Optical Spectroscopy Instrumentation Design
155
5.2.4 Photoluminescence (Fluorescence and Phosphorescence)
Molecular and atomic fluorescence are examples of widely accepted photoluminescent
phenomena that are used to characterize analytes that encompass an extremely wide and
diverse range of applications. Fluorescence and phosphorescence spectrometry are examples of photoluminescence where the quantity of interest in the measurement is the radiant
power luminesced from the sample after absorption of a monochromatic incident light.
Fluorescence involves the emission from singlet to singlet states; that is, of the same multiplicity. Phosphorescence, however, involves a radiative transition from triplet to singlet
states; that is, of different multiplicity. The probability that a fluorophore will emit a photon (i.e., the quantum yield) is the number of times that a defined event occurs per photon
absorbed by the system. Therefore the quantum yield is a combination of the absorption
probability, in the form of a cross section, and the probability that the excited state will
decay by radiative emission. Such processes are, in turn, related to the wavelength or
energy of the incident radiation and the particular energy levels inherent within the sample,
and therefore quantum (photon) yields are a function of several parameters.
The probability of absorption at a given wavelength is expressed by the molar extinction coefficient ε(λ), m
2
mol
–1
. From this one can deduce that there will be a wavelength
or wavelengths of maximum absorption and spectral shape. The microenvironment of the
fluorophore, such as the solvent, presence of other ions and molecules, fluorophore concentration, and the surrounding temperature can all affect either the absorption or fluorescing
properties of the bulk sample. For many situations, the emitted radiant power, φ l , is proportional to the absorbed radiant power. Thus,
φ
φε
φ
l
k cl k c
=
=
2 303 0
0
.
′
(5.5)
where k is dependent on the species, its environment, and the efficiency with which the
excited molecule or atom returns to its ground state (possibly via intermediate energy levels) by the emission of a photon. For low absorbances (εcl < 0.01), the luminescence radiant power is directly proportional to the absorbed power and the sample concentration.
In the experimental diagram of a fluorimeter (Figure 5.6), the lamp provides broadband
light to the excitation monochromator, which selects the excitation wavelength. Subsequent
fluorescence is resolved and detected through the emission monochromator and photomultiplier tube detector. The emission intensity of luminescent materials can strongly depend
on a range of a given material’s parameters, and some examples of these parameters are
shown in Table 5.3.
5.3 The Fluorescence Spectrometer
It is the measurement of fluorescence emission that is the fundamental quantity in all fluorescence studies. This quantity is affected not only by the dependence on other samplerelated parameters but also by a series of instrument specific effects that can have significant
155
5.2.4 Photoluminescence (Fluorescence and Phosphorescence)
Molecular and atomic fluorescence are examples of widely accepted photoluminescent
phenomena that are used to characterize analytes that encompass an extremely wide and
diverse range of applications. Fluorescence and phosphorescence spectrometry are examples of photoluminescence where the quantity of interest in the measurement is the radiant
power luminesced from the sample after absorption of a monochromatic incident light.
Fluorescence involves the emission from singlet to singlet states; that is, of the same multiplicity. Phosphorescence, however, involves a radiative transition from triplet to singlet
states; that is, of different multiplicity. The probability that a fluorophore will emit a photon (i.e., the quantum yield) is the number of times that a defined event occurs per photon
absorbed by the system. Therefore the quantum yield is a combination of the absorption
probability, in the form of a cross section, and the probability that the excited state will
decay by radiative emission. Such processes are, in turn, related to the wavelength or
energy of the incident radiation and the particular energy levels inherent within the sample,
and therefore quantum (photon) yields are a function of several parameters.
The probability of absorption at a given wavelength is expressed by the molar extinction coefficient ε(λ), m
2
mol
–1
. From this one can deduce that there will be a wavelength
or wavelengths of maximum absorption and spectral shape. The microenvironment of the
fluorophore, such as the solvent, presence of other ions and molecules, fluorophore concentration, and the surrounding temperature can all affect either the absorption or fluorescing
properties of the bulk sample. For many situations, the emitted radiant power, φ l , is proportional to the absorbed radiant power. Thus,
φ
φε
φ
l
k cl k c
=
=
2 303 0
0
.
′
(5.5)
where k is dependent on the species, its environment, and the efficiency with which the
excited molecule or atom returns to its ground state (possibly via intermediate energy levels) by the emission of a photon. For low absorbances (εcl < 0.01), the luminescence radiant power is directly proportional to the absorbed power and the sample concentration.
In the experimental diagram of a fluorimeter (Figure 5.6), the lamp provides broadband
light to the excitation monochromator, which selects the excitation wavelength. Subsequent
fluorescence is resolved and detected through the emission monochromator and photomultiplier tube detector. The emission intensity of luminescent materials can strongly depend
on a range of a given material’s parameters, and some examples of these parameters are
shown in Table 5.3.
5.3 The Fluorescence Spectrometer
It is the measurement of fluorescence emission that is the fundamental quantity in all fluorescence studies. This quantity is affected not only by the dependence on other samplerelated parameters but also by a series of instrument specific effects that can have significant