MODULE II

SPECTROSCOPIC TECHNIQUES AND APPLICATIONS

The study of interaction of electromagnetic radiation with matter is called spectroscopy.
Absorption laws
The absorption of light by molecules is governed by certain laws. They are collectively
known as absorption laws. They are
1. Beer’s law
2. Lambert’s law
3. Beer – Lambert’s law
Beer’s law
When a monochromatic light is incident on a homogeneous medium, then the intensity of
transmitted light decreases with increase in concentration of absorbing solution.
Log α c
Log = kc
Lambert’s law
When a monochromatic light is incident on a homogeneous medium, then the intensity of
transmitted light decreases with increase in thickness of the medium.
log α t
log = kt
Beer – Lambert’s law
When a monochromatic light is incident on a homogeneous medium, then the intensity of
transmitted light decreases with increase in concentration of absorbing solution as well as
thickness of the medium.
log α ct
log = εct
Where I0 is intensity of incident radiation, It that of transmitted radiation, ε is the molar
extinction (absorption) coefficient, c is the concentration and t is the thickness of the medium.
Absorbance (A)
It is the logarithm of ratio of intensity of incident radiation to that of transmitted radiation.
A = log
Transmittance (T)
It is the ratio of intensity of transmitted radiation to that of incident radiation.
T=
log = εct
-log T = εct
Derivation of Beer – Lambert’s law
When a mono-chromatic light is passed through the homogeneous medium of
concentration (C), the rate of decrease in intensity of radiation (-dI) with thickness of the
medium (dt) is proportional to the intensity of incident radiation (I) and also the concentration
(C).
∞IC
= KCdt, where K is the proportionality constant.

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Let I0 be the intensity of incident radiation and It that of transmitted radiation passing through
any finite thickness (t) of the medium, then
∫ = KC∫
= KCt
𝐼
= KCt
𝐼𝑡
This is the integrated form of Beer-Lambert’s law.
= KCt = Ct

𝐼
A= = εCt
𝐼𝑡

Where ε is the molar extinction (absorption) coefficient. It depends on the nature of medium and
also the extent of absorption.
 The absorbance of a 0.01M dye solution in ethanol is 0.62 in a 2cm cell for light of wave
length 5000 A0. If the path length of light through the sample is doubled and the
concentration is made half, what will be the value of absorbance?

A = εct
= = = 1, absorbance remains same

 A dye solution of concentration 0.04M shows absorbance of 0.045 at 530nm; while a test
solution of same dye shows absorbance 0.022 under same conditions. Find the
concentration of test solution.
A = εCt, Here ε and t are constants
=
=
C2 = 0.0195M
 The percentage transmittance of a 0.01m dye solution in ethanol is 20% in a 2cm cell for
light of wave length 5000A0. Find the absorbance and molar absorption coefficient.
A = log = εCt
A = log = 0.6989
ε= = = 34.945 M-1cm-1

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  A 50ppm standard solution of Fe3+ after developing red colour with ammonium thio-cyanate
shows a transmittance of 0.2 at 620nm. While an unknown solution of Fe3+ after developing
colour with the same amount of ammonium thio-cyanate gives a transmittance of 0.4. Find the
concentration of unknown Fe3+ solution.
-log T =εCt, where ε & t are constants.
=
=
C2= 28.4ppm
 A solution shows a transmittance of 20% when taken in a cell of 2.5 cm thickness.
Calculate its concentration if molar absorption coefficient is 12000dm2mol-1.
A = log =εct =0.6986
log =12000 XcX 2.5
c= = = 2.3296 X mol dm-3
Fundamentals of spectroscopy
Interaction of electromagnetic radiations with matter produces spectrum. Spectrum
can be explained in terms of energy levels present in the matter.
Interaction of electromagnetic radiations with atom produces atomic spectrum. Atomic
spectrum can be explained in terms of energy levels present in the atom. When an electron jumps
from one energy level to another energy level, energy can be emitted or absorbed. When the
electron jumps from higher energy level E2 to the lower energy level E1, the excess energy can
be emitted in the form of radiation of definite frequency (ν).
Δ E = E2 – E1
hν = E2 – E1
ν=
where ‘h’ is the Plank’s constant. The frequency of emitted radiation produces a bright line.
Spectrum consists of bright lines produced by the emission of electromagnetic radiation by
atoms is called atomic emission spectra or line spectra.
Types of spectrum
Based on the nature of interaction, the spectra can be broadly divided into absorption
spectra and emission spectra.
Absorption Spectra
When an atom or molecule undergoes transition from lower energy level E1 to the higher
energy level E2, absorption of energy occurs in the form of radiation of definite frequency and
the resultant spectrum is called absorption spectrum.
Emission Spectrum
When an atom or molecule undergoes transition from lower energy level E1 to the higher
energy level E2, absorption of energy occurs in the form of radiation of definite frequency and
the resultant spectrum is called absorption spectrum.
Based on the nature of the interacting species, the spectra can be divided into two categories,
atomic spectra and molecular spectra.
Atomic Spectra
Interaction of electromagnetic radiations with atoms produces transition of electrons
between the atomic energy levels and the resultant spectrum is called atomic spectra.
Molecular Spectra
Interaction of electromagnetic radiations with molecules produces transition of electrons between
the molecular energy levels and the resultant spectrum is called molecular spectra.

