Series Editor's Preface

This book on Modern Fourier Transform Infrared Spectroscopy is a

useful addition to the ComprehensiveAnalytical Chemistry series. The
work contains different chapters that cover both fundamental and
applied aspects of infrared spectroscopy. Particular attention is given
to fundamentals of vibrational spectroscopy and to the recent develop-
ments of hyphenated chromatographic techniques. In addition, the
major portion of the applications described in this book deal with
polymeric and biological materials. Chemometric interpretation and
data analysis are also described in detail in the last chapter of the book,
indicating their relevance in infrared spectroscopy. The book can be
used as an academic text and as reference book either for those with
more expertise or for those starting with this technique. Overall, the
book covers an important technique increasingly used in analytical
chemistry.
Finally I would like to thank the authors of the book for their time
and efforts in preparing their contributions. Without their engagement
this reference work on infrared spectroscopy would certainly not have
been possible.

D. Barcel6

xvii
Acknowledgements

Writing a book needs a lot of reading, careful planning and writing. The
process is time consuming and requires the assistance of people on
whom we can rely. During the process of writing this book many have
helped us with physical work, ideas and support. It is not possible to
thank everyone who has contributed to the book. However, there are
some who have contributed to elevate the quality of the book and we are
grateful for their efforts. In this respect, we would like to thank
Professor Rolf Manne, Department of Chemistry, Agder University
College for critically reviewing Chapter 4. We would like to thank Dr.
Hideki Kandori (Kyoto University) for critically reviewing parts of
Chapter 8 and Ms. Seiko Hino for polishing the English in Chapters 3,
5, 7, and parts of Chapters 8 and 9. In addition one of the authors (VGG)
would like to thank Sheila E. Rodman of Polaroid Corporation for the
collection of some of the materials used in Chapters 6, 7 and 8.
Finally, we would like to thank our families and friends who have
given us moral support and helped us through some difficult times
during the writing of the book.

xviii
Authors' Preface
Infrared spectroscopy has a history of more than a century: the charact-
eristic absorptions of functional groups in the infrared region were
known even in the 19th century; the first infrared atlas was published in
1905, twenty years before the birth of quantum mechanics. However,
even though infrared spectroscopy is a relatively old technique, it has
always been a popular technique for chemical analysis. Developments
in computer technology, sensitive detectors and accessories for new
sampling methodologies in the infrared region have made infrared
spectroscopy one of the most powerful and widespread spectroscopic
tools of the 20th and 21st centuries. The applications of infrared
spectroscopy, and of Fourier transform infrared spectroscopy (FT-IR)
in particular, are ever expanding, due to its versatile nature. The
enormous number of articles and research papers published every year
that deal with infrared spectroscopy and its applications is clear
evidence to this.
The book "Modern Fourier Transform Infrared Spectroscopy" has
been written to reflect the popularity of infrared spectroscopy in sev-
eral different fields of science. The chapters are designed to give the
reader not only the understanding of the basics of infrared spectroscopy
but also ideas on how to apply the technique in these different fields.
The book is suitable for students at graduate level as well as
experienced researchers in academia and industry. The first three
chapters deal with the fundamentals of vibrational spectroscopy. Since
spectroscopy is the study of the interaction of electromagnetic radiation
with matter, the first two chapters deal with the characteristics,
properties and absorption of electromagnetic radiation. Chapter 3
provides the basis for vibrations in molecules from both classical and
quantum mechanical points of view. The absorption of infrared
radiation by a vibration in a molecule depends on the symmetry of the
molecule and the symmetry of the vibrations. As this aspect is not
usually treated in textbooks on infrared spectroscopy, Chapter 4 deals
with the symmetry aspects of molecules and illustrates how the reader
can determine the vibrations that are infrared active.
Chapter 6 is an overview of the instrumentation used to perform the
majority of Fourier transformed infrared spectroscopic experiments

xix
today. The chapter starts with an overview of the history of FT-IR
spectroscopy from the construction of the first interferometers in 1880
to the present day and continues with a description of the components
of an interferometer and the various scanning techniques (continuous-
scan and step-scan). Chapter 7 first describes sampling techniques
used in transmission and reflection spectroscopy and then a variety of
the so-called hyphenated techniques that combine the use of FT-IR
spectroscopy with another analytical technique. Thermogravimetric
analysis (TGA/FT-IR), liquid chromatography (LC/FT-IR), gas chroma-
tography (GC/FT-IR) and supercritical fluid chromatography (SFC/FT-
IR) are the combinations discussed in this book.
Chapter 8 depicts certain applications of FT-IR spectroscopic tech-
niques to basic and industrial research. Specifically, a large portion of
the chapter deals with the characterization of polymers and polymeric
surfaces, whereas the remaining part describes applications to organic
thin films and biological molecules. One subcategory treated in detail is
the determination of molecular orientation in polymers via static and
dynamic FT-IR experiments. Another subcategory is the applications
that involve optically active materials and conducting polymers. In
addition, very significant developments have recently taken place in
the area of infrared microspectroscopy and especially in infrared imag-
ing with the introduction of focal plane array detection. Part of this
chapter is dedicated to an explanation of the experimental procedures
associated with these imaging experiments along with selected
examples from the recent literature.
Finally, Chapter 9 deals with some modern analytical methods in
infrared spectroscopy. Again, the methods described here are not very
common in books on infrared spectroscopy. The first part of the chapter
deals with chemometric techniques that can be applied to semi-quanti-
tative and quantitative analysis of infrared spectroscopic profiles. The
text is designed to give the theoretical basis of these techniques and in
particular how they can be applied to infrared spectroscopic data
profiles. In this chapter, the subject of two-dimensional correlation
spectroscopy (2D-IR) is also discussed. The principles of the technique
along with selected examples of the applications of the 2D-IR treat-
ment are presented.
Alfred A. Christy
Yukihiro Ozaki
Vasilis G. Gregoriou
April 2001

xx
Chapter 1

Electromagnetic radiation and the

electromagnetic spectrum

We have come across people talking about microwave, UV radiation,

radio waves, x-rays, radar, cosmic rays and so on. We understand the
dangers of UV radiations from the sun and the use of microwave in
heating food. What do we associate with all these different terms? We
understand without learning the physics of these different radiations
that they are associated with different energies. For example, white
light is a form of energy and it comprises a mixture or spectrum of seven
different colours, which are all visible to the human eye. All these
colours of which white light is composed have different energies in the
descending order: violet, indigo, blue, green, yellow, orange, red. A
photographic plate is readily affected by violet light, unlike red light
which has almost no effect.
From the above discussion, it is clear that the spectrum of different
radiations falls into a larger scale of spectrum where white light is a
very small part. The larger scale containing the spectrum of different
radiations is called electromagneticspectrum.
The physical properties of radiations cannot be explained by a
single theory. Some properties such as propagation of radiation
through a medium, diffraction and reflection are better explained by a
theory called wave theory and properties like momentum are better
explained by particle theory. The propagation of radiation through
space involves electric and magnetic components of the radiation and
hence the term "electromagnetic".
Spectroscopy generally deals with the interaction of electromag-
netic radiation and matter. In order to understand this interaction, we
must understand the characteristics of the electromagnetic radiation
and the matter involved in the interaction.

1
1.1 WAVE NATURE OF ELECTROMAGNETIC RADIATION-WAVE
CHARACTERISTICS AND WAVE PARAMETERS

According to electromagnetic theory, electromagnetic radiation is a

form of energy that is composed of oscillating electric and magnetic
fields acting in planes that are perpendicular to each other and to the
direction of propagation (Fig. 1.1). The oscillating electric field is simple
harmonic and propagates as waves with a velocity (c) 3x10 8 m s- in
vacuum. The propagation velocity varies with the refractive index of the
medium.
In a two-dimensional representation, the variation of the electric
field strength of an electromagnetic radiation with propagation time in
space can be paralleled to the variation of the y co-ordinate of the trace
of a particle moving around a circle of radius A with a constant angular
velocity o radians per second (Fig. 1.2).
Let us assume that OX and OY represent the positive directions of
the x and y axes and O represents the origin of these axes. Let us also
assume that at time zero the particle passes through x and then
consider the position of the particle after t seconds. The angle traversed
by the particle in t seconds is ot radians. The y co-ordinate of the
position of the particle can be given by equation

y =A sin ot (1.1)

Electric field streng~

Direction of propagation

Fig. 1.1. Oscillating electric and magnetic fields of electromagnetic radiation.

2
 ct=r/2

s-1

act=r

wt= 3r12

Fig. 1.2. Trace of a particle moving around a circle of radius A with an angular velocity
rads- 1.

At time zero (i.e. ot = 0) the particle is at x and this function is

minimum with a value 0. At time 7d/20 (i.e ot = c/2) the particle is at y
and the function is maximum with a value A, the amplitude of the
function.
At time nl/c (i.e cot = g), the particle is at Z and the function has
another minimum. Similarly the function will have another maximum
and another minimum at times 3/2co (cot = 37/2) and 2/co (ot = 2),
respectively. The particle takes 2/o seconds (remember one circle is 2i7
radians) to complete the journey through the circular path once (one
cycle). This is called the period of the motion and denoted by . The
number of cycles the particle traces through in one second is o/2t {1/
(2t/o)} and is called the frequency (v) of the motion. The frequency is
abbreviated by the symbol Hz. The angular velocity of the motion can
then be written in terms of the frequency as

co = 2v (1.2)

A two-dimensional plot of the variation of the y co-ordinate of the

position of the particle with time is shown in Fig 1.3. Equation (1.1) can
be written to include the frequency of the motion and time as

y =A sin 2vt (1.3)

3
 A

O.5 A A

>1 0--

-0.5A

-A

Time in seconds

Fig. 1.3. A two-dimensional plot of the variation of the y co-ordinate moving in a circle as
shown in Fig. 1.2.

0.SA

-0.5A

-A

Propagation distance in metres

Fig. 1.4. The propagation of electromagnetic radiation.

The propagation of the electromagnetic radiation in space is 3x108 m

s l. Figure 1.3 can be redrawn (Fig. 1.4) in a similar manner to
represent the distance of propagation of electromagnetic radiation
along the x axis.
When the x axis represents the distance, we can define some other
parameters characteristic to electromagnetic radiation. In the

4
preceding discussion, we learnt that the y co-ordinate of the particle
has zero values at times 0, n/co and 27/co. That is at distances 0, rc/lo and
2nc/o, the propagating electric field of the electromagnetic radiation
will have zero field strength. But the distance between the extreme
points, that is, the distance travelled during a complete cycle of oscilla-
tion is called wavelength (k). However, because of the symmetry of the
sine wave, the wavelength can be defined as the distance between two
similar points in the wave. The wave has a frequency v and therefore
the velocity of propagation can be written as

kv = c (in metres per second) (1.4)

This implies that the wavelength has dimension m (metre). The

inverse of the wavelength, when the wavelength is expressed in centi-
metres is called wavenumber and is denoted by v. The dimension for
wavenumber is cm -1 .

v = 1/k cm -1 (1.5)

The wavenumber can also be thought of as the number of waves in 1 cm

length. The above two equations combined give Eq. (1.6).

v = Vc (c in centimetres per second) (1.6)

The propagation distance s of an electromagnetic radiation in t seconds

is given by

s = kvt = ct (1.7)

Equation (1.7) can be combined with Eq. (1.1) to give a relationship

describing the variation of the field strength of an electromagnetic
radiation with the distance of propagation and its wavelength as
follows

y =A sin (2nrs/k) (1.8)

The flux of energy of an electromagnetic radiation along the direction of

propagation is equally divided between electric and magnetic fields.
Electromagnetic radiations are produced by accelerating electric
charges and electric charge accelerations are produced when

5
electromagnetic radiations are absorbed. Electromagnetic radiations of
different energies are produced when the energy involved in the accel-
eration of electric charges is different. We shall see later that y-rays to
microwaves are produced during oscillations of charges at different
states in matter.

1.2 QUANTUM CONCEPT AND PARTICLE NATURE OF

ELECTROMAGNETIC RADIATION

Not all properties of electromagnetic radiation can be explained by the

wave theory. Electromagnetic radiation has particle-like (corpuscular)
properties in addition to the wave properties. The corpuscular nature of
electromagnetic radiation was developed during the early 20th century
in order explain certain characteristics that classical physics failed to
explain.
Max Planck originated the quantum hypothesis to explain the
discontinuity in the energy of an oscillator of frequency v. Planck's
hypothesis suggests that the energy of an oscillator with frequency v is
not continuously variable, but restricted to integral multiples of hv as
0, lhv, 2hv, 3hv, ... nhv where hv is a quantum of energy and h is a
universal constant which is known as Planck's constant.
Albert Einstein, in 1905, related Planck's quantum hypothesis to
the photoelectric effect where electrons from metal surfaces were
ejected by ultraviolet radiation. Einstein explained that the electrons
from a metal are ejected when they receive energy at least equivalent to
their binding energy. Furthermore, the energy of the radiation must be
confined to a small region of space in order to transfer the energy
entirely to an electron and eject it instantaneously. The ejection will
not be instantaneous if the energy is spread evenly across the entire
wave front. This leads to an understanding that the energy of electro-
magnetic radiation can be considered as packets (quanta) of energy hv
(Fig. 1.5). The quantum of radiant energy was named photon by Lewis
in 1926.
The corpuscular nature of photons was explained by their
possession of momentum. The relativistic expression shown in Eq. (1.9)

Fig. 1.5. Energy packets (quanta) of photons.

6
leads to a value of hv/c for linear momentum (p) of a photon which has
no mass (m = 0).

E = m2c 4 +p2c 2 (1.9)

The scattering of photons (electromagnetic radiation) during collisions

with electrons is a clear indication that the photons possess
momentum.

1.3 UNITS OF WAVE PARAMETERS

The SI unitary system requires that the dimensions of length, mass

and time are specified in metres (m), kilograms (kg) and seconds (s),
respectively. Table 1.1 shows the units of some parameters and con-
stants we have come across during our discussion.

TABLE 1.1
Symbols and dimensions of wave parameters and related terms
Parameter Symbol Dimensions of units
Energy J (Joule) kg m2 s- 2
Wavelength m
Frequency Hz (Hertz) s-1 (cycles per second)
Velocity of light 3x108 m s- 1
Planck's constant 6.626x1034 Js

1.4 ORIGIN OF ELECTROMAGNETIC RADIATION AND

ELECTROMAGNETIC SPECTRUM: y-RAYS TO MICROWAVE

The origin of electromagnetic radiation varies widely. The universe

contains radiations ranging from y-rays to microwaves. A system that
emits radiation is also capable of absorbing that radiation. An example
of this is the absorption and emission of yellow light by sodium atoms
by which sodium is quantitatively determined by atomic spectrometry.
We use a sodium lamp that emits light at a particular frequency which

7
 visible

infrared
radiation
Y-rays

Fig. 1.6. Oscillators in an atom and a molecule.

is produced when an excited electron relaxes from an excited state to

the ground state to excite an electron from a ground state to the same
corresponding excited state in another sodium atom.
As mentioned earlier, the electromagnetic radiations originate from
oscillators arising from the acceleration of electric charges. Gamma-
rays are produced by the oscillators in the atomic nuclei; x-rays are
produced by the oscillators arising from the tightly bound electrons in
the vicinity of the nucleus of atoms; visible and ultraviolet radiations
are produced by the oscillators arising from the outer electrons in the
molecules and atoms; infrared radiation is produced by the oscillators
arising from the vibration and rotation of molecules.
Now we should be able to understand why electromagnetic radi-
ations of different dimensions are used in studying nuclear, electronic,
vibrational and rotational transitions in matter.

TABLE 1.2
Electromagnetic spectrum: yrays to radio waves, their approximate frequency and energy
range
Radiation Wavelength Frequency range Energy range per Energy range
(m) (Hz) photon (J) per mole (kJ)
-1
y-rays 5x10-12_10 -1 2
6xl103x1O20 4x10-12x10 3 24x106-1.2x10 s
16 18 - 14
x-rays 10-S-5x10 - 12
3x10 -6x101 9 2x 10- -4x 10 1200-24x10 6
8 5 16 1 8
Far UV 1.8x10 7-10 1.7x 101 -3X10 1.13x10 -2x10 18 680-1200
3.5x10- 7-1.8x10- 7
Near UV 8.6x1014-1.7x1015 5.7x10-19-1.13x 10-18 343-680
Visible 7.7x10-7-3.5x10- 7 3.9x1014-8.6x1014 2.58x10-19-5.7x10 l- 9 155-343
4 4 0 19
Near IR 2.5x10-6-7.7x10 - 7 1.2x10' -3.9x101 8x10-2 2.58x10 48-155
Mid IR 5x10-5-2.5x106 6x 1012 -1.2x1014 4x10-2_8x 10 - 2.4-48
1 2 22 -2 1
Far IR 10-3-5x105 3x10 -610 2x10- -4x10 0.12-2.4
Microwave 3x10-1 -10 -3
10 9-3x10 11 6.6x10-2-2x10 - 2 2 0.0004-0.12

8
 Electromagnetic radiation spectrum in the larger scale is given in
Table 1.2. It is difficult to define precise limits between different
divisions and the reader should understand that the divisions are
approximate and may slightly vary with the limits given in other
textbooks.

GENERAL BIBLIOGRAPHY

P.W. Atkins, Molecular Quantum Mechanics. Oxford University Press,

London, 1984.
C.N. Banwell and E.M. McCash, Fundamentals of Molecular Spectroscopy.
McGraw-Hill, London, 1994.
W. Kemp, OrganicSpectroscopy. Macmillan, London and Basingstoke, 1978.
J.H. Vander Maas, Basic InfraredSpectroscopy. Heydon & Sons, London, 1972.

9
Chapter2

Interaction of electromagnetic radiation

with matter

2.1 ABSORPTION OF ELECTROMAGNETIC RADIATION

Matter is composed of molecules or atoms. Atoms are composed of

nuclei containing protons, neutrons and electrons surrounding the
nucleus. This means that matter is full of oscillators of very different
dimensions. Any of these can be excited to a higher level using electro-
magnetic radiation of appropriate energy. For example, vibration in a
molecule containing two atoms is equivalent to a simple oscillator. This
vibration can be excited to the next vibrational level by irradiating the
molecule with infrared radiation of appropriate energy (i.e. appropriate
frequency). Here, we shall just assume that absorption takes place (Fig.
2.1) without considering the conditions for infrared absorption, which
we shall consider in Chapter 3.
If the energy of the radiation does not match the energy difference
between the excited and ground states of the molecule, no absorption
will take place. If the frequency of the radiation that is absorbed by the
molecule is v, then the energy difference between the ground state and
excited state is given by the Eq. (2.1).

Ee-Eg = AE = hv =hc =hvc (2.1)

where Ee, Eg are absolute energies of the excited and ground states.
For example, if the wavenumber of the radiation needed to excite a
vibrational mode in a molecule is 3000 cm- l, then the energy absorbed
by one molecule is

11
 I - E x.tedx

AE = hp

Fig. 2.1. Absorption of electromagnetic radiation.

AEmoiecule = hvc = 6.626x 10 -3 4 J s x3000 cm - l 3x 101 cm s- l (2.2)

= 5.963x10 - 2 0 J

If we are interested in the energy absorbed by a mole of the substance

then we have to multiply the answer by the Avogadro's number NA =
6.023x10 2 3 mol-1. That is, the energy absorbed by a mole of the
substance is

AEmole = 5.963x1020 J x 6.023x1023 mol - = 35.9x10 J mol-' (2.3)

= 35.9 kJ mol-

2.2 PRESENTATION OF DATA: A LINE SPECTRUM

In absorption spectroscopy, the information sought for is the

absorption of radiant power from a source by the sample. In infrared
spectroscopy, this is done in practice by a spectrometer; the construction
of this instrument will be dealt with in a later chapter. The idea behind
the technique is to send radiation through (or interact with) a sample
and measure the characteristics of the radiation emerging from the
sample.
Let us assume that monochromatic radiation (only one-frequency
radiation) with sufficient intensity and of a frequency that matches the
frequency of absorption is passed through a sample. A part of the
radiation will be absorbed and the intensity of the radiation emerging
from the sample, the transmitted radiation, will be of reduced

12
 100

0
v
Frequency, Hz

Fig. 2.2. A transmittance spectrum.

aW

0
V
Frequency, Hz

Fig. 2.3. A line spectrum of absorption.

intensity. If we plot the percent of the intensity of the transmitted

radiation relative to the source intensity against frequency, then the
expected plot should contain a single line as shown in Fig. 2.2.
Similarly an absorption spectrum will be as shown in Fig. 2.3.

2.3 LINE BROADENING IN INFRARED ABSORPTION SPECTROMETRY

The real absorption spectrum of a diatomic molecule, which has only a

single vibrational mode, is not a line spectrum, but an absorption
spectrum containing a broad peak with a maximum at the frequency of
absorption as shown in Fig. 2.4.
The line broadening in spectrometry arises because of character-
istics adherent to molecules in different phases and uncertainty in the

13
 1

Q
-E
11
ii
A

V
Frequency, Hz

Fig. 2.4. A broad peak representing an absorption.

energy levels due to the limited residence time of the particle in the
excited state.
In solids, liquids and gases the particle velocities are different. The
particles collide more frequently in the gas phase than in the liquid
phase. The vibrational and rotational energy levels are perturbed from
their actual values and lead to small variations in the ground state
energy levels. This implies that solids should give sharp bands. This is
true, but the bands are split because of electronic interactions. Spectral
line broadening arises also due to the Doppler effect. The infrared
measurements are made in cells where the radiation is allowed to pass
through or interact with the sample. Molecules that are in motion
towards the source will absorb radiation of higher frequency compared
with the molecules that are moving away from the source (lower
frequency).
One of the important effects that cause spectral line broadening is
the uncertainty effect. The uncertainty principle proposed by Werner
Heisenberg suggests that there is natural limitation on how precise a
pair of physical parameters can be made. In vibrational spectrometry
the uncertainty in the energy level of the excited state, AE, and
uncertainty in the lifetime of the molecule at an excited vibrational
state, At is related by the uncertainty relationship as follows.

AE At 2 h/2 (2.4)

This can be rewritten to include frequency v as follows

A(hv)At 2 h/2t (2.5)

14
i.e.

AvAt > 1/2xT (2.6)

If the residence times at the excited vibrational states are infinite then
the uncertainty in the frequency of absorption is zero and the frequency
can be determined precisely. The molecules that undergo vibrational
transitions have a finite time of residence of 10-8 s and this leads to an
uncertainty in the frequency

Av > h/(2xAt) = 1/(2x3.14x 10-8) s 108 (2.7)

This uncertainty is small compared to the radiation frequency of

infrared radiation 1012-104. This leads us to conclude that absorption
by a single vibrational mode will be a band spectrum with a finite
frequency width as shown in Fig. 2.4.

2.4 MEASURED SPECTRA OF DIATOMIC MOLECULES

The measured infrared spectra of diatomic molecules do not show a

single band as we expected but fine spectra with several fine peaks
spaced at equal intervals with a space in the middle. The fine spectra
arise from the rotational spectra of diatomic molecules. When a di-
atomic molecule is excited the rotational levels are also excited and the
rotational absorptions are superimposed on the vibrational spectra of
the diatomic molecules. The vibrational spectrum of carbon monoxide
at high resolution is shown in Fig. 2.5.

2.5 NORMAL OR FUNDAMENTAL VIBRATIONS

Molecules of a substance are in continuous motion. They move

(translation) and rotate. Each atom in a molecule assumes a new
position with time.
Each atom in a molecule can be represented by three co-ordinates (x,
y, z) in a Cartesian system. We say that the atom has three degrees of
freedom. When we consider a molecule containing N atoms, the atoms
have a total of 3N degrees of freedom. The result of the movements of

15
 o

o -

C
o
g
O C
g

C
0

o -~
CC
O
:1

C,

0O. -
o

O UC

C
o -E:

N oC
CC
g

N E

-~C
rl i
CC
M

c
I
F,
the individual atoms can be represented by the movement of the centre
of mass of the molecule.
The position of the centre of mass-the position of the molecule in
space-can then be represented by three co-ordinates, i.e., three
degrees of freedom. The molecule rotates about its centre of mass, and
the rotations about three mutually perpendicular axes passing through
the centre of mass require three more degrees of freedom. The free
movement of the atoms is restricted by the bonds between the atoms,
but they vibrate from their equilibrium positions. These are repre-
sented by the remaining 3N - 6 degrees of freedom. These vibrational
motions are called normal or fundamental vibrations. Linear molecules
need only two degrees of freedom to specify rotations of the molecules in
space. Therefore, the linear molecules have 3N - 5 fundamental vibra-
tional modes.

2.6 INFRARED SPECTRUM OF POLYATOMIC MOLECULES

Fortunately, the energy needed to excite most of the vibrational modes

in organic molecules falls in the infrared spectral frequency region
7.5x10 1 2-1.2x101 4 (250-4000 cm-l). In infrared spectroscopy, it is
tK
1350 900

- GroYp requencies - f- Fierprt--

3.0-

2.5

i 2.0

w 1.5
eI n.

kA
1.0.

0.5.

0.05 iI.
Y
I1}

.
AAA, )L
i
vI

.
I1
,
, Aid
VCLU v
.
X
. I
- V
I
,
..

4000.0 3000 2000 1500 1000 450,0

Wavenumber cm-1

Fig. 2.6. An infrared absorption spectrum of polyatomic molecule (polystyrene).

17
customary to give the excitation energy needed in terms of wave-
number v which has a directly proportional relationship with frequency
v as v = v/c. The region 4000-1350 cm-l contains group frequencies and
the region 1350-900 cm-l contains low energy vibrations. This region is
characteristic of the molecule and is called the fingerprintregion.
If the molecule is polyatomic and the radiation is polychromatic,
then the vibrational modes absorbing in the region 4000-250 cm-l will
result in considerable overlap and the plot between absorbance and
wavenumber will be a mixture of sharp and broad bands over the whole
range. Wavenumbers are usually plotted from higher wavenumbers to
lower wavenumbers and the plot is called an infrared absorption
spectrum (Fig. 2.6).

GENERAL BIBLIOGRAPHY

P.W. Atkins, Molecular Quantum Mechanics. Oxford University Press,

London, 1984.
C.N. Banwell and E.M. McCash, Fundamentals of Molecular Spectroscopy.
McGraw-Hill, London, 1994.
W. Kemp, Organic Spectroscopy. Macmillan, London and Basingstoke, 1978.
J.H. Vander Maas, Basic InfraredSpectroscopy. Heydon & Sons, London, 1972.

18
Chapter 3

Theory of infrared spectroscopy

3.1 PRINCIPLES OF INFRARED SPECTROSCOPY

When irradiated with infrared light, a molecule absorbs it under some

conditions. The energy hv of the absorbed infrared light is equal to an
energy difference between a certain energy level of vibration of the
molecule (having an energy Em) and another energy level of vibration of
the molecule (having an energy En). In the form of an equation,

hv=En-Em (3.1)

holds. In other words, absorption of infrared light occurs principally

based on a transition between energy levels of molecular vibration.
This is why an infrared absorption spectrum is a vibrational spectrum
of a molecule.
Satisfying Eq. (3.1) does not always cause infrared absorption.
There are transitions which are allowed by a selection rule (i.e., allowed
transition) and those which are not allowed by the same rule (i.e.,
forbidden transition). In general, transitions with a change in the
vibrational quantum number by +1 are allowed transitions and other
transitions are forbidden transitions. This is what is known as a
selection rule with respect to infrared absorption. Another selection
rule with respect to infraredabsorption is one which is defined by the
symmetry of a molecule. This selection rule is, in other words, "a rule
that infrared light is absorbed when the electric dipole moment of a
molecule changes as a whole in accordancewith a molecularvibration."
The two selection rules are developed from quantum-mechanical
considerations. According to quantum mechanics, for a molecule to
transit from a certain state m to another state n by absorbing or

19
emitting infrared light, it is necessary that the following definite
integral:

(P)mn = f W.tln mdQ (3.2)

or at least one of (py)mn, and (z),, which are expressed by a similar

equation is not 0, where Px denotes an x-component of the electric dipole
moment; v denotes the eigenfunction of the molecule in its vibrational
state; and Q denotes a normal coordinate (i.e., a normal vibration; see
Section 3.3) expressed as a single coordinate. Now, let us consider only
(px)mn. A distribution of electrons in the ground state changes as the
coordinate expressing a vibration changes and, therefore, the electric
dipole moment is a function of the normal coordinate Q. Hence, Px can
be expanded as follows:

PX =( )o (d /Q)oQ+ 1(82 /Q2)Q2+... (33)

2
Expressed by a displacement of atoms during the vibration, Q has a
small value. This allows us to omit Q2 and the subsequent terms in the
equation above. Substituting the terms up to Q ofEq. (3.3) in Eq. (3.2)

8 x
(p.x)m, =(Pi)oWVm dQ+ f QydQ (3.4)

is obtained. Due to the orthogonality of the eigenfunction, the first term

of this equation is 0 except when m = n holds. The first term denotes the
magnitude of the permanent dipole of the molecule. For the second
term to have a value other than 0, both (p /taQ)o# 0 and lJynQ /mdQ 0
must be satisfied. These two conditions lead to the two selection rules.
The nature of the eigenfunction permits the integral to have a value
other than 0 only when n = m 1 holds. (Considering Q2 and the
subsequent terms of Eq. (3.3) as well, we can prove that even when n =
m + 1 fails to hold, (x)mn has a value, even though small, other than 0).
The first selection rule regarding infrared absorption is thus proved.
The other selection rule, which is based upon the symmetry of a
molecule, is obtained from (JcdQ) 0 0. The relationship (pJaQ)o X 0
indicates that infrared absorption occurs only when certain vibration
changes the electric dipole moment. The vibration is infrared active

20
when (pJaQ) 0 0 holds, but is infrared inactive when (p 1 JcaQ)0 = 0
holds.
Since most molecules are in the ground vibrational state at room
temperature, a transition from the state v" = 0 to the state v" = 1 (first
excited state) is possible. Absorption corresponding to this transition is
called the fundamental. Although most bands which are observed in
infrared absorption spectra arise from the fundamental, in some cases,
we can find bands which correspond to transitions from the state v" = 0
to the state v" = 2, 3, 4 ... (i.e., overtone transitions). However, since
overtones are forbidden, overtone bands are very weak.
The horizontal axis and the vertical axis of an infrared absorption
spectrum must now be explained. A frequency is indicated along the
horizontal axis in the units of wavenumber (cm-1 ) (with higher wave-
numbers always on the left-hand side) (Fig. 3.1). On the other hand, a
transmittance T (%) (Fig. 3.1a) or an absorbance E (Fig. 3.1b) is
expressed along the vertical axis. While infrared spectra include
reflectance spectra and emission spectra in addition to absorption

(a)
80

60

40

20

0
1.0
(b)
0.8

0.6

0.4

0.2

0
4000 3200 2400 1600 800

Fig. 3.1. Examples of infrared spectra: (a) transmittance spectrum; (b) absorbance
spectrum.

21
spectra, a reflectance, an emission intensity, etc. are expressed along
the vertical axis in the case of a reflectance spectrum, an emission
spectrum, etc.

3.2 CHARACTERISTICS OF INFRARED SPECTROSCOPY

Infrared spectroscopy provides detailed information about vibrations of

a molecule. Since molecular vibrations readily reflect chemical features
of a molecule, such as an arrangement of nuclei and chemical bonds
within the molecule, infrared spectroscopy contributes considerably
not only to identification of the molecule but also to study of the
molecule structure. Furthermore, an interaction with a surrounding
environment also causes a change in molecule vibrations, and hence,
infrared spectroscopy is useful in studying the interaction, too.
Infrared spectroscopy has many uses from basic research to various
applications. Why is infrared spectroscopy useful? The answer is
simple. It is spectroscopy which probes a vibration of a functional
group. Infrared spectroscopy can be used not only for the identification
of a functional group, but also for the investigation of the chemical bond
and environment of the functional group. For example, C=O groups of
-C=C-C=O and -CH2 -CH 2 -C=O give different frequencies. Of course,
a -C=O group and -C=O .. H-O- also yield different frequencies. We
can summarize the characteristics of infrared spectroscopy as follows:
1. Using an electromagnetic wave having a low energy, infrared spec-
troscopy rarely damages a sample. In addition, infrared spectro-
scopy may be used for non-destructive analysis of a sample.
2. Infrared spectroscopy is applicable to a sample in various states,
e.g., solid, crystal, fibre, film, liquid, solution and gas. Furthermore,
measurements of infrared spectra of a sample in a solution and in a
solid state, allow us to compare its structure in the solution with
that in the solid.
3. Infrared spectroscopy uses not only so-called infrared absorption,
but can utilize infrared reflection, emission, photoacoustic spectro-
scopy as well.
4. Connection with an optical microscope, a gas chromatograph, a
liquid chromatograph or other instrument is relatively easy, which
allows hyphenated analysis.

22
3.3. MOLECULAR VIBRATIONS

Knowledge about vibrations of a molecule is crucially important for

understanding infrared spectroscopy. It is useful also for Raman and
near-infrared spectroscopy. While vibrations of a polyatomic molecule
are generally complex, according to harmonic oscillatorapproximation
(i.e., an approximation on the premise that the force which restores a
displacement of a nucleus from its equilibrium position complies with
Hooke's law; vibrations in harmonic oscillator approximation are called
harmonic vibrations), any vibrations of the molecule are expressed as
compositions of simple vibrations called normal vibrations. Normal
vibrations are vibrations of nuclei within a molecule, and translational
motions and rotational motions of the molecule as a whole are not
included in normal vibrations. In each normal vibration, all atoms
vibrate with the same frequency (normal frequency), and they pass
through their equilibrium positions simultaneously. In general, a mole-
cule which consists of N atoms has 3N - 6 normal vibrations (3N - 5
normal vibrations if the molecule is a linear molecule). Since normal
vibrations are determined by the molecular structure, the atomic
weight and the force constant, when these three are known, we can
calculate the normal frequencies and the normal modes.

3.4 A VIBRATION OF A DIATOMIC MOLECULE

As the simplest example of molecular vibrations, a vibration of a

diatomic molecule will now be considered. Being always a linear mole-
cule, a diatomic molecule has only one normal vibration (3 x 2 - 5 = 1).
Needless to say, this vibration is a stretching vibration that the mole-
cule stretches and contracts (Fig. 3.2a). We will describe the stretching
vibration in accordance with classic mechanics. Assuming that the
nuclei are masses, m1 and min, and the chemical bond is the "spring" as

m, m2

(a) (b)

Fig. 3.2. (a) A stretching mode of a diatomic molecule. (b) A model for a diatomic molecule
(two masses combined by a spring).

23
in Hooke's law (with the spring constant k) (Fig. 3.2b), we can explain
the vibration of the molecule based on classic mechanics.
Now, assume that the masses m and m2 deviate Ax 1 and Ax2,
respectively, from their equilibrium positions. Then, the potential
energy of the system shown in Fig. 3.2b is:

V = k(Ax 2 -Ax 1 )2 (3.5)

Meanwhile, the kinetic energy of the system is:

1 2 1 dx('.i>
T =-m~lx - m2 x 2 = dt) (3.6)

Now that V and T are known, motions of the system are determined by
solving Lagrange's equation of motion:

d(aT + =0 (3.7)
dt (ai ax i

However, before solving Lagrange's equation of motion, we introduce

new coordinates Q and X.

Q= pL(Ax 2 -Ax 1 ) (3.8)

+
X = mAx mAx2 (3.9)
Vm + m2

Now,

= m2 (where i is a reduced mass) (3.10)

m +M2

Q is a coordinate of the displacement of a distance between the two

masses, while X is a coordinate of the displacement of the centre of
gravity of the system. Using Q and X, the potential energy V and the
kinetic energy T are written as:

24
T 1 Q2 + 1x2 (3.11)
2 2

v= k 2 (3.12)
2 p

We substitute V and T in the Lagrange equation of motion (3.7). First,

applying to the coordinate X (xi = X), we obtain

X=0 (3.13)

This expresses a free translational motion which is not bounded by the

potential energy. On the other hand, from the Lagrange equation of
motion regarding the coordinate Q(xi = Q), we obtain

dQ +k Q =O
2
(3.14)
dt pL

From the differential equation like Eq. (3.14), we can find a solution as
follows:

Q = Q0 cos2Tvt (3.15)

Equation (3.15) implies that the system illustrated in Fig. 3.2b has a
simple harmonic motion with the frequency v and the amplitude Q0.
Substituting Eq. (3.15) in Eq. (3.14),

(-42v2 +k)Q=0 (3.16)

Finally, we find the frequency of the spring as:

v = I 1k (3.17)
2n

Since the frequency of the spring corresponds to the frequency of the

molecular vibration and the spring constant corresponds to the force

25
TABLE 3.1
Stretching frequencies and force constants of diatomic molecules

Molecule Reduced mass (p) Force constants G(v)

-2 4
(1.66x10 ) (105 dyne/cm) (cm - l )
1
(N cm )
H2 0.50 5.73 4160
HD 0.67 5.77 3631

D2 1.00 5.77 2944

35C1 2 17.50 3.21 556

N2 7.00 22.9 2331

02 8.00 11.8 1555

HF 0.95 9.17 3962
35
H C1 0.97 5.16 2886

HBr 0.98 4.06 2558

HI 0.99 3.12 2233

NO 7.46 15.9 1877

CO 6.85 19.0 2143

constant of the chemical bond, it can be seen from Eq. (3.17) that the
frequency of the molecular vibration is proportional to the square root
of the force constant and inversely proportional to the square root of the
reduced mass of the atoms. Table 3.1 shows the stretching frequencies
and the force constants of some diatomic molecules. As can clearly be
seen in the table, the stronger a chemical bond is and the smaller the
masses of atoms are, the higher the stretching frequency of a molecule
becomes.
As diatomic molecules, there are those like H2, which consist of the
same atoms (homonuclear diatomic molecules) and those like HCl,
which consist of different atoms (heteronuclear diatomic molecules). Of
these, vibrations of only heteronuclear diatomic molecules appear in
infrared absorption spectra. This is because while the electric dipole
moment of a heteronuclear diatomic molecule changes with a molecule
vibration, that of a homonuclear diatomic molecule is always 0.

26
3.5 QUANTUM MECHANICAL TREATMENT OF A VIBRATION OF A
DIATOMIC MOLECULE

Quantum mechanics allows us to describe energy levels of vibrations of

diatomic molecules. In quantum mechanics, the first step is to write
down Schridinger'sequation, HI = Ey. The second step is to solve the
equation to calculate an eigen value and an eigen function. In terms of
classic mechanics, the total energy H of a vibration of a diatomic
molecule is the sum of a kinetic energy 1/2Q 2 (Eq. 3.11) and a potential
energy (1/2)k.(Q2/p) (Eq. 3.12),

H=T +V =2 Q2 +_ Q2 (3.18)

Replacing Q with an operator -ih/2z.dldQ, H is calculated as:

h2 d 2 lk
H -- d + -kQ2 (3.19)
8n 2 dQ 2 2

Substituting this in Hy = Ey and processing the formula, a Schrod-

inger equation on harmonic oscillator of a diatomic molecule is
obtained.

d2 Y 87 2 I Q2>
+- E- k -j) =0 (3.20)
dQ 2 h2 2 )

As how to solve this equation is described in detail in a number of

books, we will explain only a result. The formula (3.20) has a solution
only to the following eigen value Ev:

E v =(V +-hv (3.21)

where V is a quantum number of a vibration (V = 0, 1, 2, ...).

An eigen function to each value of E v is expressed as:

Wv =Nv Hv(, Q)exp - (3.22)

27
where Nv denotes a normalization constant, Hv is a Hermite poly-
nomial, and a = 4i72!v/h.
Wave functions to V = 0, 1, and 2 are as follows.

v/o =(a / 7) 4 exp(-aQ 2 /2) (3.23a)

p1 =(a / 7)14 (2a)V2 Q exp(_aQ 2 / 2) (3.23b)

,2=(al/t)" 4
(1/ 2)(2aQ2 - 1)exp(-aQ 2 /2) (3.23c)

As is clearly understood from these formulas, a wave function of a

harmonic oscillator is an even function when a quantum number is an
even number but is an odd function when a quantum number is an odd
number. Figure 3.3 shows a potential energy, wave functions, v,
existence probabilities, x' and energy eigen values, E, of the
harmonic oscillator.
When we treat vibrations of diatomic molecules in accordance with
quantum mechanics, we will find different results from when we treat
the same in accordance with classic mechanics. Firstly, the lowest
vibrational energy is not 0 but Eo = (1/2)hv. Energies have discrete
values E = (3/2)hv, E2 = (5/2)hv, E 3 = (7/2)hv, ..., and an energy
difference between adjacent energy levels is always hv. Another major

(a) (b)

-Q-0- +

Fig. 3.3. A potential energy, wave functions, and probabilities of existence of harmonic
oscillator.

28
difference between a conclusion we obtain from quantum mechanics
and a conclusion from classic mechanics is the amplitude of molecular
vibrations. While classic mechanics never allows us to assume that the
amplitude, namely, the existence probability of a diatomic molecule
extends beyond a potential energy, quantum mechanics permits us to
find a slight existence probability outside a potential energy in each
state (Fig. 3.3b).

3.6 VIBRATIONS OF POLYATOMIC MOLECULES

We will consider normal vibrations of carbon dioxide (CO,; linear tri-

atomic molecule) and water (H 2 0; non-linear triatomic molecule) as
examples of the simplest polyatomic molecules. CO2 has 3x3 - 5 = 4
normal vibrations. Figure 3.4 shows the four normal vibrations 1, 3, 2a
and 2b (see Section 4.8.1 for labelling rules). The normal vibrations 1
and 2 are vibrations that two CO bonds stretch and contract in phase
(1) and out of phase (3), respectively, called symmetric and anti-
symmetric stretching vibrations. Meanwhile, the vibrations 2a and 2b
are both vibrations that the angle of OCO changes and are called
bending vibrations.While the vibrations 2a, 2b are independent of each
other, energies required for the vibrations are in principle equal to each
other, only with planes of the vibrations differing by 90 degrees from
each other. That is, the two types of vibrations have the same energy.
Such vibrations which have principally the same energy are called
degenerate vibrations.
1 0

3 _ 0

2a ?

2b DO

Fig. 3.4. Molecular vibrations of CO2: (1) symmetric stretching vibration; (3) anti-
symmetric stretching vibration; (2a, 2b) degenerate vibrations.

29
 1 9 q

V, C
2
V
3

Fig. 3.5. Molecular vibrations of water: (1) symmetric stretching vibration (vl); (2)
bending vibration (v2); (3) antisymmetric stretching vibration (V3).

To know whether the normal vibrations 1, 3, 2a and 2b are infrared

active or not, we examine a change in the electric dipole moment at
equilibrium positions (prJQ),. In the normal vibration 1, the electric
dipole moment is always 0. Hence, the normal vibration 1 is infrared
inactive. Conversely, the electric dipole moment largely changes in the
normal vibration 3, and thus it is infrared active. In a similar manner,
the normal vibrations 2a and 2b accompany a change in the electric
dipole moment, and therefore, are infrared active (see also Section 4.8.5
and Fig. 4.28). With respect to a molecule such as a CO 2 molecule which
has the centre of symmetry, a general rule holds true that an infrared
active vibration is Raman inactive and a Raman active vibration is
infrared inactive. This rule is called the mutual exclusion rule.
Water, being a nonlinear triatomic molecule, has three normal
vibrations, as shown in Fig. 3.5. The normal vibrations 1 and 3 have
different frequencies from each other, because of different H1... H2
interactions between the two (see also Section 4.7.4 and Fig. 4.23).
In the cases of CO 2 and H 20 molecules, the frequency of a stretching
vibration is higher than that of a bending vibration. This indicates that
the stretching vibration requires a higher energy than the bending
vibration.
Next, let us consider vibrations of atomic groups. Figure 3.6 shows
six vibrational modes of an AX 2 group (e.g., CH 2, NH 2). Of the six, two
vibration modes are stretching vibrations, one being a symmetric
stretching vibration and the other an antisymmetric stretching vibra-
tion. The remaining four are bending vibrations, i.e., scissoring, rock-
ing, wagging, and twisting vibrations. Of the four bending vibrations,
scissoring and rocking vibrations are bending vibrations in the plane of
CH2 (in-plane vibrations), while wagging and twisting vibrations are
vibrations which displace vertically to the plane of CH 2 (out-of-plane
vibrations).

30
 1 2 3

4 5 6

+ T + +

Fig. 3.6. Molecular vibrations of AX2 group: (1) symmetric stretching vibration; (2)
antisymmetric stretching vibration; (3) scissoring vibration; (4) rocking vibration; (5)
wagging vibration; (6) twisting vibration.

In general, group frequencies are useful to consider vibrations of a

polyatomic molecule which includes three or more atoms (see Chapter
5). Group frequencies are vibrations of particular atomic groups
(functional groups), such as rocking, symmetric and antisymmetric
vibrations of a CH2 group, C=O stretching vibration of a carbonyl group
and stretching vibration of an OH group. (Bands due to group
frequencies are called characteristicabsorption bands.) The concept of
group frequencies hold true when certain normal vibrations are sub-
stantially determined by movements of two or a plurality of atoms
(atomic group). Group frequencies play prominent roles in the analysis
of infrared and Raman spectra. A more detailed description of group
frequencies is given in Chapter 5.

3.7 QUANTUM MECHANICAL TREATMENT OF VIBRATIONS OF

POLYATOMIC MOLECULES

We will now explore vibrations of polyatomic molecules in accordance

with quantum mechanics. A kinetic energy T and a positional energy V
are expressed as:

T =2 (3.24)
2 i=1

31
 i;iQi
V2
2 i=1
(3.25)

Hence, a total energy H is:

1 n 2V 1
H=T+V= 1Qi +Z-YiQi2 (3.26)
2i 2 i=,

Replacing Q with -ih/2w.d/dQ again and calculating Ht, we can yield a

wave equation (3.27) regarding vibrations of polyatomic molecules.

h2 a 2 +i
+
87
-2 E
, CqQ2 2 -1riQ2 V =EW (3.27)

Since normal vibrations are independent of each other, the formula

above can be separated into n wave equations which, respectively,
correspond to the respective normal vibrations, an eigen value E is
expressed as the sum of eigen values E i of the respective normal
vibrations, and an eigen function yv is expressed as a product of eigen
functions yi representing the respective normal vibrations. In short,
since the formula (3.27) has the same style as formula (3.20), the eigen
value Ei is also the same as formula (3.21).
Hence, a total of vibrational energies whose frequencies are v1, v2, ...,
Vn is:

E i =(V i +1/2)hv i (3.28)

Ev =E 1 +E 2 + .+En

=iV
1 + )hv, +iV 2 +)hv 2 + +Vn +2 hv (3.29)

Figure 3.7 shows energy levels of v1, v2, and v3 modes (Fig. 3.5) of a
water molecule. In Fig. 3.7 (0, 0, 0) denotes the lowest ground state and
(1, 0, 0), (0, 1, 0), (0, 0, 1) denote fundamental levels at which v1, v2 and
v3, respectively, have a quantum number of 1. Transitions between the
lowest ground state and the fundamental levels are fundamentals.
Next, (2, 0, 0), (0, 2, 0), (0, 0, 2) denote that vj, v2, and v3 have a quantum
number of 2, respectively, and are called ouertone levels. (3, 0, 0) ... are

32
 (,,
(1 11)

12000 I_______ -(1

_ 0 1)
(0 2 1)

(O 11)

(O 0 1)
7 8000 I--____ _-(1 0 0)
0 (0 2 0)

--
- (O 1 0)

(0 0 0)
4000

A~

Fig. 3.7. Energy levels of vl, vl, and V3 modes of water.

also overtone levels. Overtones are transitions between these overtone

levels and the lowest ground state. Combination mode levels are levels,
such as (1, 0, 1) and (0, 1, 1), at which two or more normal vibrations are
excited. Transitions between combination mode levels and the lowest
ground state are called combination modes.

3.8 ANHARMONICITY

So far, we have treated molecular vibrations as harmonic oscillators.

However, except for the vicinity of the bottom of a potential energy
curve, the harmonic oscillator model is not a good model on molecular
vibrations in reality. If the harmonic oscillator model were correct, as
we can clearly see in Fig. 3.3, dissociation of molecules should never
occur, no matter how large the amplitude is. Hence, it is necessary to
consider a potential energy function V(r) (r denotes an inter-nuclear
distance) which more accurately expresses vibrations of molecules. In

33
 -Q-- o - +Q
(re)

Fig. 3.8. Morse's function.

accordance with our instinct, V(r) must be a function such that it

rapidly increases when r approaches zero but gradually comes close to a
dissociation energy, De, where r >> re (re is an equilibrium distance)
holds. As a function which satisfies this condition, Morse's function
expressed as below is well known:

V(r) = De[1 - exp(-a(r - re))] 2 (3.30)

In formula (3.30), a is a constant. Figure 3.8 shows Morse's function.

Assuming that Q = r - re is always smaller than r and expanding V(r) by
Taylor's series into a polynomial with respect to Q in the vicinity of re,

, Q+ I IQ+-
r
) re 2
~" 6( d3
6\ or~(3.31) (3. 31)

24 +-(a8r 4 ), Q4 +

Since the first term on the right-hand side is a constant term, this term
is regarded as 0. With respect to the second term as well, since V is

34
extremely small to re, the second term is also regarded as 0. Now,
ignoring the fourth and higher-order terms and applying (32V ar2 )re = k,
the following formula holds:

V(r)= 1kQ2 (3.32)

2

In other words, Morse's function is equivalent to a function which

expresses harmonic oscillator approximation in the region close to the
equilibrium inter-nuclear distance re (a second derivative on formula
(3.30) yields k = 2a 2De).
A potential energy V is generally expressed as:

V= k2 Q2 + k3Q 3 + k4Q4 + ... (3.33)

The high-order terms such as Q3 and Q4 are called anharmonic terms.

Calculating an eigen value Ev' considering up to the Q3 term, we obtain,

EV =V + hre-IV+ 2hvexe (3.34)

where v, =a / D/ 2 n. The symbol Xe is a constant called an an-

harmonic constant.
It is possible to approximately assume the degree of anharmonicity
from the value of this constant. Table 3.2 shows the values of an-
harmonic constants for major diatomic molecules. While the constant xe
has a value of 0.01 approximately, if a molecule contains a hydrogen
atom which has a light mass, the constant xe increases (Table 3.2).
Since the anharmonic constant xe holds the following relationship with
respect to a, De, etc., it is possible to calculate the shape of Morse's
function and a dissociation energy of the molecules, etc.

ha
Xe= hve
hv - ha (3.35)
4De 4 2c1*D

From formula (3.34), one can calculate a difference, AEv, between

energy levels of vibrational quantum numbers V and V + 1.

AE= hv, - 2hvexe(V + 1) (3.36)

35
TABLE 3.2
Anharmonicity constants of diatomic molecules
Molecules Anharmonicity constant

H2 0.02685
D2 0.02055
HF 0.02176
HC 0.01741
HBr 0.01706
H1 0.01720
C12 0.007081
I2 0.002857
N2 0.006122
02 0.007639
NO 0.007337

Formula (3.36) shows that the larger V is, the smaller AE v is.
In this formula, a transition u = 0 - 1 is:

AEv = hve,- 2hvexe = hve(1- 2xe) (3.37)

Hence, AE = hv, does not hold. The value Vobs (which is an observed
value) is obtained as AE v = hvob,. We will now describe a method of
calculating ve from Vob.
HCI exhibits a strong band at 2886 cm -1 due to a fundamental (V = 0
to 1) and a weak band at 5668 cm -1 due to a first overtone (V = 0 to 2). It
is possible to calculate an absorption wavenumber ve and an anharm-
onic constant xe from these observed values.
With respect to V = 0 - 1 and V = 0 -> 2,

2
AE (1) = hv( (1- x ) (3.38a)

AE, (,-,) = 2hv,(1 -3x, ) (3.38b)

Hence,

2886 = v(1-2x ) (3.39a)

36
5668 = 2v,(l- 3xe ) (3.39b)

Solving these simultaneous equations, we obtain xe = 0.0174 and ve =

2990 cm - . We must consider Ve to discuss the strength of a chemical
bond, since considering Vob, is not sufficient for this purpose.

3.9 OVERTONES AND COMBINATION MODES

It is anharmonicity that allows overtones and combination modes to be

observed. Let us consider selection rules regarding infrared spectra
once again. This time, we will consider anharmonicity on a dipole
moment.

I I Q-L-! Q2 . (3.40)
DQ )o 2y Q

() =( 4)Of edQ+ j J JnQy-dQ

(~ Ix m(3.41)
=1xlv m +Q |v

2(,9Q22 )9Q
0dQ+ Q

The third term has a value other than 0 when ( 2 pjdQ 2 ) 0 and
2
ynQQ PmdQ X 0 both hold. The latter integral has a value other than 0
when V' V and V + 2. Hence, even a first overtone is no longer
forbidden if we consider the term Q2 as well. In a similar manner,
second, third, etc. overtones are no longer forbidden as we take higher-
order terms into consideration. However, the intensities of these over-
tones are far weaker than those of fundamentals. The frequencies of
first, second, third, etc. overtones are smaller than double, triple,
quadruple of the frequencies of fundamentals, respectively. This is
because the differences between the vibrational energy levels become
narrower as the quantum number u increases, as clearly shown in Fig.
3.9 and indicated by formula (3.36). Anharmonicity excludes combina-
tion modes as well from those forbidden in a similar manner. The
intensities of combination bands are also weak.

37
3.10 FERMI RESONANCE

In some cases, overtones and combination modes are as strong as

fundamentals. This occurs when we have Fermi resonance. Fermi
resonance is developed by anharmonicity, when the frequency of an
overtone or a combination mode by chance happens to be approx-
imately the same as that of a fundamental (or when an overtone and a
combination mode have very close frequencies with each other). When
we have Fermi resonance, two relatively strong bands appear in a
region where we are supposed to observe only one strong fundamental,
one on the high-wavenumber side to the fundamental or the overtone
and the other on the low-wavenumber side to the fundamental or the
overtone. These two bands both contain contributions from the funda-
mental and the overtone. In other words, Fermi resonance occurs when
the fundamental and the overtone are mixed together because of
anharmonicity.
Figure 3.9 shows an infrared spectrum of benzaldehyde as an
example of Fermi resonance. In general, aldehyde yields a band due to
a CH in-plane bending vibration in the vicinity of 1400 cm l. An
overtone of this band is expected to appear in the vicinity of 2800 cm l,
and its frequency is close to the frequency of CH stretching vibration of
aldehyde. In the real spectrum, we find one band on a somewhat higher
wavenumber side and another band on the lower wavenumber side to
2800 cm-l (Fig. 3.9). These bands are created because of Fermi reson-
ance between the overtone of the CH in-plane bending vibration and
the CH stretching vibration.
2

000 00-__

I1
5T
10000 F
Isd
,,
I
CUN
-
I- Al~~~J'LL
-
JW L( - -

3200 2800 2400 2000 bU I&0

/cml

Fig. 3.9. An infrared spectrum of benzaldehyde as an example of Fermi resonance

38
 Let us now consider why Fermi resonance occurs. Assume we have
small terms (such as a potential energy due to anharmonicity) which
provide perturbations to two energy levels, El, E2 (El > E2). We can
calculate changes in E1 and E2 caused by the perturbations if we solve
the following formula with respect to W.

E 1 + al -W 0
=0 (3.42)
D3 E 2 +a 2 -W

where al, a 2, P are the small terms which provide the perturbations. As
we expand formula (3.42), we obtain,

W 2 -(E 1 +E 2 + 1 + a2)W +-(E

1 + a,)(E + a2)-P
2
=0 (3.43)

As we solve this formula (assuming that a, a 2, p are sufficiently

smaller than E1, E2), we obtain,

W=E1 +a 1 + D or E 2 +a 2 (3.44)
E1- E,2 E - E2

Changes in E1 and E2 are extremely small if E1 - E2 >> 0. However,

when E1 - E 2 = 0, that is, when E1 and E 2 are close to each other, a type
of resonance occurs, thereby considerably changing the values of E and
E2.
When we consider Fermi resonance, we must note that only those
having the same vibrational symmetry cause Fermi resonance between
them. For instance, although two OH stretching vibrations v1 and V3
(Fig. 3.5) of a water molecule do not cause resonance between them
since the vibrations have different symmetries from each other, their
overtones (2v1 and 2v3 ) have the same symmetry and therefore cause
Fermi resonance (see also Section 4.9.2).

GENERAL READING

Theory of molecular vibrations

G. Herzberg, MolecularSpectra and Molecular Structure I. Spectra of Diatomic
Molecules, 2nd edn. Van Nostrand, Amsterdam, 1950.

39
G. Herzberg. Molecular Spectra and Molecular Structure II: Infrared and
Raman Spectra of Polyatomic Molecules. Van Nostrand, Amsterdam,
1945.
K. Nakamoto, Infrared and Raman Spectra of Inorganic and Coordination
Compounds, 5th edn. Wiley-Interscience, New York, 1997.
E.B. Wilson, Jr., J.C. Decius and P.C. Cross, Molecular Vibrations. McGraw-
Hill, New York, 1955.
L.A. Woodward, Introduction to the Theory of Molecular Vibrations and
VibrationalSpectroscopy. Oxford University Press, London, 1972.

Infraredspectroscopy
L.J. Bellamy, The Infrared Spectra of Complex Molecules, Vol. 1, 3rd edn.
Chapman and Hall, London, 1975; Vol. 2, 2nd edn. Chapman and Hall,
London, 1980.
N.B. Colthup, L.H. Daly and S.E. Wiberley, Introduction to Infrared and
Raman Spectroscopy, 3rd edn. Academic Press, San Diego, CA, 1990.
P.R. Griffiths and J.A. deHaseth, Fourier Transform Infrared Spectroscopy.
Wiley-Interscience, New York, 1986.

40
Chapter 4

Symmetry of molecules, group theory and

its applications in vibrational
spectroscopy

In Chapter 3, we learnt that a molecule is infrared active if the dipole

moment changes during the vibrational motion and that a molecule is
Raman active if the polarizability changes during the vibrational
motion.
In the case of diatomic molecules, the number of degrees of freedom
corresponding to the vibrational motions in the molecule is 3x2 - 5 = 1.
This motion is the stretching vibration along the axis of the molecule.
We saw in Chapter 3 that the dipole moment of a heteroatomic diatomic
molecule changes during its vibration and the molecule is infrared
active, and that the dipole moment of a homonuclear diatomic molecule
does not change and the molecule is infrared inactive. We then went on
to discuss carbon dioxide molecule and identified that the normal
vibrations 2, 3a and 3b are infrared active because of the change in
dipole moment during these vibrational motions in the molecule.
Furthermore, we found that the normal vibration 1 is infrared inactive
because the dipole moment does not change during this motion..
However, by the general mutual exclusion principle, as we saw in
Chapter 3, we know that this rule holds for molecules with centre of
symmetry. That is, the vibrational motions that are infrared active are
Raman inactive and the motions that are infrared inactive are Raman
active (this must be seen as a rule of thumb not as an absolute
principle). To determine whether a motion is Raman active, we must
find out whether there is change in the polarizability during this
motion. Symmetric vibrational modes in a molecule that lead to a
change in the size of the molecule generally involve changes in the
polarizability and are Raman active. For example, symmetric stretch-

41
ing in carbon dioxide leads to a change in the size of the molecule and
involves a change in polarizability and the motion is Raman active.
For the polyatomic molecules, it is often difficult to determine
whether a mode is infrared active or Raman active. The selection rules
for infrared and Raman absorption can be arrived at by considering the
symmetry of the molecules. In order to do this, we have to learn the
symmetry aspects of the molecules, their group and symmetries of the
different molecular vibrations.

4.1 SYMMETRY OF MOLECULES: SYMMETRY ELEMENTS AND

SYMMETRY OPERATIONS

Geometrical figures such as equilateral triangles, squares, cubes etc.

possess symmetry. A visual inspection of these shapes leads us to
perceive that there are some symmetrical features in them. For
example, in a square figure cut out from a white sheet of paper, all the
sides are equal. Their diagonals bisect at the centre of the square, the
corners of the square are at equal distances from the centre, the square
can be divided into two halves along the axis passing through the mid
points of any two opposite sides or along their diagonals. Furthermore,
the figure can be rotated in several ways about its centre, along the
diagonal axes and along the axes passing through the middle points of
any two opposite sides to get the same figure without any apparent
change. We say that this figure has several symmetry elements. When
certain operations are made, the figure seems apparently unchanged.
We call these operations symmetry operations.
In polyatomic molecules, the orientation of atoms in space may
reveal certain symmetry features in the molecules. We can identify
various symmetry elements, and symmetry operations that can be
performed on molecules. Identifying symmetry elements and under-
standing symmetry operations are important to classify molecules into
different point groups, which we shall consider in the next section.

4.1.1 Identifying symmetry elements and symmetry

operations

We will take the benzene molecule as an example; it possesses all the

symmetry elements we will come across in discussing molecular
symmetry and we will try to understand the symmetry elements and

42
 6 2 Rotationby60 5
2
C6
3 4
5

Fig. 4.1. A clockwise rotation of the benzene molecule by an angle of 60 (C6) about an axis
passing through the centre and normal to the plane of the molecule.

symmetry operations. We mark the corners as 1, 2, 3, 4, 5, and 6 (Fig.

4.1). These numbers will help us to identify any changes in the orienta-
tion of the corners representing the carbon atoms in space.
Let us assume that the plane of the molecule lies on the plane of the
paper. Then we imagine an axis passing through the centre and normal
to the plane of the molecule, and consider some rotations of the figure
along this axis. A rotation of the molecule (clockwise) about this axis
through an angle of 60 (2x/6) is performed on the molecule and the new
positions of the corners are shown in Fig. 4.1. The molecule is in-
distinguishable if the numbers indicating the positions of the corners
are removed.
We can repeat the same operation five more times leaving the
molecule apparently unchanged (Fig. 4.2). After the sixth operation,
the molecule assumes its original position. The rotation axis is the
symmetry element and the rotation about this axis is the operation.
This rotation axis is called proper rotation axis (we shall see later that
there is defined another rotation axis called improper rotation axis). It
is given the symbol C and the order of the axis is written as the suffix.
The six-fold proper rotation axis in benzene can be written as C6. The
operation generated by this element has also the same symbol. The
operations generated by the C6 symmetry element are C6, C62 , C 63, C64 ,
C65 and C66. As mentioned above, the molecule assumes its original
position after the C6 6 operation. This is thus the same as doing nothing
to the molecule (C66 = E). The operation is called the identical operation
and denoted by E.
The molecule also has a three-fold (Fig. 4.3) rotation axis C,, and a
two-fold (Fig. 4.4) rotation axis C2, coinciding with the proper rotation
axis we identified earlier. The proper rotation axis of highest order is
called the principal axis. It is also important to identify equivalent

43
 6 5 4

4 6 5
5 1 6 C6 3
C6

4 2 3 ! 2 6

(b 3 (d)

6
C6I

6 4

5
5

(a) 4 C6 5 (f)
0 (e)
C6

4
C6

Fig. 4.2. Multiple C6 rotations. (a) Benzene molecule = C6 6; (b) C6; (c) C62; (d) C63; (e) C64 ;
(f) C65.

TABLE 4.1
Rotation axes, operations generated and equivalent operations
Operation
2 3 4 5 6
C, C6 C6 C6 C C 6

E
2
C, C, C3 C33
C2 C2 C22

rotations at this stage. Some of the repeated operations generated by

the C6 symmetry element may be equal or identical to the symmetry
operations generated by the other symmetry elements. For example,
the symmetry operations generated by the C6, C3 and C2 can be com-
pared. The operations C62 , C63, C64, C6 6 are equal to operations C3, C2, C32

44
 1

6
6 2 Rotation by
1200

C3 1
5 3

(a)

2 4

1 5

(c)

Fig. 4.3. C3 operations about the same axis as in Fig. 4.2.

1 Rotation by 180 4
-

C2
2 * 5

3 6
0
lnttinn iv lRl

4 1
C2

Fig. 4.4. C2 operations about the same axis as in Fig. 4.2.

and E, respectively. Table 4.1 summarises the operations generated by

the rotation axes and some of their equivalent operations. The table
shows that the distinct operationsgenerated by the C6 rotation axis are
C6 and C6 5. The rotation axis C3 generates C3 and C32, and C2 produces
only one distinct operation.

45
 A

C2
2
6 6
6 2 - .

5 3 '3
(a)
4 4

Cc
2
SC
2 \

(b)

C2 XC 2

C2 "

(c)

Fig. 4.5. (a) C2 operations about an axis lying on the plane of the molecule and passing
through two opposite corners of the molecule; (b) different C2 as above; and (c) C2
operations about an axes lying on the plane of the molecule and passing through the
middle points of two opposite sides of the molecule.

