Development and method validation for testosterone and epitestosterone in human urine samples by liquid chromatography applications.
An isocratic high-performance liquid chromatographic method for the determination of testosterone (T) and epitestosterone (ET) in human urine using liquid-liquid or solid-phase extraction (SPE) is developed and validated. The optimum separation is achieved using a Hypersil C(18) column, water-acetonitrile (57:43, v/v) as the mobile phase and UV-absorbance detection at 245 nm. The recoveries obtained for T and ET in liquid-liquid and SPE demonstrate that these procedures are interchangeable. Quantitation limits for T and ET are 8.6 and 5.4 ng/mL using solvent extraction and 7.3 and 5.7 ng/mL using SPE, respectively. The proposed method is used to evaluate the urinary T, ET, and the T/ET ratio for a healthy male population using liquid-liquid extraction, and the T and ET excretion profile for nine healthy men using SPE.
Gonzalo-Lumbreras
R
R
Department of Analytical Chemistry, Faculty of Chemistry, Universidad Complutense, 28040 Madrid, Spain.
Pimentel-Trapero
D
D
Izquierdo-Hornillos
R
R
D000328:Adult; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004845:Epitestosterone / Q000652:urine*; D006801:Humans; D008297:Male; D012016:Reference Values; D015203:Reproducibility of Results; D013739:Testosterone / Q000652:urine*
D013739:Testosterone; D004845:Epitestosterone