Advanced Analytical

Techniques
In Lecture 1
•Light is described as electromagnetic waves of energetic
particles ; photons and is usually described in terms of
wave properties and photon properties

E=hν =hc/λ =hcύ

In Lecture 2
• Radiation sources
• Ways of absorbing energy

• Which electrons get excited?

•Types of Electronic Transition

• Absorption Spectrum and its types

Radiation Sources
• Requirements:
1- Of constant intensity over the time interval during measurements.

2- Provide sufficient radiant energy over the wavelength where the

absorption is to be measured.

• Radiation sources divided in to three types:

1. Continuous sources
2. Line Sources
3. Lasers
Radiation Sources
• A beam of light carrying of only one discrete of λ is said to be:
Monochromatic Light
•A beam of light carrying radiation of several λ (s), is said to be:
Polychromatic or Heterochromatic Light.
•Generally energy per photon depends on the wavelength, the shorter
wavelength photon is higher in energy than longer wavelength
one.
If a substance is irradiated with EMR, the energy, E, of the
incident photons may be transferred to the atoms or molecules
raising them from the ground state to an excited state. This process
is called an absorption.

The energy of transition may be given by the following equation :

ΔE = Eex – Eg = hν

Where ν is the frequency of EMR at which the transition takes place ,

which is characteristic for each molecule.

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Absorbed E is lost in the form of
heat , light or molecular collision.

Absorption Emission
of light of light
increase decrease
E of E of
molecule. molecule.
Absorption Emission

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The electromagnetic spectrum and it’s interaction” Ways
of absorbing energy”
•all radiatns forms is the same and move with the speed of light, they
differ only infrequency and wavelength and the effect produce in
matter according to the following:
1. Transitional energy: Raising electrons to a higher energy level
when molecule absorbs energy in UV-visible region.
2. Vibrational energy: Raising the vibration of the constituent nuclei
when molecule absorbs energy in I.R region.
3. Rotational energy: Increasing rotation of the molecule around the
axis when molecule absorbs energy in F.I.R “Far Infrared” region.
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 A molecule may absorb light energy in three ways:

[1] by raising an [2] by increasing [3] by increasing the

electron to a higher the vibration of rotation of molecule
energy level constituent about its axis
(transitional energy) nuclei (vibrational), (rotational)
when molecule when molecule when molecule absorb
absorb absorb IR FT-IR irradiation.
visible and UV light. irradiation.

Molecular Spectra involve Electronic, Vibrational, or Rotational transitions.

• Electronic transitions • Vibrational • Rotational transitions

are vertical lines on such a plot transitions occur mostly between
since the electronic transition occur between rotational levels of the same
occurs so rapidly that the different vibrational state, although
internuclear distance can't vibrational there are many examples of
change much in the process. levels of the combination vibration
same electronic rotation transitions for light
state. molecules.
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 Kinds of Electronic Transitions
Electrons in a molecule can be classified into 2 types:
1. Closed-shell electrons (not involved in bonding; have high excitation E).
2. Outer shell electrons

Which electrons get excited?

The valency electrons in outer shell.
the outer electrons in an organic molecule may be:
1- Sigma () electrons
2- Pi () electrons electrons
3- Non-bonding electrons

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Which electrons get excited?
The outer electrons in an organic molecule may occupy one of three different
energy levels as follows:
1. Sigma () electrons: are bonding electrons which represent valence bonds formed due
to linear overlapping of the electronic clouds of S or sp orbitals. They posses the lowest
energy level “almost stable” Covalent single-bond electrons

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Which electrons get excited?
2. Pi () electrons: are bonding electrons constituting the Pi bonds “double bonds” and
result from lateral overlap “p” orbitals. They are of higher energy than sigma. double or
triple bonds

3. Non-bonding electrons : these are atomic orbitals of hetero atoms e.g. N,O, halogen or
S which do not participate in bonding. They usually occupy the highest energy level of
ground state.

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Outer electrons in an
organic molecule may
occupy one of
3 different energy levels
(σ, π or n energy level).

♣ A bond between two atoms consists of one or more molecular

orbital formed from overlap of the atomic orbitals of the electron
pair that forms the bond.

♣ Combination of two atomic orbitals gives rise to both a

bonding and antibonding molecular orbitals, The bonding orbital
has the lower energy and is thus occupied by the electrons in the
ground state.

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Types of Electronic Transition

•These transitions are:

•[1] In excited state, the σ-electrons occupy an anti-bonding energy level
(σ*) and the transition is termed: σ-σ* transition.
•[2] In excited state, π-electrons occupy an anti-bonding energy level π* and
the transition is termed: π - π* transition.
•[3] While the n-electrons may occupy σ* or π* levels to give n - σ*
transition or n - π* transition.
Arrangement of transitions according to E:

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Absorption Spectrum
When sample molecules are exposed to light having an energy
that matches a possible electronic transition within the molecule,
some of the light energy will be absorbed as the
electron is promoted to a higher energy orbital.
Plot of absorption intensity versus λ, is called
Absorption Spectrum
Absorption spectrum may be:
[a] Line spectrum

[b] Band spectrum

Absorption Spectrum
[a] Line spectrum:
occurs with atomic spectra such as sodium metal which has a sharp line at 589 nm
Absorption Spectrum
[b] Band spectrum:
occurs with molecules due to the presence of different vibrational and rotational
sublevels which the molecules may occupy on Transition to excited state.
That is rotational and vibrational
modes will be found combined with electronic
transition result in band rather than line spectra.
 UV-Visible Absorption Spectra
A spectrometer records the wavelengths at which
absorption occurs, together with the degree of absorption at
each wavelength. The resulting spectrum is presented
as a graph of absorbance (A) versus wavelength, as
in the spectrum shown below.
• For spectrophotometric analysis, measurements are made at a wavelength (λ max)
corresponding to a peak in the absorbance spectrum.

Why λmax ?
• This wavelength gives the greatest sensitivity- maximum response
for a given concentration of analyte.
• Errors due to wavelength drift and the bandwidth selected by
the monochromator are minimized because the spectrum varies
least with wavelength at the absorbance maximum.
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