52] Time-resolved fluorescence on flavins and flavoproteins

Abstract

Publisher Summary This chapter discusses the time-resolved fluorescence on flavins and flavoproteins. In this study, two methods are employed for obtaining lifetimes: phase and modulation fluorometry and single photon decay spectroscopy. In the first technique, the response to modulated light is determined, while in the second method the response to light flashes is measured. Phase fluorometry, using high modulation frequencies, has the advantage that single lifetimes can be obtained quickly with large precision. The disadvantage is that complex decays cannot be analyzed unless a whole range of frequencies is employed. The method of single photon counting is superior to phase fluorometry in two respects; it is more sensitive and it allows heterogeneous decays to be analyzed. A distinct disadvantage is that very short lifetimes can only be obtained, with reasonable accuracy, with the use of deconvolution procedures to avoid instrumental pitfalls and artifacts. Studies also reveal that the lifetime of flavin mononucleotide (FMN) fluorescence (4.7 ns at room temperature) is about a factor of two longer than that of flavin adenine dinucleotide (FAD), whereas the quantum yield of FMN is at least 10 times higher.