Stereoselective reversible ketone formation from 10-hydroxylated nortriptyline metabolites in human liver.
1. E- and Z-10-hydroxynortriptyline are major metabolites of amitriptyline and nortriptyline in man. Upon incubation with human liver microsomes or cytosol, these metabolites were oxidized to the corresponding ketones, E- and Z-10-oxonortriptyline. (+)-E- and (+)-Z-10-hydroxynortriptyline were distinctly preferred over the (-)-isomers as substrates. NADP+ supported the oxidation in cytosol, whereas in microsomes NAD+ was the best cofactor. 2. Incubation of E- and Z-10-oxonortriptyline with NADPH and cytosol resulted in the nearly exclusive formation of (+)-E- and (+)-Z-10-hydroxynortriptyline. Kinetic analysis revealed high-affinity reduction (K(m) 1-2 microM) of the two ketones and an additional low-affinity component with the E-isomer. 10-Oxonortriptyline reduction was also catalysed by rabbit, but not by rat or guinea pig liver cytosol. 3. With [4-3H]NADPH as cosubstrate, tritium was incorporated into E- and Z-10-hydroxynortriptyline preferentially from the pro-4R position. Redox cycling of (+)-E- and (+)-Z-10-hydroxynortriptyline in cytosol in the presence of NAD- and NADPH was indicated by 3H incorporation from [pro-4R-3H]NADPH. 4. Recombinant human carbonyl reductase catalysed low-affinity reduction of E-10-oxonortriptyline with preferential transfer of the pro-4S-3H of labelled NADPH. 5. Ketone reduction in cytosol was strongly inhibited by 9,10-phenanthrenequinone and dehydrolithocholic acid and moderately by other 3-oxo steroids and some anti-inflammatory drugs. 6. The high-affinity reduction of E- and Z-10-oxonortriptyline and the oxidation of the alcohols in cytosol are probably mediated by a member of the aldo-keto reductase family of enzymes.
Breyer-Pfaff
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U
Institut für Toxikologie, Universität Tübingen, Germany.
Nill
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D000328:Adult; D000368:Aged; D001711:Biotransformation; D002648:Child; D003600:Cytosol / Q000378:metabolism; D005260:Female; D006801:Humans; D006900:Hydroxylation; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008297:Male; D008745:Methylation; D008862:Microsomes, Liver / Q000378:metabolism; D008875:Middle Aged; D009249:NADP / Q000378:metabolism; D009661:Nortriptyline / Q000031:analogs & derivatives* / Q000378:metabolism* / Q000493:pharmacokinetics; D010084:Oxidation-Reduction; D013237:Stereoisomerism
C099073:10-oxonortriptyline; C013567:10-hydroxynortriptyline; D009249:NADP; D009661:Nortriptyline