METABOLISM OF AMITRIPTYLINE WITH CYP2D6 EXPRESSED IN A HUMAN CELL-LINE

1. Expressed human cytochrome P450 enzyme CPY2D6 was used to metaboliz e amitriptyline (AMI). It was established that CYP2D6 not only catalyz ed 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 of the metabolites, 10-hydroxy-AMI, NT and 10-hpdr oxy-NT, was formed. 2. Biochemical parameters of NT formation from AMI were determined, yielding K-m = 47.48 +/- 1.32 mu M; V-max = 3.95 +/- 0.11 nmol/h/mg protein. The same parameters were calculated for the f ormation of 10-hydroxy-AMI (E + Z-isomers) from AMI, yielding K-m = 10 .70 +/- 0.20 mu M; V-max = 8.99 +/- 0.47 nmol/h/mg protein. 3. The for mation of 10-hydroxy-NT from AMI proceeded primarily via NT and to a m uch lesser extent via 10-hydroxy-AMI. 4. Quantitative analyses of AMI and its metabolites were difficult to reproduce when the metabolites w ere analysed underivatized. Two derivatization procedures, acetylation and trifluoroacetylation, were employed to improve assay reproducibil ity.