Chapter 17b

Fundamentals of
spectroscopy
1

Luminescence
Fluorescence
 Radiational transition between states with the same
spin quantum number
 Lifetime is always short 10-8 – 10-4 s.
Phosphorescence
 Radiational transition between states with different spin
quantum numbers
 Lifetime is 10-4 – 102 s
Chemiluminescence
 Is produced when a chemical reaction yields an
electronically excited species that emits light upon
return to the ground state
A + B  C* + D
C*  C + h 2

1
 3

Absorption vs. Emission Spectra

Spectra arising from fluorescence and phosphorescence come at lower energies

than absorption (the excitation energy). That is, molecules emit radiation at longer
wavelengths than the radiation they absorb.

Chapter 18 4

2
 Absorption vs. Emission Spectra

In absorption, wavelength λ0 corresponds to

a transition from the ground vibrational level
of S0 to the lowest vibrational level of S1.

Absorption maxima at higher energy

(shorter wavelength) correspond to the S0
to S1 transition accompanied by absorption
of one or more quanta of vibrational energy.

After absorption, the vibrationally excited S1

molecule relaxes back to the lowest
vibrational level of S1 prior to emitting any
radiation. Emission from S1 can go to any
vibrational level of S0. The highest energy
transition comes at wavelength λ0, with a
series of peaks following at longer
wavelengths.

Emission spectra is roughly a mirror image

of absorption.
5

In emission spectrum 0 comes at slightly lower energy

than in the absorption spectrum?
The λ0 transitions do not exactly overlap. In the emission spectrum, λ0 comes at
slightly lower energy than in the absorption spectrum. Why?

A molecule absorbing radiation is initially in its electronic ground state, S0, and it
possesses a certain geometry and solvation. An electronic transition to S1 is
much faster than the vibrational motion of atoms or the translational motion of
solvent molecules. Hence, when radiation is first absorbed, the excited S1
molecule still possesses its S0 geometry and solvation. Shortly after excitation,
the geometry and solvation change to their most favorable values for the S1 state,
which lowers the energy of the excited molecule.

When the S1 molecule fluoresces, it returns to the S0 state with S1 geometry and
solvation.
This unstable configuration must
S1 with S0 geometry
have a higher energy than that of and solvation S1 with S1 geometry
an S0 molecule with S0 geometry and solvation
and solvation.
Energy

absorbed

emitted
Energy
Energy


The net effect is that the λ0

emission energy is less than the S0 with S0 geometry S0 with S1 geometry
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λ0 excitation energy. and solvation and solvation

3
 In emission spectrum 0 comes at slightly lower
energy than in the absorption spectrum?
S1 with S0 geometry
and solvation S1 with S1 geometry
and solvation
 120 pm
H H

absorbed
116.5o 31o 119o

emitted
Energy
C
Energy
O C

Energy
Excitation O
H
H
110 pm
132 pm 109 pm

S0 with S1 geometry
S0 with S0 geometry
and solvation
and solvation

Excitation and emission spectra

 Excitation spectra
 Obtained by fixing an emission wavelength and scanning
the excitation wavelength (ex)
 Emission spectra
 Obtained by scanning the emission wavelength ( em) at a
fixed excitation wavelength

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 9

Chemiluminescence

Biolumienscence
Luciferase enzyme
Luciferin + O2 → Oxyluciferin + Light
Luciferins typically undergo an enzyme-catalysed oxidation
and the resulting excited state intermediate emits light upon
decaying to its ground state
10

5
Quantum yield ()
The ratio of the number of molecules that luminesce to the
total number of the excited molecules
 For a highly fluorescent molecule  approaches 1 (100%)
 For non fluorescent molecules  = 0
kf kf
 
k f  kisc  kic  k pd  k d k f  k radiationless

kf = Rate constant for fluorescence

kisc = Rate constant for intersystem crossing
kic = rate constant for internal conversion
kpd = rate constant for pre-dissociation
kd = rate constant for dissociation

