Fluorescence and Its Measurement

Fluorescence is the radiative emission of light energy from an excited molecule for its return to ground state of same spin multiplicity, i.e., from 5i to So state. Fluorescence is a spin-allowed transition process and occurs strongly in a relatively short lime in the order of picoseconds to microseconds. The fluorescence emission spectra are almost the mirror image of the absorption spectra. Only one peak is 

The intensity of fluorescence is measured for quantitative analysis of fluorescent compounds present in different clinical and industrial samples. 

The intensity of fluorescence is directly proportional to the concentration of the fluorescent compound. If the target compound is not fluorescent, then it is converted into a fluorescent derivative by reaction with a suitable (nonfluorescent) reagent. The fluorescence emitted by the fluorescent compound is measured using a spectrofluorometer [6]. Most of the modem spectrofluorometers employ diffraction grating monochromators to select the appropriate wavelengths for maximum excitation and emission. The basic components of a fluorometer are a light source, an excitation monochromator, a sample holder, an emission monochromator, and a fluorescence detector as shown in Fig. 6.6. 

The most commonly employed lamps are medium- and high-pressure mercury lamp or xenon arc lamp, having an output covering the whole UV-Visible spectrum range. Xenon arc lamp operated stroboscopically is preferred for its continuous output. The lamp is operated in a current of air to disperse the toxic ozone formed from oxygen on exposure to UV radiation. 

The slit width of the monochromator is adjusted to select the wavelength for maximum absorption by the sample and allow its transmission for excitation of the sample. 

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