Simultaneous determination of a new hepatoprotective agent, YH439, and its metabolites, M4, M5, and M7 in plasma and urine by high-performance liquid chromatography.

A high-performance liquid chromatographic method was developed for the simultaneous determination of YH439, and its metabolites (M4, M5, and M7) in human plasma and rat urine using testosterone as an internal standard. The method involved deproteinization (plasma sample) or extraction (urine sample) followed by injection onto a C18 reversed-phase column. The mobile phase was acetonitrile-0.063 M phosphoric acidisopropyl alcohol (38:48:14, v/v/v), and the flow rate was 1.0 ml/min for the two methods. The column effluent was monitored by a UV detector set at 317 nm. The detection limits for YH439, M4, M5, and M7 in human plasma were 50, 40, 50, and 50 ng/ml, respectively, using the deproteinization method, and the corresponding values in rat urine were 50, 100, 50, and 50 ng/ml using the extraction method. No interferences from endogenous substances were found.