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Electromagnetic radiations
Radiations having electric and magnetic field associated with them are called
electromagnetic radiations. They are radio waves, micro waves, IR (far IR, middle IR, near IR),
visible light, UV light (ordinary UV, vacuum UV), X- rays, γ – rays and cosmic rays. They are
arranged in the increasing order of their frequency or the decreasing order of wave length. This is
known as electromagnetic spectrum. They do not require any medium for their propagation.
They possess wave like characteristics.
Characteristics of electromagnetic radiations
1. Wave length (λ)
It is the distance between two adjacent crests or two adjacent trough of a wave. Unit is
nanometer and A°.
1nm = 10-9 meter
1A° = 10-12 meter
2. Frequency (ν)
Number of waves which pass through a given point in 1 second is called frequency. Its unit is
Hertz or Cycles per Second (CPU).
1 Hertz = 1 Cycles per Second (CPU)
3. Velocity (v or c)
Distance travelled by a wave in 1 second is called velocity. Its unit is m/S. Velocity of all
electromagnetic radiations in space is constant and is 3 X 108 ms-1
4. Wave number (ῡ)
It is the reciprocal of wave length. Its unit is cm-1 or m-1.
ῡ= ⁄
Molecular spectra
Interactions of electromagnetic radiation with molecules produce molecular spectra. In a
molecule three types of energy levels are present. They are rotational, vibrational & electronic
energy levels. The transition between these energy levels results in the emission or absorption of
energy in the form of radiation of definite frequency. Generally large numbers of such transitions
are possible.

Within the same electronic energy level, there are large numbers of vibrational energy levels and
within the same vibrational energy level, there are large number of rotational energy levels. The
electronic, vibrational and rotational energy levels are represented by n, v, j.

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Total energy of a molecule = E ele + E vib + E rot
When a molecule emit or absorb radiation, it may results in the electronic vibrational and
rotational energy changes. All these changes are quantized and give rise to three types of
molecular spectra. They are rotational spectra, vibrational spectra and electronic spectra.
The energy involved in these transitions is in the following order.
E ele – E1 ele > E vib – E1 vib > E rot – E1 rot
Because of the change in energy level between the rotational, vibrational and electronic energy
levels, molecular spectrum is obtained at three different regions of the electronic spectra.
Vibrational Spectra ( IR Spectra)
IR spectroscopy is concerned with the study of absorption of IR radiation (800nm-
1mm), which causes vibrational transition in the molecule. When a gaseous molecule absorbs IR
radiation, it produces transition between the vibrational energy levels. Resultant spectrum is
called vibrational spectra.
The essential requirement for a molecule to produce IR spectrum is that the dipole
moment of the molecule must change during vibration. Thus vibrational spectrum is given by
hetero nuclear diatomic molecules. Polyatomic molecules with and without dipole moment also
produces IR spectrum. Homo nuclear diatomic molecules such as O2, N2, H2 etc. will not
produce IR spectrum, since they have zero dipole moment. But molecules like HCl, HBr, NO 2,
H2O, CO2, NH3, CH3Cl and SO2 can produce IR spectra and they are IR active molecules. In the
case of polyatomic molecules without dipole moment, the exciting radiations can induce an
oscillating dipole moment. Hence they are also IR active.
When a gaseous molecule having dipole moment vibrates, it generates an electric field
which can interact with electrical component of IR radiation. During this interaction
energy can be emitted or absorbed in the form of radiation of definite frequency. Thus the
vibration of the molecule produces IR spectrum.
Instrumentation of IR spectrometer

Theory of IR spectra
In order to explain the theory of IR spectra, consider the vibration of hetero nuclear
diatomic molecule. It is similar to the vibration of a simple harmonic oscillator.
Vibrational energy of simple harmonic oscillator = Vibrational energy of diatomic molecule
E vib = (v + ⁄ ) hω
Fundamental frequency of vibration, ω= √ , where ‘k’ is the force constant and it gives an idea
about the strength of the bond.
Reduced mass (µ) =
When the molecule is in the vibrational ground level (v =0), E vib = ⁄ hω. This is the lowest
vibrational energy level and it is called zero point energy.
Let us consider the vibrational transition from lowest vibrational (v1) to the next higher
vibrational energy level (v).

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Energy change during vibrational transition (∆E vib) = E v – Ev1
= (v + ⁄ ) hω - (v1 + ⁄ ) hω
= (v-v1) hω = ∆vhω
The selection rule for vibrational spectrum is ∆v = ±1
∆E vib = hω
Spectrum can be explained in terms of wave number.

