Scope :

Spectrophothometry

Chromathography

* UV-Vis Spectro
* GC (Gas Chromatography)
* IR- Spectro
* LC (Liquid Chromatography)
* AAS (Atomic Absorption
* HPLC (High Performance Liquid
Spectro)
Chroatography)
* MS (Mass Spectro)
* GC-MS
* NMR (Nuclear Magnetic Resonance)
* MRI (Magnetic Resonance Imaging)
*GC-MS

Modern Analytical Chemistry

Theory of Colorimetry

Light + Color

Colorimetry?
Each chemicals substances/componuds
(gas/solutions/solids) have characteristic
colors(Chroma).
The intensity of such a color is proportional to
the concentration of the substances/
compounds.

Internal Energy of Molecules

Etotal=Etrans+Eelec+Evib+Erot+Enucl
Eelec = Electronic transitions (UV, X-ray)
Evib = Vibrational transitions (Infrared)
Erot = Rotational transitions (Microwave)
Enucl= Nucleus spin (NMR: Nuclear Magnetic Resonance)

or
(MRI: Magnetic Resonance Imaging)

Light

What is Light
Light is a special type of wave
What we know as light or VISIBLE LIGHT is
actually a type of something called
ELECTROMAGNETIC SPECTRUM (ES).
ES:
Electromagnetic Wave (EW)
Electromagnetic Radiation (ER)
6

Electromagnetic Spectrum
The EM spectrum is the ENTIRE range of EM waves in order of
increasing frequency and decreasing wavelength.

As you go from left right, the wavelengths get smaller and the frequencies
get higher. This is an inverse relationship between wave size and frequency.
(As one goes up, the other goes down.) This is because the speed of ALL EM waves
is the speed of light (300,000 km/s).

Electromagnetic Spectrum

1A0 = 10-8cm = 0,1nm (nano meter)

Electromagnetic Spectrum

Electromagnetic Spectrum

Electromagnetic Spectrum

Electromagnetic Waves
When something creates energy it also emits
radiation. Depending on the amount of
energy, the object will emit different types of
electromagnetic radiation.
When we studied mechanical waves, they
were all transferred through a medium. What
medium is light transferred through?
LIGHT DOES NOT NEED ONE!
12

Electromagnetic Radiations
Electromagnetic waves are special in the fact
that they do not need a medium to propagate
through.
But what is creating the disturbance? What is
emitting this energy?
ELECTRONS

13

Electromagnetic Radiations
Electrons in materials are vibrated and emit energy
in the form of photons, which propagate across the
universe.
Photons have no mass, but are pure energy.
Electromagnetic Waves are waves that are made
up of these photons.
When these photons come in contact with
boundaries, E-M waves interact like other waves
would.
14

Electromagnetic Waves
Electromagnetic waves are everywhere.
Light is only a small part of them
Radios
TVs
Microwaves
Light (Visible/UV/InfraRed)

Radiation
Lasers
CD/DVD players
X-Rays

15

Ultra Violet (UV) Spectrum

Frequencies: 7.5 x 1014 - 3 x 1016 Hz
Wavelengths: 400 nm - 10 nm
Quantum energies: 3.1 - 124 eV

The region just below the visible in wavelength is called the near ultraviolet.
It is absorbed very strongly by most solid substances, and even absorbed
appreciably by air.
The shorter wavelengths reach the ionization energy for many molecules, so the far UV
has some of the dangers attendent to other ionizing radiation.
The tissue effects of ultraviolet include sunburn, but can have some therapeutic effects as well.
The sun is a strong source of ultraviolet radiation, but atmospheric absorption eliminates most
of the shorter wavelengths.
The eyes are quite susceptible to damage from ultraviolet radiation.
Welders must wear protective eye shields because of the uv content of welding arcs can
inflame the eyes.
Snow-blindness is another example of uv inflamation; the snow reflects uv while most other
substances absorb it strongly.

