[39] Direct fluorometric methods for measuring mexed-function oxidase activity

Publisher Summary

This chapter focuses on the direct fluorometric methods for measuring mixed-function oxidase activity. Ullrich and Weber developed a direct fluorometric method to measure the mouse liver microsomal cytochrome-P-450-dependent O-dealkylation of 7-ethoxycoumarin to yield 7-hydroxycoumarin. Recently, Burke and Mayer described a direct fluorometric assay to measure the O-dealkylation of ethoxyresorufin. The broad substrate specificity and ready induction of the monooxygenase has allowed the development of a large number of specific assays for the enzyme system. Many of the methods require solvent extraction of metabolites and subsequent analysis to quantitate the concentration of the metabolite. The number of manipulations required in these assay methods and the problems involved in solvent extraction of compounds of intermediate polarity complicates and extends the time required for analysis of enzyme activity. Several direct spectrophotometric and spectrophotofluorimetric assays are developed to obviate these problems, and two fluorescent assays are encapsulated in this chapter.