Primary and secondary oxidative metabolism of dextromethorphan. In vitro studies with female Sprague-Dawley and Dark Agouti rat liver microsomes.
The O-demethylation of dextromethorphan (DM) to dextrorphan (DR) is catalysed by the polymorphic CYP2D6 (cytochrome P4502D6) isozyme in man. DM is commonly used as a probe for phenotyping subjects as either poor or extensive metabolizers for the debrisoquine/sparteine oxidative polymorphism via CYP2D6. The enzyme kinetics of DM O- and N-demethylation, and the N- and O-demethylations of the primary metabolites DR and 3-methoxymorphinan (3MM), respectively, were studied in liver microsomes from female Dark Agouti (DA) rats, the poor metabolizer counterpart, and female Sprague-Dawley (SD) rats, the extensive metabolizer counterpart. The formation of metabolites was quantified by HPLC with fluorescence detection and kinetic parameters were calculated. The intrinsic clearance (Vmax/Km) of the O-demethylation of 3MM to 3-hydroxymorphinan (3OHM) was 180-fold lower in DA rats (0.11 vs 20.77 mL/hr/mg) due to a 60-fold higher Km (108.7 vs 1.76 microM) and 3-fold lower Vmax (11.5 vs 35.95 nmol/mg/hr). The kinetics for DR N-demethylation to 3OHM did not differ between rat strains. The Michaelis-Menten constant (Km) for DM N-demethylation to 3MM was similar between SD and DA rats (85.04 vs 68.99 microM); however, SD rats displayed a 2-fold higher Vmax (83.37 vs 35.49 nmol/mg/hr) and intrinsic clearance (0.96 vs 0.51 mL/hr/mg). The O-demethylation of DM to DR in SD rats showed a high and low affinity enzyme component, with the high affinity intrinsic clearance contributing 98% of the total intrinsic clearance in these rats. DM O-demethylation in DA rats was characterized by a single enzyme system. The high affinity O-demethylating enzyme in SD rats showed a 20-fold lower Km (2.5 vs 55.6 microM) and a three-fold higher Vmax (51.04 vs 16.84 nmol/mg/hr) resulting in a 66-fold higher intrinsic clearance (20.04 vs 0.31 mL/hr/mg) compared to DA rats. Quinine, dextropropoxyphene, (+/-)methadone and (+/-)propafenone were shown to be potent inhibitors of 3MM and DM O-demethylation but did not inhibit DR or DM N-demethylation at similar concentrations. SD and DA rats showed a clear strain difference in 3MM O-demethylation and DM O-demethylation. In contrast, DR N-demethylation and DM N-demethylation do not appear to be under genetic control in the female SD-DA rat model. Kinetic parameters and inhibition studies suggest that 3MM and DM O-demethylation pathways in the rat may be mediated by the same cytochrome P450 isozyme.
Kerry
N L
NL
Department of Clinical and Experimental Pharmacology, University of Adelaide, Australia.
Somogyi
A A
AA
Mikus
G
G
Bochner
F
F
D000818:Animals; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D003915:Dextromethorphan / Q000031:analogs & derivatives / Q000378:metabolism*; D005260:Female; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism; D007700:Kinetics; D008745:Methylation; D008862:Microsomes, Liver / Q000378:metabolism*; D010084:Oxidation-Reduction; D010641:Phenotype; D051381:Rats; D017207:Rats, Sprague-Dawley
D065607:Cytochrome P-450 Enzyme Inhibitors; D007527:Isoenzymes; C028657:3-methoxymorphinan; D003915:Dextromethorphan; D003577:Cytochrome P-450 Enzyme System