Spectroscopic Techniques

Various energy states associated with molecules / atoms of matters.

• At any given moment particles of matter possess different energy states. These energy
states can be changed when particles interact with photons of electromagnetic
radiation.
Possible energy states that can be occupied by particles of matter are:
1. Rotational energy states (with the smallest differences between energy
change)
2. Vibrational energy state (with greater energy difference between energy states
than rotational)
3. Electronic transition energy state (With the highest energy difference)
• The ground state is the state with lowest possible energy and is also the state of high
probability at room temperature.
What happens when a certain radiation happens to interact with a substance?
• Depending on the amount energy of photons of radiation, the energy states of particles
of a substance maybe changed through energy absorption or emission.
• But take note that energies difference between energy states are quantised just the
same as energy of photons.
• For example, for an electron to move from the ground electronic state S0 to the first
electronic excited S1 state it needs to absorb a photon of radiation with right amount
of energy (E = hV) equivalent to the energy difference between S0 and S1.

• The diagram below shows some possible energy state transition.

• Take note that rotational energy state change requires the smallest amount of energy.

Major points about energy states of particles and impact of radiation

• Particles (atoms/ molecules) at given moment may possess various energy states, of
rotational, vibrational and electronic.
• To change from one energy state to another a quantised (fixed) amount of energy is
absorbed or emitted.
• An electromagnetic radiation of the right amount of photon energy equivalent to the
energy states difference is required for a given transition.
• To transit from a higher energy state to a low energy state a radiation with photon
energy equivalent to the energy gap between energy state is emitted.
• The field of spectroscopic make use of radiations that possess the right amount energy
to cause a change in energy state.
• Examples of some transitional energy change employed is spectrometry.
1. Electronic transitions:
• Requires photons of energy from the UV and Visible regions.
• Meaning only photons of UV and visible radiations possess enough energy
for electronic transition,.
• UV- visible spectrometry techniques make use of these transitional energy
states,
•

2. Vibrational transitions:
• Requires photons of energy from the Infra-red region.
• Meaning photons of Infra-red radiations are equivalent to energy difference
between vibrational states.
• Infra-red and Raman Spectrometry techniques make use of these transitional
energy state.
3. Rotational transitions:
• Require photons of energy from microwaves
• Meaning photon energies of microwaves are equivalent to the difference in
energy of rotational states.
• Rotational spectroscopies make use of these transitional energy states.
UV- visible Spectrometry
• The use of photons from the UV and visible regions to quantitatively or qualitatively
study the compositions of some chemical compounds.
• Radiations of wavelength range between 200nm – 800nm are employed in this
technique.
• These type of radiation cause transition of electrons from low energy molecular
orbitals to high energy molecular orbitals.
• Take note electrons in molecules occupy certain probability spaces called macular
orbitals as shown in the diagram below.
• The blue arrows in the diagram below shows the most dominant electrons
transitions caused by UV- visible photons.
• This common transition is known as HOMO to LUMO transition
HOMO: Highest occupied molecular orbital
LUMO: Lowest unoccupied molecular orbital
• The other transitions shown with orange arrows requires photons of higher energy
than that of UV- Visible photons.

What type chemical compounds can be analysed through UV-visible spectrophotometry?

• Conjugated compounds ( compounds with alternating double/triple bonds and single

bonds
• Complex ions, in particular complex ions of transitional metals.
 UV -visible Spectrophotometer
• This is a Spectroscopic instrument that measure the absorbance of UV- visible
radiation by chemical compounds.
• The instrument consists of various components as shown in the flow diagram below.

UV-visible spectrum
• UV-visible spectrum is graphical representation of absorption curve for a compound
at a given wavelengths.

• The wavelength with the highest absorbance is called lambda maximum and is
different for each compound. Therefore, we can use lambda maximum to identify a
compound.
 Transmittance and Absorbance in UV- visible spectrometry
• Transmittance (T) is a fraction of incident intensity that is transmitted out
of the material T= It/I0
• Absorbance (A) is the amount light that is absorbed by a substance.
• Absorbance A and Transmittance are logarithmically related as shown
below
The Beer- Lambert s’ Law
Anther important aspect of UV- visible spectrometry is that the measured absorbance is
found to be directly proportional to the concentration of a the absorbing substance. This
relationship is known as The Beer- Lambert s’ Law.
 UV-visible absorbance standard curve
• A plot of absorbance of well-known concentrations of a substance.

