CHAPTER FIVE

UV/VISIBLE SPECTROSCOPY

UV/Vis spectrophotometer

1
 Introduction

• Ultraviolet and visible spectrometers have been in general

use for the last 35 years and over this period have become
the most important analytical instrument in the modern day
laboratory.
• In many applications other techniques could be employed
but none rival UV-Visible spectrometry for its simplicity,
versatility, speed, accuracy and cost-effectiveness.

2
 Cont…

• Most analytical measurements in the UV region occurs b/n

200 and 380 nm. The visible region occurs b/n 380 and 760
nm. Spectrometers of this type are called UV/Visible
spectrometers.
• UV/VIS measure absorption or transmittance like AAS &
AES.
• Used for identification of inorganic and organic species.
• Ultraviolet radiation stimulates molecular vibrations and
electronic transitions.

3
 Principle

• When UV or visible light passes through the compound with multiple

bonds, the transition of valence electrons from lower to higher energy
level occurs and gives rise to UV-Visible spectroscopy.

• Molecules containing π-electrons or non-bonding electrons (n-

electrons) can absorb the energy in the form of ultraviolet or visible light
to excite these electrons to higher anti-bonding molecular orbitals. 

• The more easily excited the electrons (i.e. lower energy gap between the
HOMO and the LUMO), the longer the wavelength of light it can absorb.

4
 Cont…
Electronic transitions
• Electronic transitions occur when the molecule absorbs energy
Molecular Orbital Theory

5
Example1

Example 2

6
 1 σ → σ* transition

• σ electron from orbital is excited to corresponding anti-bonding

orbital σ*.

• The energy required is large for this transition.

• e.g. Methane (CH4) has C-H bond only and can undergo σ → σ*
transition and shows absorbance maxima at 125 nm.

7
 2 π → π* transition

• π electron in a bonding orbital is excited to corresponding

anti-bonding orbital π*.

• Compounds containing multiple bonds like alkenes, alkynes,

carbonyl, nitriles, aromatic compounds, etc undergo π → π*
transitions.

• e.g. Alkenes generally absorb in the region 170 to 205 nm.

8
 3 n → σ* transition
• Saturated compounds containing atoms with lone pair of
electrons like O, N, S and halogens are capable of n → σ*
transition.

• These transitions usually requires less energy than σ → σ*

transitions.

• The number of organic functional groups with n → σ* peaks

in UV region is small (150 – 250 nm).

9
 4 n → π* transition
• An electron from non-bonding orbital is promoted to anti-
bonding π* orbital.

• Compounds containing double bond involving hetero atoms

(C=O, C≡N, N=O) undergo such transitions.

• n → π* transitions require minimum energy and show

absorption at longer wavelength around 300 nm.

10
Terms used in UV / Visible Spectroscopy

Chromophore
• The part of a molecule responsible for imparting color, are
called as chromospheres.
OR
• The functional groups containing multiple bonds capable of
absorbing radiations above 200 nm due to n → π* & π → π*
transitions.
• e.g. N=O, C=O, C=N, C≡N, C=C, C=S, etc

11
Cont…

To interpretate UV – visible spectrum following points should be

noted:

1. Non-conjugated alkenes show an intense absorption below

200nm & are therefore inaccessible to UV
spectrophotometer.

2. Non-conjugated carbonyl group compound give a weak

absorption band in the 200 - 300 nm region.

12
Cont…
O
e.g. C
Acetone which has λmax = 279 nm
H 3C CH3

• When double bonds are conjugated in a compound λmax is

shifted to longer wavelength.
e.g. 1,5 - hexadiene has λmax = 178 nm

2,4 - hexadiene has λmax = 227 nm

CH2 CH3
H2C H3C

13
Cont…

3. Conjugation of C=C and carbonyl group shifts the λmax of

both groups to longer wavelength.

e.g. Ethylene has λmax = 171 nm

H2C CH2
O
C
Acetone has λmax = 279 nm H 3C CH3
O

Crotonaldehyde has λmax = 290 nm H2C C

CH3
14
List of common chromophores and their transitions

15
Auxochrome

• The functional groups attached to a chromophore which modifies the

ability of the chromophore to absorb light , altering the wavelength or
intensity of absorption.
OR
• The functional group with non-bonding electrons that does not absorb
radiation in near UV region but when attached to a chromophore alters
the wavelength & intensity of absorption.

16
Cont…

e.g. Benzene λmax = 255 nm

OH

Phenol λmax = 270 nm

NH2

Aniline λmax = 280 nm

17
18
 Radiation Sources

• Radiation Sources produce light over a continuum of wavelengths in

molecular absorption(UV/VIS).
• Example;
 Deuterium and hydrogen lamps (160-380 nm)
 Tungsten (W) filament lamp (320-2400 nm)
 Xenon (Xe) arc lamps (200-1000 nm)
 Light emitting diodes (LED) for the visible wavelengths

19
Monochromator: disperse the radiation according to wavelength and allow
selected wavelengths to illuminate the sample.
Example: diffraction grating and prism (complex and expensive)
Beam splitter: split the beam of light of each wavelength into two halves of
equal intensities, one goes to the test sample and other to the
blank/reference sample.
Sample containers: Samples for UV/Visible spectroscopy can be solids,
liquids, or gases. Different types of holders have been
designed for these sample types.
1. Liquid and Gas Cells/Sample Holders/
Cuvettes
• Plastic, Glass, Quartz, Fused silica etc…
2. Solid Sample Holders
• Film holder 20
Detectors
• It detects the intensities of the transmitted light coming out of the cells
and compares for each wavelength and generates current proportional to
the difference in the intensities of the sample and blank.
• Modern detectors use a sensitive transducer to convert a signal consisting
of photons into an easily measured electrical signal.
Example

21
 Signal Processor and Output Device
• Modern spectrometers have digital readouts & integrated
microprocessors for purposes of calibration, control of the wavelength
drive, slit adjustment, and sometimes for the interchange of the light
sources.
• They also include interfaces to personal computers sometimes called
data stations. e.g. Analog-to-digital converters
Recorder
• It records an UV/Visible spectrograph with absorbance against the
wavelength.

22
23
Applications of UV/Visible Spectroscopy
• Qualitative & Quantitative Analysis:
– It is used for characterizing aromatic compounds and conjugated
olefins.
– It can be used to find out molar concentration of the solute under
study.
• Detection of impurities:
– It is one of the important method to detect impurities in organic
solvents.
• Detection of isomers are possible.
• Detection of functional group - presence or absence of chromophore.
• Can determine the extent of conjugation in polyenes (more the
unsaturation, more the shift towards longer wavelength).
• Determination of molecular weight using Beer’s law.

24