Rapid high-performance liquid chromatographic method for the separation of hydroxylated testosterone metabolites

摘要

A rapid high-performance liquid chromatography (HPLC) method is described for the quantitation of hydroxytestosterone metabolites.The method combines a Hypersil BDS C_(18) analytical column (10 cmX0.46 cm) and a linear mobile phase (1.25 ml/min) gradient of tetrahydrofuran-acetonitrile-water (10:10:80,v/v) changing to tetrahydrofuran-acetonitrile-water (14:14:72,v/v) over 10 min then remaining isocratic for 3 min.The total run time for the chromatographic separation of eight metabolites of testosterone is 15 min.Detection by UV is linear between 300 ng/ml and 10 #mu#g/ml with a limit of detection on column of 300 ng/ml.A method for the direct HPLC analysis of liver microsomal incubates of [~(14)]testosterone if also briefly described and when combined with the HPLC method,offers a distince advantage over previously reported methods for the rapid screening of testosterone hydroxylase activity in rat and human liver microsomes.