Spectroscopy is the most powerful tool available for the study of atomic & molecular structure and is used in the analysis of a wide range of samples

1. UV-Visible Spectroscopy
Prepared by
Prof.(Dr.) Dinesh Kumar Mehta
MMCP, (MMDU), Mullana

2. Spectroscopy
• It is the branch of science that deals with the study of interaction of
matter with light.
OR
• It is the branch of science that deals with the study of interaction of
electromagnetic radiation with matter.
Spectroscopy is the most powerful tool available for the study of atomic
& molecular structure and is used in the analysis of a wide range of
samples
UV-Vis spectroscopy
UV-Vis spectroscopy is an analytical technique that measures the
amount of discrete wavelengths of UV or visible light that are absorbed
by or transmitted through a sample in comparison to a reference or blank
sample.
Humans are able to see a spectrum of visible light, from approximately
380 nm, which we see as violet, to 780 nm, which we see as red.1
UV
light has wavelengths shorter than that of visible light to approximately
100 nm.

3. Ultraviolet (UV) region: The term ultraviolet means “beyond violet”
which is derived from a Latin word “Ultra” meaning beyond. Here the
radiation starts at the blue end of the visible light from 400nm and ends
at 200nm.
Visible region: The visible region of the electromagnetic spectrum
ranges between 400-780nm.
The visible light is otherwise called as white light or ordinary light and
is composed of different colors as seen in a rainbow.
A physical quantity is said to have a discrete spectrum if it takes only
distinct values, with gaps between one value and the next.
The classical example of discrete spectrum (for which the term was first
used) is the characteristic set of discrete spectral lines seen in
the emission spectrum and absorption spectrum of isolated atoms of
a chemical element, which only absorb and emit light at

4. particular wavelengths. The technique of spectroscopy is based on this
phenomenon.
Application of UV-VIS Spectroscopy
 Detection of impurities
 Qualitative analysis
 Quantitative analysis
 Single compound without chromophore
 Drugs with chromophoric reagent
 It is helps to show the relationship between different groups, it
is useful to detect the conjugation of the compounds
Ultraviolet and visible (UV-Vis) absorption spectroscopy is the
measurement of the attenuation of a beam of light after it passes through
a sample or after reflection from a sample surface. Absorption

5. measurements can be at a single wavelength or over an extended spectral
range
UV spectroscopy is an important tool in analytical chemistry. The other
name of UV (Ultra-Violet) spectroscopy is Electronic spectroscopy as it
involves the promotion of the electrons from the ground state to the higher
energy or excited state.

6. A single xenon lamp is commonly used as a high intensity light source
for both UV and visible ranges. Xenon lamps are, however, associated
with higher costs and are less stable in comparison to tungsten and
halogen lamps.
For instruments employing two lamps, a tungsten or halogen lamp is
commonly used for visible light, whilst a deuterium lamp is the common
source of UV light. As two different light sources are needed to scan
both the UV and visible wavelengths

7. Theory
Energy absorbed in the UV-visible region produces changes in the
electronic energy of the molecule resulting from transitions (ground
state to excited state) of valence electrons in the molecule.
Three distinct types of electrons are involved in organic molecules.
They are as follows:
a) σ electrons
b) π electrons
c) n electrons
Electronic Transitions
1. σ→ σ* transition:
An electron in a bonding s-orbital is excited to the corresponding anti-
bonding orbital and observed with saturated compounds.
The energy required is large.
For example, methane (which has only C-H bonds, and can only
undergo σ→ σ* transition transitions) shows an absorbance maximum at
125 nm.
2. n → σ* transition:

8. Saturated compounds containing atoms with lone pairs (non- bonding
electrons) like O, N, S and halogens are capable of n→ σ* transition.
These transitions usually need less energy than n → σ* transition.
They can be initiated by light whose wavelength is in the range 150 -
250 nm. The number of organic functional groups with n → σ* peaks in
the UV region is small.
3. π→ π* transition:
π electron in a bonding orbital is excited to corresponding anti- bonding
orbital π* and observed in conjugated compounds.
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.
4. n → π* transition:
An electron from non-bonding orbital is promoted to anti- bonding π*
orbital and required lower energy. 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.

9. Thus Energy needed for promoting an electron follows the order: σ > π>
n
Different Types of Molecular Orbitals
Bonding Orbital: They directly participate in bond formation between
atoms. Electrons of bonding orbital are represented as σ and π.
Non-Bonding Orbital: They do not participate in bond formation. Eg :
atoms such as oxygen, sulphur, nitrogen (valence electrons).
Anti-Bonding Orbital: They oppose bonding and electrons do not take
part in bonding. Eg : π* and σ*.

10. Principle of UV spectroscopy
UV spectroscopy obeys the Beer’s-Lambert law, which states that: when
a beam of monochromatic light is passed through a solution of an
absorbing substance, the rate of decrease of intensity of radiation with
thickness of the absorbing solution is proportional to the incident
radiation as well as the concentration of the solution.
The expression of Beer-Lambert law is-
A = log (I0/I) = ECb
Where, A = sample’s absorbance value at specific wavelength or
frequency
I0 = intensity of light incident upon sample cell
I = intensity of light leaving sample cell
C = molar concentration of solute in mol L-1
b = is the path length of sample cell (cm.)
E = molar absorptivity
From the Beer-Lambert law it is clear that greater the number of
molecules capable of absorbing light of a given wavelength, the greater
the extent of light absorption. This is the basic principle of UV
spectroscopy.