Biosynthesis of 20-hydroxyeicosatetraenoic acid (20-HETE) and 12 (S)-HETE by renal cortical microsomes of the cynomolgus monkey

Abstract

Renal cortical microsomes of the cynomolgus monkey, Macaca fascicularis, were incubated with [14C] arachidonic acid in the presence of NADPH. The two main metabolites were identified as 20-hydroxyeicosatetraenoic acid (20-HETE) and 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE) by capillary GC-MS and by chiral phase HPLC. 12(S)-HETE was formed in highly variable amounts by the very same preparation of microsomes on different occasions. The microsomal biosynthesis of 12(S)-HETE was increased several-fold by addition of Ca++, NADPH (or NADH) and ATP, and the biosynthesis was reproducible in the presence of these agents. The biosynthesis was unaffected by proadifen (SKF-525A), however, and was inhibited by nordihydroguaiaretic acid. Some preparations of renal medullary microsomes and cytosol also contained 12(S)-lipoxygenase activity, which was not stimulated by Ca2+, NADPH and ATP and thus resembled the well-known platelet enzyme. Significant formation of 12(R)-HETE could not be detected. In conclusion, 20-HETE was identified as the main cytochrome P-450-derived metabolite and 12(S)-HETE as the main lipoxygenase product in microsomes of monkey renal cortex.