UV-VISIBLE SPECTROSCOPY

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 Electromagnetic radiation
Energy of an EMR
Electromagnetic spectrum & it’s regions
 Spectroscopy & it’s types
 Colorimetry
UV-spectroscopy
 Electronic transitions
Terms used in UV-Visible spectroscopy
Absorption band & it’s types
Beer lambert’s law & it’s deviations
⓫ Reference
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Electromagnetic radiation is a type of energy that
is transmitted through space at enormous
velocities.

Radiant energy has wave nature and being associated

with electric as well as magnetic field,these
radiations are called electromagnetic radiation.

Electromagnetic radiation has it’s origin in atomic

and molecular processes.
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The field may be represented as electric and
magnetic vectors oscillating in mutually
perpendicular planes.

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The energy of an EMR can be given by the
following equation:
E=hµ

Where E= Energy of radiation

h=Planck’s constant
µ=Frequency of radiation

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Frequency(µ)= c/λ
Where c=velocity of light in vacuum
λ= wavelength

:-Hence, E=hµ

E=hc/λ

Therefore,energy of a radiation depends upon frequency

and wavelength of radiation.

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The arrangement obtained by arranging various types
of electromagnetic waves or radiations in order of
their increasing wavelegth or decreasing frequencies is
called electromagnetic spectrum.

The electromagnetic spectrum is divided into a

number of regions; these are artificial divisions in the
sense that they have been defined solely as a result of
differences in the instrumentation required for
producing and detecting radiation of a given frequency
range.
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Ultraviolet: 190~400nm
Violet: 400 - 420 nm
Indigo 420 - 440 nm
:Blue: 440 - 490 nm
Green 490 - 570 nm
:Yellow: 570 - 585 nm
Orange: 585 - 620 nm
Red: 620 - 780
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nm
spectroscop
y
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SPECTROSCOPY

Spectroscopy is the measurement and

interpretation of electromagnetic radiation
absorbed or emitted when the molecules
or atoms or ions of a sample moves from
one energy state to another energy state

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1) Atomic spectroscopy :
Here,the changes in energy takes
place at atomic level.
Eg: Atomic absorption spectroscopy, Flame
photometry

2)Molecular spectroscopy :
Here,the changes in energy takes
place at molecular level.
Eg: UV spectroscopy,colorimetry,infra red
spectroscopy
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 Absorption spectrophotometry can be
defined as the measurement of absorption of radiant
energy by various substances.It involves the
measurement of absorptive capacity for radiant
energy in the visible,UV and IR regions of the
spectrum.

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o λ- 400-800nm
o Coloured substance absorbs light of different λ in
different manner and hence get an absorption
curve
o The λ at which maximum absorption takes place
is called as λmax
o λmax is characteristic for every coloured substance
o On plotting a graph of concentration v/s
absorbance,we get a calibration curve that is
useful in determining the concentration or
amount of a drug substance in the given sample 14
solution.
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 UV spectroscopy is concerned with the study of absorption
of uv radiation which ranges from 200-400nm.
Valence electrons absorb the energy thereby
molecules undergoes transition from ground state to
excited state.
This absorption is characteristic and depends on
the nature of electrons present.
Types of electrons
σ electrons: in saturated compounds
π in unsaturated compounds
electrons: in non bonded electrons
n electrons:
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 1) σ-σ*
 σ electron from orbital is excited to
corresponding anti-bonding orbital σ*.

The energy required is large for this transition.

The organic compounds in which all the valence shell

electrons are involved in the formation of σ bond do
not show absorption in normal uv region (200-
400nm)

This transition is observed with saturated compounds.

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 Eg: Methane(CH₄) has C-H bond only and can undergo σ-
σ* transition and shows absorption maxima at 122nm.

 The usual spectroscopic technique cannot be used below

200 nm.

 To study this high energy transition,the entire

region should be evacuated (Vacuum uv region)

 Here,the excitation ocuurs with net retention of

electronic spin

 This region is less informative

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 2) π-π*
 π electron in a bonding orbital is excited
to corresponding anti-bonding orbital π*.

 Energy required is less when compared to n-

σ*

 Compounds containing multiple bonds

like
alkenes,alkynes,carbonyls,nitriles,aromatic
compounds etc undergo π-π* transition.