• It can be used to determine unknown concentrations if the absorbance is obtained

from the spectrum.
 Serf Assessment Tutorial Questions
(1) Give the wavelength range of the following spectra regions

(a) U.V
(b) Visible
(c) U.V -visible region
(2) Which type of energy transition are equivalent to the photon energy in U.V-visible
region?
(3) Which electrons molecular orbital transitions are most involved UV- visible
spectrometry.
(4) What is a monochromator?
(5) What kind of data are obtained from the U.V visible spectrum? Explain clearly what
these data represent.
(6) A certain sample was put in a spectrophotometer sample holder and irradiated
(exposed) with radiations from the UV-visible region, but no trance of absorbance
was recorded from the spectrum. What conclusion would you make from this
observation?
(7) Explain how you would obtain a standard curve of concentrations vs absorbance from
experimental data.
(8) If the absorbance (A) of radiation by particular sample solution is 0.6 . Find the
transmittance (T) of this sample.
(9) In the context of spectrophotometric analysis, what is a “blank sample”?
(10 ) In the context of spectrophotometric analysis, what is an interference?

(11 Calculate the absorbance at 530 nm for the following concentrations of potassium
permanganate in a 1 cm cuvette. Molar absorptivity constant is 2 200 L mol-1 cm-1

(a) 1.5 x 10-4 mol /L b) 9.0 x 10-5 mol/L

 Infra-red spectrophotometry

• A spectroscopic technique that involves the interaction of infra-red radiation with

matters.
• This technique is based on monitoring the absorbance/ transmittance of radiation that
causes change in vibrational state of chemical bonds of molecule.
• This technique is used to identify different types of functional groups present in a
compound.
•

Vibrational energy states of molecules

• Covalent bonds are not static but rather act like stretching springs. This property
makes covalent bonds to stretch back and forth (stretching vibrations) and also
bending vibrations.

• Molecules of substance do vibrate along their chemical bonds.

• Different chemical bonds vibrate at different frequencies based on their bond strength.
• The diagram below shows possible vibrational mode of a triatomic molecule
• The stronger the bond the greater the vibrational frequency and the weaker the
chemical bonds the lower the vibrational frequency.
• Photons from the infra-red region have enough energy (E=hv) that is exactly
matching the energy gap between bond vibrational energy states of molecules.

• The lowest vibrational energy state of a molecule is the ground state vibrational states

• IF a photon of energy equivalent to the difference in vibrational energy state is

absorbed then a molecule will move from lower vibrational state to a higher
vibrational state

• Different chemical bonds/ functional groups absorb different vibrational energies

based on their bond strength.

• Absorption of vibrational radiant energy normally occur at particular frequency/

wavenumber, which serves as the characteristic property of that chemical bond/
functional group.
 Infra-red spectrum
• Infra-red spectrum shows the wavenumber or the frequency of radiation at which a
particular functional group / chemical bonds absorbs vibrational energy. On the
spectra graph it always appears as vibrational peaks pointing downward or upward,
depending the y -axis variable.
• Each functional group has a characteristic wavenumber at which it which an
absorption bands/ peak appear.

• In general, the stronger the chemical bonds the greater the vibrational energy.
Vibrational peaks/bands of strong chemical bonds tend to appear at higher
values of wavenumbers.

• Take note that at wavenumbers, where there is no significant absorption of

infrared, most of the radiations are almost 100% transmitted through the
sample.

• When absorption of infra radiation is significant, absorption peaks appear at

those wavenumbers and the transmitted radiation is highly reduced.
• Infrared absorption locations for common functional groups are tabulated in a
table as shown below. Such a table can be used to identify various functional
groups from a given IR spectrum.
Serf assessment on I R spectrum (use the table below as you work through spectrum
activities)

Bond Functional group Absorbance (cm-1)

O–H Alcohols 3200 – 3600 / strong and broad*

O–H Carboxylic acids 2500 – 3200 / medium and very

broad*
C=O Aldehydes / ketones / 1680 – 1750 / strong and sharp
carboxylic acids/ esters

