Spectroscopy

The study of the interaction of matter with the electromagnetic spectrum

❖ The best tool to study Matter

❖ What does light do to sample?
❖ Why there is a light absorption/emission

Thermal Infrared Imaging Spectrometer (TIS)

Ref:- Fundamentals of Molecular spectroscopy,
by C. N. Banwell and E.M.McCASH
What is light or Electromagnetic radiation ?
Light as a Wave
➢ Light as a simple harmonic wave of sinusoidal character
y = A sine(2put) A = maximum amplitude of the wave
u = frequency of the wave
t = time in sec
➢ Light has both electric and magnetic components, travelling in straight line except reflected or refracted

An EM radiation can be specified by

➢ Wavelength (l in unit of length)
 α 1/l
➢ Frequency ( in Hz), number of cycles per sec  = c/l, c = 3 x 1010 cm/sec
➢ Wave number (ῡ = 1/l) in cm -1 & number of cycles/Wave per cm  = cῡ

Light as particle :- by Max Planck in 1900 and Quantization of Energy

➢ The particle component of light is called a photon (massless particle a small wave-packet of EM radiation/light).
The photon is the quantum of electromagnetic radiation.
➢ The energy (E) component of a photon is proportional to the frequency.
E = h
ie E = hc /l h = 6.626 X 10-34 Joules sec, Planks constant
 How does light interact with sample?
Classical mechanics, light interacts with charges
➢ Molecules/atoms are composed of charged particles
➢ Light (electromagnetic field) exerts force on these charges (depends on the strength of the field, the
magnitude of the charges
Quantum mechanics, EM radiation can interact with matter in two ways:
• Collision – particle-to-particle – energy is lost as heat and movement (scattering)
• Coupling – when the wave properties of the radiation and particle matches, both “couple” to get the next higher
quantum mechanical energy level
1. Absorption 2. Emission Jablonski diagram

DE = E2 – E1
DE = h joules
 = DE/h

A molecule can absorb or emit only in the multiples of Quanta

Transitions in molecules

Quantum mechanics demand discrete Energy levels in Molecule

Electronic Vibrational Rotational Translational
Energy Energy Energy energy
The entire electromagnetic spectrum can be used
Frequency,  in Hz
~1019 ~1017 ~1015 ~1013 ~1010 ~105

Wavelength, l
~.0001 nm ~0.01 nm 10 nm 1000 nm 0.01 cm 100 m

Energy (J/mol)
> 109 > 107 105 103 ~10 ~10-1 to 10-3

g-rays X-rays UV IR Microwave Radio

Nuclear excitation Core electronic Outer electronic Molecular vibration Molecular rotation Nuclear/Electro
(PET) excitation excitation nic Magnetic
(X-ray diffraction) Electronic energy Vibrational Energy Rotational energy spin Resonance
Bond breaking NMR (MRI)

Visible
➢ A molecule absorb only selected frequencies
➢ Various energy levels in the molecule are quantized (discrete)
Instrumentation for spectroscopy
Monochromators

Select wave length

• Power Source: For UV- Xe, H2 Or D2 and Tungsten Lamp. For IR- Nichrome wire, Globar, Laser source
• Monochromators: Prism, Gratings in combination with slits and mirrors
• Sample cell (Cuvette) Silica for UV-VIS and glass or Plastic, NaCl, NaBr for IR
• Wavelength Disperser: an information sorting system, spreads light out spatially according to its wavelength
• Photodetector: A transducer changing optical information into electrical information (Photomultiplier tube,
Diode array)
• Readout: digital (ADC), meter, strip chart recorder
General Set Up for single beam instrument

General Set Up for double beam instrument

Representation of spectra
100% 0%

Transmittance (%)

Absorbance
0%
100%

IR spectra of water UV spectra of water

Practical spectroscopy: Factors affecting- (Molecular Spectroscopy- by Banwell..…)

1. Width of spectral lines 100% 0%

a. Collision broadening (more severe in liquids than gases)

Transmittance (%)

Absorbance
b. Doppler broadening (in gases this effect is more)
c. Heisenberg uncertainty principle (less effect )
ℎ ℎ
δE x δt   10-34Js; h x δν x δt 
2p 2p 0%
100%
1
➔ δν  2p δ t If δ t = 10−8 𝑠, δν = 108 Hz

(Radio frequency region)

2. Intensity of spectral lines
a. Transition probability (TP) decided by selection rule
a. Allowed transition, TP  0 and
b. Forbidden transitions, TP = 0
b. Population of states (Boltzmann distribution)
N upper
= 𝑒 −∆𝐸/𝑘𝑇 , T = Temperature in K, k = 1.38 x 10−23 J𝐾 −1
N lower
Nupper = Nlower 𝑒 −∆𝐸/𝑘𝑇
Q. Calculate the ratio of molecules in the upper to those in the lower energy level when the separation between
these energy levels at 300 K is as follows
a) 11.9 J mol-1 b) 11.9 kJ mol-1 c) 119 kJ mol-1