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Eg:Alkenes generally absorb in the region
Absorption usually occurs in the ordinary
uv spectrophotometer

Absorption bands in unconjugated alkenes (170-

190nm)

Absorption bands in carbonyls (180 nm)

Introduction of alkyl group in olefinic linkage

produces bathochromic shift
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 3) n-σ*
Saturated compounds containing one hetero
atom with unshared pair of electrons(n) like
O,N,S and halogens are capable of n-σ*
transition.

These transition require less energy than σ-

σ* transition.

In saturated alkyl halides, the energy required for

transition decrease with increase in the size of
halogen atom (or decrease in electronegativity)
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 Eg:Methyl chloride has a λmax of
173nm. Methyl iodide has a λmax
of 258nm.

 This type of transition is very sensitive to

hydrogen bonding
Eg: Alcohol & amines

 Hydrogen bonding shift the uv absorptions

to shorter wavelength.
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 4) n-π*

An electron from non-bonding orbital is promoted

to anti-bonding π* orbital.

 Compounds containing double bonds

involving hetero atoms(C=O,N=O) undergo
such type of transitions.

This transition require minimum energy out of

all transitions and shows absorption band at
longer wavelength around 300nm.
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Eg:Saturated aldehydes shows both type of transitions
(n-π*, π-π*) at {low energy and high energy}
around 290 and 180 nm.

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TERMS USED IN UV-
VISIBLE
SPECTROSCOPY

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 Chromophore is defined as the nucleus or any
isolated covalently bonded group responsible for
the absorption of light radiation.

 Any group which exhibits absorption of

electromagnetic radiations in the visible or
ultraviolet region.
C=C , C=O ,NO2 etc

 Some of the important chromophores are

carbonyls,acids,esters,nitrile,ethylenic
groups. 27
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 These are co-ordinatively saturated or un-saturated
groups which themselves do not absorb
radiations,but when present alongwith a
chromophore enhances the absorbing properties of
chromophore.

 Also known as colour enhancing group.

 All auxochromes have one or more non-bonding
pair of electrons.
-NH2 ,-OH ,-OR,-COOH etc
 It extend the conjugation of a chromophore by
sharing the non-bonding electrons. 29
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 ABSORPTION &
INTENSITY
SHIFTS

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 1) Bathochromic shift(red shift)

When the absorption maxima(λmax)of a

compound shifts to longer wavelength,it is known
as bathochromic shift or red shift.

The effect is due to the presence of auxochrome or

by change of solvent.

Eg: The n-π* transition for carbonyl compounds

experiences bathochromic shift when the polarity
of solvent is decreased.
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 2) Hypsochromic shift(blue shift)

When the absorption maxima (λmax) of a

compound shifts to a shorter wavelength,it is known
as hypsochromic shift or blue shift.

The effect is due to the presence of a group

causes removal of conjugation or by change of
solvent.

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Eg:

Aniline shows blue shift in acidic medium since it

loses conjugation. Aniline(280nm) & Anilinium ion
(- 203nm).

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3) Hyperchromic effect

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4) Hypochromic effect

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SHIFTS & EFFECTS

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ABSORPTION
BANDS

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 The spectrum consist of sharp peaks and each peak
will correspond to the promotion of electron from one
electronic level to another.

 During promotion,the electron moves from a given

vibrational and rotational level within one electronic mode
to the other within the next electronic mode.

 Thus,there will be a large no of possible transitions

 Hence,not just one but a large no. of wavelengths

which are close enough will be absorbed resulting in
the formation of bands 39
1) K –Band
 K-Bands originate due to π-π* transition from a
compound containing a conjugated system

 Such type of bands arise in compounds

like dienes,polyenes and enones etc.
Compound Transition λmax(nm) εmax

Acetophenone π-π* 240 13,000

1,3-butadiene π-π* 217 21,000

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 R-Band transition originate due to n-π* transition of
a single chromophoric group and having atleast one
lone pair of electrons on the hetero atom

 These are less intense with εmax value below 100

Compound Transition λmax(nm) εmax

Acetone n-π* 270 15

Acetaldehyde n-π* 293 12

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 Such type of bands arise due to π-π* transition
in aromatic or hetero-aromatic molecules.

