title: 17 beta-Estradiol metabolism by hamster hepatic microsomes. Implications for the catechol-O-methyl transferase-mediated detoxication of catechol estrogens.
abstract: We have shown that the metabolism of 17 beta-estradiol in hamster liver microsomes is concentration-dependent. At low (< 25 microM) concentrations of 17 beta-estriol, 16 alpha-hydroxylase activity predominated, and estriol was the major metabolite. At higher concentrations (25-75 microM), 16 alpha-hydroxylation and aromatic hydroxylation at C2 contributed equally to 17 beta-estradiol metabolism. Aromatic C4-hydroxylation was maximal at 75 microM of 17 beta-estradiol and was always less than C2-hydroxylation. Dehydrogenation of the 17 beta-hydroxyl group to the ketone (estrone) was also observed, but both estrone and 2-hydroxyestrone were minor (approximately 3%) metabolites of 17 beta-estradiol, only detectable at concentrations of 50 microM and above. Catechol-O-methyl transferase (COMT) effectively converted both 2- and 4-hydroxyl-17 beta-estradiol to their corresponding monomethoxy metabolites. Effective reducing conditions are required for COMT activity, because catechol estrogens are readily oxidized to their corresponding ortho-quinones, and ascorbic acid is routinely added to assays of COMT activity. Interestingly, although ascorbic acid (1 mM) increased the recovery of 2- and 4-hydroxy-17 beta-estradiol from microsomal incubations, it decreased the recovery of the methoxy metabolites (approximately 40%). Since the enediol function of ascorbate resembles that of a catechol group, ascorbate is a substrate for COMT and probably competes with the catechol estrogens for methylation. Because previous studies describing the ability of COMT to inhibit the covalent binding of electrophilic reactive metabolites of [4-(14)C]17 beta-estradiol to microsomal protein were performed in the presence of high (100 mM) Mg2+ concentrations, we also investigated the effects of Mg2+ on 17 beta-estradiol metabolism. Concentrations of Mg2+ > 10 mM inhibited the metabolism of 17 beta-estradiol, as evidenced by i) the increased recovery of substrate; ii) a decrease in the formation of estriol, estrone, and 2-, and 4-hydroxy-17 beta-estradiol; iii) a decrease in the recovery of water-soluble metabolites when incubations were performed in the presence of glutathione (GSH) to trap the reactive electrophilic metabolites; and iv) a decrease in the amount of reactive electrophilic metabolites bound to microsomal protein. GSH also decreased the covalent binding of electrophilic metabolites of [4-(14)C]17 beta-estradiol to microsomal protein, with the concomitant formation of water-soluble metabolites. Thus, both COMT and GSH combine to limit the formation of electrophilic metabolites from 17 beta-estradiol. The relative importance of each of these pathways to the disposition of the catechol estrogens remains to be determined.
authors.affiliation: Division of Pharmacology and Toxicology, College of Pharmacy, University of Texas at Austin 78712, USA.
authors.forename: M
authors.initials: M
authors.lastname: Butterworth
authors.forename: S S
authors.initials: SS
authors.lastname: Lau
authors.forename: T J
authors.initials: TJ
authors.lastname: Monks
mesh_terms: D000818:Animals; D001205:Ascorbic Acid / Q000494:pharmacology; D002394:Catechol O-Methyltransferase / Q000378:metabolism*; D006224:Cricetinae; D004958:Estradiol / Q000031:analogs & derivatives / Q000378:metabolism*; D002393:Estrogens, Catechol / Q000378:metabolism*; D005978:Glutathione / Q000494:pharmacology; D008658:Inactivation, Metabolic; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D008647:Mesocricetus; D008745:Methylation; D008862:Microsomes, Liver / Q000378:metabolism*; D012436:S-Adenosylmethionine / Q000378:metabolism; D039181:Steroid 16-alpha-Hydroxylase
chemical_list: D002393:Estrogens, Catechol; D004958:Estradiol; D012436:S-Adenosylmethionine; C001390:2-hydroxyestradiol; D039181:Steroid 16-alpha-Hydroxylase; D002394:Catechol O-Methyltransferase; D005978:Glutathione; D008274:Magnesium; D001205:Ascorbic Acid