The in vivo metabolism of Jungle Warfare in greyhounds

Delta 6-Methyltestosterone was reported as the main active ingredient of the purported dietary supplement Jungle Warfare. This compound is structurally similar to 17 alpha-methyltestosterone, containing an additional Delta 6 double bond, and is reported to possess notable androgenic activity, raising concerns over the potential for abuse of Jungle Warfare in sport. The in vivo metabolism of Delta 6-methyltestosterone in greyhounds was investigated. Urinary phase I (unconjugated) and phase II (glucuronide) metabolites were detected following oral administration using liquid chromatography-mass spectrometry. No phase II sulfate metabolites were detected. The major phase I metabolite was confirmed as 16 alpha,17 beta-dihydroxy-17 alpha-methylandrosta-4,6-dien-3-one by comparison with a synthetically-derived reference material. Minor amounts of the parent drug were also confirmed. Glucuronide conjugated metabolites were also observed, but were found to be resistant to hydrolysis using the Escherichia coli beta-glucuronidase enzyme. Qualitative excretion profiles, limits of detection, and extraction recoveries were determined for the parent drug and the major phase I metabolite. These results provide a method for the detection of Jungle Warfare abuse in greyhounds suitable for incorporation into routine screening methods conducted by anti-doping laboratories.

Automated summary

Delta 6-Methyltestosterone has been identified as the main active compound in Jungle Warfare, a dietary supplement. It has similar structure to 17 alpha-methyltestosterone but with an extra Delta 6 double bond, leading to concerns about its potential androgenic effects and abuse in sports. In vivo metabolism studies in greyhounds revealed the presence of urinary phase I and phase II metabolites, with the major phase I metabolite identified as 16 alpha,17 beta-dihydroxy-17 alpha-methylandrosta-4,6-dien-3-one. Glucuronide conjugated metabolites were also observed but resistant to hydrolysis. This study provides a method for detecting Jungle Warfare abuse in greyhounds, useful for anti-doping laboratories.