Detection of dihydrotestosterone gel, oral dehydroepiandrosterone, and testosterone gel misuse through the quantification of testosterone metabolites released after alkaline treatment.

The natural occurrence of endogenous anabolic steroids together with their availability in different administration forms makes the detection of their misuse a great challenge for doping control laboratories. Nowadays, the detection of endogenous steroids abuse is performed by the analysis of the steroid profile. Recently, androst-1,4-dien-3,17-dione (1,4-AD), androst-4,6-dien-3,17-dione (4,6-AD), 17β-hydroxy-androst-4,6-dien-3-one (6-T), and androst-15-en-3,17-dione (15-AD) have been described as testosterone (T) metabolites released after basic treatment of the urine. In the present work, the usefulness of these metabolites has been evaluated detecting the use of three different forms of endogenous steroids in a single dose: dihydrotestosterone gel (DHT), oral dehydroepiandrosterone (DHEA), and T gel. After the independent administration of these endogenous steroids, a rise in the value of several of the ratios calculated between the tested metabolites was noticed. For DHT, a small increase was observed for the ratios 1,4-AD/15-AD, 6-T/15-AD and 4,6-AD/15-AD although only for one volunteer. Better results were obtained for oral DHEA and T gel where an increase was observed in all volunteers for several of the tested ratios. The detection time in which the misuse can be detected (DT) has been evaluated using two different approaches: (1) comparison with population based reference limits, and (2) comparison with individual threshold levels. The obtained DTs were compared with the results of previously published markers for the misuse of such substances. When using basic released metabolites, shorter DTs were obtained for DHT, similar DTs for DHEA, and the detectability was substantially improved for T gel.