Reduction of 7-alkoxyresorufins by NADPH-cytochrome P450 reductase and its differential effects on their O-dealkylation by rat liver microsomal cytochrome P450.
Antibody-inhibition experiments established that the induction of cytochrome P450c is largely responsible for the marked increase in liver microsomal 7-ethoxyresorufin O-dealkylation in rats treated with 3-methylcholanthrene, whereas the induction of cytochrome P450b and/or P450e is largely responsible for the marked increase in 7-pentoxy- and 7-benzyloxyresorufin O-dealkylation in rats treated with phenobarbital. When reconstituted with NADPH-cytochrome P450 reductase and lipid, purified cytochrome P450c catalyzed the O-dealkylation of 7-ethoxyresorufin at a rate of approximately 30 nmol/nmol P450/min, which far exceeded the rate catalyzed by either purified cytochromes P450b and P450e or microsomal cytochrome P450c. In contrast, purified cytochrome P450b and P450e were poor catalysts of the O-dealkylation of 7-pentoxy- and 7-benzyloxyresorufin. However, purified cytochrome P450b is an excellent catalyst of several other reactions, such as the N-demethylation of benzphetamine, the hydroxylation of testosterone, and the O-dealkylation of 7-ethoxycoumarin. The low rate of 7-pentoxyresorufin O-dealkylation catalyzed by purified cytochrome P450b did not reflect a requirement for cytochrome b5, and could not be ascribed to an artifact of the method used to measure the formation of resourufin. The catalytic activity of purified cytochrome P450b toward 7-pentoxyresorufin was consistently low over a range of substrate and lipid concentrations, and was not stimulated by sodium deoxycholate (which stimulates the N-demethylation of benzphatamine by purified cytochrome P450b). Evidence is presented which indicates that cytochrome P450c catalyzes the O-dealkylation of both the oxidized and reduced forms of 7-ethoxyresorufin, with perhaps a slight preference for the reduced form. In contrast, cytochrome P450b preferentially catalyzes the O-dealkylation of the oxidized form of 7-pentoxyresorufin. Conditions that favored formation of the reduced form of 7-ethoxyresorufin tended to stimulate its O-dealkylation by purified cytochrome P450c, whereas conditions that favored formation of the reduced form of 7-pentoxyresorufin decreased its rate of O-dealkylation by purified cytochrome P450b. Such conditions included a molar excess of NADPH-cytochrome P450 reductase over cytochrome P450, the presence of superoxide dismutase, and the presence of DT-diaphorase (liver cytosol).(ABSTRACT TRUNCATED AT 400 WORDS)
Dutton
D R
DR
Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas 66103.
Parkinson
A
A
D000818:Animals; D000937:Antigen-Antibody Reactions; D001555:Benzene Derivatives / Q000494:pharmacology; D002374:Catalase / Q000494:pharmacology; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D007700:Kinetics; D050356:Lipid Metabolism; D008862:Microsomes, Liver / Q000378:metabolism*; D016660:NAD(P)H Dehydrogenase (Quinone); D009251:NADPH-Ferrihemoprotein Reductase / Q000378:metabolism*; D010078:Oxazines / Q000378:metabolism*; D010084:Oxidation-Reduction; D011808:Quinone Reductases / Q000378:metabolism; D051381:Rats; D013329:Structure-Activity Relationship; D013482:Superoxide Dismutase / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism
D001555:Benzene Derivatives; D010078:Oxazines; D013481:Superoxides; C007649:ethoxyresorufin; D003577:Cytochrome P-450 Enzyme System; D002374:Catalase; D013482:Superoxide Dismutase; D009251:NADPH-Ferrihemoprotein Reductase; D016660:NAD(P)H Dehydrogenase (Quinone); D011808:Quinone Reductases; C007164:cumene hydroperoxide