Fluorescence spectrum determination

The optical train employed for photometric determinations of fluorescence depends on the problem involved. A spectral resolution of the emitted fluorescence is not necessary for quantitative determinations. The optical train sketched in Figure 22B can, therefore, be employed. If the fluorescence spectrum is to be determined the fluorescent light has to be analyzed into its component parts before reaching the detector (Fig. 28). A mercury or xenon lamp is used for excitation in such cases. 

Nalidixic acid has a strong fluorescence spectrum which has been used for its determination in biological fluids.(8)(9)(24)(26)(40)... 

Exciplex methodhas also been proposed for droplet temperature measurement. In an oxygen environment, however, the fluorescence from the exciplex is quenched by the oxygen. In addition, fuel droplets may contain aromatic hydrocarbons that can produce fluorescence emissions, masking the fluorescence spectrum of the dopants used for the temperature determination. 

The fluorescence quantum yield of a compound may be determined by comparing the area under its fluorescence spectrum with the area under the fluorescence spectrum of a reference compound whose fluorescence quantum yield is known. The spectra of both compounds must be determined under the same conditions in very dilute solution using a spectrometer incorporating a corrected spectrum capability, in order to overcome any variation in detector sensitivity with wavelength. 

The energy of the first excited singlet state can be determined from the wavelength at which the first vibrational band in the absorption spectrum coincides with the vibrational band in the fluorescence spectrum ... 

Fluorescence quantum yields are usually determined by integration of the fluorescence spectrum (and subsequent normalization using a standard of known fluorescence quantum yield in order to get rid of the instrumental factor k appearing in Eqs 3.17 or 3.18 see Chapter 6). In practice, attention should be paid to the method of integration. 

It should again be emphasized that the determination of a true fluorescence spectrum and of a quantum yield requires the use of very dilute solutions or there may be some undesirable effects. 

The relative changes in intensity of the vibronic bands in the pyrene fluorescence spectrum has its origin in the extent of vibronic coupling between the weakly allowed first excited state and the strongly allowed second excited state. Dipole-induced dipole interactions between the solvent and pyrene play a major role. The polarity of the solvent determines the extent to which an induced dipole moment is formed by vibrational distortions of the nuclear coordinates of pyrene (Karpovich and Blanchard, 1995). 

The excitation spectrum of a fluorescent material, i.e., the incident radiation spectrum required for the induction of fluorescence, is determined by the absorption spectrum of the fluorescent material, which it often closely resembles, and by the efficiency with which the absorbed energy is transformed into fluorescence. Normally, the excitation spectrum is of higher photon energy (shorter wavelength) than that of the corresponding fluorescence emission, and in sensor schemes this has an effect in the choice of preferred fluorescent agent, compatible with appropriate optical detection devices. 

Native fluorescence of a protein is due largely to the presence of the aromatic amino acids tryptophan and tyrosine. Tryptophan has an excitation maximum at 280 nm and emits at 340 to 350 nm. The amino acid composition of the target protein is one factor that determines if the direct measurement of a protein s native fluorescence is feasible. Another consideration is the protein s conformation, which directly affects its fluorescence spectrum. As the protein changes conformation, the emission maximum shifts to another wavelength. Thus, native fluorescence may be used to monitor protein unfolding or interactions. The conformation-dependent nature of native fluorescence results in measurements specific for the protein in a buffer system or pH. Consequently, protein denatur-ation may be used to generate more reproducible fluorescence measurements. 

The fluorescence spectrum of the nonsteroidal anti-inflammatory agent piroxicam 21 has been determined in a variety of solvents (Scheme 7) <1999PCP4213>. The key observations are that the molecule exists with a strong H-bond between the phenolic OH and the adjacent amide. A very high Stokes shift in the excited state was observed and attributed to the proton-transfer event (tautomerization) between the phenolic and amide oxygens (cf. 21 —>63). In the case of protic solvents, such as water, the open conformation 64 was observed. 

With an interplanar separation of 3.73 A, 4,4 -paracyclophane is the lowest member of the series to exhibit an alkylbenzene absorption spectrum and the broad structureless fluorescence spectrum of this molecule with a peak intensity at 3400 A is by definition an excimer band further separation of the aromatic rings in 4,5 and 6,6 -paracyclophanes restores the fluorescence spectrum to that of the alkylbenzenes. These observations by Rice et al.115 illustrate the critical nature of the interplanar separation in determining the extent of interaction between -electron systems in the ground and excited configurations. 

From the steady state fluorescence spectrum of indole in water a fluorescence quantum yield of about 0.09 is determined. Since the cation appears in less than 80 fs a branching of the excited state population has to occur immediately after photo excitation. We propose the model shown in Fig. 3a). A fraction of 45 % experiences photoionization, whereas the rest of the population relaxes to a fluorescing state, which can not ionize any more. A charge transfer to solvent state (CITS), that was also introduced by other authors [4,7], is created within 80 fs. The presolvated electrons, also known as wet or hot electrons, form solvated electrons with a time constant of 350 fs. Afterwards the solvated electrons show no recombination within the next 160 ps contrary to solvated electrons in pure water as is shown in Fig. 3b). 

The Practical Determination of C(0- The time-dependent fluorescence Stokes shift of the spectrum should manifest itself as (i) a rapid decay in the fluorescence intensity on the blue edge of the fluorescence spectrum, (ii) a... 

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