title: Effect of rociverine on P450-dependent monooxygenases and its N-deethylation metabolism in rat liver microsomes.
abstract: Rociverine [2-(diethylamino)-1-methylethyl cis-1-hydroxy [bicyclohexyl]-2-carboxylate] citrate (ROC) is an antispasmodic agent therapeutically active in humans at doses of 0.5-1 mg/kg. This study investigated the effect of acute administration of the drug on hepatic microsomal cytochrome P450 (P450)-catalysed drug metabolism. Only high doses (> or = 100 mg/kg) of ROC were able to induce in rats the hepatic microsomal pentoxyresorufin O-depenthylase (PROD) and 16 beta-testosterone hydroxylase activities both associated with P4502B1/2 and the erythromycin N-dimethylase (ErD) and 2 beta-testosterone hydroxylase activities both dependent on P4503A1/2. However, at 100 and 200 mg/kg of ROC, the 16 beta-testosterone hydroxylase and PROD were the most induced activities, suggesting that P4502B1/2 are the isoforms most sensitive to ROC induction. Accordingly, ROC treatment enhanced, in a dose-dependent manner, the amount of P4502B1/2 and 3A1/2 in microsomes as assayed by western blotting. The northern blot analysis of ROC-treated rat liver showed that the P4502B1/2 induction appears to be regulated at the mRNA level as in the induction by phenobarbital (PB). The oxidative metabolism of ROC with hepatic microsomes from control or PB- and ROC-induced rats resulted in a N-deethyl ROC derivative (major metabolite) and an unknown minor ROC derivative. The kinetic parameters for the N-deethylation of ROC were studied with purified P4502B1 and with microsomes from control or rats treated with various inducers (phenobarbital, ethanol, beta-naphthoflavone, dexamethasone and rociverine). It was found that phenobarbital-, dexamethasone- and rociverine-induced microsomes deethylated ROC with a Vmax about five times higher than that (0.9 nmol/min/mg protein) of control microsomes, although with a similar affinity (Km approximately 0.3 mM). In a reconstituted system, the purified P4502B1 metabolized ROC with a high deethylation rate (22 nmol/min/nmol P450). Moreover, the ROC deethylation was inhibited by compounds such as hexobarbital, metyrapone and triacetyloleandomicin, selective inhibitors for P4502B and/or P4503A enzymes. On the other hand ROC, when added in vitro, inhibited the 16 beta- and 2 beta-testosterone hydroxylases and the PROD and ErD activities. Taken together, these results indicate that the ROC-inducible P4502B and P4503A are involved in ROC deethylation. In conclusion, it has been demonstrated that ROC is a weak phenobarbital-like inducer of P450, probably able at high and reiterated doses to alter its own metabolism, at least in the rat liver.
authors: Laboratory of Genetics and Biochemical Toxicology, Istituto di Mutagenesi e Differenziamento, Consiglio Nazionale delle Ricerche, Pisa, Italy.
authors: S
authors: S
authors: Menicagli
authors: A
authors: A
authors: Lippi
authors: M
authors: M
authors: Criscuoli
authors: P G
authors: PG
authors: Gervasi
mesh_terms: D000079:Acetaldehyde / Q000378:metabolism; D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D015153:Blotting, Western; D001643:Bridged Bicyclo Compounds / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000494:pharmacology*; D003509:Cyclohexanecarboxylic Acids; D019362:Cytochrome P-450 CYP2B1; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004790:Enzyme Induction; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D009693:Nucleic Acid Hybridization; D010088:Oxidoreductases / Q000378:metabolism*; D010089:Oxidoreductases, N-Demethylating / Q000378:metabolism; D010276:Parasympatholytics / Q000494:pharmacology*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013250:Steroid Hydroxylases / Q000378:metabolism
chemical_list: D001643:Bridged Bicyclo Compounds; D003509:Cyclohexanecarboxylic Acids; D007527:Isoenzymes; D010276:Parasympatholytics; D012333:RNA, Messenger; D003577:Cytochrome P-450 Enzyme System; D010088:Oxidoreductases; D013250:Steroid Hydroxylases; D001189:Aryl Hydrocarbon Hydroxylases; D019362:Cytochrome P-450 CYP2B1; D051544:Cytochrome P-450 CYP3A; C022724:testosterone 7-alpha-hydroxylase, hamster; D010089:Oxidoreductases, N-Demethylating; D000079:Acetaldehyde; C021082:rociverine