „motors―along
the connection. This could be translated into a decentralised actuation system, allowing an
architectural skin to adapt according to specific stimuli. An integrated actuation system like
this would be very useful for adaptable folded structures. Integrated actuation would allow a
system to deploy without centralised control.
Movement and Complex Geometry

Movement AND complex geometry seem currently impossible to achieve in an
architectural solution.
The analysis of the overall system of Euglena could provide a new approach to solutions
for adaptability in building skin. Useful information to have would be the relation of the scale
of deformation of the skin stripes to the movement along the stripes. An analysis of overall
geometric changes and a model would be helpful to fully understand the process.
CONCLUSION
An overview with the organism examples and phenomena interesting for transfer from
bionanotechnology to architecture is given in Table 1. Several criteria and aspects important
for the specific example are described, and estimated values for transfer issues like scaling
and grade of abstraction are provided.
The novel approach attempted in this book chapter turned out to be fruitful for both sides.
The bionanotechnologists and the architects got input concerning the applicability of reserach
results and future research directions. This approach should be intensified and extended to
other fields. The newly erected Center of Excellence TU BIONIK at the Vienna University of
Technology (comprising 30 researchers from eight faculties) and other similar centers such as
the BIONIS network in the UK or the BIOKON network in Germany provide the experts and
the interdisciplinarity that is needed.
Current challenges in architecture that shall be met with the help of nanotechnology are:
Functional integration to improve durability and provide new functions for building
elements, for example multifunctional facades, providing optimum light conditions
and protection against corrosion, fungi and vandalism.
Improvement of customer convenience, changes in social and urban structures will
have to be dealt with, for example concerning safety issues.
The need for material and energy efficiency, together with increasing environmental
consciousness, will be important triggers for change.
Increasing activation of building elements requires integration of new functions like
sensing and actuation, for example according to environmental conditions.
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Lecture Material 23
BIOPOLYELECTROLYTE MULTILAYER
MICROSHELLS: ASSEMBLY,
PROPERTY AND APPLICATION
ABSTRACT
The layer-by-layer (LbL) self-assembly of polyelectrolyte multilayer microshells
with tailored wall thickness on a nanometer-scale range, ordered wall composition, as
well as controlled size and permeability provides new opportunities for the technological
application in areas such as synthesis of nanomaterials, sensing, catalysis, separation,
drug delivery, and bioactive species immobilization. Biopolyelectrolyte-based
microshells have gained particular attention due to their nontoxicity to cells/organisms
and environment-benign nature. This chapter highlights a recently developed
biopolyelectrolyte microshells by the use of LbL self-assembly technique. We describe
methods and materials used to fabricate the biopolyelectrolyte microshells, and key
properties of these microshells including their stability and permeability responded to
external environments. Entrapment of various species including nanoparticles, dyes and
other organic materials in the shells is also detailed. Additionally, a major part of the
chapter is also devoted to the development of multilayered microshells, where two
aspects of potential applications in drug loading/release and pollutant remediation using
the biopolyelectrolyte shells as microcarriers/microreactors are presented.
INTRODUCTION
Materials with well-defined structures in the (sub-)micrometer regions have attracted
increasing interest in recent years. Micelles, vesicles and viruses used by Nature in biological
systems, as spatially confined microcontainers, have widely been studied for the application
in areas such as biological chemistry, synthesis and catalysis [1-3]. Nevertheless, a nano-
engineered microshell (microcapsule) can not be achieved using these systems because of the
difficulties associated with the precise controllability of their size and permeability as well as
their limited stability. Therefore a great effort has been devoted to prepare size- and shape-
persistent hollow microshells with control at the nanometer level.
Polyelectrolyte microshells were introduced in 1998 by Caruso and Möhwald et al. [4].
They are hollow, micro-sized polyelectrolyte spheres fabricated by electrostatic layer-by-
layer (LbL) self-assembling of oppositely charged polyelectrolytes on the surface of a
sacrificial template core. This preparation technique is very versatile and by varying the
colloidal core or the wall components, the physicochemical properties of the shells can be
customized. Such polyelectrolyte microshells have been proven to present the tailored wall
thickness on a nanometer-scale range, ordered wall composition, as well as controlled size,

shape and permeability, and thus display a collection of great potential applications for
synthesis of nanomaterials, heterogeneous catalysis, spatially confined precipitation,
crystallization, and enzyme immobilization [5-9].
Up to now a variety of charged polymers has been successfully assembled into thin films
by the LbL technique. The most conventional polyelectrolytes for fabricating multilayered
microshells are commercially available ones such as poly(ethyleneimine) (PEI),
poly(allylamine) (PAH), poly(diallydimethylammonium chloride) (PDADMAC),
poly(styrenesulfonate) (PSS), poly(vinylsulfate) (PVS), and poly(acrylic acid) (PAA). Many
applications, for example biomedicine and bioremediation, however, require the
biodegradable and biocompatible microshells. Such particles, in practice, are of particular
interest and can avoid the toxicity to cells/organisms and the secondary contamination to
ambient ecosystem.
In this chapter we will focus on biopolyelectrolyte-based hollow microshells that allow
the manipulation of surface charge, shape and size, permeability, and mechanical stability.
After a short introduction of the LbL assembly technique, we will describe methods and
materials used to fabricate several kinds of biopolyelectrolyte microshells, and discuss their
key properties and the loading techniques of various species in the microshells as well. A
major part of this chapter will also be devoted to the updated development of the
biopolyelectrolyte microshells in practical application.
CONSTRUCTION AND PROPERTIES OF BIOPOLYELECTROLYTE

MICROSHELLS
The LbL Assembly Technique
The so-called LbL assembly technique, introduced at the beginning of the nineties by
Gero Decher, was originally based on the alternate adsorption of charged cationic and anionic
polyelectrolytes on a charged planar substrate [10]. The driving force at each step for
multilayer buildup is the electrostatic attraction. One starts with surface charge of substrate.
Polyelectrolyte molecules with opposite charge are easily adsorbed onto the surface of the
substrate. Not all of the ionic groups of the adsorbed polyelectrolyte are consumed by the
electrostatic interactions with the substrate. In this case, the charge overcompensation occurs,
and this leads to a reversal of the surface charge, promoting the adsorption of a next,
oppositely charged polyelectrolyte. Along this route, ordered polyelectrolyte multilayers are
formed by sequential deposition procedure (Figure 1). This is a rather elegant LbL template
approach attributed to several particular virtues: i) the LbL technique is capable of creating
highly tailored polymer thin films with a large variety of functional groups embedded in the
film structure; ii) no complicated instruments are needed, thus it is a simple method for
fabricating multilayers; and iii) the LbL deposition is independent of the size or shape of the
substrate, thus it can be extended not only to original planar substrate, but also to other
substrates with different shapes. In this chapter, emphasis is placed on tri-dimensional
polyelectrolyte microshells fabricated by LbL assembling of oppositely charged
polyelectrolytes on dissoluble colloidal templates, followed by additional information on

these LbL-assembled microshells.
Biopolyelectrolyte Microshells
Polyelectrolyte Micro- and Nanoshells

In 1998, a new type of polymeric microshells was introduced and became the subject of
extensive research. Their fabrication is based on the LbL self-assembly on a colloidal
substrate followed by the dissolution of this template as sketched in Figure 2. The formed
core-shell particle is a replica of the template colloid particle, where the size varying from 0.2
to 10 m has been reported. The thickness of the shell walls is controlled by the number of
assembled polyelectrolyte layers, and can be tailored on a nanometer-scale precision. Up to
now many substances, such as synthetic polyelectrolytes [7-9], lipids [11, 12], dendrimers
[13-15], enzymes [9, 16], DNA [17], empty virues [18], and nanoparticles [19] have been
incorporated as layer components to build the multilayered shells on colloidal particles. As
cores, many types of cores including organic particles (melamine formamide, MF;
polystyrene, PS), inorganic particles (CaCO3, CdCO3, MnCO3, SiO2), organic crystals,
microgels or oil droplets [20-25] have been developed, and all require specific removal
chemistry. The main advantages of these polyelectrolyte microshells are i) the use of the mild
aqueous conditions for their synthesis, ii) the use of simple building blocks along with the
possibility to introduce a high degree of multifunctionality in shell walls, and iii) selective
permeability.
Figure 1. (A) Schematic representation of the deposition of a polyelectrolyte film on a substrate. Steps 1 and
3 represent the adsorption of a polyanion and polycation respectively, and steps 2 and 4 are washing steps.
(B) Simplified molecular picture of the first two adsorption steps depicting film deposition starting with a
positively charged substrate. Counterions are omitted for clarity. The polyion conformation and layer
interpenetration are an idealization of the surface charge reversal with each adsorption step which is the basis
of the electrostatically driven multilayer buildup depicted here. (C) Chemical structure of poly(styrene
sulfonate) (PSS) (left) and poly(allylamine hydrochloride) (PAH) (right), often used as polyions to build LbL
films. (Reprinted with permission from ref. 10. Copyright 2007, The Royal Society of Chemistry)
Figure 2. Schematic illustration of the procedure for the LbL self-assembling of a colloidal substrate followed
by the dissolution of this template. Initial steps a & b involve stepwise film formation by repeated exposure
of the colloids to polyelectrolytes with opposite charges. The excess polyelectrolytes are removed before the
next layer is deposited. Upon the desired number being obtained the core is decomposed (c) resulting in a
suspension of hollow polyelectrolyte shells (d).
Polyelectrolyte multilayer microshells may find applications in very distinct fields. They
may be used as microcontainers for the synthesis or separation of materials in restricted
volume, drug delivery, and storage systems as enzymatic and catalytic microreactors, or they
can be acted as sensor. Some examples of spatially defined synthesis in the polyelectrolyte
shells constructed by the LbL method have been reviewed [26, 27], and other reports on the
loading and release of drugs or enzymes in a controlled way have also been described
elsewhere [28, 29]. Recent advances demonstrate that biodegradable polyelectrolyte
microshells would be preferred to non-degradable ones in biomedical applications and
environment-friendly processes. Degradability of polyelectrolyte multilayers was first
introduced by Picart et al. using polysaccharides, polypeptides as building blocks [30]. These
macromolecules are prone to enzymatic degradation and drug release from such multilayers
can be considered as ―t riggered release‖ [31, 32]. Moreover, as the wall of the biodegradable
polyelectrolyte microshells was observed to be extremely thin i.e. below 100 nm they could
be of interest for the intracellular delivery of drugs [33]. In the following section, we will just
briefly review some typical biopolyelectrolyte microshells and discuss their properties. The
molecular structures of biodegradable species mentioned below are summarized in Table 1.
Table 1. Chemical structures of biodegradable species used for the fabrication of the
biopolyelectrolyte microshells
Species chemical species chemical

structure structure
H O
chitosa H
CH2 OH H H NH H
2
poly-L-glutamic H
O OH
n OH
O
O
O acid *
N
n *
H NH2 CH2OH H
(CHI) H H H (PGA) O OH
NH2.HCl
poly-L- HN
poly(hydroxypropyl * n
*
arginin NH - O NH
e methacrylamide
(PARG) dimethylaminoethyl O O
* N
H
n
*
) O
O
(P(HPMA-DMAE)) N
dextran O poly-L-lysine OH O
sulfate HO OH
(PLL) N N n
H
(DEXS) SO3-
Na+
O H
H COONa
alginat H H OH
H H OH H H
OH COONa O H H
O H OH
e O O O
OH O O
O
O H COONa
sodium H OH COONa H
OH
H
H H
H H OH
(ALG)
Natural Microshells of Alginate-chitosan

Natural polysaccharides such as chitosan (CHI) and alginate sodium (ALG) have been
widely investigated for applications in coating membranes, controlled-release drug delivery
and biomaterials [34-36]. CHI is a natural cationic polymer derived from chitin N-
deacetylation. ALG is an anionic polymer composed of a naturally occurring block
copolymer of two monosaccharide units obtained from marine brown algae. The ALG/CHI
polyelectrolyte complex (PEC) systems are commonly developed as a complex planar
membrane and an ALG gel bead is coated with CHI [37, 38], which suffers some limitations
both in controlling the membrane thickness on a nanometer-scale and in characterizing the
encapsulating PEC membrane precisely. Recently, several research groups have reported a
natural and biocompatible polyelectrolyte microshell by using ALG and CHI as building
blocks with the LBL self-assembly technique [39-42]. Colloidal particles such as MF, CaCO3
particles, PS particles and microcrystals may be used as templates in these systems involved.
The fabricated ALG/CHI shells were found to have several intriguing advantages over the
conventional polyelectrolyte microshells composed of PSS/PAH or PSS/PDADMAC,
including super mechanical/chemical stabilities and enhanced structure transformation of
shells walls in response to external microenvironment [43].
As is well-known, ALG and CHI are two oppositely charged hydrophilic natural
polysaccharides. It is observed that the constructed ALG/CHI shells in the phosphate buffer
saline PBS solution display high swelling compared to the template size. This swelling
phenomenon is of great importance because it will cause the formation of a more porous
scaffold with open porosity on the wall architecture, facilitating the permeability to various
species. Indeed, spontaneous deposition of organic substances, for example polyelectrolyte
PSS, anti-cancer drugs, nanoparticles, and dye molecules within ALG/CHI shells have been
successively achieved [42-47]. Also, it is worth mentioning that the ALG/CHI shells clearly
preserve their hollow spherical morphology and possess shape persistence in different
surrounding media.
The stability and mechanical property of polyelectrolyte microshells has been studied by
confocal laser scanning microscopy (CLSM) technique on the deformation/rupture of the
shells upon incubation in PSS polyelectrolyte solutions with different concentrations [42]. A
striking finding is that all the ALG/CHI shells preserve their spherical shape intact even when
the PSS concentrations are increased upwards to 20.0 wt% (Figure 3). In contrast to the
typical PSS/PAH or PSS/ PDADMAC shells that collapse or deform at a 4 wt% or more than
4 wt% PSS concentration [19, 48], the ALG/CHI microshells exhibit good stability and
mechanical property. This observation could be explained by considering the differences in
the architecture of polymers involved. Alginate sodium is a straight-chain polysaccharide
composed of two monomers, mannuronic acid and its C-5 epimer guluronic acid. Chitosan is
a polysaccharide polymerized by N-acetyl-D-glucosamine and glucosamine. The ALG/CHI
shells are obtained stepwise by the electrostatic interaction of the carboxylic groups of ALG
chains and the amino groups of CHI, in which the cyclohexane rings are directly linked to the
carboxylic groups of ALG chains and the amino groups of CHI. The cyclohexane rings
themselves are more flexible than the methylene groups connected to the ammonium cations
in PAH and the sulfonic acid anions in PSS because of their different arrangements and
conformations in space.
Figure 3. CLSM images of hollow shells composed of (ALG/CHI) as a function of the PSS (Mw 70,000)
bulk concentrations after incubation for 60 min. The PSS concentrations represented as wt% are indicated in
the insets. FITC-albumin was used to label the shells. Fluorescence intensity from the interior of the shells
can be eliminated by reducing the mixture time of a fluorescent label reagent and a microshell suspension.
(reprinted with permission from ref.42. Copyright 2006, Elsevier)
The wall texture changes of the ALG/CHI shells occurred upon incubating in PSS
solutions were traced by fluorescence recovery after photobleaching (FRAP) measurements
directly under CLSM (see Figure 4 and [42]). 6-carboxyfluorescein (6-CF) was chosen as a
fluorescence probe. The changes in fluorescence intensity of the closed region undergo three
stages: (i) photochemical bleaching appearing as a dark center inside the shell; (ii) recovery of
the bleached fluorescence from dark to strong; and (iii) stable fluorescence intensity in the
shell interior. The figure illustrates the fluorescence intensity in the shell interior after a
bleach pulse as a function of time. Prior to incubation the fluorescence recovery time of the
shells is ca. 700 s; while after the shells are incubated in 8 wt% PSS solution, the fluorescence
recovery becomes only ca. 40 s. The reduction of recovery time indicates the formation of
large holes in the walls. Assuming the transport is diffusive across the wall of thickness d
43 nm (obtained by SFM software analysis), the diffusion coefficient can be estimated
according to a method reported in the literature [21], namely, 0.8 10 12 cm2 s 1 for the non-
incubated shells and 1.4 10 11 cm2 s 1 for the incubated shells. The enhanced permeability
after incubation in PSS was ascribed to the rearrangement of the macromolecular layer
constituents [42]. This situation originates from the external stimuli existent in the bulk
solution, which might lead to a more porous scaffold with open porosity on the wall
architecture. Apart from these, dye 6-CF molecules can enter into the produced pores, and
then be in a different environment from those permeate into the ALG/CHI microshells. As a
result, a different intensity recovery profile from ALG/CHI without PSS incubation may be
expected.
a b
c
bleach
Intensity
recovery
0 200 400 600 800 1000 1200 1400
t/s
d
bleach
Intensity
recovery
0 20 40 60 80 100 120 140 160

t/s
Figure 4. Typical photochemical bleaching and recovery of the fluorescence of fluorescein in the interior of
(ALG/CHI) before (a & c) and after (b & d) incubation in 8.0 wt% PSS. (reprinted with permission from
ref.42. Copyright 2006, Elsevier)
Figure 5. (a) TEM image of ultramicrotomed slice for NPs-loaded ALG/CHI shells; (b) cross-section SEM
image for NPs-loaded ALG/CHI shells. (reprinted with permission from ref.43. Copyright 2007, Elsevier)
In most cases the polyelectrolyte wall is impermeable to polymers, proteins and

nanoparticles. But for the microshells composed of ALG and CHI, rigid nanoparticles (NPs)
such as polystyrene (PS) or SiO2 are readily sucked into the inner volume of the shells
through a simple incubation process, as confirmed in Figure 5. The loading amount of NPs in
a shell composed of (ALG/CHI)5 was determined in two media i.e. H2O and 0.1 M NaCl. In
pure water, the encapsulated amount for the SiO2 NPs is 6.0 10 3 g/shell, and for the PS
NPs, the encapsulated amount is 0.8 10 3 g/shell. Whereas in NaCl media, the
3
encapsulated amount is 3.5 10 g/shell for SiO2 NPs and 0.5 10 3 g/shell for PS NPs.
Apparently, the loading amount in H2O is greater than that in NaCl. The addition of Na+ ions
can enhance the electrostatic interaction of the network structure composed of ALG and CHI,
which will inevitably arose the decrease of the permeability of the shell walls, and thus lead
to the reduction of the loading amount [21]. Besides, the loading amount of NPs in the shells
is dependent on the initial concentration of the NPs added, and thus also influences the size of
NP-loaded shells.
The encapsulation process of NPs in the shells of ALG and CHI is schematically outlined
in Figure 6. There are several aspects responsible for the loading process: (i) The highly
expanded shells with enlarged holes on the wall texture are permeable to PS or SiO2 NPs.
Thus, NPs can cross the layer walls with low residual charges and enter into the shells due to
the existence of a concentration gradient of NPs between the interior and exterior of the
shells, and finally establish equilibrium between NPs suspended in the surrounding solution
and within microshells. (ii) The network architecture of ALG/CHI shell walls is mechanically
stable and shape-plastic, even upon loading relatively large rigid particles. (iii) The loaded
NPs might exist in an aggregated or complex form so that the real concentration within the
interior of the microshells is lower than in the bulk solution, thus promoting efficient
encapsulation of NPs in the shells. Still we have to stress that we do not yet have quantitative
data concerning the number and size of pores which would enable the shell to operate in an
anticipated way. Nevertheless, the quest for practical applications for the ALG/CHI system is
becoming intense owing to its proven stability and mechanical property, together with
efficient loading ability to a variety of organic species and good biocompatibility to ambient
environment, and will be detailed in the third section.
Figure 6. Schematic illustration of the procedure for the encapsulation NPs in the LbL self-assembled ALG-
CHI shells. (reprinted with permission from ref.43. Copyright 2007, Elsevier)
Self-exploding Microshells
A different example of degradable microshells, so-called self-exploding microshell, was
reported by De Geest et al. where a biodegradable dextran gel core was surrounded by an LbL
polyelectrolyte membrane [29, 49-51]. Biodegradable dextran-based microgels in a micro-
meter range can be first prepared from dextran-hydroxyethyl methacrylate (DEX-HEMA) via
a radical polymerization procedure. Then, the dextran microgels as templates were coated
with negatively and positively charged polypeptides (like poly-L-glutamic acid, PGA and
poly-L-arginine, PARG) using alternate LbL deposition. The methacrylate groups in the DEX-
HEMA microgels are connected to the dextran backbone by a carbonate ester. Thus, the
microgels are facile to be degraded by the hydrolysis of the crosslinks. The degraded products
e.g. the original dextran chains and HEMA oligomers under physiological condition are
difficult to cross through the outer layer barrier, and hence leading to the increase of the inner
swelling pressure of the polyelectrolyte microshells. As a result, the microshell membrane
ruptured. The rupturing process of these microshells is displayed in Fig 7. The degradation
time of dextran microgels can be easily tailored by varying the crosslink density of the gel
network. The whole rupturing process does not need any external trigger. The dextran-based
microgels are promising for biomedical applications given the fact that DEX-HEMA is
biocompatible and proteins can be readily incorporated inside the microgels.
Very recently, De Geest et al. developed two types of new intracellularly degradable
polyelectrolyte microshells templated onto CaCO3 microparticles. The first type of
microshells was composed of poly-L-arginine (PARG) as the polycation and dextran sulfate
(DEXS) as the polyanion. In the second type of microshells poly(hydroxypropy
lmethacrylamide dimethylaminoethyl) (P(HPMA-DMAE)) and PSS were employed as the
polycation and polyanion, respectively. Instead of using dextran-based microgels as core that
described above, CaCO3 particles filled with dextran in this system were prepared by a co-
precipitation method, and then employed as templates to alternately coating of the polyanion
and the polycation using an LbL technique until the desired number of assembled multilayers
was achieved. Subsequently, the CaCO3 core was removed by complexation with EDTA to
obtain microshells filled with dextran (Figure 8). The authors demonstrate that upon
incubating kidney cells in DEXS/pARG microshell filled with dextran, polyelectrolyte
microshells containing an enzymatically or hydrolytically degradable polycation
spontaneously degrade in chosen cells, after lipid-raft-mediated uptake [50]. Further, to
enhance the cellular uptake of the microshells, a polycation was coated as the outermost layer
because most cell types exhibit a negative surface charge. Such soft shells could be attractive
for the intracellular delivery of therapeutic nucleic acids and proteins.
Figure 7. Snapshots of A1-4) (PSS/PAH)3-coated dex-HEMA-DMAEMA microgels during the rupturing of

the membrane B1-4) uncoated dex-HEMA-DMAEMA microgels. The time intenval between the snapshorts
is 15 min. (reprinted with permission from ref.29. Copyright 2007, The Royal Society of Chemistry)
Figure 8. Schematic representation of the synthesis of polyelectrolyte shells filled with FITC-dextran using
CaCO3 particles as a template. A) co-precipitation of FITC-dextran (gray lines) in CaCO3 particles (gray
dots) during the mixing of calcium chloride and sodium carbonate solutions, B) LbL coating of the CaCO3
particles, and C) polyelectrolyte shells filled with FITC-dextran obtained after dissolution of CaCO3.
(reprinted with permission from ref.51. Copyright 2006, Wiley)
Other Biopolyelectrolyte Microshells

Yang et al. prepared a hollow shell through an LbL self-assembly of DNA and poly-L-
lysine (PLL) templated onto CaCO3 microparticles.52 The DNA/PLL shells are well dispersed
in water medium and resemble the shape and size of template particles as an intact spherical
shape. The permeability of the assembled shells was investigated by fluorescence probes with
different molecular weights and it was found that the shell walls were permeable to
macromolecules with molecular weight even as high as 150 kDa. On the basis of the fact that
the hydrodynamic radius of FITC-dextran (Mw 150 kDa) is approximately 10.7 nm [53],
and nanopore diameter in multilayer shells 2 20 nm [54], it is understandable that
fluorescence probes can penetrate multilayer walls of the DNA/PLL microshells. The release
of DNA and model drug molecules was achieved by salt-triggered shell decomposition. The
salt concentration in the incubation solution was found to exert an important influence on the
release amount and rate of DNA and drug. One special feature of this system is that two
remedial agents including DNA and drug can be simultaneously encapsulated onto the
carriers, and this might improve curative effect due to the synergetic effect of DNA and drug.
Encapsulation of active species, for example proteins in the preformed microshells may
be performed via a ―pos t-load‖ procedure. To preserve the integrity of many therapeutic
macromolecules, a key task would be to design a shell wall that is non-adhesive to most
proteins. A proper choice of layer constituents for protein encapsulation was proposed in [55].
Multilayer microsized shells made of alginate and protamine do not exhibit protein adsorption
on the shell walls. The microshells possess the high loading capacity of 109 chymotrypsin
molecules per shell. The chymotrypsin embedded in the shells maintains a high physiological
activity of about 70%. The protein release of 75-85% can be achieved after 6 h incubation at
0.08 M Tris buffer solution. The release rate of chymotrypsin can be regulated by additional
adsorption of polyelectrolyte layers onto the shells with loaded protein.
Recently the use of (strept)avidin/biotin interactions as a driving force for LbL
fabrication of films as free-standing hollow shells has been described [56]. Pegylated
multilayered architectures have been created by an LbL deposition of (strept)avidin and
biotinylated PLL through avidin/biotin complexation instead of conventional electrostatic
interaction. This technique not only extends the family of microshells, but also provides new
opportunities for the generation of robust films with tailored interfacial binding and transport
properties.
The surface charge of the shells and degree of swelling of the network forming the shell
wall are expected to have key functions. A qualitative approach in this direction is achieved
by using fluorophores of various molecular weight and CLSM for detection. Fluorophores
available in low-molecular weight range (fluorscein and rhodamin 6G) and high-molecular
weight domain for example FITC-labeled dextran were chosen for this study. Micrometer-
sized hollow shells made of chitosan/chitosan sulfate were prepared by means of electrostatic
LbL technique, and then employed to test of the shell wall permeability in different media
including water, different pHs and salt-containing solution [57]. The results illustrate the key
role of charge effect for any transport of low or high molecular weight substances through the
shell wall. Oppositely charged species are bound as long as there are free binding sites. In the
absence of salt, repulsion between the highly charged shell surface and the fluorophore
dominates even if the fluorophore is a small molecule such as fluorescein. Static light
scattering results demonstrate the polyelectrolyte complexes (PEC) rule out salt effects on the
shell‘s degree of swelling. To gain more substantial and quantitative data regarding the shell
wall permeability, macromolecular fluorophores with a narrow molecular weight distribution
would be helpful detectors for exploring size effects in terms of a cutoff limit.
APPLICATION OF BIOPOLYELECTROLYTE MICROSHELLS

