The hepatic microsomal biotransformation of delta 5-steroids to 5 alpha, 6 beta-glycols via alpha- and beta-epoxides.

Abstract

Bovine liver microsomes converted four A'-steroids, cholesterol, 5-cholestene, 20-methyl-5-pregnen-3p-01, and pregnenolone, to the corresponding 5,6a-epoxides, 5,6/3-epoxides, and 5a,6/3-glycols in the presence of an NADPH-generating system, ferrous ion, and ADP.All of the metabolites were separated and isolated by high performance liquid chromatography and identified by gas chromatography-mass spectroscopy.The microsomes catalyzed hydrolysis of the a-and &epoxides of all the examined A'-steroids to the corresponding 5a,6bglycols.The A'-steroid double bond oxidation reaction was mediated by microsomal lipid peroxidation but not by P-450.The reaction required ferrous ion and was inhibited with the metal-chelating agent, EDTA.The microsomal epoxidation took place a t higher rates from the &side of the A'-steroid molecules than from the a- side to yield the a-and p-epoxides in the ratio 1:4.The obligatory intermediacy of the a-and &epoxides in the microsomal conversion of the A'-steroids was confirmed with cholesterol as the model substrate by means of the isotope-trapping method.Hepatic microsomal A'-steroid epoxide hydrolase had unique properties.The well known potent inhibitor, 3,3,3-trichloro-1-propene oxide (TCPO), of hepatic microsomal epoxide hydrolase did not inhibit but remarkably stimulated the microsomal hydrolysis of cholesterol epoxides.Under the same conditions, TCPO completely inhibited microsomal hydrolysis of xenobiotic epoxides, styrene oxide and safrole oxide.The microsomal hydrolysis of cholesterol epoxides was potentially inhibited with iminocholestanols (5,6a-imin0-5a-cholestan-3fl-01 and 5,6~-irnino-5~-cholestan-3/?-01).The aziridines, however, did not inhibit the microsomal hydrolysis of the xenobiotic epoxides.These strongly suggest the nonidentity of xenobiotic epoxide hydrolase with A'-steroid epoxide hydrolase.Endogenous A'-steroids such as cholesterol (1, 2) and pregnenolone' (3) are oxidized to their a-, /?-epoxides, and 5a,6/?-*The costs of publication of this article were defrayed in part by the payment of page charges.This article must therefore be hereby marked "aduertisement" in accordance with 18 U.S.C.Section 1734 solely to indicate this fact.' The abbreviations used for trivial and systematic names of steroids are: pregnenolone, 3P-hydroxy-5-pregnen-20-one; pregnenolone a-epoxide, 5,6a-epoxy-3P-hydroxy-5a-pregnan-20-one; pregnenolone &epoxide, 5,6/3-epoxy-3P-hydroxy-5/3-pregnan-20-one; pregnenolone 5a,6,L?-glycol, 3,8,5,6P-trihydroxy-5a-pregnan-20-one; cholesterol aepoxide, 5,6a-epoxy-5a-cholestan-3~-01; cholesterol P-epoxide, 5,6,8epoxy-5~-cholestan-3~-01; cholestanetriol, 5a-cholestane-3/3,5,6/3triol; 5-cholestene a-epoxide, 5,6a-epoxy-5a-cholestane; 5-cholestene ,&epoxide, 5,6P-epoxy-