Exp-03 Handout

Experiment 2

NAME:___________________________ BATCH:__________
ROLL NO:________________________ SEM:__________
SIGNATURE:_____________________ DATE:____________

DETERMINATION OF FLUORESCENCE QUANTUM YIELD & ESTIMATION

OF MOLAR EXTINCTION COEFFICIENT
(Anthracene and NBD-Octylamine)
AIM
To determine fluorescence quantum yield and estimate molar extinction coefficient of Anthracene and
NBD-Octylamine
THEORY
Fluorescence Quantum yield gives the efficiency of fluorescence process for particular system which
is defined as the ratio of number of photons emitted by a molecule via fluorescence to the number of
photons absorbed. There are two known methods to determine the quantum yield, (i) Comparative and
(ii) Absolute method. The experiment reported here is based on comparative method in determining the
fluorescence quantum yield. It requires one to have a standard molecule whose quantum yield is
known. Quinine sulfate will be used as standard for this experiment.
Therefore, from the fluorescence spectrum of sample and a standard, quantum yield of unknown
sample (Φsample) can be expressed in terms of known quantum yield of standard (Φstandard), which is
given below.
Fluorescence Spectrum Area of Sample
Φ Φ
Fluorescence Spectrum Area of Standard
So, optical matching should be performed before recording the fluorescence spectrum.
Molar Extinction Coefficient, (or molar absorption coefficient, or molar absorptivity), is a

measurement of how strongly a chemical species absorbs light at a given wavelength. According to
Beer–Lambert law, the probability that a photon will be absorbed in a medium is directly proportional
to the concentration of the absorbing molecule and to the thickness of the sample.
The absorbance or radiation density (RD or A) of a solution is defined by the equation:
A= log (I0/I)
where I0 is the intensity of the incident beam impinging on a cell containing the solution and I that of
the transmitted beam. The radiation density is related to the concentration of the solution as
A= εcl or
ε = A/cl (L mol−1 cm−1)
where l (cm) is the path length of the cell, c (mol/L) is the molar concentration (number of moles of the
solute dissolved per liter of the solution) of the absorbing species in the solution and ε is a constant of
proportionality called the molar extinction coefficient. Molar extinction coefficient depends upon the
wavelength of the incident radiation and is greatest where the absorption is most intense.
MATERIALS REQUIRED
NBD-Octylamine (synthesized in experiment 1), Anthracene, Quinine sulfate, conc. H2SO4,
Dichloromethane, Ethanol, Beakers, Micropipette, Fluorescence and UV-VIS spectrophotometers with
cuvette cells etc.
Experiment 2
PROCEDURE
Estimation of Molar Extinction Coefficient:
1. Prepare solution of NBD-Octylamine and Anthracene in DCM and ethanol respectively with
known concentrations.
2. Record the UV-VIS absorption spectra for both the samples.
3. Calculate Molar Extinction Coefficient, for both.
[Note: repeat the experiment for concurrency]
Determination of Fluorescence Quantum yield:

1. Dissolve few milligrams of Quinine sulfate in 0.5 molar H2SO4 and record its UV-VIS
absorption spectrum.
2. Record the following set of emission spectra for reference and compound,
a) Quinine sulfate and NBD-Octylamine
b) Quinine sulfate and Anthracene.
at the matched wavelength (described in next section)
3. Calculate the integrated fluorescence intensity (peak area) from the spectrum for reference and
compound.
4. Calculate the fluorescence quantum yield for both sets as per formula.
[Note: repeat the experiment for concurrency]
Observation, calculations and results:
a) Molar Extinction Coefficient of NBD-Octylamine:

b) Molar Extinction Coefficient of Anthracene:
c) Area of fluorescence spectrum for Quinine sulfate, NBD-Octylamine and Anthracene:
d) Fluorescence Quantum yield of NBD-Octylamine and Anthracene:
e) Include all spectra in your results:
f) Comment on the spectra and results:

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Experiment 2

DETERMINATION OF FLUORESCENCE QUANTUM YIELD & ESTIMATION

Molar Extinction Coefficient, (or molar absorption coefficient, or molar absorptivity), is a

Determination of Fluorescence Quantum yield:

Observation, calculations and results:

a) Molar Extinction Coefficient of NBD-Octylamine:

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