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  • Determination of free and deconjugated testosterone and epitestosterone in urine using SPME and LC-MS/MS.

Determination of free and deconjugated testosterone and epitestosterone in urine using SPME and LC-MS/MS.

Bioanalysis (2010-12-24)
Yanwei Zhan, Florin M Musteata, Fabien A Basset, Janusz Pawliszyn
ABSTRACT

A thin sheet of polydimethylsilosane membrane was used as an extraction phase for solid-phase microextraction. Compared with fiber or rod solid-phase microextraction geometries, the thin film exhibited much higher extraction capacity without sacrificing extraction time due to its higher area-to-volume ratio. The analytical method involved direct extraction of unconjugated testosterone (T) and epitestosterone (ET) followed by separation on a C18 column and detection by selected reaction monitoring in positive ionization mode. The limit of detection was 1 ng/l for both T and ET. After method validation, free (unconjugated) T and ET were extracted and quantified in real samples. Since T and ET are extensively metabolized, the proposed method was also applied to extract the steroids after enzymatic deconjugation of urinary-excreted steroid glucuronides. The proposed method allows quantification of both conjugated and unconjugated steroids, and revealed that there was a change in the ratio of T to ET after enzymatic deconjugation, indicating different rates of metabolism.

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Supelco
Epitestosterone solution, 1.0 mg/mL in acetonitrile, ampule of 1 mL, certified reference material, Cerilliant®
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