BIOTRANSFORMATION OF AMITRIPTYLINE BY CUNNINGHAMELLA-ELEGANS

A fungal biotransformation system as an in vitro model for mammalian d rug metabolism was investigated. Amitriptyline, a widely used antidepr essant, was effectively biotransformed within 72 hr by the filamentous fungus, Cunninghamella elegans. Eight major metabolites in HPLC eluti on order (11-hydroxyamitriptyline N-oxide, 11-hydroxynortriptyline, 11 -hydroxyamitriptyline, 10-hydroxyamitriptyline, 3-hydroxyamitriptyline , 2-hydroxyamitriptyline, nortriptyline, and amitriptyline N-oxide) we re produced at estimated molar ratios of 2:1:10:0.6:0.1:1 :2.5:0.5, re spectively. These metabolites were isolated by HPLC and identified by UV/MS analyses, as well as NMR spectroscopic analysis for most of thes e metabolites. In some cases, they were also compared with authentic s tandards. Glucose, culture age, and substrate concentration significan tly affected the extent of amitriptyline metabolism. Kinetic studies i ndicated that nortriptyline and 11-hydroxyamitriptyline were produced as initial major metabolites, The hydroxylated metabolite was excreted from mycelia, but amitriptyline and its N-demethylated metabolite, no rtriptyline, were not. An O-18(2) labeling experiment showed that the oxygen atoms in 11-hydroxyamitriptyline and 2-hydroxyamitriptyline wer e derived from molecular oxygen. The cytochrome P450 inhibitors SKF 52 5-A (1.5 mM), metyrapone (2.0 mM), and 1-aminobenzotriazole (1.0 mM) i nhibited the biotransformations of amitriptyline by 50, 75, and 95%, r espectively. A microsomal preparation was shown to catalyze the 11-hyd roxylation of amitriptyline, which was inhibited by SKF 525-A and carb on monoxide. The similarities of amitriptyline metabolism in C. elegan s and in humans and rats are discussed.