Abstract
The present work was undertaken to develop a method for the determination of testosterone (T) and epitestosterone (ET) in human urine using the technique of gas chromatography/mass spectrometry (GC/MS) with selected ion monitoring (SIM). Urinary steroids were extracted by means of SEP-PAK C18 cartridge, and were hydrolyzed with β-glucuronidase. The hydrolyzate was treated by running it through a SEP-PAK silica cartridge and a thin layer chromatography, to remove interfering materials. Furthermore, proper amounts of internal standard, 1-dehydrotestosterone (1-dehydro T) were added. Urinary steroids in extract were converted into 3-enol, 17-ditrifluoroacetate derivatives and were quantitated with a GC/SIM using each molecular ion (T, ET: m/z=480, 1-dehydro T: m/z=478). As a result, T and ET were completely separated by using a 3% SE-30 as an isolation column for GC and it became feasible to finish the analysis in less than 3min without disturbing peaks. The T values in urine measured by this method correlated well with those by radioimmunoassay (r=0.928). This method is logical and was found to be specifically suitable for the clinical laboratory test.