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Transition type in fluorescence

 *  : not fluorescent
most molecules may have bonds ruptured by this transition
 n* : weak fluorescence
 * : strong fluorescence
 Higher absortpivities

 Larger energy difference between singlet and triplet sate 

smaller kisc

Chapter 18 12

6
 Fluorescence and structure
 Compounds containing aromatic rings give the most
intense fluorescence
 Highly conjugated double bond structures fluoresce.
 Unsubstituted aromatic compounds fluoresce in solution
with the quantum efficiency increases with the number of
rings and degree of condensation.
 Simple heterocyclics such as pyridine, and furan do not
fluoresce.
 Substitution on aromatic ring
Electron donating groups – increase fluorescence
Heavy atoms, such as Cl, Br, I…. Decreases
fluorescence.

Chapter 18 13

Fluorescence and structure: 1. Congugation

*  

Fluorescence increase as conjugation increase  H

N O S N

pyridine pyrole
Not Fluorescencet
H

N N
N
Higher molar
absorptivity

quinoline indole
Fluorescent
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Because of conjugation , it became fluorescent molecules

7
Fluorescence and structure: 2. Structural rigidity

Fluorene Biphenyl
 
Rigid: not flexible to bend

O
Zn
N N
OH 2
Weak fluorescence
Enhanced fluorescence
Fluorescence increases as molecules become more rigid.
15

Fluorescence and structure: 3. substitution

 Substitution for electron donating

groups can enhance fluorescence : -
OH, -NH2, -OCH3

 Electron withdrawing groups can

decrease the fluorescence :
Such as: -Cl, -Br, -I, NHCOCH3, -NO2, -
COOH

Chapter 18 16

8
 Temperature and solvent effects
 Fluorescence decreases with
Increase in temperature
 Decreased solvent viscosity
 Enhance radiationless deactivation by external
conversion

External Conversion (ec): Deactivation of the excited electronic state may also
involve the interaction and energy transfer between the excited state and the
solvent or solute in a process called external conversion. Low temperature and
high viscosity leads to enhanced fluorescence because they reduce the number of
collision between molecules, thus slowing down the deactivation process.

17

Foluorescence and solvent effects

In solvents containing heavy atoms e.g.

CBr4
Presence of other heavy atoms e.g. Pb, I
also decreases fluorescence and enhance
phosphorescence Highly fluorescent
 Increased intersystem crossing through
vibrational levels Br

In presence of oxygen

 Paramagnetic properties of oxygen promotes
intersystem crossing and conversion of excited
molecules to triplet state. Paramagnetic Highly phosphorescent
properties tend to quench fluorescence. Heavy atom effect

18

9
 Effect of pH on Fluorescence
The wavelength and emission intensity is different for protonated and unprotonated
forms of the compound. Examples

Fluorescence
relative intensity 20 0 20 10

Changing pH shifts equilibrium, causes protonation/deprotonation , i.e., modifies

fluorescent molecule.

Chapter 18 19

Effect of concentration on Fluorescence

 The intensity of fluorescence is directly proportional to the

concentration of fluorophores within a suitable range.
 At high concentration, fluorescence intensity decreases as a
result of self quenching.

Chapter 18 20

10
Effect of concentration
F  K ( P0  P )
P
 10 bC F
P0
F  K P0 (1  10 bC ) C

F  K P0  2.303bC 
2.303bC 
2


2.303bC 
3

 ...
 2! 3! 
For 2.303bC  0.05
F  2.303 K P0bC  KC

F: fluorescence intensity
P0 is the radiant power of the beam incident on the solution and
P is its power after it passes through a length b of the medium.
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Effect of concentration

Self absorption or
self quenching

 Deviation from linearity at high concentration

Higher terms become important
Self absorption: occur when the wavelength of emission
overlaps an absorption peak
Self quenching: from collision of excited molecules.

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Determination of Se in nuts

Higher terms

Self absorption

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