Fundamental of Spectroscopy

By Avani Khristi
Assistant Professor QA
Parul Institute of Pharmacy
 Spectroscopy

• Spectroscopy:
Spectrum + Scopein

Bands of Examination
different wavelength or
Evaluation
 Spectroscopy

• SPECTROSCOPY deals with the interaction of Electromagnetic

Radiation (EMR) with matter (solid, liq., gas, solution etc) to
identify (Qualitative) and/or to estimate (Quantitative) an
analyte.
• ELECTROMAGNETIC RADIATION (LIGHT)
• EMR can be described in terms of both particles and waves
(Dual nature of light)
• Light waves consist of perpendicular and oscillating electric and
magnetic fields
 Properties of EMR

Light waves can be characterized By:

• Wavelength (, Greek lambda): Distance from one
wave peak to the next.
• Units: m, cm, m, nm or Å
• Frequency (, Greek nu): Number of peaks that
pass a given point per second.
• Units: Cycles/second or s-1 or Hertz (Hz)
• Wavenumber (σ, Greek Sigma): Number of waves per
cm.
• Units: Kaiser, cm-1
• 1 Kaiser(k) = 1 cm-1, 1kK = 1000 cm-1
• Energy (E):
• Units : Ergs, Joules, Calorie
• eV
• eV/Photon
• eV/mole of photons
• eV/Einstein
• [1 mole of photon = 1 Einstein]
• eV: It is the amount of Kinetic energy gained by a
single unbound electron when it is accelerated
through an electronic potential of 1 volt.
• 1 eV = 1.602 x 10-19 Joules
• 1 eV = 1.602 x 10-12 Joules
• 1 Joule = 107 erg
• 1 erg = 6.241 x 1011ev
• 1 cal = 4.18 Joules
• Electromagnetic radiation consists of discrete packets
of energy, which we call photons.
• Photons are the particles of light or the quanta of light.
• Each photon carries the energy, E (Joule).
E  h
Where h is the Planck’s constant (=6.626x10-34 J.s)
The all characteristics of light can be related as follows:

c
E  h  h  hc

 Regions of electromagnetic radiation
Ejection of inner
shell electrons
Molecular
processes

change in the spin of

electrons or protons
that occur

configuration
when light is Change in
absorbed in nuclear
each region
Sales

Violet
Red
Orange
4th Qtr
 Classification of
Spectra

Line Band Continuous

 Line spectrum

• line spectrum: an emission spectrum consisting of

separate isolated lines.
• an emission (of light, sound, or other radiation)
composed of a number of discrete frequencies or
energies. Atoms are involved. eg: Ca, Al, Mg, Na etc.
• Line spectra are also called atomic spectra because
the lines represent wavelengths radiated from atoms
when electrons change from one energy level to
another.
 Band spectra

• Band spectra is the name given to groups of lines

so closely spaced that each group appears to be
a band, Here Molecules are involved e.g., Aspirin,
Benzene etc.
 Continuous Spectra

• an emission spectrum that consists of a

continuum of wavelengths. e.g. sunlight
 Classification of Spectroscopy

Spectroscopy

Absorption Emission Scattering Miscellaneous

1. UV-Visible 1. Atomic 1. Raman 1. X-ray diffraction

(200-800 nm) (200-800 nm) (200-2000 nm) / X-ray
•Atomic 2. Fluorescence / 2. Turbidometry Crystallography
•Molecular Molecular (200-800 nm) (to find 3D
2. IR 3. Nephelometry structure)
(2.5 – 16 16µm)
3. ESR (107nm)
4. NMR (1011nm)
5. X-ray (20nm)
 Wave properties of EMR

• Atomic spectroscopy is the study of the

electromagnetic radiation absorbed and emitted
by atoms. Atoms are in their lowest energy state
called Ground state. (Electronic)
 Molecular Spectroscopy

• A given molecule will specifically absorb only those wavelengths which have energies
that correspond to the energy difference of the transition that is occurring.

• Thus, if the transition involves the molecule jumping from ground state A to excited
state B, with an energy difference of ΔE, the molecule will specifically absorb radiation
with wavelength that corresponds to ΔE, while allowing other wavelengths to pass

through unabsorbed.
• The atomic spectra involves only transitions of
electrons from one electronic level to another
while the molecular spectra involve transitions
between rotational and vibrational energy
levels, in addition to electronic transitions.
• Hence the spectra of molecule are much more
complicated than those of atoms.
Particle / Photon Property of an EMR
• Transitions between electronic levels are found in the UV
and Visible region.
• Transition between vibrational levels, but within the same
electronic level, lie in the near and mid-IR and can also be
observed in Raman techniques.
• Low energy transitions involving the rotation of molecules
or of portions of molecules are due to selective absorption
in the far IR and microwave region.
Born Oppenheimer approximation:
• E = Eelectronic + Evibrational+Erotational

Terms applied to EMR:

1. Bathochromic shifts: Move to lower wave number
2. Hypsochromic shifts: Move to higher wave
number
3. Hyperchromic shift: Increase in intensity
4. Hypochromic shift: Decrease in intensity
 General Principles of spectroscopy
OR
Principle interaction between drugs and EMR
• Interaction involves definite type of
energy / wavelengths
The Energy interacted is exactly the
difference between two energy levels of
molecule.
e.g. Benzene interacts with only 255 nm
wavelength.
• The interaction takes place between
comparable energy of EMR and that of
portion of matter / drug.
• e.g. that part of benzene interacts with EMR
which has same energy as of EMR.
• Benzene + UV Vis. Valance e-Conjugated double bond
• Benzene + X-rays 1st e- orbital
• Benzene + NMR Spinning nucleus
• Benzene + IR Vibrating bonds
• The interaction causes instability or excited
state having momentary existence.
Energy of excited state is inversely related with life
of excited state
Stability α 1
Energy
Greater the energy of excited state, smaller the
lifetime.
Life span at excited state if 10-15 Sec.
Due to very short excited state, the molecule
seems as in ground state, & continuous absorption
of energy takes place which ultimately gives a
spectrum of respective EMR.
The type of EMR interacted depends upon the
type of matter or more specifically
structure of that compound
• This relationship is used for Qualitative
analysis of drugs.
• e.g.: 255nm interacts only Benzene
• 266 nm interacts only riboflavin
• 279 nm interacts only phenol
The intensity of EMR interacted is directly
proportional to the quantity of drug
This relationship is used for quantitative analysis
of drugs.
• The outcome or result of interaction is
expressed in terms of spectrum or graph.
 Types of Interaction

1. Absorption:
Retaining some amount of radiations by
matter / drug.
2. Emission:
Giving out radiation after interaction e.g.
Fluorescence, Phosphluorescence
3. Reflection:
Bouncing back of EMR at smooth or
shining surface.
4.Refraction:
• Bending of EMR due to change in density of
the medium.
5. Dispersion
Spreading or differentiation of radiations
based on wavelength.
6. Scattering:
Random change in direction of EMR due to
particles / obstacles.
Two types: Elastic scattering (Turbidometry,
Nephelometry), Inelastic scattering
(Raman)
7. Diffraction:
Bending of EMR at sharp edge or aperture.
Spreading of EMR in the direction of
propagation of EMR.
Bending α 1
Wavelength
Longer wavelength bends less.
8. Polarization
Cutting of the planes of vibrations of EMR
and allowing only one plane to pass
through. (process of conversion of
unpolarized light to polarized light.
9. Interference:
Physical mixing of EMR. Depends on the
phase or symmetry, Constructive
interference and Destructive interference.