Thus the wave number of absorbed line (ῡ) = = = = √

According to Hooke’s law, 𝟏 𝐤
ῡ = 𝟐𝛑𝐜 √

K = 4π2c2ῡ2μ

Thus the frequency or wave number of absorbed line depends on

 Force constant(k)
 Reduced mass(µ)
Stretching frequency for C≡C, C=C, C‒C bond are in the following order etc.
C≡C > C=C > C‒C
Since the force constant for each type of bond follows the order.
k C≡C > k C=C > k C‒C

Different vibrational transitions are possible between the vibrational energy levels, but all
the vibrational energy levels are equally spaced with a spacing of ‘hω’. So all the lines in the
spectrum would fall in the same region. Hence vibrational spectrum is expected to consist of a
single line.
At ordinary conditions of temperature and pressure, most of the molecules are in the
vibrational ground level (V = 0). So only one transition is possible according to the selection
rule, i.e. 0 → 1 transition and the resultant lines are called fundamental lines and are highly
intense lines. But in actual practice molecular vibrations are not harmonic. Such an oscillator is
called anharmonic oscillator. For an anharmonic oscillator ΔV is rarely unity. Sometimes it is
+2 and sometimes it is +3. If ΔV = +2, then the most probable transition is 0 → 2 transition
and the resultant lines are called first overtones. They have lesser intensity than the fundamental

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lines. If ΔV = +3, then the most probable transition is 0 → 3 transition and the resultant lines
are called second overtones. They have least intensity.
 Calculate the force constant of HCl molecule, if it gives IR absorption at 3050cm-1.
(Given that atomic masses of H = 1 amu and Cl = 35 amu)
ῡ = 3050cm-1 = 3050X100 m-1
μ= X 1u = X 1.67 X 10-27 kg = 1.63X10-27 Kg.
k = 4π2c2ῡ2μ = 4π2X (3X ) 2(3050X100 m-1)2X1.63 X 10-27 kg
= 532.8Nm-1
 CO molecule absorbs at 2140cm-1. Calculate the force constant of the molecule. Given
atomic masses of C &O are 12u and 16u respectively.(1u = 1.67 X 10-27kg).

ῡ= √

μ= X 1U = X 1.67 X 10-27kg
k = 4π2C2ῡ2μ
k = 1853 Nm-1
 The vibrational frequency of HCl molecule is 2886cm-1.Calculate the force constant of
the molecule. Reduced mass of HCl is 1.63X10-27 Kg.
ῡ= √ cm-1
k = 4π2C2ῡ2μ = 4π2 X (3 X 10 8)2 (2886 X 100)2X 1.63X10-27 Kg.
k = 481.8 Nm-1
 The fundamental vibrational frequency of 12C16O is 2140 cm-1. Without calculating force
constant, find the fundamental frequency of 13C17O in cm-1.
k = 4π2c2ῡ2μ
𝝁 = X 1.67 X = 1.145 X kg

𝝁 = X 1.67 X = 1.23 X kg

𝝁
=√
𝝁

𝝁
= X √
𝝁

= 2140 X √ =2065 cm-1

 Calculate the force constant of HF molecule, if it shows IR absorption at4138 cm-1. Given
that atomic masses of hydrogen and flourine are 1u and 19u respectively. What would be
the wave number if hydrogen atoms are replaced by deuterium atoms?
k = 4π2c2ῡ2μ
μ= X 1U = X 1.66 X 10-27kg = 1.577 X 10-27kg
k = 4π2C2ῡ2μ = 4π2 X (3 X 10 8)2 (4138 X 100)2X 1.577X10-27 Kg = 958.3 Nm-1
When deuterium is substituted for hydrogen, k remains constant.

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lines. If ΔV = +3, then the most probable transition is 0 → 3 transition and the resultant lines
are called second overtones. They have least intensity.
 Calculate the force constant of HCl molecule, if it gives IR absorption at 3050cm-1.
(Given that atomic masses of H = 1 amu and Cl = 35 amu)
ῡ = 3050cm-1 = 3050X100 m-1
μ= X 1u = X 1.67 X 10-27 kg = 1.63X10-27 Kg.
k = 4π2c2ῡ2μ = 4π2X (3X ) 2(3050X100 m-1)2X1.63 X 10-27 kg
= 532.8Nm-1
 CO molecule absorbs at 2140cm-1. Calculate the force constant of the molecule. Given
atomic masses of C &O are 12u and 16u respectively.(1u = 1.67 X 10-27kg).

ῡ= √

μ= X 1U = X 1.67 X 10-27kg
k = 4π2C2ῡ2μ
k = 1853 Nm-1
 The vibrational frequency of HCl molecule is 2886cm-1.Calculate the force constant of
the molecule. Reduced mass of HCl is 1.63X10-27 Kg.
ῡ= √ cm-1
k = 4π2C2ῡ2μ = 4π2 X (3 X 10 8)2 (2886 X 100)2X 1.63X10-27 Kg.
k = 481.8 Nm-1
 The fundamental vibrational frequency of 12C16O is 2140 cm-1. Without calculating force
constant, find the fundamental frequency of 13C17O in cm-1.
k = 4π2c2ῡ2μ
𝝁 = X 1.67 X = 1.145 X kg

𝝁 = X 1.67 X = 1.23 X kg

𝝁
=√
𝝁

𝝁
= X √
𝝁

= 2140 X √ =2065 cm-1

 Calculate the force constant of HF molecule, if it shows IR absorption at4138 cm-1. Given
that atomic masses of hydrogen and flourine are 1u and 19u respectively. What would be
the wave number if hydrogen atoms are replaced by deuterium atoms?
k = 4π2c2ῡ2μ
μ= X 1U = X 1.66 X 10-27kg = 1.577 X 10-27kg
k = 4π2C2ῡ2μ = 4π2 X (3 X 10 8)2 (4138 X 100)2X 1.577X10-27 Kg = 958.3 Nm-1
When deuterium is substituted for hydrogen, k remains constant.