Biopolyelectrolyte Microshells are of particular interest for many biomedical
applications, especially for drug delivery owing to their nontoxicity and biocompatibility to
organisms. It is evidenced that drugs, for instance anti-cancer drug (doxorubicin, DOX;
daunorubicin, DNR) and photodynamic therapy drug (hypocrellin B, HB) or model drugs can
be successfully entrapped into the biopolyelectrolyte microshells. The release of drugs or
model drugs from the interior of the shells is also involved in more updated reports and partial
details have been mentioned in the above section. To avoid the content reiteration, we in this
section will focus on a kind of system constructed by ALG and CHI as shell wall composites
and describe its applications in drug loading/release and pollutant remediation.
Drug Delivery Application of ALG/CHI Microshells
In consideration of drug formulation development, the encapsulation of anti-cancer drugs

in the microshells can a provide a means of concentrating and protecting the drug molecules
in a defined volume as well as decreasing toxic side effects to normal organics. As an
example hydrophobic photodynamic therapy (PDT) drug, HB was accumulated in the
ALG/CHI shells templated onto MF colloidal particles via a nonspecific binding [39]. The
fluorescence level indicated that after HB-loaded ALG/CHI shells being incubated in MCF-7
human breast cancer cells in vitro, the intracellular uptake efficiency of the HB-
(ALG/CHI)4/cell solution was 19.4%, while the uptake efficiency of the HB-
(ALG/CHI)4/ALG/cell solution is 1.2%. The charge of outer layer of microshells is a key to
enhancing the uptake efficiency because the cell surfaces are mainly composed of negatively
charged lipids. The XTT assay further verified that the HB-loaded ALG/CHI shells possessed
high cytotoxicity after exposure to visible light. However, the use of visible light (which does
not sufficiently penetrate the skin) limits the use of HB-loaded ALG/CHI shells as a better
photodynamic therapeutic anti-cancer agent. Notwithstanding, the uniqueness of
biopolyelectrolyte mictoshells as PDT drug transporters is beginning to emerge and needs to
be fully developed.
DNR can induce apoptosis or namely, programmed cell death of tumor cells by blocking
the cell cycle and inhibiting the DNA polymerase enzyme [40, 58]. Likewise, the clinical
application of DNR has often suffered from the development of tumor cells‘ resistance and
toxicity. To solve these problems layered biopolyelectrolyte microshells made of ALG/CHI
were introduced by Peng et al. and the controlled release of DNR-loaded microshells might
be a perspective pathway [40]. The ALG/CHI shells were fabricated by a template-assisted
assembly in an LbL manner, followed by removal of CaCO3-carboxymethyl cellulose
colloidal particles. The ALG/CHI shells showed strong ability to load DNA, capable of
reaching up to several hundred times of the feeding concentration. The encapsulated DNR can
be released in pH 7.4 PBS or 0.1M HCI solution and the release follows a diffusion-
controlled procedure at the initial stage. In vitro BEL-7420 culture verified the efficacy of the
loaded DNR on induction of cell apoptosis. The animal experiment in vivo showed that the
encapsulated DNR had better ability of tumor inhabitation than the same dosage of free DNR,
although the experimental dosages could not completely inhibit the growth of the tumors.
In a similar formulation, anti-cancer drug DOX or peptide BH3 has also been entrapped
in the interior of the ALG/CHI shells under modest conditions without addition of other
reagents [47]. The mass of DOX loaded in one capsule of four alginate/chitosan layers (i.e.
the volume V = 2.5 × 10-10 cm3) is calculated as ca. 1.4 × 10-13 g, which corresponds to 1.5 ×
108 DOX molecules. Also, the release of drug in the shells is dependent on the number of
assembled layers of the shells. Colorimetric XTT cell viability assay results showed that the
drug-loaded microshells at high concentrations tested could kill cancer cells more efficiently
than free-drug alone. The binding of the microshells to human lung cancer cells would
effectively result in a higher local concentration of drug in the direct vicinity of the cells and
likely accounts for the greater cytotoxicity observed.
The Tong group recently reported several papers on using ALG and CHI as shell wall
components for the encapsulation and controlled release indomethacin (IDM, a non-steroidal
anti-inflammatory drug) microcrystals [41, 59-60]. The authors demonstrated that upon being
incubating in an enzyme pepsin solution, the multilayer ALG/CHI film was partially
destroyed because of the enzymatic degradation of CHI. By means of increasing the layer
number and raising the deposition temperature, the stability of the ALG/CHI multilayer film
to the enzymatic erosion is enhanced accordingly. These results provide effective methods to
protect the multilayer film fabricated using LbL assembly from the enzymatic erosion and to
prolong the release of the encapsulated drug.
Additionally, the ALG/CHI microshells fabricated through the LbL technique have also
successfully used as carriers for the loading and release of other distinct drugs including
ibuprofen, acridine hydrochloride and insulin [61-64].
Concluding this section we should also comment on universal applicability of the shells
composed of ALG/CHI. It has been shown that the ALG/CHI shells can be potentially
applicable as drug delivery vehicles for various active drugs. Thus, the proposed formulation
can be extended by simultaneously loading two or more kinds of drugs inside the shells
adjusted to the specific application in patients.
Pollutant Remediation Application of ALG/CHI Microshells
With the development of chemical and pharmaceutical industries, many recalcitrant

organics accumulate in water and suffer the risk of contamination of the underground sources
in an irreversible way [65]. The existence of these pollutants in excess of a few parts per
billion (ppb) in water could lead to serious health problems [66]. Great effort has been made
using widely-called ―adv anced oxidation processes‖ (AOPs) for treatment of these
recalcitrant pollutants to more biodegradable molecules or mineralization into CO2 and other
inorganics [67-69]. However, using AOPs to remove pollutants suffers from the drawback of
an excessive loss of the active species generated prior to reacting with pollutants at a lower
concentration level. In this case, a perspective solution has been proposed in by introducing
the LbL assembled biopolyelectrolyte microshells into organic species-polluted systems for
pre-adsorption, combined with Fenton reagent (one of advanced AOPs). Besides, the pre-
encapsulation of harmful compounds also provides a promising pathway to minimize the
pollution problem via a post-treatment of photocatalytical degradation after use. The
following paragraphs review these solutions that have been exploited to effectively remove
organic pollutants from waste water.
The system we propose here makes use of a natural green polyelectrolyte microshell
composed of ALG and CHI as a microreactor/microcontainer to load organic pollutants. The
loading procedure was performed under moderate conditions (room temperature, pure water
media) by adding a suspension of the ALG/CHI shells to a solution of pollutant overnight
[44, 45]. The accumulated amount of pollutants in one shell composed of (ALG/CHI)5 are 6.0
10-11g for rhodamine B (RhB) and 4.2 10-11g for fluorescein (Flu), which corresponds to
7.5 1010 RhB molecules and 6.7 1010 Flu molecules, respectively. The accumulated
ALG/CHI microshells were then redispersed in the Fenton reagent (Fe3+ and H2O2) after
simple filtration. As stated above, the ALG/CHI shells are permeable to dye molecules,
polyelectrolytes, dextran, and rigid NPs. Thus, the Fenton reagent can cross the shell wall
barrier and enter into the shells, and then react with loaded pollutants under light radiation.
CLSM images provide direct observation on the changes of the accumulated dyes inside the
shells before and after the photoreaction (Figure 9). During the whole photodegradation no
fluorescence of dye is observed in the bulk solution. This means that the dye molecules are
well located in the shell interior and that the photoreaction process mainly takes place in the
microshells. Compared with the hollow ALG/CHI shells, no apparent change in surface-wall
texture occurs upon accumulation of the shells with RhB, whereas after the photoreaction, an
obvious enhancement in the RhB-accumulating shell wall is observed (Figure 10)
Importantly, in the two consecutive recycling of accumulation and subsequent
photodegradation of pollutants, the constructed ALG/CHI microshells remain intact spherical
shape, indicating that the shells are stable against attack from highly active species generated
in a photo-assisted Fenton reaction.
Figure 9. Variations in CLSM fluorescence images of the (ALG/CHI)5 shells accumulated with RhB in the
presence of Fe3+ (2×10-4 M) and H2O2 (5×10-4 M) before (a) and after irradiation for 45 (b) and 60 min (c),
respectively. During the measurements the optical parameters of the CLSM remained unchanged. (d) TEM
image of the RhB-accumulated shells under the same condition as (c). (reprinted with permission from ref.44.
Copyright 2005, The Royal Society of Chemistry)
a b
c d
Figure 10. SFM images of the air-dried (ALG/CHI)5 shell (a), RhB-accumulating (ALG/CHI)5 shell (b), and
RhB-accumulating (ALG/CHI)5 shell after 60 min of visible radiation in a solution of Fe3+ (0.2 mm) and
H2O2 (0.5 mm) (c). d) HR-SFM image of the area 300 nm × 300 nm marked in (a) showing the surface
texture of the (ALG/CHI)5 microshell free of folds. (reprinted with permission from ref.45. Copyright 2008,
Wiley)
To enhance the catalytic activity of the reaction system and the stability of the catalysts,
species with catalytic activity are generally bound on appropriate support materials such as
neutral organic polymers, ion exchange membranes or resins, and inorganic materials (clay,
zeolites et al.) [70-73]. Furthermore, these supported catalysts can also be easily separated
and re-utilized via a simple centrifugation/filtration process [74]. By combination with the
advantages of heterogeneous catalysts and proper characteristics of hollow shells, a novel
heterogeneous Fe-immobilized polyelectrolyte microshells has been constructed by
electrostatic LbL self-assembly technique, in which natural polymers e.g. ALG and CHI
along with perchloric acid iron are used as shell wall components [75]. We find that the Fe-
immobilized shells i.e. Fe/ALG/CHI shells can efficiently accumulate the dye molecules
entering into the shells under ordinary conditions. The loading amount of dyes in the shells is
mainly attributed to the nature of internal layer of the shells themselves. The accumulated
dyes in the interior of shells can be degraded at a rapid rate by H2O2 oxidation under visible
light at a wide range of pH from acid to neutral suspension. Furthermore, the photoreaction
rate of accumulated in the Fe/ALG/CHI shells in acid media is relatively faster than that in
neutral media (Fig 11). It is of great importance here that this system can avoid little activity
for substrate degradation at pH > 4 (Fe ions precipitate) as well as difficult post-treatment of
Fe sludge after the reaction. The whole procedure for the accumulation and subsequent
photodegradation of pollutants in the shell interior is illustrated in Figure 12.
The nature of short-lived radicals formed during the photoreaction under visible
irradiation in two different media can be examined by spin-trapping ESR technique [76]. The
results show that in an acid medium HO• radicals are involved in the photoreaction process,
while in a neutral medium the photoreaction process might not be mainly HO• radical-
dependent and •OOH/O2• radicals as active oxygen intermediates might be involved in the
reaction process. In such a case, the faster photoreaction rate of pollutant defined in the
microspace in an acid solution than in a neutral solution might be ascribed to higher oxidative
activity of HO• radicals than that of •OOH/O2•− radicals involved in the photoreaction
studied [77-79]. More detailed study on the photoreaction mechanism is still needed.
The other way to minimize the pollution problem is to encapsulate imidacloprid (IMI,
one kind of insecticide) microcrystals with CHI and ALG by LbL assembly for prolonged
release, and then adsorb different kinds of photocatalysts including TiO2, SDS/TiO2, Ag/TiO2
and SDS/Ag/TiO2 for photocatalytic degradation and mineralization of nano-IMI [80]. It has
been shown that the IMI loading and encapsulation efficiency reach up to approximately 56%
and 81%, respectively. The polysaccharide shells prolong the release time of the encapsulated
IMI crystals. Among the photocatalysts SDS/Ag/TiO2 exhibits the highest photocatalytic
activity. This study establishes a model for the effective utilization of any kinds of insecticide
with microcrystal structure.
1.0
0.8
d
0.6
0
c
C/C
0.4
a
0.2
b
0.0
0 20 40 60 80 100 120 140
Irradiation time \ min
Figure 11. Photoreaction rate of accumulated dyes in two sets of conditions (a) RhB, pH 2.7; (b) AO, pH 2.7;
(c) RhB, pH 6.0 and (d) AO, pH 6.0. (reprinted with permission from ref.75. Copyright 2008, Elsevier)
Figure 12. Schematic illustration of the procedure for the visible light-assisted degradation of dye pollutants
accumulated in the preformed Fe-immobilized polyelectrolytes microshells. (reprinted with permission from
ref.46. Copyright 2008, Bentham Science Publishers Ltd)
The above-mentioned approaches have several significant advantages for the remediation
of pollutants: i) it allows the accumulation of a variety of pollutants under moderate
conditions in shells comprised of natural ALG/CHI polyelectrolytes, ii) the photodegradation
reaction can proceed in aqueous media without addition of other organic reagents, and iii) the
assembled shells possess good photostability, and iv) due to the tailored shell thickness on a
nanometer scale as well as the ordered shell composition and the versatility of the LbL
method, this technique is likely to be extended to other homogeneous/heterogeneous
photocatalytic systems for the remediation of organic pollutants in aqueous ecosystems.
CONCLUSION
The controlled LbL assembly of biopolyelectrolytes on removable colloidal particles can
be used to create multilayer microshells with customized properties. This kind of
biopolyelectrolyte microshell could become a new class of molecular transporters for various
in vitro and in vivo delivery applications as well as be used as an environment-benign
microcontainer for water treatment. This domain of research will probably be one of the most
exciting in the fields of biotechnology and medicine, driven by biopolyelectrolyte microshells
with new combinations of properties.
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Lecture Material 24
SYNTHESIS AND ELECTRON FIELD EMISSION FROM

DIFFERENT MORPHOLOGY CARBON NANOFIBERS
ABSTRACT
Nanofibers are rocking the commercialization in lighter material science. Best example is
single walled carbon nanotube materials in aerospace and aviation industry. Basically their
spatial carbon carbon conformation and configuration in the molecular make up plays
significant role in their properties. A simple quantum description is given here on atomic
structure and molecular conformation of fullerene carbon nanotube material. The synthesis,
field emission properties and applications are reviewed of nanofiber sheet structures.
INTRODUCTION
Carbon Nanofiber and Carbon Nanotube
Carbon, which belongs to group IV of the periodic table, is the lightest element in this
group, and it possesses countless interesting physical and chemical properties. Among the
different types of supports used in heterogeneous catalysis carbon materials attract a growing
interest due to their specific characteristics which are mainly: (i) resistance to acid / basic
media, (ii) possibility to control, up to certain limits, the porosity and surface chemistry and
(iii) easy recovery of precious metals by support burning resulting in a low environmental
impact. In contrast to Si, Ge and Sn, which have the same number of electrons in the
outermost shell as carbon and can only exist in cubic sp3 hybridization, carbon not only
exhibits sp3 hybridization (diamond), but also planar sp2 hybridization as in the graphite
structure and sp1 hybridization as in carbynes. Each carbon atom has six electrons which
occupy 1s2, 2s2 and 2p2 atomic orbitals. The 1s2 orbital contains two strongly bound core
electrons. Four more weakly bound electrons occupy the 2s22p2 valence orbitals. In the
crystalline phase, the valence electrons give rise to 2s, 2px, 2py and 2pz orbitals which are
important in forming covalent bonds in carbon materials. Since the energy difference between
the upper 2p energy levels and the lower 2s level in carbon is small compared with the
binding energy of the chemical bonds, the electronic wave functions for these four electrons
can readily mix with each other, thereby changing the occupation of the 2s and three 2p
atomic orbitals so as to enhance the binding energy of the carbon atom with its neighboring
atoms. The general mixing of 2s and 2p atomic orbitals is called hybridization, whereas the
mixing of a single 2s electron with one, two, or three 2p electrons is called spn hybridization
with n = 1,2,3 [1,2]. The bonding structures of diamond, graphite and nanotubes, or fullerenes
are shown in Figure 1.1. When a graphite sheet is rolled over to form a nanotube, the sp2
hybrid orbital is deformed for rehybridization of sp2 toward sp3 orbital or bond mixing.
This rehybridization structural feature, together with electron confinement, gives
nanofibers/nanotubes unique, extraordinary electronic, mechanical, chemical, thermal,
magnetic, and optical properties [3-5].
The physical reason why these nanostructures form is that a graphene layer (defined as a
single 2D layer of 3D graphite) of finite size has many edge atoms with dangling bonds,
index dangling bonds and these dangling bonds correspond to higher energy states. Therefore
the total energy of a small number of carbon atoms (30 -100) is reduced by eliminating
dangling bonds, even at the expense of increasing the strain energy, thereby promoting the
formation of closed cage clusters such as fullerenes and carbon nanotubes. For example,
diamond and layered graphite forms of carbon are well known, but the same carbon also
exists also in planar sheet, rolled up tubular, helical spring, rectangular hollow box, and nano-
conical forms. Elemental carbon in the sp2 hybridization can form a variety of amazing
structures. Apart from the well-known graphite, carbon can build closed and open cages with
honeycomb atomic arrangement. First such structure to be discovered was the C60 molecule
by Kroto et al. in 1985 [1]. Although various carbon cages were studied, it was only in 1991,
when Iijima [6] observed for the first time tubular carbon structures. Two years later, Iijima
and Ichihashi [7] and Bethune et al. [8] synthesized single-walled carbon nanotubes
(SWNTs). Actually carbon nanotubes (CNTs) are allotropes of carbon. Nanotubes are
members of the fullerene structural family, which also includes buckyballs. Whereas
buckyballs are spherical in shape, a nanotube is cylindrical, with at least one end typically
capped with a hemisphere of the buckyball structure. The nature of the bonding of a nanotube
is described by applied quantum chemistry, specifically, orbital hybridization. Nanotubes are
composed entirely of sp2 bonds, similar to those of graphite. This bonding structure, which is
stronger than the sp3 bonds found in diamond, provides the molecules with their unique
strength. Nanotubes naturally align themselves into "ropes" held together by Van der Waals
forces. Under high pressure, nanotubes can merge together, trading some sp2 bonds for sp3
bonds, giving great possibility for producing strong, unlimited-length wires through high-
pressure nanotube linking [9].
A single-wall carbon nanotube (SWCNT) is best described as a rolled-up tubular shell of
graphene sheet (Figure 1.2(a)) which is made of benzene-type hexagonal rings of carbon
atoms [10-12]. There are many possible orientations of the hexagons on the nanotubes, even
though the basic shape of the carbon nanotube wall is a cylinder. A single walled carbon
nanotube is a graphene sheet appropriately rolled into a cylinder of nanometer size diameter
[13,14]. The planar sp2 bonding, which is characteristic of graphite, plays a significant role in
carbon nanotubes. The body of the tubular shell is thus mainly made of hexagonal rings (in a
sheet) of carbon atoms, whereas the ends are capped by half-dome shaped half-fullerene
molecules. The internal diameter of these structures can vary between 0.4 and 2.5 nm and the
length ranges from few microns to several millimeters. vectors. Thus, although carbon
nanotubes are closely related to a 2D graphene sheet, the tube curvature and the quantum
confinement in the circumferential direction lead to a host of properties that are different from
those of a graphene sheet.
A multi-wall carbon nanotube (MWCNT) is a rolled-up stack of graphene sheets of
coaxial SWCNTs, with the ends again either capped by half-fullerenes or kept open. Both
SWCNTs and MWCNTs have physical characteristics of solids and are nanocrystals with
high aspect ratios of 1000 or more, although their diameter is close to molecular dimensions.
The number of walls present can vary from two (double wall nanotubes) to several tens, so
that the external diameter can reach 100 nm. The concentric walls are regularly spaced by
0.34 nm similar to the inter graphene distance evidenced in turbostatic graphite materials. The
main difference between nanotubes and nanofibers consists in the lack of a hollow cavity for
the latter. The diameters of carbon nanofiber (CNF) are generally higher than the ones
presented by nanotubes and can easily reach 500 nm. A nomenclature (n,m), used to identify
each single-wall nanotube, refers to integer indices of two graphene unit lattice vectors
corresponding to the chiral vector of a nanotube. Chiral vectors determine the directions
along which the graphene sheets are rolled to form tubular shell structures and perpendicular
to the tube axis vectors. Figure 1.2 shows the schematic representation of the construction of
a nanotube by rolling-up an infinite strip of graphite sheet (so called graphene). In Figure
1.2(a) the chiral vector Ch = na1 + ma2 connects two lattice points O and A on the graphene
sheet, where n and m are integers, a1 and a2 the unit cell vectors of the two-dimensional lattice
formed by the graphene sheets. The direction of the nanotube axis is perpendicular to this
chiral vector. An infinite strip is cut from the sheet through these two points, perpendicular to
the chiral vector. The strip is then rolled-up into a seamless cylinder. T = t1a1 + t2a2 is the
primitive translation vector of the tube [15]. The nanotube is uniquely specified by the pair of
integer numbers n, m or by its radius R = Ch / 2 and chiral angle which is the angle
between Ch and the nearest zigzag of C–C bonds. All different tubes have angles between
zero and 30o. Special tube types are the achiral tubes (tubes with mirror symmetry): when n =
m, the nanotube is called ―a rmchair‖ type (θ = 0◦) [Figure 1.2(b)]; when m = 0, then it is of
the ―zi gzag‖ type (θ = 30◦) [Figure 1.2(c)]. Otherwise, when n = m, it is a ―chi ral‖ tube and θ
takes a value between 0◦ and 30◦ [Figure 1.2(d)]. The value of (n,m) determines the chirality
of the nanotube and affects the optical, mechanical and electronic properties. Nanotubes with
|n - m| = 3q are metallic and those with |n - m| = 3q ± 1 are semiconducting (q is an integer).
Figure 1.1. Bonding structures of diamond, graphite, nanotubes and fullerenes.

Since the single wall carbon nanotube is only one atom thick and has a small number of
atoms around its circumference, only a few wave vectors are needed to describe the
periodicity of the nanotubes. These constraints lead to quantum confinement of the wave
functions in the radial and circumferential directions, with plane wave motion occurring only
along the nanotube axis corresponding to a large number or closely spaced allowed wave
Exhaustive studies concerning electronic properties of both SWCNT [16] and MWCNT
[17] are available in the literature, whereas carbon nanofiber (CNF) are often considered as
conductive substrates that can exert electronic perturbations similar to those of graphite [18].
In the case of SWCNT, studies have demonstrated that they behave like pure quantum wires
(1D-system) where the electrons are confined along the tube axis. Electronic properties are
mainly governed by two factors: the tube diameter and the helicity, which is defined by the
way in which the graphene layer is rolled up (armchair, zigzag or chiral) [8]. In particular,
armchair SWCNTs are metallic and zigzag ones display a semi-conductor behavior. Studies
on MWCNTs electronic properties have revealed that they behave like an ultimate carbon
fiber at high temperature their electrical conductivity may be described by semi-classical
models already used for graphite, whereas at low temperature they reveal 2D-quantum
transport features [17]. Nanotubes can be as small as 1 nm in diameter and as long as 100,000
nm.
These tubes are extremely strong, approaching the strength of diamond, and also dissipate
heat better than any other known material. Carbon nanofibers/nanotubes are one of the
strongest and stiffest materials known, in terms of tensile strength and elastic modulus
respectively.
Figure 1.2. Schematic representation of the construction of a nanotube by rolling-up an infinite strip of
graphite sheet.
This strength results from the covalent sp2 bonds formed between the individual carbon
atoms. Depending on how they are configured, CNFs/CNTs are good conductors of electricity
and can also act as semi-conductors for molecular electronics. CNFs/CNTs are three
dimensional as opposed to the current silicon based electronics that are two-dimensional.
They appear to be able to extend the miniaturization process by several additional orders of
magnitude over current methods. As they conduct electricity better than copper and can also
act as semi-conductors. CNFs/CNTs have very good elasto-mechanical properties because the
two dimensional arrangement of carbon atoms in a graphene sheet allows large out-of-plane
distortions, while the strength of carbon-carbon in-plane bonds keeps the graphene sheet
exceptionally strong against any in-plane distortion or fracture. Some of the properties of
carbon nanotube, which are forming a driving force for their wide range of applications, are
shown in Table -1.
Table 1. Some fundamental properties of carbon nanofiber/nanotube
Properties Value References

Average Diameter: SWNT's 1.2-1.4 nm [19]
MWCNTs 2-100 nm [20]
CNFs 10 nm - [21]
Young's Modulus ~ 1 TPa [22,23]
Maximum Tensile Strength ~ 63 Gpa [24]
Band Gap: For Metallic 0 eV [25]
For Semi-Conducting 0.18 – 1.8 eV [25]
Thermal Conductivity (Room Temp.) 3000 W/mK [26]
Carrier mobility Semi-Conducting NT 105 cm2/Vsec [27]
Maximum Current Density 109 A/m2 [28]
Turn-on field 1.5 – [29,30]
Threshold field for CNTs/CNFs 1.5 – [31,32]
History of Carbon Nanofiber and Nanotube
We provide here a brief review of the history of carbon fibers, the macroscopic analog of
carbon nanotubes, since carbon nanotubes have become the focus of recent developments in
carbon fibers. Since last decade, new carbon forms like carbon nanofibers (CNF) or nano
filaments and carbon nanotubes (CNT) have generated an interest in the scientific
community. However, it has got to be remembered that carbon nano filaments have been
synthesized for very long as products from the action of a catalyst over the gaseous species
originating from the thermal decomposition of hydrocarbons. One of the first evidence that
the nano filaments thus produced could have been nanotubes, exhibiting an inner cavity, can
be found in the transmission electron microscope micrographs published by Hillert et al. in
the year of 1958 [33]. Radushkevich et el. published clear images of 50 nanometer diameter
tubes made of carbon in the year 1952 [34]. This discovery was largely unnoticed, the article
was published in the Russian language. The production of graphite nanofibers is even older
and the first reports date of more than a century [35,36]. Their efforts were mostly directed
toward the study of vapor grown carbon filaments, showing filament growth from the thermal
decomposition of hydrocarbons. The second applications-driven stimulus to carbon fiber
research came in the 1950‘s from the needs of the space and aircraft industry for strong, stiff
light-weight fibers that could be used for building lightweight composite materials with
superior mechanical properties. This stimulation led to great advances in the preparation of
continuous carbon fibers based on polymer precursors, including rayon, polyacrylonitrile
(PAN) and later mesophase pitch. The late 1950‘s and 1960‘s was a period of intense activity
at the Union Carbide Corporation, the Aerospace Corporation and many other laboratories
worldwide. This stimulation also led to the growth of a carbon whisker [37], which has
become a benchmark for the discussion of the mechanical and elastic properties of carbon
fibers. The growth of carbon whiskers was also inspired by the successful growth of single
crystal whisker filaments at that time for many metals such as iron, non-metals such as Si, and
oxides such as Al2O3, and by theoretical studies [38], showing superior mechanical properties
for whisker structures [39]. Parallel efforts to develop new bulk synthetic carbon materials
with properties approaching single crystal graphite led to the development of highly oriented
pyrolytic graphite (HOPG) in 1962 by Ubbelohde and co-workers [40,41], and HOPG has
since been used as one of the benchmarks for the characterization of carbon fibers. While
intense effort continued toward perfecting synthetic filamentary carbon materials, and great
progress was indeed made in the early 1960‘s, it was soon realized that long term effort
would be needed to reduce fiber defects and to enhance structures resistive to crack
propagation. New research directions were introduced because of the difficulty in improving
the structure and microstructure of polymer-based carbon fibers for high strength and high
modulus applications, and in developing graphitizable carbons for ultra-high modulus fibers.
Because of the desire to synthesize more crystalline filamentous carbons under controlled
conditions, synthesis of carbon fibers by a catalytic Chemical Vapor Deposition (CVD)
process was developed, laying the scientific basis for the mechanism and thermodynamics for
the vapor phase growth of carbon fibers in the 1960‘s and early 1970‘s. In parallel to these
scientific studies, other research studies focused on control of the process for the synthesis of
vapor grown carbon fiber [42-45], leading to the more recent commercialization of vapor
grown carbon fibers in the 1990‘s for various applications. Concurrently, polymer-based
carbon fiber research has continued worldwide, mostly in industry, with emphasis on greater
control of processing steps to achieve carbon fibers with ever-increasing modulus and
strength, and on fibers with special characteristics, such as very high thermal conductivity,
while decreasing costs of the commercial products. As research on vapor grown carbon fibers
on the micrometer scale proceeded, the growth of very small diameter filaments less than 10
nm, was occasionally observed and reported [46-47], but no detailed systematic studies of
such thin filaments were carried out. Oberlin et al. clearly showed hollow carbon fibres with
nanometer-scale diameters using a vapour-growth technique [48]. The interest in fibrous
carbon has since then been recurrent and a significant boost in the research in carbon
nanostructure field coincides with the discovery of multiwall carbon nanotubes (MWNT) by
Iijima in 1991 [6]. It is likely that carbon nanotubes were produced before this date, but the
invention of the transmission electron microscope allowed the direct visualization of these
structures. Carbon nanotubes have been produced and observed under a variety of conditions
prior to 1991. The arc discharge technique was well known to produce the famed
Buckminster fullerene on a preparative scale [49] and these results appeared to extend the run
of accidental discoveries relating to fullerenes. The original observation of fullerenes in mass
spectrometry was not anticipated [5], and the first mass-production technique by Kratschmer
et al. was used for several years before realising that it produced fullerenes [49].
SYNTHESIS AND GROWTH MECHANISM OF CARBON NANOTUBE AND

