Geoscience Reference
In-Depth Information
fluorescence lifetimes include time correlated single-photon counting (TCSPC) spectros-
copy and fluorescence lifetime imaging microscopy (FLIM), which spatially resolves the
fluorescence lifetime of a sample (Lakowitz,
2006
, chapters 4 and 17). Such techniques are
not discussed here but additional information can be found in the references at the end of
this chapter.
1.3.4.3 Fluorescence Efficiency (Quantum Yield)
Several processes compete with fluorescence for the deactivation of the lowest excited
singlet state; therefore the intensity of fluorescence (
I
f
), is obtained by multiplying
I
o
(
Eq. [1.2]
) by the fraction of excited molecules that “actually” fluoresce as shown in
Eq. (1.3)
:
I
=Φ
I
(1.12)
f
F
0
Φ
f
is called the quantum yield of fluorescence or the fluorescence efficiency.
The quantum yield can range from 1, when every molecule in an excited state undergoes
fluorescence, to 0, where no fluorescence takes place. The fluorescence quantum yield can
be related to the fluorescence lifetime by
k
k
Φ
F
==
τ
R
k
(1.13)
RF
F
Therefore the fluorescence lifetime,
τ
F
, is a measure of the fluorescence quantum yield,
Φ
F
. This indicates that the rate constant for the radiative decay processes
k
R
, is con-
stant for a particular fluorophore as it is an intrinsic electronic property of the mole-
cule. Consequently the fluorescence lifetime is influenced by changes in the nonradiative
decay pathways. For example, a subsequent increase in nonradiative decay rates will
reduce the fluorescence lifetime. As such, fluorescence lifetimes are extremely sensitive
to the molecular environment surrounding the fluorophore. Therefore, this makes fluo-
rescence lifetime measurement of individual fluorophores present in complex aquatic
samples difficult to interpret.
The relationship between phosphorescence intensity (
I
P
) and analyte concentration is
similar to that for fluorescence (
Eq. [1.7]
).
I
=
ΦΦ
0
110
I
(
−
−
Î
cl
)
(1.14)
P
STP
In
Eq. (1.14)
,
I
0
,
Îc
, and
l
are as defined in Beer and Lambert's Law (
Eq. 1.6
),
Φ
ST
is the
quantum yield of intersystem crossing, and
Φ
P is the quantum yield of phosphorescence.
The former represents the fraction of excited molecules that undergo intersystem crossing
from the lowest excited singlet state to the lowest triplet state, and the latter is the fraction
of excited molecules having undergone intersystem crossing that are actually deactivated
by phosphorescence.
