Mechanism-based inactivation of the major beta-naphthoflavone-inducible isozyme of rat liver cytochrome P-450 by the chloramphenicol analog N-(2-p-nitrophenethyl)dichloroacetamide.
The effectiveness, selectivity, and mechanism of the inactivation of the major beta-naphthoflavone-inducible isozyme of rat liver cytochrome P-450 (BNF-B) by the chloramphenicol analog N-(2-p-nitrophenethyl)dichloroacetamide (pNO2DCA) have been investigated. Intraperitoneal administration of pNO2DCA to beta-naphthoflavone-treated rats at doses of 10 and 100 mg/kg resulted in 72 and 95% decreases, respectively, in the ethoxyresorufin deethylase activity of subsequently prepared liver microsomes. Similar decreases were observed in the warfarin R-6 and R-8 hydroxylase activities of the microsomes. At the lower dose of pNO2DCA, only those R-warfarin hydroxylase activities attributable to BNF-B were decreased, whereas, at the higher dose, inhibition of additional cytochromes P-450 was evident. In vitro, pNO2DCA was found to be a highly efficient inactivator of purified BNF-B in a reconstituted system. The maximal rate constant for inactivation and the apparent Km for the inhibitor were 0.52 min-1 and 2.7 microM, respectively. Inactivation of BNF-B by pNO2DCA appears to involve an impairment in electron transfer from NADPH-cytochrome P-450 reductase, as evidenced by a decrease in the NADPH- but not the iodosobenzene-supported metabolism of ethoxycoumarin by the modified enzyme. However, in the absence of substrate, there was no decrease in the NADPH oxidase activity or in the steady state level of ferrous carbonyl complex formed enzymatically. Likewise, the maximal level of isosafrole metabolite-P-450 complex formed by BNF-B was not decreased by modification with pNO2DCA, although the rate of complex formation was inhibited.(ABSTRACT TRUNCATED AT 250 WORDS)
D000081:Acetamides / Q000494:pharmacology*; D000818:Animals; D001571:Benzoflavones / Q000494:pharmacology*; D002701:Chloramphenicol / Q000031:analogs & derivatives* / Q000494:pharmacology*; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis; D004790:Enzyme Induction / Q000187:drug effects; D005419:Flavonoids / Q000494:pharmacology*; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology*; D006899:Mixed Function Oxygenases / Q000378:metabolism; D009251:NADPH-Ferrihemoprotein Reductase / Q000378:metabolism; D010084:Oxidation-Reduction; D051381:Rats; D011919:Rats, Inbred Strains; D019324:beta-Naphthoflavone
D000081:Acetamides; D001571:Benzoflavones; D065607:Cytochrome P-450 Enzyme Inhibitors; D005419:Flavonoids; D007527:Isoenzymes; C048628:1-(4-nitrophenyl)-2-dichloroacetamidoethane; D019324:beta-Naphthoflavone; D002701:Chloramphenicol; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D009251:NADPH-Ferrihemoprotein Reductase