Advantages of Fluorescence and Phosphorescence

Luminescence spectroscopy may be divided into two major areas fluorescence spectroscopy and phosphorescence spectroscopy. The differences between the two are based mostly on the time frames on which the phenomena of fluorescence and phosphorescence occur, phosphorescence decaying much more slowly (often taking several seconds) than fluorescence subsequent to excitation. Slight differences between the instrumentation used to observe fluorescence and that to observe phosphorescence take advantage of the temporal distinction between the two luminescence phenomena. Chemiluminescence is a form of fluorescence differing only in the fact that a chemical reaction as opposed to incident light generates the excited state. 

The excitation spectrum of a molecule is similar to its absorption spectrum, while the fluorescence and phosphorescence emission occur at longer wavelengths than the absorbed light. The intensity of the emitted light allows quantitative measurement since, for dilute solutions, the emitted intensity is proportional to concentration. The excitation and emission spectra are characteristic of the molecule and allow qualitative measurements to be made. The inherent advantages of the techniques, particularly fluorescence, are ... 

CL-based analytical methods have several advantages such as rapidity, specificity and cost-effectiveness one minor disadvantage is that they require semi-skilled personnel. There is no requirement for an excitation source as in fluorescence and phosphorescence, a monochromator (often not even a filter), or the use of radioactive or hazardous chemicals. Thus, CL is advantageous for routine analysis. 

A major advantage of luminescence lifetime measurements is their use for spectral assignment. Specifically, the assignment of the luminescence bands as fluorescence or phosphorescence is primarily determined via luminescence lifetime measurements. As a rule of thumb, lifetimes on the order of microseconds and longer (milliseconds, seconds) are normally indicative of phosphorescence, while fluorescence lifetimes are normally on the sub-microsecond level (nanoseconds, picoseconds etc). It should be noted, however, that fluorescence and phosphorescence lifetime values vary from one case to another, depending on the system under study as well as... 

In polymer blends the features one would like to characterize are the amount of interphase in the system and the nature of its properties. An interesting approach to this probln takes advantage of the different sorption properties of oxygen in the two polymers making up the blend. Sorption is the product of solubility times diffusion. Fluorescence and phosphorescence quenching provide a measure of sorption For the process... 

Several types of labels have been used in immunoassays, including radioactivity, enzymes, fluorescence, luminescence and phosphorescence. Each of these labels has advantages, but the most common label for clinical and environmental analysis is the use of enzymes and colorimetric substrates. 

The use of excited triplet states as probes occurred in concert with the development of fluorescent probe methodologies but have not been employed to the same extent. Phosphorescence quantum yields in solution are generally much lower than fluorescence quantum yields, and when the emission of triplet states is measured, the signal-to-noise advantage of the fluorescent probes is lost. In most cases, triplet excited states are followed by their absorption spectra using laser flash photolysis. A second reason for the scarcer reports on the use of excited triplet states probes in supramolecular systems is the fact that the laser flash photolysis technique has not been as widespread as fluorescence techniques. This situation has changed over the past decade and we expect that the number of smdies which employ excited triplet states will increase. 

The ketone group is a useful model because it can be excited selectively in the presence of other groups commonly contained in polymer chains, such as the phenyl rings in polystyrene, and so the locus of excitation is well defined. Furthermore, there is a great deal known about the photochemistry of aromatic and aliphatic ketones, and one can draw on this body of information in interpreting the results. A further advantage of the ketone chromophore is that it exhibits a number of photochemical processes from the same excited state. Thus one has a probe of die effects of the polymer matrix on these processes by determination of the quantum yields. The competing processes include (1) fluorescence (Eq. 26), (2) phosphorescence (Eq. 27), (3) the Norrish type-I reaction (Eq. 28), (4) the Norrish type-II reaction (Eq. 29), (5) photoreduction (Eq. 30), (6) the... 

Elsewhere in this volume Millhauser et al. have discussed the application of nitroxide electron paramagnetic resonance (EPR) spin labels to the study of the structure and dynamics of biopolymers. Another type of EPR spin label that also is useful for investigating biopolymer systems is provided by the photoexcited triplet state of an intrinsic chromophore, because a triplet state carries electronic paramagnetism. A major advantage of the photoexcited triplet state of an intrinsic chromophore over an extrinsic spin label such as a nitroxide adduct is the relatively small structural perturbation caused by the former, which consists only of a localized electronic excitation. Although not as widely exploited as fluorescence, the phosphorescence of proteins, originating from the photoexcited triplet state, has received a great deal of attention. EPR afficionados have a natural attraction to photoexcited triplet states that dates back to the... 

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