Rapid microsample analysis of imipramine and desipramine by reversed-phase high-performance liquid chromatography with ultraviolet detection.

A rapid and highly sensitive HPLC assay method was developed to measure small amounts of imipramine and its major metabolite, desipramine. The assay involved simple extraction procedures using clomipramine as the internal standard. The mobile phase consisted of acetonitrile (60%) and 0.01 M triethylamine in distilled water (40%) with the pH adjusted to 3.0. Separations were achieved on a C18 column and the effluent measured for UV absorption at 260 nm. The chromatographic separation was excellent, with no interference from endogenous serum constituents. This assay was suitable for measuring drug concentrations in the range of 10-1000 ng/ml using a 0.1-ml serum sample. The method was applied to a drug disposition study in transgenic mice with increased plasma alpha 1-acid glycoprotein.