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 In order to explain the features of IR spectrum, let us consider the IR spectrum of aceto
phenone. Spectrum can be divided into two regions. First region lies in between 1600 – 4000
cm1. This region is called functional group region, since the vibrational modes of most of the
functional groups occur in this region. A few broad bands in the functional group region show
the presence of a functional group. Second region lies between 600 – 1600 cm-1. A few broad
bands in this region show the presence of an aliphatic group. Large numbers of sharp bands in
this region show the presence of an aromatic group. Hence this region provides the confirmation
of identity and this region is called finger print region.

Applications of IR spectroscopy;
 The identity of unknown substance can be explained by considering the IR spectrum of
unknown substance with that of a known substance.
 This will give valuable information about molecular symmetry and force constant.
 This can be used to detect the presence of impurities, since the impurities give rise to
extra absorption bands.
 This can be used to distinguish intra molecular H-bonding & inter molecular H-bonding.
This can be done by taking the IR spectrum at different dilutions. As the dilution
increases intensity of absorption band diminishes when it is due to inter molecular H-
bonding and as the dilution increases intensity of absorption band remains unchanged
when it is due to intra molecular H- bonding.

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 Electronic (UV –Visible) Spectroscopy
Instrumentation of UV –Visible) Spectrometer

Electronic spectrum of a molecule

Electronic spectrum of molecule arises due to the transition between electronic energy
levels. Electronic transitions are highly energetic transitions. Hence it produces spectrum in the
UV – Visible region.
In a molecule three types of energy levels are present. They are rotational, vibrational &
electronic energy levels. Within the same electronic energy level, there are large numbers of
vibrational energy levels and within the same vibrational energy level, there are large number of
rotational energy levels. So electronic transitions are always followed by simultaneous
vibrational and rotational transitions. Hence we get a broad spectrum consisting of lines for
electronic, vibrational and rotational transitions.

Suppose E1(ele), E1(vib), E1(rot) be the electronic, vibrational and rotational energy of the
molecule before transition and E(ele), E (vib), E (rot) be the electronic, vibrational and rotational
energy of the molecule after transition.
Total energy of the molecule before transition (E1) = E1 (ele) + E1(vib) + E1(rot)
Total energy of the molecule after transition (E) = E (ele) + E (vib) + E (rot)
Change in energy during transition ( E) = (E - E1)
= (E (ele) + E (vib) + E (rot)) – (E1 (ele) + E1(vib) + E1(rot))
Principle of electronic spectroscopy
In order to explain the principle of electronic spectra, let us consider the Frank Condon
principle. According to Frank Condon principle, during an electronic transition internuclear
distance remains constant. Since the electrons move much more rapidly than the nucleus.
The figure shows the potential energy versus inter nuclear distance curve of ground state
and first excited state of the diatomic molecule XY. Quantum mechanically molecule is at the

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center of the ground vibrational level of ground electronic state. Electronic transitions are always
represented by a vertical line.
In figure (a), upper and lower inter nuclear distance is same. According to Frank Condon
principle, most probable transition is 0 → 0 transitions. In figure (b), upper state inter nuclear
distance is slightly lesser than the lower state. According to Frank Condon principle, most
probable transition is 0 → 2 transitions. In figure (c), upper state inter nuclear distance is slightly
greater than the lower state. According to Frank Condon principle, most probable transition is 0
→ 2 transitions. In figure (d), upper state inter nuclear distance is considerably greater than the
lower state. Here there is no specific electronic transition; instead we get some dark faded lines
and is called continuum.

Fundamentals of electronic spectroscopy

Chromophore
Any isolated covalently bonded group that shows characteristic absorption in the UV
Visible region irrespective of the fact that whether colour is produced or not.
Eg: >C=C<, ─C ≡ C ─, ─N≡ N─ etc.
Types of chromophores ζ – ζ*, 𝞹 – 𝞹*, n – 𝞹* & n – ζ*
Chromophores are of two types
1. Chromophores containing π bond undergoes 𝞹 – 𝞹* transition
Eg. Ethylene, acetylene etc.
2. Chromophores containing both π and n electrons undergoes 𝞹 – 𝞹*, n – 𝞹* transitions.
Eg. CH3─ CH2─CHO
Auxochromes
These are substituent on the Chromophore which leads to red shift. eg. NH2─, SH─, OH─ etc.

