Spectrophotometric principle and

applications

Beer-Lambert law and its applications

To know how to use Beer-Lambert law in environmental analysis of pollutants.

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 Related Books
Sl. No Name of Book
1 Introduction to Spectroscopy
(A Guide for Students of
Organic Chemistry)
2 Fundamentals of Analytical
Chemistry
(International Edition) (6th to
9th Edition)
3 Vogel’s Text Book of
Quantitative Chemical Analysis
(New Edition)
4 Introduction to Spectroscopy
Author List
Donald L. Pavia, Gary
M. Lampman, George
S. Kriz
Douglas A. Skoog,
Donald M. West, F.
James Holler
J. Mendham, R. C.
Denney, J. D. Barnes,
M. Thomas
B. K. Sharma

2
 Chapter Outcome

• To know about basics of spectroscopy.

• To know about various types of internal
energy of matter.
• Determination of concentration, structure and
geometry of the compound in a particular
sample using spectrophotometry as
spectroscopic tool.
• Application of Beer’s law in qualitative and
quantitative analysis of sample.
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 Energy of matter
• Energy is the capacity to do work
• Kinetic and potential energy
• The kinetic energy, Ek of a body is the energy the body
possesses as a result of its motion. For a body of mass
m travelling at speed ν,

• The potential energy, Ep of a body is the energy it

possesses as a result of its position.
• If a body is at a height h above the surface of the earth,
then its potential energy is mgh, where g is a constant
called the acceleration constant of free fall.

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 An insight into the energy of matter
• The total energy/internal energy
Kinetic energy is associated with liquid
and gaseous molecules
If we avoid kinetic energy

A
C

• E (vibration) is the energy of the molecule as a whole due

to interatomic vibrations, fig A
• E (electronic) is the energy associated with the transition of
electrons within various outer orbitals of the molecule
• E(rotational) accounts for the energy associated with
rotation of the molecule about its center of gravity, fig C

5
 Electronic Energy (Eelectronic) Line
spectrum

Energy level diagram of a multielectron atom Absorption transitions of helium

Atomic emission spectrum of helium 6

What is spectroscopy?
• The interactions of radiation and matter are the subject of the
science called spectroscopy.

• Spectroscopy is referred to a branch of science in which light (that

is visible radiation) was resolved into its component wavelengths to
produce spectra.

Importance of spectroscopy
• Spectra of course played a vital role in the development modern
atomic theory.

• In addition, spectroscopy has proved to be a powerful tool for the

elucidation of molecular structure as well as for the qualitative and
quantitative determination of both inorganic and organic
compounds.
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 What is spectroscopy ?
• With the passage of time, the meaning of spectroscopy has
become broadened to include studies not only with light but
also with other types of electromagnetic radiation, such as X-
ray, ultraviolet, infrared, microwave, and radiofrequency
radiation.

• Indeed, current usage extends the meaning of spectroscopic

methods still further to include techniques that do not involve
electromagnetic radiation. Examples of the last include
acoustic, mass, and electron spectroscopy.

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Electromagnetic radiation
• Electromagnetic radiation is a form of energy that is transmitted through
space at enormous velocities.
• Electromagnetic radiation consists of a wave of electric and magnetic
fields perpendicular to the direction of propagation (as in figure the x-
direction) and mutually perpendicular to each other. As shown in figure, a
plane polarized wave, with the electric and magnetic fields oscillating in
the xy- and xz-planes, respectively.

• Electromagnetic radiation can be described as a wave with properties like

wavelength, frequency, velocity, and amplitude.

• The wave model fails to account for phenomena associated with the
absorption and emission of radiant energy.

• Electromagnetic radiation can be treated as discrete packets of energy or

particles called photons or quanta.

• The energy of a photon is directly proportional to its frequency .

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 Wave
 Waves are disturbances that travel through space with finite velocity.

 Examples include the collective motion of water molecules in ocean waves

and gas particles in sound waves .

 Waves can be characterized by a wave equation, a differential equation that

describes the motion of the wave in space and time, .

 Harmonic waves are waves with displacements that can be expressed as

the sine and cosine functions.

 These concepts are used in classical physics to describe the wave character
of electromagnetic radiation

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 Wave property of electromagnetic radiation

In (a), a plane-polarized wave is shown propagating along the x

axis. The electric field oscillates in a plane perpendicular to the
magnetic field. If the radiation were unpolarized, a component
of the electric field would be seen in all planes.

In (b), only the electric field oscillations are shown. The

amplitude of the wave is the length of the electric field vector at
the wave maximum, while the wavelength is the distance
between successive maxima.
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 Sinusoidal wave
 Sinusoidal signals are smoothly varying;
there are no abrupt changes in amplitude.
The rate at which the amplitude changes
(we call this the slope) is not constant. In
fact, the slope of a sinusoid varies in a
sinusoidal fashion.

 Sinusoidal signals occur in repeating

cycles. The number of cycles that occur in
one second is equal to the frequency of the
signal in hertz (abbreviated Hz).

