Testosterone administration to mares: criteria for detection of testosterone abuse by analysis of metabolites in plasma and urine.

The researchers administered a dose of testosterone to two mares and then studied the way certain metabolites behaved in their blood and urine. They determined that testosterone glucuronide in plasma and a concentration ratio of testosterone sulfate and epitestosterone sulfate in urine can be used as indicators of testosterone abuse, with the ability to detect misuse up to a month after administration.

Research Objective

• The objective of the study was to establish a reliable method for detecting the misuse of testosterone in mares by tracking key metabolites in the blood and urine following the administration of the testosterone hexahydrobenzoate.

Methodology

• Testosterone hexahydrobenzoate was intramuscularly administered to two mares.
• The researchers then studied the time course of various metabolites in the blood and urine using a method based on gas chromatography-mass spectrometry coupled with stable isotope dilution.

Key Findings

• Testosterone glucuronide, found in plasma, was identified as the most suitable marker for monitoring exogenous testosterone up to two weeks after administration when a threshold value of 200 ng/L was applied.
• In urine, the simultaneous measurement of the concentrations of testosterone sulfate (TS) and epitestosterone sulfate (ES) created a concentration ratio (TS/ES) which proved to be independent of urine flow. This ratio served as a potential indicator of testosterone misuse 20 to 30 days after dosage, particularly when a threshold value of 8 was used.
• Moreover, when the TS/ES ratio was close to the threshold, researchers suggested considering the concentrations of 5 alpha-androstane-3 beta, 17 alpha-diol (glucuronide) and its 17 beta-isomer (sulfate) as additional indicators of testosterone abuse. Those concentrations should not exceed 50 micrograms/L.

Implications

• This research offers comprehensive insight into the metabolic response to synthetic testosterone administration in horses, thereby providing a method for detecting testosterone abuse.
• The findings are fundamental in ensuring horse racing’s integrity, as they allow for the detection of synthetic hormones up to an entire month after administration.