CARBON NANOFIBER BY DIFFERENT PROCESS
There have been major developments in the synthesis processes and characterization
techniques of CNT and CNF. It is possible to produce CNTs by a wide range of deposition
methods. The most popular deposition techniques are arc discharge [6,50], laser ablation [51],
chemical vapour deposition (CVD) [52,58-62] plasma enhanced chemical vapour deposition
[53] and solvothermal process [54-55]. The earliest approach to produce nanotubes was an arc
process [56] as pioneered by Iijima in 1991. This was shortly followed by a laser ablation
technique developed at Rice University [57]. Chemical vapor deposition (CVD) has become a
common technique to grow nanotubes in the last five years [58-62]. The figure-of-merit for
an ideal growth process depends on the application. For development of composites and other
structural applications, the expected metric is the ability to achieve controlled growth of
specified thickness on patterns is important for applications in nano electronics, field
emission, displays, and sensors. The arc process involves striking a dc arc discharge in an
inert gas (such as argon or helium) between a set of graphite electrodes [6,56]. The electric
arc vaporizes a hollow graphite anode packed with a mixture of a transition metal (such as Fe,
Co or Ni) and graphite powder. The inert gas flow is maintained at 50-600 Torr. Nominal
conditions involve 2000 - 3000 C, 100 amps and 20 volts. This produces SWCNTs in
mixture of MWCNTs and soot. In the arc-discharge synthesis of nanotubes, Bethune et al. in
1993 used as anodes thin electrodes with bored holes which were filled with a mixture of pure
powdered metals (Fe, Ni or Co) and graphite [8]. The electrodes were vaporized with a
current of 95-105 A in 100-500 Torr of He. The gas pressure, flow rate, and metal
concentration can be varied to change the yield of nanotubes, but these parameters do not
seem to change the diameter distribution. Typical diameter distribution of SWCNTs by this
process appears to be 0.7 - 2 nm. On the other hand in laser ablation method, a target
consisting of graphite mixed with a small amount of transition metal particles such as such as
Co, Ni, Fe etc., catalyst is placed at the end of a quartz tube enclosed in a furnace [57]. A
neodymium-yttrium-aluminum-garnet laser was employed to vaporize the target and helium
or argon carrier gas was flowed through the tube. Argon gas flowing through the reactor,
heated about 1200 C by a furnace, carries the vapor and nucleates the nanotubes which
continue to grow. The nanotubes are deposited on the cooler walls of the quartz tube
downstream from the furnace.
Both the above-mentioned methods were used to synthesize SWNTs in relatively large
quantities. They are based on the condensation of hot carbon gases through vaporizing solid
carbon, where temperatures of > 3000 K are initialized by either arc or laser. Due to such a
high temperature, carbon nanotubes obtained exhibit high straightness and high crystallinity.
However, depositions by both approaches are not directly on the substrates and are in a form
of either powder or mat. Thus applications require additional manipulation or processes to
deposit the CNTs on substrates. Lastly, the products obtained normally contain a large
quantity of catalyst and carbon particles so that a purification step becomes necessary.
Chemical vapor deposition (CVD) methods have been successful in making carbon fiber and
filament since more than 10-30 years ago [46,63,64], but in recent years, chemical vapor
deposition (CVD) has progressed rapidly towards the growth of carbon nanotubes and has
been adopted worldwide [65]. In this method, transition catalysts such as Fe, Co, Ni are
deposited on the substrates (silicon, quartz, molybdenum etc.). A mixture of precursors such
as hydrogen, methane, acetylene, and ammonia are flowed into the chamber. Assisted by
either direct heating or by an external source such as plasma and hot filament, carbon gases
are decomposed at the catalyst surface and carbon nanotubes are grown directly on the
substrates at temperatures between ~ 700-1400 K. The advantage of this method is that the
nanotubes can be deposited directly on the substrate, which facilitates nanotube applications
and mass production. Nevertheless, MWNTs are often obtained by CVD and only until
recently SWNT growth by CVD was possible [66]. The drawback of the catalytic CVD-based
nanotube production is the inferior quality of the structures that contain gross defects (twists,
tilt boundaries etc.), particularly because the structures are created at much lower
temperatures (600-1000 oC) compared to the arc or laser processes (~2000 oC). Several
plasma based growth techniques have been reported [67-69] and in general, the plasma-grown
nanotubes appear to be more vertically oriented than that is possible by thermal CVD. Since
the plasma is very efficient in tearing apart the precursors and creating radicals, it is also hard
to control and keep the supply of carbon low to the catalyst particles and hence, plasma based
growth always results in MWCNTs and filaments. Different CVD systems which employ
different approaches to dissociate the precursor gases have been used including thermal CVD
[70], hot filament chemical vapour deposition (HFCVD) [53], and plasma enhanced chemical
vapor deposition (PECVD) [71]. Several types of plasma systems which have been used
including dc-plasma [72], radio frequency (RF) plasma [73], microwave plasma [74], and
electron cyclotron resonance (ECR). In CVD technique, CNTs and CNFs are grown using the
catalytic decomposition of hydrocarbons over transition metal catalysts such as iron, cobalt
and nickel at temperatures ranging from 550 to 1000 oC [52]. The function of these metals is
to facilitate the decomposition of the hydrocarbon gases and the formation of the tubular
graphene structures. Chemical composition and particle size of the catalyst is expected to
crucially affect the diameter and the number of walls of the carbon nanotubes [75]. The metal
catalysts have been prepared by several methods including wet chemical solution [76], thin
metal films [71], thick metal films/substrates [77], colloids [75], and sol-gel techniques [61].
In some cases surface treatments such as wet chemical etching [67], plasma etching [77], ion
beam sputtering [62] and annealing [71] have been used to enable the formation of nano-
particles before the growth. Effect of catalyst on growth of carbon nanofibers have been
studied by Kamada et al. [78]. They have been successfully grown carbon nanofiber (CNF)
films on Pd-Se, Fe-Ni, and Ni-Cu alloy catalysts at low temperatures by a thermal chemical
vapor deposition method. Among these alloy catalysts, Ni-Cu alloy catalyst was found to be
most suitable for low temperature growth of CNF. The CNFs grown using Pd-Se catalyst
were found to have more defective structure than that obtained with the other catalysts, and
exhibited excellent field emission property with threshold field ~ 1.1 V/ m. Merkulov et al.
[79] evaporated Ni on n-type Si by E-gun. Shyu et al. deposited Fe-Ni with various
components by e-beam evaporation [80]. Kin et al. [81] prepared copper-nickel powder by
coprecipitation of the metal carbonates from mixed nitrate solutions using ammonium
bicarbonate and a sequence complex treatment process including drying, calcining and
reducing, etc. Carbon nanofibers (CNFs) were grown on a Ni–P alloy catalyst deposited on a
silicon substrate in a microwave heating chemical vapor deposition system with methane gas
at 650 oC [82]. The nanosized clusters on the clustered surface of the Ni–P alloy catalyst film
directly provided the nucleation sites for CNFs without any pretreatment before the growth of
the CNFs. The CNFs grown on the Ni–P alloy catalyst showed random orientation and it
composed of parallel graphite planes. Figure 2.1 shows scanning electron microscopy images
of CNFs grown at substrate temperature approximately 650 oC for 7 min. It reveals that the
growth rate of CNFs is related to the thickness of Ni–P alloy catalyst film. The results
indicate that the growth rate of the CNFs decreases as the thickness of the catalyst film
increases. The above phenomenon explained by the diffusion of carbon atom into the catalyst
particle. The growth of carbon nanostructures, including CNTs and CNFs, occurs by diffusion
driven precipitation of carbon atoms from the supersaturated catalyst particles [83]. The size
of catalyst particle increases and that causes the diffusion length to increase and the gradient
of supersaturation to decrease. These factors will decrease the growth rate of CNFs. So, the
thin catalyst film has a larger growth rate than the thick catalyst film. The result also proves
that diffusion of carbon through the catalyst particle is the rate-determining step in the growth
of carbon nanostructures using a Ni-P alloy catalyst.
Figure 2.1. SEM images of CNFs grown at 650 0C substrate temperature for 7 min. The corresponding
thickness of Ni–P alloy catalyst film is 20 and 40 nm in images (a) and (b) respectively [from ref. 82].
Figure 2.2 is the SEM micrograph showing the surface morphology of CNFs grown at
approximately 650 oC for 10 min. All of these SEM images show that these CNFs grown on
the catalyst film with various thicknesses have similar morphology and are not vertically
aligned but randomly tangled. The diameter of the CNFs in Figure 2.2(a)-(c) is approximately
30-70, 50-120 and 70-150 nm, respectively. The diameter of the CNFs increases as the size of
the catalyst clusters increases with catalyst film thickness. The results show that the diameter
of the CNFs is dependent on the initial thickness of the pre-deposited catalyst film. Wei et al.
[69], Yudasaka et al. [84] and Bower et al. [85] also reported same type results using Fe, Co
and Ni catalyst.
Figure 2.2. SEM images of CNFs grown at 650 0C substrate temperature for 10 min. The
corresponding thickness of Ni–P alloy catalyst film is 20, 30 and 40 nm in images (a), (b) and (c),
respectively [from ref. 82].
Vertically aligned carbon nanofiber and naotubes were synthesized by different technique
and there are different growth mechanism proposed by different groups [86-92]. The physical
and chemical characteristics of vertically aligned carbon nanofiber (VACNF) structures, in
comparison to ideal multiwalled carbon nanotubes, offer inherent processing advantages
imparted by their vertical architecture. The ability to control the VACNF growth rate is an
important practical aspect of the synthesis process because some applications require high
growth rates, whereas others would benefit from a lower growth rate but a high degree of
uniformity and control over the final VACNF length. Thus understanding the factors that
determine the growth rate is essential not only from the fundamental science point of view,
but also from the point of view of practical applications. In conventional thermal CVD the
growth of carbon nanofibers/nanotubes occurs in three main steps [63]; (i) decomposition of
the carbonaceous gas molecules at the surface of the catalyst nanoparticle, (ii) diffusion of the
resultant carbon atoms through the catalyst nanoparticle from the nanoparticle/gas interface
towards the nanoparticle/nanofiber interface due to the concentration gradient, and (iii)
precipitation of carbon atoms at the nanoparticle/nanofiber interface. Each of these steps can
be a complex process by itself [93,94] and the whole picture is not completely understood,
mainly due to the difficulty of conducting imaging and surface analysis in situ during the
growth. In thermal CVD, the carbon feedstock is the molecules of the carbonaceous gas used
in the growth process. The growth rate is determined by all of the three steps, but under
typical growth conditions appears to be diffusion limited as suggested by the equality of the
activation energies for the nanofiber growth rate and for the diffusion of carbon atoms
through the catalyst [95].
Chuang et al. [86] prepared carbon nanofibers and carbon nanotubes using CH4 as a
precursor material of carbon on Ni/Si and Ni/Ti/Si substrates at 640 C and 700 oC by thermal
chemical vapor deposition method. They have explained the growth of carbon nanofibers
(CNF) and carbon nanotubes (CNT) on Ni/Si substrate through tip growth mechanism, and
the growth mechanism of carbon nanotubes on Ni/Ti/Si substrate through root growth
mechanism [87]. Figure 2.3(a) schematically shows the solid amorphous carbon nanofiber
grown on Si substrate at 640 oC under the catalysis of Ni particle. During the heating of Ni/Si
substrate from room temperature to 640 oC, the Ni film on Si surface would agglomerate into
Ni nanoparticles, which still are in solid state. In the growth chamber the CH4 molecules are
adsorbed on the upper surface of Ni particles and decomposed into C and H atoms. These C
atoms are absorbed into Ni particles and diffuse to lower part through inside of Ni particles.
Due to the difference of both temperature and concentration of carbons between upper and
lower parts of Ni particles and the weak adhesion force between Ni particle and Si surface,
once the concentration of C atoms exceeds the saturation solubility of Ni, the C atoms would
precipitate on the lower surface of Ni particles and form carbon nanofiber. Because of the
roughness of Ni particle surface, the precipitated carbons could not form graphene planes on
the lower surface and hence only amorphous solid carbon nanofibers are obtained. Figure
2.3(b) schematically displays the hollow carbon nanofiber, i.e. carbon nanotube, is grown on
Si substrate at 700 oC under the catalysis of Ni particle. When the growth temperature is
raised from 640 - 700 oC, the Ni nanoparticles become fluid-like due to the lower melting
temperature of Ni–C alloy and the effects of size and interfacial stress between Ni particle
and carbon tube [96]. The surface of fluid-like Ni nanoparticle is much smoother than solid
Ni nanoparticle, and the diffusion of C atoms on fluid-like Ni nanoparticle surface is much
higher than on the surface of solid Ni nanoparticle. The C atoms are much easy to precipitate
on the surface of middle part, not the lower part, of Ni nanoparticle, and form the parallel
grapheme planes. Therefore, the carbon nanotubes are grown through the tip growth
mechanism. Figure 2.3(c) schematically shows the hollow carbon nanofiber, i.e. carbon
nanotube, is grown on Ti/Si substrate at 700 oC under the catalysis of Ni particle. Due to the
strong adhesion force between Ni nanoparticle and Ti film surface, the growth force cannot
push Ni nanoparticle up. Therefore, the carbon nanotubes are grown through the root growth
mechanism. When the temperature of Ni/Ti/Si substrate increased, a part of Ni diffuse
through Ti layer and into Si substrate to form NiSi and the rest of Ni would agglomerate to
form Ni particles [97]. The C atoms from the decomposition of CH4 under the catalysis of Ni
particles diffuse into Ni particles and Ti layer. When ternary alloy of Ni-Ti-C was formed at
the lower part of Ni particle due to the interdiffusion of Ni, Ti and C, the ternary alloy layer at
the bottom of Ni particle would melt due to the lower melting point of the Ni-Ti-C ternary
alloy [98]. Although the eutectic temperature of Ni-C is 1327 °C, there are many reports
[99,100] indicating that the Ni catalyst particle had melted or was behaved fluid like during
CNTs growth in the temperature of 600-900 °C due to the size effect of catalyst at nanometer
level and the interfacial effect between catalyst and carbon. The eutectic temperature of Ni-
Ti-C is 1265-1295 °C, lower than 1327 °C, hence the Ni-Ti-C alloy layer at the bottom of Ni
particle can melt due to the same reasons for Ni-C. Therefore Ni particles could sink into and
firmly adhere to the Ti layer. When C atoms in Ni particles exceed the saturation solubility,
they would form CNTs and grow up under the catalysis of Ni particles by root growth
mechanism. Once Ni particles were surrounded by CNTs, the possibility for CH4 to touch Ni
particles and decompose into C and H would decrease largely. For making CNT growth able
to continue, the C atoms supplied from Ti layer. Figure 2.4 schematically shows the layer
structure of substrates before and after CNTs growth. The three-layered Ni/Ti/Si substrate
becomes four layers structure after CNTs growth. From the AES depth profile measurement
of each element and the layer structure of sample they confirmed that the Ti interlayer
couldn't prevent the outward diffusion of Si and the inward diffusion of Ni, C and Ti;
however it can supply C atoms to continue the CNTs growth at later stage in CNT growth
process.
Figure 2.3. Schematic diagrams of one dimensional carbon growth mechanism for solid amorphous
carbon nanofiber grown through tip mechanism (a), carbon nanotube grown through tip mechanism (b),
and carbon nanotube grown through root mechanism (c) [ From Ref. 86].
Figure 2.4. Schematic diagrams for substrate (a) before CNTs growth and (b) mechanism for CNT
growth [From Ref. 87].
Plasma enhanced chemical vapor deposition technique has achieved lower growth
temperatures compared to other methods and vertical alignment of the nanotubes, which
facilitates the CNT based device fabrication with current silicon technologies. It has been
suggested [88] that in PECVD only VACNFs grown from the tip are aligned specifically due
to the presence of the plasma electric field in the growth process, whereas VACNFs grown
from the base are aligned mainly due to the crowding effect. Consequently, in the case of the
base-type growth, deterministic synthesis of isolated VACNFs is expected to be rather
difficult. Vertically aligned carbon nanofibers (VACNFs) were synthesized by direct-current
plasma enhanced chemical vapor deposition using acetylene and ammonia as the gas source
as shown in Figure 2.5 [101]. Generally in the case of PECVD growth the activation energies
were found to be different [71] and the growth rate was suggested to be limited by the supply
of carbon from the gas phase.
Huang et al. showed that the VACNF growth rate depends quite strongly upon the gas
mixture and plasma power used in the PECVD process [102]. This was attributed to changes
in the chemical composition of the excited gas species. These species include (i) simple
radicals created in the plasma as a result of direct dissociation of C2H2 and NH3 molecules
and (ii) larger radicals that form due to collision and consequent attachment of radicals to
each other as they move towards the substrate. It was suggested that changing the gas mixture
or plasma power changes the chemical distribution of the excited species. Since different
species are expected to have different decomposition rates at the catalyst surface, the
nanofiber growth rate also changes. The fact that reduction of the C2H2 content resulted in a
several-fold increase of the growth rate as well as a dramatic increase of the nitrogen content
within the nanofibers strongly suggests that the growth occurs mainly due to the species
created in the plasma, not due to unexcited C2H2 molecules.
Figure 2.5. SEM image of an array of VACNFs grown by PECVD for 5 min and of randomly oriented
CNFs/CNTs produced after the plasma was turned off and the growth run continued via purely thermal
CVD [from Ref.101].
Figure 2.5 shows the SEM image of a sample for which VACNFs were first grown using
a plasma, then the plasma was turned off, the NH3 flow was stopped, and C2H2 was flowed
over the sample that was kept at the same high temperature as during the PECVD part of the
growth (~700 oC). During the purely thermal phase, the VACNFs did not seem to increase in
length, but very long thin non-aligned CNFs/CNTs that were not present during the PECVD
part of the growth were produced. The non-aligned CNFs seemed to originate from the bases
of VACNF, where perhaps some of the catalyst was still left. This indicates that the catalytic
activities are quite different for the base- and tip-type growth modes. While the base-type
catalytic growth can produce CNFs in the purely thermal process, the tip-type growth,
requires the presence of radicals. The inability to re-grow VACNFs using only thermal CVD
confirms the idea of the feedstock for the VACNF growth consisting mainly of radicals and
not C2H2 molecules. There is another reason that a carbon shell may form around the catalyst
nanoparticles sitting at the tips of VACNFs after the plasma is turned off, which prevents
decomposition of carbonaceous species at the catalyst surface and consequently the VACNF
growth. Since some applications may require very long VACNFs, it is highly desirable to
develop controllable ways to further increase the growth rate. One way to relax the limit for
the VACNF growth rate, imposed by the process of radical diffusion towards the substrate, is
to change the radical transport mechanism from diffusive to forced flow by applying a
pressure gradient perpendicular to the substrate surface to force more radicals to impinge on
the surface [103]. The radical flux in this case will be given by F = Cv, where C is the radical
concentration and v is an average velocity towards the substrate. Thus, by increasing the gas
flow and therefore the radical velocity one can expect to achieve substantial increase in the
VACNF growth rate. The high gas flow during the growth can be produced highly conical
structures even for dense forests of VACNFs. Isolated VACNFs tend to assume conical shape
during the growth due to reactive species emerging for the discharge and attaching to the
sidewalls of the nanofibers [104]. In contrast, dense VACNF forests nominally consist of
essentially cylindrical nanofibers due to the shielding of the sidewalls by neighboring
VACNFs [79]. However, if the velocity of the incoming radicals is high enough, they will be
able to penetrate the inter-fiber space for even densely spaced VACNFs and precipitate at the
sidewalls thus forming conical structures [101]. Also the degree of conicity can be modified
by changing the gas mixture. Merkulov et al. [88] proposed the following model to explain
the vertical alignment of PECVD grown CNFs. The growth of CNFs occurs via
decomposition of the carbonaceous gas molecules at the catalyst particle surface or in the
glow discharge, diffusion of the carbon atoms through the particle, and subsequent
precipitation at the particle/fiber interface [63]. The axis of a CNF growing perpendicular to
the substrate coincides with the direction of the applied electrostatic force, resulting in a
uniform tensile stress across the entire nanofiber/catalyst particle interface, as shown in
Figure 2.6(a) and (d) Consequently, carbon uniformly precipitates across the interface and the
fiber continues to grow vertically (perpendicular to the substrate). However, if there were a
spatial fluctuation in the C precipitation at the interface, CNF growth would deviate from
vertical alignment, as shown in Figure 2.6(c) and (b). In the case of nanofibers growing from
the tip (catalyst particle at the tip), the electrostatic force produces a compressive stress at the
particle/nanofiber interface where the greater rate of growth is seen [Figure 2.6(c)]. Likewise,
a tensile stress is applied to the particle/nanofiber interface where the lesser rate of growth is
seen. These opposing stresses favor subsequent C precipitation at the interface experiencing
tensile stress and the lesser rate of growth. The net result is stable, negative feedback that acts
to equalize the growth rate around the entire periphery of the particle/nanofiber interface, and
vertically aligned CNFs are grown. The presence of the preferred direction of C precipitation
can be caused by stress-induced diffusion [105] due to the stress gradient in the catalyst
particle and possibly by the variation in the stress-dependent sticking of diffusing C atoms to
the C side of the Ni-C interface. Since the nanofiber base is attached to the substrate, the
stress created at the particle/ nanofiber interface with the greater growth rate is tensile [Figure
2.6(d)] and acts to continue the increased growth rate, thus causing the CNF to bend even
further. The inherent instability of positive feedback control systems leads to the wildly
varying CNF orientation.
Ngo et al. synthesized vertically aligned carbon nanofibers (VACNFs) using palladium as
a catalyst by plasma enhanced chemical vapor deposition (PECVD) [106]. Figure 2.7 shows
the TEM micrographs of vertically aligned carbon nanofiber grown on thick Pd catalyst. They
observed that the thick Pd films lead to a variety of growth morphologies including a hybrid
tip growth phenomenon, as well as small cone angles that are imparted by the
elongation/wetting of the inner cavity of the CNFs by the Pd catalyst.
Huang et al. reported growth of core/shell carbon nanofibers and nanotubes using metal
sulfide (FeS, CoS and NiS) as a catalyst by arc discharge technique [107]. Figure 2.8 shows
the schematic growth model of the core–shell carbon nanofibers and large-cavity carbon
nanotubes. The catalyst seed of metal sulfide come from melted metal sulfides or from the
combination reaction of evaporated metal and sulfur ions dissociated from metal sulfides. The
linkage of sulfur with metal and carbon, as well as the low-temperature environment prevents
the core materials from extruding out of the carbon nanofibers during growth. If the core/shell
carbon nanofibers are not exposed to high temperature during and after their growth, the
stuffed sulfides will remain in the carbon shell and contract and separate upon cooling.
Fig. 2.6 Alignment mechanism of carbon nanofibers. If a CNF grows vertically (along the electric-field
lines), electrostatic force F creates a uniform tensile stress across the entire catalyst particle/nanofiber
interface, regardless of whether the particle is located at the tip (a) or at the base (b). If during the
growth the CNF starts to bend due to spatial fluctuations in carbon precipitation at the
particle/nanofiber interface, nonuniform stresses are created at the particle/nanofiber interface. For the
nanoparticles at the tip (c) and at the base (d) the stresses are distributed in the opposite way, which
leads to the nanofiber alignment in the first (c) but not in the second (d) case. White ellipses indicate the
interface regions where the stresses occur [From Ref. 88].
When the temperature is high (> 1600 OC), such as in areas where the arc plasma reaches,
the metal sulfide core materials released from the carbon shells and subsequently the carbon
nanofibers will be annealed to become carbon nanotubes with large cavities, the main
component of the deposit on the bottom of the bowl-like cathode. The intensive gas flow
inside the cathode bowl and the gravity of the filled carbon nanofibers could be the force to
bring the nanofibers into the plasma environment for the annealing process. On the other
hand, in the plasma region, the catalyst seeds of metal sulfides could also lead directly to the
growth of large cavity carbon nanotubes. The growth model is similar to that of metal
catalyzed growth of carbon nanotubes and core/shell carbon nanofibers are no longer the
intermediates of the carbon nanotubes.
Figure 2.7. TEM micrographs of vertically aligned carbon nanofiber [from ref. 106].
Figure 2.8. Schematic growth mechanism proposed for the formation of the core/shell carbon
nanofibers and carbon nanotubes [From ref 107].
Carbon nanofibers have been synthesized by the thermal decomposition of acetylene with
a copper nanocatalyst derived from cupric nitrate trihydrate at a low temperature of 260 ◦C
[108]. Figure 2.9 shows the typical TEM image of as-prepared regularly helical nanofibers
with a symmetric growth mode. The copper nanoparticles changed from initial irregular
shapes to regular shapes during the growth of nanofibers.
Figure 2.9. TEM image of typical helical carbon nanofibers [From ref. 108].
Figure 2.10 shows the schematic diagram of growth mechanism of helical nanofibers.
The mechanism of morphological changes of copper catalysts and the growth of carbon
nanofibers are proposed in five steps: (1) the dehydration of cupric nitrate trihydrate, (2) the
decomposition of cupric nitrate into Cu oxide, (3) the reduction of Cu oxide, (4) the formation
of Cu nanoparticle before the growth of carbon nanofibers, and (5) the reaction with acetylene
for the growth of carbon nanofibers.
Figure 2.10. The schematic diagram of growth mechanism of helical carbon nanofibers [From ref. 108].
Hansen et al. reported that copper nanocrystals would undergo dynamic reversible shape
changes in response to changes in the gaseous environment, and the shape changes were
caused both by adsorbate-induced changes in surface energies and by changes in the
interfacial energy [109]. The gas adsorption on the surface of Cu nanoparticles and the
surface energy of different crystallographic planes of a single crystal were the main driving
force for the gas-induced surface reconstruction and reshaping of the Cu nanoparticles. In a
small metal particle, surface energies associated with different crystallographic planes are
usually different. The catalyst particles undergo surface reconstruction to form geometrical
shapes, which were able to promote the formation of carbon nanofibers with certain growth
conditions of catalysts, gas, and temperature [110,111]. The shape changes of copper
nanoparticles were induced by the adsorption of gases on the surfaces of particles. During the
reaction, the active sites of the copper nanoparticles were changed from one place to another
by following the surface reconstruction. The copper nanoparticle size has a considerable
effect on the morphology of carbon nanofibers. The helical carbon nanofibers with a
symmetric growth mode were grown on copper catalyst nanoparticles with a grain size less
than 50 nm. When the catalyst particle size was around 50-200 nm, straight carbon nanofibers
were obtained dominantly. It is reasonable to assume that it is possible to control the diameter
of carbon nanofibers by controlling the size of the catalyst particle.
A novel method for the direct growth of a single carbon nanofiber (CNF) onto the tip of a
commercially available scanning probe microscope (SPM) using Ar+ ion irradiation was
reported by Tanemura et al. [112]. This method was proposed on the basis of the
experimental fact that the Ar+ ion bombardment of carbon coated substrates induced the
formation of conical protrusions that possessed a single CNF at their tip. Commercially
available Si SPM tips were coated with carbon and then were Ar+ ion bombarded at room
temperature and at 200 oC. Figure 2.11(a) and (b) shows SEM image of a commercially
available Si SPM chip of the tip region before and after sputtering respectively. The CNF thus
grown was ~30 nm in diameter and 1.5 m in length. The length was controlled between 0.5
and 1.5 m by varying the sputtering duration.
Figure 2.11. SEM image of Si SPM chip at tip region (a) before and (b) after sputtering [From ref. 112].
The formation of ion-induced CNFs explained in terms of erosion and/or growth

processes during sputtering. In a case where CNFs are formed by the erosion process alone,
so-called ‗‗seed‘‘ materials, which differ from the surface-constituent materials, are necessary
for the CNF formation. These are known as ―l eft-standing‖ protrusions [113]. The seed
materials act as a protection against sputtering during the continuous erosion of the
surrounding surface, thus yielding the protrusions tipped with the seed materials [Figure
2.12(b)]. The protrusions thus formed must be linear in shape and possess no conical base.
TEM observation confirmed that there was no seed material on the CNF top and possessing
CNF-tipped cone structure, which disagree with those of projections formed by the erosion
process alone. This clearly suggests that the diffusion process plays a dominant role in the
formation of CNFs. Based on the TEM observations, they proposed the following growth
mechanism of ion-induced CNFs: (i) Formation of conical protrusions triggered by surface
defects such as grain boundaries and small amounts of impurities, (ii) deposition of carbon
atoms sputter-ejected from the surface onto the sidewall of the conical protrusions, and (iii)
surface diffusion of the deposited carbon atoms toward the tips during sputtering. Since the
diffusion is the thermal process, sputtering at elevated temperatures must enhance the
diffusion of deposited carbon atoms, thus yielding the longer CNF on the tip of the conical
protrusions. Van Vechten et al. demonstrated that carbon atoms readily migrated as far as ~20
mm on the surface during sputtering [114]. In addition to the thermal diffusion, the ion-
bombardment-enhanced diffusion, which is widely known to occur during sputtering, [115] is
likely responsible for the CNF growth.
One of the most successful approaches to obtain oriented arrays of nanotubes uses a nano
channel alumina template for catalyst patterning [116]. First, aluminum is anodized on a
substrate such as Si or quartz, which provides ordered vertical pores. Anodizing conditions
are varied to tailor the pore diameter, height and spacing between pores. This is followed by
electrochemical deposition of a cobalt catalyst at the bottom of the pores. The use of a
template not only provides uniformity but also vertically oriented nanotubes.
Figure 2.12. Schematic representation of the possible formation mechanism of surface projections. (a)
left-standing model based on ion etching process alone and (b) a growth model based on the diffusion
processes. [From ref. 112].
BASIC THEORY OF ELECTRON FIELD EMISSION

Field emission (FE) is based on the physical phenomenon of quantum tunneling, in which
electrons are injected from the surface of materials into vacuum under the influence of a
strong external electric field [117]. The potential barrier is rectangular when no electric field
is present, and becomes triangular when a negative potential is applied to the solid. The slope
of the latter depends on the amplitude of the local electric field E just above the surface. This
local electric field is drastically enhanced if the structure of the emitter is very sharp and
protruding (high aspect ratio) as in the case of a CNT. Compared to thermionic electron
emission and photo electron emission where electrons have sufficient energy to overcome the
potential barrier (work function ), field emitted electrons tunnel into the vacuum because of
a strongly deformed barrier under an electrical field (shown in Figure 3.1).
Figure 3.1. Potential-energy diagram for electrons at a metal surface under an applied electric field,
where a strongly deformed potential results from the combination of the applied field and the image
charges induced by the emitted electrons.
In the presence of high electric field, flat thin films of some materials emit electrons at
macroscopic fields of about 1 to 10 V/ m, although cold field emission of electrons normally
occurs only at fields of about 2 V/ m or above.
I This occurs because the thin film is an electrically nanostructured heterogeneous