 Benzene shows absorption peaks between 230-

270nm.when a chromophoric group is attached to
the benzene ring ,the B-Bands are observed at longer
wavelengths than the more intense K-Bands.
Compound Transition λmax(nm) εmax

Benzene π-π* 255 215

Phenol π- π* 270 1450

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 E-Band originate due to the electronic transitions in
the benzenoid systems of three ethylenic bonds
which are in closed cyclic conjugation.
 These are further characterized as E1 and E2 bands
 E1 band which appear at shorter wavelength is
usually more intense than the E2 band for the same
compound which appears at longer wavelength.
Compound E1 Band E1 Band E2 Band E2 Band

λmax(nm) εmax λmax(nm) εmax

Benzene 184 50,000 204 79,000

Napthalene 221 133,000 286 9,300

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BEER’S LAW
According to this law,when a beam of monochromatic
radiation is passed through a solution of absorbing species,the
intensity of beam of monochromatic light decreases
exponentially with increase in concentration of absorbing species

-dI/dc α I
LAMBERT’S LAW
Lambert’s law states that the rate of decrease of
intensity of monochromatic light with the thickness of the
medium is directly proportional to the intensity of incident
light.
-dI/dt α I

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According to beer’s law,
-dI α I
dc
The decrease in the intensity of light (I) with
concentration(c) is proportional to intensity of
incident light(I)
-dI = K.I { removing & introducing the
dc constant of proportionality
“K”}
-dI = K.dc { rearranging terms}
I
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On Integrating the
equation,
∫ ∫
- dI = K. dc
I
-ln I = K.c + b {b constant of integration}
When concentration =0, there is no absorbance,Hence
I=I₀ Substituting in equation 
-ln I₀ = K*0+b
-ln I₀ = b
Substituting the value of b in equation
-ln I = K.c-ln I₀
ln I₀ - ln I = Kc { since log A-logB = log A }
B
ln I₀ =
Kc I
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 I₀ = ekc { removing natural log }
I
I = e- {making inverse on both sides}
I₀ kc

I = I₀e-kc  [equation for beer’s law]

According to lambert’s law,

-dI α I
dt
This eqn can be simplified by replacing ‘c’ with ‘t’ in

I = I₀e-kt 

Eqn  &  can be combined to get I = I₀e-kct 48

Converting natural log to base 10 & K= k x
0.4343
I = ₁₀-kct { rearranging terms }
I₀

I₀ = { inverse on both sides}

₁₀kct I
Taking log on both
sideslog
, I₀ = Kct 
I
Here, Transmittance T = I₀ , Absorbance,A = log 1
I

T 49
 A = log 1
T
A = log 1 { Since T = I₀
} I/I₀ I
A =log I₀ 
I
Using eqn  &  , since A =log I₀ log I₀ = Kct ,
and I
A= Kct I
Instead of K, we can use ε

A = εct { Mathematical eqn for beer

lambert’s
law}
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There are 3 types of deviations usually observed

A)The real limitation of the law is that the beer’s law

is successful in describing the absorption behaviour
of dilute solutions only.

B) CHEMICAL DEVIATIONS:

 Association of molecules
This can be explained by taking the examples of
methylene blue at small concentration(10‾⁵ molar)
and at concentration above 10‾⁵molar.

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 Dissociation of molecules
This can be explained by the fact that
dichromate ions posses their maximum absorbance
at 450nm which is orange in colour .But upon
dilution,it will be dissociated to chromate ions
having maximum absorbance at 410nm which is
yellow in colour.

This law is not valid in case if the absorbing material

is coagulated into a small number of large units.

This law shows deviation if the absorbing material

at the required wavelength contains presence of
impurities.
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 This law is not applicable in case of suspension.

C)INSTRUMENTAL DEVIATIONS:

 Strict adherence of an absorbing system to this law is

observed only when the radiation used is monochromatic.

 Stray radiation,slit width also causes deviation.

Hence,the reasons for the deviation depends on

environment such as temperature,pressure,solvent,
refractive index of the sample

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Absorbance
0.800
λ=430nm
0.600 λ=570nm

0.400

0.200

0.00
0.00 4.00 8.00 12.00 16.00
concentration 56
Absorbance
0.800 ε=1000 ε=1500

0.600
ε=1750
0.400

0.200

0.00 2.0 4.0 6.0 8.0 10.0

concentration 57
Absorbance 0.0%
 2.0 0.2%

 1%

 1.0 5%

 0
 2.5 5.0 7.5 10 concentration
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