Pharmaceutical Sciences 1996, 2: 299-303 Received June 5, 1996 Accepted July 1, 1996 zyxwvutsrq 0 1996 Pharmaceutical Sciences zy Oxidative Metabolism of Dihydrocodeine in Dark-Agouti and Sprague-Dawley Rat Liver Microsomes LYNETTE C. KIRKWOOD, ROGER L. NATION, GEOFFREY D. REYNOLDS*, ANDREW A. SOMOGYIt AND LLOYD N. SANSOM zyxwvuts School of Pharmacy and Medical Sciences and *School zyxwv of Chemical Technology, University of South Australia, Adelaide 5000 and ?Department of Clinical and Experimental Pharmacology, University of Adelaide, Adelaide 5005, South Australia Abstract The oxidative metabolism of dihydrocodeine to nordihydrocodeine and dihydromorphine was studied in liver microsomes of female Dark-Agouti (cytochrome P450 2D1 (CYP2Dl) deficient) and Sprague-Dawley rats. Evaluation of microsomal metabolism in these two rat strains is a useful in-vitro model to test possible substrates of polymorphic human cytochrome P450 2D6 (CYP2D6). Nordihydrocodeine formation rates were similar in both strains. Analysis of the MichaelisMenten kinetics of dihydromorphine formation showed a significant difference (P zyxwvu < 0.05) between strainsl with respect to K, (943 zyxwvutsr pM for Dark-Agouti; 123 pM for Sprague-Dawley), V , (0.925; 2.37 pmol min- g-') and inbinsic clearance (0.986; 19.5 mL min- ' g- I). In Spragut-Dawley liver microsomes, dihydromorphine formation was suppressed by the CYP2Dl inhibitors, quinine and quinidine, at concentrations which had no effect on nordihydrocodeine formation. These in-vitro findings indicate that in rat liver microsomes the cytochrome P450 system is involved in dihydrocodeine metabolism to dihydromorphine and nordihydrocodeine and that CYP2Dl is involved in the 0-demethylation to dihydromorphine but not the N-demethylation to nordi- hydrocodeine. The results of this study are in agreement with recent in-vivo studies of dihydrocodeine metabolism in man which indicate CYP2D6 is the predominant enzyme catalysing dihydromorphine formation. Dihydrocodeine was first described in 1911 and remains in wide clinical use zyxwvutsrq as a moderately potent opioid analgesic and antitussive drug. It is structurally similar to codeine, varying only by the reduction of the C7-C8 double bond of codeine to a single bond (Fig. 1). Because of the structural similarity between dihydrocodeine and codeine it has been proposed that their pathways of metabolism may be similar (Rowel1 et a1 1983) and this has been recently substantiatedby Hufschmid et a1 (1995) who measured urinary excretion of dihydrocodeine and its metabolites after an oral dose of the drug. The latter authors reported that the drug was excreted unchanged, as dihydrocodeine conjugates and as the free and conjugated N- and O-demethylated metabolites, nordihydrocodeine, dihy- dromorphine and nordihydromorphine. The involvement of cytochrome P450 2D6 (CYP2D6) in the metabolism of dihy- drocodeine was also investigated by determining the metabolic profile of dihydrocodeine after administration of the CYP2D6 inhibitor quinidine. Almost no morphinoid metabolites of dihydrocodeine could be detected after administration of qui- nidine 2 h before dihydrocodeine intake. FIG. 1. Structure of dihydrocodeine Correspondence: L. C. Kirkwood, School of Pharmacy and Medical Sciences, University of South Australia, North Terrace, Adelaide 5000, South Australia. Five to 10 per cent of the Caucasian population lack CYP2D6 and are classified as poor metabolizers; the remainder are referred to as extensive metabolizers (Eichelbaum & Gross 1990). F r o m et a1 (1995) measured dihydromorphine for- mation after an oral dose of dihydrocodeine to six extensive and six poor metabolizers phenotyped for the CYP2D6 poly- morphism with sparteine. These authors found that dihy- dromorphine formation was one-seventh in poor metabolizers and concluded that dihydrocodeine 0-demethylation to dihy- dromorphine was catalysed mainly by CYP2D6. We have investigated the metabolism of dihydrocodeine in an in-vitro animal model for the CYP2D6 polymorphism in man. In rats, cytochrome P450 2D1 (CYP2Dl) serves similar metabolic functions to CYF'2D6 in man (Soucek & Gut 1992). The female Dark-Agouti rat lacks functional CYP2Dl (Gon- zalez et a1 1987), whereas in the Sprague-Dawley rat, and other rat strains, this isoform is mainly responsible for the metabolism of substrates affected by the debrisoquine/ sparteine polymorphism in man (Al-Dabbagh et a1 198 1 ; Larry et a1 1984; Kahn et a1 1985). Therefore, the female Dark- Agouti rat can be used as an animal model to predict which substrates might show impaired oxidation in human poor metabolizers. Indeed, liver microsomes from these rat strains have been employed to investigate codeine metabolism zy in- vitro and the results support the suitability of the model for the human poor and extensive metabolizer phenotypes for this substrate (Mikus et a1 1991). In this study, we report on the use of Dark-Agouti and Sprague-Dawley rat liver microsomes for enzyme kinetic and chemical inhibition studies to investigate the oxidative meta- bolism of dihydrocodeine and the involvement of CYP2Dl in