(ENH) material.
II Internal nanostructure creates geometrical field enhancement at or near the film or
vacuum interface, so local fields are much greater than macroscopic fields.
III Electron emission at low macroscopic field strengths is a property of all ENH
materials under appropriate conditions.
IV Field enhancement is the primary effect, but there also be the secondary effects that
contribute to facilitating emission at low fields.
A simple mathematical expression for field emission can be obtained from the
Heisenberg uncertainty principle:
∆p. ∆x ħ/2 (3.1)
where ∆p is the uncertainty of the electron momentum and ∆x is the corresponding

uncertainty in position. Considering electrons near the Fermi level, the barrier height is the
work function φ, so the uncertainty of the electron momentum ∆p = (2m φ) 1/2 and ∆x can be
obtained from equation. (3.1)
∆x =  / 2 (2m φ) 1/2 (3.2)
Thus when ∆x is of the order of the barrier width, the probability that electrons would
penetrate the barrier is high. The barrier width is
x=φ/E (3.3)
where E is the electrical field. Combining equations 3.2 and 3.3, we can obtain an estimate of
the electrical field required for electron field emission:
3
1 2
E 2(2m /  2 ) 2
(3.4)
e
A more detailed mathematical description has been done with the Wentzel- Kramers-
Brillouin (WKB) method. The emission current density (J) can be obtained as a function of
the electrical field, E, and work function, φ, which is described by Fowler-Nordheim (F-N)
equation [118]:
3
AE 2 2
(3.5)
J exp B ( y) A / cm 2
.t 2 ( y ) E
where A and B are constants with values of 1.54 x 10-6 and 6.87 x 107, respectively, and
1/ 2
( y) 0.95 y 2 , where y 3.8 x10 4 E , Both t 2 ( y ) and ( y ) are contributions
arising from the image potential, which is due to positive image charges induced by the
emitted electrons at the surface. Image charges can cause a further lowering of the barrier
height by a factor of e 2 (Figure 3.1). Under typical conditions, t 2 ( y ) and ( y ) are close
4x
to unity and normally omitted in practice. The standard physical assumptions of F-N theory
are that the metal: (i) has a free-electron band structure; (ii) has electrons that are in
thermodynamic equilibrium and obey Fermi-Dirac statistics; (iii) is at zero temperature; (iv)
has a smooth flat surface; and (v) has a local work function that is uniform across the emitting
surface and is independent of external field. It is also assumed that: (vi) there is a uniform
electric field above the emitting surface; (vii) the exchange-and-correlation interaction
between the emitted electron and the surface can be represented by a classical image
potential; and (viii) barrier penetration coefficients may be evaluated using the JWKB
approximation. In addition, the F-N equation is based on the assumption of a flat surface and
only valid at 0 K, therefore, a modified F-N equation has to be used for an irregular surface
and at different temperatures. In order to initiate field emission, an extremely large electrical
field has to be employed, which is difficult to achieve from flat surface. However, with a tip
structure, a high local electrical field can be obtained around high curvature regions. For
instance, an electrical field E at a surface of a sphere, with a radius r and a potential V, is E =
V/r. When r becomes smaller, E will become larger. Therefore, to account for the geometrical
effects on the local electric field, a field enhancement factor β is introduced in the F-N
equation as follows:
3
2
6 ( E) 2
J 1.54 x10 exp 6.87 x10 7 A / cm 2 (3.6)
E
An experimental F-N plot is modeled by the tangent, which can be written in the form
[119-121]
I S
ln( ) a (3.7)
V2 V
where a is a constant and
3
6.83 x10 7 2
d (3.8)
S
A linear relationship can be obtained when plotting ln(I/V2) vs I / V, and if we know the
work function (φ) of the material, specially for carbon nanotubes, we will assume a work
function of 5 eV, [122,123] the inter-electrode distance (d), and the slope (S) of the F-N plot,
the field enhancement factor β can be obtained.
In geometrical configurations resembling a parallel plate capacitor, the macroscopic field
EM is defined by:
EM = V/d, (3.9)
where V is the voltage applied across a gap of thickness d. The local field E is the field, close
to the emitting surface (within 1-2 nm of the surface atoms), that determines the barrier
through which field emitted electrons tunnel. The field E is some times called the barrier
field. E is typically a few V/nm, and is often significantly higher than EM. Their ratio defines
a field enhancement factor
= E / EM (3.10)
Considering the simple physical models of a ‗floating sphere at the emitter plane
potential‘ and a ‗hemisphere on a post‘ the corresponding mathematical expressions have the
form:
=m+h/r (3.11)
where r is the radius of the sphere or the hemisphere, h is its height above the emitter plane,
and m is a constant generally taken as 0, 2 or 3 [124].
FIELD EMISSION FROM CARBON BASED MATERIALS

Physically the length of CNT is several mm and diameters down to 10 Å, so the
nanotubes exhibit a very high geometric aspect ratio (h/r), and has been shown that the
applied electric field is concentrated precisely at the nanotube tips [125,126], which results in
a large field enhancement factor , typically ~10,000. In addition to the geometric field
enhancement factor shown in equation (3.6), the effect of adsorbates i.e., forming a tunneling
state at the surface [127] the tube cap and tip structures such as open or close [128,129] and
the surface morphologies and contaminations i.e. catalyst particles, amorphous carbon [130]
have been proposed as affecting their extraordinary field emission behavior. Carbon
nanotubes have the right combination of properties such as nanometer size diameter,
structural integrity, high electrical conductivity, and chemical stability that make good
electron emitters. Electron field emission from carbon nanotubes was first demonstrated by
Rinzler et al. in 1995 [131], and has since been studied intensively on various carbon
nanotube materials. Field emission properties of different types of carbon nanotubes have
been reported, including individual nanotubes [131-133], MWCNTs embedded in epoxy
matrices [134], MWCNT films [135], SWCNTs [136], aligned MWCNT films [65,137] and
hollow carbon nanotubes [132]. Random and aligned MWCNTs were found to have threshold
fields slightly larger than that of the SWCNT films and are typically in the range of 3-5 V/µm
for a 10 mA/cm2 current density [65]. These values for the threshold field are all significantly
better than those from conventional field emitters such as the Mo and Si tips which have a
threshold electric field of 50-100 V/µm. It is interesting to note that the aligned MWCNT
films do not perform better than the random films. This is due to the electrical screening
effect arising from closely packed nanotubes [138].
The case of emission from the large variety of carbon-based materials suggests that the
NEA is not a prerequisite, and more general emission models are desired for these systems.
Many models have been proposed to discuss the origin of electron emission from ta-C films
and other amorphous carbon (a-C) films. Robertson suggested that field emission at a low
electric field is due to the low electron affinity of ta-C films [139]. Other models such as
space-charge-induced band bending [140], surface dipole-controlled emission [139], field
enhancement due to the film microstructure [141] and conductive paths caused by localized
sp2 sites were also proposed [142]. Thermal annealing is an effective method to alter structure
of DLC films. Ilie et al. [143] and Carey et al. [144] proposed that the presence of sp2 clusters
within the insulating sp3 matrix could give rise to field enhancement in amorphous carbon (a-
C) films containing large defect densities (>1019 cm−3). It was proposed that the presence of
such dielectric inhomogeneity is responsible for field enhancement in these films. Since sp2
clusters will have different dielectric constants, the application of the external field will result
in local field enhancements around the clusters and will aid in the emission of electrons.
Groning et al. [142] explained the emission mechanism from DLC films in a way that, like a
freestanding conductive tip in the vacuum, sp2 bonded carbon clusters are assumed to form a
conductive channel in an insulating matrix, which leads to local field enhancement and hence
to an enhanced electron emission. Since the sp2 clusters are located at or near the Fermi level
and high concentration sp2 carbon clusters in the films play a more important role in
determining the electron emission property of the films. The emission can depend on various
parameters such as negative electron affinity [145], band gap [146], surface termination
[145,147], depletion layers [148] and film thickness [149]. In these cases, the emission can be
interpreted in terms of homogeneous films and a well-defined band structure [150]. In
amorphous carbon, the sp3 content controls the band gap and electron affinity. Nanostructured
carbon, nanocrystalline diamond, and carbon nanotubes are the types of carbon that emit at
lowest applied field. In microcrystalline diamond, emission is found to occur from grain
boundaries [151,152], that is, nm-scale sp2-bonded regions of positive electron affinity.
Similarly, emission from carbon nanotubes [153] occurs from 1 nm curved regions.
CNFs films were synthesized by plasma enhanced chemical vapor deposition about 100
nm in diameter and about 10 m in length using P doped n-type Si (100) wafers and indium
tin oxide (ITO) coated glasses as substrates [154]. Figure 4.1(a) shows the SEM image of the
CNF film on Si substrate and (b) shows the TEM image of a tip of a CNF where as (c) shows
a schematic illustration of the CNF structure. The CNFs ranged 50-100 nm in diameter and
over 10 m in length, which were randomly oriented to the substrate showed electron field
emission characteristic (as shown in Figure 4.2(d)). Since the nanofibers were grown to
random orientation, electrons can be emitted with any directions from the protrusions on the
fiber.
The threshold electric field (Eth) was estimated 2.4 V/ m, which is comparable with the
threshold field of several carbon nanostructures such as, Eth of single wall carbon nanotube
(SWCNT) by arc discharge about 2.2 V/ m, multi wall carbon nanotube (MWCNT) by
microwave plasma CVD about 1.8 V/ m and nanostructured carbon film about 3.0 V/ m
respectively [155-157]. The reason of the excellent field emission characteristics is due to the
CNF film has many protrusions which are 10 nm in width and 30 nm in length, as shown in
Figure 4.1(b). Since the nanofibers were grown to random orientation, electrons can be
emitted with any directions from the protrusions on the fiber. Aligned carbon nanofibers and
hollow carbon nanofibers were grown by micro wave ECR-CVD method using methane and
argon mixture gas at a temperature of 550 oC showed good electron field emission (Figure
4.2) [158]. The aligned carbon nanofibers give a high current density 7.25 mA/cm2 at 12.5
V/ m in comparison with the value 0.69 mA/cm2 at 12.5 V/ m of the hollow carbon
nanofibers.
Figure 4.1.(a) SEM image of the CNF film on Si substrate, (b) TEM image of a tip of a CNF, (c)
schematic illustration of the CNF structure and (d) J-E curve and in inset the corresponding F-N plot
[From ref. 154].
Figure 4.2. I-V graph of the carbon nanofiber (a) and the hollow carbon nanofiber (b) [From ref. 158].
Effect of catalysts on the field emission property was studied by Kamada et al. [78].
Carbon nanofiber films on Pd-Se, Fe-Ni, and Ni-Cu alloy catalysts have been synthesized at
low temperatures by a thermal CVD technique. Figure 4.3(a) shows the field electron
emission characteristics of the CNF films grown at 600 oC. The threshold electric fields of the
CNFPd-Se, CNFNi-Cu, and CNFFe-Ni films are estimated to be 1.1, 2.8, and 3.8 V/ m,
respectively. The CNFs grown using Pd-Se catalyst were found to have more defective
structure than that obtained with the other catalysts, and exhibited best field emission
property. It is likely that defects play a role as electron emission sites. Figure 4.3(b) shows the
threshold electric field obtained for the CNFPd-Se, CNFFe-Ni, and CNFNi-Cu films as a function
of growth temperature. The threshold electric field was not strongly dependent on the catalyst
type and decreased with growth temperature. Ilie et al. [159] reported that the surface
electronic properties introduced by defects could provide a local field enhancement to
facilitate the field emission. From this, it is suggested that the excellent field emission
property obtained for CNFPd-Se originates from numerous defects in the body of the CNFs. In
this sense, good crystallinity of the CNFs is not required to obtain good field emission
characteristics.
Carbon nanofibers (CNFs) were grown on a Ni-P alloy catalyst deposited on a silicon
substrate via MWCVD technique with methane gas [82]. The CNFs grown on the Ni-P alloy
catalyst showed random orientation and it composed of parallel graphite planes with defects
tilted from their axis. Figure 4.4 shows the electron emission current density versus electric
field (J-E) curves of CNFs with different thickness of the catalyst.
Figure 4.3. (a) Emission current density as a function of applied electric field and (b) threshold electric
field for the CNFPd-Se, CNFFe-Ni, and CNFNi-Cu films as a function of growth temperature [From
ref. 78].
For CNF grown on catalyst with thickness 20 nm, the turn-on field was approximately
0.11 V/ m with an emission current density of 10 mA/cm2 and the threshold field was 3.1
V/ m with an emission current density of 10 mA/cm2. CNF grown on catalyst thickness 30
and 40 nm, have almost the same turn-on field approximately 0.22 V/ m, but the threshold
field is 3.4 and 4.1 V/ m, respectively.
Figure 4.4. J-E curves for CNFs deposited with different thickness of catalyst and in Inset
corresponding F-N plot [From ref 82].
The excellent field emission properties of Ni-P alloy catalyzed-CNFs may be attributed to
the random and defects of CNFs. Davydov et al. [160] have also pointed out that perfectly
aligned CNTs were less efficient field emitters and had lower field enhancement than chaotic
CNTs. Some reports relating defect densities to field emission properties have also been
proposed [161,162]. The enhanced emission may originate from the defect-induced energy
bands that are formed within the band gap of graphite. The energy barrier that the electrons
must tunnel through to be emitted is reduced, so the electrons residing at these defect levels
can be emitted directly into vacuum from these bands or be transported to the surface states
for emission [163]. Obraztsov et al. [164] have also found that the field emission properties
were improved by increasing the density of structural defects. Figure 4.4 also indicates that
the field emission properties of CNFs with small diameter are better than those of the CNFs
with large diameter. A clear fluctuation of the I-V curve at higher voltages for CNFs is seen in
Figure 4.4, indicating some emission sites are damaged or destroyed. It is reasonable to
suggest that the CNFs with small diameter are more easily damaged by ion bombardment
than the CNFs with large diameter [165].
The CNF is synthesized on the iron-evaporated Si substrate by microwave plasma
chemical vapor deposition and nitrogen (N2) plasma treatment is carried out to modify the
CNF surface [166]. A reduction in the turn-on electric field is achieved by N2 plasma
treatment for the CNF and the stability of the electron emission current is also improved by
nitridation of the CNF surface. In Figure 4.5 the field emission characteristics are compared
between as grown and N2 plasma-treated samples. Potential barrier height is reduced by
nitridation of the CNF surface. The excellent field emission of carbon nanofibers and
nanotubes have stimulated their applications as electron sources in x-ray applications [137],
and field emitters in electron microscopes [132], with their environmental sensitivities of
electrical conduction.
Figure 4.5. Field emission characteristics (a) untreated and (b) N2 plasma treated CNFs [From ref. 166].
SYNTHESIS AND FIELD EMISSION PROPERTY OF DIFFERENT CARBON

NANOSTRUCTURE
Synthesis and Field Emission Property of Carbon Fibrous Films
Among the different techniques for the production of carbon nanotubes, plasma enhanced
chemical vapor deposition is a high yield and controllable method for the production carbon
nanotubes/nanofibers [167] with mass production. Thin film catalyst layers have been
successfully employed in the carbon nanofiber growth. The use of metal catalysts such as Ni,
Fe, Co, Pt and some of their alloys has been explored in an effort to control the size and
morphology of carbon nanostructures formed through the decomposition of hydrocarbons.
We have used Ni as a catalyst for the formation of carbon fibers, because carbon
nanostructure formed on Ni are more crystalline than those formed with other catalysts
[168,169]. The target used for sputtering was a Ni plate of thickness ~1 mm with a diameter
2.5 cm (with purity 99.99 %, Aldrich). The Ni target has been sputtered on Si substrates via
dc sputtering technique to produce thin film of Ni catalyst. The substrates were 10×10 mm2
cleaned Si (400) wafer. The Si substrates were etched in HF (~20%) for 5 minutes to remove
the surface oxide layer and finally cleaned in an ultrasonic cleaner. The sputtering was done
at a pressure 0.2 mbar sending argon as a sputtering gas with an inter-electrode distance 1.6
cm at room temperature for deposition time 5 minutes, which yielded a Ni film with a
thickness ~10 nm, as measured by quartz crystal thickness monitor. For sputtering, we
maintained high voltage 2.5 KV and corresponding current density was 19.5 mA cm-2. Table -
5.1 shows the deposition parameters for Ni catalyst thin films. After deposition of the
catalyzed film, the sample was immediately transferred in to the CVD chamber where
nanofibers growth has been performed.
Deposition chamber used for the synthesis of carbon fibers was made of stainless steel
(SS). The plasma was produced between two parallel plates SS electrodes as usual. The lower
plate was grounded and the upper plate was used as the cathode. The schematic diagram of
the DC-PECVD unit is shown in Figure 5.1. The deposition chamber was initially evacuated
by a standard rotary and a diffusion pump arrangement up to a base pressure of 10−6 mbar.
The substrate (Si) was placed on a molybdenum substrate holder, which could be directly
heated. When the chamber pressure attained 10−6 mbar, the Mo substrate holder was started to
heat by sending current through it. The substrate temperature could be varied by varying the
current through the Mo substrate holder, which was connected to the secondary of a step
down transformer. The temperature of the substrate was measured by a disappearing filament
type pyrometer (PYROPTO, IT65). Acetylene (C2H2) gas was used in PECVD process as a
precursor of carbon. Acetylene (C2H2) gas was allowed to flow maintaining the CVD
chamber pressure 50 mbar. Deposition was performed at 2.0 kV DC supply with
corresponding current density 25 mA cm-2 for 30 min. Substrate temperature was varied from
700 to 850oC for different set of experiment. The deposition parameters for the synthesis of
carbon fibrous thin films via PECVD have been shown in Table - 5.2.
Figure 5.1. Schematic diagram of the DC -PECVD unit.
Table 5.1. Deposition parameters used for dc-sputtered deposited Ni catalyst
Deposition parameters Corresponding values

Deposition time 5 min.
dc voltage 2.5 kV
Electrode distance 1.6 cm
Sputtering gas Argon (Ar)
Gas pressure mbar
Substrates used Si (400)
Substrate temperature 300 K
Table - 5.2. Deposition parameters used for the synthesis of carbon fibrous thin films

1. Deposition time 30 min.
2. dc voltage 2.0 kV
3. Electrode distance 1.4 cm
4. Precursor material Acetylene (C2H2)
Gas pressure 50 mbar
Substrates used Ni catalyzed Si (400)
Substrate temperature 700 - 850 0C
Figure 5.2 showed the SEM micrographs of the deposited films, which showed the
existence of carbon fibers in the films. The morphologies of the films have been changed with
the change of substrate temperature. At 700 oC substrate temperature, only particles have
been found but at 750 oC, some carbon nanofibers have also been grown. At 800 oC substrate
temperature, carbon fibers have been grown with length ~ 1000 nm and the corresponding
diameter ~ 400 nm. Finally at 850 oC substrate temperature, the best quality carbon fibers
have been grown with length ~ 2000 nm with corresponding diameter ~ 400 nm. It is clear
from these studies of substrate temperature variations that at lower substrate temperature only
particles are grown and at higher substrate temperature the morphology changes from
particles to nanotubes or fibers like structure i.e. quasi one dimensional growth takes place at
higher substrate temperature.
The electron field emission properties of the CNFs deposited on Si substrates have been
studied by our high vacuum (~10-7 mbar) field emission setup as shown in Figure 5.3. Field
emission measurements were carried out by using a diode configuration consisting of a
cathode (the film under test) and a stainless steel tip anode mounted in a liquid nitrogen
trapped rotary-diffusion vacuum chamber with appropriate chamber baking arrangement. The
measurements were performed at a base pressure of ~5 x 10-7 mbar and at different
temperature, which was controlled with a controller and measured with a thermocouple. The
tip-sample distance was continuously adjustable to a few hundred m by spherometric
arrangement with screw-pitch of 10 m. The anode-sample spacing was set at a particular
value by rotating the micrometer screw which served as an anode electrode. Field emission
current-voltage measurements were done with the help of an Agilent multimeter (model
3440-1A). Emission characteristics were registered and analyzed with the help of a personal
computer.
Figure 5.2. SEM image for different substrate temperature (a) 750 oC, oval shaped particles, (b) 800
oC, fiber like and (c) 850 oC; fibers [From ref. 168].
Figure 5.3. Schematic diagram of the field-emission apparatus [From ref. 170].
We have used the simplified F-N equation for field emission analysis. Figure 5.4(a)
shows the emission current (I) vs. macroscopic field (E) curves of carbon nanofibers thin film
deposited on Si substrate for anode-sample separation (d) of 60 m. The macroscopic field is
calculated from the external voltage applied (V), divided by the anode-sample spacing (d).
Theoretically, the emission current I is related to the macroscopic electric field E by
3
2
2 1 2 b vF
I A a tF ( E ) exp{ } (5.1)
E
where, is the local work-function, is the field enhancement factor, A is the effective
emission area, a is the first Fowler-Nordheim Constant (1.541434 x 10-6 A eV V-2), b is the
second F-N Constant (6.830890 x 109 eV-3/2 V m-1), and vF and tF are the values of the special
field emission elliptic functions [119] v and t, evaluated for a barrier height . In so-called
Fowler-Nordheim coordinates, this equation takes the form:
3
1
I (v F b 2
)
ln{ 2 } ln{ t F 2 A a 1 2
} (5.2)
E E
An experimental F-N plot is modeled by the tangent, which can be written in the form
[119-121]:
3
1
I 1 2 (s b 2
)
ln{ 2 } ln{ rA a } (5.3)
E E
where r and s are appropriate values of the intercept and slope correction factors, respectively.
Typically, s is of the order of unity, but r may be of order 100 or greater. Both r and s are
relatively slowly varying functions of 1/E, so a F-N plot (plotted as a function of 1/E) is
expected to be a good straight line. The F-N plot of our sample is shown in Figure 5.4(b). It
has been observed that the I-E curve in the present work is closely fitted with straight line.
This suggests that the electrons are emitted by cold field emission process. The turn-on field,
which we define as the macroscopic field needed to get an emission current I = 0.09 A,
(which corresponds to an estimated macroscopic current density, Jest = 14.5 A/cm2, where
Jest = I/A, A = anode-tip area) were lying in the range 2.57 to 9.71 V/ m for variation
substrate temperature films. This value is quite lower than that of nanocrystalline carbon 6.4
V/ m [171] and carbon nanofiber arrays (~3 V/ m) reported by Cao et al. [172].
According to the F-N plot (Figure 5.4(b)), the slope m (given by equation (5.4)) would
represent the combined effect of work function and enhancement of local electric field and is
given by,
3
2
m
b (5.4)
The effective work function E is related with the true work function through the
2
relation [173] E = / 3 . Using = 5 eV is the work function of CNF [174]; the field
enhancement factor was calculated from the slope of the F-N plot, lies in the range 8090 to
1945 and the corresponding effective work function C lies in the range 0.0124 to 0.0321 eV
for films deposited with different substrate temperature, which is comparable with CNT films
[173].
The plots of I-E graph for different electrode distance are shown in Figure 5.5(a) and (b)
the corresponding F-N plot. The turn-on field was found to vary in the range 6.87 to 2.87
V/ m for a variation of anode sample spacing 80 - 120 m for the carbon fibrous film
deposited at 850 0C. In the I-E graph (Figure 5.5(a)), we observed a parallel shift of curves
with respect to anode-sample separation (d) i.e., for a particular electric field the current
density increases with increasing the anode-sample separation. Zhou et al. [175] reported
similar type of observation for their -SiC nanorods. We suppose that this type of shift
observed in our sample may be due to the change in the effective emission area of the sample
for different anode-sample separation. The change of effective emission area with respect to d
may be related to the geometry of the anode.
Figure 5.4. Emission current vs. macroscopic field curves of carbon fibrous film deposited for different
substrate temperature (a) and corresponding. F-N plot (b) [From ref. 168].
In our experiment we have used a conical shape anode with tip diameter 1 mm, therefore
the lines of force immerging from the edge of the anode tip and terminating to the sample
surface are diverging in nature, whereas the lines of force immerging from the flat surface of
the tip are parallel in nature. Hence, the effective emission area of the sample increases with
increasing d as shown in Figure 5.6. Au et al. [176] performed field emission of silicon
nanowires using a spherical-shaped stainless steel probe with a tip diameter of 1 mm as an
anode. They also found a parallel shift in their I-V curve.
Figure 5.5. I-E curves carbon fibrous film for different anode-sample separation (a) and (b)
corresponding F-N plot (b) [From ref. 168].
Okano et al. [177] reported that their macroscopic current density for diamond films was
independent of the anode-sample separation. Their field emission apparatus consisted of a
parallel plate arrangement of the anode and sample, separated by spacers. So the electric lines
of force between the anode and the sample were parallel in nature, hence effective emission
area remained independent of the anode-sample spacing.
Figure 5.6. Schematic diagram for the dependence of effective emission area as an increasing function
of anode-sample separation.
Synthesis and Field Emission Property of Vertically Aligned CNFS
Vertically aligned carbon nanofibers (VACNFs) have been synthesized by direct current
PECVD technique and for the synthesis of VACNFs, Ni catalyst was deposited in thin film
form on Si substrates via RF magnetron sputtering technique. For sputtering, we maintained
RF power 200 watt and corresponding chamber pressure 0.1 mbar. We have synthesized Ni
thin film having different thickness varying from 10-20 nm, as measured by quartz crystal
thickness monitor (HindHivac, Digital thickness monitor, Model: DTM-101). The deposition
parameters for Ni catalyst have been shown in Table- 5.3. After deposition of the Ni film, the
substrates were immediately transferred into the CVD chamber where nanotube growth has
been performed. Acetylene (C2H2) gas was used in PECVD process as a precursor of carbon
and during deposition the chamber pressure was maintained at 30 mbar. Deposition was
performed at 2.2 kV DC with corresponding current density 21.5 mA cm-2 for 25 min.
Table – 5.3. Deposition parameters used for rf-magnetron sputtered deposited Ni

catalyst

RF-power 200 Watt
Electrode distance 3 cm
Sputtering gas Argon (Ar)
Gas pressure 0.2 mbar
Substrates used Si (400)
Substrate temperature 300 oK
Deposition time 4-10 min.
An atomic force microscope (AFM-NT-MDT, Solver Pro) was used to analyze the
surface topography of the grown CNFs films. Figure 5.7 shows the AFM images of the
carbon nanofibrous films deposited on Ni catalyst having different thicknesses. From the
figure it is clear that the diameter of the CNFs increase and length decreases with the increase
of Ni catalyst film thickness. The diameter and lengths of the CNFs deposited on thinner
catalyst (Ni film thickness 10 nm) are ~ 150 nm and 2.5 m whereas the diameter and length
of the CNFs deposited on thicker catalyst (Ni film thickness 20 nm) are ~ 250 nm and 1.0
m. The morphology of the catalyst film is known to play a critical role in CNF growth. So,
the thickness of the Ni catalyst film will affect the growth and the properties of CNFs. The
diameter of the CNFs decreased as the thickness of the catalyst film decreased.
Figure 5.7. AFM 3D pictures of VACNFs deposited on different thickness of Ni catalyst (a) for 10 nm,
(b) for 15 nm and (c) for 20 nm [From ref. 178].
Figure 5.8(a) shows the emission current density (J) vs. macroscopic field (E) curves for
the VACNF films deposited at a fixed anode-sample separation (d) of 120 m. Field emission
characteristics of the films were analyzed using the help of simplified Fowler-Nordheim (F-
N) theory [119-121]. The F-N plots of our sample are shown in Figure 5.8(b).
Figure 5.8. J-E graph of VACNFs for different aspect ratios (a) and corresponding F-N plot (b) [From
ref. 178].
It has been observed that all the J-E curves in the present work are satisfactorily fitted
with a straight line, which suggests that the electrons are emitted by field emission process.
The threshold field, which we define as the macroscopic field needed to get an emission
current density J = 10 A/cm2, were lying in the range 4.3 to 5.4 V/ m for VACNFs
deposited on Ni catalyst having different thicknesses. As our deposited CNFs have different
aspect ratios so the local field enhancement occurs and changes in the threshold field was
observed for different type of VACNFs. The field enhancement factor ( ) as well as emission
current density is strongly dependent on the aspect ratio of canon nanofibers (shown in Figure
5.9). Tsai et al. [82] showed that the CNFs with small diameter and many defects exhibited
excellent field emission properties than the CNFs with larger diameter.
Figure 5.9. The variation of threshold field and emission current density with aspect ratio carbon
nanofiber [From ref. 178].
Synthesis and Field Emission Property of Multiwalled Carbon Nanotubes
For the deposition of Ni catalyst in thin film form by dc sputtering process we have used
a Ni plate of thickness ~1 mm with a diameter 2.5 cm (with purity 99.99 %, Aldrich). The
substrates were 10×10 mm2 cleaned Si (400) wafer. The Si substrates were etched in HF
(~20%) for 5 minutes to remove the surface oxide layer and finally cleaned in an ultrasonic
cleaner. The sputtering was done at a pressure 0.1 mbar sending argon as a sputtering gas
with an inter-electrode distance 1.6 cm at room temperature for deposition time 5 minutes,
which yielded a Ni film with a thickness ~ 12 nm, as measured by quartz crystal thickness
monitor. For sputtering, we maintained high voltage 3.0 KV and corresponding current
density was 27.5 mA cm-2. After deposition of the catalyzed film, the sample was
immediately transferred to the CVD chamber where the synthesis of nanotubes was
performed. The deposition procedure was described in section 5.1. Table 5.4 shows the
deposition parameters for the synthesis of MWCNTs thin films via DC-PECVD technique.
Table 5.4. Deposition parameters used for the synthesis of MWCNTs thin films