Visible Light
Frequencies: 4 - 7.5 x 1014 Hz
Wavelengths: 750 - 400 nm
Quantum energies: 1.65 - 3.1 eV
The narrow visible part of the electromagnetic spectrum corresponds to the wavelengths near

the maximum of the Sun's radiation curve.

In interactions with matter, visible light primarily acts to elevate electrons to higher energy levels.
White light may be separated into its spectral colors by dispersion in a prism.

Infra Red (IR) Spectrum

Frequencies: .003 - 4 x 1014 Hz
Wavelengths: 1 mm - 750 nm
Quantum energies: 0.0012 - 1.65 eV

The range adjacent to the visible spectrum is called the "near infrared" and the
longer wavelength part is called "far infrared".
Used for communication
In interactions with matter, infrared primarily acts to set molecules into
vibration.
Infrared spectrometers are widely used to study the vibrational spectra of
molecules.
Infrared does not penetrate the atmosphere well, but astronomy in the infrared
is carried out with the Spitzer Space Telescope.

The physical Behavior of ES

1. Reflection
3. Absorption
4. Penetration/Transmission
Refraction
Dispersion
Diffraction
Interference
Polarization

Properties of Light
Electromagnetic radiation moves in waves

Wave motion of light

1- Wave Properties
The wave is described either in terms of its wavelength (),the distance
between successive maxima or minima of a wav(nm),
the frequency (v), the number of oscillation of the field per second.
The velocity of light, c, is
given by the equation:

C= V*

Speed of E/M Waves

As a Waves, we found that

=>C = V*
We also said that the speed of a wave in a certain
medium is always constant.
It has been found that the speed of E-M waves and
light is -- 3 x 108 or 300,000,000 m/s
671,000,000 mph
186,000 miles per second
We call this value c
22

c=V*
C is constant throughout the universe, as long as light
is in a vacuum.
When it is in other materials, c can change, but can
never be larger than its value in a vacuum.
Since c is constant, all of E-M waves will have a
corresponding frequency to go along with their
wavelength.

23

c = V * V = c /
Lets find the corresponding frequency ranges for a few of the
groups of E-M waves.

2003 Mike Maloney

24

2- particle properties
Electromagnetic radiation of light can be viewed as a stream of discrete
wave packets of distinct particles called photons.
The energy E of photon depends upon the frequency of the radiation

E = hV
Therefore:
h = Plancks constant (6.626 x 10-34 J s)
V = frequency of the radiation (most common units = cm-1
Energy is inversely proportional to wavelength

Range of Behavior
Electromagnetic waves are
characterized by their wavelength
or frequency.

Linked by the speed of light

The frequency is associated with

an energy.
High frequency is high energy

The common unit is the electronvolt (eV).

Work done by 1 V potential on
one electron

E h hc
hc = 1.240 keV nm

The photoelectric effect When light shines on

metals, electrons (photoelectrons) are ejected from
their surface.
A certain frequency has to be achieved or the effect does not
work
Red light will not cause
electrons to eject!

Electromagnetic Radiation
Traditional upper boundaries for types of EM radiation:
Radio waves
Microwaves
Infrared
Visible light
Ultraviolet light
X-rays
Gamma rays

(m)
1
110-3
0.7510-6
0.410-6
1.210-8
1.410-11

(Hz)
E (eV)
3108
1.2410-6
31011
1.2410-3
41014
1.65
7.51014
3.1
2.41016
1102
31019
1.2105
(highest energy)

Internal Energy of Molecules

Etotal=Etrans+Eelec+Evib+Erot+Enucl
Eelec = Electronic transitions (UV, X-ray)
Evib = Vibrational transitions (Infrared)
Erot = Rotational transitions (Microwave)
Enucl= Nucleus spin (NMR: Nuclear Magnetic Resonance)

or
(MRI: Magnetic Resonance Imaging)

Visible Spectrums
(Color)

Color/Colour??
Light can either be transmitted/ Reflected/
absorbed by the substances /compounds

