Spectroscopy

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Ultra Violet Spectroscopy
Prepared by:
Prof. Santosh Waghmare
JSPM’’s
Charak College of Pharmacy And Research
Wagholi, Pune

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• Spectroscopy- is the science that deals with
the interactions of various types of
radiation(EMR)with matter.
• Each type of EMR has both the properties of
wave as well as a particle.
• EMR can be described as a wave occurring
simultaneously in electrical and magnetic
fields and it is also described to consist of
particles called quanta or photons.

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Spectroscopy – Radiation Terminology
• Wavelength (λ) - length between two equivalent
points on successive waves
• Wavenumber – the number of waves in a unit of
length or distance per cycle - reciprocal of
the wavelength
• Frequency (ν) – is the number of oscillations of the
field per second (Hz)
• Velocity (c) – independent of wavelength – in
vacuum is 3.00 x 1010 cm/s (3.00 x 108 m/s)
• Photon (quanta) – quantum mechanics nature of
light to explain photoelectric effect

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Regions Wavelength Frequency in
wave numbers
Energy,Kcal/
mole
Cosmic rays 5 x 10 –5 nm
Gamma rays 10-3 – 0.14 nm
X rays 0.01- 15 nm
Far UV 15- 200 nm 666,667 –
50,000
1,907-143
UV 200- 400 nm 50,000-20,000 143-71.5
Visible 400 – 800 nm 25,000-12,500 71.5-35.7
Near IR 0.8 – 2.5 µm 12,500-4000 35.7-11.4
Mid IR 2.5 – 25 µm 4000-400 11.4-1.14
Far IR 0.025- 0.5 mm 400-200 1.14-0.57
Microwave 0.5 – 300 mm 200-0.033 0.57- 9.4 x
10 -5
Radio
frequency
0.3 – 10 9 m
E
n
e
r
g
y
d
e
c
r
e
a
s
e
s

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ν = c / λ -------------- [1]
• ν = frequency in Hz/cps
• c = 3 x 1010 cm/sec
• λ = wavelength in cm
E = hv --------------- [2]
• h= Planck’s constant (
• ν = frequency in Hz/cps
E = hc / λ --------------- [3]

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deca 10 da deci 10-1 d
hecto 102 h centi 10-2 c
kilo 103 k milli 10-3 m
mega 106 M micro 10-6 m
giga 109 G nano 10-9 n
tera 1012 T pico 10-12 p
peta 1015 P femto 10-15 f
exa 1018 E atto 10-18 a
zetta 1021 Z zepto 10-21 z
yotta 1024 Y yocto 10-24 y

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Types of Spectroscopy
• Ultraviolet (UV) spectroscopy uses electron transitions to
determine bonding patterns.
• Infrared (IR) spectroscopy measures the bond vibration
frequencies in a molecule and is used to determine the
functional group.
• Nuclear magnetic resonance (NMR) spectroscopy detects
spin transition in hydrogen atoms and can be used to
distinguish isomers.
• Mass spectrometry (MS) fragments the molecule and
measures the masses.

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UV spectroscopy/ Electronic spectroscopy
• UV region – 200-400 nm/
2000 - 4000Å
• It is primarily used to
measure the multiple bond
or aromatic conjugation
within molecules.
• Vacuum UV region – below
200 nm/ 100 – 200 nm
Bcos, Air absorbs radiation
in this region, so vacuum
is required to make
measurements.
Absorption of UV rays
Transitions of valence
electrons in the
molecule
Changes electronic
energy of the molecule

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 - * > n- * > -* > n- *

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1.  - *
• alkanes
• Sigma bonds are very strong, therefore
transitions require higher energy – 150 nm
• Studied in Vacuum UV region.

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2. n- *
• Saturated compounds with one hetero atom with
unshared pair of electrons.
• Saturated halides, alcohols,ethers, aldehydes,
ketones, amines etc.
• 150 – 250 nm
• In saturated alkyl halides, the energy with in the
size of halogen.
• Cl > I
• Absorption maxima for n- * tends to shift to
shorter wavelengths in the presence of polar
solvents. (hydrogen bonding)

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3. -* ( K band)
• Unsaturated compounds
• 170-190 nm
• Allowed transition-

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4. n- *( R band)
• Unsaturated compounds with one hetero atom
with unshared pair of electrons.
• Requires least energy longer λ (280 nm)
• Forbidden transition -

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Conjugated systems
• Conjugation – electron are delocalized; lower energy
required to exite electrons;
• The pi orbitals of the separate alkene groups combine to give
new orbitals.
• Bonding orbitals – 1& 2
• Antibonding orbitals- 3* & 4*
• Transition – 2-3*
• Requires lower energy than -*
• Absorption maxima will shift to longer wavelength.

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Spectroscopy – Energy Transitions
• n- *- Molar absorptivity 10 – 100 L cm-1 mol-1
• -* - Molar absorptivity 1000 to 10,000 L cm-1 mol-1
• Applications of absorption spectroscopy to organic
compounds are based upon this transitions to
excited state of *.
• Energy required for those excitation states is within
200 to 700 nm.
• Chromophores involved are unsaturated functional
groups to provide the π orbitals.
• n- * transitions are hypsochromic (shift towards
blue) with increasing polarity of solvent
• -* transition often is bathochromic (shift to red)
with
• increased solvent polarity

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• B – bands: benzenoid bands are characteristics
of aromatic and hetero aromatic compounds.
In benzene the B-band is at 256nm.
• E- band: ethylenic bands are characteristic of
aromatic systems like B bands.