1. Deposition time 30 min.
2. dc voltage 2.0 kV
3. Electrode distance 1.4 cm
4. Precursor material Acetylene (C2H2)
5. Gas pressure 30 mbar
6. Substrates used Ni catalyzed Si (400)
7. Substrate temperature 900 0K
Figure 5.10(a) shows the FESEM micrographs of the deposited films, which showed the
existence of carbon nanotubes in the films. The diameter of the carbon nanotubes are ~ 12 nm
and few micrometer in length. The transmission electron micrographs of multiwalled carbon
nanotubes have been shown in Figure 5.10(b). It could be observed that the carbon nanotubes
are multiwalled with diameter ~12 nm.
Figure 5.10. FESEM micrograph (a) and HRTEM lattice image of the MWCNT (b).
Figure 5.11(a) shows the emission current density (J) vs. macroscopic field (E) curves for
MWCNT films for different anode-sample separation (d). The F-N plot of our sample is
shown in Figure 5.11(b). The threshold fields were found to vary in the range 4.7 - 3.6 V/ m
for a variation of anode-sample separation in the range 80 - 150 m.
Figure 5.11. (a) J-E curves for the MWCNTs for different anode-sample separation (d) and (b)
corresponding F-N plots.
In the J-E graph (Figure 5.11(a)), we observed a parallel shift of curves with respect to
anode-sample separation (d) i.e., for a particular electric field the current density increases
with increasing the anode-sample separation. For example, at a field of 5 V/ m, the J values
were found to be 0.2 mA/cm2 (for d = 80 m), 1.2 mA/cm2 (for d = 110 m) and 3.7 mA/cm2
(for d = 150 m). This type of shift observed in our sample is due to the change in the
effective emission area of the sample for different anode-sample separation, which is
described in section 5.1.
EFFECT OF TEMPERATURE ON THE ELECTRON FIELD EMISSION

FROM VERTICALLY ALIGNED CARBON NANOFIBERS (VACNFS) AND
MWCNTS
The synthesis of vertically aligned carbon nanofiber (VACNF) thin films has been
described in previous section 5.2 and for synthesis of MWCNTs, Ni catalyst deposited in thin
film form on Si substrates via RF sputtering technique. The target used for sputtering was a
Ni plate of thickness ~1 mm with a diameter 2.5 cm (with purity 99.99 %, Aldrich). For
sputtering, we maintained RF power 180 watt and corresponding chamber pressure 0.2 mbar.
After deposition of the Ni film, the substrates were immediately transferred into the CVD
chamber where nanotube growth has been performed. Acetylene (C2H2) gas was used in
PECVD process as a precursor of carbon. C2H2 gas was allowed to flow maintaining the CVD
chamber pressure 40 mbar. Deposition was performed at 2.5 kV DC supply with
corresponding current density 27.5 mA cm-2 for 30 min at 700 oC substrate temperature. AFM
image (Figure 6.1(a)) of the carbon nanofibrous films shows that the vertically aligned CNFs
are having an average diameter ~ 250 nm and length 2.0 m. Figure 6.1(b) shows the FESEM
micrographs of the carbon nanotubes, which showed that the diameter of the carbon
nanotubes are ~ 20 nm and few micrometer in length. The transmission electron micrographs
of multiwalled carbon nanotubes have been shown in inset of Figure 6.1(b). It could be
observed that the carbon nanotubes are multiwalled with diameter ~ 20 nm.
Figure 6.1.(a) AFM 3D pictures of vertically aligned carbon nanofiber thin films, (b) FESEM
micrograph and in inset HRTEM micrograph of the MWCNT [From ref. 179].
For temperature dependent field emission total current density (J = JE + JT, where JE and
JT are the field current and thermionic current density respectively) given by simplified F–N
equation and Richardson equation as [180,181]:
J J E JT (6.1)
3
2
1 2 sb
J a ( E ) exp( ) ADT 2 e KT
(6.2)
E sin( )
where A is a constant about 120 A/(cmK)2, D is the average transmission coefficient of

emitter surface, T is the temperature in Kelvin, is the work function of CNF/CNT, k is the
Boltzmann constant and is the temperature correction factor, and is given by
1
2.2 (kT / q) 2
(6.3)
1.959 E
For CNF/CNT with a work function 5 eV [174,182] and temperature below 1000 K, the
value of [ sin( ] in eqn. (6.2) is always 1.0 and in our studied temperature range, (< 400
o
C), the highest contribution of thermionic emission is much smaller than the field emission
current density i.e., the measured emission property is predominated by field emission current
because below 1000 K, the thermionic emission effect is less significant than the field
emission effect [183]. Hence eqn. (6.2) reduced as
3
1
J 1 2 (s b 2
)
ln{ 2 } ln{ r a } (6.4)
E E
Figure 6.2(a) and 6.3(a) shows the emission current density (J) vs. macroscopic field (E)
curves of carbon nanofibers and MWCNTs thin films respectively, for different temperature
and corresponding F-N plot is shown in Figure 6.2(b) and 6.3(b). It has been observed that the
J-E curve in the present work is closely fitted with straight line. This suggests that the
electrons are emitted by cold field emission process. The threshold field, which we define as
the macroscopic field needed to get an emission current density J = 10 A/cm2, were lying in
the range 5.1 to 2.6 V/ m for CNFs and lying in the range 4.0 to 1.4 V/ m for MWCNTs for
the variation of temperature from 300 K to 650 K.
Figure 6.2. J-E graph of CNFs for different temperature (a) and corresponding F-N plot of CNFs (b)
[From ref. 179].
Figure 6.3. J-E graph of MWCNTs for different temperature (a) and corresponding F-N plot of
MWCNTs (b) [From ref. 179].
It is clear from the Figure 6.4(a) and (b) that with the increase of temperature, threshold
field decreases and current density increases for CNFs as well as MWCNTs. The emission
current density is strongly dependent on the work function and as well as on the aspect ratio.
For an example to get emission current density J = 5 mA/cm2, 9.5 V/ m field needed for
CNFs but for MWCNTs 4.5 V/ m field needed for the same current density at 650 K ambient
temperature. The work function of materials is temperature dependent. Therefore, the
decrease of threshold field with the increase of temperature may be due to the decrease of
work function of CNF and MWCNT films.
Figure 6.4. The variation of threshold field and emission current density with temperature (a) for CNFs
(b) for MWCNTs [From ref. 179].
Figure 6.5 shows the variation of the effective work function with temperature for CNFs
and MWCNTs. It shows that the effective work function decreases with increase of
temperature. The variation of the effective work function with temperature is consistent with
the variation of the emission current density as observed. From the quantum mechanical
tunneling phenomena, we know that the Fermi energy determines the field emission current.
The work function ( ) is given by EV - EF, where EV is the fixed vacuum level and EF is
the Fermi level [184]. For low temperature emission, Fermi level is lower and the electrons
have to transmit through a much boarder barrier as shown in Figure 6.6(a) and for high
temperature field emission, Fermi level increase so barrier width decrease as shown in Figure
6.6(b). Hence the emission current increases under same field for high temperature field
emission (as shown in Figure 6.4).
Figure 6.5. Variation of the effective work function with temperature for CNFs and MWCNTs [From
ref. 179].
Figure 6.6. Schematic diagram of the transmission of electrons from the MWCNTs at low (a) and high
(b) temperatures under applied field [From ref. 170].
As the field emission characteristic has a complicated dependence on electric field and
temperature when these two factors coexist, and cannot be explained simply by F–N or
thermionic emission model, the information of can still provide fruitful information.
According to the F-N plot (Figure 6.2(b) and 6.3(b)), the slope m (given by equation (6.5))
would represent the combined effect of work function and enhancement of local electric field
and is given by,
3
2
m
b (6.5)
The effective work function C is related with the true work function through the
2
3
relation [183] C = / . Using = 5 eV is the work function of CNF/CNT, the field
enhancement factor was calculated from the slope of the F-N plot, lies in the range 4589 to
9917 and the corresponding effective work function C lies in the range 0.018 to 0.011 eV
for the CNFs with different ambient temperature. For MWCNTs field enhancement factor lies
in the range 5838 to 11448 and the corresponding effective work function C lies in the range
0.015 to 0.010 eV. The field enhancement factor increases monotonously with the
temperature, which explains very well the increase in emission current density with
measuring temperature (shown in Figure 6.4). But physically the field enhancement factor
should depend on the geometric shape of the emitter rather than temperature. Hence there
may be other factors responsible for such temperature dependence of emission current.
Although the exact explanation of the observed temperature dependence of the emission
current needs more research, the following effects may have strong influence. The presence
of defects or surface states is predominant in nanomaterials like CNT or CNF. Wang et al.
and Xu et al. also proposed that the field emission property is related with the defect densities
[161,162]. These states might have small activation energies and when the temperature in
increased, the carriers trapped in these states are activated into the conduction band and more
emission currents results. Chen et al. [185] observed the field emission of different oriented
CNTs and they discovered that the CNTs oriented parallel to the substrate have a lower onset
applied field than those oriented perpendicular to the substrate. They also suggested that the
defect emission mechanism is a reason for the low onset electrical field. Obraztsov et al.
[164] have also found that the field emission properties were improved by increasing the
density of structural defects. The field enhancement factor also depends on aspect ratio of
carbon nanostructure. The aspect ratio (h/r, where r is the average radius and h is the length
of the tubes, respectively) of our MWCNTs is greater than that of CNFs. From our
experimental result we also see that the field emission properties of CNTs are better than
those of the CNFs with large diameter. Another possible reason is that the screening effect,
which diminishes the electric field near the CNFs. Nilson et al. proposed from their
experimental observation that to overcome screening effect the distance between nanotubes
will be 1-2 times the tube height [186]. As our deposited vertically aligned CNFs are compact
so screening effects is much remarkable. Since it is well known that MWCNTs are mostly
conductive, their conductivity should decrease with higher temperature and thus causes the
screen effect less remarkable.
APPLICATION OF CARBON NANOFIBER AND CARBON NANOTUBE

Along with the improvement of the production and characterization techniques for
nanotubes, progress is being made in their applications. MWCNTs have also exhibited
ballistic transport [187]. The reasons for high electron conductance in carbon nanotubes are as
follows; i) physical perfection: a smooth surface with no chemical dangling bonds and no
edges reducing surface states, which affects backscattering; ii) strong covalent bonds in CNT
and iii) no low-energy dislocations or defects, which also reduces backscattering and provide
stability for high current transport. Carbon nanofibers/nanotubes have many properties from
their unique dimensions to an unusual current conduction mechanism that make them ideal
components of electrical circuits. Due to their semi conducting properties,
nanofibers/nanotubes may be the building blocks for smaller, faster computers. Other
potential applications in electronics and computers include, storage devices, It was proposed
to use nanotubes as central elements of electronic devices including field-effect transistors,
single-electron transistors [188] and rectifying diodes [189] and for logic circuits [134]. The
geometric properties of nanotubes such as the high aspect ratio and small tip radius of
curvature, coupled with the extraordinary mechanical strength and chemical stability, make
them an ideal candidate for electron field emitters [129,186]. CNT field emitters have several
industrial and research applications; flat panel displays [131], outdoor displays, traffic signals
and electron microscopy. De Heer et al. [135] demonstrated the earliest high intensity
electron gun based on field emission from a film of nanotubes. The properties of carbon
nanotubes (CNTs) and the less crystalline carbon nanofibers (CNFs) have attracted
considerable interest for both scientific and technological issues [17,190]. Their impressive
mechanical properties [191], high current carrying ability [192], and field emission
performance [135] have opened the way to a number of applications such as field emission
devices [193], interconnects [194], sensors [195], super-capacitors [196], fuel cells [197] and
battery electrodes [198]. The vertical geometry of carbon nanofibers (CNFs) is particularly
useful in technologies such as nanoelectronics [89], electrodes for biosensing/stimulation
[90], nanomechanical [91], and thermal interface materials [92]. Achieving near-ohmic
contact at the nanotube-metal interface as well as investigating the affect of nanotube
crystallinity is critical for evaluating and modeling the electrical performance of on-chip
interconnects. Sim et al. reported that the carbon-nanofiber-based (CNF) ionization gas
sensing devices on plastic substrates [199]. The device is configured as diode structure with a
Cu plate and a CNF film as anode and cathode respectively. For a fixed applied voltage of
600 V, the ionization current of that device exhibits two regions of linearity with respect to
gas pressure below and above 5 Pa, suggesting that the device can be employed as vacuum
ion gauge. Ngo et al. reported that due to thermal conductance properties CNF-Cu composite
material could be use as a thermal interface material in both IC packaging and equipment
cooling applications [200]. The concept of nanothermometer using CNT was first proposed
by Gao et al. [201,202]. They used gallium filled CNT as nanothermometer and transmission
electron microscope is necessary for observation of CNT during temperature measurement.
From our experimental observation, it is proposed that the nanothermometer can be

constructed using MWCNT more easily. As the emission current vary linearly with
temperature for a particular applied electric field, so temperature can be directly measured.
The sensitivity of the nanothermometer can be adjusted by choosing the area of the MWCNT
film or appropriate applied electric field. The above study shows that the temperature
dependent field emission property of CNFs and MWCNTs have potential for development of
direct thermal-to-electrical power conversion applications. Continued improvements in the
PECVD of CNFs/CNTs and related nanostructures are indeed required to explore the
potential utility of these structures in advanced applications and future large-scale integration.
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Lecture Material 25:
CARBON NANOTUBES: A NEW ALTERNATIVE FOR

ELECTROCHEMICAL SENSORS
ABSTRACT
The goal of this chapter is to summarize the recent advances in carbon nanotubes as
a new material for electrochemical sensors. Since their discovery in 1991, carbon
nanotubes have received considerable attention in different fields. Their special geometry
and unique electronic, mechanical, chemical and thermal properties make them a very
attractive material for the design of electrochemical biosensors.
The first application of carbon nanotubes in the preparation of a sensor was reported
by Britto in 1996. Since then, an increasing number of publications involving sensors
based on carbon nanotubes (either single or multi-wall) for substrates like glucose,
lactate, alcohols, phenols, neurotransmitters, aminoacids, proteins, carbohydrates among
others, have been reported. This fact demonstrates the usefulness of carbon nanotubes for
the development of electrochemical sensors.
The advantages of carbon nanotubes for promoting electron transfer reactions -with
special emphasis in those involving biomolecules-, the different methodologies for
incorporating carbon nanotubes in sensors (either suspended in solutions, in polymeric
films or in composite matrices), the analytical performance of the resulting biosensors as
well as future prospects are discussed in this article.
GENERAL ASPECTS OF CARBON NANOTUBES

In the last years there has been an increasing interest in nanoscience, basically to
understand the behavior of structures with sizes close to atomic dimensions. Even when many
nanostructures are currently under investigation, the area of nanotubes is one of the most
active. Carbon nanotubes (CNTs) present one of the simplest chemical composition and
atomic bonding configuration, at the same time that show the most extreme diversity and
richness among nanomaterials referred to structures and associated properties [1]. Since their
discovering in 1991 by Iijima [2], carbon nanotubes have been the target of numerous
investigations due to their unique properties [1,3-6]. The outstanding structural, electronic and
mechanical properties make them a very unique material attractive for a wide range of
applications [7].
It is important to mention the crucial aspect of carbon hybridization in the properties of
the resulting material. While in diamond the sp3 hybridization originates a rigid and almost
isotropic structure, the sp2 of graphite shows planar bonds, three-fold coordinated in the
planes with weak bonding between planes and anisotropic physical properties [4, 7]. Carbon
nanotubes are built from sp2 carbon units and present a seamless structure with hexagonal
honeycomb lattices, being several nanometers in diameter and many microns in length [3, 6].
CNTs are closed structures that present two well defined regions with properties clearly
different, the tube and the cap [8]. Each end of the nanotubes is capped with half of a
fullerene-like molecule that is responsible for the diameter of the tube [9]. To obtain the
convex structure, it is necessary to introduce a positive curvature into the planar hexagonal
graphite lattice and it is done by creating topological defects, which in this case are pentagons
[6]. As it is discussed in the following sections, these caps can be opened by using different
treatments providing new alternatives for many interesting applications.
Adapted from Reference 10.
Figure 1. Schematic diagram of a single-wall carbon nanotube (SWNT) (a) and a multi-wall carbon
nanotube (MWNT) (b).
There are two groups of carbon nanotubes, multi-wall (MWCNTs) and single-wall
(SWCNTs) carbon nanotubes [3,6,10,11] (Figure 1). MWCNTs can be visualized as
concentric and closed graphite tubules with multiple layers of graphite sheet defining a hole
typically from 2 to 25 nm separated by a distance of approximately 0.34 nm [2-4]. SWCNTs
consist of a single graphite sheet rolled seamlessly, defining a cylinder of 1-2 nm diameter.
MWCNTs can be considered as a mesoscale graphite system while SWCNTs are real single
large molecules [7].
It is important to define the chiral vector of the nanotube Ch, which is given by Ch = nā1 +
m ā2 where a1 and a2 are unit vectors in the two-dimensional hexagonal lattice, and n and m
are integers as shown in Figure 2 [12]. The diameter, dt, is the length of the chiral vector
divided by ¼. Another important parameter is the chiral angle θ, which is the angle between
Ch and a1.
From reference 12.
Figure 2. Schematic diagram of the hexagonal sheet of graphite. Carbon atoms are at the vertices. The
parameters that define the nanotube structure when the sheet is ‗rolled‘ (chiral angle, chiral vector, basis
vectors a1 and a2) are indicated in the figure.
The ends of the chiral vector meet each other when the graphene sheet is rolled up to
form the cylinder. According to this, tubes of different diameters and helical arrangements of
hexagons can arise by changing the values of n and m. In other words, depending on the
values of n and m it is possible to have different nanotube structures [4,9,10]. In fact,
depending on how the two-dimensional graphene sheet is rolled up, there are three types of
carbon nanotubes, armchair, zigzag and chiral. A schematic representation of these structures
is given in Figures 3 and 4. Armchair nanotubes are formed when n = m and the chiral angle
is 30o. Zigzag nanotubes are formed when either n or m is zero and the chiral angle is 0o. All
other nanotubes are known as chiral nanotubes and present chiral angles intermediate between
0o and 30o [1, 6, 9, 12].
Figure 3. (a) Schematic honeycomb structure of a graphene sheet. SWCNTs can be formed by folding
the sheet along lattice vectors. The vectors a1 and a2 are shown. Folding of the (8,8), (8,0), and
(10,-2) vectors lead to armchair (b), zigzag (c), and chiral (d) tubes, respectively. From reference 1.
Figure 4. Schematic representation of the atomic structure of an armchair (a) and a ziz-zag (b)
nanotube. See reference 12.
The electrical properties of CNTs depend sensitively on the (n,m) indices and, therefore,
on the diameter and chirality [4, 9 ,12]. According to the m,n structural parameters values,
SWCNTs can be either a metal, semiconductor or small-gap semiconductor [1,4,9,12]. When
n=m, the CNTs are metallic. If n – m = 3 x integer, the CNTs present an extremely small
band gap and at room temperature they have metallic behavior. For other intermediate values
of n – m the behavior is that of a semiconductor with a given band gap [4,9]. This extreme
sensitivity of electronic properties on structural parameters is one of the most important
aspects of nanotubes that make them very unique. Calculations have predicted that all the
armchair tubes are metallic while the zigzag and chiral tubes are either metallic or
semiconductor depending on their diameter and chiral angle [6,13].
The combination of size, structure and topology give nanotubes important mechanical
properties such as high stability, strength and stiffness, low density and elastic deformability
with interesting surface properties (selectivity, surface chemistry). The helicity as well as the
diameter introduce important changes in the electronic density of states, given to nanotubes
unique electronic characteristics. These electronic properties open the doors to a wide range
of fascinating electronic devices applications [7,9]. Topological defects in nanotubes result in
local perturbations to their electronic structure. In this sense, the caps are more metallic than
cylinders due to the pentagonal defects. These defects also enhance the chemical reactivity of
the ends giving the possibility to open the tubes, fill them with foreign substances and
functionalize the ends [14-19].
The strength of carbon bond determines the fascinating mechanical characteristics of this
material that are superior to other known materials [20, 21]. CNTs are extremely flexible. In
fact, they can be twisted, flattened and bent into small circles without breaking. They can also
be compressed without fracture [6,7,9].
CNTs also possess interesting electrochemical properties. Several works have
demonstrated the electroactivity of CNTs due to the presence of reactive groups on the
surface [10, 22-24]. The small dimensions produce high current densities on the surface of the
electrodes allowing the study of heterogeneous process with excellent results. Ab-initio
calculations demonstrated that the improvement in the electron transfer is due to the curvature
of the tubes that originate changes in the energy bands close to the Fermi level. The presence
of pentagonal defects produced regions with charge density higher than those observed in the
region of hexagonal graphite, either in planar or in tubular structures (3-4 times higher)
demonstrating the connection between topological defects and electroactivity of CNTs [23].
As it is discussed in the next sections, lower peak potential separations and higher peak
currents are observed in the voltammetric behavior of several molecules in the presence of
CNTs. These results suggest an interesting electrocatalytic activity, associated with the
carbon nanotubes dimensions, the electronic structure and the topological defects present on
the tube surface [7, 11, 22-24].
SYNTHESIS OF CARBON NANOTUBES

The synthesis of CNTs is receiving considerable interest and the main goal is to obtain
large scale production of highly pure CNTs. There are three basic methods for synthesis of
SWCNTs and MWCNTs: electrical arch discharge, laser ablation (laser vaporization) and
chemical vapor deposition (CVD) (or catalytic decomposition of hydrocarbons) [1,7, 9,10, 25,
26].
Electric arc Discharge
It was the first method used for fabricating CNTs and it consists of establishing an
electric arc discharge between two graphite electrodes (cathode of around 8-12 mm and anode
of 6-8 mm) separated approximately 1 mm under an inert helium atmosphere. A bias of
around 10-35 V is applied to the electrodes to establish currents of 60-100 A. The most
important factor is to get stable discharge plasma. Since elevated temperatures are achieved,
the anode material is sublimated and deposited on the cathode and surrounding walls. The
inner deposit contains mainly MWCNTs mixed with polycrystalline graphite nanoparticles,
while the outer shell is composed mainly of fused graphite powders, nanoparticles and
amorphous carbon [27, 28]. It was reported that the use of single or bi-metal mixtures of Co,
Ni, Y and Fe favored the production of SWCNT [29, 30], the most efficient being the
combination Co/Y and Ni/Y.
Several are the parameters that affect the arch-discharge nanotubes production such as
gas type, pressure and flow rate, electric field strength, electrode materials and dimensions, in
addition to unquantified variables such as apparatus size and geometry, and thermal gradients
[25]. The typical production rates are around 20-100 mg/min.
Lee et al [31] proposed the use of plasma rotating electrode process to generate a more
stable discharge plasma. The goal was to distribute the micro-discharges more regularly
between the two electrodes, stabilizing, therefore, the plasma. On the other hand, the rapid
rotation produces a centrifugal turbulence that allows the vaporized material to move it
outwards and deposit on the chamber walls or close collectors rather than on the cathode
surface, producing CNTs of smaller diameters. Another avenue [32] consisted of the
discharge of the electrodes submerged in liquid nitrogen providing, in this way, an inert
environment and temperature control.
Laser Ablation
The solid graphite target was mounted in a quartz tube and placed in a temperature-
controlled oven. After vaporization of this target with a pulsed Nd:YAG laser, a carbon-based
soot was collected from the inside of the apparatus. To obtain SWCNTs, it is necessary to
dope the graphite target with transition metal catalysts. The system was improved by the
incorporation of a second laser Nd:YAG delayed slightly behind the first [33]. The
incorporation of equal parts (0.5-1.0 atom %) of cobalt and nickel powders in graphite have
demonstrated to be highly efficient for the production of SWCNTs. Eklund et al [34] have
produced SWCNTs at rates around 1.5 g/h using a 1.7 kW sub-picosecond free electron laser
(FEL). As in the case of arch-discharge, a careful evaluation of the different variables has to
be made in order to optimize the production.
Chemical Vapor Deposition
This procedure involves the pyrolysis of gas molecules with high content in carbon at
elevated temperatures in the presence of catalyst [10, 25]. There are two basic protocols, in
one of them, called supported growth process (the most used), the catalyst is prepared and
deposited on a support medium, which is inserted into a flow apparatus (a tube at atmospheric
pressure in a temperature controlled furnace) and exposed to elevated temperatures, usually
500-1100 oC for a given time. In the other protocol, called floating-catalyst growth, the
catalyst and the carbon source are injected into the system simultaneously, either in gas phase
or in aerosol. Therefore, the decomposition and reaction can occur suspended in the gas flow
or following self-deposition on a surface in the reactor. The most common support media are
metallic Si, Si wafers or different SiO2 based materials. Fe, Co and Ni have demonstrated to
be the most successful catalysts. It is important to mention that the diameter of the carbon
nanotubes is proportional to the particle size, making it possible to tune the nanotube diameter
by controlling the catalyst deposition. Even when the most widely used hydrocarbon is
acetylene; methane, ethylene, propylene and few aromatic compounds have been also
employed. The use of templates for growing the nanotubes inside porous materials has been
also described. The advantage of catalyst incorporation into the gas flow is to circumvent the
effects of catalyst deactivation by coating with pyrolized hydrocarbons. Floating catalyst
methods contribute to the production of SWCNTs, while most of other CVD-based methods
produced mainly MWCNTs.
USE OF CARBON NANOTUBES AS ELECTRODE MATERIAL

Due to their peculiar properties, carbon nanotubes have received enormous attention for
the preparation of electrochemical sensors, as it was reviewed by Zhao et al [3], Wang [35]
and Li et al. [36]. Different procedures for immobilizing the nanotubes onto electrochemical
transducers have been described and the most representative are presented in the following
sections.
Pretreatments of Carbon Nanotubes
As in the case of other carbon materials [37], some pretreatment of CNTs is necessary to
improve the electron transfer properties and/or to allow further functionalization. The
protocols are based, in general, on the oxidation of CNTs. Depending on how drastic is the
treatment, it is possible not only break the tubes but also shorten them. In all cases the ends
and side walls are modified with a high density of diverse oxygenated functions, mainly
carboxyl groups. Different schemes have been proposed and in order to facilitate the
understanding, they were agrupated in chemical and electrochemical. Spectroscopic
techniques like Raman and IR; microscopies like AFM, SEM and TEM; and electrochemical
techniques indicated that the physical structure of CNTs remains the same, although the ends
were opened and oxidized to give carboxylic groups.
Chemical Pretreatments
Oxidation in air is one of the methods for purifying CNTs. However, at the same time
that purified them, this oxidative scheme produces the activation of CNTs. Palleschi et al.
[38] proposed the oxidation at 400 oC using an air flow of 12 mL/min for 1 hour.
Sotiropoulou and Chaniotakis [39] proposed two mechanisms for activating MWCNTs grown
by CVD on Pt substrate. One of them was performed by air oxidation at 600 oC for 5 min
under air flow.
There are a large number of protocols that use acidic solutions for activating carbon
nanotubes. Solutions of sulfuric, nitric and hydrochloric acids either concentrated or diluted,
or mixtures of them have been used at room temperature or under refluxing with or without
sonication for different times. In a general scheme, once the surface was oxidized, the next
step was the careful rinsing of the oxidized nanotubes with ultrapure water, followed by the
drying step either at room temperature by exposure to air or under IR lamp, or in vacuum at a
given temperature.
Compton et al. [40] proposed the activation of MWCNTs in concentrated nitric acid at 60
o
C for 20 hours. Mao et al. [41] have reported the pretreatment of MWCNT by refluxing in 3
M nitric acid for 12 h.
A more drastic scheme proposed the acidic oxidation using a mixture of concentrated
sulfuric (98 %) and nitric acid (65 %) in a ratio 3:1 for 8 hours at 40 oC [39]. Under these
conditions the tube caps are opened and the tubules are shortened in fragments of different
length. After that, the CNTs were washed with pure water and dried at 100 oC overnight.
From reference 68.
Scheme 1. Schematic representation of the formation of SWNT assemblies.
In another protocol [42], the MWCNTs-modified electrode was prepared by using