 Real-life sinusoidal signals have

amplitudes that vary with respect to time;
however, we can also describe the horizontal
progression of the signal using degrees. The
completion of one full cycle corresponds to
an angle that varies from 0° to 360°, and the
halfway point of the cycle occurs at 180°.
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 Sinusoidal wave
Rota Sin Cosine
tion
(°)
0 0 1.0
30 0.5 0.87
45 0.71 0.71
Fig. B 60 0.87 0.5
Fig. A
90 1.0 1.0
A: linear displacement through cyclic rotation 120 0.87 -0.5
B: The graphs of the sine (solid red) and 180 0 -1.0
cosine (dotted blue) Functions are sinusoids 270 -1.0 0
of different phases 360 0 0.98

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Wave characteristics E = hν
The period (p)
of an
electromagnetic
• The amplitude (A) of an electromagnetic wave is a vector quantity that provides a measure
of the electric or magnetic field strength at a maximum in the wave.
wave is the time
in seconds for
successive
maxima or
minima to pass
a point in space.
The frequency
(ν) of an
electromagnetic
• The period (p) of an electromagnetic wave is the time in seconds for successive
wave is the
number of
oscillations that
occur in one
maxima or minima to pass a point in space.
Frequency is

• The frequency (ν, nu) of an electromagnetic wave is the number of oscillations

second (1/p).
expressed in
hertz (Hz), which
corresponds to

that occur in one second.

one cycle per
second, that is,
1 Hz = 1 s-1. The
frequency does
not change as it
passes through
Velocity, v, of
different media.
• The unit of frequency is the hertz (Hz), which corresponds to one cycle per second, that is, 1 Hz = 1 s-1.
The frequency of a beam of electromagnetic radiation does not change as it passes through different
the wave front
through a
medium
depends on
both the
medium and the
media.
• Velocity (v) of the wave front through a medium depends on both the medium
frequency.

Velocity and the

wavelength
depend on the
medium.
and the frequency
• Velocity and the wavelength depends on the medium
Wavelength, λ is
the linear
• Wavelength, λ is the linear distance between successive maxima or minima in a
distance
between two
maxima or
minima
wave
• Radiation velocity and wavelength both decrease as the radiation passes from a
vacuum or from air to a denser medium. Frequency remains constant.

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Regions of electromagnetic radiation

Regions of the UV, Visible, and IR Spectrum

Region Wavelength Range
UV 180–380 nm
Visible 380–780 nm
Near IR 0.78–2.5 µm
Mid IR 2.5–50 µm 15
The regions of the electromagnetic spectrum used in
spectroscopy and their interaction with the analyte.

Figure 24.3: Interaction of an analyte with electromagnetic radiation can result in the
types of changes shown. Changes in electron distributions occur in the UV/visible
region. The wavenumber, wavelength, frequency, and energy are characteristics that
describe electromagnetic radiation. (From C. N. Banwell, Fundamentals of Molecular
Spectroscopy, 3rd ed., New York; McGraw-Hill, 1983, p. 7. 16
 Wavelength variation in various
regions electromagnetic radiation
Wavelength Units for Various Spectral Regions
Region λ, Unit Definition
X-ray Angstrom, Å 10-10 m
Ultraviolet/Visible Nanometer, nm 10-9 m
Infrared Micrometer, μm 10-6 m

Calculate the wavenumber of a beam of infrared radiation with a

wavelength of 5.00 µm.
Solution

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Modes of molecular Vibration

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 Vibration modes of H2O

Symmetric mode of stretching

In plane bending mode

Asymmetric stretching mode

No. Of vibrations = 3N-6 = 3
For non-linear molecule
The three normal mode of vibration of H2O. N = No. of atoms

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Spectroscopic measurement principle

hν
M is in ground state
M* is an excited state, excitation

Energy level diagram

Heat is a form of electromagnetic radiation released

Figure 24-5: Po > P

(a), radiation of incident radiant power P0 can be absorbed by the analyte, resulting in a transmitted beam of lower radiant
power P. For absorption to occur the energy of the incident beam must correspond to one of the energy differences shown in
(b). The resulting absorption spectrum is shown in (c). 20
Emission and chemiluminescence spectroscopy
The term emission spectroscopy
usually refers to methods in which the
stimulus is heat or electrical energy.
chemiluminescence spectroscopy
refers to excitation of the analyte by a
chemical reaction.
In both cases, measurement of the
radiant power emitted as the analyte
returns to the ground state can give
information about its identity and
concentration.
Figure 24-4: Emission and The results of such a measurement
chemiluminescence spectroscopy. The are often expressed graphically by a
analyte is stimulated by applying heat spectrum, which is a which is a plot of
or electrical energy or by a chemical some function of attenuation of a
reaction. beam of radiation versus wavelength,
frequency, or wavenumber or
A plot of the emitted radiation as a
function of frequency or wavelength .
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Photoluminescence spectroscopy
• What is important to us is that some of the incident
radiation can be absorbed and promote some of the
analyte species to an excited state, as shown in Figure 24-
5.
• In absorption spectroscopy, absorption of photon/light is
measured as a function of wavelength. Absorption
measurements can give both qualitative and quantitative
information about the sample.
• In photoluminescence spectroscopy (see Figure 24-6), the
emission of photons is measured following absorption.
The most important forms of photoluminescence for
analytical purposes are fluorescence and
phosphorescence spectroscopy.
• The major distinction between fluorescence and
phosphorescence is the time scale of emission with
fluorescence being prompt and phosphorescence being
Figure 24-6 delayed and also related with spin change during
emission.
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