C-O Alcohols / esters / ethers 1050 – 1300 / medium

C-H Alkanes / alkenes etc 2850 – 3100 / medium

1. What wavenumber would appear on an IR peak, if the frequency of radiation absorbed by a

molecule was 2.5 x 1013 Hz?
2) ethanol (CH3CH2OH)

displayed
formula

i.r. spectrum

Bond / (Functional group) Absorption / cm-1

2) ethanoic acid (CH3COOH)

displayed
formula

i.r. spectrum

Bond / (Functional group) Absorption / cm-1

3) Ethyl Ethanoate (CH3COOCH2CH3)

H O H H

H C C O C C H

H H H

i.r. spectrum

Bond / (Functional group) Absorption / cm-1

1750

1250

3000
 Raman Spectroscopy

• Raman spectroscopy is a technique that relate the energy of inelastic scattered

radiation to the vibrational energy states of sample molecule.

Three types of scattering processes that can occur when light

interacts with a molecule.

• Since it involves the change in vibrational energy states just like the IR, the technique
is another example of vibrational spectroscopy.

• Raman spectrometry technique make use of radiations mostly from visible region and
near-Infrared.

• Unlike IR which involves the absorption of radiation to enact changes in vibrational

states, Raman spectroscopy brings about the same vibrational changes but through the
scattering of radiation.
• As you can see in the diagram above, the scattering of radiation is classified into three
types based on energy change between the incident radiation and scattered radiation.

• The diagram below shows different types of scattering with their related energy
transition.

What exactly happens during scattering of radiation.

• Most of the scattered radiation ( almost 99.99%) from the

sample are Rayleigh. These scattering effect does not involve
energy change, hence the scattered photon has the same
wavelength, frequency and energy as the incident radiation.
Such scattering is a type of elastic scattering.
 • Rayleigh scattering has no effect on vibrational states of
molecules, therefore has not affect on Raman spectrum
• The other two inelastic scattering (stoke and anti-stoke affect
the vibrational states of molecules) hence they are of Raman
effect.
• At room temperature the stoke Raman scattering has higher
probability of occurrence than anti-stoke,

What do we record during scattering?

Study the schematic illustration below to get the concept about

Raman scattering
• The peak intensity of Raman photons
• Characteristic Raman shifts of various chemical bonds

• The difference in wavenumber of incident radiation (laser) and

wavenumber of Raman scattered radiation is called the Raman
shift. ( )
• proportional to the difference between bond vibrational
energy levels.

• This is calculated by the following equation:

• Where Δν~ is the shift in wavenumbers,

λ0, is the laser wavelength, and

λ1 is the scattered wavelength.

Example of Raman spectrum

• Raman spectrum produce provide same information as that of IR.
i.e – specific bond/ functional vibrational peaks
• Each functional group has a specific Raman shift peak
• Just like IR we can make use recorded wavenumber data base to
identify the functional group.
• IR and Raman spectroscopic techniques are complementing each
other.
• Normally IR active (strong signals) molecules have weaker signals
in Raman and vice versa
• Raman spectrum for each substance is unique , hence we can use
Raman spectrum to identify substances or even determine their
purity

The diagram below represent Raman spectrum for diamond and

polystyrene.

• Pure Dimond highly uniform (identical bond connections) hence

there is only one vibrational peak for such bonds.
• Polystyrene on the other hand is a heterogeneous polymer with
different bond connectivity hence there are various vibrational
peaks
Take note that there are various methods of Raman spectroscopy such

Types of Raman Spectroscopy

• Surface Enhanced Raman Spectroscopy (SERS)
• Tip Enhanced Raman Spectroscopy (TERS)
• Surface Enhanced Resonance Raman Spectroscopy (SERRS)

*Detailed descriptions of these techniques are beyond the scope of this module. But
you can research on each to find out about the basic concepts.
 Serf Assessment Questions

1. State the major similarity of Raman and IR spectroscopic techniques.

2. State the major differences between Raman and IR spectroscopic

techniques.

3. Which type of radiation are more often used in Raman

spectroscopy?

4. What type of energy transition is associated with Raman effect?

5. Differentiate between Rayleigh and Raman scattering effect.

6. Define the term “Raman shift” and explain its major role in Ramm
spectroscopy.

7. Describe briefly the main concepts of Surface enhanced Raman

spectroscopy (SERS)

8. More energetic radiations such as UV provide stronger Raman

signals but more aften are not that recommended. Explain why?

9. State the advantages of Raman Spectroscopic technique as compared

to other spectroscopic techniques.