There are other two-fold proper rotation axes lying in the plane of
the molecule as shown in Figs. 4.5a, b and c. There are three such axes.
We give them the symbols C2 ' and C2 " to differentiate from the two-fold

46
 1 4

6 2 i 3 5

5 3 2 6

I
4 t

Fig. 4.6. Inversion operation about the middle point of the benzene molecule.

axis shown in Fig. 4.4. We use the same symbol for all three axes of each
type because of our understanding that the operations generated by
these axes are equivalent. When specifying symmetry elements, we
collect them together. There are six C2 axes lying in the plane of the
molecule. As discussed above, it is also clear that the operations C22
generate the identity (C22 = E).
Now, we shall consider other symmetry elements in the molecule.
Any point on the molecule can be inverted (reflected in the midpoint of
the molecule) through the centre of the benzene molecule without any
apparent change in the molecule (Fig 4.6). We call this symmetry
element inversion centre and the operation inversion. The symbol for
the inversion operation is i. The molecule assumes its identical position
when operated on twice with i. That is i2 = E.
The molecule has several symmetry planes. The symmetry element,
symmetry plane generates reflection of the molecule in the plane. The
symbol for the symmetry operation is . These planes of symmetry can
be differentiated as h, V or Cd. GCh is a horizontal mirror plane lying
perpendicular to the principal axis. ov is a vertical mirror plane con-
taining the principal axis which is conventionally taken as vertical.
There are three such mirror planes in the benzene molecule (Figs. 4.7a
and b). This plane contains the molecular plane of the benzene mole-
cule. d is dihedral mirror plane, a special case of a vertical plane
bisecting two C2 axes that lie perpendicular to the principal axis. In the
case of benzene there are three dihedral planes, as shown in Fig. 4.7c.
Each mirror plane generates reflection and the molecule assumes its
identical position when operated on twice with a mirror plane; that is
v2 = E, oh2 = E and 2
Cd = E.
There is another symmetry element called improperrotation axis or
rotation reflection axis. The axis generates a combined operation
consisting of an n-fold rotation followed by a horizontal reflection. The

47
 cv Cv

(a)

Fig. 4.7. (a) Reflection operation about a vertical mirror plane containing the principal
axis and passing through two opposite corners of the molecule; (b) three such vertical
mirror planes; (c) three vertical mirror planes containing the principal axis and passing
through the middle points of the opposite sides of the molecule; and (d) horizontal mirror
plane lying on the plane of the molecule.

benzene molecule has two improper rotation axes S6 and S,3 as shown in
Figs. 4.8a and b. Figure 4.8a shows the effect after the first operation. It
is important to note that the molecule is in reflected form after the

48
 6 6'
I

1'
6

5 i 3
] A
_ .
2'

4 3 a
4

I S6

(a)

1 5 5'
I

6'
6 2

3 1'
5

4 2 2'

S3

(b)

Fig. 4.8. Rotation reflection operation.

rotation and indicated by the numbers with primes. The improper

rotation axis generates operations S6 , S6 2 , S63, S 64, S65, and 566. A careful
study of the resulting molecule after each operation S6 will reveal that
some of the above operations are not distinctive. They are equivalent to
the operations generated by other symmetry elements of the benzene
molecule. The improper rotation axes and their equivalent operations
are given in Table 4.2.
As in the case of the rotation axes, the distinct operations generated
by the improper rotation axis S6 are S, and S65 . The independent
operations generated by the S 3 axis are S3 and S3 5. Identifying distinct-
ive operations generated by the symmetry elements is important in our
discussion of reducible representations.

49
TABLE 4.2
Improper rotation axes, operations generated and equivalent operations
Operation
S6 S6 S62 S6
3
S6
4
S,5 S66
C3 i C32 E

Operation
S, S, S3 2 S33 S34 S, 5 S3 6
2
C3 h C3 E

Now it is time to summarise the symmetry elements of the benzene

molecule. We have identified the identity E, rotation axes C 6, C3, C2,
3C2', 3C2", mirror planes Gh, 3v, 3 d, inversion centre i and improper
rotation axes S6 and S3. Among these symmetry elements, inversion
centre and symmetry planes generate one operation each. However,
the proper and improper rotation axes generate more than one
operation. These operations and their equivalent single operations are
given in Tables 4.1 and 4.2. We can now summarise all the symmetry
operations that can be generated by these elements as E, C6, C6 5, C3,
C32, C2, 3C2', 3C2", i, S 3 , S 35 , S6 , S6 5 oh, 3V and 3 d. We have selected
here a molecule with high symmetry and most of the molecules we will
be dealing with in this chapter will be simpler than this.

TABLE 4.3
Some examples
Figure Symmetry elements Examples

4.9a Only E CHFC1Br

4.9b E and a mirror plane CHCl2Br, (CH 3)2CHC1
4.10a and b E, C2, v' and %c" HO, HS, CH20, COC12, (CH 3 )2C=O,
CH 2C12, C6 H5 X, CH 5 N etc.
4.11a and b E, C3, 3ov NH 3, CHX3, POC13
4.12a and b E, C3 , 3C 2, Gch, 3%v and S3 BF3 , PF,
4.13a E, C. and Dcv HCN
4.13b E, C_, ooc and nCh CO 2

50
4.1.2 Identifying symmetry elements: some examples

Figures 4.9-4.13 illustrate some models of molecules possessing cer-

tain common symmetry elements. Table 4.3 summarises the symmetry
elements and some examples of molecules possessing these.

(b

Fig. 4.9. Type of molecules (a) possessing E as the only symmetry element, (b) possessing
symmetry elements E and a.

/1 C2 C2
0V'- //

(a) (b)

Fig. 4.10. Type of molecules possessing symmetry elements E, C2 , %cy'and c%".

51
 :YY

Fig. 4.11. Type of molecules possessing symmetry elements E, C3, 3 v.

C2

be

%C

C2

Fig. 4.12. Types of molecules possessing symmetry elements E, C3, 3C 2 , Gh, 3%, and S3.

52
 B

infinite number of
p vertical planes through
the principal axis

Coo-principal axis
(b)

A -

I infinite number of
vertical planes through
the principal axis

Cw-principal axis (a)

Fig. 4.13. Type of molecules possessing symmetry elements (a) E, C, and cOOv,and (b) E,
C,, ooo, and h.

4.1.3 The classification of molecules, point groups

The discussion on symmetry elements and inspection of the examples

reveal that the molecules containing a different number of atoms may
contain the same symmetry elements. For example, the molecules
listed in row 3 of the Table 4.3 have the same symmetry elements E, C2,
oyv' and ov". The molecules that have the same symmetry elements can
be shown to have several properties in common. Therefore, it is
advantageous to put all these different molecules into a specific group.
The symmetry operations corresponding to the symmetry elements of a
molecule leave at least one point invariant (unmoved). We call these
groups point groups.

53
 -

C,

C--roUDL

Fig. 4.14. Example of a molecule belonging to C2 point group.

The molecules are assigned to different point groups according to

their symmetry elements. We start with molecules with few symmetry
elements and special groups. Molecules of the types shown in Fig. 4.9a
possess only the identity element E; they belong to group C. The
molecules of the type shown in Fig. 4.9b have E and a vertical mirror
plane and they belong to group C,. We see that the molecules of the type
shown in Fig. 4.10a and b possess more symmetry elements (E, C 2, ,v'
and ov' ) and belong to group C2v. Likewise, the molecules of the type
shown in Fig. 4.11 possess E, C3, v,' and v,", v'"' and are said to belong
to the point group C3 v. We can clearly see that the classification follows
the symmetry elements. Generally, if a molecule has a Cn rotation axis
(principal axis) and n vertical planes passing through the principal
axis, then it is said to belong to the point group Cnv.
Linear molecules of the type shown in Fig. 4.13a have an infinite-
order principal axis and infinite number of vertical planes; the type
shown in Fig. 4.13b have, in addition to the above symmetry elements,
a mirror plane normal to the principal axis. These belong to the point
groups C, and Dih, respectively.
Classifying molecules into different point groups does not require
the identification of all the symmetry elements. The presence of certain
symmetry elements in a molecule implies the presence of certain other
symmetry elements. Scheme 4.1 helps us to assign molecules into
different point groups (without identifying all the symmetry elements
in several cases).

54
 no

Scheme 4.1.

TABLE 4.4

Some examples and their point groups

Molecules Symmetry elements Point group

NH 2 -NH 2 , H2 02 E, C2 (Fig. 4.14) C2

NH3 , PH3, POC13 E, Ca and 3%7 C3v

Trans CIH=ClH E, C2 , h and i (Fig. 4.15) C2h

C2H6 (staggered) E, C3, 3C2 (horizontal), 3o d (Fig. 4.16) D3d

C2H2 E, C2 , 2C 2 (horizontal) and ch (Fig. 4.17) D2h

BF3, PC15 E, C3, 3C 2 (horizontal) and Gh D3h

3
C6H 6 E, 2C6, 2C3, C2 , 3C2', 3C2", i, 2S3, 2S6 , oh, Gd D6h
3
and oh

55
 Oh

C2h- group

Fig. 4.15. Example of a molecule belonging to C2h point group.

6C
/
(-U-,
p)- 0 (K
f C
2 C2

I U
C2 I C2

D3 d-group Od

Fig. 4.16. Example of a molecule belonging to D3d point group.

4.2 GROUP THEORY AND SYMMETRY OPERATIONS AS ELEMENTS

OF A GROUP

Mathematically, a set (G) of abstract elements, on which a binary

operation o is defined is said to form a group with respect to this
operation if the elements of the group obey the following four rules.
1. The product of any two elements A and B and the square of every
element is a member of the set.

56
 D2 h-group

Fig. 4.17. Example of a molecule belonging to D2 h point group.

2. The set contains an identity element E for which EoA = A.

3. The elements follow associative law. That is (AoB)oC = Ao(BoC)
4. For each element A of the set, there exists an inverse in the set such
that AoA -1 = E.
Mathematically, the binary operation can be defined in several ways.
For the purpose of our discussion of symmetry operations, we define
this operation as a product operation (one operation followed by
another). Furthermore, to avoid confusion with the symmetry
elements, we refer the "elements of a group" as "members of the group".
Our aim in this section is to show that the set containing symmetry
operations as members forms a group. We can make use of the water
molecule with four symmetry elements for this purpose. The molecule
(Fig. 4.18) has the symmetry elements E, C2, (v') and % (v"). The
symmetry operations are also E, C2, oz and o%. We must be aware of
the difference between the symmetry elements and symmetry
operations. Symmetry operations contain all the individual operations
that can be performed about the symmetry elements (see the benzene
example).
According to the first rule the product of two symmetry operations
in the set is a symmetry operation. We shall follow the steps shown in
Fig. 4.18 to identify whether the product of the symmetry operations x
followed by C2 yields a symmetry operation. It is evident from the
illustrations that the product of the above operations leaves the atoms
in the molecule in positions that can be transformed by a single
operation. This can be simply written as follows.

57
 Xz

C2
%yz

Fig. 4.18. Symmetry operations and products of symmetry operations. water molecule is
used as an example.

C2( = oyz (4.1)

Similarly, from Fig. 4.18

CCC2 = E (4.2)

A product table (Table 4.5) can be set up to show that this is true for all
the operations. This table will also help us to investigate the remaining
rules. For example, the following two equations (the multiplication
table mentioned above will help here) confirm that the symmetry
operations follow associative law.

58
TABLE 4.5
Product table of operations
The operation performed first

E C2 aF,2 cy
E E C2 a_ 1yz

C2 C2 E ayZ
2 (z
aOxz a7 ayZ E C2
y yz aZ C2 E

= =
(yz C2)xz xz z E (4.3)

G2 (C2 Gxz) = yz yz = E (4.4)

The above equations also explain that the elements cv and 2yz have
inverses yGxand cy, respectively. This is true for all the symmetry
operations in the set.
At this stage, it is necessary to mention that the symmetry
operations may not be commutative. This means that the result of two
successive operations may not be the same if their operation order is
reversed.
The symmetry operations can be represented by several ways. If
there exists a group with other members P, Q, R and S that satisfy the
same multiplication table shown above, the group is said to be
homomorphous with the group containing symmetry operations.

4.3 MATRIX REPRESENTATION OF THE SYMMETRY OPERATIONS

In a Cartesian co-ordinate system the position vector a of a point A can

be written as the product of a row vector containing the unit vectors
along the co-ordinate axes and a column vector containing the co-
ordinates of the point.

a= (ij k) y

59
If this vector is rotated anti-clockwise by an angle (p about the z axis,
the new co-ordinates of the point can be expressed in terms of the
original co-ordinates and the angle (p.The z co-ordinate does not change
during the rotation. The new x (x') and y (y') co-ordinates can be
calculated from the projection of a onto the xy plane. They are

x' = x cosq -y sin(p

y' = x sinmp + y cosp

Z =z

These can be written as a product of a 3x3 matrix and a 3x1 column

matrix containing the original co-ordinates. Therefore, the new
position vector of the point can be written as the product of a matrix
and the original position vector.

x' cosp -sinp 0 x

y' = sin cosp OY0

rx''cos -sin(p 0 x
(i jk)y =(ij k) sinp cosw O Y
z ' 0
O 1 z

a' = D(T)a

The matrix D(T) is called the transformation matrix (operation) and in

this case the transformation is rotation by an angle p.
Similarly, symmetry operations and combinations of symmetry
operations in a point group can be relatively simplified by turning to
matrices. For a summary of matrices and their properties see the
appendix.
We again use the water molecule Cartesian co-ordinate system as
an example. The molecule lies on the XZ plane (a) containing the
rotation axis C2 along the Z axis. The H, O and H atoms can then be
represented by Cartesian co-ordinates (x1,yi,zl), (x2,y2,z 2) and (x3,y3,z,)

60
 -

C2
Oyz ayZ
N Fxz

Fig. 4.19. Water molecule in three Cartesian co-ordinate systems.

centred at H, O and H atoms respectively. Now we can consider the

effect of identity operation (E) on the molecule (Fig. 4.19).

Identity operation

x1 X1

Y1 Y1
Z, Z1
3C, X2
Y2 Y2
Z, Z2
X, X3
Y3
Y3
y3
Z,
.Z,- _z3 _

The operation does not transform any of the co-ordinates and therefore
they remain the same. Now let us consider the Cartesian displace-
ments by the operations C2 , az and Gy~ on the molecule. The effects of
the operations on the co-ordinates are shown below.

61
 C2
-
x1 -X3 X1 rXl -X 3

Y1 -Y3 Y1 -y 1 Fx1 -I Y3
-Yl
Z1 z3 Z1 Yz Z3

X2 -X2 X2 Zl x2
3C2
Y2 -> -Y2 Y2 -Y2 Y2 Y2
Z, Z2 Z2 Z2
I z2 Z2
X3 -xi X3 X3 -x 1

Y3 -Y1 YZ3 -Y 3 Y1
_Z3 L Z3 L Z3 Z3 Z1 -

If the new co-ordinates after each transformation are represented by a

column matrix (X 1,Y 1,Z,X 2,Y 2,Z 2,X 3,Y 3,Z 3), then this column matrix
representing each transformation can be written as the product of a
9x9 square matrix containing elements 0, 1 and -1; and the column
matrix representing the co-ordinates of the atoms before the
transformation.

Matrix representation for identity (E) operation

D(E): Matrix A

Xi 100000000 xr
Y1 010000000 Y1
Z 001000000 Z1

X2 000100000 X2
Y2 000010000 Y2
Z2 000001000 Z2
X3 000000100 X3
000000010 Y3

_
z3 _ _
000000001 _ _ _

62
Matrix representation for C2 operation

D(C2 ): Matrix B

X1 0 0 0 0 0 0 -1 O 0 x1
Y1 0 0 O 0 000 -1 0 Y1
Z1 0 0 0 0 000 0 1 Z1

X2 0 0 0 -1 000 0 0 X2

Y 0 0 0 0 -1 0 0 0 0 Y2
Z2 0 0 0 0 0 1 0 O 0 Z2
X3 -1 0 0 0 000 0 0 X3

Y3 0 -1 0 0 000 0 0 Y3

Z3 0 0 1 0 000 0 0 Z3

Matrix representing oy operation

D(3,,): Matrix C

X, 1 0 0 0 0 00 0 0 x1
Y, 0 -1 0 0 0 00 0 0 Yl
z1 0 1 0 0 00 0 0 Z1

Xy2 0 00 1 0 00 0 0 3C2
00 00 -1 0 0 0 0 Y2
z3
Y3
00 00 0 1 0 0 0 Z2
X3 00 00 0 01 0 0 X3
Y, 00 00 0 00 -1 0 Y3

4, 00 00 0 00 0 1 Z3

63
Matrix representation for a, operation

D(az): Matrix D
Xi 0 0 0 0 0 0 -1 0 0 x1

Y, 0 O O 000000 1 0 Y1
Z1 0 0 0 0 000 1 Z1

X, 0 O0 -1 0 0 0 0 0 X2

Y2 0 O 0 1 0 0 0 0 Y2
Z2 0 O 0000 1 000 Z2
X3 -1 0 0 0 0 0 0 0 0 X3

3 0 1 0 0 0 0 0 0 0 Y3
z3
-Z' 00 1 0 0 0 0 0 0 _Z3

A group containing the above four matrices A, B, C, D can be shown to

obey the multiplication table shown in Table 4.5 and are homomor-
phous to the C2, point group containing the symmetry operations E, C2,
a and a'.
The dimension of the matrices representing the symmetry operations
depends on the basis selected. In the above case, we have selected
Cartesian co-ordinates as the basis and therefore the matrices are 9-
dimensional.

4.4 CHARACTER OF THE SYMMETRY OPERATIONS

The symmetry operations have some symmetry characteristics. These

characteristics can be deduced from the characteristics of the matrices
representing the symmetry operations. One such characteristic is the
character of a matrix representing a particular symmetry operation.
The character is the sum of the diagonal elements (trace) in the matrix.
For example, the character of the matrix representing the identity
operation is 9. The characters of the matrices representing the sym-
metry operations C2, a(X) and 'a,z)are -1, 3 and 1, respectively. We have
used Cartesian co-ordinates as the basis for this representation and
therefore the set of characters of the symmetry operations is called the
Cartesian representation. This is written as
C 2, E C0 - 1a
r 9 -1 3 1

64
4.5 CLASSES OF OPERATIONS

Two operations P and Q of a symmetry group are said to belong to a

class if there is a third operation in the group such that

RPR-1 = Q where RR -1 = E

We say that Q is the similarity transform of P and that Q and P are

conjugate. Conjugate operations fall into the same class. It can also be
shown that the operations belonging to a class has the same character.
The ammonia molecule belongs to the C3v point group and has the
symmetry operations E, C3 , C32, I'v V", "V. By using Fig. 4.20a, we can
show that the operations C3, C3 2 belong to a class(Eq. 4.5) and ac'v, v,

Y Y3
t
t,I 3 ACJ -'2 - v ?

, I r,

(a)
1
-1L --
QD' X,

Y I I C3 X,

Y2

C3
(c)

Fig. 4.20. A two-dimensional projection of ammonia molecule and symmetry operations.

65
a"'v belong to another class (Eq. 4.6). We know that the inverse of a
mirror reflection ('v) is the same reflection ('v). Making the reflection
twice leaves the system unchanged. Let us now consider the operation
2
C3 followed by 'v,and C3 . The result is equal to the operation (a"v (Eq.
4.6).
G'v C3 o'v = C3 2 (4.5)
The operations C32 and C3 are conjugate and belong to the same class.
Similarly, the operations ('v, cy"v and "'v can be shown to belong to
another class.

C32 'v C3 = ca", where C3 is the inverse of C32 (C32 C3 = E) (4.6).

Therefore the symmetry operations of the C,3 point group can be simply
written as E, 2C3 and 3Gv. We can also work out the characters for the
symmetry operations in the Cartesian representations. A two-
dimensional projection of the molecule is shown in Fig. 4.20. The z-axis
is chosen as the principal axis passing through the nitrogen atom
perpendicular to the plane of the paper. A C3 rotation will be in the anti-
clockwise direction for the reader. The effects of operations C3 and a'v
are shown in Fig. 4.21. The effects of operations on the co-ordinates and
the matrix representations of the operations are shown below.