Light 100% Transmitted = Transparance

Light 100% Absorbed = Black
Light 100% Reflect ed = White
Light Partly absorbed, partly refected, partly
transmitted = Color

Spectroscopy &
pectrophotometry ?
Spectroscopes/Spectroscopy measure
electromagnetic emission
Spectrophotometers measure
electromagnetic absorption

Colors & Wavelengths

Visible Light

COLOR

WAVELENGTH ( in nm)

Ultraviolet

< 380

Violet

380 435

Blue

436 480

Greenish-blue

481 490

Bluish-green

491 500

Green

501 560

Yellowish-green

561 580

Yellow

581 595

Orange

596 650

Red

651 780

Near Infrared

> 780

Complimentary Colors

Complimentary Colors

Complimentary Colors

Complimentary Colors

Complimentary Colors

Chromophor
Functional Groups

Chemical Structure

Aromatic Compounds

Benzene, Toluene, Xilene, Nafthalene,

Antrasene, etc

Phenol derivatives

Phenol, ortho, metha, para

Nitrogen Group

Piridine, Qinoline, Korbazolum, etc

Nitroso

NO atau (-N-OH)

Nitro

NO2 ata (NN-OOH)

Grup Azo

-NN-

Grup Etilen

-C=C-

Grup Karbonil

-C O -

Grup Karbon Nitrogen

-C=NH ; CH=N-

Grup Karbon Sulfur

-C=S ; -C-S-SC-

Energy Transitions

Second Excitation state

E2
hf2
E1

First Excitation state

hf1

E0

Ground state

Electron transitions

Vacuum UV or Far UV
(<190 nm )

UV/VIS

 Transitions
An electron in a bonding s orbital is excited to the
corresponding anti bonding orbital.
The energy required is large.
For example, methane (which has only C-H bonds, and
can only undergo transitions) shows an
absorbance maximum at 125 nm.
Absorption maxima due to transitions are not seen
in typical UV-VIS spectra (200 - 700 nm)

n Transitions
Saturated compounds containing atoms with lone
pairs (non-bonding electrons) are capable of n
transitions.
These transitions usually need less energy than
transitions.
They can be initiated by light whose wavelength is in
the range 150 - 250 nm.
The number of organic functional groups with n
peaks in the UV region is small.

n and Transitions
Most absorption spectroscopy of organic
compounds is based on transitions of n or
electrons to the excited state.
These transitions fall in an experimentally
convenient region of the spectrum (200 - 700 nm).
These transitions need an unsaturated group in the
molecule to provide the electrons.

Chromophore

Excitation

max, nm

Solvent

C=C

171

hexane

C=O

n*
*

290
180

hexane
hexane

N=O

n*
*

275
200

ethanol
ethanol

C-X
X=Br, I

n*
n*

205
255

hexane
hexane

Orbital Spin States

For some of the excited states, there are states with a
pair of electrons having their spins parallel (in 2
orbitals), leading to total spin of 1 and multiplicities
of 3.
Total Spin

Multiplicities

Orbital Spin States

Singlet state (S):Most molecules have ground state with
all electron spin paired and most excited state also have
electron spin all paired, even though they may be one
electron each lying in two different orbital.
Such states have zero total spin and spin multiplicities of 1,
are called singlet (S) states.

Total Spin

Multiplicities

Orbital Spin States

For Triplet state (T): Under the influence
of external field, there are three values (i.e. 3
energy states) of +1, 0, -1 times the angular
momentum.
Such states are called Triplet states (T).

Selection Rules of electronic

transition

Electronic transitions may be classed as intense or weak

according to the magnitude of max that corresponds to
allowed or forbidden transition as governed by the
following selection rules of electronic transition:
Spin selection rule: there should be no change in spin
orientation or no spin inversion during these transitions.
Thus, SS, TT, are allowed,
but ST, TS, are forbidden.
(S=0 transition allowed)

Lambert Beer Law

Spectrophotometric Analysis
Spectrophotometric techniques are used to
measure the concentration of solutes in solution by
measuring the amount of light that is absorbed by the
solution in a cuvette placed in the spectrophotometer.
The spectrophotometer can measure the amount of
light or electromagnetic radiation (of certain frequency)
transmitted or absored by the solution.