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Bathochromic shift (Red shift)
Shift of absorption band towards longer wavelength region (red region) is called Bathochromic
shift. Reason for red shift is the presence of auxochrome and change in solvent.
Hypsochromic shift (Blue shift)
Shift of absorption band towards shorter wavelength region (blue region) is called Hypsochromic
shift. Reason for blue shift is the presence of removal of conjugation and change in polarity of solvent.
Hyper chromic shift
Shift in which there is an increase in intensity of absorption band is called hyper chromic shift.
Hypo chromic shift
Shift in which there is an decrease in intensity of absorption band is called hypo chromic shift.
Electronic spectrum of polyatomic molecules
In the case of polyatomic molecules, electronic transitions give rise to absorption spectra in the
UV-Visible region. From the investigations of UV-Visible spectra we get information about
various energy levels in the molecule depending on the energy of the molecular orbitals. The
electronic transitions are ζ – ζ*, 𝞹 – 𝞹*, n – 𝞹* & n – ζ* respectively.

1) ζ – ζ* transitions
The energy required for this transition is very high. Since the ζ electrons are held more strongly
in the molecule and are highly energetic. Hence the absorption band occurs in far UV region. All
saturated hydrocarbons will undergo this transition. These types of transitions occur only below
150nm. The ordinary UV spectrometers can take spectra only from 200 – 780nm. Hence
saturated hydrocarbons cannot be detected using UV-Visible spectra.

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2) 𝞹 – 𝞹* transition
Unsaturated hydrocarbons containing 𝞹 bonds can produce this type of transition. But C=C of
CH2=CH2 molecule gives absorption maxima at 169nm.So it cannot be detected using ordinary
UV spectrometer .But in compounds containing conjugated double bonds, due to the presence of
conjugated double bonds absorption occurs in visible region. In such molecules 𝞹 – 𝞹* transition
produces absorption bands in the UV-Visible region.
(eg. Butadiene, absorption occurs at 217nm). Due to this transition high intensity absorption
bands are produced in the near UV region. Benzene also produces these transitions, due to the
presence of three conjugated double bonds. In the case of lycopene red coloured of tomato, there
are eleven double bonds in conjugation, its λmax is 505 nm. For every double bond in
conjugation, there is an increment of 30 nm.
3 ) n – 𝞹* transition
Unsaturated compounds containing atoms such as O, N, S, X etc, containing lone pair of
electrons produces absorption band due to n – 𝞹* transition and the absorption bands are
observed in the visible region. (eg., aldehydes and ketones containing C=C and C ≡ C bonds)
4) n – ζ* transition
Saturated compounds containing atoms such as O, N, S, X etc, containing lone pair of electrons
produces absorption band due to n – ζ* transition and the absorption bands are observed in the
near UV region. Generally absorption takes place below 200nm. (eg., CH3OH, CH3NH2&
(CH3)2NH etc.)
UV- VIS energy level diagram of conjugated systems
The energy levels of π molecular orbitals of conjugated systems can be obtained from
Huckel’s molecular orbital theory. In such molecules, the ground state electrons occupy the low
energy Huckel molecular orbital in spin paired state. Out of the occupied orbitals, the one with
higher energy is termed as HOMO (highest occupied molecular orbital). Similarly, out of the
unoccupied orbitals, the one with lower energy is termed as LUMO (lowest unoccupied
molecular orbital). If the molecule absorbs electromagnetic radiation in the UV – VIS region,
electrons get promoted from HOMO to LUMO and the wavelength of absorption depends upon
HOMO-LUMO energy gap.
UV- VIS energy level diagram of ethylene, butadiene and hexa-triene

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 Ethylene contains a simple isolated C꞊C bond, but the other two have conjugated double
bonds. In the case of ethylene, there is one π bonding molecular orbital and one π* antibonding
molecular orbital. So π→π* transition occurs and ethylene gives an absorption maxima at 169
nm. In 1, 3-butadiene, there are two double bonds in conjugation. So it has four π molecular
orbitals (ψ1, ψ2, ψ3, ψ4) formed by the linear combination of four atomic orbitals (p orbitals). Here
ψ1 & ψ2 are the bonding π molecular orbitals and ψ3 & ψ4 are the antibonding π* molecular
orbitals. In the ground state the four π electrons occupy bonding molecular orbitals. Here the
transition takes place from HOMO (ψ2) to LUMO (ψ3) and absorption occurs at 217nm. In 1, 3,
5-hexatriene, there are three double bonds in conjugation. So it has six π molecular orbitals (ψ1,
ψ2, ψ3, ψ4, ψ5, ψ6) formed by the linear combination of six atomic orbitals (p orbitals). In the
ground state the six π electrons occupy bonding molecular orbitals. Here the transition takes
place from HOMO (ψ3) to LUMO (ψ4) and absorption occurs at 247nm.
UV- VIS energy level diagram of benzene
In benzene, there are three double bonds in conjugation. So it has six π molecular orbitals (ψ1, ψ2,
ψ3, ψ4, ψ5, ψ6) formed by the linear combination of six atomic orbitals (p orbitals). In the ground
state the six π electrons occupy bonding molecular orbitals. Here the transition takes place from
HOMO to LUMO and absorption occurs at 255nm. Here ψ2, ψ3 are degenerate orbitals and ψ4,
ψ5 are also degenerate orbitals

