Dealkylation of pentoxyresorufin: a rapid and sensitive assay for measuring induction of cytochrome(s) P-450 by phenobarbital and other xenobiotics in the rat.
The O-dealkylation of pentoxyresorufin (7-pentoxyphenoxazone) by rat liver microsomes was examined. The reaction appeared highly specific for certain phenobarbital inducible forms of cytochrome P-450 and was increased 95- to 140-fold by animal pretreatment with phenobarbital (75 mg/kg/day, four ip injections) and approximately 50-fold by Aroclor 1254 (500 mg/kg, one ip injection) while animal pretreatment with 3-methylcholanthrene (50 mg/kg/day, three ip injections) resulted in less than a 2-fold increase over the rate detected in control microsomes. It was observed that this activity, in microsomes for Aroclor-pretreated rats, was dependent on O2 and was inhibited by metyrapone and SKF 525-A, indicative of cytochrome(s) P-450 mediation in the reaction. When antibodies directed against purified cytochrome(s) P-450s were employed to inhibit the pentoxyresorufin O-dealkylation reaction, antibodies to P-450PB-B greatly inhibited the reaction (greater than 90%), while antibodies to P-450PB-C or P-450PB/PCN-E had minimal effects. Assay of hepatic microsomes from rats which were pretreated with varying doses of phenobarbital (0.9-75 mg/kg/day, four ip injections) indicated that while aminopyrine-N-demethylase activity was induced only 2-fold at the maximum dose (75 mg/kg/day), pentoxyresorufin O-dealkylase activity was induced approximately 140-fold at this dose and approximately 4-fold by a dose of phenobarbital as low as 0.9 mg/kg.
D000633:Aminopyrine N-Demethylase / Q000096:biosynthesis; D000818:Animals; D001140:Aroclors / Q000494:pharmacology; D020111:Chlorodiphenyl (54% Chlorine); D019363:Cytochrome P-450 CYP1A1; D019362:Cytochrome P-450 CYP2B1; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis*; D003640:Dealkylation; D004790:Enzyme Induction / Q000187:drug effects; D008297:Male; D008748:Methylcholanthrene / Q000494:pharmacology; D008862:Microsomes, Liver / Q000201:enzymology*; D010078:Oxazines / Q000378:metabolism*; D010088:Oxidoreductases / Q000096:biosynthesis; D010089:Oxidoreductases, N-Demethylating / Q000096:biosynthesis; D010634:Phenobarbital / Q000494:pharmacology*; D051381:Rats; D013379:Substrate Specificity
D001140:Aroclors; D010078:Oxazines; D020111:Chlorodiphenyl (54% Chlorine); D008748:Methylcholanthrene; C044943:pentoxyresorufin; D003577:Cytochrome P-450 Enzyme System; D010088:Oxidoreductases; D019363:Cytochrome P-450 CYP1A1; D019362:Cytochrome P-450 CYP2B1; D010089:Oxidoreductases, N-Demethylating; D000633:Aminopyrine N-Demethylase; C023680:dimethylaminoantipyrine N-demethylase; D010634:Phenobarbital