The identity operation E

X1 X,

Y1 Y,
Zi Z1
X2 X2
Y2 Y2
Z2 Z2
X3 X3
y3 Y3
z3
Z4 X4
y4
z4
_Z,

66
 C3

````\

Fig. 4.21. The effect of C3 and Y'v operations on ammonia molecule.

The matrix representation for the identity operation

D(E): Matrix A
100000000000
0 1 0 0 0 0 0 0 0 0 0 0
0 1 0 0 0 0 0 0 0 0 0
000100000000
000010000000
000001000000
000000100000
000000010000
000000001000
000000000100
000000000010
000000000001

67
The C3 operation

X, -X 3 cos 60 - Y3 cos 30
Y, X3 sin60-Y 3 sin30
Z, Z3
X, -X, cos 60 Y1,cos 30
Y2 X1 sin60 -Y, sin30
Z, Z1
X, -X 2 cos60-Y, cos 30
Y3 X2 sin60-Y sin30
Z3 Z2
X, -X 4 cos60-X4 cos30
Yll X 4 sin60 -Y4 sin30
z1 Z4

The matrix representation of C3 operation

D(C 3 ): Matrix B

0 0 0 0 0 0 -1/2 -3/2 0 0 0 0
0 0 0 0 0 0 -/3/2 -1/2 0 0 0 0
0 0 0 0 0 0 0 0 1 0 0 0
-1/2 3 /2 0 0 0 0 0 0 0 0 0 0
3/2 -1/2 0 0 0 0 0 0 0 0 0 0
0 0 1 0 0 0 0 0 0 0 0 0
0 0 0 -1/2 -,3/2 0 0 0 0 0 0 0
0 0 0 J3/2 -1/2 0 0 0 0 0 0 0
0 0 0 0 0 1 0 0 0 0 0 0
0 0 0 0 00 0 -01/2 /3/2 0
0 0 0 0 0 0 0 0 0 -,r3/2 -1/2 0
0 0 0 0 0 0 0 0 0 0 1

68
The c v operation

Xi -- xI

Y, Y,
Z1 Z1
X2 -X 3

Y2 13

Z2
X3 -x 2
Y3 Y2
z3 Z2
X4 -X 4

y4

Z4 Z4

The matrix representing the v operation is:

D(cv): Matrix C

--1 00 0 0 0 0 0 0 0 0 0
0 1 0 0 0 0 0 0 0 0 0 0
O 0 1 0 00 0 0 0 0 0 0
0 O O O O O -1 0 0 0 0 0
0 O O O 000000
O 1 0 0 0 0
0 O O O 0000000
O O 1 000
0 O O -1 0 0 0 0 0 0 0 0
0 O O 0 1 0 0 0 0 0 0 0
0 O O O 0 1 0 0 0 0 0 0
0 00 0 00 0 0O 0 -1 0 0
0 O O O O 000000000
O O O 1 0
0 0 0 0 0 0 0 0 0 0 0 1

69
The characters of the operations in Cartesian representation are

C3v, E 2C3 3c7v

F 12 0 2

4.6 REDUCIBLE AND IRREDUCIBLE REPRESENTATIONS

We mentioned earlier that the characters of matrix representations

depend on the basis selected. For example, Cartesian representations
of molecules belonging to a point group would give different characters
depending on the number of atoms. For example, H 2 0 and SO2
molecules give representations consisting of 9x9 matrices each; CH 2 0 a
representation consisting of 12x12 matrices; and CH2 C12 a representa-
tion consisting of 15x 15 matrices.
All these representations formed by these matrices are reducible. To
illustrate this, we shall use the matrix representations of the symmetry
operations for the water molecule in the C2,, point group using only x co-
ordinate vectors as the basis for the representation. Matrices A, B, C
and D representing the symmetry operations E, C2, oy2and z,,will then
be (the reader can work this out):

Fl 0 OJ 0 00 FO - -
A 0 1 0 B= 0 -1 0 C= o 1 0 D= 0 -1
001 0- 0 0 0 -1 0 0

The characters for this representation are 3 -1 3 -1. The above

matrices can be similarity transformed using a 3x3 dimensional matrix
P such that P- 1AP = A', P-1BP = B', P 1 CP = C', and PDP = D'. The
transformation leads to non-zero elements in the leading diagonals of
the matrices A', B', C' and D'. The characters of these matrices are the
same as the matrices A, B, C and D, respectively (see Appendix III, P).
The similarity transformation (reduction) can be continued with A', B',
C' and D' until a matrix P is not found to satisfy the above trans-
formations. The resulting matrices A', B', C' and D' will have non-zero
diagonal elements. The similarity transformed matrices A', B', C' and
D' are:

70
 (1 1 1 1)

1 0 0 1 O1
0 10 0 1 0 0
A'= 0 1 0 B'= 0 -1 0 C'= 0 1 0 D'= 0 -1 0
0 1 0 -1 0 1 0 0 -1

(1 -1 1 -1)
The corresponding diagonal elements of the matrices A', B', C' and D'
form representations that are called irreducible representations.From
the above matrices we have 1 1 1 1, and two 1 -1 1 -1 irreducible
representations.

4.6.1 Irreducible representations and character tables

A molecule belonging to a point group can have infinite numbers of

representations. But all these can be reduced to a combination of a set
of irreducible representations. Each point group has a unique set of
irreducible representations and they are presented in the character
table (see Tables 4.5 and 4.6) of the point group.
The character tables (Tables 4.5 and 4.6) need some explanation.
The top left corner of the table shows the symbol for the point group.
Different classes of operations of the point group are given on the top
row in the second column. The number appearing in front of the
operations indicates the number of group elements (equivalent
operations) in the class. All the operations belonging to a class have the
same character. The irreducible representations are labelled by the
Mulliken symbols A1, A2, B1 and B2 in the first column of the character
table. The following describes their meanings and other symbols we
will be encountering later in character tables of different point groups.
A collection of character tables is given in Appendix 2.
1. One-dimensional irreducible representations (they have character
1 or -1) are labelled as A or B. The symbol A is used for the
irreducible representation that is symmetric with respect to
rotation about the principal axis (z-axis that has the highest order of
rotation). The symbol B is used if the representation is anti-

71
TABLE 4.5
Character table of the point group Cz2

C2v E C2 xz %z
A1 1 1 1 1 X2 y22

A2 1 1 -1 -1 Rz xy
B1 1 -1 1 -1 x RX xz
B2 1 -1 -1 1 y Ry yz

TABLE 4.6
Character table of the C3v point group

C3v E 2Ca 3v _
Al 1 1 1 Z X2 +y2,2
A2 1 1 -1 Rz
E 2 -1 0 (xy) (Rx,Ry) (x2 - y 2,xy)(xz,yz)

symmetric with respect to rotation about the principal axis. Two-

dimensional representations (doubly degenerate) are labelled as E
and three-dimensional (triply degenerate) as T.
2. The subscripts 1 and 2 are used respectively depending on whether
the representation is symmetric or antisymmetric with respect to a
C 2 rotation axis lying in the plane perpendicular to the principal
axis (or with respect to a vertical plane if the C2 axis is lacking).
Primes (') and double primes (") are used respectively depending on
whether the representations are symmetric or antisymmetric with
respect to a horizontal plane of symmetry. Subscripts g (gerade-
even) and u (ungerade-uneven) are used respectively depending on
whether they are symmetric or antisymmetric with respect to
centre of inversion (i).
3. The use of subscripts in two-dimensional and three-dimensional
irreducible representations also follows certain rules but we will
take them as given in character tables.

The character table also shows the irreducible representations some

directional properties belong to. Their meanings are as follows

72
1. The transformation properties of the rotational modes of a molecule
R, R and Rz belong to irreducible representations of the group to
which the molecule belongs. These are classified under the
respective irreducible representations to which they belong.
2. The co-ordinates x, y and z in a Cartesian system or dipole moment
operator (p) or translation operator (T) transform in the same way
as the irreducible representations under the symmetry operations
of a group. The transformation properties of the Cartesian co-
ordinates under the operations of a group can be easily determined.
For example in C2 v, an x co-ordinate under the operations E, C2, ,
%z transform into x, -x, x and -x (Fig. 4.22). Therefore, the char-
acters of this one-dimensional representation are 1, -1, 1, -1 which
are those of the irreducible representation as B 1. Similarly the co-
ordinates y and z can be shown to span the irreducible repre-
sentations B2 and A.
Binary combinations of the co-ordinates (x2, y 2 , z 2, X2 -y 2, xy, xz and yz)
are also classified in the same way and placed in a separate column.
The degenerate pairs are given in brackets. We shall be using these
properties in determining whether a vibration is infrared active or
Raman active. Binary combinations can be calculated from the
irreducible representations of the co-ordinates x, y and z. For example,
in C2v, the characters of the combination x 2 -y 2 (x.x-y.y) can be calcu-
lated by using the characters of the co-ordinates x (x spans A with
characters 1, 1, 1, 1 for the operations) and y (y spans B 2 with charac-
ters 1, -1, -1, 1). The result 1, 1, 1, 1 leads to the classification ofx 2 -y2
under A.

4.6.2 Reducing representations of a point group

The process of reducing representations can be achieved by using

matrix representations of the operations as shown in Section 4.6. For
example, in the case of a water molecule, which belongs to the C2,, point
group, the matrices representing the E, C2 , =,and %o,operations can be
transformed into another set of representations by using similarity
transformations. If the matrices representing these operations are A,
B, C and D (as shown above), and if there exists a matrix P and its
inverse p-1 such that PAP = A', P-BP = B', P-1 CP = C' and Pr-DP =
D' then these can be transformed into a new set of matrices A', B', C'
and D' representing the same operations. Their characters remain the
same (see appendix on matrices). The transformation is repeated until

73
 Z
Z

C2
.y

---------- - x
~X .*--------
I
(a) -Y

Y Y

i---------- X

(b)
I

z
XZ

z z

Y ------ Y
I -

-~- ......-

(c)

Fig. 4.22. The transformation properties of the Cartesian co-ordinates under symmetry
operations.

all the matrices representing the operations are blocked out as

matrices containing non-zero diagonal elements. When such matrices
are obtained, each set of corresponding diagonal elements (a'/i, b'ii, c'ii,
and d'ii) will belong to one of the irreducible representations of the
group. Then the reducible representation can be written as the sum of
the irreducible representations.

74
 Usually, similarity transformation of matrices involves long and
complicated process involving matrices. However, the number of
irreducible representations that form a reducible representation can be
calculated using the formula below and the character table of the
relevant group.

Ni =(l/h) E NgX(R)Xi(R) (4.7)

Over all
classes

where N = number of times each irreducible representation i appear in

the reducible representation; h = order of the group-number of
distinguishable symmetry operations in the group; Ng = the number of
operations in each class; (R) = the character of the reducible repre-
sentation for the operation R; Xi(R) = The character of the irreducible
representation i for the operation R.
We shall illustrate the reduction of the Cartesian representation of
water molecule belonging to C2v point group (see Section 4.4).
The character table for the point group is shown in Table 4.5. By
considering the number of operations in each class (the number shown
in front of each operation), we can calculate how many times the
irreducible representation A appears in the above reducible
representation as follows:

N i = (1/h)YNg Xi(R)X(R)

1.E 1.C2 1.a l.cYyz

r 9 -1 3 1

A, 1

N(A1 ) = 1/(1+1+1+1)} [Ng(E)Xi(E)(E) + Ng(C 2) Xi(C 2 )X(C2 ) +Ng(xz)

Xi(xz) X(xz) + Ng(yz) Xi (Cyz)X( yz)]
5

= {1/4) [1.1.9 + 1.(-1).1 + 1.3.1 + 1.1.1]

= {1/41[9 - 1 + 3 + 1] = 3

75
 .E 1.C2 1.a_ l.a

r i9 -1 3 1

N(A2 ) = 1/4[Ng(E) xi(E)x(E) + Ng(C2 ) Xi(C 2)x(C2) + Ng(Qz) Xi(=) X(o=) +

Ng(Oyz) Xi(yZ) X(Oyz)
= 1/4[1.1.9 + 1.(1)(-1) + 1.(-1).3 + 1.(-1).1]
= 1/4[9 - 1 - 3 - 1] =

N(B 1 ) = {1/4 [Ng() (E) (E) +Ng(C 2) zi(C2)z(C2) +Ng(cz) Xi(o,) &() +
Ng(%yz) i(yz) X(%yz)]
= {1/41[1.1.9 + 1.(-1).(-1) + 1.1.3 + 1.(-1).1]
= t1/4)[9 + 1+ 3 -1] = 3

1.E 1.C2 l.a= L%,

F 9 -1 3 1

B2 1 -1 -1 1

N(B1 ) = {1/4} [Ng(E) Xi(E)X(E) +Ng(C2) zi(C2)X(C 2) +Ng(5,) Xi(Tzz) X(xz) +

Ng,(yz) Xi((Cyz) X(Gyz)]
= {1/41[1.1.9 + 1.(-1)(-1) + 1.(-1).3 + 1.1.1]
= {1/4}[9 + 1- 3 + 1] = 2

76
These reductions show that the Cartesian representation can be
reduced as linear combination of irreducible representations as

F=3A1 +A 2 +3Bl+2B2 (4.8)

Similarly, using the C3Vcharacter table, the reducible Cartesian repre-

sentation of ammonia molecules can be written as linear combination
of the irreducible representations Al, A2 and E.

1.E 2.C3 3.o3

Fr 12 2
A01 1

N(A1 ) = [1/(1+2+3)] [Ng(E) xi(E) X(E) + Ng(C3)Xi(C3) x(C 3) + Ng(ov) Xi(ov)

X(Cv)]

= 1/6)[1.1.12 + 2.1.0 + 3.1.2] = 1/6[12 + 6]

=3

1.E 2.C3 3.%y

r 12 0 2

A2 11 -1

N(A 2) = 1/(1+2+3)] [Ng(E)Xi(E)X(E) + Ng(C3)Xi(C3)X(C 3) +Ng(Gv) Xi(Cv)

X(ov)]
= 1/6}[1.1.12 + 2.1.0 + 3.(-1).2] = 1/6[12 - 6]

=1

77
 1.E 2.C3 3.c v
F i12 0 2

E 2 0

N(B1 ) = {1/(1+2+3)} [Ng(E)Xi(E) x(E) +Ng(C3)xi(C 3)x(C 3) + Ng(ayv) Xi(v)

X(Yv)]

= {1/6}[1.2.12 + 2.(-1).0 + 3.0.2] = 1/6[24]

=4

Therefore,

F=3A,+A2 +4E (4.9)

The total dimensions of the representations are equal to the number of

co-ordinates of the molecule.

4.6.3 Determining characters of operations in Cartesian

representations to obtain reducible representations

Applications of symmetry to molecular vibrations require: (1) the

determination of the group to which the molecule belongs; (2) selection
of a suitable basis for the representation of the operations; and (3)
determination of the characters of the different operations with respect
to the selected basis.
The identification of the group to which the molecule belongs was
discussed in Section 4.1.3. There are several different ways to select the
basis for the representations. However, we will restrict ourselves to
Cartesian representations. Determination of the characters of different
operations of a group on a molecule is the tricky part which needs some
simplification. In the examples shown above for water and ammonia,
the use of Cartesian representations resulted in matrices of dimensions
3Nx3N (N = number of atoms in the molecule). Working with matrices
of large dimensions is difficult. However, we could follow certain rules
that could make the character determination relatively simple: (1) only
those atoms that are not shifted during a symmetry operation
contribute to the characters (these are the atoms that contribute with

78
non-zero diagonal elements in the matrices; (2) it follows that the
identity has a character equal to 3n.
Therefore, it is enough to look for atoms that are not shifted during a
symmetry operation and sum up the numbers representing the
coefficients of the co-ordinates after the operation.

4.7 APPLICATION TO MOLECULAR VIBRATIONS

4.7.1 Vibrational motion, infrared and Raman spectra

We learnt in our earlier discussions that vibrational motions of a

molecule lead to infrared and Raman spectra of the molecule.
The infrared absorption arises when the vibrational motion of a
molecule produces an oscillating dipole. The oscillating dipole interacts
with the electromagnetic radiation of the same frequency and absorbs
it. The infrared spectrum measured by irradiating a sample with
polychromatic radiation in the range 4000-250 cm-l contains the
absorption patterns of such vibrations.
The Raman activity arises from the inelastic scattering of photons
from the radiation source by a sample. When a sample is irradiated by
monochromatic radiation of visible light, the spectrum measured in the
mid-infrared region will contain lines with different intensities. When
photons collide with molecules of the samples, some photons lose a part
of their energy and are scattered as radiation with less energy (with
longer wavelengths). These are called Stokes lines. On the other hand
molecules, which are already in an excited state, lose energy and the
photons absorb this energy and the scattered radiation will have higher
energy and would give lines at shorter wavelengths. These are called
anti-Stokes lines. A part of the incident radiation may also pass
through the sample without any change in the wavelength. This is
called Rayleigh radiation.The infrared region is less energetic than the
visible region and Stokes lines are lines with longer wavelengths than
the excitation radiation. Therefore, the Raman spectrum measured in
the mid-infrared region contains only Stokes lines.
When a molecule is exposed to electromagnetic radiation, the
electric field component of the radiation distorts the electron distri-
bution in the molecule. This distortion induces a dipole moment that is
proportional to the electric field E (Eq. (4.10)). The proportionality
constant is called polarizability.

79
p = aE (4.10)

The polarizability ca is a scalar quantity when pi and E are parallel.

Otherwise, it is a tensor quantity with nine components:

xx axy xz
IL= C ay yy ay z E
zx zy ( zz,

The polarizability a may oscillate during a vibration and hence the

induced dipole moment. The general selection rule for the Raman
activity is that the molecule should have an oscillating polarizability.

4.7.2 Vibrational motions and their symmetries

We have seen earlier that the internal motions of a molecule containing

N atoms can be described by 3N-6 degrees of freedom (or 3N-5 degrees
of freedom for linear molecules). These are called fundamental
vibrations or normal modes of vibrations or normal modes. Some of
these are infrared active, some are Raman active and some are both
infrared and Raman active.
The Cartesian representation of a molecule provides us with a
reducible representation, that can be reduced to a combination of the
irreducible representations representing all the modes of the molecule
including translation and rotation. In the case of the water molecule,
the Cartesian representation is reduced to a linear combination of
irreducible representations as

F = 3A, +A + 3B + 2B,2

The water molecule has three translational modes and three rotational
modes. A look at the character Table 4.5 shows that the translations
along the X, Y and Z axes span B, B2 and Al, respectively and rotations
about the X, Y and Z axes span B1 , B2 and A 2, respectively. By sub-
tracting these species from the total representation, the modes
representing the normal modes of vibrations can be determined.

80
Total representation: 3A, + A 2 + 3B, + 2B 2
Translational modes: Al + B, + B2
Rotational modes: A 2 + B1 + B2
Normal modes: 2A1 +B1
Two of the three normal modes of the water molecule are totally
symmetric.

4.7.3 Wavefunctions representing vibrational motion, and

their symmetries

In quantum mechanics the vibrational states of an electronic state of a

molecule are described by wavefunctions. Any vibrational wave-
function of a polyatomic molecule is a product of the wavefunctions of
the normal modes. The wavefunction of the excited state of a molecule
is also the product of the wavefunctions of all the modes of motion of the
molecule. It means that the total wavefunction describing the motion of
a molecule with N atoms in an excited state can be written as the
product of 3N-6 wavefunctions (or 3N-5 wavefunctions for linear
molecules).
P = 91929393 ... 93N-6 (4.11)
The wavefunction of the molecule in the lowest vibrational state is
similarly composed of 3N-6 wavefunctions. These wavefunctions are of
2
the gaussian type (= Ce (1 )Y2 ).
The wavefunctions describing the normal modes posses a symmetry
related to the symmetry of the normal mode. The wavefunction of a
vibrational ground state is totally symmetric (it involves squares of the
co-ordinates). In the case of a water molecule, the wavefunction
representing the vibrational ground state is of the symmetry type Al.
The symmetries of the wavefunctions representing the singly
excited vibrational states are the same as the symmetries of the normal
modes. Therefore, the symmetries of the wavefunctions of the singly
excited vibrational states of the water molecule are A1 , A1 and B2.

4.7.4 Symmetry and infrared absorption

When a molecule absorbs infrared radiation at normal temperatures

the molecule is excited from the vibrational ground state to the first
excited state (Av = 1 v = 1 - v = 0 and the absorption pattern of the

81
fundamentals falls in the mid IR region. The excitation of a
fundamental vibration involves transition dipole moment (transition
moment) which is evaluated by the integral (Eq. (4.12)) involving the
wavefunctions of the ground and excited vibrational states (pg and (Pe)
and the dipole moment 1p of the molecule. The absorption of infrared
radiation takes place only if the transition moment has a non-zero
value.

Transition moment I = p gPp ed (4.12)

pg is the conjugate wave function of pg.

The dipole moment 11 of a molecule arises from the charge
distribution in the molecule and can be resolved into three components
along x, y and z directions.
= + y + Pz (4.13)

PPed =(g(i +iy +plz) ped (4.14)

I=- E JpPi(pdr (4.15)

i-x,y,x

If one of the above three components has a non-zero value then the
normal mode is infrared active. For this to happen, the direct product of
the pg, li and (p should span the totally symmetric irreducible
representation. Since, the vibrational ground state spans a totally
symmetric irreducible representation, the requirement will be met if
the direct product between p (ii = x or y or z) and (Pe spans the totally
symmetric irreducible representation. In other words i (i = x or y or z)
and (Pe must span the same irreducible representation. The dipole
moment p is a vector quantity and the components Px, Py and pz span
the same irreducible representations as the translation co-ordinates
(Fig. 4.22) x, y and z, respectively. This means that if the normal mode
spans the same irreducible representation as one of the translational
co-ordinates then the mode is infrared active. This is true also for the
doubly degenerate (Table 4.7) and triply degenerate irreducible repre-
sentations because the direct products of all representations with
themselves contain the totally symmetric representation. For example,
in the case of point group C, the doubly degenerate species has

82
 '7

a)
b) Ct
_

= 3651 cm- V2 = 1595 cm -1

Al Al

, C)
c)
V3 = 3755 cm-

B2

Fig. 4.23. Symmetric and asymmetric stretching of water molecule.

TABLE 4.7
Direct product of doubly degenerate irreducible representations
E 2C 3 3a
E 2 -1 0
E 2 -1 0
ExE = F 4 1 0

characters x(E) = 2, X(C3) = -1 and X(ov) = 0. The direct product of the

characters r can be reduced again by using Eq. (4.7) as illustrated in
the examples above to
r =A 1 +A 2 + E (4.16)

83
4.7.5 Symmetry and Raman activity

An argument similar to the above can again explain the Raman activity
in a molecule. The polarizability is a tensor property and is expressed
by a matrix containing 9 elements as shown below. The transition
moment during absorption can be written as in Eq. (4.15)

I = (P
9 gedr (4.17)

I = (pgaE(p dT (4.18)

a; a a E-
I= ayx ayy a yz Ey (PedT (4.19)
(X Cy O z_ Ez

Because of symmetry axy = a,ycayz = azy and a = a,,. For Raman activity,
the above integral must be non-zero. This is true if one of the compo-
nents of the integration is non-zero. An argument similar to infrared
activity can be made here to determine whether one of the components
is non-zero and hence Raman activity.
This means that if one of the components of a spans the same
irreducible representation as (Pe then the mode is Raman active.
In the above sections, we have simplified the process of identifying
whether a normal mode is infrared active or Raman active. They can be
summarised as follows:
1. Identify the symmetry group to which the molecule belongs.
2. Develop the Cartesian reducible representation.
3. Reduce the representation as a linear combination of irreducible
representations of the group.
4. Identify the irreducible representations spanned by translational
and rotational modes.
5. Identify the irreducible representations spanned by the normal
modes.
6. Use the character table of the group and decide on whether these
normal modes span the same irreducible representation as one of
the normal co-ordinates or their product functions and hence
whether they are infrared active or Raman active.

84
4.7.6 Measured spectrum and band assignments

The above procedure can tell us the number of bands that might arise
when the infrared or Raman spectrum of the compound is measured.
However, they do not say anything about their assignments in the
spectrum. A procedure that could determine the symmetry types of the
different absorptions in a measured spectrum would ease the assign-
ments of the bands to different modes. Furthermore, the vibrational
modes of a molecule are determined using harmonic consideration of
the vibrations. These determinations can be found elsewhere [1].

4.8 EXAMPLES

4.8.1 Water and nonlinear molecules with the general

formulae BAB

We have used the water molecule as an example in our earlier dis-

cussions. We shall find out whether the three remaining modes of
motion representing the vibrations of the water molecule are infrared
or Raman or both infrared and Raman active. We found the symmetries
of the vibrations to be 2A1 + B2.
These are two completely symmetric modes and asymmetric modes
as shown in Fig. 4.23. A look at the character table for the C2v point
group (Table 4.5) shows that the translational co-ordinates z and y
transform in the same way as irreducible representations A, and B2,
respectively. Therefore, these modes are infrared active. Furthermore,
the product combination of the translational co-ordinates x 2-y2 (and z 2)
and yz transform in the same way as the irreducible representations A,
and B 2 , respectively. Therefore, the modes are Raman active. All three
normal modes of the water molecule are both infrared and Raman
active. The bands in the infrared spectrum coincide with the bands in
the Raman spectrum.
The infrared and Raman spectra of water in a gaseous state contain
three bands at 3755, 3651 and 1595 cm -1 . The number of bands is in
agreement with our theoretical prediction. However, these do not tell
us which of these two bands represents vibrations of A, symmetry.
Theoretical calculations show that the Raman scattered light are polar-
ised by the symmetry modes of type A. Therefore, by examining the
Raman scattered light with a second polariser, the bands can be
classified into different symmetry classes (Fig. 4.23).

85
TABLE 4.8
Fundamental vibrations of water molecule (in gaseous phase) and their characteristics
Frequency in Symmetry type Label Assignment
1
wavenumber ()/cm

3651 A,(totally symmetric) VI Symmetric stretch

1595 Al (totally symmetric) v2 Bending

3755 B2 (nonsymmetric) V
3 Asymmetric stretch

TABLE 4.9
The normal modes of some nonlinear molecules with the molecular formulae BAB
Symmetric stretch 7l Symmetric bending Asymmetric stretch
cml (Al) V2,cm-1 (Al) V3 cm-1 (B2)
IR & Raman IR &Raman IR & Raman

D2O (gas) [2] 2671 1178 2788#

H2S (gas) [3] 2615 1183 2627
H2Se (gas) [4] 2345 1034 2358
NO, (gas) [5] 1318 749 1610
ClO 2 (gas) [6] 943 445 1110

The absorption bands are labelled as v1,v2,v3, ... vn etc. in the

decreasing frequency order according to their symmetries (see Table
4.8 for water molecule). Note that it is customary in mid-infrared
spectrometry to give the frequency in wavenumber (v = cv). The
symmetric type bands are labelled first starting from the totally
symmetric type. Degenerate vibrations of mode n are labelled as vna,
,,n,etc. The normal modes of some non-linear molecules with a general
formula BAB are given in Table 4.9

4.8.2 Ammonia and pyramidal molecules with the general

formula AB 3

We can again use the reduced Cartesian representation of ammonia

molecule from Eq. (4.9). There are 12 normal modes. The modes repre-
sented by reducible representation E are doubly degenerate. The nor-
mal modes representing the vibrations can be determined as follows.

86
 a) b)

1
V1 = 3336 cm V2 = 932 cm 1
A, A1

c) d) X

1
V3a = 3414 cmt V4,, = 1628 cm

E E

Fig. 4.24. The normal modes of ammonia molecule.

Total representation: 3A1 + A2 + 4E

Symmetries of translational modes: A +E
Symmetries of rotational modes: A2 + E
Symmetries of vibrational modes: 2A1 + 2E
The molecule should have 3x4-6 = 6 normal modes of vibration. It is in
agreement with the above result where normal mode with symmetry
type E is doubly degenerate. The normal modes are shown in Fig. 4.24.
A look at the C3v point group shows that z and (x,y) belong to the
representations A and E respectively. Therefore, all the four
fundamental vibrational modes are infrared active and give four bands
in the spectrum (two of these are doubly degenerate).
Similarly, the character table confirms that these modes are also
Raman active. Here again the bands in the infrared spectrum are

87
TABLE 4.10
The normal modes of some pyramidal molecules and species with formulae AB 3

Symmetric Symmetric Antisymmetric Antisymmetric

stretch. vi cm-' bend. v2 cm-1 stretch. V,cm-' bend. v4 cm-l
(A1 ) IR & (Al ) IR & (E) IR & (E) IR &
Raman Raman Raman Raman

PH 3 (gas) [7] 2327 990,992* 2421 1121

AsH 3 (gas) [7] 2122 906 2185 1005
[C0 3]- (solid) [8] 939 614 971 489
2-
[S 3] (soln.) [9] 967 620 933 469

*Splitting in v2 is due to Fermi resonance (see Section 4.9.2).

coincident with the bands in the Raman spectrum. The normal modes
of some pyramidal molecules with general formula AB 3 are given in
Table 4.10.

4.8.3 BF3 and planar molecules with formula AX 3

The molecules belong to the D3h point group. The Cartesian represent-
ation of the molecule (Fig. 4.25) gives a reducible representation as
shown below. The characters were derived with the help of Fig. 4.25.

D 3h E 2C3 3C 2 - 2S3 3ov

Frat 12 0 -2 4 -2 2

The Cartesian representation reduces to a combinationA', + A'2 + 3E' +

2A"2 +E". Of these, translational modes spanA"2 + E' and the rotation-
al modes span A' 2 +E". Therefore, the vibrational modes span A'1 + 2E'
+ A" 2. These represent 6 normal modes of vibration (3x4-6). The
character table for the point group D,, suggests that normal modes
with symmetry A" 2 and E' are infrared active because these modes
transform the same way as the translational co-ordinates. The normal
modes with symmetry A'1 and E' are Raman active. Thus the modes
with symmetry E' are both infrared and Raman active. These modes
are shown in Fig. 4.26. The normal modes of some planar molecules
with a general formula AX3 are given in Table 4.11.

88
 C3 Zt
F
Y,
C2

0, F

VI
X,
ZI

-1

I d Xl

-Y,

II .
Fig. 4.25. The effect of symmetry operations of molecules belonging to the D3h point
group.

TABLE 4.11
The normal modes of some planar molecules and species with formulae AX 3

Symmetric Symmetric Antisymmetric Antisymmetric

stretch. v, cm- 1 bend. v2 cm- 1 stretch. V, cm- 1 bend. V4 cm-
(A' l) Raman (A" 2) IR active (E') IR & (E') IR &
active Raman Raman
BH 3 (gas) [10] 2623 1132 2820 1610
AlBr 3 (gas) [11] 228 107 450-500 93
A1F 3 (matrix) [12] 660 284 960 252

4.8.4 Planar molecules of lower symmetry with four atoms

We have discussed the normal modes of pyramidal and planar

molecules belonging to the C3 Vand D3h point groups. If one of the atoms

89
 t Ji

( F
Ae_
v1=888 cm V3a V4 a

A1

h --

v2 =708 cm-l V3 a 3b =1505 cm - V4 a= V4 b =482 cm

t

A2 E E

Fig. 4.26. The normal modes of BF3 .

ofX in the planar molecule AX3 is replaced by Y the symmetry elements

of the molecule reduce and the molecule will belong to the C2v point
group. If Y and Z replace two of the X atoms, the molecule assumes the
symmetry elements of the Cs point group. However, the number of
normal modes remains constant. Furthermore, the character tables for
the C2v and C point groups suggest that all the modes in these
molecules are both infrared and Raman active.

4.8.5 Carbon dioxide and linear molecules with the formula

XYX

The CO2 molecule belongs to the Dh point group. Using the Cartesian
co-ordinates a reducible representation of the molecule can be obtain-
ed. The characters of the operations for the reducible representation
can be determined using Fig. 4.27. The character for the identity
operation E is (E) = 9 (all the co-ordinates contribute). When the

90
molecule is rotated about the principal axis by a small arbitrary angle
4, the new co-ordinates X, Y and Z for each atom will be Yi sin4 + Xi cos4,
Yj cos4 - Xi sino and Z i (i = 1, 2 and 3), respectively. Each of the atoms
will contribute a character of 1+2coso (see Eq. (4.20)) and therefore,
(CQ) = 3(1+2coso).Similarly, for reflection on a vertical plane through
the principal axis X(av) = 3 (see Fig. 4.27d and Eq. (4.21)). The effect of
improper rotation shifts the co-ordinates of the oxygen atoms and,
therefore, it is enough to look at the effect on the carbon atom (Fig.
4.27d). The z co-ordinate changes direction and X(S?,)= -1+2coso. The
operation inversion moves the co-ordinates of oxygen atoms and
therefore, it is enough to look at the character contribution from the
carbon atom. All three co-ordinates are reversed and hence x(i) = -3.
Similarly, the (ooC 2)= -1. The Cartesian reducible representation is
shown in in the table below.
Effect of CO on one atom:
cos sin 0I l
-sino cos 0 Y1 (4.20)
]0 0 1 Z
Effect of reflection ov on one atom:
cos 2 sin2 0 Xl

Z=[sin2o -cos2 0 Y (4.21)

The reducible Cartesian representation is:

Dh E 2CQ ... Coov i 2Sj ........ ooC 2

F t 9 3+6cos 3 -3 -1+2cos -1

Since the molecule has an infinite number of vertical planes and C?,
axes along the principal axis (i.e. the principal axis is of infinite order),
reducing the above Cartesian representation is difficult. However, a
look at the irreducible representations in the character table gives us
some clue as to how the above representation can be reduced (see
Appendix II for table Dh). In order to obtain a character -1+2cos for
the operation 2S?,, the combination must involve 21-, + Hg + - - - . In

91
 Zl

C-.0

a) b)

Principal axis 2 + Ylcos 20

S.o
-.%
'x

Y=Y2 cos -X2 sin O

/ Y2

C) I

Fig. 4.27. The effect of symmetry operations on a carbon-dioxide molecule.

order to get a character +3 for moov operation and -1 for oXC

2 operation,
there must be a combination 2 + + Zg+. The total reducible repre-
sentation reduces to a combination of the irreducible representations
as g + JIg + 2u + + 2u. Of these modes, the translational and rotational
modes span Yu+ + lu and rIg, respectively. Therefore, the vibrational
modes of the carbon dioxide span Xg+ + Eu++ riu. The molecule has four
vibrational modes (note that IIu is a doubly degenerate representation)
which is in agreement with our prediction (3x3 - 5 = 4). The normal
modes of the carbon-dioxide molecule are shown in Fig. 4.28. A look at

92
 Vi=134 cm3- - _
t
v,= 1340 cm' = 2349 cm

yg+ (a) (b)

(o 2 > -9 2.V-Z 667 cm-'

Fl u
+ +
o~.- ) 'V2b= 667 cm -'

+ - (c)

Fig. 4.28. The normal modes of the carbon-dioxide molecule.

TABLE 4.12
Normal modes of some linear molecules and species with formula XYX
Symmetric stretch. v Symmetric bend. v2 Antisymmetric
cm - (Zg+ ) Raman cm - 1 (Il) IR active stretch v3 cm 1 (Zu+ )
active IR active

CS 2 (gas) [13] 658 397 1533

KrF 2 (gas) [14] 449 233 596, 580
[CuCl 2]-(s) [15] 300 109 405

the character table for point group Dh shows that the translational co-
ordinates span u,+ and Iu. The products x2 + y 2 and z 2 span g+. That is,
the anti-symmetric stretching mode and degenerate bending mode are
infrared active and the symmetric mode is infrared inactive but Raman
active. This example also illustrates the mutual exclusion principle for
molecules with a centre of inversion. The normal modes which are
infrared active are Raman inactive and those which are Raman active
are infrared inactive. The normal modes of some linear molecular
formula XYX are given in Table 4.12.

93
4.8.6 Linear molecules with formula XYZ

When one of the atoms ofX is replaced by another atom Z, the molecule
assumes symmetry elements of the Cv point group. The character
table indicates that all the three normal modes are both infrared and
Raman active.

4.9 MEASURED VIBRATIONAL SPECTRA OF MOLECULES

We have learnt in this chapter how to predict the vibrational spectra of

molecules. However, measured spectra often show more bands than the
predicted 3N-6 or 3N-5 fundamental frequencies and, in some cases,
there are fundamental frequencies that cannot be observed in the
infrared spectrum.

4.9.1 More bands due to anharmonicity-overtones and

combination bands

One of the reasons for observing more bands than expected arises due
to anharmonicity of the normal vibrations. The normal vibrations of a
molecule were assumed to be simple harmonic. However, this is not the
case. The bonds between the atoms behave like anharmonic oscillators
and transitions between vibrational ground state and higher levels
become possible. The selection rule for anharmonic oscillators is Av =
+1, +2, +3, .... The transition between vibrational ground state to the
second excited state (v = 2 - v = 0) is called first overtone (second
harmonic) and to the third excited state (v = 3 - v = 0) is called second
overtone (third harmonic). The frequencies of these transitions are not
exact multiples of the fundamental transitions. The frequencies of
these transitions decrease (i.e., V=2- < 2 V_ o)
There is also another possibility for the transition to occur to a
combined level. This can be the sum of the tones such as v, + v2 , v1 + v2
+ V3 , etc. or 2v, + v2 , v1 + 2 V2, etc. or difference tones such as v - v2,
v2 - V,, etc. These transitions normally fall in the near infrared region
of the electromagnetic spectrum. However, some of them can be
observed in the mid-infrared region. In such cases the number of
bands in the mid-infrared region exceeds the number of bands pre-
dicted by group theory.

94
4.9.2 More bands due to Fermi resonance
Two molecular vibrations may interact with each other if they have
frequencies very close to each other (30 cm-'). For example, one of the
fundamental modes and an overtone of another mode or a combination
mode may have frequencies close to each other (accidentaldegeneracy).
These vibrations may interact if their symmetries are the same and the
overtone or combined tone is enhanced. The interacting vibrations split
and the vibration with higher frequency is raised in frequency and the
vibration with lower frequency is depressed about the average of the
two vibrations. This is called Fermi resonance (see Section 3.10 for
treatment of Fermi resonance).
An example of Fermi resonance is the interaction between the
symmetric stretch (I) around 1340 cm - ' and the first overtone (2v2
1334 cm-') of the symmetric bending at 667 cm - in a carbon dioxide
molecule. The symmetric stretch has symmetry g+. The symmetric
bending is a doubly degenerate vibration with symmetry Ig. The first
overtone of this degenerate vibration splits into two sublevels with
symmetry species g+ and Ag. Fermi resonance arises because of the
interaction between the species Xg+ of the symmetric stretching vibra-
tion and the species Eg* of the first overtone (of the symmetric bending
vibration). This interaction results in two bands in the Raman spec-
trum at 1388 cm -1 (the frequency is raised about the average 1340 cm-1 )
and 1286 cm 1 (the frequency is lowered about the average 1340 cm-1 ).

4.9.3 Overlap of rotational spectrum on vibrational spectrum

The rotational spectrum of some small molecules (diatomic and tri-
atomic) falls into the same region as the vibrational spectrum of the
molecule. The fine structure of the rotational spectrum can be seen in
the infrared of the molecule. For example, the infrared spectra of
carbon monoxide (see Fig 2.5) and the ammonia molecule are
accompanied by their rotational spectra. However, this is not a problem
in polyatomic molecules because the rotational bands are very close to
each other and hardly visible in the spectra.

REFERENCES

1. G. Herzberg, Infrared and Raman Spectra. Van Nostrand, New York,

1945.
2. W.S. Benedict, N. Gailar and P.K. Plyler, J. Chem. Phys., 24 (1956) 1139.

95
 3. H.C. Allan and P.K. Plyler, J. Chem. Phys., 25 (1956) 1132.
4. D.M. Cameron, W.C. Sears and H.H. Nielsen, J. Chem. Phys., 7 (1939)
994.
5. R.V. St. Louis and B.L. Crawford, Jr., J. Chem. Phys., 42 (1965) 857.
6. A.H. Nielsen and P.J.H. Woltz, J. Chem. Phys., 20 (1952) 1878.
7. E. Lee and C.K. Wu, Trans. FaradaySoc., 35 (1939) 1366.
8. W. Sterzel and W.D. Schnee, Z. Anorg. Allg. Chem., 383 (1971) 231.
9. J.C. Evans and H.J. Bernstein, Can. J. Chem., 33 (1955) 1270.
10. A. Kaldor and R.F. Porter, J. Am. Chem. Soc., 93 (1971) 2140.
11. I.R. Beattie and J.R. Horder, J. Chem. Soc., A (1969) 2655.
12. A. Snelson, J. Phys. Chem., 71 (1967) 3202.
13. T. Wentink, J. Chem. Phys., 29 (1958) 188.
14. H.H. Classen, G.L. Goodman, J.C. Malm and F. Screiner, J. Chem. Phys.,
42 (1965) 1229.
15. D.N. Waters and B. Basak, J. Chem. Soc., A (1971) 2733.

GENERAL BIBLIOGRAPHY

P.W. Atkins, Molecular Quantum Mechanics. Oxford University Press,

London, 1984.
C.N. Banwell and E.M. McCash, Fundamentals of Molecular Spectroscopy.
McGraw-Hill, London, 1994.
M. Ladd, Symmetry and Group Theory in Chemistry. Horwood Publishing,
Chichester, 1998.
K. Nakamoto, Infrared and Raman Spectra of Inorganic and Co-ordination
Compounds. Wiley, New York, 1978.
J.D. Ronaldson and S.D. Ross, Symmetry and Stereo Chemistry. Intertext
Books, London, 1972
A. Vincent, Molecular Symmetry and Group Theory. Wiley, London, 1977.

96
Chapter 5

Group frequencies and assignments of the

infrared bands

5.1 GROUP FREQUENCIES

Analysis of normal vibrations of a polyatomic molecule is, in general,

complicated because the molecule consists of a number of atoms.
Although the vibrations of a polyatomic molecule can be calculated by
calculating a potential energy and a kinetic energy of a system, as in
the case of a diatomic molecule, the calculation is not straightforward.
The concept ofgroup frequencies helps the analysis of vibration spectra
of a polyatomic molecule. The concept of group frequencies may be
applicable when the amplitudes of nuclei in a particular functional
group are very large in a certain normal vibration, while those of the
other atoms are very small. Infrared spectroscopy is useful for
qualitative and quantitative analysis and structural investigation of a
complex molecule since there are a number of vibrational modes which
can be regarded as group frequencies. Table 5.1 summarizes group
frequencies. Group frequencies observed mainly in Raman spectra
(e.g., S-S stretching vibration) are also shown in this table.
The idea of group frequencies is beneficial even for a very complex
molecule such as protein. As an example, let us consider normal
vibrations of an amide group. Normal vibrations of an amide group
have been calculated in detail, taking N-methylacetamide (Fig. 5.1) as
a model of the amide group [1,2]. Considering a methyl group as one
atom, N-methylacetamide is a six-atom molecule, and hence, has
twelve normal vibrations (3x6 - 6 = 12). Of the twelve, the normal
vibrations shown in Fig. 5.1 are amides I, II and III modes which are
key vibrations for studying the structure of proteins. As can clearly be
seen in Fig. 5.1, amide I has a strong characteristic of C=O stretching

97
TABLE 5.1
Group frequencies

3500 3000 2500 2000 1500 1000 500

I OH stret T
I Casym bend
NH stret CI sissor
CH stret(unsaturated) CIH sym bend
CHstret(saturated) CHl:wag
m SH start _ CHbend I JC= C)
* 01 str tn P=O- stret
IC stret I PO; antisym sret
CLO stret I PO;sym stret
* C=Ostret(COOI)
* -CONH-
i C=Cstret
* C=-Nstret
_ CO, antisym stret
I NO antisym street
*N=N stret
I -CONH-
* N-N stret _ CO2 sym stret
NN atisym stret NO2 sym stret
(c:Na ~ C-N stret
Rr.mtgd(p htA C-C street
rmm~ri~
~ ~C-O street

I I Sis
3500 3000 2500 2000 1500 1000 500

Wavenumber Icni'

vibration. Meanwhile, amides II and III are coupling modes of C-N

stretching vibrations and N-H in-plane bending vibrations. Of the
three, amides I and II appear strongly in infrared spectra, and amides I
and III appear intense in Raman spectra. Amides I, II and III bands of

C\, 3 H
C-N
0t \CH3

Fig. 5.1. Structure ofN-methylacetamide and its amide I, amide II, and amide III modes.
(Reproduced from Ref. [2] with permission. Copyright (1984) Academic Press.)

98
TABLE 5.2
Secondary structures of protein and frequencies of amide I, II, and III bands
Secondary Infrared Raman
structures
Amide I Amide II Amide I Amide III

a-helix 1655-1650 -1540 1660-1645 1300-1265

3-sheet -1690, 1680-1675, -1550 1675-1665 1240-1230
structure -1640, -1630
Random coil 1655-1645 1535-1530 1670-1660 1260-1240
P-turna -1680, -1660, -1680 ,-1660, 1330-1290
-1640 -1640
310helixa 1645-1640 -1655
31helixa -1640 -1550 -1380, -1335,
-1285

aFrom Ref. [4].

proteins are generally found in the range of 1690-1620 cm - l ,

1580-1520 cm l and 1320-1220 cm -1 , respectively. The frequencies of
these modes are known to sensitively reflect secondary structures of
polyaminoacids, peptides, and proteins [3-6].
Table 5.2 shows relationships between secondary structures of
proteins and the frequencies of amides I, II and III. Measurement of
infrared (or Raman) spectra of protein allows us to estimate the
contents of secondary structures, which will be explained in Section
8.7.1.

5.2 ISOTOPE SHIFT

In analysis of infrared spectra, a shift associated with isotopic

substitution (isotope shift) gives solid assignment of bands in many
cases. In general, force constants may be assumed not to change due to
isotopic substitution, and therefore, isotope shifts lead to mass effects
alone. Calculate the magnitude of an isotope shift, taking a diatomic
molecule as an example. The frequency of a stretching vibration of a
diatomic molecule is given by Eq. (3.17). Where v' is the frequency for
replacing an atom having a mass ml with an isotope having a mass ml',
the following relation holds:

99
Fig. 5.2. Amide I', amide II', and amide III' modes of deuterium-substituted N-methyl-
acetamide. (Reproduced from Ref. [2] with permission. Copyright (1984) Academic
Press.)

v= ~ (5.1)

p' = m,' m 2/(m' + m2) holds. As Eq. (5.1) clearly shows, the larger the
difference between ml and m,', the larger is the isotope shift. Since v/v'
= 1.36 if H is replaced with D, a C-H stretching vibration of saturated
hydrocarbons, which appears in the vicinity of 2900 cm l, shifts close to
2100 cm-l. Figure 5.2 shows amides I, II and III modes of deuterium-
substituted (ND) N-methylacetamide (which will be called amides I', II'
and III'). While shifts induced by the deuterium substitution are rather
large in the amide II and III modes as NH bending vibrations con-
tribute to these two modes, amide I mode, being principally a C=O
stretching vibration, gives rise to a very small isotope shift. What
should be noted with respect to isotope shifts of polyatomic molecules is
that vibrational modes change more or less in association with isotopic
substitution, which can be clearly understood from comparison of Fig.
5.1 with Fig. 5.2. In studies of infrared spectra, 15N-substitution, 13C-
substitution and the like are often utilized in addition to deuterium
substitution. Although an isotope shift is small when such a heavy
atom is replaced, a change in a vibrational mode associated with the
isotopic substitution is also small.

5.3 HOW TO MAKE BAND ASSIGNMENTS IN INFRARED SPECTRA

In general, we observe a number of bands in an infrared spectrum of a

molecule. The first consideration to be made when analysing the
infrared spectrum is whether it is necessary to analyze the infrared
spectrum as a whole or only a part of the spectrum. For example, if an

100
objective is to examine a secondary structure of protein, the spectral
regions where the amide I and amide II bands appear may be studied
and it is not necessary to analyze the entire spectrum. On the other
hand, when we want to identify a certain material, we must analyze a
considerable portion of the spectrum.
Although there is no absolute procedure for assignments of infrared
bands, the following methods are often found effective:
1. Find group frequencies from comparison of observed frequencies
with a table of group frequencies. During the comparison, intens-
ities as well as frequencies must be noted.
2. Compare an obtained infrared spectrum with those of similar
molecules. The similar molecules do not always need to be similar in
their entirety, but rather, may be only partially similar.
3. Measure an infrared spectrum of an isotope-substituted material
which contains deuterium, 15N, 13C, etc., and compare the infrared
spectrum with an original spectrum. This method is very effective
for the identification of a band due to a particular functional group,
a particular bond, etc.
4. Measure spectra while varying a condition of a material, such as a
temperature, pH and a solvent, and compare them with an original
spectrum. For example, since amino acid residues within protein,
respectively, have unique pK values, as pH is changed around the
pKvalues, it is possible that only bands due to particular amino acid
residues will change.
5. Measure anisotropy of infrared absorption resulting from polarized
light. This method is convenient to distinguish in-plane and out-of-
plane vibrations of a planar molecule from each other.
6. Calculate normal vibrations. Although this is a traditional method,
if not combined with another scheme, it does not allow us to com-
pletely assign an infrared spectrum of a complicated molecule. This
is because of a general lack of knowledge regarding three-
dimensional structures and force constants of molecules.
7. Try chemometrics and two-dimensional correlation analysis (see
Chapter 9)
There are unique marker bands known, some unique to proteins, some
unique to nucleus acids, etc., with which we can study structures of

101
materials (amides I and II are typical examples of marker bands for
proteins). Hence, for actual analysis of an infrared spectrum of a
molecule, it is often important to first identify marker bands. Marker
bands for organic thin films, polymers, and biological molecules and the
like will be described in Chapter 8. A library search alone is often
sufficient if the objective is merely identification of a material based on
measurement of infrared spectra.
It is very convenient to know which bands appear intense in an
infrared spectrum for its analysis. The well-known principles regard-
ing intensities of infrared bands are as follows:
1. A band due to a functional group with a strong polarity appears
strongly, e.g., OH and C=O stretching vibrations. Conversely, vibra-
tions due to a bond with a weak polarity, such as C-C and S-S
stretching vibrations, appear very weakly or do not appear in an
infrared spectrum.
2. Antisymmetric stretching vibrations are stronger than
corresponding symmetric stretching vibrations. For example, COO-
antisymmetric stretching vibrations appear more strongly than
COO- symmetric stretching vibrations.
3. As a general tendency, local vibrations are strong and vibrations of
a molecule as a whole are weak. For instance, in an infrared
spectrum of polyethylene, while CH 2 rocking vibrations appear
strongly, vibrations in which a molecule as a whole stretches and
contracts (e.g. accordion vibrations) appear weakly.
4. Among bands arising from an aromatic group, those which give
strong infrared bands are ring stretching vibrations in the
1600-1450 cm 1 region and out-of-plane bending vibrations in the
900-700 cm 1 region.

REFERENCES

1. T. Miyazawa and E.R. Blout, J. Am. Chem. Soc., 83 (1961) 712.

2. Y. Sugawara, A.Y. Hirakawa and M. Tsuboi, J. Mol. Spectrosc., 108 (1984)
206.
3. H. Susi and D.M. Byler, Arch. Biochem. Biophys., 258 (1987) 465.
4. S. Krimm, in: T.G. Spiro (ed.), Biological Applications of Raman Spectro-
scopy, Vol. 1. Wiley, New York, 1987, p.l.
5. M. Jackson and H.H. Mantsch, CRC Crit. Rev. Biochem. Mol. Biol., 30
(1995) 95.
6. H.H. Mantsch and D. Chapman (eds.), Infrared Spectroscopy of Bio-
molecules. Wiley-Liss, New York, 1996.

102
GENERAL BIBLIOGRAPHY

L.J. Bellamy, The Infrared Spectra of Complex Molecules, Vol. 1, 3rd edn.
Chapman and Hall, London, 1975; Vol. 2, 2nd edn. Chapman and Hall,
London, 1980.
N.B. Colthup, L.H. Daly and S.E. Wiberley, Introduction to Infrared and
Raman Spectroscopy, 3rd edn. Academic Press, San Diego, CA, 1990.
E. Maslowsky, Jr., VibrationalSpectra of OrganometallicCompounds. Wiley,
New York, 1976.
K. Nakamoto, Infrared and Raman Spectra of Inorganic and Coordination
Compounds, 5th edn. Wiley, New York, 1997.
N.B. Nyquist, The Interpretation of Vapor-Phase Infrared Spectra Group
Frequency Data. Sadtler Research Laboratories, Philadelphia, 1984.
G. Socrates, Infrared Characteristics Group Frequencies. Wiley, New York,
1980.

103
Chapter 6

Instrumentation

6.1 HISTORY OF INFRARED INSTRUMENTATION

Since the discovery of infrared radiation by Sir William Herschel in

1800 [1], a variety of methods have been used to improve the experi-
mental techniques for measuring infrared spectra. In particular,
Herschel used a prism and a mercury thermometer to record his
observations of heat-based radiation beyond the range of the solar
spectrum. Melloni is credited with the construction of the first mid-
infrared spectrometer in 1833, after his discovery of the transparency
of NaCl in the infrared. Excellent references on the early history of
vibrational spectroscopy and the minds that shaped the field can be
found elsewhere. This chapter will deal in detail with the interfero-
metric methods of capturing infrared radiation. Older monographs
treat non-interferometric methods in great detail [2].

6.1.1 History of FT-IR instrumentation

The construction of the first interferometers dates back to 1880. Lord

Rayleigh may have constructed an interferometer at that period, but it
is Michelson who is credited with the construction of the first operable
instrument at that time. The interferometer was used in the measure-
ment of the speed of light in diverse directions. Measurements that
took place at was is now Case Western Reserve University showed that
there was no detectable difference in the speed of light in either
direction. The lack of adequate computing power was the main reason
that it took approximately eighty years for the instrument to utilize its
full potential as an analytical tool. Ferraro has recently published a
very informative account of the history of FT-IR spectroscopy [3].

105
 The first attempts to use interferometric means to measure infrared
radiation concentrated in the far infrared region of the spectrum. The
operational needs for mechanical precision and computational load are
lower in the far-infrared compared to the mid-infrared. Therefore, a lot
of the early applications and advancements took place in the field of
astronomy where far-infrared spectroscopy is used extensively.
One very important recent development that led to the widespread
use of infrared spectroscopy as a characterization tool was the intro-
duction of the first Fourier transform infrared spectrometer, the FTS
14, by the Biorad Company of Cambridge, Massachusetts in 1969.
Several developments in the 1950s and 1960s contributed to the intro-
duction of this first commercially available instrument. The work of L.
Mertz in interferometer design during 1954-56 and the development of
data reduction algorithms in 1960-65 are probably the most significant
contributors. One historical aspect that should not be overlooked was
the role of the NASA contract to Block Engineering for an instrument
with ten times the available resolution. The model 1500 (296 in the
commercial version) had 0.5 cm-l resolution and a better signal-to-
noise (S/N) ratio than the dispersive instruments of the time.
Other notable developments were the discovery of the HeNe laser
along with the introduction of better infrared detectors, analog-to-
digital (A/D) converters and minicomputers. In 1966 a one-foot laser
with a built-in power supply was available to be used in the first FT-IR
spectrometer. In addition, pyroelectric bolometers in the form of the
deuterated triglycine sulphate (DTGS) detector also became available.
Their major advantage was that their bandwidth was compatible with
the rapid-scan frequencies. With respect to computing power there was
a remarkable development from the PDP-1 in 1960 to the DG Nova in
1969.
After the introduction of this instrument many new instruments
from various manufacturers have appeared and tremendous progress
has been achieved in the years following 1969. A list of the advance-
ments will undoubtedly include the very small footprint of the modern
instruments, quadrature detection with forward and backward scan-
ning, digital signal processing, diagnostic features, low powered air-
cooled sources, the flexible design of the research-grade instruments, the
multiple spectral ranges, the very high spectral resolution, and the
tremendous progress in FT-IR software. In addition, one of the most
important developments was the 'rediscovery' of step-scan interfero-
metry, a subject that will be extensively dealt with in this book.

106
 What is the future of infrared instrumentation? Without a doubt,
any technological breakthrough will eventually find its way to a com-
mercial design with time. Improvements in performance, new features
and capabilities, usability and (hopefully!) a reduction in cost should all
be expected.

6.2 COMPONENTS OF AN FT-IR SPECTROMETER

The components of an FT-IR spectrometer primarily include the source

interferometer, the source of radiation, the detector and other optical
elements (beamsplitters, mirrors, etc.). In addition, data manipulation
also takes place in the adjacent computer station. It is beyond the scope
of this book to give a detailed account of all the elements involved;
instead, an attempt will be made to cover in more detail important
components of these designs.

6.2.1 Sources

The source of infrared energy in an infrared instrument does not

depend on the type of instrumentation used to detect the radiation.
Both dispersive instruments and Fourier transformed instruments can
use the same types of infrared sources. Therefore, a general overview of
the available technology will be reviewed here. The more typical
sources are the Globar source and the Nernst glower, even though
nichrome coils have also been used in the past. Nichrome coils operate
at lower temperatures and therefore have a lower emissivity. Finally,
mercury arc lamps are used most frequently for experiments dealing
with the far-infrared region of the spectrum. The first two types will be
discussed in more detail here.

GlobarSource
This source is made out of silicon carbide (SiC) and it has metallic leads
at the ends which serve as electrodes. The application of electric
current results in the generation of heat, which yields radiation at
temperatures higher than 1000C. Water cooling is required for this
type of source because the electrodes need to be cooled [4]. This extra
level of complexity makes this source less convenient to use and more
expensive. Figure 6.1 shows the ratio of the globar source to a 900C
blackbody.

107
 Prisms:
4.2. 0 NaCI
. 4.2. Ramsey and Alishouse & KBr
Q CsI
3.4

s
1.8 Calculated after Silverman

1.0

0.2 I I i
2.l 6.0 10.0 14.0 18.0 22.0 26.0 30.0 34.0 38.0
Wavelength (m)

Fig. 6.1. Globar versus a 900C blackbody. (Reproduced from Ref. [7] with permission.
Copyright 1968, Pergamon.)

I.o

0.9

W 0.8

E
K0.7

0.6
0.
-255 K Stewart and Richmond

n_ I II I I I I I I I I I
1 3 5 7 9 11 13 15
Wavelength (im)

Fig. 6.2. Spectral emissivity of the globar source. (Reproduced from Ref. [7] with
permission. Copyright 1968, Pergamon.)

One other advantage of this source is its high emissivity down to 80

cm - ', making it useful in the far-infrared region of the spectrum. Figure
6.2 shows the spectral emissivity of a typical globar source [5]. These
values are only representative and are expected to change considerably
with use. Recently, a new low power air-cooled ceramic source has been
introduced into the modern FT-IR instruments [6]. This source has the
advantage that no watering cooling is necessary, making their instru-
ments portable and easier to maintain.

Nernst glowers
This infrared source's element is a mixture of yttrium and zirconium
oxides and has an emission spectrum that resembles that of a black

108
 z

3
W

Wavelengt (m)

Fig. 6.3. Ratio of a Nernst glower to a 900C blackbody. (Reproduced from Ref. [8] with
permission. Copyright 1978, Office of Naval Research.)

body at 1800 K. It is an insulator at room temperature and becomes a

conductor after it is preheated. It used to be popular but it has a
number of disadvantages, the biggest being its short lifetime and
mechanical instability. The spectral characteristics of a Nernst glower
versus a 900C blackbody can be found in Fig. 6.3 [7].

6.2.2 Infrared detectors

One of the most important elements of an infrared spectrometer is the

component responsible for the detection of infrared energy. Typically,
the description of a detector is not limited to the responsive element
which changes the incoming radiation into an electrical signal, but it
also includes the physical mounting of the element, like the windows,
the apertures, the Dewar flasks, etc. Together, they form what is called
a detector [8].
There are two general classes of infrared detectors. One class com-
prises the thermal detectors and the other class the photon detectors. As
the name suggests, thermal detectors operate by sensing fluctuations
in the temperature of an absorbing material as a result of exposure to
the incoming radiation. The other category, the photon or as often
called quantum detectors are sensitive to changes in the quantity of
free-charged carriers in the solid, brought by the interaction with the
external radiation.

109
Thermal detectors
Thermal detectors rely on four different processes to achieve detection
of infrared radiation:
1. The bolometric effect. This effect relies on the change in the elect-
rical resistance of the responsive element due to temperature
changes produced by the absorbed infrared radiation. This change
in resistance is detected by conventional techniques.
2. The thermovoltaic effect. In this case, the heating of the junction
between two dissimilar materials produces a measurable voltage
across the leads.
3. The thermopneumatic effect. A very common thermal detector, the
Golay detector, relies on this phenomenon [9]. In the case of thermo-
pneumatic detectors, a gas-filled chamber that contains an infrared
absorbing element is exposed to infrared light. Absorption of energy
by the element generates heat, which heats up the gas in the
chamber. The consequent increase in the pressure of the gas results
in the distortion of a thin flexible mirror on the other end of the
sealed gas chamber. This distortion is sensed by an independent
optical system. Golay detectors have been extensively used as far-
infrared detectors, even though they had problems with their mech-
anical integrity at one point. Figure 6.4 shows a cross-section of a
typical Golay cell [10].
4. The pyroelectric effect. In this process the radiation increases the
temperature of a crystalline material. The result is a change in the
electrical polarization of the crystal surface and the generation of an
electric field.

TARGET:
THIN
BSORBING
FILM
IR
TRANSMITTIN
WINDOW ELASTIC
MEMBRANE

GAS FILLED
CHAMBER

Fig. 6.4. Cross-section of the Golay cell. (Reproduced from Ref. [10] with permission.
Copyright 1976, Institute of Optics.)

110
Photon detectors
The other category of infrared detectors is the quantum or photon
detectors. In photon detectors, incident infrared photons result in the
production of free charge carriers in the responsive element. No serious
temperature change in the element takes place during this process.
The above category can be further divided in four underlying processes:
1. Photoconductive effect. The principle behind this effect is that a
change in the number of incident photons reaching a semicond-
ucting material changes the number of the free charge carrier in the
materials. Since electrical conductivity is directly proportional to
the number of these charge carriers, it can be used to deduce the
number of incident photons on the semiconductor.
2. Photovoltaic effect. In this case, a change in the number of incident
photons on a semiconductor p-n junction results in a change in the
voltage generated by the junction. Figures 6.5 and 6.6 show the
energy band models for unilluminated and illuminated p-n
junctions.
3. Photoelectromagnetic effect. In this case, the separation of the
charge takes place via the use of a magnetic field. The charge
separation produces a voltage that is directly proportional to the
number of incident infrared photons.
4. Photoemissive effect. In this case, an incident photon is absorbed by
the surface and gives up its energy to a free electron. This electron
can escape the surface and in the case that the surface is in an
evacuated chamber equipped with an anode and an eternal circuit,
electric current is detected.

Band
_.-----------Fermi Level

P-l n-Region

Unilluminated p-n junction

Fig. 6.5. Energy band model for an unilluminatedp-n junction. (Reproduced from Ref. [4]
with permission. Copyright 1978, Office of Naval Research.)

111
 Conduction Band

h -' h

Illuminated p-n junction

Fig. 6.6. Energy band model for an illuminated p-n junction. (Reproduced from Ref. [4]
with permission. Copyright 1978, Office of Naval Research.)

6.3 DETECTOR NOISE

There are several contributors to the noise (the fluctuation in signal

intensity for a steady radiation field) of an infrared detector. In infra-
red spectroscopy, the detector noise is most often much higher than any
other noise source. In addition, it has usually a thermal origin.
Therefore, the majority of infrared detectors do not operate at room
temperature.
Johnson noise: this type of noise is generated in resistors due to the
random thermal motion of the charge carriers. As the temperature of
the element increases there is a concurrent increase in the average
kinetic energy of the carriers, which results in an increased electric
noise voltage. This is the reason this type of noise is also called thermal
noise.
Shot or Schottky noise: this is random noise that has to do with the
statistical fluctuations of the photon fluxes. It has its origin in the
discreteness of electrical charge.
Both of the above types of noise are 'white' types of noises. This
means that they are independent of the frequency all the way out to the
cut-off frequency. Other types of noise exist that are dependent on the
frequency of the incoming radiation. The most important of these types
is the 1/f noise. Its mechanism is not well understood but, as the name
implies, its magnitude is reversibly proportional to the frequency of the
radiation.

112
 In addition, electronic components associated with the detector
contribute to the noise. Pre-amplification is always required for any
type of detection system.

6.4 PERFORMANCE OF AN INFRARED DETECTOR

The performance of an infrared detector is measured by a set of figures

of merit. The responsivity of the detector is the output of the electrical
signal to the incident radiation power. The noise equivalent power
(NEP) is the level of incident infrared signal that produces a signal-to-
noise ratio of one. Detectivity is defined as the reciprocal of the noise
equivalent power (NEP). The normalized detectivity D* is a widely used
figure-of-merit and it includes the area of the detector and frequency
bandwidth of the measurement.
Thermal detectors show a 'flat' detectivity response throughout the
entire spectral region. In contrast, photon detectors have higher
detectivity but over a limited spectral range. Figure 6.7 shows the

10

lA I I
1_1_~~~'
I
N~-I

8
in i.. 1~~,___
2 4 6 8 10 12 14 16 18
Wavelength (rim)

9109
i

10 3
2
X 10 10 10 10
10 Chopping Frequency (cps)

Fig. 6.7. Plots of D* versus wavelength for (a) thermocouple versus (b) an InSb detector.
(Reproduced from Ref. [4] with permission. Copyright 1978, Office of Naval Research.)

113
 l
10'

1011

1010

10
9

1081

Wavelength (m)

Fig. 6.8. D*(X) values for a number of commercially available quantum detectors.
(Reproduced from Ref. [4] with permission. Copyright 1978, Office of Naval Research.)

response for a thermocouple detector as compared with the spectral

response for an InSb photon detector. In addition, Figure 6.8 shows the
plots of the normalized detectivity D* for a collection of commercially
available detectors.

6.4.1 MCT detector

One of the most widely used infrared detectors is the mercury cadmium
telluride (MCT) detector. This is a photon detector which needs to
operate at liquid nitrogen temperatures of 77 K. Figure 6.9 shows the
spectral response of the commercially available detectors as a function
of wavelength. On the other hand, Fig. 6.10 depicts the frequency
response of this detector's D*. Essentially, the response is 'flat' from
about 103 Hz to 106 Hz. Furthermore, the alloy composition deter-

114
 I I
10
M 10

2
-
Z
M
0
- 10

,
9

4
t I
6 8
I
10 12 14
I I'
1 1(6
Wavelength (m)

Fig. 6.9. Plot of detectivity versus wavelength for MCT detectors. (Reproduced from Ref.
[4] with permission. Copyright 1978, Office of Naval Research.)

..
. _

1 10
2 10
RF

109

2 In
-8 . . ... . . .. '
........
I ..
I ~{J I
.
I~~
I{
1
{ II~
..
- 2
103 10 4 105 106 107
Frequency (Hz)

Fig. 6.10. Plot of D* versus frequency for an MCT detector. (Reproduced from Ref. [4]
with permission. Copyright 1978, Office of Naval Research.)

W
W
C
0..
10 i. IB\ C
a:
. i
t
O

C:
- Spectral Response
is a Function of x
I i l l I , i l
6 8 10 12 14 16 18
Wavelength (. m)

Fig. 6.11. MCT detector. Effect of alloy composition to the spectral responsivity
characteristics. (Reproduced from Ref. [4] with permission. Copyright 1978, Office of
Naval Research.)

115
 x

Fig. 6.12. Wavelength cut-off for an MCT detector versus alloy composition at 77 K
(Reproduced from Ref. [4] with permission. Copyright 1978, Office of Naval Research.).

mines the spectral response of the detector element. Figure 6.11 shows
the spectral responsivity for three different alloy compositions
(Hg,,CdxTe).
It is evident that manipulation of the alloy composition results in a
detector which is tailored to particular needs for wavelength sensi-
tivity. The spectral response of the material is determined by the
energy gaps between the various energy levels in the material. There-
fore, the energy gap in an MCT alloy is related to the ratio of HgTe to
CdTe. Figure 6.12 shows the wavelength cut-off for this ternary alloy
system.

6.5 OTHER COMPONENTS

6.5.1 Beamsplitters and mirrors

The typical kind of beamsplitters in the mid-infrared region on com-

mercially available instruments is of the germanium (Ge) on potassium
bromide (KBr) substrate type. Recently, semiconductor film beam-

116
t5

D
Wavelength (m)

Fig. 6.13. Reflectance of some common metallic films used as mirrors. (Reproduced from
Ref. [12] with permission. Copyright 1957, Wissenschaftliche Verlagsgesellschaft mbh.)

splitters for infrared spectrometers have been described which are

formed from self-supporting semiconductors, including carbon films.
Preferably, the beamsplitters are formed from silicon, germanium, or
diamond films [11]. In addition, Figure 6.13 shows the reflectance for
some commonly used mirror surfaces in the infrared region [12].

6.6 DISPERSIVE INSTRUMENTS

Nowadays, dispersive instruments are used only in selected applica-

tions due to the fact that interferometric instruments offer distinct
advantages for most applications. However, there are places where
dispersive instrumentation is still used when the response at one
wavelength or a short range of wavelengths is sought [13].
The basic components of a dispersive spectrometer are the same as
in a Fourier transform instrument with the exception of the inter-
ferometer. Any differences are within the elements and are the result of
the different ways that the source radiation is detected. For instance,
sensitive thermocouple detectors are commonplace in dispersive
instruments, whereas they are not appropriate for rapid-scanning

117
instruments [14]. In a dispersive instrument a monochromator is used
in the place of the interferometer. Before 1950, the monochromator was
a rock salt prism for use in the mid-IR region of the electromagnetic
spectrum and later was replaced by a diffraction grating. Older mono-
graphs provide excellent background information on the operation and
maintenance of dispersive infrared instruments and the reader is
recommended to consult them [15,16].

6.7 MICHELSON INTERFEROMETER

Most commercial interferometers are based on the original Michelson

design of 1891 [17]. Interferometers record intensity as a function of
optical path difference and the produced interferogram is related to the
frequency of the incoming radiation by a Fourier transformation. The
principle of a Michelson interferometer is illustrated in Fig. 6.14. The
device consists of two flat mirrors, one fixed and one free to move, and a
beamsplitter. The radiation from the infrared source strikes the beam-
splitter at 45. The characteristic property of the beamsplitter is that it
transmits and reflects equal parts of the radiation. One classic type of
beamsplitter, useful in the mid infrared spectral region, consists of a
thin layer of germanium (refractive index, n = 4.01) on an infrared
transparent substrate (e.g., KBr). The transmitted and reflected beams
strike the above described mirrors and are reflected back to the beam-
splitter where, again, equal parts are transmitted and reflected. As a
consequence, interference occurs at the beamsplitter where the

RADIATIO
FROM SOt MOVING MIRROR

BE

TO DETECTOR
Fig. 6.14. Block diagram of a Michelson interferometer.

118
 Un

Z l
M
UI
U
C I~
I

I
Z'
_
. .
$---------
U '
0X

o
Qc

"o00 SZS9 s=a -at i2 s sea s 5 7Z'- 75

0 TA POINTS

Fig. 6.15. Typical interferogram showing centre-burst region.

radiation from the two mirrors combine. As shown in Fig. 6.14 when
the two mirrors are equidistant from the beamsplitter constructive
interference occurs for the beam going to the detector for all wave-
lengths. In this case, the path length of the two beams in the inter-
ferometer are equal and their path difference, called the retardation
(6), is zero.
The plot of detector response as a function of retardation produces a
pattern of light intensity versus retardation, commonly referred to as
the interferogram. The interferogram of a monochromatic source is a
cosine function. Equation (6.1) describes the above relationship:

1(6) = B(v) cos(27v6) (6.1)

where v is the wavenumber in cm-l and 6 is the optical path difference,

or retardation. The Fourier transform of the above expression is a peak
at the frequency of the monochromatic radiation. I is the intensity in
the output beam as a function of retardation (6), and B is the intensity
as a function of radiation frequency (v). In contrast, the interferogram
of a polychromatic source can be considered as the sum of all cosine
waves that are produced from monochromatic sources. The polychrom-
atic interferogram has a strong maximum intensity at the zero

119
retardation point where all the cosine components are in phase, as can
be seen in Fig. 6.15. This point is also known as the centreburst point
[18]. The expression for the intensity of the interferogram of a poly-
chromatic source as a function of retardation is described by Eq. (6.2):

1(6) = [[B(v)[1+ cos(2cv6)]] /2]dv (6.2)

Thus, in Fourier transform interferometry the data are "encoded" by

the interference produced by the retardation and then "decoded" by the
Fourier transform to yield the desired intensity signal as a function of
frequency (or wavelength).

6.8 ADVANTAGES OF INTERFEROMETRY

Two kinds of multichannel advantages exist in Fourier transform

interferometry, compared with a dispersive instrument in which only a
very narrow band of frequencies is observed at a time. The first-and
biggest practical advantage of Fourier transform spectroscopy-is the
simultaneous detection of the whole spectrum at once; it is called the
Fellgett or multiplex advantage [19,20]. Even though a factor of ca. 2 in
signal strength is lost because half of the beam is reflected back to the
source, the multichannel advantage is nevertheless 10 4 or higher. That
is, theoretically an interferometer can achieve comparable signal-to-
noise to a dispersive monochromator 104 faster.
In addition, the so-called Jacquinot or 'etendue' advantage exists.
This advantage is associated with the increase in source throughput
[21]. During dispersive detection the throughput is severely limited by