Applications of UV – Visible spectroscopy

 In medical lab test for determining cholesterol and blood sugar.
 For the characterization of dyes and colourants
 For the detection of aromatic compounds
 Detection of impurities. ( Benzene is present as an impurity in cyclohexane can be
detected)
 Write three points of comparison between UV & IR spectrum.
UV-VIS spectroscopy IR-spectroscopy
Electronic spectrum is due to Vibrational spectrum is due to the vibrational
the electronic transitions in motions of atoms and molecules.
atoms and molecules.
It arises by the interaction of It arises by the interaction of EMR with
EMR with matter matter.
EMR in the UV-Vis region is EMR in the IR region is responsible for
responsible for electronic electronic spectrum
spectrum

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2) 𝞹 – 𝞹* transition
Unsaturated hydrocarbons containing 𝞹 bonds can produce this type of transition. But C=C of
CH2=CH2 molecule gives absorption maxima at 169nm.So it cannot be detected using ordinary
UV spectrometer .But in compounds containing conjugated double bonds, due to the presence of
conjugated double bonds absorption occurs in visible region. In such molecules 𝞹 – 𝞹* transition
produces absorption bands in the UV-Visible region.
(eg. Butadiene, absorption occurs at 217nm). Due to this transition high intensity absorption
bands are produced in the near UV region. Benzene also produces these transitions, due to the
presence of three conjugated double bonds. In the case of lycopene red coloured of tomato, there
are eleven double bonds in conjugation, its λmax is 505 nm. For every double bond in
conjugation, there is an increment of 30 nm.
3 ) n – 𝞹* transition
Unsaturated compounds containing atoms such as O, N, S, X etc, containing lone pair of
electrons produces absorption band due to n – 𝞹* transition and the absorption bands are
observed in the visible region. (eg., aldehydes and ketones containing C=C and C ≡ C bonds)
4) n – ζ* transition
Saturated compounds containing atoms such as O, N, S, X etc, containing lone pair of electrons
produces absorption band due to n – ζ* transition and the absorption bands are observed in the
near UV region. Generally absorption takes place below 200nm. (eg., CH3OH, CH3NH2&
(CH3)2NH etc.)
UV- VIS energy level diagram of conjugated systems
The energy levels of π molecular orbitals of conjugated systems can be obtained from
Huckel’s molecular orbital theory. In such molecules, the ground state electrons occupy the low
energy Huckel molecular orbital in spin paired state. Out of the occupied orbitals, the one with
higher energy is termed as HOMO (highest occupied molecular orbital). Similarly, out of the
unoccupied orbitals, the one with lower energy is termed as LUMO (lowest unoccupied
molecular orbital). If the molecule absorbs electromagnetic radiation in the UV – VIS region,
electrons get promoted from HOMO to LUMO and the wavelength of absorption depends upon
HOMO-LUMO energy gap.
UV- VIS energy level diagram of ethylene, butadiene and hexa-triene

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 Ethylene contains a simple isolated C꞊C bond, but the other two have conjugated double
bonds. In the case of ethylene, there is one π bonding molecular orbital and one π* antibonding
molecular orbital. So π→π* transition occurs and ethylene gives an absorption maxima at 169
nm. In 1, 3-butadiene, there are two double bonds in conjugation. So it has four π molecular
orbitals (ψ1, ψ2, ψ3, ψ4) formed by the linear combination of four atomic orbitals (p orbitals). Here
ψ1 & ψ2 are the bonding π molecular orbitals and ψ3 & ψ4 are the antibonding π* molecular
orbitals. In the ground state the four π electrons occupy bonding molecular orbitals. Here the
transition takes place from HOMO (ψ2) to LUMO (ψ3) and absorption occurs at 217nm. In 1, 3,
5-hexatriene, there are three double bonds in conjugation. So it has six π molecular orbitals (ψ1,
ψ2, ψ3, ψ4, ψ5, ψ6) formed by the linear combination of six atomic orbitals (p orbitals). In the
ground state the six π electrons occupy bonding molecular orbitals. Here the transition takes
place from HOMO (ψ3) to LUMO (ψ4) and absorption occurs at 247nm.
UV- VIS energy level diagram of benzene
In benzene, there are three double bonds in conjugation. So it has six π molecular orbitals (ψ1, ψ2,
ψ3, ψ4, ψ5, ψ6) formed by the linear combination of six atomic orbitals (p orbitals). In the ground
state the six π electrons occupy bonding molecular orbitals. Here the transition takes place from
HOMO to LUMO and absorption occurs at 255nm. Here ψ2, ψ3 are degenerate orbitals and ψ4,
ψ5 are also degenerate orbitals

