Metabolism of amitriptyline with CYP2D6 expressed in a human cell line

Abstract

1. Expressed human cytochrome P450 enzyme CPY2D6 was used to metabolize amitriptyline (AMI). It was established that CYP2D6 not only catalyzed ring 10- hydroxylation of AMI, but also mediated its N -demethylation to nortriptyline (NT), as well as the formation of 10-hydroxy-NT from NT. When the metabolism of AMI by CYP2D6 was repeated in the presence of quinidine, none ofthe metabolites, 10-hydroxyAMI, NT and 10-hydroxy-NT, was formed. 2. Biochemical parameters ofNT formation from AMI were determined, yielding Km 47 48 1 32 mu M; Vmax 3 95 0 11 nmol h mg protein. The same parameters were calculated for the formation of 10-hydroxy-AMI (E Z-isomers) from AMI, yielding Km 10 70 0 20 mu M; Vmax 8 99 0 47 nmol h mg protein. 3. The formation of 10-hydroxy-NT from AMI proceeded primarily via NT and to a much lesser extent via 10-hydroxy-AMI. 4. Quantitative analyses of AMI and its metabolites were difficult to reproduce when the metabolites were analysed underivatized. Two derivatization procedures, acetylation and trifluoroacetylation, were employed to improve assay reproducibility.