the area of the entrance slit. Even though the interferometer has an
entrance aperture of its own, its throughput advantage ranges from 10
to 250 over the infrared frequency range. This was the reason that FT-
IR spectra of astronomical sources, where very weak astronomical
emission sources are present, were produced even before the Fast
Fourier Transform (FFT) was invented [22].
A third practical advantage of interferometry is the so-called
Connes or registration advantage. Connes advantage stems from the
ability of interferometry using a monochromatic source (e.g. a helium-
neon (HeNe) laser in today's spectrometers) to accurately and precisely
index the retardation, resulting in a superior determination of the

120
retardation sampling position. For example, if the above mentioned
HeNe laser ( = 632.8 nm) is used, zero crossings in the visible inter-
ferogram occur at intervals of 632.8/2 nm = 0.3164 mm. Because the
Nyquist theorem demands at least two sampling points per cycle, the
highest infrared frequency that would satisfy the Nyquist criterion is
15,804 cm -l. For mid-IR use, sampling at every other zero-crossing (1
-
kHeNe intervals) produces a maximum Nyquist frequency of 7902 cm l.
Connes advantage allows tremendous reproducibility of interferogram
sampling and data storage. This results in full realization of signal-to-
noise problems from repeated scans and it is particularly useful for the
dynamic experiments that will be discussed later in this book [23,24].

6.9 APODIZATION

The amplitude of the side lobes which appear adjacent to absorption

bands in the Fourier transform of an interferogram can be drastically
reduced if a mathematical manipulation is performed. This treatment
is called apodization, from the Greek word ano6os (without feet). This
mathematical treatment is necessary because the Fourier trans-
formation is performed over finite limits, even though the theoretical
expression for the interferogram's intensity involves infinite limits.
Therefore, when the interferogram is truncated, this sudden cut-off
results in the appearance of oscillations around the sharp spectral
features (absorption bands) in the transform.
When the interferogram is multiplied by the apodization function,
the transform is essentially free of side lobes. Two of the most popular
and effective apodization functions are the triangular and the Happ-
Genzel functions. The amplitude of the first side lobe using triangular
apodization is larger than that of the Happ-Genzel function, but the
opposite is observed for the subsequent lobes. In general, it can be
stated that Happ-Genzel apodization is quite similar to triangular
apodization and for most situations they give comparable results [25].
Both types of apodization were used at different times in the work
reported in this dissertation.
Overall, it can be stated that the biggest drawback of apodization is
the worsening of the spectral resolution, since the contributions of the
extremes of the interferogram wings are reduced. Therefore, a trade off
exists between the reduction in spectral distortion and the worsening of
resolution.

121
6.10 RESOLUTION

Resolution is defined as the minimum distinguishable spectral

interval. The maximum retardation determines the resolution of the
scan. The maximum optical resolution achievable by a particular FT-IR
spectrometer is given by (Dma) - cm l, where D.ma is the maximum
optical path difference attainable by the interferometer. Figure 6.16
shows the effect of different resolution on the appearance of single
beam spectra. The spectra shown in this figure have been offset for
clarity. Figure 6.16a shows the open-beam background spectrum of the
unpurged spectrometer recorded with a resolution of 2 cm - l (Dm~ = 0.5
cm). It is clear that the rotational lines of vapour water are well
resolved. In contrast, Fig. 6.16b shows the open-beam spectrum
acquired with 16 cm - l resolution (Dm, = 0.0625 cm). Vapour water
absorptions are not resolved due to the lower resolution.
Dynamic methods have the potential to increase spectral resolution
beyond the above limit due to the existence of the possibility of different
responses of the components of highly overlapped bands. This possi-

a) Ca

E
M
Mt
e

U
=

. b)
'A

3600 '100 2600 2100 1600 1'0O 600

Wavenumbers

Fig. 6.16. (a) Open-beam background, 2 cm-l; (b) open-beam background, 16 cm 1 .

122
bility will be further discussed when dynamic infrared experiments
will be presented.

6.11 PHASE CORRECTION

The non-ideality of the beamsplitter in a real interferometer results in

the introduction of sine components to an interferogram which, in
principle, should consist only of cosine components. Equation 6.3 shows
the modified relation for the intensity of the interferogram:
+o

1(5) = [[B(v)[l + cos(2gv6 + DBS(v))]] / 2]dv (6.3)

where FBs(V) is the wavelength dependent phase shift introduced by the

beamsplitter.
Phase correction is the mathematical procedure to remove the sine
components from the interferogram. The Fourier transform of a com-
plete double-sided interferogram provides the correct power spectrum,
without any phase correction, since the ambiguity does not affect the
magnitude. However, when a single-sided interferogram is computed,
some knowledge of the phase is required in order to compute the true
spectrum [26]. Two of the most popular phase correction routines used
in single-sided interferograms are the Mertz algorithm and the
Forman algorithm. In the Mertz routine, the largest data point in the
interferogram is assigned as the zero retardation point and the
amplitude spectrum is calculated with respect to this point. A short
double-sided interferogram is measured and its corresponding phase
array is used to phase correct the entire single-sided spectrum. The
Forman correction is essentially equivalent to the Mertz routine but it
is performed in the retardation space [27,28]. Modifications to the
Mertz phase correction have appeared in the literature and were
originally applied to the vibrational circular dichroism (VCD) spectra
[29]. The result of these modifications is that the phase spectrum does
not change sign if a quadrant boundary is crossed.
As an alternative, a "stored" phase array can be used to produce
proper phase correction for the transformed interferograms. This
phase array is calculated from a double-sided reference interferogram.
The procedure relies on the fact that the beamsplitter phase does not
change from scan to scan.

123
6.13 EFFECTS OF MIRROR MISALIGNMENT

Mirror misalignment in an interferometer produces a lowering in the

energy on the higher energy end of the spectrum (transform). This can
be seen in Fig. 6.17 which illustrates three single beam intensities. The
degree of alignment varies in these three scans, and this is evident on
the high energy portion of the spectrum. These continuous-scan results
can be compared to step-scan phase modulation results obtained by
dithering the moving mirror with piezoelectric transducers.

Continuous-scanFT-IR
Most commercially available FT-IR spectrometers use the continuous-
scan mode of operation, where the moving mirror is scanning at
constant velocity. This type of scanning works very well for routine
measurements. In the continuous-scan mode of interferometry the
laser fringe counter is used to sense the accuracy of the scanning
velocity. If a deviation is sensed, correction signals are generated that

Wavenumbers
Fig. 6.17. Effect of mirror misalignment on the appearance of single-beam transmission
spectra.

124
assure the proper operation (constant velocity). The consequence of this
mode of operation is that each infrared wavelength (), is modulated at
its own particular Fourier frequency, given by Eq. (6.4):

f(k) = 2v/ (6.4)

where v is the mirror velocity.

Continuous-scan FT-IR is the technique of choice when static
spectral properties are determined. Co-addition of successive scans
increases the signal-to-noise ratio (S/N) by a factor proportional to t,
where t is the time that the signal is averaged at each collection point.

Step-Scan FT-IR
In step-scan FT-IR data are collected while the retardation is held
constant or is oscillated about a fixed value. Therefore, in order to apply
the technique to mid-infrared and shorter wavelength measurements,
a method for controlling the retardation and implementing a special
sampling rate comparable to that achieved in modern continuous-scan
instruments is required. In recent years several different control
methods have been reported. However, all of these basically rely on the
use of the HeNe laser fringe pattern to generate the control signal and
to determine the step size [30]. The biggest advantage of the step-scan
mode is the separation of the time of the experiment from the time of
the data collection.
Two types of experiment are possible with step-scan interferometry.
One type is the time-domain or time resolved experiments where data
are collected as a function of time at each mirror position. Sorting of the
data results in interferograms that contain spectral responses at differ-
ent times. The event under study must be a repeatable process in order
for the experiment to work.
The other type of experiment capable with step-scan is the so-called
frequency domain or synchronous modulation experiments. In these
experiments, there are two ways to modulate the intensity of the
infrared radiation in order to generate step-scan interferograms. One
way is to use amplitude modulation (AM) which can be achieved by
means of a chopper. When a chopper is used for intensity modulation, a
lock-in amplifier is used to detect the signal before digitization occurs.
The technique has the drawback that the signal is riding on top of a
large DC offset, which has to be subtracted before any meaningful data
can be obtained. This can be done by either calculating the average

125
value of the interferogram and subtracting it from each sample point
before the Fourier transform takes place or by setting the lock-in
amplifier offset to zero, far from the interferogram. Even though the
latter technique eliminates the problem of reduced dynamic range, the
technique is still susceptible to DC drift.
Another way to achieve modulation of the radiation is by phase
modulation (PM). Phase modulation is achieved in some step-scan
instruments by a low amplitude oscillation of the moving mirror along
the light path, but any other way of producing phase-difference
modulation is acceptable. PM results are superior to AM results by at
least a factor of 2 in S/N, when the experiment is detector-noise limited.
This improvement stems from the fact that the PM interferogram is
essentially the first derivative of the AM interferogram, therefore
source intensity fluctuations and other variations of the beam intensity
will cancel out [31]. Another parameter associated with PM modulation
experiments is the so-called 'phase modulation characteristic' [32].
This refers to the connection between the amplitude of the phase
modulation and the wavelength region of maximum modulation
efficiency. For the mid-IR region, a PM modulation amplitude of 2 XHeNe
(zero-to-peak) is appropriate (maximum modulation at 2300 cm-l).
In contrast to the continuous-scan method, the advantages of step-
scan operation include the ability, as mentioned above, to apply
virtually any modulation frequency to the infrared radiation and to
carry out multiple modulation experiments. Since the frequency of
modulation is not a function of any retardation velocity (e.g., mirror
scan speed), they have no dependence on radiation wavelength. In
addition, the use of lock-in amplifier detection or digital signal pro-
cessors (DSP), provides a high degree of noise rejection, analogous to
the Fourier filtering effective in the continuous scan mode. Another
advantage of lock-in amplifier or DSP detection is the easy retrieval of
the signal phase. This is possible due to the fact that the beamsplitter
(instrumental) phase is identical for the in-phase and quadrature (900
out of phase) components of the signal. These components are easily
obtained as outputs of a two-phase lock-in amplifier. As a result, not
only the magnitude M, but also the phase D can be easily obtained by
following Eqs. (6.5) to (6.8):

M = (2 + Q2 )1/ 2 (6.5)

D = arctan(Q/I) (6.6)

126
I=Mcos (6.7)

Q = M sine (6.8)

The signal-to-noise (S/N) ratio is increased by staying longer at each

data collection point. The two modes of operation, step-scan and
continuous-scan, should produce identical results under conditions of
detector-limited noise. The time resolution of the step-scan technique is
limited only by the rise-time of the detector, by the electronics
(especially by the A/D converter), and by the signal strength. Therefore,
it is capable of measuring various relaxation processes that occur in the
sub-microsecond regime and are closely associated with molecular-
scale phenomena.

6.14 FOURIER TRANSFORMATION AND ITS USE IN FT-IR

INSTRUMENTATION

The breakthrough in the application of interferometry to spectroscopy

came with the discovery by Cooley and Tukey of the fast Fourier
transform (FFT) algorithm in 1964 [33]. The FFT procedure takes
advantage of several properties of the discrete FT, which is somehow
redundant in nature [34]. The FT case can be represented as an n-
vector (n points interferogram) which must be multiplied by an nxn
matrix, each row of which is a discrete representation of a complex
sinusoid. The result of the multiplication is an n-vector, which is the
transformed spectrum. A straightforward approach requires n2
operations, where operations are complex multiplications followed by
complex additions. Since the nxn matrix is highly ordered and cyclical
it can be readily factored. IfNis chosen such that it is an integral power
of two, then extra advantage may be realized by calculating the FT on a
digital computer [35].
As an example, the Fourier transform of a 2048 point vector
requires (2048)2, or 4.2 million multiplications. The FFT algorithm
reduces this amount to (2048) x log(2048), for a total of 24233 multi-
plications. Obviously, the great advantage of FFT can be appreciated as
the number of data points gets larger and larger. Today's personal
computers can calculate an array similar to the one described above in
a fraction of one second.

127
REFERENCES

1. W.F. Herschel, Phil. Transact. Roy. Soc., 90 (1800) 284.

2. C.N.R. Rao, Chemical Applications of Infrared Spectroscopy. Academic
Press, New York, 1963.
3. J.F. Ferraro, Spectroscopy, 14(2), 1999.
4. W.L. Wolfe and G.J. Zissis, The Infrared Handbook. Office of Naval
Research, Department of the Navy, Washington, DC, 1978.
5. J.C. Morris, Comments on the measurement of the emmitance of the
globar radiation source. J. Optical Soc. Am. (Washington, DC), 51 (1961)
758.
6. Nicolet Instruments, private communication.
7. W.Y. Ramsey and J.C, Alishouse, A comparison of infrared sources. Infra-
red Physics (Pergamon, Elmsford, NY), 8 (1968) 143.
8. W.L. Wolfe and G.J. Zissis, The Infrared Handbook. Office of Naval
Research, Department of the Navy, Washington, DC, 1978, Chapter 11.
9. M.J.E. Golay, Rev. Scient. Instrum., 18 (1949) 347.
10. M. Hercher, Detectors and Lasers, in: Contemporary OpticalEngineering.
The Institute of Optics, Rochester, NY. 1976
11. A. Simon, J. Gast (Bruker Analytische Messtechnik GmbH, Germany.
Patent written in German. Application: DE 92-4242440 921216
12. G. Hass and A.F. Turner, in: M. Auwarter (Ed.), Coatings for Infrared
Optics. Wissenschaftliche Verlagsgesellschaft mbh, Stuttgart, 1957.
13. I. Noda, A.E. Dowrey and C. Marcott, Appl. Spectrosc., 42 (1988) 203.
14. R. White, Chromatographyl/FourierTransformInfraredSpectroscopy and
its Applications. Marcel Dekker, New York, 1990.
15. C.N.R. Rao, Chemical Applications of Infrared Spectroscopy. Academic
Press, New York, 1963.
16. A.L. Smith, Applied Infrared Spectroscopy, Fundamentals, Techniques,
andAnalytical Problem-Solving. Wiley-Interscience, New York, 1979.
17. A.A. Michelson, Phil. Mag., Ser. 5, 31 (1891) 256.
18. P.R. Griffiths and J.A. de Haseth, Fourier Transform Infrared Spectro-
scopy. Wiley, New York, NY, pp. 407-425, 1986.
19. P. Fellgett, Thesis, Cambridge University, Cambridge, UK, 1951.
20. P. Fellgett, J. Phys. (Radium), 19 (1958) 187.
21. P. Jacquinot, J. Opt. Soc. Am., 44 (1954) 761.
22. J. Connes and P. Connes, J. Opt. Soc. Am., 56 (1966) 896.
23. J. Connes, Optical Instruments and Techniques. Oriel Press, Paris, 1970.
24. R.A. Palmer, Spectroscopy, 8(2) (1993) 26.
25. A.G. Marshall and F.R. Verdun, FourierTransforms in NMR, Optical, and
Mass Spectrometry. Elsevier, Amsterdam, 1990.
26. L. Mertz, InfraredPhys., 7, (1967) 17.
27. M.L. Forman, J. Opt. Soc. Am., 56 (1966) 978.

128
28. M.L. Forman, W.H. Steel and G.A. Vanasse, J. Opt. Soc. Am., 56 (1966) 59.
29. C.A. McCoy and J.A. de Haseth, Appl. Spectrosc., 42 (1988) 336.
30. R.A. Palmer, Spectroscopy, 8(2) (1993) 26.
31. J. Chamberlain, InfraredPhys., 11 (1971) 25.
32. J. Chamberlain and H.A. Gebbie, Infrared Phys., 11 (1971) 57.
33. J.W. Cooley and J.W. Tukey, Math. Comput., 19 (1965) 297.
34. C.J. Manning, Ph.D. Thesis, Duke University, 1991.
35. M.L. Forman, J. Opt. Soc. Am., 56 (1966) 978.

129
Chapter 7

Sampling techniques and applications

PART A. DIRECT TECHNIQUES

In recent years there have been remarkable theoretical and

experimental advances in a variety of measuring methods for infrared
spectroscopy, which have made it possible for anyone to try various
infrared measurement methods relatively easily [1-6]. While trans-
mission methods are most often used for measurements of infrared
spectra of general samples, the attenuated total reflection (ATR)
method, diffuse reflectance (DR) method, reflection-absorption (RA)
method, photoacoustic spectroscopy (PAS), infrared microspectroscopy,
and emission spectroscopy, etc., are also often employed.

7.1 TRANSMISSION SPECTROSCOPY

Transmission methods are suitable for infrared measurements of

liquid (solutions) samples, powder samples and gases. Practical
measures for the intensities of infrared bands in a transmission
spectrum are given by Lambert-Beer's law. Before we explain the law,
let us define transmittance, T, and absorptivity, A, When a parallel
light beam enters transparent material with the length of d cm (as
shown in Fig. 7.1), transmittance, T, is defined as follows:

T=ItIIo (7.1)

where It and Io are the intensities of the parallel beam at the positions
of incidence and exit, respectively. Transmittance, T, is the proportion
of the intensity of transmitted light to that of incident light. On the

131
 x x+dx
! - /::

Io It >

Fig. 7.1. Absorption of light by a transparent material.

other hand, absorptivity, A is the proportion of the intensity of

absorbed light (a) to that of incident light; namely

A = I/Io (7.2)

If one assumes that the transparent material does not scatter light and
is non-fluorescent, the following equation holds:

It+a =Io (7.3)

Therefore

A+T=I (7.4)

Now we will develop Lambert-Beer's law which explains the degree of

decrease in the intensity of incident light. If the intensity of incident
light decreases from i to i + di when the light proceeds from x to x + dx in
the transparent material, the degree of decrease in the intensity (di) is
considered to be proportional to the intensity of the incident light (i),
the concentration of a sample, c (mol dm-3 ), and the thickness of the
sample (dx):

-di = c*ic dx (7.5)

where e* is a proportional constant. If we make definite integral of Eq.

(7.5) from 0 to d

132
- i =d c|cf dx (7.6)

Thus

I t = Io exp(-cdc*) (7.7)

Equation (7.7) is a mathematical representation of Lambert-Beer's

law, which means that the intensity of incident light decreases expo-
nentially. Lambert-Beer's law is usually used in the form of a common
logarithm. Equation (7.7) can be expressed as follows in terms of a
common logarithm:

It = Io.lO0 d (7.8)

where

8 = 0.434* (7.9)

If we take the common logarithm of Eq. (7.8)

log(IJIt) = log(l/T) = cds (7.10)

log(IJI t) is defined as absorbance, which is proportional to the

concentration, c, of the sample and the path length, d, of a cell. The
proportional constant is a measure of the strength of absorption at a
particular frequency and is called molar absorption coefficient. The
unit of 8 is (mol dm- cm) - l, namely, M-l cm-l (in SI units, it is m2 mol-).
c is specific for each sample (the values of usually range from 10 to
105). One can calculate the concentration from the measurement of the
absorbance of a sample, if one knows e and d (usually, d is taken as 1
cm). In contrast, one can determine from known d and c.
When we use a transmission method on a liquid sample, the selec-
tion of window materials (which transmit infrared light) and the length
of a cell are important. During selection of window materials, we must
note a usable wavenumber range, a refractive index, and solubility in
water. Table 7.1 summarizes the usable wavenumber range, the
refractive index, and the solubility in water of representative window
materials. Among the window materials for non-aqueous solutions,
KBr is most often used. KBr has a wide usable wavenumber range

133
TABLE 7.1
The usable wavenumber range, the refractive index, and the solubility in water of
representative window materials for infrared transmission spectroscopy

Window Usable wavenumber Refractive Properties

material range/cm - 1 index

CaF 2 75000-1000 1,40 insoluble in water

BaF 2 67000-800 1,45 insoluble in water
KBr 43000-400 1,52 soluble in water and alcohol
CsBr 42000-250 1,65 soluble in water
CsI 42000-200 1,72 soluble in water
NaCl 40000-600 1,50 soluble in water
KC1 33000-400 1,47 soluble in water
ZnSe 20000-500 2,42 insoluble in water
KRS-5 12000-350 2,35 insoluble in water, poisonous
Si 10000-100 3,42 insoluble in water
Ge 5000-400 4,00 insoluble in water

(4000-340 cm-l) and is inexpensive; furthermore, the refractive index

of KBr (n = 1.52) is close to that of a solution. Although we can use only
above 1000 cm -1, insoluble CaF 2 (n = 1.39) is proper for an aqueous
solution (n = 1.33). KRS-5 (n = 2.37) is often used for infrared measure-
ments in lower wavenumber regions. However, KRS-5 is poisonous and
requires an attention.

7.1.1 Liquid (solution) samples

Infrared spectra of liquid samples are measured using a fixed cell for
liquid or an assembled cell. Fixed cells and assembled cells are
basically the same in that a spacer having a certain thickness is
disposed between two window materials. While a fixed cell is
advantageous in that the cell has a clearly defined thickness and is
airtight, it is not easy to clean this type of cell. Conversely, although an
assembled cell is easy to clean, this type of cell is not airtight in general.
When our sample is a solution sample with high viscosity and a low
vapour pressure, it is also possible to measure a spectrum only with the
solution sample held between two window materials.

134
 Since water exhibits strong infrared bands, a path length of a cell
must be 10 upm or shorter to measure an infrared spectrum of an
aqueous solution by a transmission method. If we use heavy water
(D 2 0) instead of light water (H 2 0), strong water bands (-3500 cm -l ,
-1650 cm- ) show a downward shift. Therefore, it is effective to use
deuterium water; one measures spectral regions in the vicinity of 3500
cm -1 and 1650 cm-l. The ATR method is often also used to measure
infrared spectra of aqueous solutions. A variety of liquid transmission
cells are commercially available. Figures 7.2a and b show expanded
views of the micro demountable flow-thru cell and heated demountable
cell kit, respectively.

(a)

(hi

Fig. 7.2. Expanded views of the micro demountable flow-thru cell (a) and heated
demountable cell kit (b). (From catalogue of Nicolet.)

135
7.1.2 Powder samples

To measure an infrared spectrum of a powder sample by a transmission

method, the KBr disc method is usually used. Since a particle diameter
is too large when we use a powder sample as it is, incident light is
irregularly reflected and, as a result, we cannot obtain a good spec-
trum. Therefore, it is necessary to crush the sample into pieces each
having a diameter of 1-2 pm in advance.

7.1.3 KBr method

When using the KBr method, crush and mix 1-2 mg of a sample and
approximately 100 mg of KBr powder in an agate mortar, introduce the
mixture into a tabletting equipment, and tablet. With this method, the
following three points should be noted:
1. Since it is impossible to obtain good tablets if KBr or a sample
contains moisture (i.e., resultant tablets become opaque), remove
the moisture as much as possible.
2. If the sample is not sufficiently crushed, the tablets produced will
become opaque or the quantity of transmitted light will decrease
(due to light scattering). Hence, the crushing must be sufficient.
3. K+ or Br and cations or anions contained in the sample sometimes
exchange with each other. Further, crushing, pressurization, etc.
sometimes causes denaturation of the sample.
To confirm that a situation like (3) has not occurred, it is desirable to
measure infrared spectra of the same sample by other methods (e.g.,
the nujor method) in parallel with the KBr method. When the amount
of the sample is small, use micro-tabletting which allows one to make a
tablet having a diameter of 1 mm.

7.2 INTERNAL AND EXTERNAL REFLECTION SPECTROSCOPY

7.2.1 Attenuated total reflection (ATR) method

As its name suggests, the ATR method is a type of reflectance method

and has been used mainly for surface analysis and analysis of bulk
materials and aqueous solutions [1-6]. In the ATR method, a sample is
placed on a prism which has a larger refractive index than that of the
sample, as shown in Fig. 7.3, and infrared light is introduced to the

136
 anterior capsule
epithelium

nucleus
ncex - posterior capsule
cortex

ATR prism (ZnSe)

Fig. 7.3. An ATR prism and an eye lens on it. (Reproduced from Ref. [7] with permission.
Copyright (1998) Society for Applied Spectroscopy.)

prism such that the infrared light is reflected totally at an interface

between the prism and the sample [7]. At this stage, to develop total
reflection, the incident angle 0 must be larger than the critical angle Oc.
Where the refractive indices of the prism and the sample are nI and n2,
respectively, the critical angle is defined as:

Oc = sin-l(n2 /nl) (7.11)

Total reflection discussed here is not reflection whereby the incident

light is totally reflected at the interface, but reflection whereby the
incident light penetrates inside the sample once to a certain extent
before being reflected. That is, although the incident light is reflected
100% in a wavelength range in which the sample does not absorb light,
in a wavelength range in which the sample absorbs it the reflectance is
decreased depending on the strength of the absorption. Hence, if we
measure the intensity of reflected light in a certain wavelength range,
we can obtain a spectrum that resembles a transmission spectrum.
In the ATR method, the penetrationdepth c is important. The depth
dp is expressed as follows:

dK (7.12)
2x in 2 0 _ )2)

where kl denotes the wavelength of the light within an ATR crystal. As

is clear from the Eq. (7.12), the depth dp is determined by the incident
angle, the wavelength, and further, by a ratio of the refractive index of

137
a sample to the refractive index of the ATR crystal. Since the depth dp is
at most a few gm in the infrared region, an ATR spectrum provides
information regarding a surface and around the same. In the ATR
method, the contact between a sample and an ATR prism must be very
smooth. Figure 7.3 shows an example of non-destructive analysis of an
eye lens by the ATR method [7].
The ATR method is very suitable to obtain infrared spectra of
aqueous solutions. Figures 7.4a and b show infrared spectra of water
measured with the transmittance method and ATR method, respect-
ively [8]. A strong feature which appears in the range of 3600 to 3200
cm-l is due to the OH stretching vibrations, while a band in the vicinity
of 1640 cm-l is assigned to the HOH bending vibration. A major
problem in studying aqueous solutions comes from these two bands.
Although the spectrum (Fig. 7.4a) is obtained using a very thin liquid
cell (whose thickness is 10 lpm), we can find a strong band with
absorbance of 1.5 or higher. The ATR method considerably weakens the
band due to the OH stretching vibrations. This is because the shorter
the wavelength, the smaller the penetration depth dp (Eq. (7.12)). In
this manner, with the ATR method, we can suppress the intensity of
the strongest band of water, and therefore, data processing for sub-
tracting the infrared spectrum of water is relatively easy.
Figure 7.5a shows an ATR infrared spectrum of oxidized cyto-
chrome c (pH 9.3, 10wt%) in a phosphate buffer. In Fig. 7.5b, a

0
cd

0
E:

4000 3600 3200 2800 2400 2000 1600 1200 800

Wavenumber/cm -'

Fig. 7.4. Infrared spectra of water measured with the transmittance method (a) and ATR
method (b). (Reproduced from Ref. [8] with permission. Copyright (1994) Tokyo Kagaku
Dojin.)

138
 Z
C3

C
EX

Wavenumber/cm -'

Fig. 7.5. (a) An ATR/infrared spectrum of oxidized cytochrome c (pH 9.3, lOwt%) in a
phosphate buffer. (b) An ATR/infrared spectrum of the phosphate buffer solution.
(c) Difference spectrum obttained by subtracting spectrum (b) from spectra (a).

spectrum of the phosphate buffer solution is presented. Note that the

spectrum of the protein solution is close to that. Hence, in order to
extract useful information from the spectrum shown in Fig. 7.5a, we
have to subtract the spectrum of buffer (Fig. 7.5b). Figure 7.5c shows a
resulting spectrum that is obtained by subtracting the spectrum of
buffer. A simple guideline for determining if the water spectrum is
successfully subtracted is to see whether the range of 2400 to 1700 cm - '
is flat.

7.2.2 Prisms and accessories for ATR spectroscopy

ATR spectroscopy has long been used for various samples from aqueous
solutions to bulk materials [1-6]. A variety of prisms and accessories
for ATR spectroscopy have been developed and are commercially avail-
able. They are divided into two groups: single reflection type with a
hemicylinder crystal and multiple-reflection type with a trapezoidal
crystal. Figure 7.6 depicts optical schematics of several commercially
available ATR accessories. In FT-IR spectroscopy, the multiple-reflec-

139
tion setups are more popular than single reflection setups because in
the former one can control the ATR signal intensity easily by adhering
samples on both sides of an ATR prism or by changing the size of
samples. The energy loss by the multiple-reflection can be compensated
by the increase in the number of acquisition or the use of a MCT

(b)

(c)

Fig. 7.6. Various accessories for ATR spectroscopy: (a) single-reflection variable-angle
hemicylinder; (b) multiple-reflection single-pass crystal; (c) circle ATR cell for liquid
samples.

TABLE 7.2
The usable wavenumber range, the refractive index, and the properties of representative
window materials for ATR spectroscopy
Window Usable wavenumber Refractive Properties
I
material range/cm index

ZnSe 20000-400 2,4 suitable for aqueous solution

KRS-5 20000-300 2,40 poisonous
AS 2 Se3 12500-800 2,80 suitable for aqueous solution,
fairly fragile
Si 5000-1600 3,40 fragile
Ge 5000-900 4,00 high refractive index, suitable
for aqueous solution, fragile
--

140
 (a) (b)

i
I
i
II

Fig. 7.7. (a) Accessory for micro ATR; (b) in situ ATR accessory.

detector. The ATR cell shown in Fig. 7.6b is very suitable for films,
solids, and liquids, while the cell shown in Fig. 7.6c is designed for
liquid samples. Table 7.2 summarizes properties of selected optical
materials used for ATR spectroscopy. Recently, micro ATR cells and in
situ ATR accessories have made marked progress; Figs. 7.7a and b
show their examples, respectively.

7.2.3 Examples of ATR infrared studies

Two examples of ATR infrared studies are introduced here. Another

example will be described in Section 9.7.3. Figure 7.8a-c show ATR
spectra of water and the anterior and posterior portions of the lens
capsule of a 3-month-old albino rabbit lens on an ATR prism [7]. The
spectra of the capsules are very close to that of water, but one can see
weak features in the 1600-1000 cm-l region that are not assigned to
water. Figures 7.9a and b present difference spectra that were obtained
by subtracting the spectrum of water from the spectra of the lens
capsule [7]. The thickness of the capsules of the rabbit lens is about
2-20 pm, while the penetration depth of the ATR prism in the
1700-1000 cm-l region is less than 1 pm. Therefore, all the ATR signals

141
 0031

w
0
z

0
o
(,

1
WAVENUMBER /cm-

Fig. 7.8. ATR/infrared spectra of water (a) and the anterior (b) and posterior (c) portions
of the lens capsule of a 3-month-old albino rabbit. (Reproduced from Ref. [7] with
permission. Copyright (1989) Society for Applied Spectroscopy.)

in Fig. 7.9a and b may be due to type IV collagen, a major component of

the capsule that envelops the lens. Figure 7.9c shows an ATR spectrum
of purified type IV collagen in an aqueous solution. It is noted that the
ATR spectra of the rabbit lens (Figs. 7.9a and b) are very close to that of
type IV collagen in the aqueous solution (Fig. 7.9c). Bands at 1638 and
1553 cm-l are due to amide I and amide II of type IV collagen, respect-
ively. Medium features at 1080 and 1033 cm -l are assigned to C-O
stretching modes of collagen [7]. The frequencies of amide I and II
suggest that type IV collagen in the capsules assumes a secondary
structure that is very close to 3 helix [7]. This ATR study is a good
example demonstrating the potential of ATR spectroscopy in investi-
gating the structure of biological molecules in situ.
ATR/IR spectroscopy has considerable promise in on-line monitor-
ing [9,10]. In general, an ATR immersion probe is used as a sending
head, which allows one to monitor reaction occurring within about 1 ipm
of the surface of an ATR prism. The ATR method has two advantages in
chemical process analysis. One is that the immersion probes have little
effect from bubbles and suspensions in reaction mixture. Another is
that the ATR method is applicable to samples that do not transmit
enough light. Figure 7.10a shows the results of on-line monitoring of

142
 WAVENMMBR /cm-1

Fig. 7.9. (a) The difference spectrum between spectrum (a) and spectrum (b) in Fig. 7.8.
(b) The difference spectrum between spectrum (a) and spectrum (c) in Fig. 7.8. (c) ATR/
infrared spectrum of Type IV collagen from human placenta in aqueous solution (Repro-
duced from Ref. [7] with permission. Copyright (1989) Society for Applied Spectroscopy.)

the esterification of acid anhydride and diol measured by an ATR in

situ setup [10]. The esterification is one of the most basic reactions in
chemical industries. In the esterification by measuring the acid value
and hydroxyl group value at any time the terminal point of the reaction
must be predicted. It can be seen from Fig. 7.10a that as the reaction
proceeds, a broad feature near 3400 cm l due to the OH stretching
mode of alcohol decreases and a sharp band at 1700 cm-l assigned to
the C=O stretching mode of ester increases. Figure 7.10b illustrates
correlation for the acid value between the laboratory data and the ATR/
IR data [10].

7.2.4 Reflection-absorption spectroscopy

Reflection-absorption spectroscopy (RAS) is a powerful technique for

measuring IR spectra of thin films on metal [1,2,8,11,12]. Figure 7.11
illustrates the principle of RAS. Let us consider the behaviour of
polarized light impinging on a metal surface. When light impinges on

143
 (a)

0.5 initiation of reaction

-0.010 d
_
4000 3600 3200
!
N.2800 --
r
2400
41 1

200oo 160
fA"
1200 725
Q

,iXjl
C
0.5 _

in the middle of reaction

-0.010
46)00 3600 3200 2800 2400 2000 16 1200 725

0.5
termination of reaction

--(0 fi 0003600 200 2800 200 2000 16t0 _,

---- O WO 320 280 240 200 160 12-0 72

Wavenumber cm '

(b)

120 Regression Statistics

o00 RSO.=0.99
80 Std. Err.=3.4

40 6167.0
- 20
~~40l ~~~~3729 36.1
26.6
cJ P~~~~~26 25.9
0 20 40 60 80 100 120
acid value iAoraor4data)

Fig. 7.10. (a) The results of on-line monitoring of the esterification of acid anhydride and
diol measured by an ATR in situ setup. (b) Correlation for the acid value between the
laboratory data and infrared data. (Reproduced from Ref. [10] with permission.
Copyright (1995) Kogyo Gijutsu Co.)

the metal surface, an electric field is generated near the surface. The
intensity of the electric field depends upon the incident angle and the
direction of polarized light. When perpendicular-polarized light is used,
the electric vector of the incident light and that of the reflection light
cancel each other out because the phase of the incident wave is shifted
by 180 due to the effect of free electrons in metal. As a result, there is

144
Fig. 7.11. Behaviour of two kinds of polarized light which is incident onto a metal surface:
(a) perpendicular polarization; (b) parallel polarization.

almost no standing wave. In contrast, when parallel-polarized light is

applied, the electric vector of the incident light and that of the
reflection light strengthen each other, generating an electric field
perpendicular to the metal surface. The strength of this standing wave
increases as the incident angle approaches 90 .
Figure 7.12 shows the dependence of the absorption factors for the
parallel (Ap) and perpendicular (As) polarization at the wavenumber of
maximum adsorbed layer [12]. An optimal incident angle that gives the
highest sensitivity changes with metals and the wavenumber of incid-
ent light. Usually the optimal incident angle is 85-89 . The higher the
reflectivity of the metal, the larger the optimal incident angle.
When parallel-polarized light impinges on a thin film absorbed on a
metal surface, the standing wave interacts with the molecules in the
film and the light is absorbed. The change in reflectance in the spectral
region where the light is absorbed can be represented by the following
equation [8]:

)
-1 - 1 cs(7.13)
n Cos0

Here, AR and Ro are a change in reflectance induced by the presence of

the thin film and reflectance in the absence of the film, respectively. n1
and n2 are refractive indices of medium (such as atmosphere) and that

145
 x

0 20 40 60 80
Angle of incidence, degrees

Fig. 7.12. Dependence of the absorption factors for the parallel (Ap) and perpendicular
(As ) polarization at the wavenumber of maximum adsorbed layer. (Reproduced from Ref.
[12] with permission. Copyright (1993) John Wiley &Sons.)

thin film, respectively. 0, a, and d represent an incident angle, an

absorption coefficient, and a thickness of the thin film, respectively. In
the case of transmission spectroscopy, the ratio of a change in the
intensity of transmitted light in the presence of the sample (A) and the
intensity of the incident light (Io) can be represented by the following
equation:

rlIo = -ad (7.14)

Comparison of Eqs. (7.13) and (7.14) reveals that the sensitivity of RAS
is higher by (4n1 3 sin20)/(n2 3 cosO) than that of transmission
spectroscopy [8]. The term 4nl3 sin 2 0 represents the dependence of the
intensity of standing wave appearing on the metal surface upon the
incident angle, while the term 1/cosO is concerned with a sample area
irradiated by the incident light.
RAS has been widely used for studies of thin films such as
Langmuir-Blodgett films, thin polymer films, coating films and
epitaxial layers on silicon wafers [12]. RAS is very useful not only for
investigating chemical composition and molecular structures but also

146
for molecular orientations. Good examples of applications of RAS are
given in Section 8.6.

7.3 DIFFUSE REFLECTANCE SPECTROSCOPY

A diffuse reflectance method is a measurement method for measuring

reflected light (diffuse-reflected light) which exits a sample surface
after impinging upon the sample surface and thereafter getting
reflected, transmitted while refracted, and scattered repeatedly (Fig.
7.13) [1,2]. When we irradiate a powder sample with light, while a
portion of the light is regularly reflected at a powder surface, the
remaining majority enters the powder and diffuses. During the process
of diffusion, since light having particular wavelengths is absorbed by
the sample, if we measure the intensity of the diffuse-reflected light at
various wavelengths, we obtain a spectrum that is similar to a trans-
mission spectrum.
The intensity of a diffuse-reflectance spectrum is generally
expressed by a Kubelka-Munk equation such as:

K (1-R )2
f(R ) (7.15)
S 2R

where K, S, and R, are an absorption coefficient, a scattering coefficient

and an absolute diffuse reflectance, respectively. Further, f(R.) is
called an K-M (Kubelka-Munk) function. Since K is in proportion to a

I I

I:D;incident light
diffuse reflection light
S: regular refrection light

Fig. 7.13. Schematic representation of regular reflection and diffuse reflectance for a
powder sample.

147
 YA
'1 F

incident light diffuse reflectance

light

o
d
t
1I t J+dJ
dy t
I+dI t/

Fig. 7.14. Incident light and diffuse-reflected light in a powder layer (Kubelka-Munk
theory).

molar absorption coefficient, , and a concentration of a sample c (K =

yc where y is a proportional constant), if the scattering coefficient is a
constant value, the K-M function should be in proportion to the sample
concentration. To measure the absolute diffuse reflectance is not
realistic; we therefore measure a relative diffuse reflectance R instead
(which is a ratio of the intensity of reflection from the sample to the
intensity of reflection from a standard substance).
Let us yield a Kubelka-Munk equation. Assuming that monochro-
matic light is incident upon a powder layer which has a thickness d, as
that shown in Fig. 7.14, from the top along they-axis, the incident light
propagates in the sample in the negative direction along the y-axis and
diffuse-reflected light propagates in the positive direction along the
same axis. A decrease, dI, in the intensity I of the light which propa-
gates within a thin layer portion dy in the negative direction and a
decrease, dJ, in the intensity J of the light which propagates in the
positive direction are expressed as follows:

-dI = (K+ S) Idy - SJdy (7.16a)

dJ = - (K+ S) Jdy + SIdy (7.16b)

The first term on the right-hand side of Eq. (7.16a) denotes an intensity
decrease due to absorption and scattering of the incident light, while
the second term on the same side denotes a contribution from back

148
scattering light. Equation (7.16b) can be interpreted in a similar
manner. Now, if(S + K)/S = 1 + K/S = a, Eqs. (7.16a) and (7.16b) can be
re-written as follows:

-dl/Sdy = aI- J (7.17a)

dJSdy = -aJ+I (7.17b)

Diving Eq. (7.17a) by I and Eq. (7.17b) by J and adding the two
equations, we obtain

dr/Sdy = r2 - 2ar+ 1 (7.18)

Integrating this equation, we obtain

(r2 -2ar +1)-ldr =Sldy (7.19)

In this equation, the values of r when y = 0 and y = d hold are Rg (a

background reflectance) and R (the reflectance of the sample),
respectively. If an integral of Eq. (7.19) is expressed by the relationship
as below

dr
dr ~dr 1 (7.20)
2
(r - 2ar + 1) r + (2ar - 1)2 }{r -(2ar - 1) 2})

the integral is solved simply as:

{R, -a -(a 2 1)V2 {Rg -a +(a 2 _ 1)21 =2sd(a2 1)2 (7.21)

In12 2sd(a _ 1)"" (7.21)
{Rg -a -(a 2 -1)2 ){Rs -a +(a 2 -1)1/ 2

Since we can ignore Rg for an extremely thick sample (i.e., Rg = 0 when d

= ~), Eq. (7.21) is simplified as follows:

{-a -(a 2 _1)'2){R -a +(a2 _1)l/2 =0 (7.22)

If we solve Eq. (7.22) with respect to R,

149
R = 1 1 (7.23)
a+(a2 -1) / 2 1 +K + (KlS)+2 2K s}.3

We can obtain the Kubelka-Munk equation (7.15), if we solve this

equation above with respect to KIS.
To carry out qualitative and quantitative analysis using the diffuse
reflectance method, we need to use an equation which links the
intensity of diffuse-reflected light and an absorption characteristic of a
sample. An equation that is often used is the following equation which
is yielded from the Kubelka-Munk equation.

K = cosh(log(R)) -1 (7.24)

Equation (7.24) is illustrated in Fig. 7.15. Although K/S (which is

related to the absorption characteristic of the sample) and log(l/R)
(which is related to the intensity of diffuse-reflected light) are not in a
linear relationship with each other, in a narrow KIS range we can
regard that the two are approximately in a linear relationship with
each other. When we analyze an infrared diffuse-reflectance spectrum,
log(1/R) is usually indicated along the vertical axis.
Problems with the diffuse-reflectance method are an influence by
regular reflection and a scattering coefficient. The former is influenced
by the size, the shape and an absorption coefficient of powder. In
general, the smaller a powder diameter is, the weaker the regular

0
s

log (l/R)
Fig. 7.15. Relationship between log (1/R) and K/S derived from Kubelka-Munk equation.

150
reflection light becomes. The latter is influenced by the diameter, the
shape, a filling density, etc., of power. Hence, infrared diffuse-reflect-
ance spectra are usually measured for samples mixed with KCl or KBr
powder to reduce the influence of regular reflection. For a sample that
interacts with KCl or KBr, diamond powder and silicon powder are also
employed as a diluent.
The Kubelka-Munk theory is based on a continuum model. It does
not consider the particle nature of the powdered samples. The scatter-
ing coefficients is affected by packing density and particle size and
hence affects the resulting infrared spectrum. Studies have been
undertaken to investigate the effect of particle size, size distribution,
packing density and the orientation of the sample cup [13-15].
Generally, the spectra of the same sample vary 15-30% (in absolute K-
M units) when run by several people [16]. Griffiths and co-workers
demonstrated the effect of pressure on the diffuse reflectance infrared
spectrum of powdered samples. They concluded that the duration of
pressure applied is also an important factor for spectral reproduc-
ibility. They obtained a spectral reproducibility of 3% relative standard
deviation after subjecting the sample to high pressure (order of tons per
square inch) over a period of 15 min.
The problem arising from the factors adherent to the physical
nature of the sample was approached by Christy et al. [15,17] in two
different ways.
1. For a sample with a narrow range of particle size distribution, the
packing was carried out by an automatic packing machine to
eliminate variations arising from packing pressure and time.
2. The sample cup was rotated slowly during the spectral measure-
ments to eliminate the inhomogeneity of the sample arising from
the different shapes and sizes of the sample particles.
However, these techniques are not widely used because of practical
problems. Attempts have also been made to modify the Kubelka-Munk
function to include particle size. The Kubelka-Munk function was
shown to possess an excellent inverse relationship with particle size
(see Fig. 7.16) of the samples [18]:

f(R) oc lid (7.25a)

where d is particle diameter.

151
 0.37

0.30

0.23

; 0.16

.0
0.09

0.02

3280 3120 2952 2784

Wavenumber cm'

Fig. 7.16. Diffuse reflectance spectra of 4%(w/w) samples of mono-disperse polystyrene

-1
spheres in KBr in the region 3300-2700 cm .

By introducing particle size in the Kubelka-Munk function (Eq.

(7.25b)), quantitative determinations could be made using the diffuse
reflectance technique [18].

f(R) = (KISd (7.25b)

However, the above equation must be taken strictly as an empirical

relationship because there is no theoretical justification for the
inclusion of the particle size in the Kubelka-Munk function.
The diffuse reflectance spectrometry is applied to a wide variety of
solid samples. The samples that can be ground can be easily mixed with
ground KBr powder for spectral measurement. Furthermore, samples
that are dark like coal and kerogen are comfortably measured using the
diffuse reflectance technique (see Fig. 7.17). Samples that are difficult
to grind, for example asphaltenes can be measured using a technique
used by Christy et al. [19].
In this technique the asphaltene sample is dissolved in a low boiling
liquid such as dichloromethane. A background spectrum is measured

152
 a
E

I0
2e

4U
a0
WU

Wavenumber cm-I

Fig. 7.17. Diffuse reflectance spectra of three different coke samples with their volatile
matter content (VMC) values.

with firmly packed KBr powder. Then the asphaltene solution can be
carefully dropped onto the surface of the KBr powder. The sample
penetrates the particles and settles on the surface of the particles.
Since the diffuse reflectance technique is a near surface technique, a
small amount of the sample from a few drops of the solution is more

153
 TRANSMITTANCE
. . SPECTRUMi I I i
TRANSMITTANCE SPECTRUM

U)

z
I Ii I I 200 I i I 1 In
Z 40Do 3000 2000 1600 1000 6 To

v
DIFFUSE REFLECTANCE SPECTRUM
id

4000
A, _~~~~~~~~~~~~~~
3000 2000 1600 1000 600 1/cm
WAVE NUMBER

Fig. 7.18. Diffuse reflectance spectrum of asphaltene (obtained by deposition technique)

and absorption spectrum of the same amount of sample in KBr using the transmission
technique.

than enough to acquire a good quality diffuse reflectance spectrum

(Fig. 7.18).
The sampling technique in diffuse reflectance spectrometry has also
been modified to suit high temperature and high-pressure studies.
Thermal dehydration and decomposition reactions have been carried

154
out [2--23]. An example using high temperature in diffuse reflectance
infrared spectrometry is given in Chapter 9.

7.4 PHOTOACOUSTIC SPECTROSCOPY

When a sample is irradiated by monochromaticlight it is absorbed and

converted into heat. This heat is propagated into the gas surrounding
the solid and causes a variation in pressure. If the gas is contained to
surround the sample, then the pressure variation can be detected as an
acoustic signal. The photoacoustic spectroscopy is based on this signal.
In photoacoustic spectroscopy, the sample is placed in a cell con-
taining a gas. A microphone is attached to the cell for the detection of
the photoacoustic signal. A sketch is shown in Fig. 7.19.
Rosencwaig and Gersho [23] have discussed the theory of the photo-
acoustic effect with solids. The theory was developed considering the
diffusion of heat generated by the absorption of electromagnetic
radiation by a solid sample.
When infrared radiation strikes the surface of a sample that has an
absorption coefficient cm-l, the radiation intensity decays
exponentially as

I=I e (7.26)

where I is the incident energy and I is the energy at depth x. The

optical absorption depth of the sample is p = 1/P cm. If the radiation is

Zig

CGas

Microphone
/ / I' / //
zzr

F=-o+lb)
Fig. 7.19. Sketch of the setup used in photoacoustic measurements.

155
modulated sinusoidally at o rad s-l, then the intensity of radiation at a
depth x is

I = Io (1 + cosot) e- (7.27)

Absorption of infrared radiation at a depth x produces heat energy PI

per unit volume (heat density).

PI = X PIo (1 + cosot) e- > (7.28)

The heat generated then decays exponentially as

PI e (7.29)

where a is the thermal diffusion coefficient. The heat decay can then be
written as
X
4 1Io (1 + cost) e e - = X pIo (1 + cosot) e- (P+"' (7.30)

The diffusion depth of the heat generated is p, = 1/a cm. The diffusion
depth can be related to the physical parameters Ks, p,, Cs as

Ps = (2KJpsCsc o) (7.31)

where Ks is the thermal conductivity, p, is the density, and Cs is the

specific heat of the sample.
The oscillating heat is then dissipated to the gas that is in contact
with the solid. Equation (7.31) shows that the heat generated at depth x
is oscillating.
By considering the sample thickness from x = 0 to x = -(l + lb) and
gas column length from x = 0 to x = +lg, Rosencwaig and Gersho [23]
have solved the thermal diffusion equation for the system, temperature
distribution in the cell, the periodic heat variation in the gas and the
acoustic pressure wave produced in the cell. The equations are too
complex to deal with here. However, the solution for the incremental
change in pressure in the cell that is responsible for the production of
acoustic signal is shown in Eq. (7.32). The term Q varies with the
opaqueness of the solid sample and the gas in the cell.

6P(t) = Q e- [ (wt -h/4)] (7.32)

156
Rosencwaig and Gersho [23] have divided the solids into six different
categories depending on their optical opaqueness to explain the
dependency of Q and the signal: three cases for optically transparent
and three cases for optically opaque solids according to their relative
magnitude of the thermal diffusion length (p,), as compared to the
thickness of the solid () and optical absorptionlength (p).

(a) For optically transparentsolids where pa > 1, the cases are

1. Thermally thin solids where p, >> 1 and p, > p.
2. Thermally thin solids where ps >> 1 and p < pp.
3. Thermally thick solids where p. < 1 and yp << pp.
For thermally thin solids the signal is proportional to Pl and varies as
co- 1 and depends on the properties of the backing material. For thick
solids the signal depends on Pp,. It means the acoustic signal depends
only on the light absorbed within the thermal diffusion length ps.

(b) For optically opaque solids where 1<<

1, the cases are
1. Thermally thin solids where p >> 1 and p, >> pp.
2. Thermally thick solids where ps < 1 and pi > pp.
3. Thermally thick solids where p, << 1 and p, < pp.
For thermally thin solids, the signal is independent of P and depends on
the thermal properties of the backing material and varies as m-1. For
thermally thick solids in case 2 the signal is independent of and
depends on the thermal properties of the solid and varies as o-l. For
thermally thick solids in case 3 the signal is proportional to 1ps and
dependent on the thermal properties of the solid and varies as -3/2.
When a photoacoustic signal is plotted against wavelength (wave-
number) it gives a spectrum that is similar to the absorption spectrum
of the material.
This is because the photoacoustic signal approximates Beers law
[24]. The theory proposed for the photoacoustic effect of solid materials
has been reviewed by Monahan and Nolle for particulate solid material
[25]. This and several other investigations into the photoacoustic effect
of particulate solid materials show that the effect is dependent on the
particle size of the bulk [26-28]. The photoacoustic signal was shown to
be inversely proportional to the particle size when the samples are
diluted in non-absorbers [24,26-28]. The similarity in the signal
dependency on the particle size of the photoacoustic spectra and

157
reflectance spectra led to the Kubelka-Munk type treatment of the
photoacoustic signal [24,26-28].
Most of the samples are susceptible to photoacoustic detection in the
infrared region [24]. The photoacoustic infrared measurements are
made using carbon black as reference sample. The technique needs
almost no sample preparation and can be used in measuring the
spectra of samples in solid, liquid and gaseous phases. The technique
provides an alternative means of acquiring optical spectra of opaque
solid materials.
The applications of photoacoustic spectroscopy in the infrared
region has increased significantly since the late 1970s. With the high
throughput and multiplexing advantages of modern FT-IR spectro-
meters, photoacoustic spectroscopy can be used as a routine analytical
technique in the mid-IR region. Vidrine [24] has applied the technique
in the mid-infrared region to a wide variety of solid and liquid samples
(Fig. 7.20).

PART B. HYPHENATED TECHNIQUES

The combination of different analytical techniques to tackle a problem

yields the so-called hyphenated techniques. Obviously, the desired
result is the coupling of two or more techniques without sacrificing the
performance of either one. In recent years, coupled or hyphenated
techniques have become popular and have been successfully applied to
the solution of many analytical problems [29,30]. HPLC-MS, tandem
MS-MS, GC-FTIR, TLC-FID, GC-MS, etc. are some of the more
common hyphenated techniques. An overview of the most widely used
combinations that involve FT-IR spectroscopy in polymer analysis
today will be discussed here.

7.5 TGA/FT-IR

The coupling of thermogravimetric analysis (TGA) and Fourier trans-

form IR spectroscopy (FT-IR) is a good practical example of such an
instrumental approach for solving specific analytical problems. This
hyphenated technique, TGA/FT-IR, provides a quantitative assess-
ment of the decomposition process via the thermogram, and an identifi-
cation of the decomposition products from the infrared signatures of

158
 C

Co

-
'-S:
I l, E
.
t-mooL

CJ)'-W __. r,
ow
-wa

0o

C-E

0
r,

.50

I= V

Xi: r 0-
r wI
I
iL r- I
Owi
0 ,
- 0I
m- ,I
1
~nr
(uN-O nOO rO
w_-
YV NN- c'1
LLJ D
I
a j. a, tO0;,-
i:-
r,aT'.
.N stU)....
.- nt ~.
NT
Lil
I
-',U.......
uI
-meo zzz>sIJ

0o0. OO 00

159
the evolved gases. The gases are transferred from the TGA instrument
by means of a heated transfer line to avoid the possibility of conden-
sation. With such a combination, the sample is introduced into the TGA
instrument without any form of chemical or physical modification. The
application of the sequential infrared analysis adds a new dimension to
thermogravimetric analysis by adding specificity, which it otherwise
lacks. An alternative way of looking at the combination is to consider
the TGA instrument as a sample handling or sample treatment front-
end to the FTIR spectrometer, where one can make full use of the
interpretive and diagnostic characteristics of infrared spectral
analyses [31].
There are many examples in the literature where the combination
of TGA-FTIR has been used successfully to study a variety of materials.
In one such study, the effect of aliphatic amines on the mechanism of
the thermal decomposition of polybenzoxazines was investigated [32].
Figures 7.21 and 7.22 show the TGA thermograms of the methyl-dimer
and the amyl dimer, and Fig. 7.23 shows the corresponding FT-IR
spectra of the evolved gases. In this work it was also proposed that the
Mannich base in polybenzoxazines plays a significant role in the ther-
mal degradation of polybenzoxazines. Figure 7.24 shows the proposed
mechanism of Mannich base cleavage in the benzoxazine dinner. The
contribution of hydrogen bonding to the degradation mechanism of the
Mannich base was also examined.
Another example involved the characterization of weathered seal-
ants [33]. In that study, TGA/FT-IR results were compared for silicone
and polyurethane unweathered and weathered sealants (6000 h ex-
posure time). The results indicated that the TGA/FT-IR combination is
useful in the determination of the degradation changes occurring in the
sealants due to weathering. In another study, the characterization of
amine-activated epoxies as a function of cure took place using TGA/FT-
IR [34]. It is known that the physical and chemical properties of cured
diglycidyl ether of bisphenol A (DGEBA) are greatly affected by the
initial and final cure temperatures and cure schedule. These properties
are also affected by the deviation from the stoichiometric ratio of curing
agent used. Analysis of a previously cured epoxy for these parameters
has usually involved large samples and a greater amount of time. In
the experiment described here, a cured epoxy was studied as it
decomposed. During the TGA/FT-IR experiment, evolution profiles for
specific gases were obtained, as well as the normal TGA weight loss
profiles. Using this information, both the cure schedule and epoxy/

160
 .-A
12U 6

100 5{

Z 80
3 al
;60

40
1 '
20 0-
0 -I
0 50 100 150 200 250 300 350 400
Temperature (C)

Fig. 7.21. TGA thermograms of methyl-dimer (solid symbols) and amyl dimmer (open
symbols). (Reproduced from Ref. [32] with permission. Copyright (1998) J. Wiley &Sons.)

8
D
51

Wavenumber (cm'1)
Fig. 7.22. Infrared spectra of evolved gases from degrading amyl-dimer. (Reproduced
from Ref. [32] with permission. Copyright (1998) J. Wiley & Sons.)

activator cure ratios could be established from the analysis of the cured
polymer. The particular material studied, a DGEBA polymer cured
using a primary cycloaliphatic diamine, showed a curing mechanism
similar to that obtained using an aromatic diamine. However, the
decomposition behaviour was more reminiscent of an epoxy cured by
using an aliphatic diamine system. This work demonstrated that a
cured polymer could therefore be characterized in terms of both therm-
al history and activator-resin ratio in a single TGA/FT-IR experiment.

161
 0
4
<

4( D0
Wavenumber (cm'l)

Fig. 7.23. Infrared spectra of evolved gases from degrading methyl-dimer. (Reproduced
from Ref. [32] with permission. Copyright (1998) J. Wiley & Sons.)

OH OH

CH3 CH, CH,

CCH CI H

IH~

CH3
6 cHCj-IN-CH

CH, CH,

Fig. 7.24. Mechanism of Mannich base cleavage in benzoaxazine dimmer. R is the amine
substituent. (Reproduced from Ref. [32] with permission. Copyright (1998) J. Wiley &
Sons.)

162
 In another study, the identification of the decomposition products of
carbon fibre-reinforced phenylethynyl-terminated polyimide compo-
sites (PETI/IM7) resulted in a proposed three-step mechanism for the
decomposition [35]. The assigned glass-transition temperature and the
mechanism for the thermal decomposition of PETI/IM7 were obtained
using DSC, DMA and TGA/FTIR/MS techniques. The assigned glass-
transition temperature was shifted to a much higher temperature after
the sample was cured. TG/MS/FTIR results indicated that water and
NMP were released from the fresh sample during the staging step.
Furthermore, NMP and CO2 were the two major products released from
the sample during the curing step.
TGA/FTIR and isothermal TGA (IGA) was used in a study of poly-
imide thermal oxidative stability to obtain simultaneous identification
and relative quantification of the materials evolved during decompo-
sition in either air or nitrogen [36]. Two high-temperature stable
addition-cured polyimides and two aromatic condensation polyimides,
all four containing fluorinated connecting linkages in the dianhydride
monomers were compared. Figure 7.25 shows the structure of the 3F
and 6F thermoplastics and the PMR-II-50 and VCAP-75 uncrosslinked
thermosets. The TGA-FTIR technique allowed for the simultaneous

3FDA/PPDANPA n=115 {3F)

6FDAIPPDA/PA n-113 (6F)

PMR-II-50 n-9

VCAP-75 n-14

Fig. 7.25. Structures of the 3F and 6F thermoplastics and the PMR-II-50 and VCAP-75
uncrosslinked thermosets. (Reproduced from Ref. [36] with permission. Copyright (1999)
J. Wiley & Sons.)

163
 Temperature,'C

Fig. 7.26. TGA curves for the 3F and 6F polyimides. (Reproduced from Ref. [36] with
permission. Copyright (1999) J. Wiley & Sons.)

Air __ Nitroaen

E
o 0.6
o
0.8 U.

0.6
i----L-,
8 0.4 0.4
Ma

o 0.2' 0.2

0 I 0

350 450 550 650 750 350 450 550 650 750
Temperature, C Temperature, C

3F F-
6F PMR-11-50 - VCAP-75

Fig. 7.27. Evolution profiles of carbon dioxide in air and nitrogen. (Reproduced from Ref.
[36] with permission. Copyright (1999) J. Wiley & Sons.)

identification and relative quantification of evolved decomposition

products (CO,, CO, ArNCO, and CHF3 ) of the four polyimides degraded
in air or nitrogen. Isothermal TGA-FTIR (IGA-FTIR) was also done in
air to determine the relative rate of product evolution at a constant
temperature. Figures 7.26 and 7.27 show the thermograms and the
evolution of carbon dioxide in air and nitrogen. Similar profiles were
obtained for all the evolved gases. As a result, the probable initial

164
 __3
*ffe
0 X .

I
Carbon Monoxide
1
Trifluoromethane and
1
a = Carbon Monoxide
and Carbon Dioxide some Hydrogen Fluoride b+ c = p Phenylene Diisocyanate
b +d = Phenyl Isocyanate

Fig. 7.28. Probable initial decomposition events and identified evolved degradation
products. (Reproduced from Ref. [36] with permission. Copyright (1999) J. Wiley &Sons.)

decomposition events and identified evolved degradation products are

depicted in Figure 7.28.
Other authors were able to exactly assign the volatile components
measured by IR spectroscopy to the decomposition stages detected in
TGA. The thermal decomposition of elastomer sealing rings and of the
thermal behaviour of vinyl flooring material were the two systems
studied [37]. In another study, the thermal degradation of styrene-
sulphonic acid blends and copolymers of styrene was studied [38]. The
blends had enhanced thermal stability relative to virgin polystyrene
but there was no enhancement in thermal stability for the copolymers.
Therefore, it was concluded that it was necessary to have adjacent
sulphonic acid groups to permit the formation of a graphite-like char-
acter which can provide thermal protection to the polymer. It was
therefore necessary to have a good match in degradation temperatures
of the two components if one is to have significantly enhanced thermal
stability.
The utility of TGA/FTIR was demonstrated in an investigation of
several polymeric systems including graft co-polymers of acrylonitrile-
butadiene-styrene (ABS), poly(methylmethacrylate), and styrene-buta-

165
diene block copolymer blends [39]. One of the findings of this work was
that additives may interact with poly(methylmethacrylate) by co-
ordination to the carbonyl oxygen to a Lewis acid and the subsequent
transfer of an electron from the polymer chain to the metal atom or by
the formation of a radical which can trap the degrading radicals before
they can undergo further degradation. When an inorganic char-former
is grafted onto a polymer, there is a good correlation between TGA
behaviour in an inert atmosphere and thermal stability in air. How-
ever, the same cannot be said when the char is largely carbonific.
The effect of two novel epoxypropanecarbazole-based dyes on the
thermal stability of PVC has also been reported [40]. TGA/FTIR was
used to study the effect of the metal on the thermal decomposition of
metal clusters doped poly(acrylic acid) (PAA) and poly(methacrylic
acid) (PMAA) which were prepared by radical decomposition. The
colloids were obtained by condensation of the metals and acrylic acid at
77 K using several metals such as Au, Ag, Pd, Cu, Bi, Sn, Sb, Ge, Ga, In,
Cd and Zn. The presence of the metal clusters avoided the formation of
larger molecular weight polymers. A complete study of the thermal
degradation between 50 and 550C was performed, and the decompo-
sition temperatures were obtained from the second derivatives. In
PAA, the highest decomposition temperature corresponds to the un-
doped polymer, 546C, but in PMMA, the Pd-doped polymer increases
the decomposition temperature from 467 to 469C for the lowest molec-
ular weight. Infrared spectra of the evolved gases indicated that the
decomposition products are mainly MAA and alkenes [41].
A study on thermal degradation of polymeric sulphonic and phos-
phonic acids and their sodium salts was reported in which the
identification of the volatile products and decomposition residues was
key to proposing a mechanism for the degradation [42]. TGA/FTIR
studies used to confirm the thermal degradation mechanism for co-
polymers based on long-chained diol dimethacrylates and BIS-GMA/
TEGDMA 2,2-bis [4-(2-hydroxy-3-methylacrylolyl-oxypropoxy)phenyl]
propane/triethyleneglycol dimethacrylatel [43]. Thermal degradation
studies of polyacrylonitrile [441, poly(styrene-g-acrylonitrile) [45],
poly(N-isopropylacrylamide) [46] and poly(methyl methacrylate)
blended with propyl ester phosphazene [47] were also reported. In the
case ofpolyacrylonitrile, cyclization of the polymer proceeds before any
mass is lost and the driving force for cyclization is the formation of
aromatic rings. The extent of cyclization was controlled by the presence
of head-to-head linkages within the polymer. Ammonia and hydrogen

166
cyanide are the first gases lost and schemes are proposed to account for
their formation. Oligomers are lost from the uncyclized portion of the
polymer lying between the cyclized portions and the amount of non-
volatile fraction is largely determined by the extent of oligomer loss. A
detailed mechanism is presented to account for the observed formation
of the volatile products and the structural changes that occur in the
residue.
TGA-FTIR was also used to identify possible mechanisms of
thermal degradation of Nafion H and Nafion/PMMA blends [48]. A
study of the phase transition and thermal stability of Xydar and Zenite
liquid crystalline polymers was reported [49]. The mechanism of the
thermal decomposition of liquid crystal polymer Vectra B950, a random
liquid crystalline copolymer of ester amide, in both air and in nitrogen
was also studied by the same group [50]. A single-stage decomposition
process was found when heating in nitrogen atmosphere and the
decomposition is due to the ester-linkage rupture. However, a double-
stage decomposition process was found when heating in air, and that
the decompositions are mainly due to the ester linkage rupture for the
first decomposition step and the oxidative reaction for the second
decomposition step. Annealing slightly changes the decomposition
components occurring in the early stage of thermal degradation.
TGA/FTIR has also found application in studies of flammability and
fire-retarding properties of polymeric materials. The effect of various
flame retardants on a PP/PE copolymer was studied using TGA/FTIR
[51]. Specifically, a PP/PE copolymer was successively flame retarded
using Mg(OH) 2 , then using brominated trimethylphenyl indane
associated with Sb 20 3 (Br/Sb), and finally using blends of equal weights
of this last combination with Mg(OH) 2 or talc-containing, non-hydrated
fillers. The decompositions of the pure and the additive-containing
copolymer were studied by TGA/FTIR. It was found that a good
correlation existed between the maxima of Gram-Schmidt curves and
the derivatives of TGA curves. The coupling of techniques shows that
the incorporation of the Br/Sb flame retardant limits strong exothermic
phenomena due to sample ignition. In the case of Mg(OH) 2 associated
with Br/Sb, the decomposition of the hydrated mineral occurs at a lower
temperature than the reaction between brominated trimethylphenyl
indane and Sb2 O3. This delays the action of Br/Sb flame retardant
towards higher temperatures improving the thermal stability of the
polymer. A good agreement is also found between TGA-FTIR con-
clusions and fire resistance tests carried out on standardized samples.

167
 0

-20
e

.t -60
.J

-80

-100
0 500 1000 1500 2000 2500 3000 3500 4000 4500 5000 5500 6000 6500
Time (s)

Fig. 7.29. TGA curves of PP/PE copolymer containing flame retardants. (Reproduced
from Ref. [51] with permission. Copyright (2000) Elsevier.)

3
25-
T
2
1.5
W 1
o0.5
0

-0.5
2000 2500 30010 3500 4000 4500 5000 5500 6000 6500 7000
Time (s)
- ComnbuStes -COZ -- H20

Fig. 7.30. Gram-Schmidt curves of PP/PE with Mg(OH) 2 evolving gases in air. (Repro-
duced from Ref. [51] with permission. Copyright (2000) Elsevier.)

Figure 7.29 shows the TGA curves of PP/PE copolymer containing

flame retardants and Figure 7.30 shows the Gram-Schmidt curves of
PP/PE with Mg(OH) 2 evolving gases in air.
The flammability of a series of polyurethanes based on 2,4-toluene
diisocyanate (TDI), with different amounts of 3-chloro-1,2-propanediol
(CPDO), has been reported [52]. Flammability of the polymers was
determined using the oxygen index (OI) method. The influence of
CPDO on the thermal decomposition parameters (e.g., initial decompo-
sition temperature (IDT), char residue at 500C, and DTG maximum

168
Fig. 7.31. FT-IR spectra of a characteristic sample after 3.0 min. (Reproduced from Ref.
[52] with permission. Copyright (1998) J. Wiley & Sons.)

drA_
- A _

99.0 Fv-( -
Il-ijr~-
00.0

T0.

%T 95.0

94.0

910

92.0

91.0

io.,
fv,~
__
..
I
00O
2 000 1000 7'0
CW1

Fig. 7.32. FT-IR spectra of a characteristic sample after 5.3 min. (Reproduced from Ref.
[52] with permission. Copyright (1998) J. Wiley & Sons.)

temperature) and on the (OI) values were all discussed. Figures 7.31
and 7.32 show the FT-IR spectra of a characteristic sample after 3.0
min and after 5.3 min, respectively. Figure 7.33 shows the correlation
between the OI and IDT and T30%. Some correlations between the

169
 ---
200
-L

T,
IDT [C
[9C1 o
. - 275
106 -

- 270

192 -

- 265

188 -

184 -
i_. - 260

-266

- 250
ran
I I I I I -
20 21 22 23 24 26 26
01

Fig. 7.33. Correlation between the OI and IDT and T30 %. (Reproduced from Ref. [52] with
permission. Copyright (1998) J. Wiley &Sons.)

thermogravimetric data and OI values were found, thus making it

possible to apply the TGA method for certain flammability predictions
of modified polyurethanes.
TGA/FTIR has been applied to the study of the solid-phase transi-
tions of the anti-tumouragent, AG337 [53]. Loss of water was observed
in the range of 20-150 and HC1 was lost in the range of 200-250 . This
hyphenated technique has also been applied to studies of supported
solid acid catalysts. The deactivation and regeneration of sulphated
zirconia was probed in an effort to improve catalytic properties [54].
The deactivation of sulphated zirconia catalysts in the isomerization of
n-butane was also studied [55]. Nafion and Nafion/(silicon oxide)-based
nanocomposites were investigated [56]. The decomposition processes of
CoMgAl-hydrotalcite (CoaMgbAl(OH)c(TA)dnH 2 O) clays intercalated
with terephthalate anions ([C 6H 4(COO)2] 2 - or TA2- ) was studied by a
variety of methods including TGA/FTIR [57]. It was found that the as-
prepared samples undergo different decomposition pathways when
they were heated in air and in nitrogen atmospheres respectively;
particularly the collapse of the layered structure, i.e., dehydroxylation
and the thermal decomposition of TA2 - were overlapped within a

170
 (a)

_-- C.Hyv

H20

U)

r- C02
-1
(b)

r
G.Hy
B 3

H20

C02

0 200 400 600 800

Temperature (C)

Fig. 7.34. Integrated absorbance of the evolved gases with air as the carrier gas.
(Reproduced from Ref. [53] with permission. Copyright (2000) American Chemical
Society.)

narrow temperature range (20-30C) in air, showing a vigorously

exothermic effect. However, the two thermal processes were distinctly
separated in N2. Each thermal process lasted over a rather wide
temperature range (100-200C). Figure 7.34 shows the integrated
absorbance of the evolved gases with air as the carrier gas, and Fig.
7.35 shows the integrated absorbance of the evolved gases with N2 as
the carrier gas.
Carbon nanoparticlesand short tubules were also produced in the
solid state from the intercalated TA2 - anions during the decomposition
of the hydrotalcite-like compounds in N 2. These carbon nanomaterials
are multiwalled and close-ended with a diameter of 10-35 nm and a
length of 20-200 nm. Pathways of the nanocarbon formation as well as

171
 (a)

CxHy

H20
A

Zjj C02

(b) la C
C
ra
C
CxHy

H20

I I
C02

0 300 600 900

Fig. 7.35. Integrated absorbance of the evolved gases with N2 as the carrier gas.
(Reproduced from Ref. [53] with permission. Copyright (2000) American Chemical
Society.)

catalytic function of cobalt oxides generated from the above thermal

processes were further investigated with various instrumental
methods. Finally, the characterization of poly(ureidosilazanes) and
their pyrolytic conversion to ceramic materials was also reported [58].

7.6 LC/FT-IR

The combination of liquid chromatography with infrared spectroscopy

has found extensive use primarily in polymer characterization. The
advantage of combining liquid chromatography with FTIR spectro-
metry is the specific detection or identification of materials.

172
 GiTlmMk (IR tspwanQ

Twit

.rnMnuu
"In rnevyv)

Opt ad c5e on dlac

Fig. 7.36. LC/FT-IR technique.

There are two main approaches: one is to use a flow cell arrange-
ment and continuously record the infrared spectra; the other is to
eliminate the solvent by deposition of the eluent on a substrate and
evaporating the solvent. A recent review [59] focused mainly on the