Applications of UV – Visible spectroscopy

 In medical lab test for determining cholesterol and blood sugar.
 For the characterization of dyes and colourants
 For the detection of aromatic compounds
 Detection of impurities. ( Benzene is present as an impurity in cyclohexane can be
detected)
 Write three points of comparison between UV & IR spectrum.
UV-VIS spectroscopy IR-spectroscopy
Electronic spectrum is due to Vibrational spectrum is due to the vibrational
the electronic transitions in motions of atoms and molecules.
atoms and molecules.
It arises by the interaction of It arises by the interaction of EMR with
EMR with matter matter.
EMR in the UV-Vis region is EMR in the IR region is responsible for
responsible for electronic electronic spectrum
spectrum

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 iii) It is soluble in most of the organic solvents.
iv) It is highly volatile and can be easily removed from the system
In the NMR spectra, we get a reference peak corresponding to TMS. The position of peaks in the
NMR spectra is measured relative to that of reference compound and is expressed in terms of
chemical shift (δ)

Shift in position of NMR signals due to shielding and de-shielding of protons by

the circulation of electrons is called chemical shift.

Chemical shift can be calculated by the expression,

Chemical shift,δ = X 10 6ppm
υo –operating frequency
υsample– frequency of sample
υTMS – frequency of reference (TMS)
Generally chemical shift value is in between 0 and 10 ppm. There are two scales
are used for the measurement of chemical shift. They are δ - scale & η - scale. These two
scales are related to each other by the expression,
η = 10 - δ

In η – scale, signal for TMS is obtained at 10ppm. But in δ – scale, TMS signal is obtained at 0
ppm.
When a molecule is placed in a magnetic field, the electrons are forced to circulate about its own
protons or nearby protons and this circulation produce an induced magnetic field. The rotation of
electrons about its own protons produces an induced magnetic field, which opposes the applied
field. Thus the field felt by the proton is diminished and the proton is said to be shielded.
Shielding shift the absorption upfield. If the protons are rotated about nearby protons, then the
induced magnetic field either opposes or reinforces the applied field. If the induced magnetic
field opposes the applied field, proton is said to be shielded. If the induced magnetic field
reinforces the applied field, then the proton feels higher field strength and the proton is said to be
de-shielded. De-shielding shifts the absorption downfield. Chemical shift for different kinds of
protons are measured relative to that of the reference peak (TMS signal). The NMR signals for
different types of protons in a molecule appear at different field strengths with respect to TMS
signal. This difference in absorption position of protons with respect to TMS signal is called
chemical shift.

 Calculate the frequency of operation of NMR instrument, if particular proton of δ value

4.2 shows a difference in frequency 1260 Hz from the TMS.
Chemical shift, δ = X
4.2 = X 106

V0 = X 106

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 = 300 X 106 Hz = 300MHz
 CHCl3 protons show a shift in frequency of 728 Hz from TMS signal in a 100MHz NMR
instrument. How much would be the shift in frequency for the same proton from TMS in
a 300MHz NMR instrument?
Chemical shift, δ = X =

Chemical shift, δ is constant

= = 2184 Hz

 If gyromagnetic ratio of 13C is of that of 1H. What is the frequency must be irradiated to
take 13C NMR spectrum if same instrument take 1H NMR spectrum at 300MHz.

υ = ( ) H0, Here field strength H0 is constant.

= ⁄

𝛎2 = = 75 MHz

Factors affecting chemical shift

i) Electronegativity

If the electronegativity of substituents attached to the protons increases, then electron

density around the protons decreases and the protons feels higher field strength & the protons
are said to be de-shielded. In the case of methyl halides, protons of CH3F are least shielded
(most de-shielded) and those of CH3I (2.2ppm) are most shielded(least de-shielded).
Compounds CH3F CH3-O-CH3 (CH3)3N CH3-CH3
δ of CH3 protons 4.3ppm 3.2ppm 2.2ppm 0.9ppm
ii) Cumulative effects of electronegative substituents

If the number of electronegative substituents attached to the protons increases, then

the protons feels higher field strength and the proton is said to be de-shielded. For example
consider the chemical shifts for various chlorinated methane derivatives,

Compounds CHCl3 CH2Cl2 CH3Cl

δ of CH3 protons 7.3ppm 5.3ppm 3.1ppm

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iii) Distance from the electronegative atom
If the distance from the electronegative atom substituted in the hydrocarbon chain increases,
electron density around protons increases and shielding increases so chemical shift value
decreases.
Compound H3C – CH2 – CH2Cl
Chemical shift (δ)(ppm) 1 1.4 3.4
iii) De-shielding

In de-shielding, induced magnetic field reinforces the applied magnetic field.

Therefore de- shielding increases the chemical shift (δ) values. In the case of aromatic protons
and olefinic protons δ value is high due to the de- shielding effect.
Low resolution NMR Spectrum of CH3-OH

In methanol (CH3-OH) there are three identical CH bonds and one OH bond are present. The
chemical environment of CH bonds and one OH bond are different. Since oxygen is more
electronegative than carbon. So electron density around -OH proton is lesser than that around
CH3- protons. As a result, CH3- protons are more shielded than –OH proton. That means
shielding constant (ζ) is greater for CH3- protons. ζCH ˃ ζ OH
3

Shielded proton feels lower field strength. So observed magnetic field (B) is greater for –OH
proton.
B OH ˃ B CH3

That means actual field felt by the CH3- protons is lesser than that felt by –OH proton. Hence on
increasing the external magnetic field, -OH proton will comes into resonance earlier than CH3- protons.
As a result NMR signal for –OH proton will be observed at a lower field strength and that of CH3-
protons will be observed at higher field strength.
The area under each peak is directly proportional to the number of protons. Thus the area under the signal
of CH3- protons will be three times larger than that under –OH proton.