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Chromophore- is a covalently unsaturated group
responsible for electronic absorption.(e.g., C=C,
C=O, NO)

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Types of Chromophore
Independent : presence of single chromophore is
sufficient to impart colour like
Azo group -N=N-, Nitroso group -NO
Dependent : when more than one chromophore is
required for producing colour like C=O, C=C
Auxochrome- is a saturated group with nonbonded electrons
which, when attached to a chromophore, changes the
wavelength as well as the intensity of an absorption.
Auxochromes are usually either basic or acidic
e.g. -NH2, -OH, -NHCH3, -SO3H, -COOH, -COCH3, -OCH3

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Types of auxochrome
1. Bathochromic group NO2
NO2
NH2
Colourless Pale Yellow Dark Yellow
2. Hypsochromic group
The groups which lighten the colour of chromophore
Acetylation of -OH group ------- -OCOCH3
Acetylation of -NH2 group ------ -NHCOCH3
shows blueshift

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When light falls on homogeneous medium, a portion of the incident
Light is reflecte, a portion is absorbed and the the rest is transmitted.
Io = Ia + It + Ir
Ir is only 4% so can be eliminated.
Io = Ia + It

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Lambert’s law
• When monochromatic light
passes through a transparent
medium, the rate of decrease
in intensity with the thickness
of the medium is proportional
to the intensity of the light.
• Intensity of the emitted light
decreases exponentially as the
thickness of the absorbing
medium increases
arithmetically.
• More the thickness, more will
be the absorption.
• Absorption coefficient is defined
as the reciprocal of the
thickness requires to reduce
the light to 1/10 of its intensity.
-dI/db = kI (1)
I = intensity of radiation
b= thickness of medium
k= proportionality factor
Integrating eqn. 1, & putting I= Io
when b = 0,
ln Io/It = kb (2)
Or,
It = Ioe-kb (3)
By changing natural log to common
log,
It = Io x 10 –0.4343kb = Io x 10-Kb (4)
K = k/2.3026 and is absorption
coefficient.
K = 1/b

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Beer’s law
• Intensity of a beam of monochromatic light
decreases exponentially as the concentration of the
absorbing substance increases arithmetically.
• It= Ioe-k’c
= Io x 10-0.4343k’c = Io x 10-K’c (5)
Combining eqn. 4 & 5,
It = Io x 10-abc (6)
Log (Io/It) = abc
A = abc
A = Log Io/It
ε= A/ bc
10 ε = A 1%
1cm x mol.wt.
T= It/Io

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Term Symbol Formula Older terms
Absorbance A A= abc Extinction E,
Optical Density
OD,
absorbancy
Transmittance T It/Io Transmission
Opacity Io/It
% transmittance % T 100 T % transmission
Absorptivity a A/bc Extinction
coefficient,
Absorbancy index
Molar absorptivity ε A/bc
(concentration
in moles)
Molar extinction
coefficient

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Deviations from Beer’s law
Real or true
deviation
Instrumental
deviation
Chemical
deviation
• Only dilute solns.
• Conc. Solns.
Change in RI
Each molecule does
not absorb in same
manner.
+ve & -ve deviation
• stray radiation reaching
detector
•Fluctuation of source
•Sensitivity changes
in detector.
•Defect in detector
amplification system
•Broad band pass
(wide slit width)
• Ionization
• Dissociation
•Reaction with solvent

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Instrumentation
Source Monochromator Sample
Solvent
Detector
Deuterium
lamp
Filters
• absorption filters
• interference
filters
Quartz
Fused silica
Barrier layer
cell
Tungsten
Lamp
Prisms
• cornu type
• littrow type
95%
ethanol
Phototube
Xenon arc
lamp
Gratings Photomultiplier
tube
(PMT)
Diode array

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Sources
1. Deuterium & hydrogen lamp (160 to 350 nm)
• D2 + Ee D2* D’ + D’’ + hv
• Shape of aperture between two electrodes, constricts the discharge to
a narrow path, and produces intense ball of radiation.
• Life = 500 hrs
2. Tungsten filament lamp (350 to 2500nm)
• High operating temp. (3500 K)
• (Tungsten filament + iodine) Quartz envelope
• Long life (iodine)
• I2 + gaseous W = WI2 (volatile)
strikes the filament
decomposes, and redeposits tungsten
3. Xenon arc lamp (200 to 1000 nm)

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Filters
1. Absorption filters:
• Absorb certain portions of
spectrum
• Coloured glass (thermally
stable)
• Effective bandwidth- 30 to
250 nm
2. Interference filters
• optical interference
• transparent dielectric
(CaF/MgF)
• λ = 2 t η / n

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Prisms
Cornu prism Littrow prism

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Gratings
1200 to 1400 grooves/ mm

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Sample Holder
• Quartz or Fused Silica Cuvettes
Optically transparent

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Solvents
For selecting a solvent for UV/VIS experimentation two main
things should be considered
• Transparency of the solvent
• Effect of solvent on absorbing system
Most commonly used solvent is 95% ethanol

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Detectors
Barrier layer cell/ photovoltaic cell

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Phototube

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PMT

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Diode array detector

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Double beam spectrophotometer

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Applications
• Qualitative Analysis
• Quantitative Analysis