Principles of Fluorescence Spectroscopy

328 QUENCHING OF FLUORESCENCE
P8.3. Quenching in the Presence of Cyclodextrins: Figure
8.73 shows iodide quenching of naphthalene with 3 mM
β-cyclodextrin. Figure 8.74 shows iodide quenching of
naphthalene with 1% v/v benzyl alcohol with increasing
amounts of β-CD. Assume the unquenched lifetime of
naphthalene is 45 ns. Explain the results.
Figure 8.73. Quenching of naphthalene by iodide in the presence of
3.00 mM β-cyclodextrin. Revised and reprinted with permission from
[308]. Copyright © 1990, American Chemical Society.
Figure 8.74. Quenching of naphthalene by iodide in the presence of
benzyl alcohol (1% V/V) at various η-CD concentrations. Revised and
reprinted with permission from [308]. Copyright © 1990, American
Chemical Society.
P8.4. Separation of Static and Dynamic Quenching of
Acridone by Iodide: 176 The following data were obtained
for quenching of acridone in water at 26CE KNO 2 is used
to maintain a constant ionic strength, and does not
quench the fluorescence of acridone.
M KI M KNO2 F0 /F
0.0 1.10 1.0
0.04 1.06 4.64
0.10 1.00 10.59
0.20 0.90 23.0
0.30 0.80 37.2
0.50 0.60 68.6
0.80 0.30 137.0
A. Construct a Stern-Volmer plot.
B. Determine the dynamic (K D ) and static (K S ) quenching
constants. Use the quadratic equation to obtain
K D and K S from the slope and intercept of a plot of
K app versus [Q].
C. Calculate the observed bimolecular quenching constant.
The unquenched lifetime τ 0 = 17.6 ns.
D. Calculate the diffusion limited bimolecular quenching
constant, and the quenching efficiency. The diffusion
constant of KI in water is 2.065 x 10 –5 cm 2 /s
for 1 M KI (Handbook of Chemistry and Physics,
55th Edition).
E. Comment on the magnitude of the sphere of action
and the static quenching constant, with regard to the
nature of the complex.
P8.5. Separation of Static and Dynamic Quenching: The following
table lists the fluorescence lifetimes and relative
quantum yield of 10-methylacridinium chloride (MAC)
in the presence of adenosine monophosphate (AMP). 18
[AMP] (mM) τ (ns) Intensity
0 32.9 1.0
1.75 26.0 0.714
3.50 21.9 0.556
5.25 18.9 0.426
7.00 17.0 0.333
A. Is the quenching dynamic, static, or both?
B. What is (are) the quenching constant(s)?
C. What is the association constant for MAC-AMP
complex?
D. Comment on the magnitude of the static quenching
constant.
E. Assume the MAC–AMP complex is completely
nonfluorescent, and complex formation shifts the
absorption spectrum of MAC. Will the corrected
excitation spectrum of MAC, in the presence of nonsaturating
amounts of AMP, be comparable to the
absorption spectrum of MAC or the MAC-AMP
complex?