Determination of total and free amitriptyline and nortriptyline in human plasma by RP-HPLC / 中国药理学通报

ABSTRACT

AIM To develop a sensitive, specific and reliable reversed phase high performance liquid chromatographic method(RP\|HPLC) to determine the total and unbound(free) concentrations in human plasma of amitriptyline and its major metabolite, nortriptyline. METHODS\ The assay involved a simple extraction procedure. The mobile phase consisted of acetonitrile and distilled water(30∶70, V/V ), containing triethylamine(0 5%) and orthophosphoric acid(0 3%), pH 3 1. Separation was achieved on the C18 ODS column and the effluent was measured for UV absorption at 240 nm. RESULTS The calibration curves were linear in the range of 4～400 ?g?L -1 for total concentration, and in the range of 4～64 ?g?L -1 for free concentration for both amitriptyline and nortriptyline. The lowest limits of detection were 4 ?g?L -1 for both compounds. The absolute recovery rates were 102 0%?3 77% for amitriptyline and 99 3%?7 13% for nortriptyline. The precision values(RSD) of intra day and inter day for both amitriptyline and for nortriptyline were determined to be