MWCNTs (obtained by CVD) previously treated by refluxing in concentrated nitric acid for
about 5 hours, filtered and washed with pure water until neutral pH and then dried under
vacuum.
Palleschi et al. [38] proposed an additional oxidation of CNTs previously oxidized in air
flow by dispersing in 6.0 M HCl for 4 hours under ultrasonic agitation followed by washing
until neutral pH and drying. The same group also presented another treatment by dispersing
CNTs in 2.2 M nitric acid for 20 hours at room temperature (under ultrasonic agitation the
first 30 min), then washing with distilled water to neutrality and drying in oven at 37 oC.
Luo et al. [43] proposed the dispersion of CNTs in 4.0 M HCl for 4 hours under
ultrasonic agitation to eliminate metal oxide catalysts. After rinsing carefully with water until
neutral pH and drying, the CNTs were dispersed in 60 mL mixture of concentrated nitric acid
plus sulfuric acid (1:3) with ultrasonic agitation for 4 hours in a water bath followed by
washing until neutral pH and finally drying.
Hu et al. [44] performed pretreatments by soaking the CNTs in 5 M nitric acid,
ultrasonically dispersing for 6 min and drying under IR lamp oven for 4 h at 45 oC. Then, the
CNTs were diluted with a large amount of water and a little Triton X-100 to increase
solubility, sonicated until getting a black solution which after filtering gave the nanotubes.
The picture displays in Figure 5 clearly shows the absence of the caps in CNTs after
performing the pretreatment.
From reference 44.
Figure 5. SEM micrograph of the carboxyl-modified CNTs.
Hu et al. evaluated the functionalization of CNTs using different chemical pretreatments

[45]. They oxidized the CNTs in two ways, by using mixed acids for 10 min at 65 oC in a
water bath, and by soaking them in nitric acid under sonication for 10 or 20 min. An increase
in the resistivity was found for longer times in the acidic solution and by using mixture of
acids.
Electrochemical
Different schemes of electrochemical pretreatments have been proposed. In general, they
depend on the system under investigation. For instance, for the oxidation of dopamine, 18
cycles between -1.00 V and 1.50 V at 1.0 V/s in a 0.050 M phosphate buffer pH 7.40
demonstrated to be the optimum pretreatment for obtaining the best voltammetric behavior of
dopamine [46]. However, for the oxidation of amitrole, 75 cycles between -1.0 and 1.5 V at
1.0 V/s in 0.050 M phosphate buffer solution pH 7.4 were necessary to obtain the best
analytical signal [47].
In some cases a combination of several pretreatments was used to improve the electron
transfer. For instance, Palleschi et al. [48] proposed a treatment consisting of an oxidation
step at 400 oC in the presence of O2 for 1 h, then dispersion in 6.0 M HCl for 4 hours under
ultrasonic agitation. Another pretreatment proposed by the authors was the oxidation of CNTs
in a 2.2 M HNO3 solution for 20 h at room temperature with sonication the first 30 min. After
that, once the CNTs were immobilized on the electrode electrochemical pretreatments were
performed by preanodization and precatodization at 1.70 V and -1.50 V for 3 min each in a
phosphate buffer solution (0.2 M pH 7.0). Another scheme also proposed by Palleschi‘s
group [49] was the combination of the typical chemical oxidation in oxygen flow or in the
presence of nitric acid and electrochemical pretreatment by cycling with increasing potentials
range in 0.5 M sodium sulfate between 0.2 y 0.1 V or between 0.5 and –0.1 V at 100 mV/s for
5 min, depending on the compound under investigation, followed by a succesive cycling in a
wider potencial range up to 1.8 V y -0.4 V several cycles.
Strategies for the Preparation of CNTs-modified Electrodes
One of the problems for the preparation of sensors based on the use of carbon nanotubes
is their insolubility in usual solvents. Therefore, it is necessary to disperse them in an
adequate medium. Several strategies have been proposed for the immobilization of carbon
nanotubes on electrochemical transducers, the most significant are summarized below.
Dispersion in Different Solutions
Acidic Solutions
CNTs have to be dispersed in an adequate solution before immobilizing on a given
electrode. In general, the procedures are based on casting the electrode, usually glassy carbon
or gold, with a drop of the given dispersion and drying under different conditions.
In one case [50] the electrode was prepared by casting a polished and clean glassy carbon
electrode GCE with 10 µL of a solution obtained by dispersion of MWCNTs in concentrated
sulfuric acid (1 mg/mL) followed by drying at 200 oC for 3h and careful rinsing.
Wang et al. [51] proposed the preparation of MWCNT-GCE by casting a polished GCE
with 20 µL of a concentrated nitric acid solution containing 2 mg/mL MWCNTs followed by
a drying step at room temperature for 30 min.
N,N-Dimethylformamide
In general, the immobilization of CNTs on electrodes by dispersing in N,N-dimethyl
formamide (DMF) consists of polishing the electrode, casting it with the CNTs suspension in
DMF and drying to evaporate the solvent.
Li et al. [52, 53] have reported the preparation of CNTs-modified GCE by casting the
GCE with 15 µL of a SWCNTs suspension (armchair structure, prepared by arch-discharge
and purified by oxidation in air) in DMF (0.1mg/mL). The drying step was performed under
an IR heat lamp.
Pang et al. [54] proposed the use of a SWCNT-modified GCE prepared by casting the
polished GCE with 2 µL of the black suspension of SWCNT in DMF (1mg/mL), followed by
heating under an infrared lamp to remove the solvent.
Nafion
Wang et al. [55] have reported on the ability of the perfluorosulfonated polymer Nafion
to solubilize SWCNTs and MWCNTs. They proposed that a homogeneous, well-distributed
solution of Nafion/CNTs was obtained with 0.5 % v/v (in 0.05 M phosphate buffer pH 7.4)
and 5% v/v polymer solution for SWCNTs and MWCNTs, respectively. A polished GCE was
modified with 20 µL of a 2 mg/mL CNTs solution in 0.5 % w/w Nafion. The coating was
allowed to dry at room temperature for 2 hours. The dispersion was prepared by agitation in
an ultrasonic bath for 10 min and in this way, it was stable for 3-4 days.
Mao et al. [41] reported the use of a CNT-modified GCE prepared by casting the GCE
with 5 μL of a CNTs dispersion prepared in 0.5 % methanolic Nafion solution and followed
by the evaporation of the solvent for 10 min.
Chitosan
Multiwalled carbon nanotubes were solubilized in an aqueous solution of the biopolymer
chitosan (CHIT) [56]. A 0.50 w/v % CHIT stock solution was prepared by dissolving
chitosan flakes in hot (80-90 °C) aqueous 0.05 M HCl. The solution was cooled at room
temperature, and its pH adjusted to 3.5-5.0 using a concentrated NaOH solution. CNTs were
solubilized in CHIT solutions (0.50-3.00 mg of CNT mL-1) by ultrasonic agitation for 15
minutes. An aliquot of 30 µL of the colloidal solution of CNT-CHIT was placed on the
surface of GCE and dried for 2 hours at room temperature.
Other Media
Compton et al. [57] proposed the immobilization of CNT on basal plane pyrolitic
graphite electrodes using different approaches. One of them was done by dispersing the
powder in acetonitrile and then casting the electrode. The solvent was eliminated by
evaporation.
Other methodology proposed the preparation of MWCNTs-modified electrode by casting
a gold surface previously cycled between 0.0 and 1.5 V in 0.5 M sulfuric acid with 5 µL of a
black suspension of CNTs prepared by dispersing the oxidized MWCNTs in double distilled
water (0.5 mg/mL) [42]. The electrode was then dried under vacuum at about 50 oC.
Hu et al [58] have proposed an amperometric sensor by casting a GCE with 10 µL of the
dispersion of MWCNTs in water in the presence of dihexadecylhydrogenphosphate (DHP)
(1mg of CNTs in 5 mg DHP and 5 mL water, under sonication). The solvent was allowed to
evaporate at room temperature in air.
Incorporation in Composite Matrices Using Different Binders
Teflon
The advantages of using an electrode based on the dispersion of CNTs in a Teflon binder
was reported by Wang et al. [59]. The CNT/Teflon composite was prepared by hand-mixing
(with spatula) CNTs and granular Teflon for 10 min and then packed the paste into the
electrode cavity.
A MWCNT/Teflon electrode was prepared by hand-mixing the desired amounts of the
CNT with granular Teflon for ten minutes [60]. The portions were packed in a Teflon tube (2
x 2 mm cavity), smoothed on a weighing paper and rinsed with distilled water.
Bromoform
Hill et al. [61] have reported on the feasibility to use carbon nanotubes as electrode
material, by packing oxidized CNTs into a glass capillary in bromoform, nujol, deionised
water or mineral oil. When necessary, bromoform was eliminated by backing the packed
nanotubes in an oven at 80-100 oC. Britto and co-workers [62] have proposed the preparation
of an electrode based on the dispersion of CNTs with bromoform and packing inside a glass
tube.
Mineral Oil
Rivas and Rubianes [46] reported for the first time the advantages of a new composite
material prepared in an easy, fast and very effective way by dispersing MWCNTs within
mineral oil as binder (60.0/40.0 % w/w) . The resulting carbon nanotube paste electrode
(CNTPE) retained the properties of the classical graphite carbon paste electrode (CPE) such
as the feasibility to incorporate different substances, the low background currents the easy
renewal of the surface and composite nature. Electrodes with oil content smaller than 40.0 %
w/w were difficult to pack into the Teflon tube. Pastes with 50.0 % w/w showed similar
behavior. CNTPE prepared with short (1-5 microns length) and long MWCNTs (5-20
microns length) of 20-50 nm diameter demonstrated to be highly useful as detectors in flow
systems [63]. The content of mineral oil was an important variable in the preparation of these
carbon nanotubes composites and even when no substantial differences were observed
between the electrodes, those prepared with long carbon nanotubes (55.0 % w/w) and mineral
oil (45.0 % w/w) allowed to obtain less noisy and more reproducible signals.
Palleschi et al. [38] reported on a composite electrode by mixing SWCNTs and mineral
oil, 60/40 % w/w. SEM pictures showed a surface topography more uniform than in the case
of CPE with the nanostructures embedded inside the oil binder.
Wang et al. [64] proposed the use of CNTPE for the detection of homocysteine. The
composite was prepared by mixing MWCNTs of 5-20 nm length with oil in a ratio 3:2
(CNT/oil) and then placed in a plastic pipette tip (0.5 mm diameter).
Magno et al. [65] reported an amperometric biosensor prepared by mixing MWCNTs
with mineral oil (60/40 % w/w) followed by the covering with a polymer obtained by
electropolymerization of a 1.0 mM 3,4-dihydroxybenzaldehyde (3,4-DHB) (in 0.1 M
phosphate buffer solution pH 7.0) at 0.3 V for 1 min.
Wang and coworkers [66] reported on the use of CNTPE containing Cu as a detector for
Capillary Electrophoresis for the determination of carbohydrates compounds. The electrode
was prepared by hand mixing mineral oil, MWCNT and copper powder in a weight ratio
1:1:2 (carbon/oil/Cu).
Inks
Another interesting strategy proposed by Wang et al. [67] was the preparation of a
composite material based on the dispersion of CNTs in ink, in a mode similar to that for
preparing graphite screen printed electrodes (SPE). The electrodes were fabricated following
two strategies. In one case they were prepared using the same ink as that for preparing the
SPE, obtaining a very efficient combination of the advantages of thick-films sensors with the
excellent electrochemical properties and analytical performance of CNTs. The other way for
preparing these screen printed electrodes consisted of mixing 60 mg CNTs with 500 µL of
isophorone solution containing 2 % (w/v) PVC, 2 % (v/v) DBE-4 and 2% (v/v) DBE-5 (DBE-
4: dibasic ester containing 98.4 % dimethyl succinate and 0.3 % dimethyl glutarate; DBE-5:
dibasic ester containing 99 % dimethyl glutarate and 0.4 % dimethyl succinate) until
homogeneous aspect. The ink was printed on alumina ceramic plates. After that, the resulting
electrodes were cured for 1 h at 150 o C and then allowed to cool down at room temperature.
SEM pictures showed a microporous structure of flake-shaped particles non-uniformly
distributed.
Immobilization on Pyrolitic Graphite Electrodes

Compton et al. [40] proposed the immobilization of MWCNTs on basal plane pyrolitic
graphite electrodes by abrasively attaching CNTs on the electrode surface by gently rubbing a
polished electrode on a filter paper containing 2 mg MWCNTs for 1 min.
Luo et al. [43] proposed an electrode prepared by intercalation of CNTs in a pyrolitic
graphite electrode previously polished with emery paper and alumina slurries and sonicated
with water. It was performed by grounding the dry graphite electrode on a weighing paper
containing a suitable amount of CNTs powder to intercalate them on the graphite surface by
mechanical force and adsorption.
Using other Methodologies

Liu et al. [68] proposed the use of SWCNTs previously cut by oxidation in a mixture of
acidic solutions under sonication. Once the carboxyl groups were generated, the assemblies
were prepared on the top of the gold surface modified with monolayers of 11-amino-N-
undecylmercaptan. The covalent attachment between the carboxyl residues and the amino
groups was performed by using dicyclohexylcarbodiimide as condensing agent.
Figure 6. Electron microscope images. (A) Vertically aligned multiwalled CNT arrays with length
about 1 µm. (B) Collapsed CNT arrays after purification process. (C) CNT arrays with SOG after
purification and tip opening process. (D) High-resolution transmission electron microscope image of an
opened CNT end. See reference 69.
Another interesting avenue was the use of MWCNTs arrays grown by CVD using Fe as
catalyst and C2H4. SEM and TEM showed that CNTs were relatively straight with ends
closed and metal-encapsulated catalyst [69]. After an extensive oxidative pretreatment by
heating in air and acid oxidation to open the nanotubes and generate carboxylic groups, the
CNTs collapse in most cases. Therefore, the authors proposed the use of a film of spin-on
glass (SOG) to fill the gaps between the individual CNTs. The spin-on glass film with 15 %
w/w methyl groups bond to Si atoms of Si—O backbone was deposited at a spin speed of
3000 rpm, followed by curing at 400 oC for 4 hs under positive pressure of argon (Scheme 2).
Therefore, the SOG film provides the structural support to the carbon nanotubes and serves as
a dielectric material insulating the individual CNTs. Figure 6 shows SEM images of the
vertically aligned CNTs array(A), collapsed CNTs array before depositing SOG film (B),
CNTs array with SOG after oxidative treatment (C) and TEM image of an open-ended CNTs.
Scheme 2. Fabrication and oxidative pretreatment of carbon nanoelectrode arrays for functionalization.
See reference 69.
Lin et al [70] proposed a similar methodology by preparing the electrodes modified with
CNTs. The nanotubes were obtained by CVD using Ni as catalyst and once they were grown,
an Epson 828 epoxy-based polymer with an MPDA curing agent was spin-coated on the
substrate covering the half of the CNTs. The protruding part of them was eliminated by
polishing. The oxidation of the CNTs was performed by electrochemical oxidation in 1.0 M
NaOH at 1.5 V for 90 s. Once the carboxylic residues were obtained, they were activated in
the presence of carbodiimide (EDC) and N-hydroxysuccinimide (NHS).
The self assembled layer-by-layer of polyelectrolytes on CNTs previously functionalized
was also proposed [71]. CNTs were synthesized on a grid for TEM by CVD using metallic
catalysts. The negative charges were originated onto the surface of carbon nanotubes by
adsorbing a pyrene derivative.
Poly(diallyldimethylammonium (PDDA) and polystyrene sulfonate (PSS) were
alternatively adsorbed (starting with the polycation) from 30 mM aqueous solutions for 1
hour (first bilayer) or for 18 min for the subsequent layers. The studies were performed by
TEM and elemental analysis. A uniform coverage was obtained even for nanotubes of
diameter as small as 2 nm. The pyrene derivatives do not modify the morphology of CNTs,
they just provide them a negative charge density. Some studies with Confocal Fluorescence
Microscopy were also performed using colorants of opposite charge to the layer under
investigation. It was found that after every adsorption step a charge inversion was obtained
despite the interpenetration of the layers.
Figure 7. Scheme of the CNTs modification using layer-by-layer electrostatic self-assembly of

polyelectrolytes. See reference 71.
Figure 8. TEM image of gold nanoparticles self-assembled on the surface of MWNTs through 17-(1-
pyrenyl)-13-oxo-heptadecanethiol. See reference 72.
Guo et al. [72] reported on the non-covalent modification of SWCNTs by using an

interesting scheme. The SWCNTs were previously refluxed in concentrated nitric acid,
washed with water and dried. The SWCNTs were first converted to acylchloride in SOCl2 and
then they reacted with didecylamine at 90-100 oC for 96 hours. After extranting the nanotubes
and removing the solvent they were ready to use. In a typical experiment, 3 mg of the
interlinker 17-(1-pyrenyl)-13-oxo-heptadecanethiol (PHT) was incubated with 10 mL solution
of modified SWCNT in toluene (0.5 mg/mL). The PHT binds on the surface of SWCNT
mainly through π-π interactions between the pyrenyl units of PHT and the side-wall of
modified SWCNT. Since the PHT presents a thiolated residue, the gold nanoparticles can be
bond to the architecture and obtain MWCNT densely coated with gold nanoparticles,
converting this architecture in a good platform for further biosensors designs. Figure 8 shows
a TEM image of the gold nanoparticles immobilized at MWCNTs through PHT linker.
Another avenue for immobilizing CNTs was the dispersion of CNTs in cyclodextrin [73].
The electrode was prepared in the following way, 1 mg of CNTs was dispersed in 10 mL β-
cyclodextrin (CD) (2% aqueous solution) to give a 0.1 mg/mL solution. An aliquot of 7 µL
was dropped on the previously polished, sonicated and dried under IR lamp graphite
electrode. The electrode modified in this way was dried under IR lamp. As it is discussed
below, the presence of cyclodextrin is not only useful for immobilizing the CNTs but also for
improving the molecular recognition of the resulting structure.
Wang et al. [74] have made an interesting comparison of the electrochemical response of
glassy carbon electrodes modified with MWCNTs prepared by arc-discharge (ARC) and
chemical vapor deposition (CVD). The MWCNTs were dispersed in different media: Nafion,
concentrated nitric acid and DMF. The electrochemical performance of the resulting
electrodes was evaluated using potassium ferricyanide, NADH and hydrogen peroxide. They
found that the electrocatalytic activity, the background current and the overall
electroanalytical performance are highly dependent on both, the method used for preparing
the CNT and the dispersing agent. The lowest capacitances were obtained with CNTs
prepared by ARC while the best amperometric detection of the redox markers was obtained
with nanotubes prepared by CVD and using a DMF as dispersing agent. It is very important
to evaluate the connection between the preparation conditions of carbon nanotubes and the
performance of the resulting electrode material.
ELECTROCHEMICAL BEHAVIOR OF DIFFERENT COMPOUNDS

AT CNTS-BASED ELECTRODES. SENSING PROPERTIES
Cytochrome C and Azurin
Hill et al. [61] have reported the electrochemical behavior of proteins at CNTs mixed
with mineral oil, deionised water, nujol or bromoform and packed within a glass capillary.
They studied the electrochemical response of cytochrome c and azurin and found that these
proteins can be immobilized on and within the packed opened nanotubes without
denaturation.
Similar study was performed with cytochrome c using bare GCE, GCE modified with
untreated CNTs and GCE modified with treated CNTs [53]. While no response was observed
at the GCE and an irreversible behavior was obtained at the unactivated SWCNT-modified
GCE; a quasi reversible bevahior was observed at the activated SWCNT-modified GCE, with
a peak potential separation of 73.7 mV. These results point out the advantages of CNTs in the
electron transfer reaction. A linear relationship between peak current and cytochrome c
concentration was found between 3.0 x 10-5 M and 7.0 x 10-4 M with a detection limit of 1.0 x
10-5 M.
Dopamine and Related Compounds
Britto and co-workers [62] have reported a dramatic improvement in the electrochemical
behavior of dopamine using a composite prepared by dispersion of CNTs in bromoform.
Cyclic voltammograms at this electrode showed a Ep of 30 mV, demonstrating the
reversibility of the dopamine/dopaminequinone redox couple at this material. The electrode
was also challenged by immersing it in brain tissue homogenates and no change in the
voltammetric response for dopamine was observed under these conditions.
Rubianes and Rivas [46] demonstrated the advantages of a composite material prepared
by mixing MWCNTs and mineral oil (CNTPE) on the electrochemical behavior of different
biomolecules. The voltammetric signal for dopamine, ascorbic acid, dopac and uric acid
largely improved at the composite containing CNTs. Figure 9 shows cyclic voltammograms
obtained at 0.100 V/s for 1.0 x 10-3 M ascorbic acid (A), uric acid (B), dopamine (C) and
dopac (D) at composite carbon electrodes containing only graphite (40.0 % w/w), graphite
(30.0 % w/w) and 10.0 % w/w MWCNTs and only MWCNTs (40/60 carbon/oil). In all cases,
larger voltammetric peak currents and lower overvoltages for the oxidation of the different
compounds was obtained. For instance, the peak potential separation for dopamine and dopac
decreased 133 and 313 mV at CNTPE compared to CPE, while the overvoltages for the
oxidation of ascorbic acid and uric acid decreased 230 and 160 mV, respectively.
Palleschi et al. [38] reported the use of a composite electrode obtained by mixing
SWCNTs and mineral oil in a ratio 60/40 % w/w (carbon/oil). Comparable background
currents were obtained with CPE and CNTPE when CNTs were not pretreated, increasing in a
factor of 100 after CNTs pretreatment. The authors reported a significant improvement on the
electrochemistry of dopamine, serotonin, 5-Hydroxytryptamine and other compounds like
caffeic acid, ferricyanide, sodium hexachloroiridate (III) and catechol.
Palleschi et al. [49] also compared the voltammetric behavior of Pt, GCE and CNTPE
(60/40 % w/w ratio) using inorganic and organic redox couples. The capacitances were 30.7
and 5.6 µF/cm2 for CNTPE and CPE, respectively. Both, untreated and pretreated CNTPEs
showed resistances similar to those of Pt and CPE, while GCE exhibited the largest one. The
redox behavior of dopamine and other compounds such as hexacyanoferrate,
hexachloroiodate (III), hexaminruthenium (III), p-methylaminophenol sulfate and
ferrocenemonocarboxylic acid were evaluated at the different electrodes. CNTPE showed
good electroactivity with all molecules, especially using CNTs pretreated with nitric acid. The
oxidation of CNTs produced an increase in the capacitive current making even worst the
electrochemical response. After 15 days exposed to air the signal decreased just 30 %.
Figure 9. Cyclic voltamograms for 1.0 x 10 -3 M ascorbic acid (A), uric acid (B), dopamine (C) and
dopac (D) at diferent electrodes: (- - -) CPE; (. . .) CPE containing 10% w/w MWCNTs; (———)
CNTPE (40/60 % w/w). Supporting electrolyte: 0.050 M phosphate buffer solution, pH 7.40. Scan rate:
0.100 V s-1. See reference 46.
Luo et al. [43] described an electrode based on the incorporation of CNTs for the
simultaneous determination of dopamine, ascorbic acid and serotonin. By using differential
pulse voltammetry, dopamine could be detected in the range of 0.5 to 10 µM in the presence
of 5 µM serotonin and 0.5 mM ascorbic acid, with a detection limit of 0.1 µM. For serotonin,
the signal was linear between 1.0 and 15 µM in the presence of 5 µM dopamine and 0.5 mM
ascorbic acid, with a detection limit of 0.2 µM. No interference was reported on the
determination of 5 µM dopamine and 5 µM 5-hydroxytryptamine in the presence of 10-fold
excess dopac, 5-fold excess uric acid, 200-fold excess oxalate and 500-fold excess glucose.
Excellent reproducibility was also reported. No interference was observed in the
determination of dopamine and serotonin in brain of rabbit.
The study of the groups involved in the redox behavior of CNTs as well as the
electrochemical behavior of dopamine and related compounds was also reported [75]. A GCE
was modified by casting a suspension of nitric acid containing SWCNTs (prepared by arc-
discharge). The studies were performed by XPS and IR, demonstrating the participation of the
carboxylic groups in the redox behavior of the electrode, which were reduced to CH2OH
coupled with four electrons. At a scan rate of 0.1 Vs-1, the Epc and Epa were -0.126 and -0.024
V vs SCE in a Britton-Robinson solution pH 6.9. The resulting electrode demonstrated to
have highly catalytic activity towards several biomolecules such as dopamine, epinephrine
and ascorbic acid. At the SWCNT-film modified GCE, there is an important shifting of the
dopamine oxidation peak potential. The current was linear from 1.0 x 10-6 M to 2.0 x 10-4 M
dopamine.
Bai et al. [76] studied the electrochemistry of empty nanotubes and nanotubes filled with
toluene after casting NTs in a gold electrode. MWCNTs were shortened and then filled with
toluene. The empty nanotubes and toluene-filled nanotubes were dispersed with ultrasonic
agitation in ethanol forming a 0.1 mg/mL solution. 15 µL of this solution were dropped on a
polished gold surface and the solvent was then allowed to evaporate. The authors proposed
that the filling of CNTs with toluene may affect the electronic properties of nanotubes,
improving the charge transfer of dopamine and epinephrine. A linear relationship between
peak current and dopamine concentration was obtained between 5.0 x 10-6 and 3.0 x 10-4 M
while the detection limit was 3.0 x 10-7 M. In the case of epinephfrine, an improvement in the
voltammetric behavior was also obtained.
Li. N. et al [52] reported a study about the voltammetric behavior of dopac at GCE
modified with SWCNTs. The behavior of dopac drastically changes at the CNTs-modified
GCE, with important increase in the peak current in comparison with GCE, indicating a
noticeable increment in the heterogeneous rate constant. A linear relationship between peak
current and dopac concentration was found between 1.0 x 10-6 and 1.2 x 10-4 M, while the
detection limit was 4.0 x 10-7 M. Dopac can be determined at CNTs-modified GCE in the
presence of 3-methoxy-4-hydroxyphenylacetic acid (HVA). Dopac can be also detected in the
presence of 5-hydroxy-tryptamine since both oxidation peaks are clearly distinguishable.
Pang et al. [54] studied the electrochemical behavior of L-dopa at SWCNT-modified
GCE. Before starting, the electrode was immersed for 120 s in the L-dopa solution. L-dopa
showed an irreversible behavior at bare GCE with peak potential separation of 161 mV. On
the contrary, a quasi reversible behavior with peak potential separation of 55 mV was
obtained at the SWCNTs-modified electrode. Experiments performed by differential pulse
voltammetry showed a linear range between 5.0 x 10-7 and 2.0 x 10-5 M L-dopa and a
detection limit of 3.0 x 10-7 M.
Li et al. [77] reported the electrocatalytic oxidation of norepinephrine at a GCE modified
with SWCNTs. The electrode showed a very good reproducibility and stability. A linear
relationship was obtained between the oxidation peak current and norepinephrine
concentration between 1.0 x 10-5 and 1.1 x 10-3 M and the detection limit was 6.0 x 10-6 M.
The electrocatalytic activity of the SWCNTs-modified-GCE was also demonstrated with
dopamine, epinephrine and ascorbic acid.
Another approach to determine epinephrine was proposed by Compton et al. [40]. They
used MWCNTs abrasively attached to the basal plane pyrolytic graphite (bppg). Despite an
important increase in the background currents was observed in the presence of CNTs, the
electrode demonstrated a good performance. A decrease of 300 mV in the oxidation
overvoltage for epinephrine and a significant increase in the associated peak current was
obtained with the CNT-bppg in comparison with the bppg electrode. Amperometric detection
of epinephrine performed with a rotating disk electrode at 0.25 V showed a linear range from
0.1 µM to 0.1 mM, with a detection limit of 0.02 µM. The electrode demonstrated to be
highly stable since after 20 min at 0.25 V in a 40 µM epinephrine solution the response
remained almost constant. Another interesting fact was the very good resolution of ascorbic
acid and epinephrine oxidation peaks, with a difference of 220 mV in the oxidation peak
potentials, at variance with the response at bare graphite electrode where just one wave was
obtained as a result of the overlapping of the two processes. The electrode showed very good
short and long term stability.
Homocysteine and Related Compounds
Wang et al. [64] proposed the use of CNTPE for the detection of homocysteine.
Voltammetric experiments of 160 µM homocysteine at CNTPE showed a well defined signal
at 0.28 V that reached a maximum at 0.64 V. This peak current depended linearly with the
square root of the scan rate. On the contrary, at CPE only a slight increase in the oxidation
current was obtained at 0.40 V, with no peak current definition. A linear relationship between
the voltammetric current and homocystein concentration at 0.64 V was observed between 20
and 180 µM, with a detection limit of 17.3 µM. Amperometric experiments were also
performed at a potential of 0.70 V and a linear relationship between steady-state currents and
homocystein concentration was obtained between 5 and 50 µM with a detection limit of 4.6
µM. The authors also evaluated the response of different thiolated compounds like cysteine,
glutathione and N-acetylcysteine and the results revealed that in all cases the oxidation at
CNTPE occurs at lower potentials than at the classical CPE with substantially higher currents.
Another electrochemical method for the sensitive determination of homocystein was
reported by Mao et al. [41]. They employed a CNT-nafion-modified GCE. Two processes
were observed at this electrode in the presence of 1.0 mM homocystein, one at 0.0 V and the
other at around 0.35 V. The first one was attributed to the catalytic oxidation of homocystein
mediated by the oxygen-containing molecules present at the oxidized CNTs and the other one
to the direct oxidation of homocystein at the CNT-modified GCEs facilitated by the CNTs.
The amperometric response at 0.0 V was very fast, reaching the steady state in 10 s, with a
detection limit of 6.0 x 10-2 µM. The flow injection response was more stable at the CNT-
Nafion-GCE than at GCE with a very small decrease of the signal after 50 min. Standars
deviations of 2.3 % were obtained after 20 injections of 0.80 µM homocystein. The response
was very reproducible, with negligible changes electrode-to-electrode, low charge currents
and good conductivity.
Carbohydrates
The direct oxidation of glucose in alkaline solutions by using MWCNTs attached to the
surface of a GCE by using conductive silver paint was also proposed [78]. The electrode, of
an effective area of 0.0474 cm2, was used without any pretreatment. A substantial decrease in
the oxidation overvoltage for glucose was observed in a highly alkaline medium,
demonstrating the catalytic activity of CNTs towards the oxidation of glucose. A linear
relationship was observed between the steady-state current at 0.20 V and glucose
concentration between 2.0 and 11.0 mM, with a sensitivity of 4.36 µAcm-2 mM-1 and a
detection limit of 1.0 µM. The sensitivity remained without changes even in the presence of
chloride, indicating that under these conditions there was not poisoning of the electrode. The
CNTs-modified GCE resulted very stable and useful as electrochemical detectors, although,
the selectivity needs to be improved for further application.
The use of MWCNT composite electrodes containing Cu as a detector for capillary
electrophoresis determination of carbohydrates compounds was also reported [66]. The
oxidation of sucrose, galactose and fructose at Cu-CNTPE started at potentials around 0.2 V
lower than at Cu electrodes allowing in this way a highly sensitive detection of different
sugars using NaOH solution pH 12.5 for the amperometric detection. Compared to Cu or
CNTs alone, this new material displayed a substantially greater promotion of the oxidation of
carbohydrates and, consequently, significatively higher sensitivity with detection limits of 20
µM glucose and 25 µM gluconic acid.
NADH
Wang et al. [50] have reported on the highly catalytic activity of a GCE coated with
MWCNTs towards NADH oxidation. A decrease of 490 mV in the oxidation overvoltage
compared to GCE was obtained, allowing the detection of NADH at low potentials. The
response was very stable, since after 60 min at 0.60 V the signal for 5 x 10-3 M decreased just
a 10 %. A fast response (8-10 s) was obtained under conditions that at bare GCE would have
been impossible to get.
Another interesting work proposed by Wang et al. [67] showed the advantages of CNTs-
SPE. Cyclic voltammograms for NADH among other compounds such as hydrogen peroxide,
potassium ferricyanide and catechol showed higher electrochemical activity at CNT-SPEs
than at SPEs, especially evident for the first two compounds.
Rubianes and Rivas [79] reported that the oxidation of NADH at CNTPE started at -
0.100 V, that is, 0.300 V less positive than at CPE due to the catalytic effect of carbon
nanotubes. The CNTPE demonstrated an effective short term-stability since even after 15 min
at 0.400 V, the oxidation signal of 1.0 x 10-5 M NADH decreased less than 20 %. In similar
experiments at CPE the signal decreased more than 80 %.
Aminoacids
Wang et al. [80] proposed the amperometric detection of non-electroactive aminoacids at