flow cell method. This review states that flow-cell LC-FTIR methods
have rather poor detection limits but can be useful for the specific and
quantitative detection of major constituents of mixtures. On the other
hand, solvent-elimination techniques provide much better sensitivity
and enhanced spectral quality which is essential when unambiguous
identification of low-level constituents is required. The LC-transform
(Lab Connections) is one of the most common methods for solvent
elimination [60]. It consists of two modules, a sample collection module
and a scanning module accessory. The fist module collects the eluent
from the column as a thin trail around the perimeter of the sample
collection disk. The solvent is removed by using a nozzle with a heated
nebulizer. At the same time, the sample is deposited on the collection
disk [61]. The collection disk is made out of germanium and is placed on
a rotating stage inside the scanning module. This module consists of a
scanning controller and of the necessary optics that couple the infrared
light back to the FT-IR spectrometer. Figure 7.36 shows such an
instrument.

173
 The first demonstration of identifiable infrared spectra obtained
from buffered (volatile and nonvolatile buffers) mobile phases using the
monodisperse aerosol generator interface for combining liquid
chromatography with FT-IIR (MAGIC-LC/FT-IR) spectrometry was
described in 1990 [62]. Ammonium acetate, a volatile buffer, was used
to buffer an 80:20 acetonitrile:water mobile phase to pH 5.0. Caffeine
was deposited from this buffered mobile phase, and the spectrum was
used as a reference to compare with caffeine spectra obtained from
nonvolatile buffered mobile phases. The two nonvolatile buffers used
were potassium hydrogen phthalate (KHP) and potassium dihydrogen
phosphate (KH2PO 4 ). The KH 2PO4 was used to buffer an aceto-
nitrile:water mobile phase and a methanol:water mobile phase, where-
as the KHP buffer was used only in a methanol:water mobile phase.
Samples of caffeine were deposited from each of the above buffer
systems along with the nonvolatile buffer. IR spectra of caffeine were
obtained by spectral subtraction of previously stored buffer spectra
from the caffeine:buffer spectra. The resulting spectra were identical to
a caffeine reference spectrum.
Various examples in the literature show the power of the combina-
tion of liquid chromatography with infrared spectroscopy. Useful solu-
tions to problems of determining polymer composition as a function of
molecular weight for a range of polymers have been illustrated by the
technique. One study focused on the use of a solvent-evaporative
interface in conjunction with a GPC-viscometer chromatograph and a
FTIR spectrometer in order to provide functional-group information as
a function of molecular weight [63]. The GPC-viscometer/solvent evap-
orative interface/FTIR system was applied to a variety of polymer and
coatings systems as a tool for product problem solving and elucidation
was presented. In addition, examples of the use of the solvent evapora-
tive interface to elucidate compositional heterogeneity of copolymers
are illustrated. The potential use of the solvent evaporative interface in
GPC/LC cross fractionation studies for very fine elucidation of polymer
compositional heterogeneity was also explored.
This hyphenated technique has also been used in the character-
ization of asphalt binders based on chemical and physical properties
[64]. The chemical composition and physical properties of unmodified
and polymer modified asphalts were studied using a variety of tech-
niques including GPC \ FT-IR. Two viscosity-based asphalt grades and
two polymers (styrene-butadiene-styrene and styrene-ethylene-
butylene-styrene) that were used to modify asphalt were studied. The

174
combination of GPC and FTIR was an excellent approach for finger-
printing and quality control of polymers and asphalt binders. In
addition, the theological properties of asphalt binders were good
characteristics for determining the optimum polymer concentrations
for effective modification.
A thermospray was modified and used to couple HPLC to FTIR
spectrometry, which was applicable to both normal- and reversed-
phase HPLC [65]. Column effluents from the HPLC system were
desolvated by thermospray and solutes were deposited as individual
spots on a moving stainless steel belt substrate, which continuously
transferred the analytes into the diffuse reflectance (DRIFT) accessory
of the FTIR, enabling identification of deposited solutes by measure-
ment of the IR spectrum. The thermospray temperature and thermo-
spray height were shown to influence the deposition of solutes. By use
of a heated external nitrogen gas flow, desolvation of the reversed-
phase HPLC eluents was improved.
A coupled GPC/FT-IR system was also developed to measure short
chain branching as a function of molecular weight in polyethylenes and
ethylene copolymers in relatively short time scales. Careful selection of
the IR detector, use of a low volume flow-through cell with a large
optical path length and selecting GPC conditions to maximize the
polymer concentration in the cell enabled the characterization of poly-
mers with very low average comonomer concentrations. A method for
calibration of the infrared detector was presented and results for a
series of polyethylenes of known average co-monomer content, VLDPE,
LLDPE, MDPE and broad molecular weight polyethylenes were also
presented to illustrate the capability of the system. The quality of the
data from the GPC/FTIR can be assessed with results on the same
polymers obtained using other fractionation techniques. It was found
that reliable results could be obtained above MW of approximately
10,000. However, at low molecular weights where chain end corrections
become large for infrared measurements, values were confirmed with
measurements obtained using NMR spectroscopy.
In another study, where this time solvent elimination did not take
place, infrared spectroscopy was proposed as a molecular specific
detection system for HPLC in an aqueous phase, focusing on the
chromatographic separation of sugars in beverages. The separation
was achieved with an isocratic HPLC setup using an ion exchange
column with Ca 2 ' serving as the counterion. The FT-IR detection of the
C-O bands in the mid-IR between 1000 and 1200 cm -1 was performed

175
 re

0.03

8 0.02
C LUCOSE

0.01

OSE

1400 1300 1200 1100 1000 900

wavenumber cm"

Fig. 7.37. Three-dimensional plot of a standard solution containing 10 mg/ml each of

sucrose, glucose and fructose. (Reproduced from Ref. [65] with permission. Copyright
(1997) American Chemical Society.)

0.006

0.005

0.004

; 0.003

f 0.002

1400 1300 1200 1100 1000 900

wavenumrnber
cm'

Fig. 7.38. FT-IR spectra extracted from the peak maxima of HPLC peaks. (Reproduced
from Ref. [65] with permission. Copyright (1997) American Chemical Society.)

in real time with a 25 pm flow cell without elimination of the solvent.

Characteristic FT-IR spectra of the common sugars sucrose, glucose,
and fructose in concentrations of 1 mg/ml were recorded during the
separation process. The calibration of these compounds in the 5-100
mg/ml range resulted in a linear correlation with a standard deviation
of 0.11, 0.07, and 0.11 mg/ml for sucrose, glucose, and fructose, respect-

176
 0

0 5 10 15 20 25 30
retention time Imin

Fig. 7.39. HPLC-FTIR traces of a taurine-containing soft drink. (Reproduced from Ref.
[65] with permission. Copyright (1997) American Chemical Society.)

0.010

0.008

0.006

0.004

0.002

-0.001
1400 1300 1200 1100 1000 900
wavenumber /cmn

Fig. 7.40. Taurine (A) and ethanol (B) spectra extracted from the HPLC-FTIR run of a
taurine containing soft drink. (Reproduced from Ref. [65] with permission. Copyright
(1997) American Chemical Society.)

ively. Figure 7.37 shows a three dimensional plot of a standard solution

containing 10 mg/ml each of sucrose, glucose and fructose, whereas Fig.
7.38 shows the FT-IR spectra extracted from the peak maxima of HPLC
peaks.
The method was, furthermore, applied to the analysis of nine soft
drinks and fruit juices containing between 6 and 97 mg/ml of each
carbohydrate.The accuracy of the method was confirmed by standard

177
 la
<
le

Analysis time

Fig. 7.41. Average baseline-corrected chromatogram. (Reproduced from Ref. [66] with
permission. Copyright (1997) American Chemical Society.)

ion exchange HPLC with refractive index detection. The average

deviation from the reference method was in the range of 0.5-0.9 mg/ml.
Furthermore, the method was able to identify and quantify minor
components in beverages, such as taurine (4 mg/ml) and ethanol (0.4
mg/ml). Figure 7.39 shows the HPLC-FTIR traces of a taurine-con-
taining soft drink whereas Figure 7.40 the taurine and ethanol spectra
extracted from the HPLC-FTIR run of a taurine containing soft drink.
For many situations, chemometric methods are used to enhance the
data analysis. In one such study, the analysis of a reaction product by
HPLC coupled with Fourier transform spectroscopy LC/FTIR origin-
ates a series of overlapping peaks [66]. The applicability of the
orthogonal projection approach, the fixed size window evolving factor
analysis approach, and the evolving factor analysis approach for the
resolution of those overlapping peaks into individual chromatograms
and infrared spectra is discussed. The reaction product under study
was the result of the reaction of a straight-chain alkyl alcohol with
citric anhydride. The results are evaluated by identifying the different
compounds present using the spectral characteristics of the resolved
pure compound spectra and the chemistry of the system. Figure 7.41
shows the average baseline-corrected chromatogram and Fig. 7.42 the

178
 Wavenumber (cm-i)

Fig. 7.42. Pure component spectra of one of the clusters. (Reproduced from Ref. [66] with
permission. Copyright (1997) American Chemical Society.)

pure component spectra of one of the clusters. Analysis of the infrared

spectra suggested that the main compound of cluster 3 is an ammo-
nium salt of dialkyl citrate ester.

179
 LC-FTIR was also used to assist in the complete structure eluci-
dation of a globular protein, -lactoglobulin [67]. Other efforts in this
area have focused on improvements in instrumentation. For example, a
heated gas flow modified thermospray was developed which facilitates
the delivery of evaporated eluents for IR analysis using diffuse
reflectance [68]. Another report discussed the use of a temperature
programmed packed capillary liquid chromatograph coupled to an off-
line FTIR spectrometer for the analysis of the antioxidant, Irgafos P-
EPQ [69].
Liquid chromatography (LC) was coupled semi-online to FTIR
spectrometry using a spray-jet assembly interface to eliminate the LC
eluent prior to IR detection [70]. The usefulness of the LC-FTIR system
in the identification of closely related compounds in complex mixtures
was demonstrated by the analysis of a chlorinated pyrene sample
which contains a number of chloropyrene isomers and congeners.
Characteristic FTIR transmission spectra of all constituents were
recorded. Since most of these compounds were not analyzed by IR
before, spectral assignment was mainly based on the empirical
Hansen-Berg rules for substituted pyrenes. The identification limits
for the chloropyrenes typically were 5-10 ng.

7.7 GC/FTIR

This is the most commonly used technique in coupling a chroma-

tographic analysis to an FT-IR detection system. The technical
development that made GC/FTIR so widespread was the introduction
of the gold-coated glass 'lightpipe' [71,72]. This design is based on a
small-volume gas cell which is constructed to match the elution
volumes of the GC peaks. In addition, the fact that the carrier gases
used as a mobile phase do not absorb in the infrared region has helped
the application of the technique. The detection limits achievable with
this technique are at the nanogram level, thanks to improvements in
capillary column technology.
GC/FTIR methods were used to establish identification limits for
volatile organics in blood [73]. These methods were based on the purge-
and-trap extraction technique. FTIR identification limits were
measured for a number of volatile organic compounds. The FTIR
identification limits, ranging from 0.01 mg/l for ethyl acetate, methyl-
ethyl ketone, and sevoflurane to 24 mg/I for methanol, generally
allowed the detection of volatile-substance exposure at a lower level

180
than is acutely toxic. Quantitative calibration data were presented for
selected substances, based on the FID response, which shows that the
method is also amenable to quantitative analysis. The throughput of
the method without additional automation is five samples per day, the
purge-and-trap stage being the limiting factor.
In another study, a GC/FTIR/MS technique was utilized for the
determination of unusual amidine products obtained by the sublima-
tion of amino acids in the presence of silica catalyst [74]. Furthermore,
the identification of individual naphthenes in a mixed hydrocarbon
sample was made using GC/FTIR with neural network and regression
analysis [75]. Specifically, the goal was to differentiate between cyclo-
pentane- and cyclohexane-containing compounds in mixed aliphatic
hydrocarbon samples. A set of single-ring model compounds was used,
with acyclic aliphatic hydrocarbons used as counter-examples. The GC-
FTIR method was envisioned as a means of determining individual
naphthenes in a sample.
Mathematical techniques have also been employed to improve GC/
FTIR spectra. In one such study, it was found the commonly used co-
addition of spectra to a relative intensity level of 40% of the GC peak
does not lead to the optimal improvement in S/N of the resulting
composite spectrum for either simulated Gaussian-shaped or experi-
mentally obtained asymmetric GC bands. The optimal intensity level
for co-addition is a function of the shape of the GC band and the ratio of
the number of background to sample scans used in generating the
individual IR spectra. The authors also introduced the use of classical
least-squares (CLS) techniques as a superior method to improve the S/
N of the composite analyte spectrum. Using CLS methods, spectra
included in generating the composite spectrum can be a small fraction
of the maximum intensity in the GC peak while still resulting in S/N
improvements. The theoretical S/N of the composite spectrum using
CLS methods is always as good as or better than that achieved with the
co-addition method. The improvements achieved in S/N when CLS
methods are used can be more than a factor of 2 greater than results for
the traditional co-addition method for the cases considered. Also,
increasing the number of background to sample scans was a very
convenient method to improve the S/N of the composite spectrum
obtained by either method. The results for GC/FTIR are also generally
applicable to LC/FTIR, SFC/FTIR, and TGA/FTIR for bands that
contain a single analyte [76]. In addition, a set of the semi-empirical
methods was tested to find the best auxiliary tool for the GC/FTIR

181
spectroscopy-mass spectrometry identification of cyclic amide-type
compounds [77].
Finally, the pyrolysis of aquatic humic substances was studied by
pyrolysis GC/FTIR [78]. The humic substances studied gave similar
pyrolysis products, but in varying proportions. Many of the pyrolysis
products (e.g., methanol, acetone, alkylbenzenes, cyclopentane,
aliphatic and aromatic organic acids, acetamide, pyrrole and phenols)
were identified by their FTIR spectra using a digital library for
automatic comparison. Some of the compounds were related to lignin
fragments, which form a large part of the humic substances investi-
gated. Other products gave hints as to the involvement of tetrapyrroles,
fatty acids, furanoses, and amino compounds in the structure of humic
macromolecules.

7.8 SFC/FTIR

The use of supercriticalfluid chromatography (SFC) is growing very

rapidly and is a very active area of research nowadays. A supercritical
fluid is a material that is held above its critical temperature and
pressure. These fluids show a behaviour between a liquid and a gas and
the interest in them comes mainly from environmental reasons. Their
use resembles the use of organic solvents in extraction methods, there-
fore one can imaging the advantages of using a typical supercritical
fluid such as CO2 instead of organic solvents.
Applications of supercritical fluids range from extractions and
chromatography to solvents for reaction chemistry. They are also used
in the preparation of new materials. A resent review covered many
aspects of the field in detail [79]. Spectroscopic monitoring is important
in all of the above cases and vibrational spectroscopy is particularly
useful in this context because the vibrational spectrum of a given
molecule is usually quite sensitive to the environment of that molecule.
Thus, vibrational spectra are excellent probes of conditions within the
fluid. The authors showed examples that included the use of super-
critical Xe as a spectroscopically transparent solvent for chemistry and
for supercritical fluid chromatography with FTIR detection of analytes
as well as the use of supercritical CO 2 .
In another study, packed-column supercritical fluid chromato-
graphy with a mobile-phase gradient of CO2 and methanol was
performed on a mixture of 8 sulphonamides [80]. The analytes were
detected and identified with FT-IR. A fixed-integral restrictor afforded

182
the decompression zone between the column and germanium disk
where the analytes were deposited.
The comparison of direct-deposition supercritical fluid and gas
chromatography/Fourier transform infrared spectra to condensed-
phase library spectra also took place in an attempt to bring attention to
potential pitfalls [81]. In particular, comparisons of spectra from direct-
deposition (DD) capillary gas chromatography (GC) and supercritical
fluid chromatography (SFC)/FTIR measurements of two quinones with
C2 symmetry axes and several barbiturates to spectra from condensed-
phase libraries of the corresponding compounds were reported. The
best spectral search results were obtained when the eluites were
deposited on an amorphous substrate, such as ZnSe. A small number of
polar, H-bonding compounds align with each other or with a crystalline
substrate. Different crystalline forms of some polymorphic analytes
can also yield ambiguous identifications. These effects produce enough
differences in the DD GC/FTIR and SFC/FTIR spectra to cause
occasional incorrect identifications when the spectra are searched
against KBr-disk library spectra.
Polydimethylsiloxanes modified by the introduction of ethylene
oxide units were characterized using a variety of SFC coupled
techniques including SFC/FTIR [82]. The aim of this work was to form
sufficiently water soluble siloxane compounds. In another study
essential oil constituents of hops (Humulus lupulus) were characterized
using SFC/FTIR techniques. Infrared spectra of these constituents
were taken as films deposited on AgCl disks and compared with those
obtained after chromatographic separation in the IR flow-cell with
supercritical carbon dioxide. Spectra from AgCl disks were comparable
to those in supercritical CO,, but in supercritical CO2 most of the bands
appeared approx. 8-10 cm-l to higher wavenumbers. Open-tubular
SFC-FTIR analysis of the essential oil of four different hop varieties
was performed. The SFC-FTIR chromatograms showed differences in
the location and relative intensity of the peaks depending on the
variety, which was further confirmed by the FTIR spectra.
Also, a supercritical fluid extraction method for the analysis of
nonionic surfactants in a washing powder is presented. By varying the
extraction conditions, it is possible to fractionate the extracted
materials according to their polarity. The subsequent analysis and
identification of the nonionic surfactants by SFC/FTIR or mass
spectrometry detection did not require any additional sample
preparation. Most nonionic surfactantswere extracted by SFE without

183
 CO,

gas
d)

integral restricto erature

or

germanium disk

Fig. 7.43. Configuration of the nozzle assembly and the integral restrictor. (Reproduced
from Ref. [85] with permission. Copyright (1997) Elsevier.)

using a modifier to enhance the polarity of the supercritical CO2 . Other

substances can then be extracted using supercritical CO2 with 5%
MeOH as modifier [83].
Furthermore, SFC/FTIR was used to simultaneously detect
aromatic and aliphatic nonvolatile compounds of microwave susceptor
packaging [84]. Microwave susceptor packaging is designed to brown
and crisp food in a microwave oven. During microwave cooking, the
package can reach temperatures above 260C. Because of these high
temperatures and the multilayer construction of the packages, many
types of chemicals can potentially migrate into the food. SFC/FTIR
spectroscopy in series with flame-ionization detection provided a
unique solution to the simultaneous detection of the above compounds.
The authors demonstrated the utility of SFC for determining these
potential migrants in solvent extraction of several microwave sus-
ceptor packaging materials. The extraction of one package studied
contained 2-(2-butoxyethoxy)ethanol and several aliphatic chemicals
whose FTIR and mass spectra are characteristic of aliphatic ketones.
The LC-Transform, routinely used for LC/FTIR analysis, was
shown to perform very well for SFC/FTIR separation of Irganox 1076
[85]. In particular, the separation of Irganox 1076 with 5% methanol
modified CO 2 was demonstrated. High quality spectral data were

184
 IR spectrum Jflrganox 1076

I
4
.S
-

4000 3000 2000 1000

Wavenumters (cm-1)

Fig. 7.44. FTIR spectrum of Irganox 1076 collected using the configuration shown in Fig.
7.43. (Reproduced from Ref. [85] with permission. Copyright (1997) Elsevier.)

obtained throughout the 4000-700 cm-l range with no interference

from methanol. The only modification to the commercially available
interface for SFC/FTIR was the insertion of a fixed integral restrictor
into the spray nozzle. Figure 7.43 shows the configuration of the nozzle
assembly and the integral restrictor. The modified CO 2 mobile phase is
evaporated during the spray process, and the resulting chromatogram
is deposited as a continuous track on an IR transparent (e.g., usually
germanium) flat surface (e.g., the sample collection disk). This track
can then be scanned in a spectrometer to obtain spectra of the discrete
sample components. Figure 7.44 depicts the FTIR spectrum of Irganox
1076 collected via this procedure.
A direct deposition system for SFC/FTIR analysis of environmental
analytes was also evaluated [86]. A direct-deposition FTIR system was
evaluated for applicability to gas chromatography and supercritical
fluid chromatography of environmental analytes. A 100-pjm internal
diameter fused-silica transfer line was used for GC, and a 50-pm
transfer line with an integral restrictor was used for SFC. Minimum
identifiable quantities for GC/FTIR ranged from 0.5 to 2.0 ng. Some of
this sensitivity improvement can probably be attributed to the smaller
internal diameter GC column used in this work and the ability to
program the direct-deposition sample plate, thereby compensating for
changes in analyte elution volume across the GC temperature ramp.
Figure 7.45 shows the GC/FTIR spectra of DDT, dioctyl phthalate and
fluoranthene. In addition, excellent SFC/FTIR chromatograms was
obtained for poly(ethylene glycols) of average molecular weights of 400,
600, 1000, and 1500 respectively. Figure 7.46 shows the SCF/FTIR
spectra of poly (ethylene glycol) at two different times. The decreasing
ratio of-OH to -CH stretching modes as the retention time increases is

185
 l -

0004-

0.002-

"I\
to)
0.004

CU
r_
to
A 0.002

0.000

U.uu - tc)

0.004-

0.000-
1I
3000 2600 2200 1600 1400 1000
Wavenumber (cmrl)

Fig. 7.45. GC/FTIR spectra of DDT, dioctyl phthalate and fluoranthene. (Reproduced
from Ref. [86] with permission. Copyright (1994) American Chemical Society.)

consistent with the increasing ration of recurring molecular chain

residue to terminal unit -OH.
Chemometric methods are also commonly used in SCF/FTIR
spectroscopy [87]. In one such study principal component analysis was
applied to the rapid discrimination of extractable compounds that are
indigenous to papers and nonindigenous compounds based on their

186
 0.12
(a)

0.00
w
0
z
r

.:
0.20

(b)

0.00 - __ /
A ~nu ~J
lIII I.U LJ
4000 3000 2000 1000
-
t
WAVENUMBER (cm ')

Fig. 7.46. SCF/FTIR spectra of poly(ethylene glycol) at two different times. (Reproduced
from Ref. [86] with permission. Copyright (1994) American Chemical Society.)

infrared signatures. This method was applied with online supercritical

fluid extraction and SCF/FTIR to yield a fully automated analysis of
compounds that can be extracted from very complex matrixes.
Finally, the analysis of subnanogram quantities of analytes was
demonstrated using 600 pg of caffeine [88]. The minimum identifiable
quantity for caffeine (a strong IR absorber) obtained with this interface
was 600 pg (injected), whereas the level of detection was at 1-10 ng for
weaker absorbers. Spectra over the entire mid-infrared region of
compounds separated with a mobile phase of carbon dioxide modified
with two percent methanol were recorded. The interface shows linear
behaviour over two orders of magnitude for both the area under a
strong absorption band in the IR spectra vs. injected quantity and for
the area under a peak from the functional group chromatogram versus
the injected quantity.

187
7.9 INFRARED MICROSPECTROSCOPY

7.9.1 FT-IR microscope and redundant aperturing

Infrared microspectroscopy is a technique where microscopic technique

is used in combination with infrared spectroscopy. It has been revived
after the introduction of modern FT-IR spectrophotometers and
sensitive detectors such as mercury-cadmium-telluride(MCT) and has
been proved to be one of the powerful analytical tools both in analytical
and industrial applications [89,90]. Infrared microspectroscopy
provides the analytical chemist not only with the visual viewing of the
material under the microscope but also with the chemical information
of the part of the material that is being viewed in the form of its
infrared spectrum.
The instrumentation in infrared microspectroscopy is specially
designed to use with infrared radiation. Optics in an ordinary micro-
scope do not transmit infrared radiation. To avoid this problem,
reflecting type optics are used. Most of the infrared microscopes use
Cassegrain type reflecting objectives. These objectives can be used
either as a condensing lens or as an objective lens.

liable aperture

granian Objective

Samnle

Condenser X . i

Variable aperture

Fig. 7.47. Sketch of an FT-IR microscope in transmission and reflectance mode.

188
 5amnlD sujrf2ce

a)

b)

ndant Aperture
I I
Fig. 7.48. The redundant aperturing technique: (a) sample surface with two different
chemical compositions; (b) redundant aperture size that is suitable for the analysis of
these materials.

Modern FT-IR microscopes are made to perform the following

functions: (a) irradiation of a micro sample with infrared radiation; (b)
collection of the radiation emerging from the sample; (c) imaging the
collected radiation on to a detector (usually mercury-cadmium-
telluride detectors) that is connected to the microscope and FT-IR
instrument; and (d) allowing the user to select the precise area he/she
wants to measure.
Figure 7.47 shows a general lay out of an FT-IR microscope in
transmission and reflection mode. The microscope can also be used as a
visible microscope. This function allows the user to select the area of
interest for analysis. The visible and IR optical paths are made to be
collinear and parfocal to ensure identical optical paths. This also
ensures the user, the area selected for FT-IR measurement and the
area actually measured are the same. As mentioned above, the FT-IR
microscopes employ reflecting mirrors (Cassegrainian optics) for the
optical elements instead of lenses in the visible light microscopes. The
aperture placed before the cassegrainian objective gives a better
sample definition. In transmission mode, the infrared radiation is
transmitted through the sample and condensed by a cassegrainian

189
condenser and imaged through another aperture. This aperture block
diffracted light that is strayed from the adjacent sample area. This
technique is called redundantaperturing(Fig. 7.48).
Modern infrared microscopes give users the opportunity for
measuring infrared spectrum of a prescribed area of a sample either in
transmittance mode or reflection mode. Furthermore, when thin layers
of materials are present on completely reflecting bases, reflection-
absorption techniques can be used to measure the spectrum. Here, the
specular reflection is minimum and the returning radiation will be
attenuated by penetrating the sample twice. The spectrum can be used
in the same way as a transmittance or absorbance spectrum. There are
accessories available that can be connected to infrared microscopes to
measure infrared spectra using the attenuated total internal
reflectance technique. The samples are measured using a suitable
reference. For example, when samples are deposited on reflecting
bases, the base itself can be used as the background.
Several applications involving infrared microspectroscopy are
discussed in Chapters 8 and 9.

REFERENCES

1. P.R. Griffiths and J.A. deHaseth, Fourier Transform Infrared

Spectroscopy. Wiley, Chichester, 1986.
2. H.A. Willis, J.H. van der Maas and R.G.J. Miller (Eds.), Laboratory
Methods in VibrationalSpectroscopy, 3rd edn. Wiley, Chichester, 1987.
3. N.J. Harrick, Internal Reflection Spectroscopy. Interscience, New York,
1987.
4. F.M. Mirabella, Jr. (Ed.), Internal Reflection Spectroscopy: Theory and
Application. Marcel Dekker, New York, 1993.
5. M.W. Urban, Attenuated Total Reflectance Spectroscopy of Polymers;
Theory and Practice. American Chemical Society, Washington, 1996.
6. M. Claybourn, Infrared Reflectance Spectroscopy of Polymers. Global
Press, 1998.
7. Y. Ozaki, F. Kaneuchi, T. Iwamoto, M. Yoshiura and Iriyama, Appl.
Spectrosc., 43 (1989) 138.
8. M. Tasumi, S. Ochiai, H. Kato, Y. Furukawa and K. Masutani, Principle
andPracticeof FT/IR Spectroscopy, 2nd edn. Tokyo Kagaku Dojin, Tokyo,
1994, p. 105 (in Japanese).
9. D.W. Vidrine, Mid-Infrared Spectroscopy in Chemical Process Analysis,
in: J.M. Chalmerr (Ed.), Spectroscopy in Press Analysis. Sheffield
Academic Press, Sheffield, 2000, p.9 6.

190
10. N. Mitsui, H. Higashiyama and M. Watari, Keiso, 38(12) (1995) 46 (in
Japanese).
11. W.G. Golden, Fourier Transform Infrared Spectroscopy, J.R. Ferraro and
L.J. Basile (Eds.), Vol. 4. Academic Press, New York, 1987, p. 315.
12. M.W. Urban, VibrationalSpectroscopy of Molecules and Macromolecules
on Surfaces. Wiley Interscience, New York, 1993.
13. M.P. Fuller and P.R. Griffiths, Anal. Chem., 50 (1978) 1906.
14. I.M. Hamadeh, S.A. Yeboah, K.A. Trumbull and P.R. Griffiths, Appl.
Spectrosc., 38 (1984) 486.
15. R.A. Velapoldi, J.E. Tvedt and A.A. Christy, Rev. Sci. Instrum., 58 (1987)
1126.
16. S.A. Yeboah, S.H. Wang and P.R. Griffiths, Appl. Spectrosc., 38 (1984)
259.
17. A.A. Christy, J.E. Tvedt and R.A. Velapoldi, Rev. Sci. Instrum., 59 (1988) 3.
18. A.A. Christy, O.M. Kvalheim and R.A. Velapoldi, Vib. Spectrosc., 9 (1995)
19.
19. R.L. White and A. Nair, Appl. Spectrosc., 44 (1990) 69.
20. C. Bremard, C. Depecker, H. Des Grousilliers and P. Legrand, Appl.
Spectrosc., 45 (1991) 1278.
21. R.L. White and J. Ai, Appl. Spectrosc., 46 (1992) 93.
22. A.A. Christy, E. Nodland, A.K. Burnham, O.M. Kvalheim and B. Dahl,
Appl. Spectrosc., 48 (1994), 5.
23. A. Rosencwaig and A. Gersho, J. Appl. Phys., 47 (1976) 1.
24. D.W. Vidrine, Appl. Spectrosc., 34 (3) (1980) 314.
25. E.M. Monahan and A.W. Nolle, J. Appl. Phys., 48 (8) (1977) 3519.
26. J.J. Freeman and R.M. Friedman, J. Phys. Chem., 84 (1980) 315.
27. R.S. Davidson and D. King, Anal. Chem., 56 (1984) 1409.
28. C.Q. Yang and W.G. Fateley, J. Mol. Struct., 146 (1986) 25.
29. B.W. Cook, Spectroscopy (Eugene, Oreg.), 6(6) (1991) 22-26.
30. K. Norton, A.J. Lange and P.R. Griffiths, J. High Resolut. Chromatogr.
14(4) (1991) 225-259.
31. S. Materazzi, Appl. Spectrosc. Rev., 32(4) (1997) 385.
32. Hong Yee Low and H. Ishida, J. Polym. Sci., PartB: Polym. Phys., 36(11)
(1998) 1935-1946.
33. R.M. Paroli and A.H. Delgado, Can. Polym. Mater. Sci. Eng., 69 (1993)
139.
34. J.D. Johnson, D.A. Compton, R.S. Cass and P.L. Canale, Thermochim.
Acta, 230 (1993) 293.
35. Y. Tang, Y. Xie, W.-P. Pan and A. Riga, Thermochim. Acta, 357-358 (2000)
239-249.
36. M.J. Turk, A.S. Ansari, W.B. Alston, G.S. Gahn, A.A. Frimer and D.A.
Scheiman, J. Polym. Sci., Part A: Polym. Chem., 37(21) (1999) 3943-
3956.

191
37. F. Hoffmann, R. Riesen and J. Foreman, Am. Lab. (Shelton, Conn.), 32(1)
(2000) 13-14, 16-17.
38. Q. Yao and C.A. Wilkie, Polym. Degrad. Stab., 66(3) (1999) 379-384.
39. C.A. Wilkie, Polym. Degrad. Stab., 66(3) (1999) 301-306.
40. K. Pielichowski, I. Hamerton, J. Pielichowski and P. Stanczyk, Eur.
Polym. J., 34(5/6) (1998) 653-657.
41. G. Cardenas, C. Munoz and H. Carbacho, Eur. Polym. J., 36(6) (2000)
1091-1099.
42. D.D. Jiang, Q. Yao, M.A. McKinney and C.A. Wilkie, Polym. Degrad.
Stab., 63(3) (1999) 423-434.
43. K. Pielichowski, D. Bogdal, J. Pielichowski and A. Boron, Thermochim.
Acta, 307(2) (1997) 155-165.
44. T.J. Xue, M.A. Mckinney and C.A. Wilkie, Polym. Degrad. Stab., 58(1-2)
(1997) 193-202.
45. T.J. Xue and C.A. Wilkie, Polym. Degrad. Stab,. 56(1) (1997) 109-113.
46. H.G. Schild, J. Polym. Sci., PartA: Polym. Chem., 34(11) (1996) 2259-
2262.
47. B-L. Denq, Y-S. Hu, W-Y. Chiu, L-W. Chen and Y-S. Chiu, Degrad. Stab.,
57(3) (1997) 269-278.
48. M.L. Mittleman, J.R. Thomsen and C.A. Wilkie, Polym. Mater. Sci. Eng.,
63 (1990) 957-961.
49. T-S. Chung and X. Jin, Polym. Eng. Sci., 40(4) (2000) 841-856.
50. T-S. Chung, M. Cheng, P.K. Pallathadka and S.H. Goh, Polym. Eng. Sci.,
39(5) (1999) 953-962.
51. J.-P. Gibert, J.-M. Lopez Cuesta, A. Bergeret and A. Crespy, Polym.
Degrad. Stab., 67(3) (2000) 437-447.
52. K. Pielichowski, J. Pielichowski and A. Prociak, Pol. J. Appl. Polym. Sci.,
67(8) (1998) 1465-1471.
53. S. Rastogi, I. Zamansky, S. Roy, P. Tyle and R. Suryanarayanan, Pharm.
Dev. Technol., 4(4) (1999), 623-632.
54. B. Li, R. Marcus and R.D. Gonzalez, Prepr. - Am. Chem. Soc., Div. Pet.
Chem., 44(4) (1999) 430-433.
55. B. Li and R.D. Gonzalez, Appl. Catal., A, 165(1/2) (1997) 291-300.
56. Q. Deng, C.A. Wilkie, R.B. Moore and K.A. Mauritz, Polymer, 39(24)
(1998) 5961-5972.
57. Z.P. Xu and H.C. Zeng, J. Phys. Chem. B, 104(44) (2000) 10206-10214.
58. D. Seyferth, C. Strohmann, N.R. Dando and A.J. Perrotta, Chem. Mater.,
7(11) (1995) 2058-2066.
59. G.W. Somsen, C. Gooijer and U.A.T. Brinkman, J. Chromatogr., A, 856(1/
2) (1999) 213-242.
60. J.N. Willis and L. Wheller, Polym. Mater. Sci. Eng., 69 (1993) 120.
61. M.X. Liu and J.L. Dwyer, Appl. Spectrosc., 50 (1995) 349.

192
62. R.M. Robertson, J.A. de Haseth and R.F. Browner, Appl. Spectrosc., 44(1)
(1990) 8-13.
63. T. Provder, M. Whited, D. Huddleston and C-Y. Kuo. Prog. Org. Coat., 32
(1997) 155.
64. J.B. Wei, J.C. Shull, Y.J. Lee and M.C. Hawley, Int. J. Polym. Anal.
Charact., 3(1) (1996) 33.
65. M.A. Mottaleb, Anal. Sci., 15(1) (1999) 57-62; R. Vonach, B. Lendl and R.
Kellner, Anal. Chem., 69 (1997) 4286-4290..
66. F. Cuesta Sanchez, B.G.M. Vandeginste, T.M. Hancewicz and D.L.
Massart, Anal. Chem., 69(8) (1997) 1477-1484.
67. V.E. Turula, R.T. Bishop, R.D. Ricker, J.A. de Haseth, J. Chromatogr.,A,
763(1-2) (1997) 91-103.
68. M.A. Mottaleb, Anal. Sci., 15(1) (1999) 57-62.
69. I. Bruheim, P. Molander, E. Lundanes, T. Greibrokk and E. Ommundsen,
J. High Resolut. Chromatogr., 23(9) (2000) 525-530.
70. U.A.Th. Brinkman, N.H. Velthorst and T. Visser, Anal. Chim. Acta, 290(3)
(1994) 269-276.
71. L.V. Azarraga, Appl. Spectros., 34 (1980) 224.
72. R.P. Griffiths, J.A. deHaseth and L.V. Azarraga, Anal. Chem., 55 (1983)
1361.
73. I. Ojanpera, K. Pihlainen and E. Vuori, J. Anal. Toxicol., 22(4) (1998)
290-295.
74. V.A. Basiuk, Spectrochim. Acta, Part A, 55A(2) (1999) 289-298.
75. J.D. Jegla, P.R. Griffiths and D.P. Klein, Prepr.Am. Chem. Soc., Div. Pet.
Chem., 41(4) (1996) 663-666.
76. D.M. Haaland, E.V. Thomas and D.S. Blair,Appl. Spectrosc., 47(10) (1993)
1612-1619.
77. V.A. Basiuk, Spectrochim. Acta, PartA, 55A(2) (1999) 289-298.
78. R. Kuckuk, W. Hill, P. Burba and A.N. Davies, Fresenius'J.Anal. Chem.,
350(7-9) (1994) 528-532.
79. M. Poliakoff, S.M. Howdle and S.G. Kazarian, Angew. Chem., Int. Ed.
Engl., 34(12) (1995) 1275-1295.
80. M. Ashraf-Khorassani, M.T. Combs, L.T. Taylor, J. Willis, X. Liu and C.R.
Frey, Appl. Spectrosc., 51(12) (1997) 1791-1795.
81. K.L. Norton, A.M. Haefner, H. Makishima, G. Jalsovszky and P.R.
Griffiths, Appl. Spectrosc., 50(9) (1996) 1125-1133.
82. U. Just, D. Jones, R.H. Auerbach, G. Davidson and K. Kappler, J. Bio-
chem. Biophys. Methods, 43(1-3) (2000) 209-221.
83. R.H. Auerbach, K. Dost and G. Davidson, J. AOAC Int., 83(3) (2000)
621-626.
84. E.M. Calvey, T.H. Begley and J.A.G. Roach, J. Chromatogr. Sci., 33(2)
(1995) 61-65.

193
85. S.H. Smith, S.L. Jordan, L.T. Taylor, J. Dwyer and J. Willis, J.
Chromatogr., A 764(2) (1997) 295-300.
86. D.F. Gurka, S. Pyle, R. Titus and E. Shafter, Anal. Chem., 66(15) (1994)
2521-2528.
87. J. Yang, E.J. Hasenoehrl and P.R. Griffiths, Vib. Spectrosc., 14(1) (1997)
1-8.
88. K.L. Norton and P.R. Griffiths, J. Chromatogr., A, 703(1/2) (1995) 503-
522.

194
Chapter 8

Applications of infrared spectroscopy in

basic and industrial research

For many years now, infrared spectroscopy has enjoyed universal

acceptance as a chemical characterization tool across almost all the
disciplines of science. The usefulness of the technique can be attributed
to its ease of use, the cost of the instruments, and the plethora of
applications that come from the use of different sampling techniques. It
should, however, be pointed out that mass spectroscopy and nuclear
magnetic resonance (NMR) are nowadays the primary identification
tools for structural elucidation of organic compounds, with vibrational
spectroscopy playing a complementary role.
Therefore, the greatest value of infrared characterizations comes
from technicalities associated with the ease of use of the technique. Due
to the number of different subjects that can be covered in this topic we
are forced to concentrate our attention on a few selected examples.
These examples deal mainly with the use of FT-IR spectroscopic tech-
niques to polymeric materials [1], organic thin films and biological
materials,though applications of other types of materials will also be
presented.

8.1 POLYMER APPLICATIONS

8.1.1 Infrared characterization of polymers and polymeric

surfaces

Infrared spectroscopy has been used extensively in polymer character-

ization for a long time, providing information on chemical nature,
isomerization, conformational order, state of order, and orientation

195
 [2,3]. The use of vibrational spectroscopy as a characterization tool to
measure orientation in polymers has been reviewed recently [4].
Some of the most recent examples of the use of infrared spectroscopy
in the characterization of polymeric materials will be presented here.
In such examples, FT-IR spectroscopy has been applied as a bio-
diagnostic tool of polymer implants and tissue surfaces [5]. In this
particular study, surface analysis allowed the determination of the
specific molecular compounds and structures most appropriate for
long-term compatibility in humans. Important information associated
with the bioinertness or bioactivity of implants was obtained from the
spectral features of the polymer material used, including the level of
polymerization.
In another study, fluorocarbon compounds based on vinylidene
fluoride copolymers and bisphenol AF were prepared to determine the
network-forming structures of the cured materials [6]. Sections from
key stages of processing were taken and their FT-IR spectra were
recorded. These spectra established directly, for the first time, that
bisphenol AF served as the crosslinker during cure. Additionally,
persistent unsaturation was formed on the elastomer backbone after
crosslinking. It was also observed that curing for extended periods of
time produced no observable effect on the network. Furthermore, post-
curing reduced residual hydrofluoric acid in the compound and
resulted in the appearance of new absorptions at 2851 and 2920 cm l,
respectively, which are indicative of amorphous regions of poly(vinyl-
idene fluoride). These data served as an indicator in the understanding
of the fracture behaviour and long-term performance of this class of
materials.
Also the use of segmented polyurethanes as biomedical implant
materials was studied by vibrational spectroscopic probes combined
with measurements with angular dependent XPS or ESCA [7]. These
data provided a detailed description in the surface composition of
Biomer and Avcothane which are commercially available biomedical
grade polymers. In addition, the surface composition of the model
system polydimethyl siloxane (DMS) was also studied. Both attenuated
total reflectance (ATR) and photoacoustic (PA) techniques were
utilized. The authors were successful in elucidating the depth of
segregation of DMS blocks in Avcothane as well as the presence of DMS
in the very top surface of Biomer. It was found that this combination of
techniques worked better in the understanding of these complex
polymer surfaces.

196
 In another study, the thermal degradation of CO-ethylene-
propylene alternating copolymers was followed by FTIR spectroscopy
[8]. The infrared spectra of solid samples, performed in inert atmos-
phere and under high vacuum, were recorded as a function of time at
different temperatures. These data indicated that the reaction process
consisted of intra or intermolecular hydrogen transfer, yielding an enol
and a small quantity of unsaturated species. At temperatures above
the melting point, scission of the polymer chain occurred and the
product had large number of unsaturated fragments.
In another study, the radical grafting reaction of maleic anhydride
into poly(propylene oxide) (PPO) was followed by infrared spectroscopy.
PPO is widely used in the preparation of thermoplastic elastomers,
surfactants, and additives. A protection of -OH end-groups of PPO was
realized by acetylation in order to prevent side-reactions of these
groups with anhydride. FT-IR spectroscopic studies were able to follow
the extent of the grafting reaction due to the appearance of a character-
istic shift in the carbonyl region of cyclic anhydrides to higher wave-
numbers [9].
The copolymers of tetrachloroethylacrylate (TeCEA) and penta-
fluorophenylacrylate (PFPA) and of TeCEA and pentafluoro-
phenylmethacrylate (PFPMA) were examined with regard to their
applicability as core materials for optical waveguides [10]. In
particular, their thermal properties as a function of polymerization
conditions were investigated and optimized by addition of crosslinkers.
The polymerization reactions were followed by FT-IR spectroscopy,
which was able to successfully quantitate the unsaturation level in
these polymers. In addition, optical characteristics such as material
dispersion and attenuation of the polymers were also studied. The
fundamental demands on waveguide polymers are high optical
transmission at the near-infrared region around 1300 and 1550 nm, the
adjustability of the refractive index of the core polymer versus the
cladding polymer, and finally sufficient thermal stability of at least
70C. Both the examined copolymers show low absorption at 1300 and
1550 nm of 0.2 and 0.7 dB/cm respectively. The refractive index of
TeCEA/PFPA copolymer is tunable between 1.464 and 1.518 at 1300
nm. In addition, the TeCEA/PFPMA copolymer is tunable between
1.469 and 1.518 at the same wavelength.
In another study, the first attempt to investigate polymer-
surfactant interactions in gelling and non-gelling aqueous mixtures of
a nonionic cellulose ether and a surfactant by means of vibrational

197
 w
Z
z

400 600 800 1000 1200 1400

FREQUENCY (cm-1)

Fig. 8.1. IR absorption (dashed line) and Raman (solid line) spectra of solid EHEC at
room temperature. (Reproduced from Ref. [11] with permission. Copyright (1999)
American Chemical Society.)

spectroscopy was recorded [11]. In particular, a series of aqueous

solutions of ethyl hydroxyethyl cellulose (EHEC) with addition of
anionic surfactant sodium dodecyl sulphate (SDS) of different con-
centrations was investigated by FTIR absorption techniques. Figure
8.1 shows the IR absorption and Raman spectra of solid EHEC at room
temperature. The data showed that, even in the solid state, i.e., below
the gel point, interactions between the polymer and the surfactant
were present. Figure 8.2 shows the IR absorption spectra of EHEC
aqueous solutions (4 wt%) with different SDS concentrations at room
temperature. In addition, both bound and free surfactant molecules
were detected. This interaction, which cannot be characterized as
chemical, occurred mainly between the side chains of the polymer and
the sulphonic acid groups of SDS. Figure 8.3 shows the infrared spectra
of several EHEC/SDS/water systems in sol and gel states. Above the gel
point, a new type of interaction appears, which mainly involves the
S0 3 - groups and water molecules. The intermolecular interactions
were studied versus the changes of both temperature and polymer-
surfactant compositions and a possible model for the gelation process
was discussed.
This study found that there was an interaction between EHEC and
SDS even at temperatures below the gel point, i.e., in the solution state,
though the effect was weak and can hardly be characterized as a
chemical interaction. However, when the temperature rose, a new type
of interaction appeared in the system that could be associated with

198
 w
C
W
zZ
cf
o
0
O0
co

1000 1050 1100 1150 1200 1250 1300 1350 1400 1450
WAVENUMBER (cm-')

Fig. 8.2. IR absorption spectra of EHEC aqueous solutions (4 wt%) with different SDS
concentrations (at room temperature). (Reproduced from Ref. [11] with permission.
Copyright (1999) American Chemical Society.)

enhanced chain mobility. The most prominent one was the interaction
between the SO, groups of SDS and the glucose rings, which led to an
essential decrease of the mobility of the rings and also to their
deformations. Finally, clear changes in the state of water occurred with
gelation. In the gel state the hydrogen bonds between water molecules
become stronger. Moreover, the degree of H-bond strengthening
increased with the level of SDS addition, which suggested that this
effect was inspired by the S03 groups.
Furthermore, the effect of interelectrode spacing on the properties
of hydrogenated amorphous Si (a-Si:H) films grown by RF plasma
enhanced CVD method with control of dusty plasma conditions by
heating both the electrodes was studied [12]. The formation of pre-
cursors responsible for gas phase polymerization itself was thought to
be controlled by preheating of the source gas mixtures. Optimization of
the interelectrode spacing for film characteristics was carried out for
this novel deposition technique that combined cathode heating and
preheating of the source gases. The films were characterized by infra-
red spectroscopy along with absorption and reflection measurements in
the visible and near-infrared regions, and ESR spectroscopy.

199
 I
U
C
r
aC
C
G
0

O0

Wavenumber (cm- 1)
Fig. 8.3. IR absorption spectra of several EHEC/SDS/water systems in sol (solid line) and
gel (dashed line) states. Letters denote the sample type (see Table 8_1). (Reproduced from
Ref. [11] with permission. Copyright (1999) American Chemical Society.)

In another study, poly(8-quinolyl acrylate) and the polymers of the

complexes of 8-quinolyl acrylate with CuBr, NiBr2, CoBr 2 and uranyl
acetate were prepared and characterized by elemental analyses,
electronic and vibrational spectroscopic studies and magnetic moments
[13]. Furthermore, by measuring the shift of satellite infrared peaks it
was possible to determine the value of interactive bond elongation in
dilatons [14]. Measurement of the satellite intensity permitted
determination of the concentration of regularly formed sequences in
polymers. Investigation of the dependence of this concentration on
temperature, time, and load permitted study of the regularities of

200
formation kinetics of dilatons. These spectroscopic data indicated that
dilatons generally originated in the surface layers of polymer solids.

8.1.2 Phase behaviour of polymer blends [15]

In one representative study, poly(ethylene oxide) (PEO) and poly(vinyl-

phenol) (PVP) interacted quite strongly in the solid state via the
formation of hydrogen bonds [16]. The interaction is specific at the
molecular level and centres around the hydroxyl group of PVP. High-
resolution FTIR and 13C NMR spectroscopy provided convincing evi-
dence that the infrared absorption of the -OH group and the phenolic
carbon resonance of PVP are sensitive to the presence of the semi-
crystalline PEO. It was postulated that the free electron pairs of the
ether O of PEO provided the complementary site for H bonding with the
OH proton of PVP. Melting, crystallization, and single glass transition
behaviour were all discussed from a phase-diagram viewpoint and
represent macroscopic responses that are driven by molecular level
associations. Flory-Huggins analysis of the melting point depression
phenomenon suggested that the thermodynamic interaction para-
meter, X2, per monomer unit of PEO is -1.5 in the vicinity of the
melting temperature. This implied that the energetics of mixing are
exothermic and contribute favourably to concentration dependent
miscibility in the amorphous phase. Thermodynamic analysis of melt-
ing point depression via the X12 parameter was consistent with the
infrared results, which revealed that the interaction between the two
dissimilar polymers is stronger than the self-association of hydroxyl
groups in PVP.
The blending between poly(methyl methacrylate) (PMMA) and
ferroelectric (vinylidene fluoride-trifluorethylene) [P(VDF-TrFE)]
copolymer chains was investigated by FTIR spectroscopy over the full
range of composition, for the copolymer with 50 mol% of trifluor-
ethylene (TrFE) [17]. The FTIR spectra revealed an absorption band at
1643 cm l, characteristic of the blend and absent in the individual
constituents. This band was attributed to the interaction of the
carbonyl group of the PMMA side chains with the disordered helical
chains present in the amorphous region of the P(VDF-TrFE). The
consequences of adding PMMA onto the formation of the all trans
conformation of the copolymer chains was also investigated and the
effects of thermal heating on the spectra were relevant only for the
samples where the ferroelectric semicrystalline phase was present.

201
8.1.3 Deuteration studies

Isotopic exchange and direct deuteration in particular, are common

techniques in IR spectroscopic studies directed at elucidation of poly-
meric structure [18,19]. Frequency shifts upon isotopic substitution are
expected due to the mass dependence of the vibrational frequency in
the simple harmonic oscillator model. These frequency shifts can be
predicted with reasonable accuracy from the changes in the atomic
masses alone, since the force constants will remain essentially
unchanged.
Krimm's rule [20,21] is an approximation rule that applies to
hydrogen-deuterium substitution

VJVk = [1-(ATi/pT)]- /2 (8.1)

where k and vk, are the zero order frequencies of the kth vibrations for
-H and -D groups, respectively; T is the total kinetic energy; ZAT is
the change in kinetic energy upon isotope exchange' and Eis the ratio of
isotopic to normal mass, respectively.
For the case of polyethylene, Table 8.1 shows the predicted versus
the actual frequency for deuterated polyethylene (PE). The FT-IR
spectrum of low density/perdeuterated high density (LDPE/d*-HDPE)
is shown in Figure 8.4. These values compare very well with those
calculated by the application of Krimm's rule.

8.1.4 Orientation measurements

Orientation in polymers can be measured by a variety of techniques,

such as x-ray diffraction, NMR, birefringence, polarized fluorescence,

TABLE 8.1
Predicted versus actual frequencies for deuterated PE

Wavenumber ratio observed (CH2/CD2) predicted Vibrational assignments

1.342 1.349 asymmetric stretching
1.372 1.379 symmetric bending
1.341 1.349 bending
1.384 1.379 rocking

202
 W
U
Z

VI
C
--
t

WAVENUMBERS

Fig. 8.4. LDPE/d*-HDPE FT-IR spectrum; 2 cm-l resolution.

Raman depolarization, sonic techniques and infrared dichroism. The

latter technique is one of the most frequently applied tools for the
characterization of anisotropy in polymers. Most polymer systems are
subjected to the application of stress during manufacturing. The stress
is either applied in one direction (uniaxial stretching), or it can be
applied along two perpendicular directions (biaxial stretching). The
elucidation of the molecular mechanisms that take place during
elongation is of great importance to the polymer industry [22].
Classic rheo-optical studies emphasize the direct relationship
between the perturbation and the spectral response. In addition,
dynamic FT-IR spectroscopy provides the element of time in the
individual spectral responses of different parts of the molecules with
respect to the external perturbation. Before the discussion of the rheo-
optical studies in polymeric systems, a brief introduction to the theory
of infrared linear dichroism will take place.

8.1.5 Infrared linear dichroism

In general, maximum absorption takes place when the electric vector is

parallel to the transition moment of the specific normal mode

203
(functional group) and no absorption will take place when the electric
vector is perpendicular to the transition moment. The absorption of
each mode is proportional to the square of the dot product of the electric
E and transitionmoment M vectors, according to the equation

I = (EM)2 cos 2 O (8.2)

where k is a proportionality constant and 0 is the angle between the two

vectors.
In the case of a polymer macromolecule, the finally observed
absorbance A is the sum of the intensity contributions from all the
structural units of the polymer (n)
2
A = k '(EM) dn (8.3)
n

The effect of the anisotropic distribution of the transition moments

with respect to the direction of the electric vector E of the polarized
radiation is characterized by the dichroic ratioR

R =All/A1 (8.4)

where Al and A are the absorbances measured with radiation

polarized parallel and perpendicular to the stretching direction. The
value of R can range from zero (where there is no absorption in the
perpendicular direction) to infinity (no absorption in the parallel
direction). For random orientation, R = 1. If R is greater than 1 the
band is called a parallel band; if R is smaller than 1 it is called a
perpendicular band. Therefore, several kinds of information can be
acquired from the knowledge of R, namely, the elucidation of the
molecular geometry by the determination of the transition moment
directions of particular functional groups with respect to the molecular
axis and unambiguous assignment of various modes to specific
symmetry types of the normal modes.
An important parameter for every absorption band is the so-called
structuralabsorbanceAo

A o = (A +Ay +Az)3 (8.5)

which represents the absorbance of the band without the contributions

due to the orientation of the polymer. For uniaxially oriented sample,

204
 preferred orientation

tentvector

X
Fig. 8.5. A schematic representation of the distribution of the molecular chains and their
corresponding dipoles with respect to the draw axis.

produced by stretching in one direction, the structural absorbance

becomes
Ao = (A + 2A)/3 (8.6)
In practice, the orientation is never perfect and this effect can be
simulated by supposing that, on average, all the molecular chains are
displaced by the same angle from the preferred orientation (stretching
axis). Figure 8.5 is a schematic representation of the distribution of the
molecular chains and their corresponding dipoles with respect to the
draw axis. The Herman orientationfunction F is expressed by Eq. (8.7):

F= [3 < cos2 0 > - 1]/2 (8.7)

where 0 is the angle between the draw direction and the local molecular
axis chain. This orientation function can be related to experimentally
measured quantities, such as the dichroic ratio R of the absorption
band according to Eq. (8.8):

F = (R - 1)(R + 2)/(R - 1)(R + 2) (8.8)

where Ro = 2 cot 2a is the dichroic ratio for perfect uniaxial order. As a

varies from 0 to /2, Ro varies from infinity to zero. No dichroism is
observed at the so-called magic angle (for a = 5444 ' , Ro becomes unity).

205
 Some examples of the application of linear dichroism techniques to
polymers will be presented here. In one such study, linear dichroism
spectra in the mid-infrared region of fluorene aligned in stretched
polymers and nematic liquid crystals were compared with earlier
experimental and theoretical investigations [23]. Linear dichroism
spectra of molecules aligned in such anisotropic solvents were simple to
obtain, especially for small and medium-sized molecules. Most
molecules obtain a satisfactory alignment by these methods and the
spectra were simple to interpret. The information obtained from these
spectra about vibrational transition moment directions was often
crucial for vibrational assignments in molecules like fluorene. Even
high quality calculations are still unable to provide safe assignments
for all the fundamental vibrations in fluorine, as long as they are only
compared with traditional spectra. Linear dichroism spectra provided
a separation of the experimental information according to symmetry
classes. This procedure reduced the assignment puzzle drastically. In
the present case the result was an almost complete and safe assign-
ment of all symmetry-allowed fundamentals of fluorene. In a similar
study, samples of aligned molecules may be produced by using stretch-
ed polymers as anisotropic solvents [24]. Although the molecular
alignment is rarely perfect and the orientation distribution function is
not known, an exact, simple, and useful mathematical description of
the partial alignment was possible. The aligned samples could be
studied by optical spectroscopy with linearly polarized light in order to
gain information on the properties of the molecular alignment, on the
structure of the solute molecules and their interaction with the solvent,
or on spectral assignments of electronic and vibrational transitions in
the solute molecules. It is usually assumed that the alignment of solute
molecules in stretched polymer sheets is uniaxial around the stretching
direction, even in thin polyethylene sheets. The validity of this assump-
tion was investigated and confirmed through a series of measurements,
using different angles between the direction of the linearly polarized
light beam and the plane of the stretched sheet. The degree of align-
ment of solutes in stretched polyethylene is known to increase when
the temperature is lowered. A systematical study was carried out of the
alignment of a solute molecules in stretched polyethylene as a function
of temperature. This study showed that the main change in alignment
takes place at temperatures around -10 . The change was associated
with large improvements in alignment within the crystalline regions of
polyethylene. As a practical consequence, the improved alignment,

206
which was previously obtained by cooling to LN 2 temperatures, may be
easily available at more convenient temperatures.

8.1.6 Dynamic FT-IR procedures

The majority of the dynamic and time resolved experiments with

modern FT-IR instruments take place with the use of step-scan inter-
ferometry. Since a lot of the examples that will be presented later
involved the use of dynamic FT-IR techniques, a short description of
these techniques will take place here.
For the purposes of this book, dynamic infrared spectroscopy is
defined as the use of infrared spectroscopy to monitor a time-dependent
process. The study of the dynamics of the vibrational excitation/de-
excitation process itself is outside the scope of this work. However,
since the time-scale of this process is of the order of 10-13 s or less,
changes in the IR spectrum can be used to monitor the dynamics of
slower processes, within the practical limits of the speed of the detector
and electronics and the strength of the signal.
Operationally, dynamic spectroscopy can be divided into experi-
ments which use the impulse-response technique ("time-resolved"
spectroscopy) and those which use synchronous modulation techniques
("phase-resolved" spectroscopy) [25]. In the first case the dynamic
response to a perturbation is monitored as an explicit function of time;
in the second case the phase and magnitude of the response with
respect to that of the perturbation are measured. It has been previously
stated that these two types of experiment are actually closely related
(by the Fourier transform) and may be considered as limiting cases of a
general modulation experiment which uses n frequencies [26]. For the
impulse-response experiment n - c, while for the synchronously
modulated experiment, n = 1.
Until recently, dynamic infrared spectroscopy has been restricted to
the study of either relatively slow processes or limited wavelength
ranges. Dispersive spectrometers with point by point data collection or
very slow scanning offer access to broad wavelength range but require
very long data collection times to achieve this and are seriously limited
by low throughput. However, for synchronous modulation experiments,
over limited wavelength ranges, quite good results for time-resolution
in the us range have been achieved in reasonable times by use of
dispersive IR [27-30]. Tunable laser radiation using either a gas phase
laser (such as CO) or a diode laser is another possible approach to

207
dynamic IR spectroscopy. The high intensity per spectral bandwidth of
the laser make such techniques excellent from the point of view of both
spectral and temporal resolution, but the requirement to make
measurements essentially point by point (i.e., without any multiplex
advantage) and the limited tuning ranges available, restrict the
general utility of such techniques.
The synchronous modulation step-scan FT-IR spectra are acquired
most of the time through a double demodulation experiment. In these
experiments, the phase modulation of the IR beam, produced by the
"jitter" of the moving mirror is used as a carrier frequency for the
intensity modulation induced by the electric field reorientation of the
liquid crystal sample, or at the mechanical stress modulation
frequency.
The amplitude of the phase modulation can be varied between a few
nm and several pm, in order to maximize the efficiency of modulation in
the spectral range of interest [31]. For example, an amplitude of 2 XHeNe
(1.26 pm) is used for full mid-IR spectroscopy. The infrared beam is
initially passed through optical filters that remove optical frequencies
outside the useful range, thus reducing the level of noise in the
acquired spectra. Undersampling may be used in order to reduce
acquisition time. In this technique, the correct combination of optical
filters and sample spacing eliminates the unwanted effect of aliasing
(folding) [32], while at the same time producing the desired spectral
range. The infrared beam also passes through a gold wire-grid polari-
zer which allows only light polarized parallel to the initial orientation
of the liquid crystal director to reach the sample. The dynamic spectra
thus show only changes in the infrared absorption which are associated
with the reorientation of the transition dipoles in response to the
mechanical perturbation [33]. The acquisition time used to be sub-
stantial for this kind of experiment, due to the fact that the changes
under investigation are very small (on the order of 10 4 absorption
units). Modern research-grade FT-IR spectrometers equipped with the
step-scan option and DSP collection electronics reduce the acquisition
time considerably.

8.1.7 Fourier transform dynamic infrared spectroscopy

The use of Fourier transform interferometric techniques for dynamic

vibrational spectroscopy offers the combination of broad free spectral
range with relatively high spectral resolution and fast data acquisition

208
times. However, again, until recently, harnessing the multiplex and
throughput advantage of FT-IR to dynamic measurements has been of
limited success except for low resolution measurements and/or
monitoring relatively slow processes.
There are, in fact, several ways to perform dynamic FT-IR measure-
ments. First, if the lifetime of the event under investigation is > 10x the
shortest scan period of the conventional continuous-scan interfero-
meter, each interferogram can be considered to be instantaneous.
However, even though scan rates of > 50 Hz have recently been
achieved on commercial instruments (using bi-directional scanning),
this is always at the expense of resolution and, in any event, can only be
applied for a minimum of -20 ms time resolution. In addition, this is a
technique limited strictly to the impulse/response mode.
Another continuous scan approach, which is applicable to synchro-
nous modulation experiments is to scan the mirror slowly enough that
the highest Fourier frequency generated in the spectral bandwidth of
interest is more than 10x lower than the external modulation applied
to the sample. This method has been successful, but it requires an
interferometer of exceptional stability [34]. Even so, it is not practical
for external modulation frequencies of < 400 Hz, except in the far-IR.

8.1.8 Step-scan dynamic FT-IR

As an alternative to the continuous scan mode of interferometry, the

data may be collected in the step-scan mode, in which the retardation is
changed incrementally and data are collected while the retardation is
held constant. As previously stated, step-scan interferometry is more
universal in its application than is the continuous-scan method since it
can be applied without fundamental restrictions, to either synchronous
modulation or impulse/response experiments. Since the retardation is
constant while data are collected (or, as in some cases, the retardation
is modulated about a fixed valve), the spectral multiplexing is un-
coupled from the time-domain. Furthermore, step-scan FT-IR offers
both conceptual and practical simplicity since the experimental para-
meters can generally be changed independently and with ease.
Although the step-scan mode of interferometry predates the
continuous-scan mode by decades, until recently it has not been widely
available to experimenters in a form suitable for routine use outside
the far-IR [28]. Both types of time-resolved experiment, either impulse-
response or synchronous modulation experiments, can be performed

209
using the step-scan mode of operation. For impulse/response experi-
ments data are collected as an explicit function of time at the desired
intervals after each impulse. The perturbation/impulse is repeated at
each step as many times as necessary to achieve the desired signal to
noise. Data from all times are sorted by time and transformed to
produce the time-resolved spectra.

8.1.9 Step-scan impulse-response experiments

The development during the last decade of modern step-scan interfero-

metry instrumentation has allowed FT-IR to be applied to the study of
time-dependent phenomena in ways not previously possible, because of
the problems of uncoupling the spectral multiplexing from the temp-
oral domain in the continuous-scan FT-IR mode [35]. Specifically, the
time regime from tens of nanoseconds to tens of milliseconds has been
accessible to time-domain measurements to only a very limited degree
with continuous-scan instrumentation and not at all for modulation-
demodulation (frequency-domain) experiments in this time range. The
step-scan technique not only works very well in this time regime and
for slower phenomena, but is only prevented from application to faster
processes by the signal strength, the speed of available detectors, the
intensity of sources, and the speed and sophistication of the electronics.
As in the synchronous modulation experiments, it is necessary that
the response of the system to the perturbation in the impulse-response
time-resolved, or "time-domain" mode should be perfectly reversible, so
that any desired number of repetitive pulses can be used to achieve the
necessary signal averaging. In these experiments a signal from the
spectrometer externally triggers the pulse generator, which then
produces a voltage step across the cell and maintains this voltage for a
time along with respect to the response time of the sample. As stated
above, repetitive pulses (separated by a suitable recovery interval) at
each interferometer position are used for signal averaging. The result-
ing data are then sorted by time to produce individual interferograms
for each time t, which are then transformed to give the time-resolved
spectra.
Figure 8.6 shows the data collection scheme for the simplified case
of one excitation pulse for each retardation step. The data are sorted
vertically to produce the time-resolved interferograms. The time inter-
vals are usually equal, but this is only an experimental convenience,
not a fundamental requirement.

210
 Retardation Time Intervals
60 to*, tl, t2, . .....tn

1 to*, t, t2 , . . ., tn

$n to*, tl, t2, . . ., tn

Fig. 8.6. Data collection scheme for step-scan impulse-response experiment.

An example of the application of a infrared time resolved technique

will briefly take place here. In this study, a time-resolved infrared
spectrometer in the spectral region of 700-4000 cm-l was constructed
with a resolving power of 50 ns and detection limit of 10-6 [36]. Iso-
merization of retinal by light irradiation was examined to suggest an
isomerization mechanism from trans form to 3:1 mixture of 13-cis and
9-cis forms within < 50 ns. The time-resolved IR spectra of N,N-
dimethylamino-4-benzonitrile in polar BuOH and nonpolar hexa-
decane with and without oxygen bubbling gave conclusions that the
2096 cm-l band appearing only in polar solvent with a life of several ns
and no oxygen effect was due to CN stretching vibration of the excited
singlet state of twisted intramolecular charge transfer structure. A
2040 cm-l band appearing also in polar solvents with longer life in the
order of 100 ns and disappearing by oxygen bubbling was due to CN
stretching vibration of the excited triplet state.

8.2 APPLICATIONS TO LIQUID CRYSTALS AND LIQUID CRYSTAL

POLYMERS

Thermotropic liquid crystal polymers have awakened a great deal of

interest in the past decade from both technological and scientific points
of view. Great emphasis was placed on the modification of physical and
thermal properties, and analysis of the corresponding structure-
property relations [37]. Vibrational spectroscopy is a useful tool in the
characterization of these materials, and the type of information
available through infrared spectroscopy relates to crystallinity and
polymorphism, phase transitions and orientational behaviour.

211
 Efforts to investigate the responses of liquid crystals to applied
electric field using vibrational spectroscopy started in 1981, using
attenuated total internal reflectance (ATR) spectroscopy [38]. In
addition, the time course of the reorientation is most suitably studied
by the use of dynamic vibrational spectroscopy. Both time-resolved
Raman spectroscopy [391 and dynamic infrared spectroscopy [40] have
been applied to the electric-field induced reorientation of nematic
liquid crystals. Coles and Tipping used microsecond time-resolved
Raman spectra of a nematic liquid crystal as a function of the applied
electric field [41]. Kaito et al. used fast FT-IR scanning with milli-
second time resolution to study the time-dependent polarized infrared
absorption of a nematic liquid crystal [42]. Toriumi et al. published the
first stroboscopic FT-IR data with sub-millisecond time resolution in
1988 [43]. In 1991, Gregoriou et al. published the first synchronous
modulation data on the reorientational behaviour of a nematic liquid
crystal in response to an AC electric field [44].

8.2.1 Dynamic IR spectroscopy of polymers

Excellent reviews of the relevant instrumental and theoretical back-

ground of polymer deformation and relaxation studies by simultaneous
Fourier-transform IR spectroscopic and mechanical measurements
exist in the literature [45,461. In the first review, the vibrational
spectroscopy of stressed polymers, orientational measurements using
infrared dichroism, deuteration, and experimental results for thermo-
plastics and elastomers are discussed.
Dynamic rheo-optical IR techniques promise an exiting future for
vibrational spectroscopy as a tool on polymer research [47]. The earlier
work by Noda et al. [48] has been successfully adapted to interfero-
metric measurements using step-scan FT-IR techniques [49]. Noda et
al. [50] performed a dynamic infrared linear dichroism study of high
density and low-density polyethylene films near the -transition
temperature. It was found that a different deformation mechanism
operates above and below T. Specifically, a negative dynamic
dichroism at 1473 cm7l and a positive dynamic dichroism at 1463 cm -l
were observed at 32C. This observation was interpreted to mean that
above T, the orthorhombic crystallographic b axis reorients parallel to
the direction of applied strain, while the crystallographic a axis
reorients perpendicular to the strain direction for both HDPE and

212
LDPE. The dynamic dichroism of both bands changes sign at -50C
(below Tp), indicating that the dynamic reorientation directions for the
crystalline a and b axes are shifted below To. In this earlier work with
dispersive instrumentation switching between spectral regions was a
difficult task, requiring a substantial investment in time.
Lefebre et al. have performed infrared measurements on the PS/
PPO compatible blend in terms of static uniaxial strain above the glass
transition temperature [51]. Evidences that the PPO and PS chains
orient in a different way were found, in spite of the compatible nature of
the blend. The PPO orientational behaviour does not depend on PPO
concentration in the concentration range that was studied (0-35%)
while PS orientation regularly increased up to 25% PPO and then
remained constant. The authors suggested two explanations for this
behaviour. Either the increase of the PPO concentration results in an
increase of the knots of the physical network, or the relaxation of the PS
chains is hindered by PPO chains. The first explanation is not
supported by their experimental results, since the PPO orientation
remained constant as the concentration increased.
In a similar study, side chain liquid crystalline polyurethanes are a
new class of materials that show promise for mechano-optic applica-
tions. The rich morphology afforded by these materials also provided a
chance to understand the interplay between polyurethane morphology
and liquid crystalline ordering. In this study, the response of a poly-
urethane with liquid crystals pendant to the soft segments to an
applied strain using Fourier Transform Infrared (FT-IR) linear
dichroism was detected. It was found that this complex material
followed trends established in the literature for both side chain liquid
crystalline homopolymers and segmented polyurethanes. At low
strains, the soft segments aligned with strain inducing an orientation
in 'lone' hard segments. Up to strains of 40%, the LC mesogens align
with the strain field and the hard segments in hydrogen bonded
domains align perpendicular to the field. At strains above 40%, a