Lambert - Beers Law

source

cuvette

slit

detector

P0

Transmittance
I
dI
T

kcdb
I0
I0

I0

I0

b
dI
kc db
0
I0

I
I
ln( ) kbc 2.303 log( )
I0
I0

I
log( ) log T A bc
I0
k

2.303

Quantitative Analysis
Beers Law
A=ebc
e: the molar absorptivity (L mol-1 cm-1)
b: the path length of the sample
c :the concentration of the compound in
solution, expressed in mol L-1

The absorption process

How does matter absorb radiation
When polychromatic light (white light), which contains the whole
spectrum of wavelengths in visible region, is passed though an object
will absorb certain of the wavelengths, leaving the unabsorbed
wavelengths to be transmitted. These residual transmitted wavelengths
will be seen as a color. This color is complementary to the absorbed
colors.

Absorption is a process in which chemical species (atom, ion or molecule) in a

transparent medium selectively attenuate certain frequencies of EMR

Absorption spectrum is a plot of the amount of light absorbed by a

sample as a function of wavelength.
At room temperature most substance are in their lowest energy or
ground state. When an atom, molecule or ion absorbs EMR it is
promoted to higher energy states or excited states.
The excited state is a transition one and the species soon looses the
energy it gained and returns to its ground state by relaxation process
either as heat of collision or sometimes emits radiation of specific
wavelength.

 When a molecule interacts with photons of UV or VIS

radiation excitation of electrons takes place to higher
electronic energy level at any of its vibrational level.
Eex-Eg= h of the photon absorbed.
UV / VIS radiation cause electronic transition which is accompanied
by vibrational and rotational.
If the compound subjected to IR radiation vibrational and rotational
transitions in ground state occure.
Rotational transitions alone can be brought about by microwave.

 Ultraviolt and visible radiations have sufficient energy to cause

trnsitions of the outermost or valence electrons.
If large amount of energy is absorbed by certain substance, bonds may
be ruptured and new compounds are formed photolysis.
This may occur upon absorption of far Ultraviolt as its energy is
sufficiently high to exceed the energy of formation of certain bonds.
The total energy of a molecule is given by
Etotal=Eelectronic+Evibrational +Erotational

A = -logT = log(P0/P) = bc
T = P/P0
Compound x has a unique at different
wavelengths
Works for monochromatic light
%T= 100xT
: is molar absorptivity and it is unique for a
given compound ( L.mol-1. cm-1)
b : is path length (cm)
c : concentration (M)

Instruments for measuring the absorption of U.V. or visible radiation are made up of the
following components;

INSTRUMENTS
SOURCES (UV AND VISIBLE)
WAVELENGTH SELECTOR (MONOCHROMATOR)
A SAMPLE CELL
DETECTOR
A DEVICE TO READ OUT THE RESPONSE OF THE DETECTOR

schematic diagram of a single-beam UV-Vis. spectrophotometer

(Abs)
Exponential curve relationship

(Abs)
Linear curve relationship

Absorbance

concentration

0.161

0.332

0.493

0.656

0.806

1.191

7.5

1.554

10

Calibration Curve
- Linear calibration curves
- non linear calibration curves
Linear Calibration curve of Beers law

Deviations from Beers law

1- at high concentrations
2- at low concentrations

Applications

Absorption of Radiant Energy

Wave-particle Nature of Radiant energy:
Light and other forms of radiant energy have a dual nature (wave
and particle)
Electromagnetic radiation is a type of energy that is transmitted
through space as a transverse wave at enormous velocity.
It takes numerous forms known as electromagnetic spectrum.
The electromagnetic
spectrum include gamma ray, X-ray,
ultraviolet (UV), visible, infrared (IR), microwave and radio-wave
radiation.