CNTs-GCE and at Ni-CNTs-GCE. The CNTs previously treated with nitric acid were
immobilized onto the GCE using a 1 % v/v Nafion as dispersing agent. The detection was
performed either in a stirred NaOH solution or in flow system (1.0 mL/min) by applying a
potential of 0.55 V. While no response was observed at unmodified GCE for several non
electroactive aminoacids like arginine, histidine, lysine, asparagine, methionine and
phenylalanine, an excellent response was obtained at GCE modified with CNTs at potentials
higher than 0.3 V. Figure 10 shows amperometric recordings at 0.55 V in 0.1 M NaOH for
successive additions of histidine (A), asparagines (B) and methionine (C) at unmodified (a)
and SWCNT-modified GCE (b). The advantages of the presence of CNTs are clear, since no
response is observed at the bare GCE, while a fast and well defined signal is obtained at the
SWCNT-modified GCE. Some differences were found using SWCNTs or MWCNTs and
even with MWCNTs prepared by ARC or CVD. CNTs prepared by CVD demonstrated to be
more active. At electrodes prepared with SWCNTs the oxidation started a lower potentials
compared to the ones prepared with MWCNTs. In the case of Ni-CNTs-GCE, the nickel was
deposited at -2.0 V for 3 min from a 5 mM nickel sulfate solution prepared in an acetate
buffer pH 4.5. Then the electrode was transferred to a 0.1 M NaOH solution and the potential
was cycled 20 times from -0.9 to 0.9 V at 100 mV/s to ensure complete formation of the
nickel hydroxide layer. The presence of nickel hydroxide improved even more the
amperometric response of different aminoacids like arginine, hystidine, lysine, asparagines,
methionine and phenylalanine by complex formation, with detection limits in the order of 10-5
M. An electrocatalytic process is produced at the Ni-CNTs as a consequence of the reduction
of the newly formed NiO(OH) in the presence of aminoacids.
Figure 10. Current–time response for successive 10 µM additions of histidine (A), asparagine (B), and
methionine (C) at unmodified (a) and SWCNT-modified (b) glassy carbon electrodes. Operating
potential: + 0.55 V; supporting electrolyte: sodium hydroxide (0.1M, pH 13); stirring rate: 300 rpm.
See reference 80.
Uric Acid
A selective response for uric acid was obtained at β-CD-CNT-GE depending on the
nature of the cyclodextrin used [73]. The oxidation current for uric acid at β-CD-CNT-GE
was several times higher than the corresponding at α-CD-CNT-GE, indicating that the
structure of β-CD facilitates the capture of uric acid. On the contrary, α-cyclodextrin interacts
better with ascorbic acid than with uric acid. Using a β-CD-CNT-GE it was possible to detect
ascorbic acid in the presence of uric acid due to the significant peak potentials separation
(about 360 mV). The resulting electrode was stable for 4 days and the reproducibility was 1.5
%. A linear relationship between peak current (from differential pulse voltammograms in 0.2
M acetate buffer pH 4.5) and uric acid concentration was obtained from 5 x 10-5 to 5 x 10-7 M
UA. The detection was 0.2 µM. The oxidation of CNTs did not show any advantage in uric
acid quantification.
Other Compounds
Liu et al. [68] proposed the fabrication and characterization of chemically assembled
SWCNTs on gold surfaces and reported the advantages of using this electrode to study the
electron transfer of some reactions. While no response for [Ru(NH3)6]Cl3 was observed at the
thiolated gold electrode, a response similar to that at bare gold was obtained in the presence
of CNT evidencing the important improvement in the electron transfer originated by the
presence of CNTs.
Hu et al. [58] reported a well defined peak at 0.68 V for 5.0 x 10-6 M indole-3-acetic acid
(an important hormone present in plants) in pH 2.0 phosphate buffer at GCE modified with
MWCNT dispersed in DHP [58]. The oxidation peak current of the indole-3-acetic acid
increased gradually with the amount of MWCNT-DHP dispersed at the GCE up to a volume
of 15 µL. For higher volumes the thickness of the layer blocked the electron transfer. The
oxidation peak current presented a linear relationship with the concentration of the hormone
from 1 x 10-7 M to 5 x 10-5 M, with a detection limit of 2 x 10-8 M and a reproducibility of 4.3
% for 36 measurements of 5 x 10-6 M. The authors extended the use of this sensor to the
determination of the hormone in gladiola, apple and phoenix leaves showing a very good
agreement with HPLC determinations.
The detection of the anthracycline daunomycin by accumulation at MWCNTs-modified
GCE was also reported [81]. Daunomycin was accumulated at open circuit for 3 min and then
it was reduced by differential pulse voltammetry from -0.10 to -0.90 V. The reduction peak
current at -0.526 V was linear with the concentration of daunomycin in the range from 2 x 10-
8
to 1 x 10-5 M, the detection limit being 8 x 10-9 M. The relative standard deviation for 2 x
10-7 M daunomycin was about 6%. The method was successfully used for the determination
of the analyte in urine samples of cancer patients.
The MWNTs-GCE was also used to study the electrochemical behavior of brucine by
cyclic voltammetry and square wave voltammetry [82]. The current for brucine at the
modified electrode increased linearly with the concentration in the range between 1 x 10-6 to 1
x 10-4 M with a detection limit of 2.0 10-7 M.
The adsorption of molecular oxygen on CNTs was studied by Collins et al [83] and they
found that the electrical properties of CNTs are very sensitive to oxygen adsorption. Kong et
al proposed the use of nanotubes for developing miniaturized sensors for the detection of gas
molecules at room temperature [84]. The mechanism for the NO2 was a physisorption close to
chemisorption, the oxidizing NO2 molecule takes 1/10 of an electron charge from the
nanotube, increasing the hole carriers and enhancing the conductance for the p-type
nanotubes. In the case of NH3, there is a physisorption, the Lewis base donates a small
amount of electrons to nanotubes and reduce the hole-carriers, decreasing the conductance
[84].
USE OF CNTS FOR THE DEVELOPMENT OF

ELECTROCHEMICAL BIOSENSORS
A biosensor is basically a chemical sensor with two main components, a biorecognition

layer encharged of the biomolecular recognition of the analyte and a transducer that is
responsible for the conversion of the biorecognition event into an useful electrical signal [85].
Enzymes, antigens, antibodies, nucleic acids, receptors and tissues have been used as
biorecognition elements. According to this element, it is possible to separate in enzymatic
(involving a biocatalytic event) and affinity (involving an affinity event) biosensors [86].
Enzymatic biosensors are connected with the use of enzymes as biorecognition element,
while affinity biosensors involve the use of nucleic acids, antibodies, antigens or receptors.
Concerning transducers, they can be optical in their different modes, piezoelectric, thermal
and electrochemical. In the case of electrochemical biosensors, the electrical signal obtained
as a consequence of the interaction analyte/biosensing layer can be displayed as a given signal
depending on the electrochemical transduction mode. The nature of the electrode is very
important for the transduction process. In this sense, CNTs represent an important alternative
for the transduction event due to their excellent electronic properties [10].
Enzymatic Electrochemical Biosensors
The first enzymatic electrode was proposed by Clark and Lyons more than 40 years ago
[87]. Since then, electrochemical biosensors based on the use of enzymes as biorecognition
layer have received considerable attention due to the advantages they possesses as a result of
the very efficient combination of the biocatalytic activity of enzymes with the highly sensitive
electrochemical transduction.
Several strategies for immobilizing proteins on CNTs modified electrodes have been
proposed. The step of immobilization is critical, since the enzyme has to remain as much
active as possible in order to perform an efficient biorecognition of the substrate. The other
aspect to consider is that the transducer where the protein will be immobilized has to allow a
fast charge transfer to ensure a rapid and sensitive response. Therefore, it is important to take
into account that the noncovalent functionalization of the sidewalls of SWCNTs is the best
way to preserve the sp2 nanotube structure and their electronic characteristics.
Immobilization of Proteins on Carbon Nanotubes

A very interesting discussion about the adsorption of proteins on CNTs was reported by
Sun et al. [88]. The authors used ferritin as a model and evaluated the natural affinity of
purified SWCNTs towards this protein. CNTs were dispersed first in phosphate buffer
solution pH 6-7 by sonication for 1 h. The protein was then added to the solution and allowed
to interact for 24 h while stirring. The authors suggest that the presence of defects on the
surface of CNTs may play a very important role in the interaction with ferritin and that the
nonspecific interactions between the protein and CNTs include hydrogen bonding and
electrostatic and hydrophobic interactions. One important aspect to consider for the
development of biosensors is that the functionalization of CNTs with hydrophilic polymers or
with oligomeric polyethylenglycol moieties made the protein adsorption more difficult. The
authors also discussed about the affinity of SWCNTs by amine groups. Considering that
proteins are very rich in these groups, they could play a very important role in the interaction
protein-SWCNT. When the interaction is performed in the presence of carbodiimide the

immobilization of ferritin became more favorable.
Davis et al. [89] reported an important work regarding the immobilization of
metalloproteins and enzymes on oxidized, purified and vacuum-annealed SWCNTs in
aqueous solution. AFM experiments showed that the immobilization is mainly physical,
without need for covalent activation or electrostatic interaction. In fact, cytochrome c at pHs
below the isoelectric point and ferritin at pHs above the isoelectric point showed an important
adsorption obtained just by stirring the nanotubes dispersion (0.03 mg/mL) in dilute protein
solutions (50-100 µg/mL) for a given time (2-20 h). GOx could be also adsorbed in a very
efficient way, as the picture showed in Figure 11 indicates. The enzyme immobilized
remained active since in the presence of 0.5 mM ferrocenemonocarboxyic acid and glucose,
an important catalytic current was obtained (Figure 12).
Figure 11. AFM image of a glucose oxidase-modified SWCNT. See reference 89.
Dai et al. [90] reported a very interesting work introducing a simple and general approach
for noncovalent functionalization of the sidewalls of CNTs for further immobilization of
ferritin, streptavidin and biotinyl-3,6-dioxaoctanediamine in a very efficient way. The first
step was the noncovalent functionalization of SWCNTs by irreversible adsorption of a
bifunctional molecule, 1-pyrenebutanoic acid, succinimidyl ester onto the hydrophobic
surfaces of SWCNTs dispersed in DMF or methanol.
This molecule interacts in a very stable way in aqueous solution through the aromatic
rings with the basal plane of graphite via π-stacking with the sidewalls of SWCNTs. The
succinimidyl residues are highly reactive to nucleophilic substitution by primary and
seconday amines of proteins or other molecules. SWCNTs were incubated in a
pyrenebutanoic acid, succinimidyl ester solution (6 mM in DMF or 1 mM in methanol) for 1
h at room temperature followed by careful rinsing in pure DMF or methanol. The proteins
were then immobilized by incubation in aqueous solution for 18 hours at room temperature,
rinsed thoroughly in pure water for 6 hours and then dried.
Figure 12. Voltammetric response of a GOx-SWCNT-modified glassy carbon electrode in the absence
(red) and presence (blue) of 0.5 mM ferrocene monocarboxylic acid. The catalytic response (green)
after the addition of 50 mM glucose is also shown. See reference 89.
The enzymatic activity of -chymotrypsin was evaluated in composites of poly(methyl

metacrilate) with different carbon materials [91] demonstrating that the incorporation of
SWCNTs into enzyme-polymer composites results in active and stable polymeric films. The
release of the protein from the composite was evaluated measuring the enzymatic activity in
the supernatant in contact with the composite. The results showed that in the case of
SWCNTs the leaching of the protein from the composite was lower. This fact was attributed
to the union of the protein to the CNTs. The effect of other polymers such as polystyrene and
poly(lactic acid) was also analyzed and the leaching of the protein was significant in the
absence of SWCNTs. Only the hydrophobic ones (poly(methyl metacrilate) and polystyrene)
promoted the protein adsorption. The increase of CNTs percentage in the composite also
produces higher -chymotrypsin retention.
Figure 13. 1-Pyrenebutanoic Acid, Succinimidyl Ester 1 irreversibly adsorbed onto the sidewall of a
SWCNT. See reference 90.
Figure 14. Scheme of the different steps involved in the fabrication of aligned shortened SWCNT
arrays for direct electron transfer of enzymes such as microperoxidase MP-11. See reference 92.
Gooding et al. [92] presented a strategy for studying the electron transfer properties of
redox enzymes like microperoxidase 11 attached to the end of aligned SWCNTs. A
polycrystalline gold electrode previously cleaned in 0.05 M sulfuric acid was derivatized with
cysteamine by interacting for 5 hours with a 1 mM cysteamine ethanolic solution.
Subsequently, this modified-gold surface was immersed for 4 hours in a dispersion of

oxidatively shortened SWCNTs in 1 mL of DMF containing 0.5 mg of
dicyclohexylcarbodiimide to convert the carboxyl group located at the end of shortened CNTs
into active carbodiimide esters. The SWCNTs were aligned normal to the electrode surface.
Then, microperoxidase was attached to the free ends of the tubes by incubation in a 0.5
mg/mL microperoxidase in HEPES solution pH 7.5 at 4oC overnight. The schematic
representation of the protocol is displayed in Figure 14.
Since the iron center of the protein was not shielding, the electron transfer occurred and
cyclic voltammograms showed a voltammogram with a redox couple with E1/2 at -390 mV.
The coverage obtained from the reduction peak area was 35 pmolcm-2, well correlated with
the the value of 32 pmolcm-2 estimated from the AFM images. In summary, the SWCNT
normal aligned can act as molecular wires to allow the electrical communication between the
electrode and redox proteins covalently attached to the ends of SWCNTs.
Fructose Biosensor
Magno et al. [65] reported an amperometric biosensor for fructose using an electrode
prepared by dispersion of MWCNT within mineral oil (60/40 % w/w) and covered by a
polymer obtained from the electropolimerization of dihydroxybenzaldehyde. The fructose
dehydrogenase was immobilized on different membranes placed on the top of CNTPEs and
then covered with an additional polycarbonate membrane (0.03 µm pore size) to prevent
fouling and microbial attack.
Glucose Biosensor
Glucose biosensors have received a lot of attention due to the importance that the fast,
sensitive and selective glucose determination presents in the diagnostic and control of
diabetes, one of the most important diseases of this century. Different strategies involving
CNTs have been proposed for the development of glucose biosensors, and the most
representative are included below.
The suitability of CNTPE for developing highly sensitive glucose enzymatic biosensors
by incorporation of glucose oxidase (GOx) within the composite matrix was illustrated by
Rubianes and Rivas [46]. The resulting enzymatic electrode allowed the highly sensitive and
selective determination of glucose even without redox mediators, metals or anti-interferents
layers due to the important electrocatalytic effect of carbon nanotubes on the reduction of
hydrogen peroxide. Figure 15A shows the amperometric recordings obtained at -0.100 V at
CPE-GOx (a) and CNTPE-GOx (b) for successive additions of 2.0 mM glucose. Almost no
response is observed at the graphite composite electrode. On the contrary, a fast and sensitive
response was observed at CNTPE-GOx due to the catalytic activity of CNTs towards
hydrogen peroxide. Figure 15B shows the corresponding calibration plots. The sensitivity
obtained with CNTPE-GOx was 43 times higher than that obtained with the CPE-GOx. A
linear range was obtained from 2.0 mM to 25.0 mM glucose. A negligible interference was
observed even for large excess of ascorbic acid, uric acid and acetaminophen.
Glucose oxidase [55] was also immobilized at a GCE modified with CNTs dispersed in
Nafion by dipping the electrode in a solution containing 19.5 mg/mL GOx and 5 mg/mL
glutaraldehyde for 4 hours at room temperature, followed by 6 immersions in 0.5 % v/v
Nafion solution for 1 hour and a final one in a 5.0 % v/v Nafion solution for 1 hour. The
important electrocatalytic activity of this CNTs towards the oxidation and reduction of
hydrogen peroxide allowed the very sensitive glucose quantification at -0.050 V where no
interference of ascorbic acid, uric acid and acetaminophen was found. The linear range was
between 2 and 20 x 10-3 M glucose.
GOx was also immobilized on Pt modified with chemically oxidized SWCNTs through
covalent attachment using EDC [93]. The amperometric experiments at 0.40 V gave a
sensitivity of 18.7 mAM-1cm-2, a linear range up to 12 mM glucose and an apparent Km of
13.1 mM. The biosensor showed a good stability, keeping a 90 % of the activity after 4
months.
Dekker et al. [90] demonstrated that semiconduncting SWCNTs can be used for the
development of biosensors. GOx was immobilized on SWCNTs by using a linking molecule,
which binds on one side to the SWCNTs through Van der Waals interaction and on the other
side to the enzyme through an amide bond. To do so SWCNTs deposited on a silicon wafer
were left in 2.3 mg/mL 1-pyrenebutanoic acid succinimidyl ester in DMF for 2 h while
stirring, washed with clean DMF, left in 10 mg/mL GOx for 18 h, and washed in pure water
for 6 hours. From AFM images they concluded that GOx is inmobilized specifically on the
modified SWCNTs. The conductance of the semiconducting SWCNT was measured as a
function of the liquid-gate voltage. The attachment of GOx significantly decreased the
conductance of the semiconducting SWCNT as a result of the change in capacitance of the
tube because GOx immobilized on the surface of a SWCNT inhibits the movement of ions
close to the tube. GOx-coated semiconducting SWNTs show strong pH dependence,
indicating the possibility to use these sensors to measure pH changes down to 0.1 units. After
addition of glucose the conductance increases. Therefore, it can be used as an excellent
nanosize pH sensor and glucose.
Wang et al. [94] reported the use of CNTs-composite for continuous measurement of
glucose with excellent selectivity, high sensitivity, wide linear range, fast response, long-term
and thermal stability and oxygen independence. The glucose microsensor was prepared by
dispersing GOx directly within CNTs and graphite and packing the resulting biocomposite
into a 21-gauge needle. The CNT-GOx biocomposite was prepared by mixing 2.5 mg CNTs
and 7.5 mg graphite powder for 5 min, followed by the addition of 2 mg GOx and mixing for
additional 5 min. The resulting mixture was packed into a 300-μm polyimide tubing that was
inserted into a 21-gauge needle. Once the surface was smoothed, it was coated with 1 %
Nafion solution by three 10 s dipping. The CNTs demonstrated to have an important effect
upon the sensing behavior. In fact, the response increased rapidly up to 25 % w/w CNT and
then more slowly. The selected biocomposite composition was 21% w/w CNT, 17% w/w
GOx and 62% w/w graphite. The thickness of the Nafion layer demonstrated to have an
important effect on the sensitivity, a thicker Nafion film was accompanied with lower
sensitivity and wider linear range. After 80 days the response decreased just 20 %. To assess
the CNT-induced enzyme stabilization, the thermal stability was examined during storage at
90º C. It was found that when GOx was incorporated within CNTs, the thermoresistance
dramatically increased. In fact, the response decreased just 20 % and 25 % after 3 and 24
hours at 90 oC, respectively. Another interesting advantage is that packing the CNT/GOx
biocomposite within the needle facilitated the monitoring of glucose under severe oxygen
deprivation.
Figure 15. (A) Amperometric recordings obtained at CPE-GOx (a) and at CNTPE-GOx (b) for
successive additions of 5 mM glucose. The content of GOx was 10.0% w/w in both electrodes. (B)
Calibration plot obtained from amperometric recordings for successive additions of 2 mM glucose.
Working potential: - 0.100 V. Supporting electrolyte: 0.050 M phosphate buffer solution, pH 7.40. see
reference 86.
The effect of the incorporation of Pt nanoparticles in SWCNTs-Nafion film on GC and

on a carbon fiber electrodes (CFE) for developing a sensible glucose biosensor was reported
by Luong et al. [95]. GCE was polished, electrochemically activated and then modified with
an aliquot of a solution containing the CNTs (2 mg SWCNTs in a mixture of 100 µL of
Nafion and 900 µL of Pt nanoparticles). Platinum nanoparticles were in electrical contact
with GCE and CFE through the SWCNTs, enabling the composite structure to be used as an
electrode. The enzymatic electrode was prepared by dropping 3 µL GOx solution in 50 mM
phosphate buffer solution pH 7.2 (20 mg/mL). Subsequently, glutaraldehyde (3.0 uL 2.5 %)
was applied on the resulting electrode for the cross-linking. Under these conditions the
detection limit was 0.5 µM and the response time was 3 s. A linear range was observed from
0.5 µM to 5mM glucose with a sensitivity of 2.11 µAmM-1. A similar response was observed
using CFE as substrate. Maximum levels of AA, UA and acetaminophen did not show any
interference.
Another glucose enzymatic electrode was prepared by covalent attachment of glucose
dehydrogenase (GDH) to the CNT-CHIT films using glutaraldehyde (GDI) to cross-link the
enzyme [56]. Due to the electrocatalytic properties of CNTs, NADH could be oxidized at
potentials 0.300 V less positive than at the GCE, allowing a very sensitive and selective
determination of glucose and a linear range up to 300 μM with a sensitivity of 80 mA M-1 (in
0.05 M phosphate buffer solutions pH 7.4). The performance of the bioelectrode has allowed
the successful determination of glucose in urine.
Palleschi et al [96] presented a Prussian Blue (PB) modified with SWCNTs as an
efficient platform for developing enzymatic electrodes. The PB was synthesized in the
presence of SWCNTs, starting from K3Fe(CN)6 and FeCl3. The paste was prepared 60/40
w/w CNTs/mineral oil by hand mixing in a mortar and packed in a Teflon tube. Voltammetric
and amperometric parameters were compared between PB-graphite and PB-CNTs. PB-
CNTPEs showed a slower kinetics and did not improve the analytical performance of the
sensors towards hydrogen peroxide. PB-CNTPEs showed a linear range for glucose between
0.1 and 50 mM glucose. The advantage of the PB-CNTPE was the stability at very basic pH,
attributed to the peculiar structure of SWCNTs.
The authors investigated the use of aligned carbon nanotubes as platform for the
production of a conducting polymer-glucose oxidase based biosensor [97]. The aligned CNT
films were prepared by pyrolizing Fe(II)-phthalocyanine under Ar/H2 at 900ºC. GOx
immobilization was performed during pyrrole polymerization in a solution containing NaClO4
and GOx. The resulting material showed excellent electrocatatalytic properties towards the
oxidation of hydrogen peroxide. SEM pictures showed that the polymer was not only located
in the intertube spacing, but also on the top of the mat.
Wallace et al. [98] developed a new material consisting of an aligned, highly orientated
carbon nanotubes array in three dimensions, coated by a layer of polypyrrole, which allowed
the immobilization of enzymes, during its polymerization onto the nanotubes array. The
immobilization of GOx was performed by electrochemical oxidation of pyrrole (0.10 M) in a
buffer solution pH 7.45, containing 2 mg/mL GOx and 0.10 M NaClO4 at 1 V for 1 min at 10
ºC. The presence of the enzymes was confirmed by the increase in the N/C and O/C ration in
the XPS spectra of the Ppy/CNTs array before and after the GOx immobilization. The glucose
response of this bioelectrode was 10-20 times higher than that for a corresponding flat gold
electrode. The 3-D structure of CNTs provides a good template for a large enzyme loading in
an ultra thin polymer layer (<10nm).
In another work, MWCNTs were vertically adhered to a gold film and the glucose
biosensor was obtained by immersing in a 0.1 M phosphate buffer solution containing GOx to
obtain a Au-MWNTs-GOx [99]. Figure 16 shows an scheme of the biosensor. The MWCNT-
based biosensor exhibited a fast and sensitive response. The effect of opening the tubes on the
response of the biosensor was also evaluated. The nanotube caps were opened by treating
them with a mixture of HNO3 and HF. Once the tubes were opened, GOx could enter into the
hollow of MWNTs, increasing in this way the amount of enzyme immobilized. On the other
hand, when the MWCNTs were treated with the acidic solution the generated carboxylic
groups give a hydrophilic environment that allows the adsorption and insertion of the enzyme
into the cavity of the CNTs while preserving its functionality. As a consequence of that, the
stability of the biosensor largely increased. In fact, after 4 months storage at 4oC, the activity
remained in a 86.7 % of the original value.
Willner‘s group proposed an interesting approach for the construction of a glucose
biosensor based on the use of CNTs [100]. Oxidized SWCNTs were covalently attached to a
thiolated gold surface through the generated carboxylic functions. The other end of the
SWCNT was covalently attached to FAD. Therefore, when the apo-GOx was immobilized, it
could be reconstituted on the edge of SWCNTs, and in this way the electrons could be
transported long distances.The length of the CNTs was the responsible for controlling the rate
of the process.
Figure 16. Schematic illustration of the MWNT-based biosensor for glucose detection. See reference
99.
Lactate Biosensor
Rubianes and Rivas [79] proposed the immobilization of lactate oxidase (LOx) into the
CNTPE. The amperometric response to lactate was linear up to 7.0 x 10-3 M with a detection
limit of 3.0 x 10-4 M.
The increase in the activity of lactate dehydrogenase (LDH) on a GCE modified with a
SWNTs film, compared to a bare GCE was also proposed [101], by using
chronoamperometry in a convective system. The LDH-SWNT-modified GCE was more
active than the plain GCE. The irreversible catalytic oxidation of NADH was demonstrated
by the 150 mV shifting of the anodic peak potential towards more negative values, compared
to the plain GCE. The authors evaluated the SWNTs-modified electrode performance in the
reduction of pyruvate using NADH as coenzyme in 50 mM Tris-HCl buffer solution pH 7.5,
by chronoamperometry in a forced convection system.
Phenols and Catechols Biosensor