N upper
= exp(-∆E/kT)], T = Temperature, k = 1.38 x 10−23 J𝐾 −1
N lower
= exp(-11.9/ 1.38 x 10−23 J𝐾 −1 x 300K) =
= exp(-11.9 𝑥 103 / 1.38 x 10−23 J𝐾 −1 x 300K) =
= exp(-119 x 103 / 1.38 x 10−23 J𝐾 −1 x 300K) =
c. Pathlength or Concentration of sample (Beer-Lambert Law)
“When a monochromatic light passes through an absorbing medium, the intensity of light gets decreased
exponentially with the thickness and concentration of the medium as the light gets absorbed by the medium”
➢ The decrease in intensity of the light with thickness (l) is proportional to intensity of
the incident light (I0)
➢ The extend of absorption depend on
➢ Concentration of the sample (c in mol/L) and
➢ The length (l in cm) with which the light passes through the medium

𝑰
The intensity of the transmitted light, % 𝒐𝒇 𝑻𝒓𝒂𝒏𝒔𝒎𝒊𝒕𝒕𝒂𝒏𝒄𝒆 (% 𝑻) = 𝟏𝟎𝟎 𝑰𝟎 = 100T
I = I0 10 -  c l 𝟏 𝟏𝟎𝟎
𝑰
A = log = log
𝑻 %𝑻
T = Transmittance = = 10 - c l
𝑰𝟎
A = 2 log 10 - log %T
𝑰𝟎
𝒍𝒐𝒈 = 𝜺𝒄𝒍 = 𝑨 (Absorbance)
𝑰 A = 2-log %T
A = -log T log %T = 2-A
➢ The Absorbance (A) is also called Optical density
➢ 𝜺 = Molar absorptivity / extinction coefficient in cm2 mol−1 (L mol−1 cm−1)
➢ (𝜺) represents the ability of the molecule to absorb light, ∣ 𝜺 ∣ is the transition probability
Deviations from Beer lamberts law Beer’s law suggests that
➢ a calibration curve is a straight line,
Absorbance, A = 𝜺𝒄𝒍 = -log T ➢ a y-intercept of zero,
➢ a slope of εl .

Limitations of Beer lamberts law

Fundamental (High Concentration > 0.01M):
➢ Molecules are in close proximity (electrostatic interaction)
➢ Cause deviation in absorptivity co-efficient
➢ Changes in refractive index
Chemical interactions :
➢ Shift of equilibrium, association, dissociation, etc
Instrumental factors.
Requirement of Monochromatic radiations (usually a small band width)
➢Polychromatic radiation always gives a negative deviation
➢The effect is smaller if the value of ε is essentially constant over the wavelength range
➢Always measure the absorbance maxima
Stray radiation
Arises from imperfections in the wavelength selector that allow light to enter the instrument and reach the detector
without passing through the sample, Like reflections, scattering etc
1. A solution of a compound shows a transmittance of 0.800 in a 1.00 cm cell at 525 nm. Calculate the transmittance in
a 5.00 cm cell at the same wavelength.

A = -log T =  c l
T1= 0.8, l1 = 1 cm
log 𝑇1 𝑙1
= T2= 0.8, l2 = 5 cm
log 𝑇2 𝑙2
log 0.8 1
=
log 𝑇2 5
log 𝑇2 = 5𝑥 log 0.8 = −0 ⋅ 5
𝑇2 = anti log( −0 ⋅ 5) = 0.316
Q. The molar absorptivity for a complex formed between bismuth (III) and Thiourea is 9.32X10 3 L cm-1 mol -1
at 470 nm. Calculate the range of permissible concentrations of the complex if the absorbance is to be no less
than 0.10 and not greater than 0.90 when the measurements are made in 1.00 cm cells.
Amin = 0.10; Amin = 0.90
A=cl
 = 9.32X10 3 L cm-1 mol -1
Cmax = A/  l
Cmin and Cmax = ?
l = 1.00 cm
Cmax = A/  l Cmin = A/  l
= (0.90/(9.32X10 3 L cm-1 mol -1 X 1.00 cm) = (0.1/(9.32X10 3 L cm-1 mol -1 X 1.00 cm)
= 9.6 X 10 -5 mol L-1 = 1.07 X 10 -5 mol L-1

Q. If the molar absorptivity of 1,10-phenanthroline is 12,000 litre mole-1 cm-1 and minimum detectable
absorbance is 0.004. Calculate the detectable minimum concentration for a 5.00 cm path length.

ε = 12,000 litre mole-1 cm-1; According to Beer – Lambert’s law A = εcl

A = 0.004; l = 5.00 cm; c = A/ εl
c=? 0.004
=
12,000litre mol−1 cm−1x 5cm

= 6.66 x 10-8 mol/Lit