rearrangement of the ordering was found that resulted in the smectic
layers and the hard segments aligning parallel to the field. In addition,
dynamic FT-IR experiments showed that the viscoelastic reorientation
of various segments of the macromolecule could be monitored as a
function of the applied strain. For the polyurethane under study, the
cyano band was used to follow mesogen movements, and the urethane
carbonyl to track the hard segment. Evidence were presented for two
types of hard segments: those involved in hydrogen bonding within

213
hard domains, and those found in 'lone' hard segments in the soft
matrix. Evidence were also found for two types of mesogens: those
found in smectic layers, and those not involved in smectic ordering at
the hard domain interface. The hard domains and the smectic layers
had strong viscous components to their mechanical response. The 'free'
mesogens and the 'lone' hard segments, on the other hand, exhibited a
more elastic response. A model was proposed to represent these
findings, and reflections on the cooperative movement of the different
macromolecular components of the polyurethane were offered [52,53].

8.3 APPLICATIONS TO OPTICALLY ACTIVE MATERIALS

8.3.1 Organic light emitting diodes (OLEDs)

Electroluminescentdevices based on organic low molecular weight (e.g.,

Alq 3) and polymeric materials (e.g., PPV) are recently attracting much
attention mainly due to applications as large area light-emitting
devices (OLEDs). Generally, these devices are thin-film single-layer or
multilayer structures composed of a hole transport and an emitting and
an electron transport material sandwiched between two electrodes.
OLED devices are generally fabricated utilizing vapour deposition
techniques or film-casting techniques from solution.
The structure and the correlation between intermolecular inter-
actions and optical properties in various such systems have been
investigated with infrared spectroscopy among other methods. Some
selected examples will be presented here.
In one such study, the authors reported the presence of carbonyl
moieties as defects, formed during the thermal conversion of a
precursor to poly(p-phenylenevinylene) (PPV) [54]. The increase in
carbonyl groups was correlated with a dramatic reduction of PPV
photoluminescence. If the conversion is carried out in a reducing
atmosphere, e.g., 15% hydrogen in nitrogen, and the amount of
carbonyl moiety was substantially reduced and the photoluminescence
intensity of the polymer increased as much as five-fold.
In another study, thin films of a cross-linkable hole-conducting
monomeric triarylamine for use in organic light emitting diodes were
examined [55]. The rate of photo-crosslinking and the overall polymeri-
zation yields were measured using real-time FTIR spectroscopy. The
electronic properties were characterized in a typical diode configura-

214
 OC.H, OCH,,
OC,H, _0 CH,

H,C, O, H,,Co OOo

H,C,O H,,C,O O
0
"'v" rCH, 2 o c
OC,H, 0 OC
OH,,

H'Cka---o OC,H,,
OC,H, , XCH,,
H,,C,O. 2b
OH"CO
'a i' NC
H,C, o-" "O- ' o

3 ON C H, ' OC,H,,
0/ "
OCH,, HC _
0

H~C'-o 0CM *CO-' M-~

O - c
H'C-O H,,CCO 0

OCH, 0 OC, H ,,

Fig. 8.7. Chemical structure of the mono-1, bis-2a-c and tris acrylate 3,4 derivatives of
hexaalkoxytriphenylenes. (Reproduced from Ref. [56] with permission. Copyright (1999)
American Chemical Society.)

tion using InSn oxide and Al as the contacts. The current passed
through the devices was limited by the injection of holes into the semi-
conducting polymer layer by tunnelling. Cross-linked layers withstood
approximately 20 times higher currents than non-cross-linked layers
before dielectric breakdown occurred.
In a similar study, new hexaalkoxytriphenylenes having one, two, or
even three lateral attached acrylate moieties as polymerizable groups
were synthesized and characterized for use as novel insoluble hole
transport materials in organic LEDs [56]. Figure 8.7 shows the chemi-
cal structure of the mono-1, bis-2a-c and tris acrylate 3,4 derivatives of
hexaalkoxytriphenylenes. The conditions for the photopolymerization
of these monomers in thin film were evaluated and tested. The
bisacrylates and trisacrylateswere used to build insoluble networks.
When a mask was used during the irradiation, patterned films were
prepared. The polymeric reaction was controlled by GPC and FTIR
spectroscopy. Figure 8.8 shows the spectroscopic data that verify the

215
 0
co

o
al

1C
wave numbers [cm']

Fig. 8.8. Spectroscopic verification of the photo-cross-linking reaction in thin films by FT-
IR (16 scans; 4 cm - ' resolution, films on KBr): (a) trisacrylate 4 before polymerization;
(b)photopolymerized film of 4; (c) reference spectrum of a polymer derived from solution
polymerization of 1. (Reproduced from Ref. [56] with permission. Copyright (1999)
American Chemical Society.)

photo-cross-linking reaction in thin films. The networks and patterned

structure were also confirmed by UV spectroscopy, surface profiles, and
SEM photographs. Since hexaalkoxytriphenylenes are known as excel-
lent photoconductors, the photopolymerized films were used as hole
transport layers in two layered OLED with Alq3 . Finally, Figure 8.9
illustrates the OLED characteristic of a two-layer device with Alq3 as
emitting/electron transport layer and a cross-linked film as a hole-
transportlayer.
Polymer light-emitting diodes, based for example on MEH-PPV, are
known to be susceptible to oxidative degradation [571. This leads to loss
of conjugation, i.e. lower carrier mobility and higher operating voltage,
and to the formation of carbonyl species, i.e. to luminescence quench-
ing. In-situ FTIR revealed that ITO can act as the source of O. To
explore further the mechanism of oxidation and to provide guidance for
its elimination, the authors have studied the behaviour of MEH-PPV
LEDs prepared with a variety of conducting polymer anodes including
polyaniline and polythiophene derivatives cast from various solvents
and with various molecular and polymeric dopants. In all the cases
examined, polymer anodes led to significant improvement in lifetime
over devices with ITO as the anode contact. Also, in contrast to the
variability observed for ITO anodes, conducting polymers with

216
 voltage [V]
-15 -10 -5 0 5 10 15 20 25

10'

OTo
E

' 10'

C lo"

L ,
2 10

electric field [10 e V/cm]

Fig. 8.9. LED characteristic of a two-layer device with Alq3 as emitting/electron transport
layer and a cross-linked film of 3 as hole-transport layer ITO/3 (35 nm)/Alq 3 (35 nm)/Al
(200 nm). (Reproduced from Ref. [56] with permission. Copyright (1999) American
Chemical Society.)

polymeric dopants yield consistently good devices with power

efficiencies of approximately 0.5% at 5 V and brightness >1000 cd/m 2 .
Anodes prepared with small molecular dopants are more variable and
exhibit short-term behaviour which suggested interfacial electro-
chemistry. The authors described the device characteristics in the
context of a model of hole-dominated bipolar charge injection with
Langevin recombination.
In another study, poly(p-phenylenevinylene) (PPV) was derived
from a sulphonium salt precursor by ion beam irradiation [58]. A
quadrupole mass spectrometry analysis of the evolved species showed
rapid loss of HC1 and tetrahydrothiophene groups during irradiation
with 100 keV Ne +, indicating precursor degradation. Rutherford back-
scattering spectrometry confirmed the reduction of the sulphur and
chlorine content in the PPV film, whereas infrared spectroscopy
showed that the vibration mode at 2940 cm - l for the sulphonium group
has vanished for a 2 x 1016 ion effluence. The appearance of the trans-
vinylene peaks, at 3024 and 965 cm-l in PPV indicated the full con-
version of the precursor into the conjugated polymer for this effluence.
The correlation between a narrower optical band gap and the by one
order of magnitude higher conduction of a film implanted with Na + ions
with respect to a Ne + irradiation showed the doping effect induced by an
implantation with electronically active species.

217
8.3.2 Conducting polymers

In the area of the application of FT-IR techniques to study conducting

polymers, excellent reviews on the theory of polarons, bipolarons, and
solitons and use of vibrational spectroscopy in the studies of self-
localized excitations and charge transfer in conducting polymers exist
[59]. Geometry of polymer chains and changes induced by doping;
electronic states of polarons, bipolarons, and solitons; electronic
absorption spectra of poly(p-phenylene) and detection of radical anions
in p-oligophenyls were some of the issues addressed.
In addition, the first vibrational spectroscopic investigation of a
novel non-aqueous proton conducting polymer gel electrolyte consist-
ing of a PMMA matrix and a solvent mixture (ethylene carbonate (EC)/
propylene carbonate (PC) or EC/PC/N,N-dimethylformamide (DMF))
with a dissolved organic acid (benzoic or salicylic acid) took place [60].
The protonic conductivity of the gels was of the order 10-4-10 - 5 S/cm at
room temperature. It was found that the conductivity was proportional
to the degree of dissociation of the acid, the latter determined from
Raman spectroscopic data, and that the degree of dissociation
depended on the properties of the solvent mixture. Finally, the
relationship between the proton conductivity and the solvent-diffusion
dynamics was also studied.
Electronic absorption and vibrational spectroscopies of doped
conjugated polymers, whose ground states are nondegenerate were
also studied [61]. These studies have concluded that polarons are the
major species generated by doping in most nondegenerate conjugated
polymers such as polythiophene, poly(p-phenylene), and poly(p-phenyl-
enevinylene), in contrast with the previous view that bipolarons are the
major species.

8.3.3 Composites and nanocomposites

FT-IR spectroscopy has also been used in the study of composite

materials and nanocomposite films. In one such example, infrared
spectroscopy was used to monitor the formation of thin films of photo-
sensitive hybrid organic-inorganic glass on silicon via the solution
sol-gel method [62]. Glasses consisted of photoinitiator,methacryloxy-
propyltrimethoxysilane, methacrylic acid, and zirconium oxide. Clear,
low optical loss films were obtained, indicating nanophase homo-
geneity in the samples. The nanocomposite films were suitable for

218
fabricating optical components such as ridge waveguides and Bragg
diffraction gratings. The increase in the refractive index of the glass
relative to the surrounding material during photolithographic
processing was identified as a key material parameter in device
fabrication. Accordingly, electronic and vibrational spectroscopy were
used to provide insight into the structural changes that occur when
glasses were irradiated with continuous narrow band 4.9 eV and pulsed
6.4 eV light. Arguments were advanced, linking the changes in refract-
ive index to collateral densification leading to volume compaction of the
silicate network during organic free-radical polymerization. This was
shown by following the time evolution of relevant infrared absorption
bands. Free silanol and unreacted methoxysilane were consumed in the
process. Matrix densification was indicated by shifts to lower wave-
numbers in the transverse optical phonon mode associated with
decreasing Si-O-Si bond angles of the asymmetric stretching
vibration. Growth in the Si-O-Si framework was observed through
increased intensity in this infrared absorption. Similar behaviour was
observed for films irradiated with 6.4 eV light from an excimer laser. A
phase mask in combination with pulsed 6.4 eV light was used to
inscribe a 1.5 mm, high-reflectivity polarization-independent Bragg
grating into a ridge waveguide. The high reflectivity is thought to arise
from a periodic modulation of the volume compaction of the matrix.
Overall, the organic component of the glass confers unique properties
on the material that allows it to be densified even with 4.9 eV light. By
comparison, sol-gel silica with no organic component must be densified
at nearly twice the photon energy.
Di(carboxystyryl)benzene was self-assembled with a Zn complex in
THF on SiO2 and Si substrates to form thin films that exhibited blue-
green luminescence [63]. FT-IR spectroscopy of a 56 nm film on Si
showed characteristic absorption bands at 1600 cm-l, 1543 cm-l, and
1412 cm-l consistent with a powder sample. The refractive index (n)
was 1.66 at 633 nm. Multilayer growth proceeded by a 15 Angstrom
increase after initial surface coverage. These films were pursued for the
preparation of self-assembled films for electroluminescence
applications.
In a similar study, high-energy milling provided an effective and
environmentally conscious method for nanosizing Si [64]. Colloidal
suspensions of nano-sized Si were demonstrated and used for the
fabrication of high refractive index nanocomposites. Si nanoparticles
with average sizes of 20-40 nm and size distributions of approximately

219
25% were separated from milled powder via sonication and centri-
fugation. These nanoparticles were analyzed using TEM, dynamic light
scattering, x-ray diffraction and UV-visible/FTIR spectroscopy.
Formation of stable colloids was in part attributed to a thin surface-
oxide layer. The decrease in the average particle size caused a blue shift
in their absorption spectrum, thus increased the transparency in the
red part of the visible region. These Si nanoparticles were used to
fabricate high refractive index nanocomposites, with refractive indexes
<3.2, when dispersed in gelatin.
Langmuir-Blodgett (LB) films composed of the mixture of an
amphiphilic polymer containing azobenzene (Az) side chain (6Az10-
PVA) and 4'-pentyl-4-cyanobiphenyl (5CB) were prepared to mimic the
2-dimensional contacting region of the LC/Az interface of the command
surface which photochemically switches the LC alignment. UV-visible
absorption and FT-IR spectroscopic measurements were carried out
under illumination [651. These procedures allowed separate and simul-
taneous evaluations of the static state and dynamic molecular motions
of both Az and LC molecules which probably reflect the initial trigger-
ing step of the domino-mode response of the LC. The spectroscopic data
indicated the induction of reversible perpendicular/tilt orientational
changes of both the Az side chain and 5CB molecules upon alternative
irradiation of 365 and 436 nm light. Thus, 6Az10-PVA/5CB hybrid LB
film was regarded as a satisfactory interface model of a command
surface that promotes the homeotropic/planer alignment switching.
From the time courses of the photoisomerizationof Az and the orienta-
tional change, the molecular tilt was not governed only by the trans/cis
ratio of Az unit, but was strongly process-dependent (forward or back
process), indicative of involvement of strong molecular cooperativity.
Photoinduced vibrational bands of poly(3-octylthiophene) were
studied at room temperature by using time-resolved FTIR spectroscopy
(TR-FTIRS) in the nanosecond to microsecond time domain [66]. A
photo-bleach occurred in the deformation band of :C-S-C: at 1463-
1419 cm-l, and a few transient absorption bands occurred at lower
frequency (e.g., approx. 1290 cm-l). The transient absorption band at
1290 cm-l showed a signal exponential formation occurring in <200 ns,
and a double exponential decay process, with lifetimes of 53 and 788 ps.
Adding Fe2 03 nanoparticles into the polymer composite significantly
enhanced the photoinduced signal, indicating the interaction between
polymer and nanoparticle. The dynamics of these photoinduced species
on the nanosecond to microsecond time scale were discussed.

220
 In addition, vibrational spectroscopy was used for assessment of
new materials for the guided tissue regeneration (GTR) technique [67].
Implants applied in the healing of periodontal defects using this tech-
nique have to meet stringent requirements concerning their chemical
as well as physical properties. At present the implants prepared from
two layers membranes differing in porosity in their outer and inner
layers were studied clinically. Composite plates consisted of three
layers: a poly(lactic acid) film, carbon fibres coated with polylactic acid
and carbon fabric. Analysis of the infrared spectral data of samples
treated in Ringer solution allowed the description of the phenomena
resulting from the composite degradation. It was shown that material
biostability was related to the presence of carbon fibres.
Furthermore, the use of a new cathode material based on a compo-
site of an organosulphur compound, 2,5-dimercapto-1,3,4-thiadiazole
(DMcT), poly(aniline), and catalytic amounts of copper ion in secondary
lithium cells was also reported [68]. After failure of these cells, replace-
ment of the lithium anode restored the original capacity. Vibrational
spectroscopy of the copper/DMcT system indicated that addition of
Cu(II) oxidized and complexed DMcT and thus allowed more solvent to
be evaporated from the films. Furthermore, it was found that the redox
processes in the film were greatly stabilized by copper ion, likely due to
a mixture of catalytic and conductive effects.
Pyrrole can be polymerized within montmorillonite clays via chem.
means using Fe3+ and Cu 2+ as the oxidizing species [69]. The resultant
composite had properties of both the conducting polymer and the host
material. Vibrational spectroscopy, thermal analysis and conductivity
data all indicate that polypyrrole was present in the interlayer region
of the clays used. Electrochemically the conducting polymer-clay com-
posite showed promise for both sensor and electrolysis applications.
Cyclic voltammetry was studied for ascorbic acid oxidation at carbon
paste electrode and conducting polymer-clay composites/carbon paste
electrodes.
Finally, correlations were studied between dielectric, vibrational,
spectroscopic and heological parameter variations during cure of a
thermoset formulation of Araldite MY 0510 and 4,4'-methylene
dianiline [70]. Reaction kinetics values obtained from dielectric and
from spectroscopic results were in excellent agreement. Gelation and
vitrification times determined by dielectric and theological measure-
ments were also found to agree well, despite the empirical nature of
such correlations. A characteristic pattern was noticed in plots of

221
imaginary impedance as a function of reaction time, which can be used
to identify gelation and vitrification during network formation. The
realization of the full potential of dielectric impedance spectroscopy in
monitoring the progress of chemophysical changes in reactive poly-
mers, however, hinges upon a development of fundamental scientific
correlations between dielectric and chemorheological phenomena
during cure.

8.4 APPLICATIONS OF INFRARED MICROSPECTROSCOPY

Since the introduction of the first FT-IR microscope interface by Bruker

Instruments in 1983 the technique has progress to the point that very
beautiful chemical images are nowadays generated from a step-scan
FT-IR microscope equipped with a focal plane array detector. Many
reviews exist in the literature that can familiarize the reader with the
subject. In these reviews, applications are discussed in the area of
forensic science, materials science, art restoration and the biological
science [71,72].
The coupling of imaging modalities with spectroscopic techniques
adds additional dimensions to sample analysis in both the spectro-
scopic and spatial domains [73]. The particular ability of infrared
imaging to explore the spatial distribution of chemically distinct
species demonstrates the versatility and diversity of spectroscopic
imaging. The particular spectroscopic imaging instrument integrated
several infrared focal-plane arrays with a Michelson step-scan inter-
ferometer, generating high-fidelity and high spectral resolution mid-
infrared spectroscopic images. The instrumentation produced multi-
dimensional, chemically specific images, while simultaneously obtain-
ing high resolution spectra for each detector pixel. The spatial
resolution of the images approached the diffraction limit for mid-
infrared wavelengths, while the spectral resolution was 4 cm-l. Data
derived from a variety of materials, particularly biological samples,
illustrated the capabilities of the technique for readily visualizing
chemical complexity and for providing statistical data on sample
heterogeneity.
Another approach utilized the use of the infrared scattering from
the tip of an atomic force microscope. This technique overcomes the
problem that the spatial resolution is diffraction-limited to a scale of
about half the wavelength, or about five micrometers in the infrared

222
spectral region [74]. The scanning near-field optical microscope, how-
ever, can reveal sub-wavelength detail because it uses near-field
probing rather than beam focusing. Therefore, the use of the aperture-
less approach to scanning near-field optical microscopy was demon-
strated in an attempt to obtain contrast in vibrational absorption on a
scale of about 100 nm, about one-hundredth of a wavelength. Infrared
scattered light was recorded from the tip of an atomic force microscope
scanned over a composite polymer film. At the boundary between
different polymers contrast changes were observed owing to changes in
vibrational absorption. The contrast was strongly enhanced in the near
field of the probe tip, which we interpret as evidence of surface-
enhanced infrared absorption. When extended to multi-wavelength
operation, this approach should enable imaging of chemical compounds
at nanometre resolution.
Another different approach toward mid-infrared spectroscopic
imaging microscopy was introduced in which instrumentation was
designed about an InSb multichannel, focal-plane array detector and a
variable-bandpass dielectric filter [75]. The system could be configured
for either macroscopic or microscopic applications, and high-fidelity,
chemically specific images were acquired in real time. With the
dielectric filter used in this assembly, continuous tuning was provided
for the 4000-2320 cm-l spectral region with spectral resolution of
approximately 35-18 cm -l at the extremes of this wavelength interval.
The functioning of the imaging microscope was demonstrated with
samples includingpolystyrene microspheres,preparations of lipids and
an amino acid embedded in KBr disks, and a tissue sample derived
from a coronal slice of a monkey cerebellum.
Another recent study concentrated in reducing the noise of a chemi-
cal imaging experiment. Temporal resolution of fast FTI-R imaging is
limited by rapid degradation of data quality, due to increased noise,
with faster image acquisition [76]. Various coaddition schemes were
presented, meant to reduce noise and improve the quality of images
acquired from such systems. The application of the proposed schemes
allowed for improved signal-to-noise ratio (SNR) characteristics in the
resulting data. Figure 8.10 shows the single-beam spectrum from a
pixel with KBr disks in the beam path and an absorbance spectrum of
the polymer film. On the other hand, Figure 8.11 shows the collection
and coaddition scheme to obtain absorbance images with a higher SNR
using fastest single-beam image collection and the coaddition schemes
to coadd pixels with the same true absorbance values.

223
Fig. 8.10. The single-beam spectrum from a pixel with KBr disks in the beam path and an
absorbance spectrum of the polymer film. The profile with lower noise is obtained by
coadding 256 pixels illustrating noise. (Reproduced from Ref. [76] with permission.
Copyright (1999) Society for Applied Spectroscopy.)

These schemes were tested by monitoring the dissolution of a poly-

mer film [poly(a-methylstyrene)] by a low-molecular weight solvent
[methyl iso-Butyl ketone (MIBK)]. Figure 8.12 shows an image of a
polymer/air interface and a comparison of spectra obtained from a
pixel. Pseudo coaddition improved the SNR by approximately 45%. A
total acquisition time of about 100 s was achieved, allowing the dis-
solution process to be monitored by using image acquisitions separated
by 3 min. Low noise concentration profiles, linear solvent penetration
rate, and polymer dissolution rates were all measured. Figure 8.13
shows the dissolution of a poly(a-methyl styrene) film by MIBK diffus-
ing from right to left. The images at the top plot E(x,y, 1600) for
absorbance and the images at the bottom plot E(x,y, 1730) for the
solvent. Detection limits of approximately 5% and quantification limits
of approximately 20% were achieved by using optimal coaddition
strategies. This result represented an order of magnitude improvement
over untreated data.

224
 A

I FastFTIR
Absorbance Image
n co-added Data Sets

N Pseudo-Data Sets (N+I) Collected Data Sets

//
(Higher Order)

Linear Sampling
(Higher Order)
Fig. 8.11. (A) Collection and coaddition scheme to obtain absorbance images with a
higher SNR using fastest single-beam image collection. (B) Coaddition schemes to coadd
pixels with the same true absorbance values. (Reproduced from Ref. [76] with
permission. Copyright (1999) Society for Applied Spectroscopy.)

Correlative x-ray photoelectron spectroscopy (XPS) and Fourier

transform infrared (FT-IR) studies of the complex heterogeneous
structure of 50:50 poly(vinyl chloride)lpoly(methyl methacrylate) (PVC/
PMMA) polymer blends were also shown [77]. The comparable lateral

225
 Y

A F

Fig. 8.12. (A) An image of a polymer/air interface with ROI used for statistical analysis.
(B) A comparison of spectra obtained from a pixel on the FPA using described parameters
compared to a spectrum of the film obtained by a rapid-scan FT-IR spectrometer.
(Reproduced from Ref. [76] with permission. Copyright (1999) Society for Applied
Spectroscopy.)

226
 j'2
1I 41

rr
. tell
V
=

j, 5's I-
, 1:'M
* S '. 1
4
' --
,_
1', 91,"
r, I 'i ti
',u ,'
I
.,,':,

its
X
,': Ari
: A'*
I =am
nr
uto lyme 20
- Ij - U m11 t-o7 E Z-I , 1111

Polymer FJ(xy, 1600) 200 Ful Solvent E(x,y,1 730)

Uo _nMWr A 0.25 2- teo L(

Fig. 8.13. The dissolution of a poly(a-methyl styrene) film by MIBK diffusing from right to
left. The images at the top plot E(x,y, 1600) for absorbance and the images at the bottom
plot E(x,y, 1730) for the solvent. (Reproduced from Ref. [76] with permission. Copyright
(1999) Society for Applied Spectroscopy.)

resolution and parallel imaging capabilities of both techniques allowed

for a direct comparison of surface (XPS) and bulk (FT-IR) measure-
ments of polymer blends. To eliminate substrate influence and film-to-
film differences, the same areas on the polymer films were analyzed by
both methods. The effect of PMMA molecular weight on surface
separation and surface segregation was evaluated by using six blends
with a constant PVC molecular weight and a PMMA molecular weight
varying from 75 to 2132 kDalton. Imaging capabilities of both methods
were used for a qualitative comparison of the heterogeneous structure
of the blends, while a quantitative comparison of the bulk and surface
compositions of the same areas of the samples used small-area
spectroscopy from XPS and FT-IR. On the basis of the quantitative
analysis, it was concluded that surface segregation of PMMA increases
with increasing molecular weight. The determination of both surface
and bulk properties of complex heterogeneous samples is important for
a more complete understanding of the structure of complex films.
The application of infrared microscopic imaging to the study of bone
disease and fracture healing was also demonstrated. Samples of
normal and osteoporotic human iliac crest biopsies were prepared and
examined. Figure 8.14 shows an optical micrograph of the major
mineral-containing areas of bone. Two spectral parameters, one that

227
 - 1600 mm -

Trabecular Bone - Osteon

Marrow
- Cortical Bone
Nuclei

4-*00400 p-m

"Mineral Rings" -
Single
Haversian canal" Osteon

Fig. 8.14. Optical micrograph of the major mineral-containing areas of bone. The top
figure shows the spatial relationship between the cortical region, which is part of the
cylindrical structure forming the outer shell of compact bone, and the inner region
containing trabecular bone and marrow. The bottom figure shows a single osteon in
which mineral grows in 'tree-ring'-like fashion around the central blood vessel
(Haversian canal). (Reproduced from Ref. [78] with permission. Copyright (2000) Society
for Applied Spectroscopy.)

monitors the extent of mineral (hydroxyapatite) formation in the tissue

and another that monitors the size/perfection of the crystals, were
compared in the samples generated from normal and pathological
tissues. Figure 8.15 shows a series of spectra acquired from a 300 mm
wide region of normal human trabecular bone. The average mineral
levels in the osteoporotic sample were reduced by 40% from the normal.
In addition, the crystal size/perfection was substantially enhanced in
the disease state. The applicability of infrared imaging techniques to
the study of therapeutic intervention was also investigated in a study of
the effects of estrogen therapy on fracture healing in rat femurs.
Femurs were examined by IR microscopic imaging four weeks after
fracture. IR imaging showed that the mineral level was enhanced in
estrogen-treated samples. In addition, the crystals were larger/more
perfect in the treated specimens. These data demonstrate the utility of
IR spectroscopic imaging for the study of pathological states of hard

228
 U
I
I0
U

WA 40 2
r,
0
>

I
To
To

t
Wavenumber (cm-') I

Fig. 8.15. Series of spectra acquired from a 300 mm wide region of normal human
trabecular bone. (Reproduced from Ref. [78] with permission. Copyright (2000) Society
for Applied Spectroscopy.)

tissue. Finally, Figure 8.16 shows the infrared images of the index of
mineral crystallinity/perfectionand a histogram of this quantity for an
estrogen treated site in a fractured rat femur and for an untreated site.
Fourier-transform IR microspectroscopy was used to study bone
mineralization processes in an in vivo model and in enamel in osteo-
genesis imperfecta [78]. The ability of this technique to map new bone
formed in implanted macroporous calcium phosphate biomaterial from
sections was reported for the first time. This technique allowed the
correlation of the microstructure of bone formation in the in vivo model
with modifications in carbonate and phosphate environments of the
mineral phases during maturation. Analysis on enamel sections
revealed changes in the mineral environment of carbonate and
phosphate ions and probably in the size of the enamel crystals. These
modifications contributed to the fragility of enamel in osteogenesis
imperfecta. The infrared functional group imaging of a part of the
implanted biomaterial and the bone ingrowth provided the visualiza-
tion of chemical modifications occurring in biomaterial implants at 20
pm spatial resolution. The use of this technique, in conjunction with
appropriate sampling methods and data analysis should provide furth-
er insight into the molecular structure of mineral phases of calcified

229
 --
119-

I
m:-
11
CD

.4
To r. : Z
= I
z; 00

'Y'"
, I 11 .2
111
400 pm 1(030)/1( 1020)

,~K
ii-

10

$s .40
E 0 10

40

C) 12
I 12
I -
1(030)/1( 1020)
400 pm

Fig. 8.16. Infrared images of the index of mineral crystallinity/perfection and a histogram
of this quantity for an estrogen treated site in a fractured rat femur (bottom) and for an
untreated site (top). (Reproduced from Ref. [78] with permission. Copyright (2000)
Society for Applied Spectroscopy.)

tissues and help to elucidate mineralization processes, skeletal dis-

orders and properties of the biomaterials used as bone substitute.
The distribution of chemical species and the degree of orientation in
semicrystalline polymer systems have also been studied using fast
Fourier transform imaging [79]. A variety of poly(ethylene glycol)
systems, including pure polymer, high- and low-molecular weight
blends, and blends with amorphous polymers, were studied. It is shown
that fast FTIR imaging can be used to determine the distribution of
species with different molecular weights and can be used to determine
the degree of segregation of different components in blends with
amorphous polymers. Additionally, by employing an infrared polarizer,

230
the degree of orientation was determined in these systems by the
generation of spatially-resolved dichroic ratio images.
Imaging spectrometry enables passive, stand-off detection and
analysis of the chemical compounds of gas plumes and surfaces over
wide geographic areas [80]. The authors described the use of a long-
wavelength infrared imaging spectroradiometer, comprised of a low-
order tunable Fabry-Perot etalon coupled to a HgCdTe detector array,
to perform multispectral detection of chemical vapour plumes. The
tunable Fabry-Perot etalon used in this research provides coverage of
the 9 . 5 -1 4 -pm spectral region with a resolution of 7-9 cm-l. The etalon-
based imaging system provided the opportunity to image a scene at
only those wavelengths needed for chemical species identification and
quantification and thereby minimized the data volume necessary for
selective species detection. The authors present initial results using a
brassboard imaging system for stand-off detection and quantification
of chemical vapour plumes against near-ambient temperature back-
grounds. Model calculations were presented comparing the measured
sensitivity of the sensor to the anticipated signal levels for two
chemical release scenarios.
In another study, a 64x64 Mercury-Cadmium-Telluride (MCT)
focal-plane array detector attached to an FT-IR microscope was used to
spectroscopically image 8-pm-thick cross-sections of wheat kernels in
the fingerprint region of the IR spectrum [81]. After fast-Fourier trans-
formation of the raw image interferograms, the data can be displayed
either as a series of spectroscopic images collected at individual wave-
lengths, or as a collection of infrared spectra obtained at each pixel
position in the image. Image contrast is achieved due to the intrinsic
chemical nature of the sample at each pixel location in the image.
Individual cell layers near the outer portion of the wheat kernel, as well
as the primary root within the germ, can be clearly differentiated in the
IR images as a result of this enhanced chemical contrast.
Micro-imaging spectrometers incorporating focal plane array (FPA)
detection require careful demarcation of cold shield aperture size for
both optimal performance and prevention of errors. One study explored
the effects of changing the diameter of the cold shield aperture on the
intensity and spatial homogeneity of the incident radiation [82]. A
uniform polystyrene film was repeatedly imaged by using cold shields
of varying aperture sizes. It was shown that a smaller than optimal
aperture size led to image edge clipping, resulting in an inefficient use
of the array, lower overall signal, spectral distortions, and higher noise

231
characteristics. Use of an aperture size larger than required caused a
decrease in the effective dynamic range of measurements, resulting in
higher noise levels. The advantages and necessity of optimizing
imaging spectrometer performance by employing a cold shield with an
appropriately sized aperture were discussed.
The penetration of chemical reagents through human hair after
bleaching has been spatially characterized using IR microspectroscopy
with a synchrotron source [83]. Chemical imaging of hair cross-sections
before and after bleaching was achieved with high contrast, using the
peptide and lipid mid-infrared absorption bands which are charac-
teristic of hair. The ability to make images using functional groups as a
contrast mechanism can be applied to studies of other chemical groups,
if present, in the structure of the hair. In this study it was shown how
the penetration of an organically active reagent in the hair structure
could be quantified with a spatial resolution of few microns. These
results demonstrated that synchrotron infrared microscopy is a power-
ful tool for characterizing chemical interactions of hair samples with
specific cosmetic materials.
Infrared spectra of breast tumour cell lines and breast tumour
tissues have been measured. Infrared measurements of tumour cells
revealed that approximately fifteen cells are necessary to obtain
spectra of good signal-to-noise ratio using an IR microspectrometer
equipped with a conventional IR thermal source [84]. Comparative
studies of human breast tumour cell line suspensions demonstrated
that MCF-7 cells and drug-resistant NCI/ADR cells could be differenti-
ated based on their infrared signatures. The most striking differences
between MCF-7 and NCI/ADR were found in features assigned to CH2
and CH3 stretching vibrations of lipid acyl chains and PO,- stretching
vibrations of nucleic acids. To assess the potential of IR spectroscopy
for the diagnosis of breast tumour tissues, thin sections of tissue were
mapped by FTIR microspectroscopy. The spectra of these maps were
analyzed using functional group mapping techniques and cluster
analysis and the output values of the different approaches were then
reassembled into IR images of the tissues. A comparison of the infrared
images with the standard light microscopic images of the correspond-
ing areas suggested that: (i) chemical mapping based on single band
intensities was an easy way to detect microscopic fat droplets within
tissue; (ii) the comparison of IR images based on band intensities at
1054 and 1339 cm-l provided information on tissue areas containing
tumour cells; and (iii) cluster analysis of the spectra was superior to the

232
single band approach and more appropriate for differentiation between
tissue types.
In an different approach, using synchrotron radiation as an ultra-
bright infrared source, the authors were able to map the distributions
of functional groups such as proteins, lipids, and nucleic acids inside a
single living cell with a spatial resolution of a few microns [85]. In
particular, the changes in the lipid and protein distributions in both the
final stages of cell division and also during necrosis were mapped.
FT-IR microspectroscopic maps of unstained thin sections from
human melanoma and colon carcinoma tissues were obtained on a
conventional IR microscope equipped with an automatic x, y stage [86].
Mapped infrared data were analyzed by different image re-assembling
techniques, namely functional group mapping ('chemical mapping')
and, for the first time by cluster analysis, principal component analysis
and artificial neural networks. The output values of the different
classifiers were recombined with the original spatial information to
construct images whose colour or grey tones were based on the spatial
distribution of individual spectral patterns. While the functional group
mapping technique could not reliably differentiate between the differ-
ent tissue regions, the approach based on pattern recognition yielded
images with a high contrast that confirmed standard histopathological
techniques. The new technique turned out to be particularly helpful to
improve discrimination between different types of tissue structures in
general, and to increase image contrast between normal and cancerous
regions of a given tissue sample.
Infrared absorption spectroscopy was used with other scattering
and imaging techniques to elucidate the interface reactions leading to
permanent chemical bonding ofjoined hydrophilic wafers upon anneal-
ing, and to uncover the thermal evolution of H-decorated defects in H-
implanted Si wafers [87]. Detailed mechanisms were proposed whereby
the role of micro-voids as gathering sites for H2 is highlighted and the
kinetic interplay between defect formation/evolution, H passivation of
internal structures and molecular H2 formation was critical for
exfoliation to occur.
The demand also of smaller device dimensions drives the need to
improve the lithographic and the metrological tools to produce them
[88]. Characterization of the image formation during the lithographic
process is key to any process control effort. Scanning probe microscopy
(SPM) on exposed, unbaked and baked, undeveloped photoresist show-
ed morphological details of the image formation process unachievable

233
with other techniques. The use of micro-FT-IR spectroscopy was in-
vestigated for latent image chemical analysis. Both of these techniques
were used in the study of the dependence of the latent image of a
negative novolac-based chemically amplified resist, SAL 605 by
Shipley, with post-exposure bake (PEB) conditions. The objective of the
experiment was to understand how the thermal properties of the resist
and the linking reaction taking place were related to each other during
PEB. Experimental results indicated that resist from unexposed
regions diffused into the exposed resist during PEB. SPM results show
that this diffusion increased as the PEB temperature rose above the
oxide glass transition temperature of the unexposed resist. These
results showed that the linker component of the resist, hexamethoxy-
methylmelamine, was identified as one of the resist components that
diffused into the exposed regions during PEB.
Traditional methods of cell wall analysis have provided valuable
information on wall composition and architecture, but, by having to
rely on the use of bulk samples, have averaged out this intrinsic
heterogeneity. FTIR microspectroscopy addresses this problem by
providing chemical information from an area as small as 10x 10 pm of a
single cell wall fragment or area of a tissue section that has been
imaged with a microscope accessory. The authors have used FTIR
microspectroscopy as a powerful and extremely rapid assay for wall
components and putative cross-links. The spectra were sensitive to
polymer conformation, and the use of polarizers in the microscope
accessory allowed the orientation of particular functional groups to be
determined, with respect to the long axis of elongating cells. The
spectra constituted species and tissue-specific 'finger-prints', and the
use of classical discriminant analysis may provide the opportunity for
correlating spectral features with chemical, architectural or rheologi-
cal wall properties. Spectral mapping of an area of a specimen allowed
the morphological features resulting from cell growth and differenti-
ation to be characterized chemically at the single cell level. In addition,
the fidelity of the spectral images was determined by the pixel number
of the focal-plane array [89].
In another study, an instrument was described that simultaneously
recorded images and spectra of materials in the infrared fingerprint
region using a long-wavelength infrared focal-plane array detector, a
step-scan Michelson interferometer, and an IR microscope [90]. With
the combination of step-scan Fourier transform Michelson interfero-
metry and arsenic-doped silicon Si:As focal-plane array image

234
detection, an infrared spectroscopic imaging system was constructed
that maintained both an instrumental multiplex and multichannel
advantage and operates from approximately 4000 to 400 cm-l. With
this method of mid-infrared spectroscopic imaging, the fidelity of the
generated spectral images recorded through the microscope was solely
determined by the number of pixels on the focal-plane array detector,
and only a few seconds of data acquisition time were required for
spectral image acquisition. This seamless combination of spectroscopy
for molecular analysis and the power of visualization represented the
future of infrared microscopy.

8.5 APPLICATIONS TO INDUSTRIAL PROCESS

Recently, a new method for simultaneous determination of vulcanized

rubber additives by FT-IR using partial least-squares regression for
multivariate calibration was developed [91]. The effect of various wave-
number ranges and the use of the absorbance and first-derivative
spectral modes on performance were studied by applying the method to
three different sample batches containing several additives in different
proportions, all of which were resolved with satisfactory results.
In another study, a simpler spectrometer design was employed for
industrial process. A Michelson type interferometer was used where
the moving mirror was suspended by two fluxes and driven by a coil
actuator [92]. Displacement of the mirror was monitored using a much
smaller transducer with a better thermal stability than the convention-
ally used HeNe laser. The beamsplitter is a CaF 2/Si and a thermo-
electrically cooled PbSe is used as the detector. The spectral range was
from 5000 to 1800 cm-1 with resolution better than 8 cm -l .
Furthermore, the applications of FT-IR spectroscopy to industrial
processes was greatly benefited by the use of mid-infrared optical fibres
[93]. The ability to make measurements at a remote site or as a reaction
occurs offers a significant advance in these types of analyses. Mid-
infrared fibres are used to transmit radiation outside of the spectro-
meter, to the sample, and then to the detector. A section of the fibre,
with the protective cladding removed, is used as the sampling device.
In this decladded region the fibre, acting as an internal reflectance
element, contacts the sample and provides the chemical information for
analysis. In one example, the use of a fibre to monitor the progress of a
curing reaction in thermoset composite materials where the fibre was
imbedded in the matrix was studied.

235
 Chalcogenideglass fibres based on sulphide, selenide, telluride and
their rare earth doped compounds are being actively investigated
worldwide [94]. Great strides have been made in reducing optical losses
using improved chemical purification techniques, but further improve-
ments are needed in both purification and fiberization technology to
attain the theoretical optical losses. Despite these problems, current
single-mode and multimode chalcogenide glass fibres are enabling
numerous applications. Some of these applications include laser power
delivery, chemical sensing, imaging, scanning near field microscopy/
spectroscopy, fibre infrared sources/lasers, amplifiers and optical
switches.

8.6 APPLICATIONS TO ORGANIC THIN FILMS

In recent years, functional organic thin films have received keen

interest in the field of molecular electronics because of an increasing
awareness that functional organic thin films exhibit a variety of
interesting functions [95-97]. A number of molecular devices have been
proposed that are based on organic thin films such as Langmuir-
Blodgett (LB) films with nonlinear optical properties, photovoltaic
cells, piezoelectric and pyroelectric devices, resistance and conducting
materials, and chemical and biological sensors. To sufficiently under-
stand the functions of functional organic thin films, it is necessary to
investigate the arrangement and orientation of molecules in a film, and
further, structures (e.g., conformations, chemical bonds, intermolecu-
lar interactions, electronic states) and the like. In addition, knowledge
about the relationship between functions and structures is essential to
designing of a new organic thin film. While methods of exploring the
structure of an organic thin film include x-ray diffraction, atomic force
microscopy, ultraviolet and visible spectroscopy, fluorescence spectro-
scopy, ESR, infrared spectroscopy, Raman spectroscopy, etc., the
infrared spectroscopy we describe in this section can be said to be
prominent in terms of the diversity and quantity of information and the
simplicity of measurement [95-99].

8.6.1 Infrared spectra of Langmuir-Blodgett films

We will mainly describe here infrared studies on LB films which are

attracting the greatest attention among a variety of organic thin films.
In the following, LB films such as those shown in Fig. 8.17a and b will

236
 (a) Y-type

(b) Y'-type
Fig. 8.17. Structure of (a) Y-film and (b) Y'-film.

237
 C n H 2n+1

n=12; Dodecyl-TCNQ
n=15; Pentadecyl-TCNQ
n=18; Octadecyl-TCNQ

Fig. 8.18. Structure of 2-alkyl-7,7,8,8-tetracyanoquinodimethane.

be referred to as the Y-film and the Y'-film, respectively. When used for
structural investigations on LB films, infrared spectroscopy is
advantageous in the following points.
1. It is possible to measure a spectrum non-destructively at room
temperature under a normal pressure.
2. Operations for measurement of spectra are relatively easy.
3. It is possible to measure a spectrum of even a one-layer LB film.
4. Since an infrared spectrum can be measured for an LB film, a
solution, a solid and a crystal, one can compare a structure of a
sample in the LB film with structures of the sample in other states.
5. Various types of infrared measurement methods (transmission
method, the ATR method (Section 7.2.1), the RA method (Section
7.2.4), a surface-enhanced method, etc.) may be applied.
As a vibrational spectrum sensitively reflects the arrangement of
atomic nuclei and nature of chemical bonds within a molecule, or an
interaction between the molecule and a surrounding environment it is
suitable very much to study the molecular aggregation, orientation,
and structure in an LB film. Knowledge obtained from infrared spectra
of LB films are summarized as follows.
1. Orientation of molecules; whether hydrocarbon chains and chromo-
phores are perpendicular to a substrate or tilted with respect to the
substrate normal, etc. It is also possible to quantitatively estimate a
tilt angle [100-102].
2. Sub-cell packing of hydrocarbon chains [103,104].
3. Conformations of hydrocarbon chains; whether hydrocarbon chains
have trans-zigzag structure or partially contain gauche forms, etc.
[105,106].

238
4. Structures of chromophores; the conformation, chemical bonds, and
electronic states of chromophores and interactions between
molecules, etc.
5. Interactions between the substrate and the first layer.
Infrared spectroscopy is useful not only for structural characterization
of an LB film, but also usable to assess whether the LB film has a high
quality based on the knowledge (1) to (3).

8.6.2 What we can learn from infrared spectra of LB films

We will describe in more detail what we can learn from infrared spectra
of LB films, citing an example of an LB film of 2-octadecyl-7,7,8,8-
tetracyansquinodimethane (octadecyl-TCNQ) (see Fig. 8.18). Figure
8.19 shows infrared spectra of octadecyl-TCNQ in a powder, in a bromo-
form solution, and in a ten-layer LB film (the Y-film) [107]. In general,
when we measure an infrared spectrum of dye molecule with a long
hydrocarbon chain such as octadecyl-TCNQ, we typically observe infra-
red bands due to the hydrocarbon chain and bands due to the chromo-
phore. As the former, we can expect bands due to CH3 degenerate
stretching, CH3 symmetric stretching, CH 2 antisymmetric stretching,
CH 2 symmetric stretching, CH 2 scissoring, and CH2 rocking vibrations.
In the bottom spectrum of Fig. 8.19, bands at 2955, 2918, 2847, 1462 and
1417 cm-l are assigned to CH3 degenerate stretching, CH2 antisym-
metric stretching, CH 2 symmetric stretching and CH 2 scissoring
vibrations (CH 2 scissoring vibrations appear as a doublet). Although a
band due to CH3 symmetric stretching vibrations should appear in the
vicinity of 2875 cm-l, this band is weak and therefore cannot be
recognized in the spectrum [107]. Furthermore, a band arising from the
CH 2 rocking vibration is generally expected to appear in the vicinity of
725 cml. Infrared bands due to the chromophore portion can be classi-
fied into bands assigned to in-plane and out-of-plane vibrations. Bands
at 2223, 1546, and 1530 cm-' in the spectrum of the LB film are all
bands due to in-plane vibrations of TCNQ portion, and assigned to C-N
stretching, C=C stretching and C=C stretching vibrations, respectively
[107]. In general, for analysis of an infrared spectrum of an LB film, we
usually identify bands due to a hydrocarbon chain first, and thereafter
look for bands arising from a chromophore. However, it is sometimes not
easy to distinguish a band due to CH2 scissoring vibrations from a band
due to the chromophore. In such a case, we may be able to distinguish

239
 1 | h x

W
ILI
z

a:
co
m
0
C,)
4

t1

4000 3600 3200 2B00 2400 2000 1600 1200

WAVENUMBER(cm')

Fig. 8.19. Infrared transmission spectra of octadecyl-TCNQ in a powdered micro-

crystalline state (top), octadecyl-TCNQ in a bromoform solution (middle), and 10-layer
LB film of octadecyl-TCNQ deposited on both sides of a CaF 2 substrate (bottom).
(Reproduced from Ref. [1071 with permission. Copyright (1991) American Chemical
Society.)

them by measuring a spectrum of chromophore only which does not

have a hydrocarbon chain.
Now, what kind of information does a spectrum such as that shown
in the bottom of Fig. 8.19 provide? First, we can obtain information
regarding the conformation of a hydrocarbon chain from the
frequencies of CH 2 antisymmetric and symmetric stretching bands
[105,106]. These bands are known to appear in the vicinity of 2918 cm -1
and 2848 cm - l, respectively, when the hydrocarbon chain assumes
trans-zigzag structure but shift to the higher-wavenumber side if the
hydrocarbon chain contains some gauche forms. Hence, the result

240
shown in Fig. 8.19 tells us that the hydrocarbon chain of octadecyl
TCNQ has trans-zigzag conformations while in the LB film and solid
powder but contains a considerable number of gauche forms while in a
solution [107].
We can learn about subcell packing of hydrocarbon chains from
bands due to CH 2 scissoring mode [103,104]. The CH 2 scissoring
vibrations appear as a doublet at 1471 and 1462 cm-l when the
hydrocarbon chains take orthorhombic subcell packing, but as a single
band at 1467 cm -l when the chains assume hexagonal subcell packing.
Since bands due to a CH 2 scissoring vibration appear as a doublet in
the top and bottom spectra of Fig. 8.19, it is considered that the
hydrocarbon chains of octadecyl-TCNQ assume orthorhombic subcell
packing both in the solid powder and the LB film (in the spectrum of the
solution in Fig. 8.19, the CH 2 scissoring vibration appears as a singlet
band as it is naturally expected). However, special care must be taken
for the LB films of octadecyl TCNQ where the hydrocarbon chains
assume interdigitated and non-interdigitated parts. Morita et al. [108]
assigned the two bands at 1471 and 1462 cm-l of the LB films of
octadecyl-TCNQ to the CH2 scissoring modes of non-interdigitated and
interdigitated parts of the hydrocarbon chain.
If one wishes to study the molecular orientation in an LB film, one
must compare an infrared transmission spectrum with an infrared RA
spectrum (Chapter 7.2.4). Let us introduce a simple example of
comparison between a transmission spectrum and an RA spectrum.
Figure 8.20 shows a transmission spectrum and an RA spectrum of
seven-layer LB films of cadmium stearate [102]. We can readily notice
the remarkable differences in the intensities of infrared bands between
the two spectra. It is these differences in the intensities that allow us to
discuss the molecular orientation in an LB film.
Now, let us consider which bands will appear strongly in the
transmission spectrum on an assumption that the molecular axis of
cadmium stearate is nearly perpendicular to a substrate (see Fig. 8.21).
In the case of a transmission method, since an electric vector of an
infrared ray is parallel to the substrate, strong bands are those due to
vibrations whose transition moments are perpendicular to the molecu-
lar axis, such as CH 2 antisymmetric and symmetric stretching vibra-
tions (2919 and 2851 cm-l in Fig. 8.20, bottom), COO- antisymmetric
stretching vibration (1543 cm l) and CH 2 scissoring vibrations (1473
and 1463 cm-l), whereas bands whose transition moments are parallel
to the molecular axis, such as COO symmetric stretching vibration

241
 v, COO-

0.02 vsCH2 vCH2 E

P
V c2 ll I I C00-l IcoCH2
r

I 3-fi - 1 -8
R00
- 0 0 - _1800
_ __ _1200
_
I
Co
Transmission
vaCH2
0
co
Co
CHCOO
<g
77A77727 A\CH2
i I I , . i k F q q r
3000 2800 1800 1600 1400 1200
WAVENUMBER(cm- )

Fig. 8.20 Infrared RA and transmission spectra of seven-layer LB films of cadmium

stearate on silver and ZnSe plates, respectively. (Reproduced from Ref. [102] with
permission. Copyright (1990) American Chemical Society.)

(1433 cm -1 ) and CH2 wagging vibrations (which appear as a series of

bands in the 1350-1200 cm-1 region), should be rarely observed. The
transmission spectrum in Fig. 8.20 strongly supports the hypothesis
described above. According to the surface selection rule in infrared RA
spectroscopy [100,101], molecular vibrations whose transition
moments are perpendicular to a substrate surface appear strongly in
an RA spectrum. As we have assumed above, if the molecular axis of the
hydrocarbon chain is perpendicular to the substrate, COO- symmetric
stretching and CH2 wagging vibrations (when a hydrocarbon chain has
trans-zigzag conformation, as all CH 2 groups vibrate in the same plane
and couple with each other, a series of bands (band progression)
appears which expresses phase differences of the vibrations) should
appear as strong bands in the RA spectrum. Figure 8.20 exactly shows
this in reality, and accordingly proves that the hypothesis above is
correct.
One may be able to quantitatively discuss the molecular orientation
based on comparison of the intensity of each band between the trans-
mission spectrum and the RA spectrum. From the comparison of the
two spectra shown in Fig. 8.20, Umemura et al. [102] calculated the
molecular orientation in the LB film of cadmium stearate as shown in
Fig. 8.21.

242
 I

-18
- 7o
- 83'

IIIIIIIIIIIIITT11TTT777TTTTTT7
,
1111 .. 1 ............
Substrate

Fig. 8.21 Calculated tilt angles in an LB films of cadmium stearate. (Reproduced from
Ref. [102] with permission. Copyright (1990) American Chemical Society.)

Figures 8.22a and b compare infrared transmission and RA spectra

of three-layer LB films of octadecyl-TCNQ [107]. Of particular note in
Fig. 8.22 is that the intensities of the bands due to in-plane TCNQ
modes are much stronger in the RA spectrum than in the transmission
spectrum (it should be noted that three-layers of octadecyl-TCNQ are
deposited on both sides of a CaF2 substrate). Therefore, it seems that
the TCNQ plane is nearly perpendicular to the substrate surface.
Transition moments of the two C=C stretching bands at 1547 and
1531 cm-l are parallel with the molecular axis of the TCNQ chromo-
phore and perpendicular to it, respectively. The relative intensity of the
two bands is reversed between the transmission and RA spectra, but it
should be noted that both have comparable intensities in the two
spectra. Therefore, it may be concluded that the molecular axis of the
TCNQ chromophore is neither parallel nor perpendicular to the
surface, being in an intermediate direction [107]. The intensities of CH2
antisymmetric and symmetric stretching bands are comparable
between the transmission and RA spectra, suggesting that the alkyl
chain is tilted considerably with respect to the surface normal.

8.6.3 Three examples of infrared studies of LB films

In the following part of this section three examples of infrared studies

of LB films are discussed. For LB films of dodecyl-, pentadecyl-, and

243
 LU
z

0
M
C,)
M

4000 3600 3200 2800 2400 2000 1600 1200 800

WAVENUMBER(cm")

Fig. 8.22. A comparison of (a) infrared transmission and (b) RA spectra of three-layer LB
films of octadecyl TCNQ. (Reproduced from Ref. [107] with permission. Copyright (1991)
American Chemical Society.)

octadecyl-TCNQ, not only molecular orientation and structure, but also

molecular aggregation, morphology, and thermal behaviour have been
explored by infrared and ultraviolet visible (UV-Vis) spectroscopy and
atomic force microscopy [109-113]. The following conclusions could be
reached about the morphology and thermal behaviour of the LB films of
alkyl-TCNQ: (i) The LB films of octadecyl-TCNQ consist of numerous
platelet micro-crystal domains, which have the layered assembly
formed by bi-molecular layers with a thickness of 3.7 nm. A periodic
arrangement of octadecyl-TCNQ molecules with a period of 0.85 nm
can be observed inside the domains. The domains in the first layer (4.3
nm) are thicker than those above the first layer (3.7 nm). In the case of
the first layer, the direct interaction between the substrate and
octadecyl-TCNQ molecules plays an important role in determining the
thickness. (ii) A one-layer LB film of octadecyl-TCNQ shows gradual
thermally-induced structural changes while its multi-layer films show
abrupt changes. The order-disorder transition in a multi-layer LB film
of octadecyl-TCNQ with the longer even-numbered hydrocarbon chain

244
 0

0
r
C,
0 .0
o
(A en
.0

Wavenumber/cm-1 Wavenumber/cm-1

(d) 1471
CD I 1 1462
a
0
e
ID
o
a II
en
C
.0 I
E!~ ~ I

1580 1560 1540 1520 1500 1500 1480 1460 1440

Wavenumber/cm-1 Wavenumber/cm-1

Fig. 8.23. Enlargement of time-dependent changes in the infrared RA spectrum of a one-

layer LB film of pentadecyl-TCNQ on a gold-evaporated glass slide. (a) 3000-2800 cm - 1
region; (b) 2250-2190 cm-l region; (c) 1580-1500 cm-1; (d) 1500-1440 cm-1 region. The
spectra were obtained from 6 to 131 min after the film deposition. (Reproduced from Ref.
[108] with permission. Copyright (2000) American Chemical Society.)

occurs at a higher temperature than that in the corresponding LB film

of dodecyl-TCNQ with the shorter even-numbered hydrocarbon chain.
A multi-layer LB film of pentadecyl-TCNQ with the odd-numbered
hydrocarbon chain shows a transition temperature similar to the
corresponding film of dodecyl-TCNQ.
Infrared spectroscopy has also been employed to investigate aging
effects on molecular orientation and structure in LB films of dodecyl-,
pentadadecyl-, and octadecyl-TCNQ [108]. Figures 8.23a-d show time-
dependent changes in the 3000-2800, 2250-2190, 1580-1500, and
1500-1440 cm- l regions of the infrared RA spectrum of a one-layer LB
film of pentadecyl-TCNQ, respectively [108]. The spectra were meas-
ured between 6 and 131 min after the film deposition. It is noted in the

245
3000-2800 cm - region that the bands due to the CH2 antisymmetric
and symmetric stretching modes increase with time. This suggests that
the alkyl chain is nearly perpendicular to the substrate surface in the
LB film just after the film deposition, but that it becomes tilted
gradually with time.
The relative intensity of the two bands at 1471 and 1462 cm-
assigned to CH2 scissoring modes of non-interdigitated and inter-
digitated parts of the alkyl chain, respectively, changes as a function of
time (Fig. 8.23d). This result indicates that the proportion of the
interdigitated parts increases with time probably because of the
evaporation of water molecules. Figures 8.23b and c reveal that the
intensities of the C=C and C=C stretching bands increase during the
time course and that the relative intensity of the two bands at 1547 and
1531 cm 1 varies. These observations lead to the conclusion that the
TCNQ plane becomes more perpendicular with respect to the substrate
surface and the molecular axis of the TCNQ chromophore becomes
more tilted with respect to the surface normal with time. Based upon
the observations in Fig. 8.23, Morita at al. [108] have proposed a
possible model for time-dependent orientational changes in a one-layer
LB film of pentadecyl-TCNQ. Figure 8.24 depicts the model [108].
One-layer LB films of dodecyl-TCNQ also show similar time-
dependent changes, but the changes are much smaller than those for
the films of pentadecyl-TCNQ. One-layer LB films of octadecyl-TCNQ
do not show appreciable time-dependent infrared spectral changes.

Fig. 8.24. Possible model for time-dependent (a) orientational and (b) morphological
changes in a one-layer LB film of pentadecyl-TCNQ. (Reproduced from Ref. [108] with
permission. Copyright (2000) American Chemical Society.)

246
The differences in the aging effects among the one-layer LB films of the
three kinds of alkyl-TCNQ may be caused by the differences in the
strength of the hydrophobic interaction between the interdigitated
alkyl-chains and in the degree of three dimensional microcystal growth
in the one-layer LB films.
Ikegami et al. [114] studied structural changes in Langmuir (L) and
LB films of 2-methyl-5-octadecyl-N, N'-dicyanoquinonediimine
(C,,MeDCNQI) induced by charge-transfer (CT) reaction at the air-
water interface by use of infrared and UV-Vis spectroscopy. Compari-
son of UV-Vis spectra of pure C,,MeDCNQI L films with those of
C, 8MeDCNQI-CuI mixed L films suggests that CT reactions take place
at the air-water interface in the latter case. These L films were
deposited onto solid substrates as LB films by the horizontal lifting
method. Polarized infrared and UV-Vis spectra and X-ray diffraction
patterns of the LB films indicate that an interdigitated bi-layer struct-
ure of the pure films changes into a mono-layer structure for the CT
films.
Figure 8.25 shows infrared spectra in the 3200-2700 cm-l region of
LB films of pure C,,MeDCNQI and the 1:5 mixture of C,,MeDCNQI
and CuI [114]. The antisymmetric and symmetric stretching bands
appear at 2918 and 2848 cm - l, with the line widths of 14 and 11 cm-l,
respectively, in the spectrum of pure C,,MeDCNQI film. The line
widths of these bands reflect the mobility of alkyl chain. Therefore, it is

0
'o
a
ro

n
C,
0
.0
co
a
n

3.2 3.1 3.0 2.9 2.8 2.7

wavenumber (103 cm-1 )

Fig. 8.25. Structure of Cis8MDCNQI and polarized infrared spectra in the 3200-2700
cm -1 region of 56-layer LB films of (a) pure CMe,DCNQI and (b) a 1:5 mixture of
CNeDCNQI and Cul. Incident angle is 60 .

247
 wavenumber (103 cm- 1)
-1
Fig. 8.26. Polarized infrared spectra in the 2300-1300 cm region observed for 56-layer
LB films of (a) a pure C18MeDCNQI and (b) a 1:5 mixture of C1 8MeDCNQI and Cul. The
angle of incidence is 60 . (Reproduced from Ref. [114] with permission. Copyright (2000)
American Chemical Society.)

concluded that alkyl chains of C,,MeDCNQI molecules are closely

packed with highly ordered trans conformation in the LB film of
C,8 MeDCNQI. The corresponding bands are observed at 2921 and 2851
cm-' with the line widths of 22 and 14 cm-', respectively, for the CT
film, indicating that the alkyl chains in the CT film are rather loosely
packed with non-negligible content of the gauche conformation.
Figure 8.26 depicts the tilting-incident polarized infrared absorp-
tion spectra of an LB film of C,,MeDCNQI and a CT film of
C,sMeDCNQI and CuI [114]. In Fig. 8.26a, bands due to C=N (a'v 5 and
a'v6), C=C (a'vs), C=N (a'vg), and C-C (a'vl2) stretching modes are
observed ; the vibrational modes are numbered as a'vm, according to
Lunardi and Pecile [115]. The CT degree in the LB film of C 18MeDCNQI
and CuI was estimated to be -1 by the downward shifts of the C=C and
C=N stretching bands [114]. It is noted in Fig. 8.26 that all the infrared
bands observed for the CT film are stronger with s-polarized light and
that the a'v8 and a'v,,2 bands for the LB film are more intense with p-
polarized light. It is also of note that the C-N stretching bands and the
electronic excitations show stronger intensity with the s-polarization
for both films. Therefore, it seems likely that the long axis of the
DCNQI plane preferentially lies in the film plane in both films and its
short axis is nearly perpendicular to the film plane in the pure film and
lies in the film plane in the CT film.

248
 Ikegami et al. [114] estimated the orientation of the alkyl chain and
chromophore in the LB films of C 18MeDCNQI and C, 8 MeDCNQI-CuI
based upon the method proposed by Chollet and Messier [101]. Chollet
and Messier showed that it is possible to take into account the multiple
reflection due to the thin films and to calculate the out-of-plane
orientation order parameter of a vibrational mode, S = <3cos 20-1>,
where 0 is an angle between the transition dipole moment and the
normal axis of the film plane, as

S=2-3(1+ n2B
nln32C)

B= os n+ Cos A() -cos cos Ap(O)

3 (8.9)
nl +-I

C = sin 1sin/3{A(0) + A (0)

where nl, n2, and n3 denote the refractive indexes of air, a LB film, and a
substrate, respectively; Ap (y) and As () are the absorbance due to
the transition moment with the angle of incidence being yl. w2 and W 3
denote an angle of reflection in the LB film and that in the substrate,
respectively.
Ikegami et al. [114] calculated the S parameter for the alkyl chain
by the following equation:
S (chain) = -S(va(CH 2)) - S(vs(CH2)) (8.10)
where S (chain) is the S parameter for the alkyl chain and S(Va(CH 2)) -
S(vs(CH 2)) are those of CH 2 antisymmetric and symmetric stretching
modes, respectively. The averaged tilt angle of the alkyl chain was
calculated to be 40 and 50 , respectively, for the pure LB and CT films.
Similarly the averaged tilt angles of the long and short axes of the
DCNQI group were estimated to be about 75 and 30 , respectively, in
the pure LB film, and those in the CT film were calculated to be both
about 70 .
Taking account of all information obtained from infrared spectro-
scopy and x-ray diffraction patterns, Ikegami et al. [114] proposed
possible averaged structures of the LB and CT films as shown in Fig.
8.27. As in the case of the LB films of alkyl-TCNQ, an interdigitated bi-
layer structure was considered for the LB films of C, 8MeDCNQI.
Hasegawa et al. [116] investigated a hydrogen bonding network
formed between accumulated LB films of barbituric acid and

249
 ,

(b)

II
84A-
-------~ iW3-'Al
~ ~ ~ ~ 6
A '.4 -
(c)
Cu 1120 /
- ; 3. t te

_- - - - - -

Fig. 8.27. Schematic illustration of(a) C1,MDCNQI molecular shape (rotational freedom
is assumed for the connecting point between the C18MeDCNQI group and alkyl chain); (b)
possible structure of pure LB films of Cs1MeDCNQI; (c) possible structure for CT LB films
of the CMeDCNQI-Cul mixture. (Reproduced from Ref. [1141 with permission.
Copyright (2000) American Chemical Society.)

triaminotriazine derivatives (C,,BA and 2 C 8sTAZ) by use of infrared

spectroscopy. They deposited the LB films at various surface pressures
on a gold-evaporated glass slide covered with a deuterium cadmium
stearate (CdSt-d3 5 ) mono-layer. The layer configuration of IR//C,,BA/
2C,,TAZ/CdSt-d 35 /Au gave the most interesting results. The C,,BA
layer was deposited at various surface pressures on the 2 C,,TAZ mono-
layer. Figure 8.28 shows the structures of C,,BA/2 C,,TAZ and RA
spectra of accumulated C,,BA/2 C,,TAZ LB films deposited on CdSt-d 35
monolayer on a gold-evaporated glass slide. At a low surface pressure
(5 and 10 mN m-') of C,,BA monolayer, an Amide I (C=O stretching)
band at 1740 cm -l arising from C1sBA almost disappears, and instead, a
strong band appears at around 1700 ml. This observation suggests
that the C=O group interacts with TAZ moiety through strong
hydrogen bonding. A marked change takes place at 15 mN m-l; a new
band appears at 1755 cm- '. This band becomes a dominant band when
the surface pressure is increased to 20 mN m-l. The band at 1754 cm-l
is characteristic of carbonyl group of BA, and it appears at 1754 cm-l

250
 m-1

0 0

H'OA N--N
C ,-RA or. T7
-lI'- "'18^'4 Wavenumber I cm'' Wavenumber I cm'

Fig. 8.28. Structure of C18BA and 2G8TAZ and RA spectra of accumulated C1 8BA and
2G8TAZ LB films deposited on Cd stearate-d 35 monolayer on a gold-evaporated glass
slide. The C18BA monolayers were prepared at various surface pressures on 2G8 TAZ
monolayer that was prepared at the fixed surface pressure, 20m Nm 1. (Reproduced from
Ref. [116] with permission. Copyright (2000) American Chemical Society.)

when BA molecules are dispersed in an argon matrix. This band shows

a downward shift by about 60 cm-l, when BA is in the solid state at 20K
due to hydrogen bonding formation. Therefore, it is very likely that the
accumulated layer has free C=O groups at 15 and 20 mN m -l .
With the increase in the surface pressure great changes are also
observed in the C-H stretching vibration region. At low surface
pressure both CH2 asymmetric and symmetric stretching bands are
very weak. When the surface pressure is increased to 15 mN m-l, the
intensities of the bands increase largely. It is likely that the alkyl
chains tilt from the surface normal above 15 mN m-l. The CH 2 asym-
metric and symmetric stretching bands appear at 2919 and 2850 cm- '
irrespective of surface pressure. This indicates that the alkyl chains
are highly organized and the molecular configuration is trans-zigzag.
The N-H stretching band shifts from 3144 and 3112 cm l upon
going from 5 to 20 mN m -l . This result suggests that the hydrogen
bonding through the N-H group becomes stronger with the surface
pressure.
The tilt angles of the alkyl chain in the accumulated layer and the
under layer as a function of the surface pressure were calculated by a

251
 I
Jj
O
30 .under-layer
25

I 20

C
accumulated layers

ao . ' I I -
0 5 10 15 20 25
Surface pressure of C BA monolayer I mN m"-'

Fig. 8.29. Tilt angle changes of alkyl chains in the accumulated layer and the underlayer
shown by solid and dashed lines, respectively. (Reproduced from Ref. [116] with
permission. Copyright (2000) American Chemical Society.)

procedure proposed by Hasegawa et al. [116]. The procedure can take

anisotropic optical constants into account. This allows one to take any
molecular orientation into account to yield expected absorbance. For
detailed description, readers are referred to the literature. Figure 8.29
illustrates changes in tilt angles of alkyl chains in the accumulated
layer and the under layer shown by solid and dashed lines, respectively
[115]. Figure 8.30 shows schematic models of the accumulated layer at
a low and a high surface pressure of C,,BA, respectively [116]. At low
surface pressures, the alkyl chains stand almost perpendicularly to the
film surface. The chains stand perpendicularly also in the side view. In
the same view BA and TAZ planes are a little tilted because of the
configuration of the nitrogen atom. It is particularly noted that C,,BA
and 2C 1 8TAZ molecules are complementarily hydrogen bonded and
there is no free NH2 or C=O group.
The alkyl chain tilts 10 or 12 from the surface normal (Fig. 8.30b)
at high surface pressure [116]. The coupling mechanism of the C,,BA
and 2C,,TAZ molecules is largely different from that at low surface
pressure. The hydrogen bonding coupling is imperfect at high surface
pressure; one of the C=O groups in a BA ring remains as a free C=O

252
 (a)

C18BA

2C18TAZ

Front view of
BA I TAZ: (BA at 5 and 10 mNm-1) Side view

5';

(b)

C18BA

free

2C18TAZ

/.
Front view of -r
BA I TAZ: (BA at 15 and 20 mNm-1) Side view

Fig. 8.30. Schematic views of the accumulated layers. The images at (a) low surface
pressure and (b) high surface pressure are drawn. (Reproduced from Ref. [116] with
permission. Copyright (2000) American Chemical Society.)

253
group. It is very likely that the band at 1755 cm-l arises from this free
C=O group.
The highly condensed monolayer is formed when the surface
pressure of the C,,BA monolayer is high. It is difficult for the highly
condensed monolayer to incorporate into the condensed 2C,,TAZ mono-
layer deeply. In other words, the interaction between the two layers
becomes weak. Therefore, the molecular aggregation by lateral hydro-
phobic interaction plays a dominant role to lead the film to have a close-
packed structure; the tilted stance is known to be most stable. Thus,
the conclusion that the molecular tilting stance about 10 with all trans
conformation seems to be reasonable [116].

8.7 DEVELOPMENTS IN INFRARED SPECTROSCOPY OF BIOLOGICAL

MOLECULES AND MATERIALS

Infrared studies of biological molecules and materials have a long

history [117-133]. As early as 1911, W.W. Coblentz [117] pointed out
that infrared spectroscopy had considerable promise in the studies of
biological samples. One can find a number of papers published in the
1950s reporting even medical applications of infrared spectroscopy
[118,119]. In the 1960s infrared spectroscopy was extensively em-
ployed to investigate the secondary structure of proteins [119]. The
relationship between the secondary structure elements of proteins and
