Optimizing the extraction and analysis of DHEA sulfate, corticosteroids and androgens in urine: application to a study of the influence of corticosteroid intake on urinary steroid profiles

Abstract

A method of detecting and quantifying dehydroepiandrosterone (DHEA) sulfate, corticosteroids, and androgens has been developed. All of the compounds were first extracted from urine using solid phase extraction (SPE), enzymatically hydrolyzed, and separated into three samples using a second SPE. A DHEA sulfate sample was acetylated and re-extracted using SPE for purification before analysis. Corticosteroid samples were oxidized and re-extracted using liquid/liquid extraction for analysis. Androgen samples were acetylated and re-extracted using SPE prior to analysis. The extraction and analysis methods were investigated and optimized. Analyses were performed with gas chromatography/mass spectrometry (GC/MS) and gas chromatography/flame ionization detection (GC/FID). The entire procedure was then applied to the study of urine profiles of healthy volunteers and patients treated with corticosteroids. The results showed that the quantities of androgens found in patient urines were lower than in those of healthy volunteers. In addition, other metabolites were detected in patient urines.

Analysis of conjugated steroid androgens: Deconjugation, derivatisation and associated issues

“…Hot acid is generally now considered unreliable, in particular for DHEA sulfate due to low recoveries caused by steroid decomposition [37,38]. However, a recent study utilised hot acid in a one step hydrolysis-derivatisation procedure for DHEA sulfate (recovery not reported) [39].…”

Development and application of stable carbon isotope analysis to the detection of cortisol administration in cattle

“…Pujos et al 8. proposed a co‐extraction of DHEA sulphate and glucocorticoids after β ‐glucuronidase hydrolysis, followed by separation on a C 18 column (500 mg).…”

“…The stationary phase OASIS HLB was adapted from the work of Pujos et al .,8 which described purification on HLB (30 mg) cartridges. At first, all the potential relevant endogenous reference compounds such as 17 α ‐hydroxyprogesterone and 11‐desoxycortisol and the metabolites of cortisol, e.g.…”

“…Derivatisation with bismethylenedioxy‐pentafluoropropionate was employed by Furuta et al .,4 who demonstrated the good GC behaviour of cortisone and cortisol derivatives. An alternative was proposed more recently by Pujos et al .,8 consisting of the oxidation of all hydroxylated functions with potassium bichromate in sulphuric acid. This approach presents a double advantage.…”

“…A significant difference in the isotopic composition between these so‐called endogenous reference compounds (ERCs) and metabolites highlights the illegal administration of natural hormones. Only few papers dealing with this approach in food‐producing animals are available, concerning testosterone administration5–7 and, more recently, estradiol administration 8. The literature is more extensive for androgens and estrogens in humans, including some papers dealing with cortisol control in athletes' urine 8–10…”