Synthesis of 3α,6β-Dihydroxyandrostan-17-one 3-Glucuronides for the Detection of Testosterone Misuse

The convenient synthesis of two 3-glucuronide conjugates of 6β-hydroxyandrosterone (6OH-And-3G 2) and 6β-hydroxyetiocholanolone (6OH-Etio-3G 3) is reported. The conjugates are identified as long-term markers of endogenous androgenic anabolic steroid abuse in doping control studies. Their resistance to enzymatic hydrolysis by E. coli β-glucuronidase means they may be missed by conventional GC-MS analysis protocols. One promising strategy to quantify these markers is by direct methods, which requires sufficient quantities of high purity reference materials. Our convergent synthesis features concurrent stereoselective introduction of 5α/5β hydrogen and 6β-hydroxy groups on the steroid skeleton, and a mild Schmidt trichloroacetimidate glycosylation strategy.