Since the product of the oxidation of phenols and catechols catalyzed by polyphenol
oxidase (PPO) is the corresponding quinone, Rubianes and Rivas evaluated the influence of
MWCNTs on the electrochemical behavior of hydroquinone at carbon composite electrodes
with different content of CNTs [79]. It was found that as the percentage of MWCNTs
increases, the response becomes more reversible, decreasing the peak potential separation and
increasing the associated currents. Consequently, the response for dopamine was 12 times
more sensitive at CNTPE-PPO than at CPE-PPO and the detection limits for dopamine and
phenol were 1.0 x 10-6 M and 1.0 x 10-7 M, respectively. The study of the electrochemical
behavior as well as the application of the resulting biolectrodes for the determination of
phenols, catechols and alcohols in real samples was also illustrated.
Hydrogen Peroxide Biosensor

Shi et al [102] have studied the catalytic activity of MWCNTs-horseradish
peroxidase(HRP)-GCE towards hydrogen peroxide. Three microliters of a solution of 2.0
U/mL HRP and 0.5 % w/w BSA in PBS were dropped on the MWCNTs-GCE followed by
cross-linking in a closed vessel contained 25 % glutaraldehyde and water vapor for 20 min
and dried at room temperature for 1 h. The presence of CNTs had allowed the sensitive and
selective determination of hydrogen peroxide. The latter immobilization of oxidases like GOx
and LOx made it possible the detection of glucose and lactate from the hydrogen peroxide
enzymatically generated. The working potential was -300 mV and the flow rate of 2.0
µL/min.
Zhao et al. [103] proposed the immobilization of myoglobin on the surface of MWCNT-
modified GCE. They found that the protein immobilized can catalyze the reduction of
hydrogen peroxide. The pH selected for the analysis was 4.0 and under these conditions, the
response of the biosensor was very fast, reaching the 90% in 6 s. The relationship between
current and hydrogen peroxide concentration was linear up to 330 µM, the reproducibility
was 5.9 % and the detection limit was 4.2 µM under anaerobic conditions. The voltammetric
response did not change significantly after 1 month at 4 oC or 1 week stored in air.
Zhao et al. [104] reported the electrochemical behavior of myoglobin on MWCNT-GCE

and the potential application for the development of a nitric oxide biosensor. Myoglobin was
immobilized on the acid-treated-MWCNTs-GCE by dipping the electrode into 0.24 mM
myoglobin solution (in acetate buffer pH 5.6) over 72 hours. After that, the electrode was
removed, washed with water and stored at 4 oC. Electrochemical impedance spectroscopy
experiments demonstrated that the charge transfer resistance for a redox marker couple
(potassium ferro/ferricyanide) increased for electrodes prepared by longer immersion times of
the protein. Cyclic voltammetry of the resulting electrode containing myoglobin showed a
couple of reversible peaks due to the reduction of the Fe(III)-myoglobin and the reoxidation
in the reverse scan. In the presence of NO, a new cathodic peak appears at around -0.8 V in
phosphate buffer pH 7.0 due to the reduction of NO that could be assisted by the electroactive
myoglobin. Amperometric experiments performed at -0.8 V show a linear relationship
between steady-state current and NO concentration up to 18 µM. Zhao et al [105] have also
proposed the direct electron transfer between the strongly adsorbed cytochrome c and
MWCNTs-GCE. Based on these results, it was possible to detect hydrogen peroxide in the
range 2-420 µM and a detection limit of 1.02 µM.
Another work [106] reported the covalent attachment of enzymes onto the ends of
vertically oriented SWCNTs. The ends of orthogonally arrays of shortened SWCNTs can be
linked to electrochemically active heme proteins through traditional bioconjugate chemistry.
Proteins were attached to the end of SWCNTs by using the known chemistry of EDC to
promote amide linkages between carboxyl nanotubes and lysine residues of the proteins. The
authors showed that myoglobin (Mb) and horseradish peroxidase (HRP) covalently linked to
SWNT exhibited quasi-reversible FeIII/FeII voltammetry and sensitive response to H2O2.
Monolayers of vertically aligned, shortened SWNTs were assembled on ordinary pyrolitic
graphite electrodes from DMF dispersions onto an underlying composite bed of Nafion
ionomer and Fe3+ precipitated hydroxide. Voltammetric peaks of the enzymes attached onto
SWCNTs forests were stable, and did not decay during repetitive multiple scans. This
behavior is congruent with covalent attachment of proteins to the carboxylate-bearing ends of
SWNTs. The nanotubes forest behaves electrically similar to a metal, conducting electrons
from the external circuit to the enzymes. The oriented SWCNTs forests have the potential of
being fabricated as ultamicroelectrodes on the nanometer scale, offering the future possibility
of multielement nonobiosensor arrays. Detection limits of 70 nM and 50 nM for hydrogen
peroxide were found at Mb- and HRP-modified electrodes.
A new electrode material based on the use of CNTs for studying the direct electron
transfer of hemoglobin [107] was also proposed. The electrode was fabricated in the
following way, a 100 µm diameter Pt microelectrode was first chemically etched to obtain a
cavity of tens of µm depth and then ground the etched tip on a flat plate with CNTs until the
microcavity was filled. In the case of the material containing hemoglobin, the CNT powder
was mixed with 100 µL 0.1M acetate buffer pH 5.4 containing 50 mg/mL hemoglobin and
0.1 M KCl and then dried at 1oC under N2 flow. In the presence of hydrogen peroxide a well
defined peak appeared between -0.5 and -0.8 V, indicating the catalytic activity of the
hemoglobin immobilized. Therefore, it was possible to perform the detection of hydrogen
peroxide at -0.8 V. The response was fast, with stable value after 7 seconds. The current was
linear with the hydrogen peroxide concentration from 2.1 x 10-4M to 9 x 10-4 M and a
detection limit was 9 x 10-6 M.
Alcohol Biosensor
[59] A composite biosensor was prepared by adding the desired amount of alcohol
dehydrogenase (ADH) and NAD+ cofactor to a 50/50 % w/w MWCNT/Teflon composite.
The electrocatalytic properties of MWCNT towards NADH and the biocatalytic properties of
ADH are not impaired because of the presence of Teflon. A marked decrease in the
overvoltage for the oxidation of the liberated NADH was shown.
Rubianes and Rivas also proposed the immobilization of ADH (12.0 % w/w) into a
CNTPE in the presence of NAD+ (12.0 % w/w) [79]. Based on the excellent electrocatalytic
properties of MWCNTs towards the oxidation of NADH, a very fast and sensitive response
for ethanol was obtained at CPE-ADH-NAD+.
Electrochemical DNA Biosensors
Electrochemical DNA biosensors are based on the use of nucleic acids or analogues as
biorecognition element and electrochemical techniques for the transduction of the physical
chemical signal. Two aspects are essential in the development of hybridization biosensors,
sensitivity and selectivity. Traditional methods for detecting the hybridization event are too
slow and require special preparation. Therefore, there is an enormous interest in developing
new hybridization biosensors, and the electrochemical represent a very good alternative
[108].
An electrochemical DNA hybridization biosensor basically consists of an electrode
modified with a single stranded DNA called probe [109]. Usually the probes are short
oligonucleotides (or analogues such as peptide nucleic acids). The first and most critical step
in the preparation of an electrochemical DNA biosensor is the immobilization of the probe
sequence on the electrode. The second step is the hybrid formation under selected conditions
of pH, ionic strength and temperature. The next step involves the detection of the double helix
formation by using a methodology able to obtain an electrical signal that clearly demonstrates
the specific recognition of the complementary DNA among others.
Another interesting aspect of sensors containing DNA as biorecognition layer is the
detection of chemical and physical damage produced on DNA. In this case, it is necessary to
immobilize double stranded DNA at the electrode surface to build the probe. The second step
consist of the interaction of the immobilized DNA layer with the given damage agent under
controlled conditions and the last step is the transduction of the signal, either from the
nucleobases signal or from the redox signal of the damage agent [109].
Pedano and Rivas [110] reported the adsorption and electrooxidation of oligo and
polinucleotides at a CNTPE. Figure 17 shows the chronopotentiometric signals obtained in
acetate buffer solution (0.200 M pH 5.00) after 5 min accumulation at 0.200 V in 5.0 ppm
ssDNA (A) and 2.00 ppm oligonucleotide (21 bases) (B) solutions at untreated CPE (A,a:
B,a) and untreated CNTPE (A,b; B,b). Almost no signal was obtained at the classical graphite
paste electrode. On the contrary, a very well-defined signal at 1.06 V was obtained at the
CNT-based composite (29 and 61 times larger for the ssDNA and the oligonucleotide,
respectively), demonstrating the advantages of this material. No shifting in the peak potential
was obtained, indicating that the main effect of CNTs occurred in the adsorption of the
nucleic acids. The state of the surface demonstrated is very important for the adsorption and
further electrooxidation of nucleic acids. A pretreatment of 1.3 V for 20 seconds in a 0.200 M
acetate buffer solution pH 5.0 demonstrated to be the most effective. An interaction mainly
hydrophobic was reported for the interaction DNA-CNTPE.
Figure 17. Chronopotentiometric signals obtained in a 0.200 M acetate buffer solution pH 5.00 after 5
min accumulation at 0.200 V in a 0.200 M acetate buffer solution pH 5.00 containing 5.0 mg/L ssDNA
(A) and 2.00 mg/L oligoX (B) at CPE (60.0% w/w graphite powder and 40.0% w/w mineral oil) (A, a;
B, a), and at untreated CNTPE (60.0% w/w carbon nanotubes and 40.0% w/w mineral oil) (A, b; B, b).
Stripping current: 8.00 µA. Initial potential: 0.500 V. see reference 110.
Hamers et al. [111] reported the covalent attachment of DNA to CNTs previously
oxidized and derivatized with amino residues through a linker that promotes the attachment of
nucleic acids and a more efficient dispersion of CNTs in solution.
The immobilization of calf-thymus DNA molecules on multi-walled CNTs activated with
EDC and NHS by immersion in a phosphate buffer solution containing 2 mg/mL of ssDNA
for several hours was also reported [42]. Cyclic voltammograms showed that the gold
electrode containing NTs present a large background current, apparently capacitive. In the
presence of DNA, the peaks due to the carboxylic groups on CNTs disappeared and the
background current drastically decreased. The authors also performed impedance experiments
to demonstrate the presence of the nucleic acid at the electrode surface. The measurements
were performed at open circuit using 10 mM potassium ferricyanide/ferrocyanide plus 0.1 M
NaCl, for frequency ranges between 0.1 Hz to 100 kHz and amplitude of 5 mV. The presence
of the DNA layer (either single or double stranded) made more difficult the access of the
redox couple to the electrode due to the charge repulsion, therefore, decreasing the currents,
and increasing the peak potential separation and charge transfer resistance (Figures 18 and
19). EDC/NHS demonstrated to be necessary to obtain an effective immobilization of DNA,
three hours interaction being the optimum interaction time. For instance, the changes in the
charge transfer resistances were very small when the NA was immobilized in the absence of
EDC and NHS. The electrode was also used for studying the interaction with ethidium
bromide (EB), a known intercalator of DNA. Therefore, the immobilized DNA keeps their
recognition properties and the EB effectively can intercalate. The impedance experiments
showed an increase in the resistance from 16.02 to 56.19 kΩ in the absence and after 18 min
EB interaction, respectively.
Immunosensing Schemes
Immunosensors are based on high affinity reactions antigen/antibody. Several strategies

can be used to immobilize the recognition element, either the antigen or the antibody,
depending on the protocol. The detection of the recognition event uses the same principle as
the enzymatic immunoassay. In general, an enzyme is coupled to the recognition layer (the
antigen or antibody) and, to quantify the antigen/antibody interaction, the enzymatic reaction
is developed in the presence of the substrate and electrochemically detected [118].
An amperometric biosensor based on the adsorption of antibodies onto perpendicularly
oriented assemblies of single wall CNTs called forest was proposed [119]. The SWCNTs
were functionalized with carboxyl residues and shortened by sonication in a mixture of
sulfuric and nitric acids for 4 hours at 70 oC. The graphite electrode was dipped in 1mg/mL
Nafion solution for 15 min and then immersed in a 5 mg/mL FeCl3.6H2O for 15 min. After
washing, the electrode was placed in a sonicated suspension of SWCNTs in DMF for 30 min,
washed with methanol and dried. The immobilization of the anti-biotin antibody was
performed by incubation for 3 hours on the surface of SWCNTs. After adequate washing, the
electrode was blocked with 2 % bovine serum albumine in PBS. The detection of horseradish
peroxidase bound to biotin was evaluated by using a rotating disk electrode at -0.3 V, 2000
rpm with 1 mM hydroquinone and 400 µM hydrogen peroxide. The detection limit was 2.5
nM and the linear range was up to 25 nM. Unlabelled biotin was detected in a competitive
approach with a detection limit of 16 µM. The platform could be stored in a humidic aerobic
chamber at 4oC for one week without significant changes (16.7 vs 17.3 µA).
Tzeng et al. [121] reported a work dealing with the study of the influence of different
chemical treatments of CNTs vertically aligned on a Si substrate on the adsorption of
antibodies. Further application for the interaction with bacteria was also proposed.
Bianco et al. [122] presented the functionalization of CNTs with two polypeptides
employing two different strategies. In both cases, the CNTs were previously modified to
expose amine groups. In the first methodology, a model peptide was condensate on the CNTs
surface through a liker in DMF. In the second case, the peptide was the B-cell epitope from
the foot and mouse disease virus (FMVD). The immobilization was performed employing
succinimide. 1H NMR experiments showed that in both cases the peptides keep its
conformational structure. Immunological assays coupled to Surface Plasmon Resonance
(SPR) methodology showed that the adsorbed peptide was recognized by its polyclonal
antibodies being this an indication that the adsorption process did not produce significant
conformational changes in the secondary structure of the peptide. Preliminary experiments of
immnunogenicity employing FMDV-CNTs indicated that the conjugate produces strong anti-
peptide antibody immunization in mice. This peptide modification could be applied for
immunologic test offering several advantages such as higher accessibility to the peptide and
the possibility to adsorb different epitopes, allowing in this way higher diagnostic accuracy.
From reference 42.
Figure 18. (A) Cyclic voltammograms for MWCNTs-modified gold electrode treated with 2 mg/mL
dsDNA with EDC/NHS activation at different times in the presence of 10 mM K4Fe(CN)6 and 10 mM
K3Fe(CN)6 containing 0.1 M NaCl. (a) 0 h; (b) 0.8 h; (c) 1.8 h; (d) 2.8 h; (e) 4.8 h. (B) Changes of the
anodic peak current of Fe(CN)6-3 / Fe(CN)6-4 redox couple at MWCNTs-modified electrode

immobilized with dsDNA (■) and ssDNA (●) for different time intervals.
From reference 42.
Figure 19. Nyquist plots (Zim vs. Zre) for different electrodes in 10 mM K4Fe(CN)6 +10 mM K3Fe(CN)6
+ 0.1 M NaCl aqueous solutions. (a) MWCNTs-modified gold electrode; (b,c) MWCNTs-modified
gold electrode treated with 2 mg/mL dsDNA with and without EDC/NHS activation, respectively; (d,e)
MWNTs-modified gold electrode treated with 2 mg/mL ssDNA with and without EDC/NHS activation,
respectively.
Wang et al. [112] reported a significantly higher sensitivity for DNA oxidation at CNTs-
GCE compared to GCE, although the oxidation occurs at more elevated potentials, indicating
that CNTs promotes the interfacial accumulation more than the electron transfer. After 3-min
accumulation, the guanine oxidation signal was 17-fold higher than that obtained at the bare
GCE. The CNT-GCE was also used as a detector for the hybridization event performed using
the two-surface hybridization scheme. In this way, the selected biotinylated probe was
immobilized at the magnetic beads covered with streptavidin, followed by the hybrid
formation and transduction. The analytical signal was obtained from the guanine residues
after digestion. The presence of CNTs allowed an efficient way to amplify the label-free
electrochemical detection of DNA hybridization. This amplified guanine response was used
to develop a new hybridization scheme for detecting a sequence of BRCA1 breast cancer
gene. Performing the digestion of the DNA sample in the presence of copper, another
interesting alternative, well defined hybridization signals were obtained in the range 50-250
gL-1 after 20 min hybridization. The detection limit was 40 ng/mL, 2 ng or 100 fmol in the
50 L hybridization solution.
Wang et al. [113] reported the ultrasensitive electrical biosensing of DNA by using
CNTs. Carbon nanotubes were used in two directions, for the recognition and for the
transduction event. For the biorecognition event, alkaline phosphatase was bond to CNTs
through a carbodiimide linker, with coverage of around 9600 enzyme molecules per CNT.
Once the hybrids are formed, the alkaline phosphatase bond to CNTs catalyzes the formation
of the enzymatic product α-naphtol that can be electrochemically detected. The sensitivity,
improved almost 104 times in the presence of CNT modified with alkaline phosphatase. If
CNTs are also used in the transduction step by modifying the glassy carbon used as
transducer, a second amplification was obtained (30 fold), in connection with the
accumulation of the enzymatic product α-naphtol in the presence of nanotubes. A detection
limit of 1 fg/mL, or 54 aM, or 820 copies or 1.3 zmol in the 25 μL sample was obtained after
20 min accumulation.
Wang et al. [114] also proposed an attractive alternative for the amplification of DNA
hybridization based on the ultrasensitive stripping-voltammetric detection of the dissolved
CdS tags coming from the SWCNTs (Figure 20). The nanotubes were previously treated with
acetone in an ultrasonic bath for 1 h and dried in air for 15 h to eliminate the free acetone.
The activated nanotubes were dispersed in toluene and then mixed with the oligonucleotides
modified with CdS for 4 hours with shaking. The biotinylated probe 1 was added to the
streptavidin assay plate at room temperature and the hybridization took place (in a 750 mM
NaCl/150 mM sodium citrate) after addition of the probe 2-coated-SWCNT-CdS-streptavidin
previously obtained by mixing streptavidin and the biotinylated oligonucleotide called probe
2. After 1 h interaction the CdS was dissolved by addition of 1 M nitric acid. The
electrochemical determination was performed in 0.2 M acetate buffer solution pH 5.2 using a
mercury film-glassy carbon electrode in connection with square wave voltammetry to
measure the cadmium obtained. Under these conditions the detection limit at -0.60 V was 40
pg/mL or 330 amol in the 50 μL solution.
From reference 114.
Figure 20. Schematic representation of the analytical protocol of the biosensor: (a) Dual hybridization
event of the sandwich hybridization assay, leading to the capture of the CdS-loaded CNT tags in the
microwell; (b) dissolution of the CdS tracer; (c) stripping voltammetric detection of cadmium at a
mercury-coated glassy carbon electrode. P1, DNA probe 1; T, DNA target; P2, DNA probe 2.
Another work reported the immobilization of nucleic acids on CNTs previously opened
by oxidative treatment and protected by a spin-on-glass film [69]. Once the carboxyl residues
were obtained they were activated with EDC and NHS and the nucleic acids were
immobilized (Scheme 3). The hybridization event in this case was evaluated by using a
peptide nucleic acid sequence as probe and a target containing a fluorescent residue.
From Reference 69.
Scheme 3. Schematic representation of the immobilization of nucleic acid to the functionalized CNT
array.
Dai and He [115] proposed a new way for detecting the hybridization event by using
aligned CNT-DNA probes immobilized on the tip and wall of plasma aligned CNTs. The
CNTs were treated in acetic acid-plasma, followed by grafting the probe DNA sequence
through the carboxyl functions generated during the pretreatment and the amino terminal of
the DNA located at 5´-phosphate position in the presence of EDC. The detection of the
hybridization event was performed through the amperometric response ferrocene-
carboxaldehyde used as a label of the target DNA.
Fang et al. [116] described the direct detection of the hybridization event by combining
the advantages of CNTs electron transfer properties with the incorporation of nucleic acids as
dopants during the polypyrrole formation. The sensing layer was obtained by immobilization
of oxidized MWCNTs in DMF on GCE, followed by the incorporation of DNA within the
polypyrrole film during the electropolymerization process. The hybridization was performed
at 42 oC for 30 min and the analytical signal was obtained from the decrease of impedance,
especially in the region of high frequencies. The authors attributed this behavior to the higher
conductivity of dsDNA compared to ssDNA. The detection limit was 1.0 x 10-8 M.
Yao et al. [117] reported the electrostatic assembly of calf thymus DNA on oxidized-
MWCNTs-modified gold electrode using a cationic polyelectrolyte as linker. The nanotubes
films were prepared by dropping the suspension of MWCNTs in water (0.1 mg/mL) and then
evaporated under vacuum at 50 oC. The electrodes were dipped in 1.0 mg/mL PDDA for 1
hour and then in DNA for around 30 min. The multilayer film was formed by alternate
immersion in the polycation and DNA. Cyclic voltammetry in 1/15 M phosphate buffer pH
4.5 containing 0.1 M KCl showed a couple at 0.035 and 0.128 V (cathodic and anodic peaks)
related to carboxylic acid groups, responsible for the negative charges. An interesting study of
the multilayer formation using Quartz Crystal Microbalance (QCM) and Electrochemical
Impedance Spectroscopy is described. The amount of DNA immobilized at the PDDA-
MWCNT-modified gold electrode was calculated from QCM experiments, being 517.6 ng. A
much smaller amount of DNA was immobilized at bare gold electrode or at gold electrode
modified with MWCNTs, demonstrating the importance of the presence of PDDA.
Electrochemical Impedance was performed using 10 mM potassium ferricianyde and
potassium ferrocyanide, a frequency range from 0.1 Hz to 100 kHz and potential amplitude of
5 mV. The resistances for charge transfer decreased when the gold electrode was covered
with MWCNs, increasing after the immobilization of PDDA and DNA due to the non-
conductivity of these molecules. On the other hand, the negative charge of DNA can also
prevent the access of the marker to the electrode surface. The resistance increased with the
concentration of DNA and also with the number of bilayers. The authors used the electrode
for detecting the interaction of DNA with chlorpromazine, which interacts with dsDNA
through the intercalative association of the planar tricyclic phenothiazine ring system into the
double helix. By QCM it was possible to calculate the binding constant, which is 8.41 x 104
M-1 and the binding site equal to 4.70.
CONCLUSION
In summary, CNTs have demonstrated to be an excellent material for the development of
electrochemical sensors. The state of CNTs surface is a very important aspect and several
pretreatments have been proposed. They possess the main goal to oxidize CNTs improving
the electron transfer and making them more accessible for further derivatization. In general,
CNTs are broken and converted into open tubules rich in oxygenated functions, process that
can be detected by different spectroscopic and electrochemical techniques.
A large number of protocols for the preparation of electrodes based on the use of CNTs
have been proposed. As in the case of pretreatments, in general, the selection of a given
scheme will depend on the system under investigation. For instance, composites are easier to
obtain from the point of view of the preparation time, while some others like the modification
of GCE with CNTs dispersed in Nafion, are very convenient because in one step it is possible
not only to immobilize the CNTs but also to have a barrier for interferents or a platform for
further modifications.
The combination of the uniqueness of CNTs with the powerful recognition properties of
enzymes, nucleic acids and antibodies, and the known advantages of the electrochemical
techniques, represents a very good alternative for the development of biosensors able to
address the new biosensing challenges of the future.
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[121] Pantarotto, D.; Partidos, C. D.; Graff, R.; Hoebeke, J.; Briand, J.-P.; Prato, M.; and
Bianco, A. (2003). J. Am. Chem. Soc 125, 6160-6164.
CONCLUDING REMARKS
The research field of nanomaterials and bioapplications is widening and

growing very fast. In last decade has documented the rampant success of new
technology applied in almost every field of agriculture, medicine, science,
technical developments in electrochemical, chemical, physical, aerospace,
oceanography, earth science, military, and pharmaceutics industrial applications.
With keeping pace in the scientific development it has become necessary to
introduce the curriculum of nanoscience and bioapplications at undergraduate and
graduate levels in college or university education. Different sources have become
now available in public domain or web pages and news related with the
developments in nanobioscience. For trainers, teachers and educators, it is
mendatory to keep them updated with growing knowledge of nanotechnology
with upcoming bioapplications. The nanomaterials have wider applications to
utilize in social, health, and everuy wake of daily life.
The present lecture series is compiled from different available resources and
it is hoped that students, researchers and scientists will be benefited in their
interest in nanobioscience. The selection of nanomaterials and their applications
is arbitrary and unlimited. The limited information will cater the needs of
graduate researchers. With this hope we shall continue to keep adding new
information, new scientific and research material periodically.
Rakesh Sharma, Ph.D

Research Professor,
Center of Nanotechnology and Biotechnology,
Florida State University, Tallahassee, Florida 32304
Web page: http://myprofile.cos.com/rakesh
Avdhesh Sharma,Ph.D
Professor,
Nanotechnology Divison,
Electrical Engineering Department,
MP Agriculture and Technical University,
Udaipur, Rajasthan, India 313001
Web page: http://www.ctae.ac.in/index.php?page=faculty-2

Nano Materials Lectures

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Web-based Selected Lecture Series

Nanomaterials and Bioapplications

UNIT I: NANOMATERIALS AND PROPERTIES

STATUS: PRIVATE MATERIAL ONLY FOR DISTRIBUTION TO STUDENTS IN COURSE

Lecture material compilation Editors:

Cited material sources for students to read:

NSTI 2007,2008 and 2009 Conference CDs

UNIT I: NANOMATERIALS AND PROPERTIES

UNIT II: NANOMATERIAL SYNTHESIS AND CHARACTERIZATION

UNIT III: BIOAPPLICATIONS IN MATERIAL SCIENCE

Concluding Remark 681

STATUS: PRIVATE CIRCULATION FOR EDUCATION AND ACADEMIC PURPOSE ONLY

WHAT ARE NANOMATERIALS? BIOAPPLICATIONS

INTRODUCTION AND LAYOUT OF LECTURES

superconductor: Nanotubes have been shown to be superconducting at low temperatures.

5. ELECTRONIC DESIGN AND DESIGN AUTOMATION

QUANTUM TRANSPORT IN CARBON NANOTUBES

Keywords: Drude model; carbon nanotubes; electrical conductivity; drift mobility;

Eq.(1) is standard in physics of semiconductors and constitutes a relevant element of

where h is the Planck constant.

where v Fn is the magnitude of the quantized Fermi velocity.

PHYSICAL CHARACTERISTICS OF CARBON

Figure 1. The schematic diagram of the ECR-CVD system [26].

where , , , , and are dimensionless parameters and they are

A general solution to Eq. (5) can be expressed as

c1 cos c2 sin c3 cosh c4 sinh (8)

6TiCl4+8NH3 6TiN+24HCl+N2 (14)

RESULTS AND DISCUSSION

Figure 3. A SEM image of the Ni catalyst.[26].

Figure 10. Raman patterns of the Co-filled CNTs film [27].

Figure 12. EDXS of the Co-filled CNTs [27].

Figure 13. J-E properties of the Co-filled CNTs film [27].

The computational results of fundamental frequencies of carbon nanotube are shown in

300 Euler beam model

Timoshenko beam model

Chiral angle Aspect ratio Thermal effect Frequency

7.59 0.3 375.1

Figure 16(a). Growth rate vs. electric field strength [30].

Figure 16(b). Growth rate vs. oxide height [30].

[3] Trends in Nanotechnology Research, Edited by Eugene V. Dirote, Nova Science

HOW GAS-CARBON NANOTUBES SURFACE

BSSE basis set superposition error

GGA-DFT Generalized-gradient approximation DFT

Further Symbols and Acronyms

Motivation - Combining Nanotechnology and Surface Science

The combination of nanotechnology and surface science toward heterogeneous catalysis

(1) Heterogeneous catalysis is a multi-billion-dollar industry ($11.2 bil. in 2006) [41] of

(4) Nanoscience is highly inter-disciplinary; therefore, the results obtained in surface

Advantages of Carbon Nanotubes as Supports in Heterogeneous Catalysis

Monolayer CNT Samples: Drop-and-Dry Technique

Variations in Sample Preparation Techniques for Surface Chemistry Studies

TDS features very similar to those of ozone or high-temperature ―act

• A CNT/silica sample obtained as outlined above is typically well covered (Figure 1)

Oriented (Lateral/Vertical) CNT Samples

Commercially Available CNT Materials

CNTs are commonly produced by 1) LASER ablation, 2) arc discharge, or 3) catalytic

high pressure CO disproportionation) [118] consist of mixed (⅓-metallic and ⅔-

Table 2. Studies of gas-surface interactions conducted at ultra-high vacuum conditions

Adsorbates Techniques Ref.

Adsorbate Ref. Technique HOPG

The projects summarized in section 5 are conducted with HiPco powder-based

BRIEF LITERATURE SURVEY OF HOPG - HIGHLY ORDERED

Adsorption Kinetics on HOPG

Water and Hydrogen