the frequencies of amide I and amide II bands was found in the early
1960s. Infrared spectroscopy was also applied to explore the hydrogen
bonds of nucleic acid bases. Before NMR became a powerful tool for
investigating the structures and functions of biological molecules and
materials, infrared spectroscopy had been a major technique for
structural studies of biological molecules [119].
In 1971 one very famous book on biological applications of infrared
spectroscopy was published [119] in which can be found many inter-
esting applications even to microbiology and medicine. However, in the
1970s infrared spectroscopy did not receive keen interest from
biological scientists because NMR and Raman spectroscopy, which are
very suitable for aqueous solutions or physiological conditions, were
developing rapidly. The revival of infrared studies on biological mater-
ials happened in the 1980s because of the rapid development of FT
techniques which opened up a variety of possibilities for infrared
measurements; for example, infrared measurements under physio-

254
logical conditions, time-resolved measurements, and measurements of
micro samples became popular [121-131].
Infrared spectroscopy has several advantages for the studies of
biological molecules and materials over other techniques such as NMR,
CD, fluorescence, Raman, and X-ray crystallography [121-131]. First,
it is possible to measure high-quality spectra of biological molecules in
a variety of states such as aqueous solutions, films, crystals, and
organic solutions. Second, very small amounts of samples are enough to
obtain good spectra and no or very little pretreatments are requested
for the infrared measurements. Infrared spectroscopy is free from the
light-scattering (CD) or fluorescence (Raman) problem, and does not
depend on the molecular weight of biological molecules (NMR).
Another important feature of infrared spectroscopy is the potential of
slow and first kinetic studies of biological molecules [127].
Today, biological applications of infrared spectroscopy may be
divided into three categories [122]. One is structural studies of bio-
logical molecules such as proteins, lipids, nucleic acids, and biological
pigments [123,124,126,127,130,131]. Another is investigations of
dynamics and excited states of biological molecules [125-127]. Repre-
sentative examples of this category are studies of protein dynamics,
mechanism of photosynthesis, and light-induced mechanism of bact-
eriorhodopsin; time-resolved infrared spectroscopy and low-temp-
erature infrared spectroscopy are usually employed for these
investigations. Yet another category is biomedical applications of
infrared spectroscopy [128,129,132]. This category involves wide-
spread researches and applications from non-destructive identifi-
cations of bacteria, in situ determination of blood sugar to diagnosis of
cancer cells. In this section, representative examples selected from
each category are introduced.

8.7.1 Infrared spectra of proteins

Most infrared studies of proteins are concerned with the conformation

of peptide backbones, but there are some other interesting and import-
ant infrared studies such as those on structures and functions of
chromophoric groups like hemes and retinals, the microenvironments
of amino acid residues like cysteine residues (Cys) and the hydrogen
bonding of water molecules in proteins [121-127,130,131,133-135]. We
discuss here infrared studies of the secondary structure of proteins;

255
infrared studies of photoreceptor proteins will be described in Section
8.7.6.
Infrared spectroscopy is powerful in investigating the conformation
of the peptide backbones of proteins because the frequencies of amide I
and amide II bands are very sensitive to the conformation. Thus, by
analyzing the amide I and/or amide II band regions, one can estimate
the percentage of each structure element like a-helix, p-structure and
random coil structure [123,124,130-135]. However, detailed quanti-
tative analysis of amide I and amide II bands is not always an easy task
because a number of bands due to various secondary structures overlap
heavily. Table 5.2 summarizes the relationship between the frequency
of amide I band and the secondary structure. For the analysis of the
overlapping amide bands, various spectral analysis methods have been
proposed [124,126,130-133]. Most are frequency-based methods which
rely on peak assignments in either second derivative or deconvoluted
spectra. Recently, chemometrics [132] and two-dimensional (2D)
correlation spectroscopy [133] have also been introduced to explore the
amide I band region. The second derivative, deconvolution and 2D
correlation spectroscopy, enhance band resolution, enabling one to
identify the different structures in a protein and also to monitor
structural variations induced by protein denaturation. Calculation of
difference spectra is also very useful to probe conformational changes
caused by perturbation (for example, temperature, pH, and detergent)
because the resultant difference spectrum only yields bands that are
related to the groups involved in the conformational changes. For
quantitative analysis curve-fitting is often employed [124]. This anal-
ysis usually provides a very good estimate of the secondary structure
which is in good agreement with that obtained by x-ray crystallography
and CD spectroscopy. However, it must be kept in mind that the curve-
fitting method needs some assumption. For example, it assumes that
the amide bands arising from different secondary structures have the
identical molar absorptivities. Moreover, the curve-fitting analysis
requests precise assignments of all the component bands and the initial
choice of input parameters (the number of component bands and their
frequencies and widths).
The potential of 2D correlation spectroscopy in the analysis of amide
I and II regions is described in Section 9.7. The chemometrics method
needs to use a calibration matrix of the infrared spectra of proteins of
known x-ray structure [132]. Among various multivariate methods,
partial least squares (PLS) and factor analysis (FA) are employed for

256
 1750 1700 1650 1600 1550 1500
Wavenumber (cm-')

(b)

20C
1750 1700 1650 1600 1550 1500
Wavenumber (cm-')

Fig. 8.31. Temperature-dependent spectral changes of deconvoluted infrared spectra of

human transferrin receptor in D2 0. (a) pH 7.4, (b) pH 5.6. (Reproduced from Ref. [134]
with permission. Copyright (1994) Elsevier.)

the quantitative analysis of infrared spectra of proteins. This method

encounters difficulties when the spectral properties of the unknown
protein are not involved in the properties of the spectra within the
calibration set.
The amide I band region is particularly useful for monitoring
structural changes caused by protein denaturation. Figures 8.31 and
8.32 show an example of infrared studies of protein denaturation [134].
Figure 8.31 compares effects of temperature on deconvoluted spectra of
human transferrin receptor in D 2 0 between extracellular pH (pH 7.4)
and endosomal pH (pH 5.6). The quantitative analysis of the amide I
region shown in Fig. 8.31 by Hadden et al. [134] indicated that the
protein consists of 56% a-helix, 19% P-sheet, and 14% turns at extra-
cellular pH.

257
 -1 -
pH 7.4 PHcm 7.4
1617 l 1640cm-
1640cm-1
1617cm 1655cm 1683 cm1
-14cm 1549cm

,AAAAAAAAAAA.&A A AA . . M ask

..
(I
(U

n
0
.0
III 60 so i M A
iiii 9.

----As _----. !. -----. ------------------

I I
II iI I I Ittttt11
0 40 60 80 60 an n
Temperature (C)
H 5.6
- 1
140cm61-1
-1 - 1 -1
pH 5.6 1617 cm 1640cm 1655cm 1683cm 1549cm

/AA :.-... .

a,
C
(b) ................
1,
co
(U
-eP
0 UI
'

-----__ _-IU

20 40 60 80 60 40 20
Temperature (C)

Fig. 8.32. Temperature-dependent intensity variations of selected bands in deconvolved

spectra of human transferrin receptor in D2 0. (a) pH 7.4, (b) pH 5.6. (Reproduced from
Ref. [134] with permission. Copyright (1994) Elsevier.)

Figure 8.32 compares temperature-dependent intensity variations

of selected bands in the deconvoluted spectra of transferrin receptor at
pH 7.4 and pH 5.6 in D2 0 [134]. It can be seen from Fig. 8.32 that
transferrin receptor at pH 7.4 undergoes thermal denaturation at the
sharp midpoint temperature around 71 C while the thermal stability is
reduced by approximately 15C at endosomal pH (pH 5.6).

258
8.7.2 Infrared spectra of chlorophylls

Infrared spectra of chlorophylls (Chl) have been actively studied since

the 1950s [136]. One recent progress in the infrared studies of Chl is
that infrared spectra of Chl in highly dilute solutions (-10- 6 M) can be
measured [137]. Figure 8.33 shows infrared spectra of Chl-a in water-
saturated carbon tetrachloride solutions of 8x10 -2 M, 3x10 -3 M, 5x10 -4
M, 6x10 -5 M, and 9x10 - 6 M. [137]. Note that the highly dilute solutions
of Chl-a provide the infrared spectra with fairly high signal-to-noise

2000 1800 1600 1400

WAVENUMBER (cmr )

Fig. 8.33. Infrared spectra of Chl-a in water-saturated carbon tetrachloride solutions of:
-3 4 -5
(a) 8x 10 - 2 M; (b) 3x10 M; (c) 5x 10 M; (d) 6x 10 M; (e) 9x 10-6 M. (Reproduced from Ref.
[137] with permission. Copyright (1993) American Society for Photobiology.)

259
ratio. The advantage of infrared spectroscopy in Chl-a research is that
it gives intense bands due to C=O stretching modes of the ester groups
and the 9-keto group, which play key roles in forming dimers and
oligomers of Chl-a [138-141]. Bands near 1736, 1693, 1654, and 1608
cm-l1 are assigned to a C=O stretching mode of the ester groups, a C=O
stretching mode of the free 9-keto group, a C=O stretching mode of the
9-keto group which coordinates with the Mg atom of another Chl-a
molecule, and a methine-bridge stretching mode (IR I-band),
respectively [139].
In concentrated (above 10 - 3 M) carbon tetrachloride solutions Chl-a
forms a five-coordinated dimer in which the 9-keto group of one Chl-a
coordinates to the Mg atom of another Chl-a. It seems, therefore, that
the bands at 1693 and 1654 cm-l in Fig. 8.33 are due to the free and
coordinated 9-keto groups of the dimer, respectively. With the decrease
in the concentration of Chl-a, the intensity of the band at 1654 cm - l
decreases while that at 1693 cm-l increases concomitantly; the former
is almost missing in Fig. 8.33e. These observations led Okada et al.
[137] to conclude that the monomer is predominant in the dilute water-
saturated carbon tetrachloride solutions.
The coordination number of the Mg atom can be determined by the
frequency of IR I band [139]; the IR I band appears near 1608 cm- when
the Mg atom of Chl-a assumes a five-coordination with one axial ligand
whereas it is observed near 1597 cm-l when it takes a six-coordination
with two axial ligands. The IR I band is identified near 1609 cm-l in the
spectra of the concentrated solutions (Fig. 8.33a,b), suggesting that the
Mg atom is five-coordinated. The frequencies of the IR I band are not
reliable in the spectra of the dilute solutions because a strong band
near 1550 cm-l due to carbon tetrachloride makes accurate subtraction
of the solvent spectrum from the Chl-a spectra difficult in the
1600-1500 cm-l region.

8.7.3 Infrared spectra of a model compound for

phospholipids

Infrared spectroscopy is an excellent tool for investigating the struct-

ure and phase transitions of fatty acids, phospholipids, and biomem-
branes because the frequencies of CH2 antisymmetric and symmetric
stretching bands are very sensitive to the conformation of hydrocarbon
chains and those of C=O stretching bands reflect the strength of the
hydrogen bonds of the C=O groups [126,130].

260
 0
H
II
C
\ / I H 2' 4' 6
O-\ 7 0H 0 CH~ CH, CH2

C-C- 5C-CH2 C1 CH 2 CH2 CH2

/ H 1 6 11 ' ' 7'
K-C 0 8' lo' 12'
14' 16'
\ CH2 CH2 CH CH2 CH2
H /\ / \ / \2
, \ / \ /
CH 2 CH2 CH2 CH2
9' 1
11' 13' 15'

(b)
0.
I-
1587
2
uj 1585
2 1583
1.
1581
Q:
1579 -
0
1577
1575 . ..
l. . . l
_
I I
20 25 30 35 40 45 50 55 60

Temperature/C

Wavenumber/cm 'l
Fig. 8.34. (a) Temperature-dependent spectral changes in the 1800-1500 cm 1 region of
potassium salt of ascorbic palmitate (APK) in a deuterated aqueous solution (0.1 M). (b)
The frequency of the C=C stretching band versus temperature. (Reproduced from Ref.
[106] with permission. Copyright (1981) National Research Council of Canada.)

Figure 8.34 shows temperature-dependent spectral variations in

the 1800-1500 cm - ' region of potassium salt of ascorbic palmitate
(APK) in a deuterated aqueous solution (0.1 M) [106]. Bands near 1740
and 1580 cm -1 are due to the C=O and C=C stretching modes, respect-
ively. It is noted that the C=O stretching band shows a downward shift
with broadening as temperature is increased. This observation
suggests that the hydrogen bond of the C=O group becomes stronger
with temperature [140]. The C=C stretching band shows an upward
shift and Fig. 8.34b plots its frequency versus temperature. The plot
reveals that the phase transition of APK from gel or core gel to micelle
takes place near 47-48C.
Figure 8.35a depicts the 3000-2800 cm -1 region of the same spectra
as those in Fig. 8.34a. The CH 2 antisymmetric and symmetric stretch-

261
 (a) (b)
r I
U.
I I I I I i I
2853.5 - 8.0

2852 ..
a
U
0
E - .* ..
0
-E
2851.5 . 6.0'

2850.5 5 0

3000 2960 2920 2880 2840 2800 20 25 30 35 40 45 50 55 60

t
Wavenumber/cm Temperature/C
-1
Fig. 8.35. (a) The 3000-2800 cm region of the same spectra as those in Fig. 8.34 (a). (b)
The frequency (circles) and bandwidth (triangles) of the CH 2 symmetric stretching band
versus temperature. (Reproduced from Ref. [106] with permission. Copyright (1981)
National Research Council of Canada.)

ing bands are observed near 2920 and 2850 cm l, respectively. It is well
known that when the alkyl chain is highly ordered (trans-zigzagconfor-
mation), the CH 2 antisymmetric and symmetric stretching bands
appear at 2918 and 2848 cm-l, respectively, while if conformational
disorder (gauche forms) is included in the chain, they shift upward up
to 2926 and 2856 cm-l depending upon the content of gauche con-
formers [105,106].
In Fig. 8.35a, both bands due to the CH 2 antisymmetric and sym-
metric stretching modes shift upward with broadening with temper-
ature [106]. Figure 8.35b plots the frequency of the CH2 symmetric
stretching band and its bandwidth versus temperature [106]. It is of
note that the frequency changes markedly near 47-48C and that the
bandwidth varies at slightly lower temperature. The frequency shift of
the CH 2 symmetric stretching band indicates that the conformation of
the alkyl chain of APK changes from a trans-zigzag structure to a
structure with some gauche conformers upon the phase transition. The
band broadening occurs even below 40C, suggesting that the fluidity of
the alkyl chain increases at much lower temperature than the phase
transition temperature. The frequency shifts of the two CH, stretching
bands and those of the C=O and C=C stretching bands happen in the
same temperature range, and thus the change in the strength of the
hydrogen bond of the C=O group is linked with the conformational
change in the alkyl chain [106].

262
 In order to monitor the structural change of a particular part in the
alkyl chain the deuteration of that part is very useful because the CD2
stretching bands (2300-2000 cm- l region) are observed separately from
the CH 2 stretching bands [142]. Infrared spectroscopy has also been
applied to studies of phase transition of biomembrane in living bacteria
[143].

8.7.4 Infrared spectra of intact bacterial cells

Infrared spectra of proteins, Chls, and phospholipids are examples of

infrared spectra of basic biological molecules. Let us examine infrared
spectra of rather complicated biological materials. As an example of
such biological materials, infrared spectra of bacteria are described
here.
The infrared measurements of bacteria are not difficult. In fact,
even in the 1950s and 1960s infrared spectra of a number of bacteria
were measured and it was pointed out that infrared spectroscopy has
potential in the identification of bacteria [118,119]. However, it was the
middle of the 1980s that systematic infrared investigations of
characterizing intact bacteria started [129,144]. Figure 8.36a and b

2,
a,
E2

,:

at

-1
Wavenumber [cm ]

Fig. 8.36. Infrared spectra and their second derivative spectra of gram-positive and
gram-negative bacteria. (A) Staphylococcus aureus, PS96; (B) Pseudomonas
chlororaphis,American Type Culture Collection (ATCC) 17809. (Reproduced from Ref.
[144] with permission. Copyright (1991) VCH.)

263
shows infrared spectra of gram-positive and gram-negative bacteria,
respectively. Their second derivative spectra are also shown in Fig.
8.36. Table 8.2 summarizes proposed band assignments for infrared
spectra of bacteria (from Ref. [144]). These band assignments are based

TABLE 8.2
Proposed assignment of some bands frequently observed in infrared spectra of bacteria
(from Ref. [144])

Band Frequency Relative Assignment*

numbering (cm) intensity*
-3500 m vOH stretching
-3200 m-s vNH stretching (Amide A) of proteins
2959 w va(CH 3) stretching of methyl
2934 vw va(CH 2) stretching of methylene
2921 m va(CH 2) stretching of methylene in fatty acids
2898 vw vCH stretching of methine
2872 w vs(CH 3) stretching of methyl
2852 m v,(CH2) stretching of methylene in fatty acids
1 1741 w vC=O stretching of esters
1715 vw vC=O stretching of esters, carbonic acids
1695 w different amide I band components resulting
1685 w from antiparallel pleated sheets and -turns
1675 w of proteins
2 -1655 s amide I of a-helical structures
3 -1637 s amide I of P-pleated sheet structures
4 1548 s amide II band
5 1515 m "tyrosine" ring vibration band
1498 w "pheylalanine" ring vibration band
6 1468 w-m 6(CH2 ) bending of methylene
7 -1400 m m,(COO-) stretching of carboxylates
1310-1240 w amide III band components of proteins
8 1250-1220 w-m va(PO2-) stretching of phosphodiesters
1084-1088 w-m v,(PO2) stretching of phosphodiesters
9 1200-900 m C-O-C
720 vw p(CH 2) rocking of methylene
900-600 w "fingerprint region"
*m = medium; s = strong; v = very weak; w = weak; s = symmetric; a = asymmetric or
antisymmetric, v = stretching; 6 = deformation; p = rocking.

264
upon infrared spectral analysis of isolated substances and pure
compounds such as proteins, lipids, and nucleic acids.
Infrared spectra of bacteria can be divided into several spectral
regions [129,144]. The 1800-1700 cm -l region involves bands due to C=
O stretching modes of phospholipids. The 1700-1500 cm-l region is
dominated by the strong amide I and amide II bands. A few weak
features arising from amino acid residues are also observed in this
region (see Table 8.2). The region between 1250 and 1200 cm-l shows
medium bands due to a PO,- antisymmetric stretching mode of
phosphodiesters. This region provides information about DNA, RNA,
and phospholipids. The 1200-900 cm-l region involves a number of
weak to medium features arising from C-O-C and C-O stretching
modes of polysaccharides and those from a PO,- symmetric stretching
mode of phosphodiesters. One can identify a number of bands in the
second derivative spectra in this region. This region is most sensitive
and selective for the differentiations of bacteria down to the strain and
even serotype level [144]. The region between 900 and 600 cm-l is
referred to as the 'bacterial fingerprint region' because it exhibits many
weak, but extremely characteristic features [144]. Different species
and strains of bacteria have different chemical composition and chemi-
cal structures and each species or strain show a unique infrared
spectrum. The recent development of FT-IR instruments together with
the availability of powerful personal computers has led to a variety of
infrared studies of bacteria [144]. Current infrared spectroscopy
enables differentiation at different levels, classification, and identifica-
tion to genus, species or strain levels of bacteria. Infrared spectroscopy
is also used for the detection and identification of particular cell compo-
nents and for the investigation of growth-dependent phenomena and
characterization of cell-drug interactions [144].

8.7.5 Application of infrared microspectroscopy and imaging

to biological sciences

By combining an infrared spectrophotometer with a microscope,

molecular information can be obtained with a spectral resolution of -10
pm [71,145,146]. Infrared microspectroscopy has recently been applied
extensively to biological sciences and medicine because of the
capability of in situ chemical analyses of microscopic samples. One of
the most exciting developments in infrared microspectroscopy is infra-
red imaging technology (Section 8.4).

265
 Figure 8.37A shows an example of an infrared imaging study of a
biological tissue [71]; it is a cross-section of an unstained rat retina
(image width, 500 pm) viewed with an infrared microscope/spectro-
meter with all-reflecting differential interference contrast optics.
Figures 8.37B and C depict infrared spectra obtained from the outer
segment which is rich in lipid materials and from the outer nuclear cell
layer, respectively. In both spectra, bands due to amide I and II are
observed very strongly. A band near 1700 cm-l is much stronger in
spectrum B than in spectrum C. A band at 1235 cm - is assigned to a P=
O stretching mode arising from the nuclei. Lipid chain length, branch-
ing, and glycolipids are investigated by comparing the intensities of
bands due to C=O stretching, CH3 stretching, CH 2 stretching, and
H-C-OH group vibrations. The degree of the unsaturation may be
estimated from the CH stretching band at 3015 cm-l on the carbon that
is attached to the C=C band. In this way the molecular chemistry of the
individual retina layer has become possible by combining infrared
spectroscopy and microscopy [71]. Various kinds of pathological tissues
have been subjected to infrared microspectroscopy studies [145]. For
example, cancerous tissues, Alzheimer's disease plaques, and diseased
arteries have been investigated by infrared microspectroscopy.
Infrared microspectroscopy combined with artificial neural networks
has been applied to the diagnosis of cervical cancer. This topic will be
outlined in Section 8.7.7.

IPL
INL
OPL
ONL
iS
OS
Fig. 8.37. (A) A cross-section of an unstained rat retina (image width, 500 pm) viewed
with an infrared microscope/spectrometer with all-reflecting differential interference
contrast optics. (B and C: see opposite page,)

266
 3500 3000 2500 2000 1500 1000
Wavenumber (cm-l)

Abs C

3500 3000 Z500 2000 1500 1000

Wavenumber (cm-1)

Fig. 8.37 (continued). Infrared spectra obtained from (B) the outer segment, and (C) the
outer nuclear cell layer. (Reproduced from Ref. [71] with permission. Copyright (1999)
American Association for the Advancement of Science.)

267
8.7.6 Structure and function of bacteriorhodopsin studied by
low-temperature infrared difference spectroscopy

The photoreceptor proteins have been very attractive targets for infra-
red spectroscopy [125,147,148]. The proteins can be divided into two
groups: those that use light as energy source and those that employ
light as information source. The proteins in photosynthesis systems,
bacteriorhodopsin, and rhodopsin belong to the former while visual
pigments, phytochrome, and yellow proteins belong to the latter. Time-
resolved infrared spectroscopy and low-temperature infrared spectro-
scopy are very suitable to explore the light-induced mechanism of these
photoreceptor proteins.
As a representative example of infrared studies on photoreceptor
proteins, this section describes low-temperature infrared spectroscopy
studies on the light-induced mechanism for proton pumping of
bacteriorhodopsin [148]. Bacteriorhodopsin, a protein present in the
purple membrane of Halobacterium salinarium, performs uni-
directional transport of protons across the membrane by use of light
energy absorbed in the retinylidene chromophore bound to lysine 216
(Lys 216) through the protonated Schiff base. Figure 8.38a depicts
seven helices of bacteriorhodopsin with important residues for the
function and the structure of its photointermediates are shown in Fig.
8.38b. The intermediates shown in Fig. 8.38b are emerged by the light-
induced isomerization of the all-trans retinal. The photocycle is

f,*; WLtim

Fig. 8.38. (a) Seven helices of bacteriorhodopsin with a retinal bound to the -amino
group of Lys216 via a protonated Schiff base. Opposite page: (b) Photocycle of bacterio-
rhodopsin. (Reproduced from Ref. [148] with permission. Copyright (1997) Japanese
Biochemical Society.)

268
completed in less than 10 ms. One proton moves from the Schiff base to
asparagine 85 (Asp 85) in the L-to-M process and another from Asp 96
to the Schiff base in the M-to-N process.

BR
Light
Nas

0
-, K

I
N 3

I"

g. z

Fig. 8.38. (b) Caption opposite.

269
 I I I I I I I
0.02 C=O of COOH
O-H of H2 0 C=O of petide
f-3486 1 A L
148-1
0.00

3643
-0.02

-0.04 I 0A04 -II 0

I I I I lII I I
3800 3600 3400 3200 3000 1800 1600 1400 1200 1000 800

wavenumber(cm 1 )

Fig. 8.39. A low-temperature infrared difference spectrum between L and BR of bacterio-

rhodopsin (Reproduced from Ref. [148] with permission. Copyright (1997) Japanese
Biochemical Society.)

Of particular interest in studying bacteriorhodopsin is that its

discrete intermediate states can be investigated over a picosecond to
millisecond range. One can apply low-temperature infrared spectro-
scopy as well as time-resolved infrared spectroscopy to explore the
chemical process occurring in the photocycle [148]. Difference spectra
may be measured at 80K for K, 170K for L, 230K for M, and 274K for N
(the spectrum of O can only be obtained by time-resolved method).
Figure 8.39 shows a L minus BR spectrum of bacteriorhodopsin
[148]. As is evident from Fig. 8.39, infrared studies provide information
about the protonation states of carboxylic acid residues, hydrogen-
bonding changes in water, peptide carbonyls, and others. Let us focus
our attention on the 3750-3450 cm - l region where OH stretching bands
of water molecules and an NH stretching band of tryptophan 182 (Trp
182) are expected to appear. Figure 8.40 shows infrared difference
spectra in the 3750-3450 cm - ' region between the unphotolyzed state
(BR) and the intermediate (K, L, or M). Of particular note in Fig. 8.40
is that the absorbance of each water band corresponds to one or a few
water molecules [149]. In other words, one can investigate the struc-
ture and environment of a water molecule from a frequency of the OH
stretching band [148,149]. Water molecules play very important roles
in the intramembrane signalling mediated by its hydrogen-bonding.

270
 K-BR

L-BR

] 3607 3577

M-BR

3643

3700 3600 3500

wavenumber(cm 1 )

Fig. 8.40. Low-temperature infrared difference spectra in the 3750-3450 cm- 1 region of
bacteriorhodopsin between photointermediates (K, L, M) and unphotolyzed state (BR).
(This figure was prepared by H. Kandori.)

For example, in L the Schiff base forms strong hydrogen-bonding with

a water molecule coordinated with Asp85. This structure leads to
transfer of the Schiff base proton to Asp85 in the L-to-M process,
triggering proton release from glutamic acid (Glu 204) to the extra-
cellular surface. In addition, a string of hydrogen-bonding mediated by
internal water molecules and peptide carbonyls in helices B and C, and
Trp182 in helix F induces structural changes around Asp96, which may
bring about the structural variations occurring later during the M-to-N
process.
In the K minus BR spectrum of Figure 8.40 bands due to water are
very weak but large spectral changes take place in the OH stretching
band region of the L minus BR spectrum. Three bands at 3643, 3607,
and 3577 cm-l are assigned to a water molecule present close to the
unprotonated Asp85, that close to Asp96, and that close to the peptide
carbonyl of valine 49 (Val 49), respectively, based upon comparison of
the spectrum with those of D85N, D96N, and V49M mutants [150].

271
 Pro'

Asp85

Fig. 8.41. Hydrogen-bonding network from Asp85 to Asp96. (Reproduced from Ref. [1481
with permission. Copyright (1997) Japanese Biochemical Society.)

Figure 8.41 illustrates a hydrogen-bonding network from Asp85 to

Asp96 medicated by the above molecules, a long-range interaction
between Asp85 and Asp96 148]. The low-frequency shifts of the three
water bands in the L minus BR spectrum reveal that all of the water
molecules form stronger hydrogen-bonding upon the L formation. The
most important part includes the water molecule that is linked to
Asp85, Asp212, and the Schiff base, inducing distortion in the retinal.
This structure provides the strong hydrogen-bonding of the protonated
Schiff base. The intervening water molecule may also play an import-
ant role in the separation of the positive change of the protonated Schiff
base and the negative charge on Asp85 [151]. The proton transfer may
become feasible by such a distorted structure by decreasing the pKa
value of the Schiff base.
An intense band at 3486 cm-l is due to an NH stretching mode of
Trp182, showing that its indole ring located in the hydrophobic envi-
ronment has a specific interaction with the 9-CH 3 group of the retinal
chromophore in L [151]. The structural change in Trpl82 favours the
proton transfer from the Schiff base to Asp85 in the L-to-M process.

272
 Upon conversion to M the proton transfer from the Schiff base to
Asp85 occurs, collapsing the strong hydrogen-bonding of the Schiff base
N-H with the protein and directing the lone pair of the nitrogen of the
unprotonated Schiff base to the side opposite from Asp85. In this way,
the distortion in the retinal is abolished. This can be one of the switch
reactions in the unidirectional proton pumping. The high-frequency
shifts of the OH stretching bands in the M minus BR spectrum are
good evidences for the weaker hydrogen-bonding in M.
Similar studies were carried out also for rhodopsin [148]. For this
sort of studies polarized infrared spectroscopy is also powerful because
bacteriorhodopsin is oriented in purple membrane [153]. It was found
that an O-H group in bacteriorhodopsin has an angle of 60 with
respect to the surface normal.
Another interesting study by Kandori et al. [154] is to use an S-H
stretching mode of cysteine as a hydrogen-bonding probe in bacterio-
rhodopsin. An S-H stretching band is located in the 2580-2525 cm-l
region where other vibrations are absent. They substituted threonine
89 (Thr89), which is present in the 'proton channel' of bacterio-
rhodopsin for Cys (T89C) and observed the S-H stretching bands in the
difference spectra. From the S-H stretching frequency, they concluded
that the distance between the sulphur of Cys89 and the oxygen of
Asp85 becomes much closer in K, probably resulting from the chromo-
phore motion in the restricted protein environment [154].

8.7.7 Diagnosis of cervical cancer by infrared

microspectroscopy and artificial neural networks

Papanicolaou (Pap) smears are the current means for the initial screen-
ing of cervical cancer. However, this method is not always highly
accurate, with a reported 20% false-negative rate. The screening pro-
cess involves a microscopic search by naked eyes, which is time-
consuming, fatiguing, and reliant on human judgment. Thus, a more
reliable and automated means of cancer screening has been desired
[155].
It has been shown by several research groups that infrared spectro-
scopy has great potential in the diagnosis of cancer, and in particular
cervical cancer [155,156]. Infrared spectroscopy does not depend upon
morphological observations but directly monitors molecular structure
and changes in cellular chemistry. Thus, it may lead to earlier detec-
tion of abnormalities.

273
 2

1.5

0.5

To

1700 1500 1300 1100 900

Wavenumber (cm-' )

Fig. 8.42. Infrared spectra of cervical cells showing various stages of disease state
(Reproduced from Ref. [155] with permission. Copyright (1998) C.M.B. Association.)

In order to group the infrared spectra of cervical smears into ab-

normal and normal classifications, multivariate statistical analysis or
artificial neural networks (ANN) must be applied to the infrared
spectra. Romeo et al. [155] have demonstrated that ANN has some
advantages over ordinary multivariate statistical analysis in the data
treatment for biomedical materials. The multivariate statistical
analysis depends greatly upon data consistency for accurate perform-
ance while biomedical objects, in particular those exhibiting a disease
state, lack consistency because of variations in symptoms and degrees
of severity of the given disease state. ANN shows better performance
than multivariate statistical analysis probably because non-linear
statistical analysis of the data tolerates considerable amounts of
imprecise or incomplete data [155].
Figure 8.42 compares representative infrared spectra obtained for
samples diagnosed as normal and dysplasia by biopsy [155]. The
spectrum of cultured HeLa cells is also shown in Fig. 8.42. The spectra
consist of contributions from proteins, nucleic acids, lipids and carbo-
hydrates. The 1700-1500 cm-l region is dominated by amide I and II
bands due to the proteins while the 1300-900 cm l region contains
bands due to C-O stretching modes of the proteins and carbohydrates
and those assigned to the antisymmetric and symmetric phosphate
(PO2) vibrations arising from the phosphodiester linkages in nucleic
acids. Bands at 1047 and 1025 cm- ' due to C-O stretching modes of
glycogen and those at 1244 and 1082 cm-l assigned to P0 2 - anti-

274
 PC2 Scores
0.4
n
0.3

0.2

0.1

0O

-0.1-

-0.2-

.0 ., -
-v r.t.1 cores

-1.0 -0.5 6 0.5 1.0

Fig. 8.43. Two principal component scores plot of data bank used to train the neural
network. (Reproduced from Ref. [155] with permission. Copyright (1998) C.M.B.
Association.)

symmetric and symmetric stretching modes, respectively, show

marked changes between the spectra of representative normal and
representative dysplastic.
Infrared spectra of 88 normal and 32 abnormal (mild to severe
dysplasia) cervical smear samples were subjected to principal compo-
nent analysis. Figure 8.43 depicts the resultant two principal
component (PC) scores plot obtained when the infrared data was
reduced to only 7 wavenumber values (1450, 1400, 1244, 1150, 1080,
1050, and 1026 cm- ) [155]. It can be seen from the scores plot that
although most of the samples are separable, there are two obvious
atypical spectral samples of the abnormal set that appear in the normal
group. These were retained in the data base for the ANN analysis.
For the ANN analysis the infrared spectra were randomized to
construct eleven data sets of training, validation, and testing sets each
containing 80, 20, and 20 samples, respectively. Training was carried
out systematically, starting with a simple architecture and gradually
expanding the number of hidden nodes and layers. Table 8.3 sum-
marizes thirteen kinds of ANN architectures that could differentiate
between the normal and abnormal cervical smears [155]. Having
generated these architectures, it was of importance to ascertain which

275
TABLE 8.3
All architecture types used (from Ref. [1551)

Architecture no. Architectural configuration* No. of weights

1 7:1:1 10
2 7:2:1 19
3 7:3:1 28
4 7:4:1 37
5 7:1:1:1 11
6 7:1:2:1 13
7 7:1:3:1 15
8 7:2:1:1 20
9 7:2:2:1 23
10 7:2:3:1 29
11 7:3:1:1 28
12 7:3:2:1 35
13 7:3:3:1 40

*Architectural configuration: number of nodes in input layer:hidden layer/:output layer.

data sets and architectures yield the best learning. For this purpose
twenty samples with known biopsy results and previously unseen by
the networks, were selected and assigned an output according to
normality. This external data set of samples consisted of 10 normal and
10 abnormal samples. Comparison between the expected and actual
outputs for each of the 20 samples in the external testing set was made
to determine the network performance.
A two PC scores plot of the external testing set obtained by archi-
tecture of 7:4:1 is shown in Fig. 8.44 [155]. The plot demonstrates a
separation between the abnormal and normal samples, with samples
19 and 20 (mild dysplasia) closer to the normal/abnormal divide. In
order to increase the ability of the network to make generalizations and
predictions, it is necessary to train the network on hundreds of samples
from each category, ranging from normal, through the three dysplastic
categories to cancerous. This study indicated that infrared spectro-
scopy coupled with ANN may provide an objective and automated
screening technique for the diagnosis of cervical cancer.

276
 II Pr are

0.4 O #20

0.2
1
o0 a) 1
U
0 I, I
1 11
o0 #19
-0.2 1
1

-0.4

-0.8 -0.6 -0.4 -0.2 0 0.2 0.4 0.6

Fig. 8.44. Two principal component scores plot of external testing set. (Reproduced from
Ref. [155] with permission. Copyright (1998) C.M.B. Association.)

REFERENCES

1. C.D. Craver and T. Provder (Eds.), Polymer Characterization-Physical

Property, Spectroscopic, and Chromatographic Methods. Advances in
Chemical Series No. 227, American Chemical Society, 1990.
2. R.S. Stein, J. Appl. Phys., 32 (1961) 1280.
3. W.M. Grim III, J.A. Graham, R.M. Hammaker and W.G. Fateley, Am.
Lab., 16(3) (1984) 22.
4. J.L. Koenig, Spectroscopy of Polymers. American Chemical Society, 1991.
5. N.I. Afanasyeva and R.F. Bruch, Surf. Interface Anal., 27(4) (1999)
204-212.
6. A.N. Theodore and R.O. Carter III, J. Appl. Polym. Sci., 49(6) (1993)
1071-1080.
7. G.L. Grobe III; J.A. Gardella Jr., W.L. Hopson, W.P. McKenna and E.M.
Eyring, J. Biomed. Mater. Res., 21(2) (1987) 211-229.
8. G. Conti and A. Sommazzi, J. Mol. Struct., 294 (1993) 275-278.
9. T. Rische, S. Zschoche and H. Komber, Macromol. Chem. Phys., 199(8)
(1998) 1539-1545.
10. M. Johnck, L. Muller, A. Neyer and J.W. Hofstraat, Eur. Polym. J., 36(6)
(2000) 1251-1264.
11. D. Ostrovskii, A.-L. Kjoniksen, B. Nystroem and L.M. Torell, Macro-
molecules, 32(5) (1999) 1534-1540.
12. D. Das, S. Chattopadhyay, A.K. Barua and R. Banerjee, J. Appl. Phys.,
78(5) (1995), 3193.

277
13. A.Z. El-Sonbati and M.A. Diab, Polym. Degrad. Stab., 22(4) (1988)
295-302.
14. V. Vettegren, V. Teubner-Texte Phys. 9 (Prog. Polym. Spectrosc.), (1986),
158-66.
15. M.W. Urban and C.D. Craver (Eds.), Structure-Property Relations in
Polymers: Spectroscopy and Performance. Advances in Chemical Series
No. 236, American Chemical Society, 1993.
16. C. Qin, A.T.N. Pires and L.A. Belfiore, Polym. Commun., 31(5) (1990)
177-182.
17. L.O. Faria and R.L. Moreira, J. Polym. Sci. B, 38 (2000) 34-40.
18. H.W. Siesler and K. Holland-Moritz, Infrared and Raman Spectroscopy of
High Polymers. Marcel Dekker Inc., New York, NY, 1980, pp. 243-292
19. R. Zbiden, Infrared Spectroscopy of High Polymers. Academic Press, New
York, NY, 1964.
20. S. Krim, J. Phys. Chem., 32 (1960) 1780.
21. S. Krim,Adv. Polym. Sci., 2 (1960) 51.
22. L. Monnerie, FaradaySymp. Chem. Soc., 18 (1983) 57.
23. T. Thormann, M. Rogojerov, B. Jordanov and E.W. Thulstrup, J. Mol.
Struct., 509(1-3) (1999) 93-104.
24. F.R. Steenstrup, K. Christensen, C. Svane and E.W. Thulstrup, J. Mol.
Struct., 408 (1997) 139-148.
25. V.G. Gregoriou and R.A. Palmer, Macromol. Symp., 94 (1995) 75-95.
26. C.J. Manning, J.L. Chao and R.A. Palmer, Rev. Sci. Instrum., 62 (1991)
1219.
27. I. Noda, A.E. Dowrey and C. Marcott, J. Polym. Sci. Polym. Lett. Ed., 21
(1983) 99.
28. I. Noda, A.E. Dowrey and C. Marcott, Polym. Prep. (Am. Chem. Soc., Div.
Polym. Chem.), 24(1) (1983) 122.
29. I. Noda, A.E. Dowrey and C. Marcott, Polym. Prep. (Am. Chem. Soc., Div.
Polym. Chem.), 25(2) (1984) 167.
30. I. Noda, A.E. Dowrey and C. Marcott, Appl. Spectrosc. 42, 203 (1988).
31. A.W. Mantz, Appl. Spectrosc., 30 (1976) 459; A.A. Garrison, R.A.
Crocombe, G. Mamantov and J.A. de Haseth, Appl. Spectrosc., 34 (1980)
399.
32. P.R. Griffiths and J.A. de Haseth, Fourier Transform Infrared
Spectroscopy. Wiley, New York, NY, 1986, pp. 407-425.
33. V.G. Gregoriou, C. Marcott, I. Noda, A. Dowery and R.A. Palmer, J. Polym.
Sci.-B Polym. Phys., 31 (1993) 1769.
34. F. Ozanam and Chazalviel, Rev. sci. Instrum., 59(2) (1988) 242.
35. R.A. Palmer, J.L. Chao, R.M. Dittmar, V.G. Gregoriou and S.E. Plunkett,
Appl. Spectrosc., 47(9) (1993) 1297-310.
36. M. Hashimoto, T. Yuzawa and H. Hamaguchi, Jasco Rep., 37(3) (1995)
27-33.

278
37. G. Ellis, C. Marco, J. del Pino, J. Lorente, M.A. Gomez and J.G. Fatou, Vib.
Spectrosc., 9(1) (1995) 49-56.
38. A. Hatta, Mol. Crys. Liq. Cryst., 74 (1981) 195.
39. M.L. Forman, W.H. Steel and G.A. Vanasse, J. Opt. Soc. Am., 56 (1966) 59.
40. V.G. Gregoriou, J.L. Chao, H. Toriumi and R.A. Palmer, Chem. Phys. Lett.,
179 (1991) 491.
41. H.J. Coles and J. Tipping, Nature 316 (1986) 136; Mol. Cryst. Liq. Cryst.
Lett., 2 (1985) 23.
42. A. Kaito, Y.K. Wang and S.L. Hsu, Anal. Chim. Acta, 189 (1986) 27.
43. H. Toriumi, H. Sugisana and H. Watanabe, Jpn. J. Appl. Phys., 27 (1988)
L935.
44. V.G. Gregoriou, J.L. Chao, H. Toriumi and R.A. Palmer, Chem. Phys. Lett.,
179 (1991) 491.
45. H.W. Siesler, Adv. Polym. Sci., 65 (1984), 1-77
46. M.W. Urban and T. Provder (Eds.) Multidimensional Spectroscopy of Poly-
mers. ACS Symposium Series No. 598, American Chemical Society, 1995.
47. I. Noda, Chemtracts, Macromol. Chem., 1 (1990) 89.
48. I. Noda, A.E. Dowrey and C. Marcott, J. Polym. Sci. Polym. Lett. Ed., 21
(1983) 99.
49. R.A. Palmer, R.M. Dittmar, V.G. Gregoriou and J.L. Chao, Polym. Prepr.,
32 (1991) 673.
50. I. Noda, A.E. Dowrey and C. Marcott, J. Mol. Struct., 224 (1990) 265.
51. D. Lefebvre, B. Jasse and L. Monnerie, Polymer, 22 (1981) 1616.
52. B.R. Nair, V.G. Gregoriou and P.T. Hammond, Polymer, 41 (2000) 2961.
53. B.R. Nair, V.G. Gregoriou and P.T. Hammond, J. Phys. Chem. B., 104
(2000) 7874.
54. F. Papadimitrakopoulos, K. Konstadinidis, T.M. Miller, R. Opila, E.A.
Chandross and M.E. Galvin, Chem. Mater., 6(9) (1994) 1563-1568.
55. D. Muller, M. Gross, K. Meerholz, T. Braig, M.S. Bayerl, F. Bielefeldt and
O. Nuyken, Synth. Met., 111/112 (2000) 31-34.
56. A. Bacher, C.H. Erdelen, W. Paulus, H. Ringsdorf, H.-W. Schmidt and P.
Schuhmacher, Macromolecules, 32(14) (1999) 4551-4557.
57. J.C. Scott, S.A. Carter, S. Karg and M. Angelopoulos, Proc. SPIE-Int. Soc.
Opt. Eng., 3002 (1997) 86-91.
58. J. Davenas, V. Massardier and V.H. Tran, Nucl. Instrum. Meth. Phys.
Res., Sect. B, 112 (1996) 120-124.
59. Y. Furukawa, M. Tasumi and G. Zerbi (Eds.), Mod. Polym. Spectrosc.
(1999), 207-237.
60. H. Ericson, C. Svanberg, A. Brodin, A.M. Grillone, S. Panero, B. Scrosati
and P. Jacobsson, Electrochim. Acta, 45(8/9) (2000) 1409-1414.
61. Y. Furukawa, J. Phys. Chem., 100(39) (1996) 15644-15653.
62. K. Saravanamuttu, X.M. Du, S.I. Najafi and M.P. Andrews, Can. J.
Chem., 76(11) (1998) 1717-1729.

279
63. D.L. Thomsen III, J. Mwaura and F. Papadimitrakopoulos, Polym. Prepr.
(Am. Chem. Soc., Div. Polym. Chem.), 39(2) (1998) 1120-1121.
64. F. Papadimitrakopoulos, P. Wisniecki and D.E. Bhagwagar, Chem.
Mater., 9(12) (1997) 2928-2933.
65. T. Ubukata, T. Seki, S. Morino and K. Ichimura, J. Phys. Chem. B, 104(17)
(2000) 4148-4154.
66. J. Wang, B. Zou, X. Hong and D.M. Collard, Synth. Met., 113(3) (2000)
223-226
67. M. Blazewicz, M.C. Gajewska and C. Paluszkiewicz, J. Mol. Struct.,
482-483 (1999) 519-524.
68. J.M. Pope, T. Sotomura and N. Oyama, Batteries for Portable Applications
and Electric Vehicles. Proc. Electrochem. Soc., 97-118 (1997) 116-123.
69. P.W. Faguy, W. Ma, A.J. Lowe, W.P. Pan and T. Brown, J. Mater. Chem.,
4(5) (1994) 771-772.
70. J. Mijovic, S. Andjelic, B. Fitz, W. Zurawsky, I. Mondragon, F. Bellucci and
L. Nicolais, J. Polym. Sci., PartB: Polym. Phys., 34(2) (1996) 379-388.
71. D.L. Wetzel and S.M. LeVine, Science, 285(5431) (1999) 1224-1225.
72. M.C. McCann, L. Chen, K. Roberts, E.K. Kemsley, C. Sene, N.C. Carpita,
N.J. Stacey and R.H. Wilson, Physiol. Plant. 100(3) (1997) 729-738.
73. L.H. Kidder, I.W. Levin and E.N. Lewis, Fourier Transform Spectroscopy.
AIP Conf. Proc., 430 (1998) 148-158.
74. Max-Planck-Institut fur Biochemie. Nature (London), 399(6732) (1999)
134-137.
75. E.N. Lewis and I.W. Levin, Appl. Spectrosc., 49(5) (1995) 672-8.
76. R. Bhargava, T. Ribar and J.L. Koenig, Appl. Spectrosc., 53(11) (1999)
1313-1322.
77. K. Artyushkova, B. Wall, J. Koenig and J.E. Fulghum, Appl Spectrosc.,
54(11) (2000) 1549-1558.
78. S. Bohic, D. Heymann, J.A. Pouezat, O. Gauthier and G. Daculsi, C. R.
Acad. Sci., Ser. III, 321(10) (1998) 865-876; R. Mendelsohn, E.P.
Paschato, P.J. Sherman, A.L. Boskey, Appl. Spectrosc., 54 (2000)
1183-1191.
79. C.M. Snively and J.L. Koenig, J. Polym. Sci., PartB: Polym. Phys., 37(17)
(1999) 2353-2359.
80. W.J. Marinelli, C.M. Gittins, A.H. Gelb, B.D. Green, Appl. Opt., 38(12)
(1999) 2594-2604.
81. C. Marcott, R.C. Reeder, J.A. Sweat, D.D. Panzer and D.L. Wetzel, Vib.
Spectrosc., 19(1) (1999) 123-129.
82. R. Bhargava, D.C. Fernandez, M.D. Schaeberle and I.W. Levin, Appl.
Spectrosc., 53(12) (2001) in press.
83. J.-L. Bantignies, G. Fuchs, G.L. Carr, G.P. Williams, D. Lutz and S.
Marull, Int. J. Cosmet. Sci., 20(6) (1998) 381-394.
84. H. Fabian, R. Wessel, M. Jackson, A. Schwartz, P. Lasch, I. Fichtner, H.H.

280
 Mantsch, D. Naumann, Infrared Spectroscopy: New Tool in Medicine.
Proc. SPIE-Int. Soc. Opt. Eng., 3257 (1998) 13-23.
85. N. Jamin, P. Dumas, J. Moncuit, W.-H. Fridman, J.-L. Teillaud, G.L. Carr
and G.P. Williams, Proc. Natl. Acad. Sci. USA, 95(9) (1998) 4837-4840.
86. P. Lasch and D. Naumann, Cell. Mol. Biol. (Paris), 44 (1998) 189.
87. M.K. Weldon, V.E. Marsico, Y.J. Chabal, A. Agarwal, D.J. Eaglesham, J.
Sapjeta, W.L. Brown, D.C. Jacobson, Y. Caudano, S.B. Christman and
E.E. Chaban, Semiconductor Wafer Bonding: Science, Technology, and
Applications. Proc. Electrochem. Soc., 97-36 (1998) 229-248.
88. L.E. Ocola, F. Cerrina and T. May, J. Vac. Sci. Technol., B, 15(6), (1997)
2545-2549.
89. L.H. Kidder, I.W. Levin, E.N. Lewis, V.D. Kleiman and E.J. Heilweil, Opt.
Lett., 22(10) (1997) 742-744.
90. E.N. Lewis, L.H. Kidder, J.F. Arens, M.C. Peck and I.W. Levin, Appl.
Spectrosc., 51(4) (1997) 563-567.
91. M. Blanco, J. Coello, H. Iturriaga, S. Maspoch and E. Bertran, Appl.
Spectrosc., 49(6) (1995) 747-753.
92. E. Herrala, P. Niemela and T. Hannula, In-Process Opt. Meas. Ind.
Methods. Proc. SPIE-Int. Soc. Opt. Eng., 1266 (1990) 86-90.
93. N.A. Wright, R. Curbelo, D.A.C. Compton and S.L. Hill, Infrared Fiber
Opt. Proc. SPIE-Int. Soc. Opt. Eng., 1048 (1989) 153-160.
94. J.S. Sanghera and I.D. Aggarwal, J. Non-Cryst. Solids, 256/257 (1999)
6-16.
95. A. Ulman, An Introduction to Ultrathin OrganicFilms. Academic Press,
New York, 1991.
96. G.G. Roberts, Langmuir-Blodgett Films. Plenum Press, New York, 1990.
97. T. Nakamura, Handbook of Organic Conductive Molecules and Polymers,
Vol. 1. (H.S. Nalwa, Ed.). Wiley, Chichester, 1997, p. 727.
98. T. Takenaka and J. Uemura, in: J.R. During (Ed.), Vibrational Spectra
and Structure, Vol. 19. Elsevier, Amsterdam, 1991, p. 215.
99. J.D. Swalen, Annu. Rev. Mater. Sci., 21 (1991) 373.
100. T.G. Greenler, J. Chem. Phys., 44 (1969) 310.
101. P.-A. Chollet and J. Messier, Thin Solid Films, 99 (1983) 197.
102. J. Umemura, T. Kamata, T. Kawai and T. Takenaka, J. Phys. Chem., 94
(1990) 62.
103. R.G. Snyder, J. Mol. Spectrosc., 7 (1961) 116.
104. M. Tasumi and T. Shimanouchi, J. Chem. Phys., 43 (1965) 1245.
105. J. Umemura, D.G. Cameron and H.H. Mantsch, Biochim. Biophys. Acta,
602 (1980) 32.
106. H. Sapper, D.G. Cameron and H.H. Mantsch, Can. J. Chem., 59 (1981)
2543.
107. M. Kubota, Y. Ozaki, T. Araki, S. Ohki and K. Iriyama, Langmuir, 7 (1991)
774.

281
108. S. Morita, K. Iriyama and Y. Ozaki, J. Phys. Chem., B, 104 (2000) 1183.
109. J. Umemura, T. Kamata, T. Kawai and T. Takenaka, J. Phys. Chem., 94
(1990) 62.
110. S. Terashita, Y. Ozaki and K. Iriyama, J. Phys. Chem., 97 (1993) 10445.
111. Y. Wang, K. Nichogi, K. Iriyama and Y. Ozaki, J. Phys. Chem., 100 (1996)
368.
112. Y. Wang, K. Nichogi, K. Iriyama and Y. Ozaki, J. Phys. Chem., 100 (1996)
374.
113. S. Morita, K. Iriyama and Y. Ozaki, J. Phys. Chem. submitted.
114. K. Ikegami, M. Lan and T. Nakamura, J. Chem. Phys., 112 (2000) 881.
115. G. Lunardi and C. Pecile, J. Chem. Phys., 95 (1991) 6911.
116. T. Hasegawa, Y. Hatada, J. Nishijo, J. Umemura, Q. Hao and R.M.
Leblanc, J. Phys. Chem., B, 103 (1999) 7505.
117. W.W. Coblenz, Bull. Natl. Bur. Stand. (US), 7 (1911) 619.
118. K.P. Norris and J.E.S. Greenstreet, J. Gen. Microbiol., 19 (1958) 566.
119. F.S. Parker, Application of Infrared Spectroscopy in Biochemistry,
Biology, and Medicine. Plenum Press, New York, NY, 1971.
120. F.S. Parker, Application of Infrared, Raman, and Resonance Raman
Spectroscopy in Biochemistry. Plenum Press, New York, NY, 1983.
121. R.E. Hester and R.B. Girling (Eds.), Spectroscopy of Biological Molecules.
Royal Society of Chemistry, London, 1991.
122. H.H. Mantsch and D. Chapman (Eds.), Infrared Spectroscopy of Bio-
molecules. Wiley, New York, 1996.
123. S. Krimm and J. Bandekar, Adv. Protein Chem., 38 (1986) 181.
124. H. Susi and D.M. Byler, Arch. Biochem. Biophys., 258 (1987) 465.
125. W. Mantele, Trends Biochem. Sci., 18 (1993) 197.
126. R.J.H. Clark and R.E. Hester (Eds.), Biomolecular Spectroscopy, PartA
and PartB. Wiley, Chichester, 1993.
127. F. Siebert, in: H.H. Mantsch and D. Chapman (Eds.), Infrared Spectro-
scopy of Biomolecules. Wiley, New York, 1996, p. 83.
128. M. Jackson and H.H. Mantsch, in: H.H. Mantsch and D. Chapman (Eds.),
Infrared Spectroscopy of Biomolecules. Wiley, New York, 1996, p. 311.
129. D. Naumann, D. Helm and C. Schulz, in: F.G. Priest, A. Ramos-
Cormenzana and B.J. Tindall (Eds), BacterialDiversity and Systematics.
Plenum, New York, 1994, p. 67.
130. P.I. Haris and D. Chapman, in: H.H. Mantsch and D. Chapman (Eds.),
InfraredSpectroscopy of Biomolecules. Wiley, New York, 1996, p. 239.
131. M. Jackson and H.H. Mantsch, CRC Crit. Rev. Biochem. Mol. Biol., 30
(1995) 95.
132. Y. Ozaki and K. Murayama, in: Infrared and Raman Spectroscopy of
Biological Materials. Marcel Dekker, New York, 2000, p. 515.
133. Y. Ozaki and I. Noda, in: R.A. Meyers (Ed.), Encyclopedia of Analytical
Chemistry: InstrumentationandApplications. Wiley, Chichester, 2000, p. 322.

282
134. J.M. Hadden, M. Bloemendal, P.I. Haris, I.H.M. Stokkum, D. Chapman
and S.K.S. Srai, FEBS Lett., 350 (1994) 235.
135. H. Torii and M. Tasumi, in: H.H. Mantsch and D. Chapman (Eds.),
Infrared Spectroscopy of Biomolecules. Wiley, New York, 1996, p. 1.
136. K. Ballschmiter and J.J. Katz, J. Am. Chem. Soc., 91 (1969) 2661.
137. K. Okada, K. Uehara and Y. Ozaki, Photochem. Photobiol., 57 (1993) 958.
138. H. Scheer (Ed.), Chlorophylls. CRC Press, Boca Raton, FL, 1991, p. 1097.
139. M. Tasumi and M. Fujiwara, in: R.J.H. Clark and R.E. Hester (Eds.),
Advances in Spectroscopy, Vol. 14. Wiley, Chichester, 1987, p. 407.
140. M. Lutz and W. Mantele, in Ref. [138], p. 855.
141. K. Uehara, T. Tachibana, M. Tsunooka and Y. Ozaki, Photochem.
Phytobiol., 62 (1995) 496.
142. D.G. Cameron. H.L. Casel, H.H. Mantsch, Y. Boulanger and I.C.P. Smith,
Biophys. J., 35 (1981) 1.
143. D.G. Cameron, A. Martin, D.J. Moffatt and H.H. Mantsch, Biochemistry,
24 (1985) 4355.
144. D. Naumann, C.P. Schultz and D. Helm, in: H.H. Mantch and D. Chapman
(Eds.), Infrared Spectroscopy of Biomolecules. Wiley, New York, 1996, p.
279.
145. CellularMol. Biol. Special issue for infrared microbiology (1998).
146. M.D. Schaeberle, I.W. Levin and E.N. Lewis, in: H.-U. Gremlich and B.
Yan (Eds.), Infrared and Raman Spectroscopy of Biological Materials.
Marcel Dekker, New York, 2000, p. 231.
147. E. Nabedryk, in: H.H. Mantsch and D. Chapman (Eds.), InfraredSpectro-
scopy of Biomolecules. Wiley, New York, 1996, p. 39.
148. A. Maeda, H. Kandori, Y. Yamazaki, S. Nishimura, M. Hatanaka, Y.-S.
Chon, J. Sasaki, R. Needleman and J.K. Lanyi, J. Biochem., 121 (1997)
399.
149. A. Maeda, J. Sasaki, Y. Shichida and T. Yoshizawa, Biochemistry 31
(1993) 462.
150. Y. Yamazaki, S. Tuzi, H. Saito, H. Kandori, R. Needleman, J.K. Lanyi and
A. Maeda, Biochemistry, 35 (1996) 4063.
151. S. Scheiner and X. Duan, Biophys. J., 60 (1991) 874.
152. Y. Yamazaki, J. Sasaki, M. Hatanaka, H. Kandori, A. Maeda, R.
Needleman, T. Shinada, K. Yoshihara, L.S. Brown and J.K. Lanyi, Bio-
chemistry 34 (1995) 577.
153. H. Kandori, N. Kinoshita, Y. Shichida and A. Maeda, J. Phys. Chem. B,
102 (1998) 7899.
154. H. Kandori, N. Kinoshita, Y. Shichida, A. Maeda, R. Needleman and J.K.
Lanyi, J. Am. Chem. Soc., 120 (1998) 5828.
155. M. Romeo, F. Burden, M. Quinn, B. Wood and D. McNaughton, Cellular
Mol. Biol., 44 (1999)179.
156. P.T.T. Wong, R.K. Wong and K. Fung, Appl. Spectrosc., 47 (1993) 1058.

283
Chapter 9

Modern data analytical methods for

infrared spectroscopy

Infrared spectrometry is the most universally useful of all analytical

techniques [1] available for material characterisation. Quantitative
analysis using infrared spectrometry involves the measurement of the
infrared spectrum first. There are several different sampling
techniques available for this purpose depending on the availability and
physical state of the sample. Development in infrared instrumentation
and analytical methods such as diffuse reflectance, total internal
reflectance, specularreflectance, photo acoustic, GC-IR, LC-IR, infrared
microspectrometry, etc. have made this possible.
The mid-infrared profiles arise from the intensities of absorption of
the fundamental frequencies and describe chemical structure, which
are indirectly related to the physical and chemical properties of the
systems analysed.
When a sample is measured, the absorbance of a sample or a
functionality measured at a particular wavelength is proportional to
the concentration of the sample or the functionality measured (Beers
law). The traditional quantitative analysis in infrared spectrometry is,
in general, based on this relationship.

9.1 UNIVARIATE APPROACH IN INFRARED SPECTROSCOPY

As mentioned above, the success of quantitative analysis using the

univariate approach in infrared spectrometry is based on the relation-
ship between absorbance and concentration of the sample. A precise
quantitative determination using univariate technique is possible in
cases where one can isolate an absorption arising from a functional
group and relate it entirely to the concentration of the sample. The

285
accuracy of the analysis increases with the increase in the proportion-
ality constant (molar absorptivity) of the functionality at that wave-
number.
Most textbooks on infrared spectrometry deal with quantitative
analysis using univariatetechnique and the approach to the technique
can be found elsewhere.

9.2 MULTIVARIATE APPROACH IN INFRARED SPECTROSCOPY

9.2.1 The need for multivariate approach and types of

multivariate systems

If the infrared spectra obtained in chemical systems, whether static or

dynamic, contain an isolated non-overlapping absorption of a partic-
ular functional group that it is necessary to quantify then one variable
approach will be successful. However, in mixtures where the absorp-
tions arise from contributions from different functional groups, the
quantification of a particular functional group using the one-variable
approach becomes difficult and sometimes impossible. Quantitative
analysis using infrared spectra produced by these types of systems
requires some intelligent procedures.
In many industrial applications measurements are made on systems
containing several different chemical components, for example, on calib-
ration standard mixtures of components for quantitative prediction of
components in unknown samples. The infrared spectra measured on the
calibration mixtures will contain contributions from the components in
the mixtures. The spectral profiles vary with the quantities of different
components present. A procedure that can model the spectral variations
with the change in the quantities of components and predict the compo-
nents quantitatively in unknown samples could reduce the analysis
burden on industrial laboratories. The separation of chemical compo-
nents using other analytical techniques and quantification of these
components individually increases the cost of quality control.
A chemical system can be dynamic or static. For example a reaction
that is studied by using infrared spectroscopy can produce several
hundred spectra over a period of time showing the changes taking place
in the system. These spectra represent the total spectra of the
reactants and products in the mixture. Likewise, a chemical system
may be static and contain several components. These components can

286
change in concentration in the system. For example, a packaging
laminate contains several layers of polymer material. The concen-
trations of the components change across the depth of the laminate due
to the inter-diffusion of the components. An infrared analysis across
the depth of the layer will produce spectra that reflect the concentra-
tion of the components present at a particular depth.
In certain applications one might need to characterise samples by
using similarity of their spectral profiles in order to classify them into
different classes. In these cases one needs tools that can compare the
whole infrared profiles of the samples measured.
If one is interested in studying these systems using infrared spect-
rometry then the use of procedures suitable for resolving the spectra
into their contributing components and their concentrations are
needed. The procedures and techniques required to solve problems
involving most of the above types of systems involve the handling of
matrices with several thousands of entries. These put very high dem-
and on the capacity of the computers. The past decade has witnessed
rapid growth in both computer hardware capacity and the development
of new methodologies for the resolution of analytical profiles and
quantification of mixtures.
Statistical procedures that can be used in applications of the types
mentioned above are widely available from several vendors. We will not
discuss any particular multivariate package in general because all
these packages are based on the same mathematical and statistical
procedures.
In order to understand the approach used in handling the tasks
mentioned above, a theoretical treatment of the relevant chemometric
techniques is given in Sections 9.2.3 to 9.2.4.

9.2.2 Presentation of data

In most chemometric techniques data are presented in the form of

matrices so that the software can handle them and process the model-
ling needed in particular applications. In mid-infrared spectroscopy,
the spectral profiles are presented as absorbances (or percentage
transmittances) against wavenumbers. It is obvious that one should
use wavenumbers as the variables. For example, if a spectrum is
measured over m wavenumbers then the spectrum can be represented
by a lxm matrix. Here, the matrix elements are absorbances at
different wavenumbers (Fig. 9.1).

287
 The absorbances can be represented as a vector and it is customary
to give all the vectors as column vectors in multivariate data analysis.
A column vector x' can represent the data from the above object. The
prime indicates the transpose of the vector x.
If this spectral profile is going to be correlated to a property y
(dependent variable or response variable) then the measure of the
property is represented by a single value y. The variables x1 i are called
independent variables (or predictor variables). This can be extended to
several measurements n (objects). As shown above, each measurement
can be represented as follows as a row in a matrix of dimension nxm.

Y1 Xll X12 X1 3 X1 4 Xlm

Y2 X2 1 X2 2 X2 3 X2 4 X2m

Y3 X31 X3m

Y4

Yn - Xnl Xn2 Xn3 Xn4 Xnm

The matrix representing the dependent variables contain n entries (n

rows). Likewise the spectral profiles matrix contains n rows each
representing absorbances over m variables. The whole set of dependent
variables can be represented by a column vector y. The independent
variables from the n objects can be represented by a matrix X
containing n rows and m columns. In our general notation, the data
matrix X is a matrix with object vectors (Xk'; k = 1, 2...m).
If a relationship between the x variables and y variables is desired,
one sets up a mathematical equation of the type shown below.

y=f(x) +E (9.1)

y=Xb +E (9.2)

288
 -

F -

f0 a, bsorbance

4000 3000 i 2000 loo1000 400

Wavenumbers cm

I[XI X12 X13 X14 . . Xlm

Fig. 9.1. An example of an infrared absorption spectrum and the presentation of

absorbances in a row matrix.

Here, b is an mx 1 matrix containing m coefficients and E is the residual

matrix of nxl.

Y1 X11 X1 2 X 13 X1 4 . . . . . X b el
Y2 X21 X2 2 X 23 X24 ..... X2m b2 e2

Y X3 1 .. . X3m b3 e3

Y4

Yn Xnl Xn2 xn3 Xn4 n.....x en

The solution for b can be found mathematically as follows.

b =X-y (9.3)

X -1 is the inverse of the data matrix X.

289
There are three possible cases from the above representations:
1. When the number of samples and variables are equal (n = m) then
there is a unique solution for b provided that X has a full rank.
2. If the number of samples is less than the number ofvariables (n < m)
then there are infinite number of solutions for b.
3. If the number of samples is more than the number of variables (n >
m) then a solution for b can be obtained by multiple linear
regression (minimising the length of the residual vector E (/el )).
The solution (Eq. (9.6)) is obtained by forming the generalised
inverse of X.

XTy =XrXb (9.4)

T
(X)-'X y = (XTx)-XTX b (9.5)

b = (XX)-X T y (9.6)

We have to remember at this stage that only non-singular (the

determinant is non-zero) square matrices have inverses. The technique
shown above in multiple linear regression is to make the data matrix X
into a square matrix (nxn matrix) by multiplying by its transpose X .
However, the inverse of this product matrix can exist only when the
resulting matrix is non-singular (or has full rank; see appendix).
When the measurements are made in infrared spectroscopy, it can
be seen that there is a lot of redundant information in the data. It
means that there is 'collinearity'in the data profiles and the underlying
variables (latent variables) responsible for the variation in the data are
fewer than m. Modern multivariate techniques such as principal
component analysis (PCA) and partial least squares calibration (PLS)
are developed to handle data with such problems.
In multivariate data techniques, the aim is to decompose the data
matrix into the product of two matrices that can give us information on
variables and objects. The criterion for the decomposition varies
depending on the type of model one is trying to build with the data
measured.

290
 ... k01J
Fig. 9.2. Decomposition of the data matrix X into score matrix T and loading vector
matrix P. The figure illustrates the data matrix containing five measurements reduced to
the product of three score vectors and three loading vectors. Dimensionality of the
matrices X, TPT and E are the same.

9.2.3 The data matrix X and its decomposition

The data matrix X can be written as the sum of A matrices of rank 1

(Eq. (9.7)) and a residual matrix E.

X=X 1 +X 2 +X3+ ... +XA+E (9.7)

The decomposed matrices can be written as the products of score vector

ta and a loading vector pa (Eq. (9.8)).

X = tlpl'+t2P2' + t3' ... + tAPA' + E (9.8)

Equation (9.8) is illustrated in Fig. 9.2.

X = TPT +E (9.9)

9.2.4 Graphic representation of spectral profiles

The data matrix containing the spectral profiles of n objects and m

variables can be viewed in two different ways (1) objects in variable
space and (2) variables in object space. In the first case the objects (n)
are represented as data points in M dimensional (m orthogonal
dimensions) variable space. In the second case the variables (m) are
represented as data points in n dimensional (n orthogonal dimensions)
object space. Graphic representations of more than three dimensions
are impossible on paper. A plot obtained for the objects in the variable
space displays quantitatively the relationships between the objects,
and the variables plotted in the object space displays the relationships

291
between the variables. All the information contained in the data matrix
can be explained by these two plots.

I Column vector (variable vector)

Row vector (object vector) -> X1 1 X1 2 X 13 X1 4 . . . Xm I

X2 1 X2 2 X23 X24 .. X2m

X3 1 . . . . . X3m

Xnl X2 X,3 Xn4 .... . Xnm

9.3 PRINCIPAL COMPONENT ANALYSIS (PCA)

Principal component analysis is one of the several multivariate

techniques to decompose the total data matrix into data matrices
containing information regarding the system (objects and variables) we
are dealing with. The criterion used for the decomposition of the data
matrix X is to extract latent variables in the direction of maximum
variance in the data; that is, the first latent variable-the first princi-
pal component (PC 1)-is a linear combination of the original variables
that explain the largest variance in the data X. The information
explained by this latent variable is then removed from the data matrix.
The second latent variable-the second principal component (PC2)-is
then extracted in the direction that explains maximum variance in the
rest of the data. These two principal components are orthogonal to each
other. The decomposition of the data matrix X into principal compo-
nents represents an ordinary least-squares solution which minimises
the residuals E. One can extract several principal components to
explain as much as possible variance in the data. The sum of the
residuals goes down with the number of principal components
extracted and reaches a minimum and then increases with the

292
extraction of more principal components (overfitting) [2]. The number
of principal components needed to explain the information in the data
set could be calculated by a process called cross-validation [2].
If the data matrix is column centred (i.e., each variable is adjusted
in relation to the average of the variables), the origin of the variable
space moves to the point represented by the averages of the variables in
the data matrix. The principal components then pass through this
point.
We understand that the principal components are linear combi-
nations of the M original variables. The projections of objects on the
principal components (latent variables) are called scores. The scores of
the objects on the principal components can be calculated by the scalar
product between the unit vector (Wa) along the respective principal
component and the object vector.

ta =Xwa (9.10)

All these scores are extracted in the score matrix T. The scores describe
the similarities between the objects. These scores can be presented in
the form of a projection plot on the plane containing PC1 and PC2, or
PC2 and PC3, or PC1 and PC3. These plots are called score plots (Fig.
9.3). The samples that are similar group together in the score plots. The
samples that are atypical to the other samples in the set will be isolated

X2

xlCos P

PC1

Fig. 9.3. Scores are obtained by projecting object vectors onto the principal components.

293
in the score plots and can be easily identified. These samples are called
'outliers'.
The interpretation of a single latent variable and of the features of a
latent variable model is possible through the connection to the original
variables. This information is best displayed in loading plots. The co-
ordinates of a variable in a loading plot are its loadings on the principal
components. A loading plot displays the contribution of a variable to a
model directly (proportional to the square of the distance from the
origin). Variables that are lying near the origin contribute little and
variables that are lying far away from the origin contribute more in the
discrimination of the samples. Variables located in the same direction
of a principal component carry similar information.
Scores and loadings can be displayed simultaneously in plots called
biplots [3]. These plots give information on the similarity between the
samples and the variables that are responsible for the discrimination of
the samples.

9.4 MULTIVARIATE CALIBRATION

9.4.1 Partial least-squares (PLS) calibration

In many applications, one wants to relate some measured property y to

a spectral profile x of intensities. The task is performed by measuring
several pairs of(y, x) and subsequently constructing a model such that
y=fAx) (9.11)
The model is commonly calculated using least-squares procedures
under the constraint of linear models. When there is only one property
to be correlated with the spectral profile the PLS model building
process is called PLS1, and PLS2 when there are more than two. In
PLS1, the data is modelled in two stages, namely compression and
calibration. In the data compression stage the spectral data are
modelled in terms of a set of common latent regression factors {tl, t 2, t3
... tA) (scores). These are n dimensional orthogonal column vectors for a
data set with n samples and m spectral variables. These factors
describe the major variations in the spectral data and at the same time
are relevant for predicting the dependent variable. The compression
and calibration steps can be written as follows for column centred
spectral and response variables (see Fig. 9.4):

294
 x I UI r I

Fig. 9.4. A figure showing the decomposition of a data matrix X in principal component
analysis and partial least-squares calibration. The details regarding the decomposition
in these techniques are given in the text.

X= t 1 +t 2 + t3p 3 ...
' + tAPA' +E (9.12)

y = tlql+tq 2+t 3 q3 ... + tAqA +F (9.13)

In the PLS1 algorithm, the vectors t, p and q are calculated starting

with an equation relating the spectral data and dependent variable.
X=yw' +E' (9.14)
wl is a column vector (weightings) and is estimated as follows:

w 1'=y'X/ y'X (9.15)

w, is then normalised to unity. The estimated w, is then used to

calculate t, Pl and q1 [4]. These estimates are again used to calculate
the residuals of spectral and dependent variables. The process is
repeated untilA factors which minimise the prediction error are found.
The prediction of an unknown sample from the spectral profile xi
proceeds as follows. First score t is found using the estimate for w1 and
equation

Xi = tl w' + el (9.16)

Then the next score t2 is calculated from the solution of the above
equation and the estimate for w2. The process is repeated until the Ath
factor. The dependent variable is predicted by

y = tq, + t2 q 2 + t3 q 3... + tAqA (9.17)

295
Alternatively the same prediction can be written as

y =Xb (9.18)

In PLS calibration a generalized inverse X+ is constructed as

b =X'y (9.19)

and the calibration coefficients vector b is determined by Eq. (9.20) [5]

b = W'(PW')-1 q (9.20)

The parameters b are estimated so as to predict the property y as well

as possible, for instance by means of cross validation [2]. By use of the
parameters b the propertyy can be predicted from the infrared profiles.
The predicted values of the property y for the training set samples is
obtained by inserting Eq. (9.19) into Eq. (9.18):

y =XXy (9.21)

If there is a strong relationship between the property y and the intens-

ities at some wavelengths, the predicted and the measured property y
will be similar and the correlation coefficient between measured and
predicted y will approach 1.

9.4.2 An application of multivariate calibration (PLS):

determination of coal maturity (rank)

Maturity or coal rank is used extensively by geochemists to charac-

terise coal and kerogen samples [6]. Several methods have been used to
determine maturity, including vitrinite reflectance, spore colour esti-
mation, isotope ratios and chemical analysis such as biomarkers [7].
Coal consists mainly of three types of macerals: vitrinite, exinite,
and inertinite. There is a correspondence between these three maceral
groups and the chemical composition. Coal petrologists and chemists
have linked the coal rank with maceral reflectance measurements [8].
Vitrinite reflectance is the most widely used method for maturity
determinations. However, the analysis is time consuming and sub-
jective [9]. Additionally, vitrinite is only one of the organic components
in the sample and is not the major oil precursor. Assessing maturity by

296
measuring the major oil precursors or determining the chemical
composition of geological specimens should provide more meaningful
and accurate characterisation for petroleum geological purposes.
Fourier-transform infrared spectroscopy was early taken into use
for determination of maturity of kerogen [10-11] and coal [12]. The
black or dark brown colour of the samples made it very difficult to
analyse the samples by traditional transmission techniques. The dif-
fuse reflectance technique then became popular for studying coal rank
[13-14]. In these studies, specific functional group regions assigned to
aromatic, aliphatic and C-H bending and stretching modes were
correlated with chemical C/H ratios and rank. Fredericks et al. [15]
used factor analysis to correlate specific parts of the FTIR spectra with
various chemical and physical factors.
In this example, randomly selected, vitrinite-rich coal samples from
different parts of the world varying in vitrinite reflectance and in
geological age were subjected to petrological, spectrometric and multi-
variate analysis.
Twenty-five vitrinite reflectance measurements were made on each
of the collected samples in the usual manner and then averaged [8].
The standard deviations for a selection of low-ranked coals (vitrinite
reflectance 0.38-1.08) ranged from 0.03 to 0.13 vitrinite reflectance
units, with an average of 0.06.
Diffuse reflectance spectra of the samples were recorded in the
range 4000-600 cm-l using 64 scans and a resolution of 4 cm-l. Three
different spectra for each coal sample were averaged to give one
spectrum for each sample. Each spectrum, consisting of 3401 data
points, was transformed into Kubelka-Munk format [16].
Typical FT-IR spectra are given in Fig. 9.5 for five coal samples
having different rank and vitrinite reflectance. It can be seen that the
aliphatic/aromatic ratio, as determined by the ratio of the peaks at
2950 and 3050 cm l, decreases with increasing rank. It is difficult to
draw qualitative conclusions about the other peaks, probably because
of the interference from minerals.
The wavenumber variables were reduced by approximately a factor
of 10 through a maximum-entropy data reduction process [17-18]. The
reduction process provides smaller matrices for computers to handle
with concomitant decrease in the computing memory and time required
and an increase in speed.
After rejecting the abnormal and outlier calibration samples a
cross-validated calibration model was established between the spectral

297
 4000 3000 2000 1600 1200 600
Wavenumber cm'

Fig. 9.5. Typical FT-IR spectra of coal samples in Kubelka-Munk format.

profiles and the vitrinite reflectance values. The total prediction error
was then computed at the end and the number of PLS components
giving the minimum prediction error was determined.
Forty-six coal samples with vitrinite reflectances ranging from 0.38
to 1.08 were analysed. A total of five of these 46 samples were rejected,
leaving 41 coal samples for the calibration.
Multivariate calibration on the 41 samples gave an absolute
prediction error of 0.09 for determining the vitrinite reflectance of this

298
 C

0 1 2 3 4 5 6 7 8 9
Number of PLS components

Fig 9.6. The progress of prediction error with the number of principal components
extracted.

population. This prediction error is of the same order of magnitude as

the average standard deviation of the corresponding vitrinite reflect-
ance measurements (s = 0.06). This low prediction error is probably the
best that can be obtained unless the measurement uncertainty in the
vitrinite reflectance can be lowered.
The model developed by cross-validation gave an optimum
prediction using seven PLS components as evident by the minimum in
Fig. 9.6, a plot of the Standard Error of Prediction (SEP) [2], vs. the
number of PLS components. A plot of predicted vitrinite reflectance vs.
measured vitrinite reflectance (Fig. 9.7) confirms the good performance
of the model.
