Determination of a new active steroid by high performance liquid chromatography with laser-induced fluorescence detection following the pre-column derivatization

A sensitive analytical method for the determination of a new active steroid, butane acid-(5-androsten-17-one-3β-ol)-diester (A1998), was developed by high performance liquid chromatography with laser-induced fluorescence detection following the pre-column derivatization with dansylhydrazine. The calibration curve for A1998 derivatization was found linear in the dynamic range from 0.025 to 5.0 μg/ml, with the precision less than 6% (CV) and the mean extraction efficiency greater than 92%. In 200 μl of plasma samples the limit of quantitation was as low as 0.025 μg/ml with a signal-to-noise ratio of 10. This assaying was further applied to the determination of the pharmacokinetic parameters of A1998 in rats with an intravenous injection of A1998. Values for clearance for elimination, volume of distribution at steady state and terminal half life in the above case were determined as 50.3 ± 1.1 ml/min kg, 1329.0 ± 111.0 ml/kg and 44.0 ± 2.7 min, respectively.