High Resolution NMR Spectrum

If we take the high resolution NMR Spectrum of molecule, we can see a multiplet
instead of a single peak. It is due to a phenomenon called spin-spin splitting.
Spin -spin splitting (spin-spin coupling)
The interaction between spins of neighbouring nuclei in a molecule can cause splitting of
lines in the NMR spectrum. This is called spin -spin splitting (spin-spin coupling)
In order to explain spin -spin splitting (spin-spin coupling), let us consider the high
resolution spectrum of CH3-CH2-Br. This compound contains two types of protons, so two
signals are expected in the low resolution NMR spectrum. But in high resolution NMR spectrum,

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we do not get singlet for each type of proton, instead a group of peaks (multiplet) are observed.
For CH3- protons, singlet will split into a triplet with intensity ratio 1:2:1 and for CH2 ˂ protons,
singlet will split into a quartet with intensity ratio 1:3:3:1. This can be explained by considering
the spin orientation of each type of protons.

The spin of CH2˂ protons can couple with adjacent CH3- protons in three different ways
relative to the external field. Hence the field felt by the CH3- protons can be modified in three
different ways. That means NMR signal due to CH3- protons will split into three (triplet).
Similarly the spin of CH3- protons can couple with adjacent CH2˂ protons in four
different ways. Hence the field felt by the CH2˂ protons can be modified in four different ways.
That means NMR signal due to CH2˂ protons will split into four (quartet).
Area under the broad NMR and the multiplet remains the same. Spin – spin coupling is
independent of the applied field strength. The spacing between peaks of multiplet is the measure
of spin – spin coupling and is called spin – spin coupling constant (J).
We can easily determine the relative intensity of multiplet using Pascal’s triangle method.

Applications of NMR spectroscopy

i) Structural elucidation of many organic and inorganic compounds.
ii) NMR spectroscopy is used to study keto-enol tautomerism.
iii) It is used in the study of isotopes.
iv) MRI technique is based on the principle of NMR.
Magnetic Resonance Imaging (MRI)

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 MRI is the medical technique used in radiology to get the detailed pictures of internal body
parts. MRI uses a very strong magnetic field, radio frequency wave and a computer to produce
the detailed pictures of body parts such as soft tissues, bones etc. These images can be examined
in the computer monitor, transmitted electronically, printed or copied to a CD.
Principle:
It is the medical version of NMR. This method uses magnetic properties of hydrogen
present in the body cells and its interaction with external magnetic field to produce highly
detailed pictures of internal body parts. In order to perform MRI scanning, a very strong
magnetic field of 0.5 – 1.5T is required
Applications
 Physicians use MRI for the treatment of diseases like tumors of abdomen, liver cirrhosis,
heart problems etc.
 Measuring physiological functions like neuromuscular skeletal functions.
 Magnetic resonance angiography is used for getting pictures of arteries.
NMR Spectra of compounds
1. Predict NMR spectrum of CH3-CHCl-CH3(2–chloro propane)

2. Write splitting pattern in the NMR spectrum of CH2Cl-CH2-CH2Cl (1, 3- dichloro propane)

3. Give number of signals, peak ratio and multiplicity of different sets of protons in the nmr
spectrum of 1-bromopropane.
In 1-bromopropane, three types of protons are present. So three signals are expected. The
Br attached CH2- has lesser electron density, so signal is obtained far away from TMS. (i.e, at
higher δ value)

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4. How can you distinguish NMR spectrum of CH3CH2Cland CH3CHCl2applying the concept of
spin – spin splitting
NMR spectrum of both the compounds has two peaks. In CH3-CH2Cl, the -CH2Cl is attached to
the more electronegative Cl atom and is less shielded, so its signal is obtained away from TMS
signal. But CH3- is more shielded, so its signal is obtained near to TMS. But in high resolution
spectra each singlet is splitted into a multiplet due to spin - spin splitting. In CH3-CH2Cl, the
CH3- signal is splitted into triplet with intensity ratio 1:2:1, whereas in CH3-CHCl2, the CH3-
signal is splitted into doublet with intensity ratio 1:1. But in both cases -CH2Cl & -CHCl2, we get
a quartet with intensity ratio (1:3:3:1).

5. Draw the high resolution nmr spectrum of ethanol (99.99%pure)

6. Draw the high resolution NMR spectrum for ethanol containing impurities such as water,
acetic acid etc.

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7. Predict the number of signals, their relative positions and their multiplicities for ethyl
benzoate.

Prepared by Raji Sankar, Assistant Professor

Dept. of mathematics & Basic sciences,
Sree Buddha College of Engineering, Pattoor

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