Determination of amitriptyline and nortriptyline in human liver microsomes with reversed-phase HPLC in vitro

AIM: To develop a method for simultaneous determinations of amitriptyline (Ami) and its metabolite nortriptyline (Nor) in human liver microsomes. METHODS: An incubation buffer containing microsomes, NADPH-generating system, and Ami, after termination of enzyme reaction and desipramine (Des) as internal standard (IS), was extracted with diethy ether and separated on a reversed-phase ODS column. Detection was achieved at 242 nm by ultraviolet detector. RESULTS: No potential interfering peaks were found. Ami and Nor gave rapid elution and baseline resolution. The linear curves of both analyses ranged 0.02 - 10 nmol and the limit of detection was 0.01 nmol. The recovery (94 % - 101 %) had good precision with relative s of < 8.3 %. CONCLUSION: This:method is rapid, sensitive, and simple for studying the metabolism of Ami and Nor.