The above results, obtained with coal samples from different parts
of the world, produce one model that is generally applicable to vitrinite-
rich coal samples with maturity within the range defined as the oil
window (0.5-1.2). Thus, with the developed model and the combination
of diffuse reflectance FTIR spectroscopy and PLS, we can predict the
coal rank over the important oil production window on coal samples
from different locations. The values of rank obtained are equivalent to
those determined by vitrinite reflectance measurements, and are
obtained with more objectivity, greater efficiency and less sample
preparation.

299
 I / / I I (
O
1.0 0
n
Cd
Q
Q 0
e) 0
U
Q 0 00o0
C 0.8
,) o. a 0
0
Q2) 000
Cd

0 0
: 00
0s
0 0
- 0. 6
'6
._ O

0.4
0. 4 l
I

0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1

Measured vitrinite reflectance

Fig. 9.7. A plot showing the correlation between the measured and predicted vitrinite
reflectance of the coal samples used in the analysis.

9.5 TARGET PROJECTIONS

In certain applications, it will be of interest to find out which wave-

numbers in the infrared profiles are most closely correlated to the
property y. This can be done directly by calculating the covariance
between measured y and the intensities at each wavenumber, i.e. by
calculating the covariances ry,x as
ry,, =ytX (9.22)
For spectral profiles the covariances can be plotted as a covariance
graph [19]. However, a better approach is to calculate the covariance
between the predicted values of the property y and the intensities at
each wavenumber:
ry, = y tX (9.23)
The result of the so-called target projection can be plotted as a graph to
show the connection between a spectral profile andy. The point of using
predicted instead of measured values of the property y in the
correlation, is that wavenumbers of the infrared profiles with both high

300
predictive ability and high correlation with y are given increased
importance compared to those that has only a good correlation. This
can be shown by inserting Eq. (9.18) in Eq. (9.23):

ry, = bTXTX (9.24)

The product XTX is the variance-covariance matrix for the infrared

profiles. Equation (9.24) can be expanded to provide
m n m

ry,i bjExkikj =bjrxixj; i -,2,...,m

j=1 k=l j=l

From Eq. (9.25), it is clear that the covariation ryxi between the pre-
dicted values ofy and the intensities at a wavelength y is calculated as
a weighted sum of covariances ri,xj between intensities at two wave-
lengths i andj. The regression coefficients bj are used as weights. This
shows that the target projection procedure weights the importance of
prediction (bj) and correlation (rxi,xj) so that wavelengths that are
important for predicting the property y have large variance (sensi-
tivity) and are well correlated with other predictive wavelengths will be
highlighted in the target projection plots. This is exactly the wave-
lengths that are important for the interpretation of the structural
descriptor in relation to the property y [19].
Equation (9.25) further shows that for wavelengths where the
covariance ry,i = 0, the variance in the intensities are probably zeros
since it is quite improbable that a linear dependence should exist to
make this correlation exactly zero. Thus, target projection can be used
to find wavenumbers that gave the same intensity independent of the
property we are examining.
The target projection plots are easy to interpret and can be used to
name factors influencing a system. Furthermore, for systems where
variation in the multivariate data with a given dependent variable is
small, target projection can amplify the changes in the target
projection plots.
In infrared spectroscopic data, the profiles are generally broad and
overlapping. In many complex chemical systems the infrared spectra
are featureless and contain very few broad bands. The changes in the
spectra with external dependent factors are small and sometimes
undetectable by visual inspection. Target projection analysis is very
beneficial in such systems for interpretational purposes.

301
9.5.1 Understanding the dehydration process and band
assignment of the overtone vibration of the water of
crystallization of calcium oxalate monohydrate

Calcium oxalate was among the first compounds to be studied by

thermogravimetric analysis and it has been used as a standard for
thermal analysis [20,21]. The diffuse reflectance infrared spectra of
calcium oxalate monohydrate is shown in Fig. 9.8. The bands repre-
senting the H-O-H stretchings of the crystal water in calcium oxalate
is broad over the range 3700-2600 cm - '. It exhibits five bands in the
characteristic O-H stretching region. Band assignments of these peaks
made by Petrov and Soptrajanov [22] are given in Table 9.1.
Calcium oxalate crystal structure contains two non-equivalent
oxalate ions and provides two different environments for the water
molecules in the crystal structure. Furthermore, each water molecule
has one of their OH groups involved in much stronger hydrogen bond-
ing than the other. This should give rise to four O-H stretching bands.

L -

o -

(D
I

Z I
z

I
-o
_
Li
w R
4~~~~~~~~~~~~~ ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
Mo
1,-) r

O I
I 1 i i _n I
oo00 3820 3240 2860 2480 2iOO ;720 i340 960 580
NVENUMBE

Fig. 9.8. A diffuse reflectance spectrum of calcium oxalate monohydrate in KBr (2% w/w).
(Reproduced from Ref. [69] with permission.)

302
TABLE 9.1
Infrared band assignments of the OH stretching vibrations of the water molecules in
the calcium oxalate monohydrate crystals

CaC 2O4 .H2 0 Absorption (cmn1 ) Assignment

3486 v(OH) (2)
3428 v(OH) (1)
3336 v(OH) (1)
3250 26(OH) (?)
3058 v(OH) (2)

The fifth band, the lowest in intensity appears around 3258 cm - l . This
is due to the overtone of the HOH bending mode reinforced by Fermi
resonance [22]. The first and the last in the group (3486 and 3058 cm -l )
are due to one type of water molecule (type 2, as denoted by Petrov and
Soptrajanov) and the remaining (3428 and 3336 cm-1) are due to the
other type of water molecules (type 1). The oxalate stretching vibration
bands appear around 1627, 1320 cm-l and bending vibrations band
appear around 782 cm-l .
Several authors have investigated the dehydration mechanism of
calcium oxalate monohydrate [20-21,23-24]. However, none of them
were able to demonstrate that there are two water molecular environ-
ments present in the crystal and their order of elimination during
heating. In this application we will show that all these are possible in
combination with target projection analysis.
A Nicolet 800 FT-IR spectrophotometer and a diffuse reflectance
accessory manufactured by Spectra-Tech, USA, were used for the
spectral measurements. A high temperature-high pressure chamber
(also from Spectra-Tech) was placed in the diffuse reflectance accessory
in place of the ordinary sample cup. Calcium oxalate sample prepared
as a 2% w/w in finely ground KBr was placed in the sample cup and
heated to 80C and held isothermally for 30 min to eliminate physically
absorbed water from calcium oxalate and KBr. Then it was heated at a
rate 5C/min. and spectra were scanned at regular intervals. Each
sample spectrum measured during heating was ratioed with the
corresponding background (KBr measured under identical conditions)
spectrum and the resulting relative reflectance spectrum [25] was
transformed into Kubelka-Munk format. The area under the OH

303
 *10-s

.a
aD
.W

l
Wavenumber cm-'

Fig. 9.9. The infrared spectral profiles of the OH stretching vibrations of 2% (w/w)
calcium oxalate monohydrate in KBr measured at different temperatures. (Reproduced
from Ref. [691 with permission.)

stretching bands was integrated. The area under OH stretchings was

tested for linearity in advance and it can be used as a measure of the
water molecules in the crystal.
The diffuse reflectance infrared spectra acquired in the tempera-
ture range 80-155C, the dehydrationprofiles obtained using the inte-
grated area under the OH stretching peaks and the first derivatives of
the dehydration profiles of calcium oxalate monohydrate sample (2% w/
w) at 5C/min heating rate (rate of dehydration profiles after data
smoothing) are shown in Figs. 9.9, 9.10 and 9.11, respectively.
The derivative curves clearly show that there are at least two
reactions taking place during the dehydration. At a minimum, there is
a first water release step occurring between 90 and about 125C and a
second release step occurring between about 125 and 150C. This
observation is the first one of this kind. The shape of the reaction rate
profiles suggests two different environments for the water molecules in
the calcium oxalate monohydrate crystal structure, and may corresp-
ond to these different types of water molecules.

304
 1.0
- -- --
--
.- -- '

0.8 i
W
e 0.6 I ~/.
I
d

= 0.4

0.2

N n

80 102.5 125 147.5 170

Temperature oC

Fig. 9.10. The dehydration profiles of the 2% calcium oxalate monohydrate sample in
KBr. (Reproduced from Ref. [69] with permission.)

0.04

a
'1'.
" 0.02

A1
A / I ~ , - X t
0.0 I / is_. .c a! _ _
80 102.5 125 147.5 170

Temperature oC

Fig. 9.11. The rate of dehydration of 2% calcium oxalate monohydrate in KBr. (Repro-
duced from Ref. [69] with permission.)

In order to differentiate between the two different types of water

molecules, we divided the dehydration rate profiles (derivative data) of
2% calcium oxalate sample (at 5C/min) into two subsets A and B (see
Fig. 9.11).
These two subsets indicate the temperature ranges of the dehydra-
tion where one can expect different types of reactions to take place.
These temperature intervals in these two subsets were identified as
80-120 and 120-133C. The raw spectral profiles of the water stretch-
ing vibrations in the temperature intervals 80-120 and 120-133C

305
 *10--

0.000 qI
-0.080

-0.160
-0.240

2 -0.320
is -0.400
0.00
: -0.07

-0.14
-0.21

-0.28
-0.35
3857 3517 3177 2837 2497
Wavenumber (cm-l)

Fig. 9.12. Target projection plots obtained with dehydration spectral profiles in the
temperature range: (a) 80-120C; and (b) 120-133C. (Reproduced from Ref. [69] with
permission.)

were calibrated against their respective temperatures and maximum

correlating factors were obtained for each of the subsets by target
projection [26-27]. These plots are shown in Fig. 9.12a and b,
respectively.
Target projection plots show the relative variation of each variable
with temperature and each of the plots is an expression of the
behaviour of the spectral data with increase in temperature. Zero line
in the target plots separate the spectral profiles that is correlating
positively and negatively with increase in temperature. The spectral
profiles shown in Fig. 9.12 are below the zero line. This indicates that
the peaks decrease in intensity (due to loss of water stretching vibra-
tions) with increasing temperature, which is also obvious from Fig. 9.9.
However, if the water molecules disappear at the same time and at the
same rate then the target projection plots will be exact mirror images of
the original spectral profiles. The figures show that this is not the case.
Figure 9.12a shows without any doubt that the peaks at 3428 and 3336
cm -1 , which are due to type 1 water molecules, disappear at a faster rate
than the type 2 water molecules (compare the depletion profiles of the

306
peaks at 3486 and 3428 cm-'). The plot in Fig. 9.12b shows the dis-
appearance of the type 2 water molecules at a slightly faster rate than
the type 1 (observe that the peak at 3486 cm-l has a maximum
depletion profile). However, a higher rate of depletion of the peak at
3058 cm ' together with the peaks at 3428 and 3336 cm-l was not
expected. This may be an indication that there is a peak relating to type
1 water molecules under the peak at 3058 cm-1 . They may be having
overlapping maximums and seen as one peak in the infrared spectrum.
Furthermore, these plots clearly show that the dehydration of both
types of water molecules takes place at the same time but with
different rates. Obviously, the type 1 water molecules are attached to
the crystal structure with weaker hydrogen bonds than the type 2
water molecules. This is in agreement with the crystal structure
determination of calcium oxalate monohydrate by Cocco et al. [28-29].
Petrov and Soptrajanov [22] in their analysis of the infrared
spectrum of calcium oxalate monohydrate assigned the weak band
(peak at 3258 cm-l) for an overtone of the bending vibrations of one of
the types of water molecules. They were unable to assign the band to
any specific type of water molecules because of the strong oxalate
stretchings appearing in the same region as the bending modes of
water molecules. Our target projection plots indicate that this band
also decreases in intensity with temperature. This is reasonable
because this overtone arises from one type of water molecules. A close
analysis of Fig. 9.12 reveals that this overtone has a slow rate of
disappearance during the first part of the dehydration (Fig. 9.12a) and
a higher rate during the second part of the dehydration. This leads us to
confirm that the overtone arises from the type 2 water molecules.
The approximate maximas obtained with the dehydration rate
profiles indicate that these occur around 0.3 and 0.75 conversion. These
show that the two different types of water molecules are equimolar.

9.6 DATA ANALYSIS AND RESOLUTION BY ALTERNATING LEAST

SQUARES

If N spectra were obtained with mixtures containing A chemical

components at M wavenumbers, they would define a two-way data
matrix X of size N by M. If the components do not interfere with each
other, one can express each spectrum as linear combinations of the
spectra of the contributing components (Eq. (9.26)).

307
 A
X =CS T +E =Zcsi +E (9.26)
i=1

where A is the number of chemical components. ST is the spectral

matrix of the pure components (of dimension AxM) and C is the
concentration matrix of dimension NxA. The experimental noise is
expressed by the matrix E. The superscript, T, implies transposition of
a column vector into a row vector. By applying Eq. (9.26), we assume
that each measured spectrum adds up contributions from A pure
species with concentrations defined by the concentration profiles ci, i =
1, 2, ..., A} and spectra by the spectral profiles {si, i = 1, 2, ..., A}. As
mentioned above, the equation is valid for chemical components that do
not react or interfere with each other. However, if there are inter-
actions between different species, then one has to include a set of
factors that model the non-linearity caused by the interactions. Then
we can write Eq. (9.26) with additional factors that model the inter-
actions.
A A'

X =C *S +E =CisiT + E'Ci s'T +E (9.27)

i=1 i=1

The number of factors (A') needed to model the interactions, is

dependent upon spectral similarity, similarity of spectral changes due
to the interactions, as well as the noise level in the data set. Both
additive and multiplicative errors like baseline shift and intensity
variation due to, e.g., different path length for each spectrum, will
contribute to the noise.
The resolution of the component spectra and concentrations were
accomplished by using the ALS [5,30] procedure. With the constraint of
non-negative concentration and spectral profiles, the iterative process
of calculating the least square estimate of the spectral profiles in S* is
given by

S *T = (C*TC*)- 1 C*TX (9.28)

and the estimate of the concentration profiles in C* by

1
C* =XS*(S*TS*) (9.29)

308
For every cycle, negative intensities are set to zero. Prior to entering
into a new cycle, the concentration profiles are normalised to sum to
one as shown in Eq. (9.30). If one has any selective information
regarding spectra or concentration profiles, the calculated values are
substituted for by the selective information.
The relative concentrations for each component can be calculated,
taking into account that they should sum to one, by:
A
Cb = cib i =1 (9.30)
i=1

and the constants necessary to scale the concentration profiles are

found by least squares as

b = (CTC)-iCT1 (9.31)

9.6.1 Resolution of infrared spectra and concentration

profiles of the components of a multilayer laminate

Polymers and plastics play a very important part in the life of 21st
century man. A wide variety of things are made using plastics and
polymers. Thin sheets containing multilayers of polymer components
are used as packaging material in industry. The following example is to
illustrate the analysis of such a polymer laminate using infrared
microspectroscopy and alternatingleast squares. Application of chemo-
metric techniques to the infrared microspectrometric data acquired
from the laminate can reveal the spectra of individual layers and their
concentration profiles. Furthermore, the chemical changes taking
place at the interfacial regions can also be detected and their chemical
information can be extracted in the form of the layer's infrared spec-
trum. The chemical changes taking place over a period of time can be
monitored by comparing the infrared spectra of the layers at regular
intervals. This will help the industry in determining the life span of the
laminate.
The problem at hand is a static multicomponent system because
there are no dynamic changes in the concentrations of the components
in the system when the measurements are made. The chemical compo-
sition varies across the cross-section of the laminate and this variation
does not change during the analysis.

309
 12x100o pm apenure

/
r
..,.......--.-
.----
. .....1 ... ..1.11-1.
I laminate layers

-...
_----- I...............
.1.......
....
.1...... I.
...... -.I.. . . .--. ...... ......
.11.
1:x-.: .

Fig. 9.13. A sketch showing the redundant aperturing technique used in the analysis of
polymer laminate.

I
.9

3500 3000 2500 2000 1500 1000

Wavenumber cm-'

Fig. 9.14. A stack plot showing the infrared microspectroscopic spectra acquired by using
the redundant aperturing technique. (Reproduced from Brune et al., Surface Character-
ization, 1997, with permission.)

310
 The multilayer laminate sample was prepared by cutting a 5-pm
thick cross-section using a microtome (Reichert-Jung, model 2050-
Leica).
The sample was then mounted between NaCI windows in a com-
pression cell (Spectra-Tech, Inc.). A small crystal of KBr was also
placed in the same cell and this was used for collecting the background
spectrum. The spectra of the laminate sample were collected at inter-
vals of 2 lam, with a 12x100 pm2 sample area defined by redundant
aperturing technique (Fig. 9.13). A total of 256 scans were co-added at a
resolution of 8 cm-'. A total of 52 spectra of the laminate was collected
in this way.
The infrared microspectrometric data profiles of the 52 spectra (Fig.
9.14) were subjected to multiple component analysis using alternating
least squares regression (ALS).
The stack plot in Fig. 9.14 shows the presence of at least three
components. The components arising from interactions and other
underlying components are difficult to visualise in the data set. The
analysis by alternating least squares resulted in five real components.
The infrared spectra of the components are shown in Fig. 9.16 and their
concentrations are given in Fig. 9.15.
The components 1, 2, 4 and 5 are carbonated poly(vinyl chloride),
poly(vinyl acetate), polyethylene and poly(vinyl dichloride) respect-
ively. The component number 5 is an interaction product between
poly(vinyl acetate) and polyethylene. The depth span of the compo-
nents is 40, 20, 34 and 24 pm for the components 1, 2, 4 and 5,
respectively. The interaction product (component 3) has a double
distribution and spans about 60 pm.

Step-number

Fig. 9.15. The concentration profiles of the components resolved. The total concentration
profiles across the laminate are normalised to unity.

311
 Wavenumber, cm-'

Fig. 9.16. The resolved infrared spectra of the components in the laminate sample.

312
9.6.2 Determination of the equilibrium constant and
resolution of the HOD spectrum by alternating least squares
and infrared analysis

When water and deuterated water are mixed, a disproportionation

reaction takes place and an equilibrium established with the product
HOD as shown below

H 2 0(1) + D20(1) = 2 HOD(1) (9.32)

The equilibrium constant for the equilibrium is given by the equation

K = [HOD] 2/{ [H2 0] [D2 0] } (9.33)

This equilibrium between water and deuteratedwater was first studied

by Topley and Eyring [31] in 1934. The equilibrium in gaseous phase
has been studied by several investigators [32-43]. The theoretical
value of K for the equilibrium in gas phase has been determined from
statistical mechanical calculations using measured and theoretical
vibrational frequencies and anharmonicity constants obtained from
infrared spectroscopy [34]. The best theoretical value (K = 3.85) for the
equilibrium in the gaseous phase was determined by Wolfberg et al.
[41] This led to a theoretical value of 3.88 for the liquid phase equilibria
[44].
The interest in the theoretical determination of K was due to the
difficulty in determining the equilibrium constant by experiment. This
is because the species HOD can only exist in the presence of H 2 0 and
DO, and the analysis of HOD in the presence of H20O and DO back-
ground is difficult.
The use of infrared spectroscopy in the study of H 2 0 and D20O has
been almost absent because of the strong absorptions of the OH/OD
fundamental stretching vibrations. These vibrations generally give
very broad absorptions due to extensive intermolecular bonding in the
bulk of the liquid and created difficulty in obtaining the pure infrared
spectrum of HOD by subtraction routines.
When the modern sampling techniques such as total internal
reflectance became available [45], spectra with reasonable intensities
of absorptions that are suitable for quantitative analysis could be
obtained. However, the resolution of the infrared spectrum of HOD
required the use of the statistical equilibrium constant of 4 [46]. This

313
was needed to calculate the concentrations of the components in the
mixture so that subtraction of H 2 0Oand D 2 0Ospectra could be made.
In this application, the alternating least squares technique was
used to resolve the infrared spectra of H 2 0O,D 2 0Oand HOD present in
the equilibrium mixture, their concentrations and to determine the
equilibrium constant K for the reaction. Furthermore, the resolved
HOD spectra was used in assigning the bands.
The change in the concentrations in the equilibrium mixture was
achieved by changing the proportions of water and deuterated water.
A macro circle cell manufactured by Spectra-Tech was modified in
our laboratory to suit our experimental set up [47]. Samples were taken
approximately to suit previously calculated amounts that could allow
the analysis of the mixture in the range that is gradually changing from
mole ratio 1 - 0 (for water) and 0 - 1 (for D 20 sample). The experi-
ment started with a particular amount of water in the cell. The equili-
brium concentrations were changed by adding deuterated water
gradually in the cell. The D 2 0 sample was measured in the cell alone to
achieve mole fraction 1 for D2O.
A Nicolet 800 FT-IR spectrophotometer equipped with a medium
band MCT detector was used to acquire the infrared spectra. A total of
100 scans were made each time in the range 4000-650 cm-l at a
resolution of 1 cm- '. The spectra were then transformed into log(l/R).
The infrared spectra of pure water, deuterated water and a mixture
of water and deuterated water are shown in Fig. 9.17. The infrared
spectra of the pure components and mixtures in log(l/R) format were
transferred to a PC for processing and data handling. The spectral
profiles were subjected to alternating least squares technique.
With the assumption that the hydrogen bonded structures in water
do not change upon isotopic dilution, it appeared that we needed only
three components in order to describe the equilibrium between H2 O,
HOD and D2O.
Collecting the measured mixture spectra in a matrix X, the matrix
can be expressed as a product of a concentration matrix C and a
spectral matrix S, as shown in Eq. (9.26) (omitting the experimental
error matrix, E). The dimensions of the matrices are as given in Eq.
(9.26). The concentration profiles of the equilibrium mixtures can be
obtained by Eq. (9.29).
During the iterative procedure, for every cycle, negative intensities
in the spectra were set to zero. In addition to the non-negativity
constraints used by Maeder and Zuberbuehler [30] and Karjalainen

314
 I

DO
1
WAVENUMBER, cm-

Fig. 9.17. The infrared spectra of pure water, deuterated water and a mixture of the two
in log(l/R) format. (Reproduced from Ref. [31] with permission from Society for Applied
Spectroscopy.)

[48] for the concentration profiles, we imposed the constraints of zero

concentration of D 2 0Oand HOD when only H2 0Owas present and zero
concentration of H2O and HOD when only D2O was present. Prior to
entering into a new cycle, the concentration profiles were normalised.
Red shifts of the peaks were observed during the addition of deuter-
ated water to the water in the cell. This shift is due to the isotopic
effects on peak positions upon dilution. This created problems in using
only three components in the ALS procedure. In order to compensate
for the shift an additional component was included in the iteration. The
concentration profiles and the shift-factor were normalised to unit
length before entering into a new cycle. The iterative process was
terminated when the change in the difference between the measured
and reconstructed mixture spectra was less than 0.01%.
The first four score vectors obtained from a PCA-decomposition of
the bending region (1800-1000 cm-l), were used as seeds for the con-
centration profiles and the shift-factor.
The resolved concentration profiles, normalised to unit length, for
H 2O, D2 0Oand HOD are plotted vs. mole fraction added H 2 0O,in Fig.
9.18a. Together with the concentration profiles the shift-factor's

315
 z0

z
0
C)

MOLE FRACTION ADDED D2 0

Fig. 9.18. (a) The resolved concentration profiles together with the shift factor's variation
with isotopic composition. (b) The molar concentration profiles of H2 0, D2 0 and HOD.
(Reproduced from Ref. [31] with permission from Society for Applied Spectroscopy.)

316
 -
i
6

5.5

5 ~~~~i . .
4.5

4 , AI

3.5

3 IY

2.5

0.3 0.35 0.4 0.45 0.5 0.55 0.6 0.65 0.7

MOLE FRACTION ADDED D2 0

Fig. 9.19. The equilibrium constant K vs the molar fraction of added D2 0. (Reproduced
from Ref. [31] with permission from Society for Applied Spectroscopy.)

variation with isotopic dilution is plotted. The molar concentrations of

HO, D 2 0 and HOD are plotted in Fig. 9.18b. The molar concentrations
were obtained from a least-square fit between the normalised con-
centration profiles and the known total concentration. The predicted
molar concentrations were used to calculate the equilibrium constant
according to Eq. (9.33). The average equilibrium constant was calcu-
lated to 3.86 + 0.07, based on the concentrations between 30 and 70%
water. We avoided the values in the other regions in order to reduce the
error in the equilibrium constant. A plot of K vs. molar fraction of added
H 2 0 is given in Fig. 9.19. There is an excellent agreement both with
values determined by experiment [39,40] and theory (K = 3.88) [44].
With the additional shift-factor incorporated in the iterative
process, the resolved spectra of H2 0 and D2 0 fit almost perfectly with
the measured spectra of pure H 2 0 and DO. The resolved spectra for
H 2 0 and D2 0 correlates 99.99% with the measured spectra of the two
pure analytes.

317
 z

WAVENUMBER, cm-

Fig. 9.20. The resolved spectrum of HOD and the shift factor. (Reproduced from Ref. [31]
with permission from Society for Applied Spectroscopy.)

The resolved HOD-spectrum is shown in Fig. 9.20 together with the

spectral part of the shift-factor. There are three main bands appearing
in the HOD-spectrum at approximately 3385 (VoH), 2490 (oD) and 1450
cm-l (v2). In addition there are two weaker bands at 2930 (2v 2) and 1850
cm-l (analogue to the association band in normal water (v2 + VL)).
Isotopic dilution impose spectral shift in the bending regions of D 2 0O
and H 2 0O.The shift is too extensive to be exactly portrayed by only one
form factor and leads to the observed small derivative-like bands in the
HOD spectrum in the vicinity of 1640 and 1210 cm 1. The resolved HOD
spectrum has the same spectral features as the HOD spectrum
resolved by Mar6chal [46].

9.7 TWO-DIMENSIONAL CORRELATION SPECTROSCOPY

The direct observation of a correlation between an infrared band and a

Raman band, the examination a correlation between a certain band
and other bands in the same spectrum, etc. are now possible by general-
ized two-dimensional (2D) correlation spectroscopy, which we will
describe in this section. In addition, this method allows one to highlight

318
various information which cannot be extracted easily from a ordinary
one-dimensional spectrum.

9.7.1 Principle of two-dimensional correlation spectroscopy

One may associate 2D NMR above anything else with 2D spectroscopy.

It is true that 2D NMR is an essential tool today to analyze NMR
spectra of complex compounds. However, 2D spectroscopy is not neces-
sarily limited to NMR. In recent years, more and more people have
started using 2D spectroscopy in various areas related to spectroscopy.
Two-dimensional correlation optical spectroscopy, in particular, which
was proposed by Noda [49-51] about ten years ago, is attracting
increased attention as a new method for analyzing an infrared
spectrum. The truth is that 2D correlation optical spectroscopy has an
extremely wide variety of applications, ranging from various
spectroscopic analysis including infrared spectroscopy to even x-ray
diffraction.
As shown in Fig. 9.21 [52], 2D correlation optical spectroscopy
requires to expand a spectrum in both the X and Y axis directions (the

vvavenumDer, v1

Fig. 9.21. An example of a 2D correlation spectrum. 2D infrared-Raman heterospectral

correlation map generated from temperature-dependent spectral variations of N-
methylacetamide in pure liquid. (Reproduced from Ref. [52] with permission. Copyright
(1996) American Chemical Society.)

319
spectra to be drawn in the X and Y axis directions may be the same as
each other or different from each other) and examines correlations
between bands which appear in the expanded spectra. Studying the
correlations, we can more clearly note spectral features (e.g., over-
lapping bands) which cannot be easily extracted from an one-
dimensional spectrum. Although the basic idea of 2D correlation
optical spectroscopy is similar to that of 2D NMR, the methods of
calculating correlation spectra are different [49-51]. In 2D correlation
optical spectroscopy, a dynamic cross-correlation between intensity
variations in bands induced by an external perturbation is calculated
to thereby obtain a 2D correlation spectrum.
We will explain 2D correlation spectroscopy in easier words with
reference to Fig. 9.22 [51]. To obtain a 2D correlation spectrum, first of
all, we must externally apply a certain perturbation (e.g., a time
change, a temperature change, a concentration change) to our system
of interest [51-55]. Subjected to the perturbation, components
contained in the system generally respond differently from each other.
To observe the responses, the system is irradiated with an electro-
magnetic wave. In other words, a series of spectra are measured. Now,
assume that we applied a time change. In this case, we can obtain time-
dependent spectra. As we will explain using formulas, to obtain 2D
correlation spectra, it is necessary to calculate dynamic spectra (Fig.
9.22). Based on the calculated dynamic spectra, we thereafter calculate
2D correlation spectra.
While Fig. 9.22 shows a thermal change, a chemical change and
various other changes as an external perturbation, 2D correlation
spectroscopy, when initially proposed, could be applied only to an
infrared signal which changes sinusoidally with time [49,50]. Although
very effective for studying a system which is applied with a small
external mechanical or electric perturbation as in the case of stretching
a polymer film [56], the initial 2D correlation spectroscopy was sub-
Mechanical, electrical,
Perturbation ' chemical, magnetic,
t.. optical, thermal, etc.

Electro-magnetic
probe (eg, IR, UV) Dynamic
System spectra

Fig. 9.22. A general scheme for constructing generalized 2D correlation spectra. (Repro-
duced from Ref. [51] with permission. Copyright (1993) Society for Applied Spectroscopy.)

320
jected to a restriction that a change of a dynamic spectral intensity with
time (waveform) must be a simple sinusoidal wave. Hence, applications
of the initial 2D correlation spectroscopy were rather limited.
To remove the above restriction and further generalize 2D correla-
tion spectroscopy, Noda [51] proposed generalized 2D correlation
spectroscopy based on a new mathematical algorithm in 1993. The new
2D correlation spectroscopy is applicable to any waveforms, and hence,
usable to various types of perturbations [51-55]. Further, the new
calculation method is readily applicable to a variety of spectroscopic
methods. In addition, generalized 2D correlation spectroscopy can be
easily developed into hetero 2D correlation spectroscopy such as
infrared-Raman and infrared-near infrared.

9.7.2 Synchronous and asynchronous correlation spectra

We will describe the principles of generalized 2D correlation spectro-

scopy in more detail. Assume that we apply some perturbation which
changes with time to a system. In this case, we obtain a series of spectra
y(v, t) (where v denotes an infrared wavenumber, a Raman shift, etc.)
which change during a time period from -T/2 to T/2. Meanwhile,
dynamic spectra y(v, t)] are calculated by subtracting a reference
spectrum y(v) from the series of spectra yt(v, t)}

y(vt)y(v,t) = v)...-T...............T / (9.34)

..................... and the other

Although the selection of the reference spectrum is somewhat

arbitrary, in general, an average spectrum as follows is used

Y(v)= 12 y(v,t)dt (9.35)

In the case we use the formula (9.35), the dynamic spectra are devi-
ations from the average of the spectra. Let us explain dynamic spectra
by showing an actual example. Figure 9.23(A) shows temperature-
dependent changes in near-infrared spectra of Nylon-12 [57]. In the
region from 6000 to 5500 cm-l, we can find bands due to first overtones
of CH2 stretching vibrations of the alkyl chain of Nylon-12. Since the
conformation of the alkyl chain changes with temperature, the

321
 A

wavenumber, v

Nylon 12 (30*C - 150GC)

Fig. 9.23. (A) Temperature-dependent near-infrared spectra obtained from 30 to 150C in

the 6000-5500 cm- 1 region of Nylon-12. (B) Dynamic near-infrared spectra calculated
from original near-infrared spectra shown in (A). (Reproduced from Ref. [57] with
permission. Copyright (1997) American Chemical Society.)

intensities of the first overtones may change accordingly. This, how-

ever, is not very clear in Fig. 9.23A. As soon as we calculate the
dynamic spectra (induced by the temperature changes in this example),
we can find the spectra as shown in Fig. 9.23B and immediately tell
which bands change [57].

322
 Now, in order to obtain generalized 2D correlation spectra, it is
necessary to Fourier-transform the dynamic spectra measured in the
time-domain into the frequency domain. The following is Fourier
transform of dynamic spectral intensity changes y(v1 , t) observed at
some spectral variable v1.

Y(Co) = y(v,te itdt (9.36)

= ylRe(o) + iIm ())

In the formula, yRe () and Y Im (io) denote a real part and an imaginary
part, respectively, of the complex Fourier transform of y(v1, t). The
Fourier frequency co represents the individual frequency component of
the time-dependent variation of (vl, t). In a similar manner, Y2 *(co), the
conjugate of the Fourier transform of dynamic spectral intensity, y(v 2,
t), at spectral variable v2 is expressed as:

Y2 (o) =| Y(v2,t e+itdt (9.37)

= YIRe(o) -iYLm(o)

Once we find the Fourier transform, Y,(co) and Y* (co) of the dynamic
spectra in the time domain measured at v1 and v2, respectively, we can
calculate the complex 2D correlation intensity between them by the
following formula.

X(v i(),v Y 2* ()don (9.38)

Equation (9.38) is a formula for calculating a dynamic cross-correlation

between spectral bands. The formula (9.38) consists of a real part 'D(vl,
v2) and an imaginary part i(v 1 , v2) as shown in the formula (9.39):

X(V1, V2 ) =( (v 1, V2 ) + i(V ,1 V2 ) (9.39)

The real part 4)(vl, v2) and the imaginary part i(v,, v2) are called the
synchronous and asynchronous correlation spectra of the dynamic
spectral intensity variations, respectively. In other words, these repre-
sent that time-dependent changes in the spectral intensities at the two
frequencies v1, v2 are in-phase to each other (synchronous correlation

323
intensity) or out-of-phase to each other (asynchronous correlation
intensity) [51].
While we consider time-dependent changes as perturbation in
relation to the formulas (9.34) through (9.39), since classic time series
analysis allows us to replace time with any other continuous variables,
it is possible to calculate 2D correlation functions corresponding to
various types of external stimuli such as a temperature change, a pH
change and a pressure change instead of a time change.
We will now explain the synchronous and asynchronous correlation
intensities with reference to schematic diagrams. Figures 9.24a and b
show 1D(v,, v2) and T(v,, v2) as two-dimensional contours which are
called synchronous and asynchronous correlation spectra, respectively
[51]. In the synchronous correlation spectrum, there appear on the
diagonal line a few peaks called auto-correlation peaks which corres-
pond to v = v2. The larger a band intensity change in response to an
external perturbation (such as a temperature change), the stronger the
intensity of an auto-correlation peak is. An auto-correlation peak
always has the positive sign. Needless to say, a band having a strong
intensity does not necessarily shows a strong auto-correlation peak.
Hence, bands which overlap each other in a one-dimensional spectrum
may be observed as separate bands in 2D correlation spectra because of
different levels of responses to a perturbation.
Peaks located at the off-diagonal positions of the synchronous
spectrum are called cross peaks. The existence of a cross peak at (vl, v2 )
in the synchronous spectrum means that two bands at v, and v2 change
in a similar manner to each other in response to a certain perturbation.
A cross peak has the positive or the negative sign. A cross peak has the
positive sign when the intensities of both bands increase or decrease
with a perturbation. When one band increases while the other
decreases, a cross peak shows the negative sign.
An asynchronous correlation spectrum provides complementary
information to information from a synchronous correlation spectrum.
Of course, an asynchronous correlation spectrum does not show an
auto-correlation peak. A cross peak at (v1, v2 ) in an asynchronous
correlation spectrum means that the intensities of two bands at v, and
v2 exhibit out-of-phase responses to a certain perturbation. For
example, a cross peak appears when the intensities of two bands
change at different temperatures. If an intensity change at v occurs at
a higher temperature than an intensity change at v2, a cross peak
located above the diagonal line has the negative sign. On the other

324
 I
0
c
.5
M

m
U)

rn

Spectral variable, v,

Spectral variable, v

Fig. 9.24. (a) Synchronous and (b) asynchronous 2D correlation spectra constructed from
dynamic spectra. One-dimensional reference spectrum is also provided at the top and
side of the 2D map. (Original figures were prepared by Noda.)

hand, if the former occurs at a lower temperature than the latter, the
sign of the cross peak is positive. This rule, however, is reversed if D(v,
v2) < 0. One of the major features of generalized 2D correlation spectros-
copy lies in asynchronous correlation spectra. This is because we can
clarify in which order the intensities of various bands change if we
analyze asynchronous correlation spectra.

325
9.7.3 What we can learn from 2D correlation spectroscopy

The advantages of generalized 2D correlation spectroscopy are sum-

marized as follows [51-68].
1. Enhancement of apparent spectral resolution of overlapped bands.
2. Band assignments through observations of correlations between the
bands.
3. Studies of inter and intra-molecular interactions through selective
correlation between bands.
4. Probing the specific order in which the intensities of various bands
change.
Thus far, generalized 2D correlation spectroscopy has been applied, for
example, to time-, temperature-, pressure-, concentration-, pH-, and
phase angle-dependent spectral variations in the fields of infrared,
Raman, near-infrared, visible, mass and fluorescence spectroscopy
[51-681. Generalized 2D correlation spectroscopy has been utilized not
only for basic research but also for applications such as those in
biomedical sciences and food sciences. As an example of 2D correlation
spectroscopy studies, we will describe a 2D infrared correlation
spectroscopy study on the secondary structure of proteins using
hydrogen-deuterium (H-D) exchange [58].
As described in Section 8.7.1, infrared spectroscopy has long been
used as a valuable tool for qualitative and quantitative estimation of
the secondary structure of proteins. Although the assignment of the
components of the amide I band to secondary structure such as a-helix
and 3-sheet has been the object of much effort, it is still a matter of con-
troversy. Two-dimensional infrared correlation spectroscopy enhances
the spectral resolution of the amide I and II regions and makes possible
to assign some of the amide I and II bands to given conformations.
Nabet and Pezolet [58] reported a 2D infrared correlation
spectroscopy study of the secondary structure of myoglobin. They used
H-D exchange of the amide protons as an external perturbation to
generate the 2D synchronous and asynchronous spectra [58]. Because
of the fact that the amide protons associated with each conformation
are not exchanged simultaneously, the contributions from different
conformations to the amide bands may be separated. The analysis of
synchronous and asynchronous maps of myoglobin show that this
method is very useful to unravel the different component bands under
the poorly resolved amide I, II, and II' bands of proteins.

326
 They prepared thin films of myoglobin of microgram quantity on an
attenuated total reflection (ATR) crystal [58]. The H-D exchange was
induced by hydrating the films with a flow of nitrogen containing D2 0O
vapour. In general, there are two kinds of amide groups as to the
kinetics of deuteration; some amide groups that are readily accessible
to water are exchanged rapidly at the beginning of the deuteration
process, whereas those involved in structures that are less accessible to
the solvent show a slower exchange kinetics. Thus, in order to separate
more efficiently the fast kinetics from the slower ones, different
sampling time domains were used.
Figure 9.25 shows A synchronous and B asynchronous 2D infrared
correlation spectra of myoglobin calculated from the first 10 spectra
recorded during the H-D exchange process [58]. In the amide I region of
the synchronous correlation map for the rapidly exchanging protons
(Fig. 9.25A) three correlation peaks appear at 1675, 1640, and 1615
cml. These amide I components are assigned to the -turns, random
coil, and intermolecular P-sheets, often found in aggregated proteins,
respectively. Therefore, it is very likely that the amide groups associ-
ated with these three conformations are exchanged first during the
deuteration process. The strongest peak in the synchronous map is
observed in the amide II region at 1530 cm-l, while in the amide II'
region two major peaks can be identified at 1440 and 1350 cm-1 . The
asynchronous map of myoglobin for the rapidly exchanging protons
develops two cross peaks at 1675-1640 cm-l and 1640-1615 cm-l in the
amide I region, confirming that the three peaks at 1675, 1640, and 1615
cm -1 appearing in the synchronous map are ascribed to three different
conformations.
Figure 9.26 depicts the corresponding synchronous spectrum calcul-
ated from 10 spectra obtained approximately 1 h after the beginning of
the H-D exchange process [58]. It shows one autopeak at 1655 cm - , a
frequency that is generally assigned to the amide I mode of the a-helix
conformation. Thus, it seems that the amide protons of the a-helix
conformation are exchanged more slowly than those associated with
intermolecular -sheets, random coil, and 3-turns [58]. The other intense
peaks observed at 1545 and 1345 cm- l may be assigned to the amide II
and amide II' modes of the a-helices of myoglobin, respectively.
The synchronous spectrum for the slow exchanging system (Fig.
9.26) also shows a weak component at 1625 cm-l. This component could
be due to the 3-sheet conformation. Since the random coil and the -
turn structures do not develop the amide I components in the

327
 50 1250 1350 1450 1550 1650 1750
A Wovenumbers, vt B Woverurmbers,

Fig. 9.25. (A) Synchronous and (B) asynchronous 2D infrared correlation spectra of
myoglobin calculated form the first 10 spectra recorded during the H-D exchange
process. (Reproduced from Ref. [58] with permission. Copyright (1997) Society for Applied
Spectroscopy.)

S
Ike
3
E
c3
C}

13
-A Wovenurnbers,

Fig. 9.26. Synchronous 2D infrared correlation spectra of myoglobin calculated from 10

spectra measured approximately 1 h after the beginning of the H-D exchange process.
(Reproduced from Ref. [58] with permission. Copyright (1997) Society for Applied
Spectroscopy.)

328
synchronous map calculated for the long time domain, the H-D
exchange rate for the P-sheet structure seems to be slower than those
for the random coil and -turn structures.

ACKNOWLEDGEMENTS

Some of the text and figures are reprinted from: D. Brune et al., Surface
Characterization, 1997, pp. 410-424 (with permission from Wiley-
VCH). F.O. Libnau and A.A. Christy, Determination of equilibrium
constant and resolution of the HOD spectrum by alternating least-
squares and infrared analysis,Appl. Spectros., 49 (10) 1995 1431-1438;
A.A. Christy, E. Nodland, O.M. Kvalheim, A. Burnham and B. Dahl,
Determination of kinetic parameters for the dehydration of calcium
oxalate mono hydrate by diffuse reflectance FT-IR spectroscopy. Appl.
Spectros., 48 (5) (1994) 561-568 (with permission from Society for
Applied Spectroscopy).

REFERENCES

1. R.S. McDonald, Infrared spectrometry. Anal. Chem., 56 (1984)

349R-372R.
2. H. Martens and T. Naes, Multivariate Calibration. Wiley, Chichester,
1989.
3. K.R. Gabriel, Biometrica, 58 (1971) 953.
4. H.A. Martens, Multivariate calibration: Quantitative interpretation of
non-selective chemical data, Dr. Techn. Thesis, Technical University of
Norway, Trondheim, 1985.
5. S. Wold, A. Ruhe, H. Wold and W.J. Dunn III, SIAM J. Sci. Stat.
Computat., 5 (1984) 735-743.
6. F.L. Staplin, W.G. Dow, C.W.D. Milner, D.I. O'Connor, S.A.J. Pocock, P.
van Gijzel, D.H. Welte and M.A. Ytikler, How to Assess Maturation and
Paleotemperatures, Short Course Number 7, SEPM, Box 4756, Tulsa, OK
74104, USA, pp. 1-289.
7. B.P. Tissot and D.H. Welte, PetroleumFormationand Occurrence-A New
approachto Oil and Gas Exploration.Springer Verlag, Berlin, Heidelberg,
New York, 1984.
8. E. Stach, M.Th. Mackowsky, M. Teichmiiller, G.H. Taylor, D. Chandra
and R. Teichmuiller, Stach's Textbook of CoalPetrology, 3rd edn. Gebrtider
Borntraeger, Berlin, 1982.
9. D.H. Dembicki, Jr., Geochim. Cosmochim. Acta, 48 (1984) 2641.

329
10. P.L. Robin and P.G. Rouxhet, Geochim. Cosmochim. Acta, 42 (1978) 1341.
11. T.V. Verheyen and R.B. Johns, Geochim. Cosmochim. Acta, 45 (1981)
1899.
12. T.V. Verheyen, R.B. Johns and R.J. Esdaile, Geochim. Cosmochim. Acta,
47 (1983) 1579.
13. M.P. Fuller, I.M. Hamadeh, P.R. Griffiths and D.E. Lowenhaupt, Fuel, 61
(1982) 529.
14. R.E. Anacreon, Application of diffuse reflectance for analysis of coal
samples, Perkin-Elmer IR Bulletin, IRB-92, Instrument Division,
Norwalk, CT, pp. 1-29.
15. P.M. Fredericks, J.B. Lee, P.R. Osborn and D.A.J. Swinkels, Appl.
Spectrosc., 39 (1985) 303.
16. P. Kubelka and F. Munk, Ein Beitrag zur Optik der Farbanstriche.
Zeitschriftfur Technische Physik, 11a, (1931) 593-601.
17. W.E. Full, R. Ehrlich and S.K. Kennedy, J. Sed. Petrol., 54 (1984) 117.
18. T.V. Karstang and R.J. Eastgate, Chemom. Intell. Lab. Syst., 2 (1989) 209.
19. O.M. Kvalheim and T.V. Karstang, Chemom. Intell. Lab. Syst., 7 (1989) 39.
20. R.L. White and J. Ai, Appl. Spectrosc., 46 (1992) 93.
21. L.A. Sanchez and M.Y. Keating, Appl. Spectrosc., 42 (1988) 1253.
22. I. Petrov and B. Soptrajanov, Spectrochim. Acta, PartA, 31 (1975) 57.
23. C. Bremard, C. Depecker, H. Des Grousilliers and P. Legrand, Appl.
Spectrosc., 45 (1991) 1278.
24. E.S. Freeman and B. Carrol, J. Phys. Chem., 62 (1958) 394.
25. A.A. Christy, R.A. Velapoldi, T.V. Karstang, O.M. Kvalheim, E. Sletten
and N. Telnaes, Chemom. Intell. Lab. Syst., 2 (1987) 199.
26. O.M. Kvalheim and T.V. Karstang, Chemom. Intell. Lab. Syst., 2 (1987) 235.
27. A.A. Christy, O.M. Kvalheim, F.O. Libnau, G. Aksnes and J. Toft, Vib.
Spectrosc., 6 (1993) 1.
28. G. Cocco, Atti Acad. Naz. Lincei, Rend. Classe Sci. Fis. Mat. Nat., 31 (1961)
292.
29. G. Cocco and C. Sabelli, Atti Soc. Toscana Sci. Nat. PisaMem. PV Ser. A,
69 (1962) 247.
30. M. Maeder and A.D. Zuberbuehler, Anal. Chim. Acta, 181 (1986) 287.
31. B. Tropley and H. Eyring, J. Chem. Phys., 2 (1934) 217.
32. H.C. Urey, J. Chem. Soc., (1947) 562.
33. A.J. Narten, Chem. Phys., 41 (5) (1964) 1318.
34. J.W. Pyper and F.A. Long, J. Chem. Phys., 41 (1964) 2213.
35. A. Narten, J. Chem. Phys., 42 (1965) 814.
36. R.A. Weston, Jr., J. Chem. Phys., 42 (1965) 2635.
37. L. Friedman and V.J. Shiner, Jr., J. Chem. Phys., 44 (1966) 4639.
38. J.W. Pyper, R.S. Newbury and G.W. Barton, Jr., J. Chem. Phys., 46 (6)
(1967) 2253.
39. A.J. Kresge and Y. Chang, J. Chem. Phys., 49 (1968) 1439.

330
40. V. Gold, Trans. FaradaySoc., 64 (1968) 2270.
41. M. Wolfsberg, A.A. Massa and J.W. Pyper, J. Chem. Phys., 53 (8) (1970)
3138.
42. J.W. Pyper and L.D. Christensen, J. Chem. Phys., 62 (7) (1975) 2596.
43. G. Jancso and W.A. Van Hook, Chem. Rev., 74 (1974) 689.
44. D.V. Fenby and A. Chand, Aust. J. Chem., 31 (1978) 214.
45. A. Lorber, Anal. Chem., 58 (1986) 1167.
46. Y. Marechal, J. Chem. Phys., 95 (8) (1991) 5565.
47. F.O. Libnau, A.A. Christy and O.M. Kvalheim, Appl. Spectrosc., 49 (10)
(1995) 1431.
48. E.J. Karjalainen, Chemom. Intell. Lab. Syst., 7 (1989) 32.
49. I. Noda, Bull. Am. Phys. Soc., 31 (1987) 520.
50. I. Noda, Appl. Spectrosc., 44 (1990) 550.
51. I. Noda, Appl. Spectrosc., 47 (1993) 1329.
52. I. Noda, Y. Liu and Y. Ozaki, J. Phys. Chem., 100 (1996) 8674.
53. I. Noda, A.E. Dowrey, C. Marcott, Y. Ozaki and G.M. Story, Appl.
Spectrosc., 54 (2000) 236A.
54. Y. Ozaki and I. Noda, Two-dimensional Correlation Spectroscopy. Am-
erican Institute of Physics, New York, 2000.
55. Y. Ozaki, in: J.M. Chalmers and P.R. Griffiths (Eds.), Handbook of Vibra-
tional Spectroscopy. Wiley, Chichester, 2001, in press.
56. I. Noda, A.E. Dowrey and C. Marcott, in: G. Zerbi (Ed.), Modern Polymer
Spectroscopy. Wiley-VCH, Weinheim, 1999, pp. 1-32.
57. Y. Ozaki, Y. Liu and I. Noda, Macromolecules, 30 (1997) 2391.
58. A. Nabet and M. Pezolet, Appl. Spectrosc., 51 (1997) 166.
59. I. Noda, Y. Liu, Y. Ozaki and M.A. Czarnecki, J. Phys. Chem., 99 (1995)
3068.
60. S.J. Gadalleta, A. Gericke, A.L. Boskey and R. Mendelsohn, Biospectros-
copy, 2 (1996) 353.
61. M. Muller, R. Buchet and U.P. Fringeli, J. Phys. Chem., 100 (1996) 10810.
62. I. Noda, Y. Liu and Y. Ozaki, J. Phys. Chem., 100 (1996) 8665.
63. K. Ataka and M. Osawa, Langmuir, 14, 951 (1998).
64. Y. Nagasaki, T. Yashihara and Y. Ozaki, J. Phys Chem., B, 104 (2000)
2846.
65. M.A. Czarnecki, P. Wu and H.W. Siesler, Chem. Phys. Lett., 283 (1998) 326.
66. C.P. Schultz, H. Fabian and H.H. Mantsch, Biospetrosc., 4 (1998) 19.
67. L. Smeller and K. Heremans, Vib. Spectrosc. 19 (1999) 375.
68. B. Czarnik-Matusewicz, K. Murayama, R. Tsenkova and Y. Ozaki, Appl.
Spectrosc., 53 (1999) 1582.
69. A.A. Christy, E. Nodland, O.M. Kvalheim, A. Burnham and B. Dahl,
Determination of kinetic parameters for the dehydration of calcium
oxalate mono hydrate by diffuse reflectance FT-IR spectroscopy. Appl.
Spectros., 48 (5) (1994) 561-568

331
Appendix I

Physical constants, conversion factors

and atomic masses

A. Some physical constants

Quantity Symbol Value Units

Speed of light c 2.99792x103 ms - 1

Planck Constant h 6.62608xO134 Js
Avogadro constant NA 6.02214x10 2 3 mo1-1
1.66054x10- 2 7
Atomic mass unit u kg
Electron mass me 9.10939xl10-3 kg
Proton-mass mp 1.67262x10 27 kg
-2 7
Neutron mass ms 1.67493x10 kg
9
Elementary charge e 1.60218x10-1 C
Gas constant R 8.31451 JK - 1 mol - 1

B. Conversion factors

Quantity Conversion factor

1 eV 1.60218x10-19J
1 cal 4.184 J
1 atm 101.325 kPa
1A 10-1 m
-
1 Nm l 103 gl

333
C. Some atomic masses

Element Mass x10-2 7 kg

Hydrogen (H) 1.6738

Carbon (C) 19.9450
Nitrogen (N) 23.2587
Oxygen (0) 26.5676
Phosphor (P) 51.4332
Sulphur (S) 53.2369
Chlorine (Cl) 58.8867
Iodine (I) 210.7299

334
Appendix II

Some character tables and point groups

Cs E GxE
A' 1 1 x,yR z xy,z,xy
A" 1 -1 zR,R,Ry xzyz

Ci E i
Ag 1 1 R,RyRz x ,y z ,xy,xz,yz

A. 1 -1 Xy,z

C2 lE C2(z)
A i 1 ZR z x2,y 2 z2,xy
B 1 -1 xyRx,y xz,yz

C2v and C3v are found in the text of Chapter 4.

C4, E 2C4(z) C2 2(,y 2Gd

A1 1 1 1 1 1 z X2 +y2,z2
A2 1 1 1 -1 -1 Rz
2
B1 1 -1 1 1 -1 _y2
B2 1 -1 1 -1 1 xy
E 2 0 -2 0 0 (xy) (Rx,Ry) (yz,xz)

335
C5 E 2C 5(z) 2C 52 5y v 4= 72
2 2
41 1 1 1 1 z z ,X2 +y

A2 1 1 1 -1 Rz
E1 2 2cos4 2cos2 0 (xy) (R,Ry) (yz,xy)
2
E2 1 2cos2 2cos 0 x y2,xy

C,, E 2C, ... ooC V

+ (A1 ) 1 1 ... 1 z x2 +y2,z2

Z- (A2) 1 1 ... -1 Rz
n1(El) 2 2cos4 ... 0 (x,y) (Rx,Ry) (yzxz)
A (E2) 2 2cos25 ... 0
(Eg) 2 2cos35 ... 0 (x2 -y 2xy)

C2h E C2 i ah
2
Ag I 1 1 1 Rz 2,2,y

Bg 1 -1 1 -1 Rx,Ry xy
Au 1 1 -1 -1 z yz,xz
B, 1 -1 -1 1

D2 E C2 (z) C2 (y) C2 (x)

2 2 2
A 1 1 1 1 Rz z ,X ,y

B1 1 -1 1 -1 ZRz xy
B2 1 1 -1 -1 y,Ry xz
B3 1 -1 -1 1 x,Rx yz

D3 E 2C3 3C2 '

Al 1 1 1 z2x 2 + y2
A2 1 1 -1 ZR z

E 2 -1 0 (xy),(RxRx) (x2-y2 ,xy),(yz,xz)

336
D4 E 2C4 C2 2C 2 ' 2C2"

A1 1 1 1 1 1 z2,x2+y
A2 1 1 1 -1 -1 z,R
B1 1 -1 1 1 -1 x2_y 2
B2 1 -1 1 -1 1 xy
E 2 0 -2 0 0 (xy),(R R,) (yz,xz)

2
D2 d E 2S4 (z) C2 2C 2 ' (x) 3d
2
A1 1 1 1 1 1 z2,x2+y
A2 1 1 1 -1 -1 R
2
,B1 1 -1 1 1 -1 x2y
B2 1 -1 1 -1 1 z xy
E 2 0 -2 0 0 (xy),(R,,R v) (yz,xz)

D3d E 2C 3 3C2 ' i 2S 6 3cd

Alg 1 1 1 1 1 1 z2+y 2
A2g 1 1 -1 1 1 -1 Rz
Eg 2 -1 0 2 -1 0 (Rz,Ry)
Alu 1 1 1 -1 -1 -1 2_y 2
A2 u 1 1 -1 -1 -1 1 z xy

iE, 2 -1 0 -2 1 0 (x,y) (yz,xz)

D2h E C2(z) C2(y) C2 (x) i oy cz y

2
Ag 1 1 1 1 1 1 1 1 z2,x2y
Big 1 1 -1 -1 1 1 -1 -1 Rz xy
B2g 1 -1 1 -1 1 -1 1 -1 Ry xz
Bg 1 -1 -1 1 1 -1 -1 1 R yz

AU 1 1 1 1 -1 -1 -1 -1
Bl1 1 1 -1 -1 -1 -1 1 1 z
B2. 1 -1 1 -1 -1 1 -1 1 y
B3 . 1 -1 -1 1 -1 1 1 -1 z

337
D3
,1 E 2C3 3C2' Ch 2S 3 3c,

A1 ' 1 1 1 1 1 1 z2
A2 1 1 -1 1 1 -1 R,
E' 2 -1 0 2 -1 0 (x,y) (x2-y2 ,xy)
Al" 1 1 1 -1 -1 -1
A2 " I 1 -1 -1 -1 1 z
E" 2 -1 0 -2 1 0 (Rx,RY) (yz,xz)

D6h E 2C6 2C 3 C2 3C 2' 3C 2 " i 2S 3 2S 6 , 3, d 3V =

lg 1 1 1 1 1 11 1 11 1 2,X2+y2
A2g 1 1 1 1 1 1 1 1 1 -1 -1 R
Big 1 -1 1 -1 1 -1 1 -1 1 -1 1 -1
B2g 1 -1 1 -1 -1 1 1 -1 1 -1 -1 1 (yz,xz)
Ig 2 1 -1 -2 0 0 2 1 -1 2 0 0 (R~,Ry) (x2-y2,xy)
2 -1 -1 2 0 0 2 -1 -1 2 0 0
Alu 1 1 1 1 -1 -1 -1 -1 -1 -1

Blu 1 -1 1 -1 1 -1 -11 -1 1 -1 1
lu

lu 2 1 -1 -2 0 0 -2 -1 1 2 0 0 (x,y)
2 -1 -1 2 0 0 -2 1 1 -2 0 0

D.h E 2CO ... ooa, i 2S, ... moC

Z(Ag) 1 1 1 1 1 1 z2,x2+y2
g(A 2g) 1 1 -1 1 1 -1 R
[lg(Elg) 2 2cosl 0 2 -2cos 0 (RRy) (yz,xz)
Ag(E2g) 2 2cos2? 0 2 2cos2o 0 (x2 -y2 ,xy)

,S(Alu) 1 1 -1 -1 -1 -1
XZ(A 2 u) 1 1 -1 -1 -1 1
F[I(E1u) 2 2coso 0 -2 2coso 0
A(E 2 u) 2 2cos2o 0 -2 -2cos2o 0 (XY)

338
Appendix III

Matrices

A. A matrix

A matrix is an array of elements of the following form. The horizontal

sets of elements are called rows and the vertical sets of elements are
called columns. A matrix is represented by a single symbol.

all a, 2 a 13 all a,.

a2 l a2 2 a23 a2j a2m

A = [aj] = ail ai 2 ai3 ai3 aim

ani an 2 an an anm

The element aij is the entry belonging to the ith row andjth column of
the matrix. The above matrix has n rows and m columns. These are
called dimensions of the matrix. The above matrix is said to be an nxm
matrix. When the dimensions are equal, the matrix will have an equal
number of rows and columns (m = n) and the matrix is a square matrix
with m 2 entries. Matrices follow certain rules and we shall learn more
about their behaviour in the following sections.

B. Matrix addition

Matrices of same dimensions can be added. IfA and B are two matrices
of the same dimensions then the sum of the matrices is a matrix of the

339
same dimension as A and B. The resulting matrix C will contain entries
cij which are given as cij = a + b.
For example, the addition of matrices A and B are shown below

A= 0 1 -
1 2 0

0 -1 2
B= 1 2 0
-1 -1 0

1 -1 3
A+B= 1 3 -1
0 1 0

Matrix addition is commutative. It means that whether we add matrix

B with A or A with B, the resulting matrix is the same.

A+B=B+A=C

C. Scalar product of a matrix

A matrix A can be multiplied by a real number k. The multiplication

can be written as kA. The resulting matrix is of the same dimension as
A and contains elements kaij as entries. For example 3A is

0 3
3A= 0 3 -3
3 6 0

D. Matrix subtraction

The subtraction of a matrix B from A can be considered as an addition a

A+ (-1)B. Matrices of the same dimensions can be subtracted from each

340
other. The resulting matrix is a matrix of the same dimension. The
matrix would contain elements cij = aj - biv if B is subtracted from A or
di = bj - aij ifA is subtracted from B. It is easy to see that the result of
the subtraction is not the same. For example, the subtractions would
result into the following matrices.

1 1 -- -1 -1 I
A-B=fi -1 B -A = 1
2 3 0 -2 -3 0

E. Matrix multiplication

Two matrices C and D can be multiplied if the number of columns in

the matrix C is the same as the number of rows in D. For example if C is
an nxm matrix then D has to be an mxk matrix. The product of the
matrices C and D is an nxk matrix.
If the resulting matrix is E then the element ej is given by

ei = Zcikdkj
k=l

where m is the number of columns in the matrix C and number of rows

in the matrix D. There is no product between two matrices that does
not satisfy the requirement above.
For example, if

C=[' 2] and D =[ -1 1]
then the product E = CD is

CD = 5 2

For example, the product between matrices A and B given above is

341
 0 1 0 -1 2 1 0 2
AB= 0 1 -1 1 2 0 = 2 3
12 0 -1 -1 0 2 3 2

The product between B and A is

0 -1 2 1 0 1 2 3 1
BA= 1 2 = 1 -1 1 2 -1
-1 -1 0 1 2 0 -1 -1 0

As we can see from above, the product of two matrices is not generally
commutative.

F. Identity matrix

All square matrices have an identity matrix of the same dimension.

The identity matrix has elements i, = 0 when i j and 6ij = 1 when i =j.
The identity matrix is denoted by the symbol I.
The identity matrix for all 3x3 matrices is then

-1 0 O
I= 0 1 0
0 I

The product of a square matrix with its identity matrix is the square
matrix itself. That is IA = A.

0 1 0 1 1 0 1
IA= 0 1 0 0 1 -1 0 1 -1=A
0 0 1 1 2 0 1 2 0

One can also show that the product of any square matrix multiplied by
its identity matrix yields the same square matrix.

0 1 1 0 0 1 0 1
AI= 0 1 1 0 = 0 1 -1=A
1 2 0 0 1 1 2 0

342
G. Transpose of a matrix

A matrix can be transposed so that the elements in the rows of the

matrix become columns of a matrix. For example, the matrix D can be
transformed into a matrix so that the elements dij become dji of the new
matrix. The new matrix is the transpose and is denoted by D'.

D = [i -1
2 3 1
] D'= l-1 3]

H. Determinant of a matrix

Certain products of the elements compute determinant of a matrix. It

can only be evaluated for a square matrix (number of rows = number of
columns). For example the determinant of the matrix

xL
X= 11 X 12 is computed as x11 x22 - x21 xl 2 and is denoted by

det(X)= XI = Xl l X12
x2 1 22

where x22 and x12 are called co-factors of x1 and x21. When the number of
elements increases, the evaluation of the determinant becomes tedious.
For a 3x3 matrix

Z1 1 Z1 2 Z13

Z = Z2 1 22 Z23

LZ31 Z32 Z33

the determinant can be evaluated by the sum of the products between

the elements of a particular row or column and their co-factors.
The determinant is then

343
det (Z) = z 11 Z 11 + 21 Z 21 + z31 Z31 = Zll (- 1)+l Z2 2 Z23 +
z32 Z33

2 (1)2+1 Z12 Z13 Z31 (_1)3+1 Z12 Z13

Z3 2 Z33 Z22 Z23

- Z1 1 Z1 1 + 12 Z1 2 + z1 3 Z13

The determinant of matrix A = 01 1

isthen(A)+
det1(0-1)

is then det (A)=A=11(0 + 2) -0 + 1(0-1)} = 1

I. Cofactor

The co-factor of an element zij in a square matrix Z of dimension nxn is

the determinant of the matrix of dimension [(n-1)x(n-1)] obtained by
deleting the ith row andjth column from the matrix Z. The cofactor is
then ()i+j I[zi]1. The co-factor is denoted by Zij.

J. The co-factor matrix

The co-factor matrix of a square matrix Z of dimension nxn is the

matrix obtained with all the determinants of the co-factors of the
elements in the matrix Z. The cofactor matrix is denoted by [Z,,]. The
co-factor matrix of Z given above is then

Z2 2 Z23 Z2 1 Z2 3 Z2 1 Z2 2 -

Z3 2 Z33 31 Z33 Z3 1 Z3 2

Z1 2 Z13 Z1 1 Z1 3 Zll Z1 2

Z32 Z3 Z31 33
33 Z3 1 Z3 2

Z Z13 _Z 11 Z13 Zll Z1 2

Z2 2 Z23 Z2 1 Z2 3 Z2 1 Z2 2

The co-factor matrix of the matrix given above is then

344
 11 -1 0 -1 0 1
20 1 1 21 2 -1 1
0 1 I1 111;
-]2 0
01
o:2
1 Il
1 2l = -2
11 0 -1
-1
1
-2
1

11 -1 0 X 0 1

K. Inverse of a matrix

Inverse A -1 of a non-singular (non-zero determinant) square matrix A

is a non-singular square matrix that satisfies the following

A-1A = I

Inverse of a matrix A can be evaluated from its determinant and the

transpose of the co-factor matrix.

A- 1 I(1/det (A)][Aij]'

2 2 -1
= 1 -1
-1 -2

L. Solution of simultaneous equations

The knowledge of matrices can be applied to solve simultaneous

equations. For example consider the following set of equations.

X1 + x3 = 3

X2 -X 3 = -3

x1 + 2x2 = 5

The above equations can be written as follows

345
X1 + OX2 + x 3 = 3

OX1 + x 2 - X3 = -3

x, + 2 x2 + OX3 = 5

In matrix form the equations can be written as

0 1 -1O x2= -3

The matrix at the right hand side is the same as matrix A given above.
So the equations can be written as

Ax = y where x and y are x21 and -3, respectively.

X3 -1

The equations can be solved by multiplying the equation by the inverse

of A. That is

A-l Ax = A-l y

Ix =A-l y

x =A-ly

That is, the solution for the above equations is

x = A-ly

x2 = -1 1 -3 =-1
X3 -1 -2 1 -1 2

that is, x1 = 1, x2 = -1 and x, = 2.

346
M. Eigen values

If P is a square matrix of order n, then the k matrix [P - IX] is called the

characteristic matrix of P. The determinant IP - /I1 is called the
characteristicdeterminant of P. The expansion of the above determi-
nant expressed as a polynomial of degree n in X is called the character-
istic function of P. The equation f(X) = 0 is called the characteristic
equation of P and its roots are called the characteristicroots or eigen
values.
For example, for the matrix

[-2 2]'

the determinant becomes

1-k 1
-2 2-X}

This leads to the equation (1 - X)(2 - X) + 2 = 0.

The solution to the above equation gives values = 3 and X = 1.
These are the eigen values of the matrix P.

N. Eigen vectors

For any square matrix P of order n, the eigen value equation can be
written as
Px = Xx
where x is a lxn matrix and the X one of the eigen values. There are n
solutions to this equation corresponding to n eigen values. This vector x
is called an eigen vector of the matrix P. The eigen vectors can be found
by solving the above equation.

O. Trace (character) of a matrix

If A is a square matrix, the trace of A is defined as the sum of the

elements on the principal diagonal. The trace is denoted by tr(A). The
trace of matrix A given above is

347
tr 0 1 -1=2
112 -_

P. Similarity transformation of a matrix

If Q is a non-singular square matrix then the product RQR - = F is

known as a similarity transformationof Q by R. The matrices Q and R
are similar. Their determinants, eigen values and traces are equal.

tr(RQR 1 ) = tr(F)

For example, if Q is

0 -1 2
1 2 (the same as B above) and R is
-1 -1 0

O1 -1 (the same as A above), then

1 2 0

1 0 1 0 -1 2 2 2 -1 -2 -4 1
RQR - = A BA- 1= 1 -1 1 2 0 -1 -1 1 = 1 11
1 2 0 -1 -1 -1 -2 1 -1 -33

det(ABA - l) = det(B) = 2

tr(ABA -1 ) = tr(B) = 2

eigen value equations ofABA - and B are the same as ;3 - 22 + 3R -2 =

0.

348
Q. Diagonalisation of matrices

A non-singular square matrix Q can be diagonalised by another matrix

R by similarity transformation (RQR - 1 = G) such that the resulting
matrix G has only non-zero diagonal elements. These diagonal
elements are the same as the eigen values of the matrix. The columns of
the matrix Q are the same as the eigen vectors corresponding to the
eigen values taken in the same order.
Diagonalisation by similarity transformation is used in reducing
the matrices representing operations of a point group. To illustrate
this, let us use the matrices A, B, C, and D used in Section 4.6. The
matrices A and C are in the diagonal form. The matrices B and D are
the same. They have eigen values X = -1, -1 and 1. When the matrices
are diagonalised they have -1, -1 and 1 as diagonal elements. The
diagonalisation of the matrix B and D can be done by finding the eigen
vectors of the matrices B and D. These matrices have one diagonal
element (-1) in the middle row each. Therefore it is enough to diagon-
alise the first and the last rows

B=D= -1 0
-1 0 0

For k = -1

[0 -- 1 [ 01[X

2
The1 normalizing condition leads
= to +

The normalising condition leads to x2 + 1

2x 2 = 1 x = +1I and z = 1/2.

For k = 1

349
01 -ol=-,Lo l0x
-1 -iL oo z]

with normalising condition as above, we get x = +1/F2 and z = -1/42.

The matrix Q that diagonalises the matrices B and D can be written
as

o -1/2] o 1/2]-
Q/ 0
Q1/ -1 0 and Q =1 0 -1 0
o 1/ j Q l= 2 o 1/J2

-1/ 2 0 -1/ 2- 0O o -1- 1/ 0 -1/-

QBQ - = 0 -1 O -1 O O -1 0
L1/ 0 11 2 I-1 0 O / 0 1/]

= -1 0
0 0 -1

Therefore, the matrices A, B, C and D representing the symmetry

operations E, C2, %a,and %z are reduced to

1 0 O, 1
0 0 1 0 0 0O 0 i
0 1 0,O -1 i 1 0 and -1 0 , respectively.
OO 1O 0 - 0 1 0 0 -1

350
Index
absorbance, 21, 133, 285
absorption spectrum, 18
absorptivity, 131
accidental degeneracy, 95
acrylonitrile- butadiene-styrene
(ABS), 165
alkyl chains, 252
allowed transition, 19
alternating least squares, 309
amide I, 142,255
amide II, 142,256
amplitude, 3
anharmonic constant, 35
anharmonic terms, 35
anharmonicity, 33, 37,38
anionic surfactant sodium dodecyl
sulphate, 198
anisotropic distribution, 204
anti-Stokes lines, 79
anti-symmetric stretching vibrations,
29
anti-tumour agen, 170
apodization, 121
arsenic-doped silicon, 234
ascorbic palmitate, 261
assembled cells, 134
asymmetric stretching, 239
asynchronous correlation spectra,
321,325
attenuated total reflection (ATR)
method, 131, 136, 141, 196
auto-correlation peak, 324
azobenzene, 220
bacterial cells, 263
bacteriorhodopsin, 255, 268
barbituric acid, 249
beamsplitters, 107, 116
bending vibrations, 29
benzaldehyde, 38
biological materials, 195
biomarkers, 296
biomembranes, 260
biplots, 294
bisacrylates, 215
bolometric effect, 110
bone disease, 227
breast tumour, 232
cadmium stearate, 241, 242
caffeine, 187
calcium oxalate, 302
carbon nanoparticles, 171
cassegrain type, 188
cellulose ether, 197
centreburst point, 120
cervical cancer, diagnosis of, 273
chalcogenide glass, 235
character table, 71
characteristic absorption bands, 31
charge-transfer reaction, 247
chemometrics, 256
351
chlorophylls, infrared spectra of, 259
chromophores, 238
chromophoric groups, 255
classical least-squares, 181
coal rank, 296
CO-ethylene- propylene alternating
copolymers, 197
colon carcinoma, 233
combination modes, 33, 37
commutative, 59
concentration, 285
conducting polymers, 218
conjugate operations, 65
Connes advantage, 120
continuous-scan FT-IR, 124
continuum model, 151
corpuscular, 6
cross peaks, 324
crystallinity, 212
curve-fitting, 257
cycle, 3
cysteine residues, 255
degenerate stretching, 239
degenerate vibrations, 29
dehydration profiles, 304
dependent variables, 288
detectivity, 113
detector, 109
detector responsivity, 113
deuterated triglycine sulphate
(DTGS), 106
deuterated water, 313
deuteration studies, 202
di(carboxystyryl)benzene, 219
diatomic molecule, 23
dichroic ratio, 204
diffuse reflectance (DR) method, 131,
147, 285
diffuse-reflected light, 148
diffusion depth, 156
dioctyl phthalate, 185
dipole moment, 82
352
direct deposition, 185
dispersive instruments, 117
dispersive spectrometers, 207
distinct operations, 45
dynamic FT-IR procedures, 207
dynamic IR spectroscopy of polymers,
212
dynamic spectral intensity, 321
elastomer sealing rings, 165
electric dipole moment, 20
electroluminescence, 219
electromagnetic spectrum, 1
emission spectroscopy, 131
epitaxial layers, 146
essential oil constituents, 183
etendue advantage, 120
exinite, 296
external perturbation, 320
fast Fourier transform, 127
fatty acids, 260
Fellgett advantage, 120
Fermi resonance, 38,95
fingerprint region, 18
fixed cells, 134
fluoranthene, 185
focal-plane arrays, 222
forbidden transition, 19
force constant, 25
Forman algorithm, 123
Fourier transform, 323
Fourier transform dynamic infrared
spectroscopy, 208
frequency, 25
frequency domain, 125
FT-IR instrumentation, history of,
105
functional group, 22,285
fundamental vibrations, 17,80
fundamentals, 32
gauche conformation, 248
gauche forms, 240
GC/FTIR, 180
Globar source, 107
Golay detector, 110
ground vibrational state, 21
group frequencies, 18, 31, 97
Halobacteriumsalinarium,268
harmonic oscillator approximation,
23
harmonic oscillator model, 33
harmonic vibrations, 23
heavy water, 135
hemes, 255
Herman orientation function, 205
Hermite polynomial, 28
hexaalkoxytriphenylenes, 215
homomorphous, 59, 64
human melanoma, 233
hydrocarbon chains, 238
hydrogen bonds, 199
Hz, 3
identical operation, 43
improper rotation axis, 47
impulse-response technique, 207
incident light, 148
independent variables, 288
inertinite, 296
infrared absorption, 19
infrared active, 20
infrared detectors, 109
infrared inactive, 21
infrared light, 19
infrared linear dichroism, 203, 212
infrared microspectroscopy, 131
infrared spectra of a model
compound for phospholipids, 260
infrared spectra of chlorophylls, 259
infrared spectra of intact bacterial
cells, 263
infrared spectra of proteins, 255
infrared spectroscopy, 22
interferometer, 105
inversion centre, 47
irreducible representations, 71
isotope shift, 99
isotopic exchange, 202
Jacquinot advantage, 120
Johnson noise, 112
KBr method, 136
Krimm's rule, 202
Kubelka-Munk equation, 147, 150
Lambert-Beer's law, 131
Langmuir-Blodgett films, 146, 220,
236
LC/FT-IR, 172
light-emitting diodes, 216
linear dichroism, 203
linear momentum, 7
liquid crystal polymers, 211
liquid crystals, 206
liquid samples, 134
lithium cells, 221
loading plots, 294
low-temperature infrared
spectroscopy, 255
MCT detector, 114, 188, 231
Mertz algorithm, 123
methacrylic acid, 218
methacryloxy-
propyltrimethoxysilane, 218
Michelson interferometer, 118
micro-crystal domains, 244
mirror misalignment, 124
mirrors, 107, 116
molar absorption coefficient, 133
molar absorptivity, 286
mole, 12
molecular electronics, 236
molecular orientation, 245
molecular vibration, 19, 23
353
monochromatic radiation, 12
Morse's function, 34
Mulliken symbols, 71
multiple linear regression, 290
multiplex advantage, 120
mutual exclusion rule, 30
Nernst glower, 107
noise equivalent power, 113
non-aqueous solutions, 133
normal coordinate, 20
normal frequency, 23
normal modes, 23
normal vibrations, 17, 23, 80, 97
normalized detectivity, 113
object space; 291
order, 43
order-disorder transition, 244
organic light emitting diodes, 214
organic thin films, 195
orientation measurements, 202
overfitting, 293
overtones, 21, 33, 37
parallel-polarized light, 145
partial least squares, 290, 294
particle theory, 1
penetration depth, 137
pentafluorophenylacrylate, 197
pentafluorophenylmethacrylate, 197
period of the motion, 3
perpendicular-polarized light, 144
phase transitions, 260
phospholipids, infrared spectra of a
model compound for, 260
phosphonic acids, 166
photoacoustic signal, 155
photoacoustic spectroscopy (PAS),
131
photoacoustic techniques, 196
photoconductive effect, 111
photoelectromagnetic effect, 111
354
photoemissive effect, 111
photoinitiator, 218
photoisomerization, 220
photon, 6
photon detectors, 109,111
photosynthesis, 255
photovoltaic cells, 236
photovoltaic effect, 111
Planck's constant, 6
point groups, 53
polarizability, 79
polarized light, 144
poly(acrylic acid), 166
poly(ethylene glycol), 230
poly(ethylene oxide), 201
poly(methacrylic acid), 166
poly(methylmethacrylate), 165, 201
poly(propylene oxide), 197
poly(vinylphenol), 201
polyacrylonitrile, 166
polyatomic molecules, 29, 97
polyethylene, 206
polymer blends, 225
polymer characterization, 195
polymeric sulphonic acids, 166
polystyrene microspheres, 223
polyurethanes, 168
predictor variables, 288
principal axis, 43
principal component analysis, 290,
292
prisms and accessories for ATR
spectroscopy, 139
product operation, 57
proper rotation axis, 43
proportionality constant, 286
propyl ester phosphazene, 166
protein dynamics, 255
proteins, infrared spectra of, 255
Pseudomonaschlororaphis,263
pyroelectric devices, 236
pyroelectric effect, 110
pyrolysis products, 182
quantitative analysis, 285
quantum, 6
quantum mechanics, 19
random coil structure, 256
Rayleigh radiation, 79
reducible representations, 70
redundant aperturing, 189
reflection absorption, 131, 143, 238,
241
refractive index, 2, 133
registration advantage, 120
response variables, 288
retardation, 119
retinals, 255
rocking vibrations, 30, 239
rotation reflection axis, 47
Schottky noise, 112
Schrodinger's equation, 27
scissoring, 30,239
score plots, 293
secondary structure, 142
selection rule, 19
SFC/FTIR, 182
silicone, 159
similarity transform, 65
simple harmonic, 2
size distribution, 151
spectrometer, 12
spectroscopic imaging instrument, 222
spectrum, 1
specular reflectance, 285
Staphylococcus aureus, 263
step-scan dynamic FT-IR, 209
step-Scan FT-IR, 125
step-scan impulse-response
experiments, 210
Stokes lines, 79
stretching vibration, 23, 138
structural absorbance, 204
styrene-butadiene block copolymer
blends, 165
subcell packing, 241
supercritical fluid chromatography,
182
surfactant, 184, 197
symmetric stretching, 239
symmetric stretching vibrations, 29
symmetry elements, 42
symmetry of a molecule, 20
symmetry operations, 42
symmetry plane, 47
synchronous correlation spectra, 321,
324
synchronous modulation, 125, 207
target projection plots, 306
target projections, 300
tetrachloroethylacrylate, 197
TGA/FT-IR, 158
thermal conductivity, 156
thermal detectors, 109, 110
thermogravimetric analysis, 302
thermoplastics, 163
thermopneumatic effect, 110
thermovoltaic effect, 110
thin films, 236
time-domain experiments, 125
time-resolved experiments, 125
time-resolved infrared spectroscopy,
255
total internal reflectance, 285
trans conformation, 248
transferrin receptor, 258
transition moment, 82,204
translation, 15
transmittance, 21,131
transmitted radiation, 12
trans-zigzagstructure, 240
triaminotriazine derivatives, 250
trisacrylates, 215
twisting vibrations, 30
two-dimensional correlation
spectroscopy, 256, 318
355
univariate technique, 285 wave theory, 1,6
wavelength, 5
variable space, 291
wavenumber, 5, 286
vinylidene fluoride copolymers, 196
weathered sealants, 159
vinylidene fluoride-trifluorethylene,
window materials, 133
201
vitrinite, 296
x-ray diffraction, 319
wagging vibrations, 30, 241
water molecules, 255 zirconium oxide, 218

356