The use of bioassays to detect designer androgens in sports supplements
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Androgens are the most widely abused prohibited substances in sports. Detection of androgen abuse in sports relies on using sensitive gas chromatography tandem mass spectrometry-based techniques. These techniques require knowing the structure of the test compound in order to detect it. The last 15 years has seen the emergence of steroids with novel structures, termed designer steroids, which can bypass detection. In recent years, many of these designer steroids have appeared in sports supplements. There is limited data on the safety and efficacy of designer steroids. Numerous studies report that consumption of sports supplements containing designer androgens are associated with a number of adverse health effects, including cholestatic jaundice. Furthermore, it is often not known if these designer androgens have beneficial anabolic activity. The overall hypothesis of this thesis was that designer steroids contained within sports supplements are potent androgens. The main aim of this thesis was to assess the androgenic and anabolic activity of sports supplement-derived designer steroids using reporter gene androgen bioassays and a C2C12 myoblast cell model. Additionally, the Australian sports supplement market was screened for undeclared androgenic substances. Chapters 3 and 4 investigated the androgenic bioactivity of 22 designer steroids utilising in vitro androgen bioassays. Chapter 3 aimed to assess the intrinsic androgenic bioactivity of the designer steroids using the Saccharomyces cerevisiae-based yeast androgen bioassay. It was determined that 45% of the sports supplements had strong androgenic activity. Chapter 4 tested these designer steroids in the HuH7 cell line to mimic hepatic metabolism. This chapter showed that several of these strong androgens remained potent or were activated into more potent androgens after metabolism. Further, several intrinsically strong androgens were deactivated. Chapter 5 assessed the anabolic potential of several potent designer androgens in a C2C12 myoblast cell line. This study demonstrated that five androgens which had strong AR bioactivity, also demonstrated a high anabolic potential, with significant increases in myotube hypertrophy, nuclei accretion and MHC expression. Finally, Chapter 6 investigated the presence of undeclared androgenic substances in sports supplements available to the Australian market. Using the yeast and HuH7 androgen bioassays, it was shown that 5.3% (6/112) of the supplements had androgenic activity. In conclusion, this thesis demonstrates that sports supplements contain potent androgens, and should be of concern to the general Australian population and athletes, due to the potential health risks associated with androgen abuse, and the potential for testing positive in a doping test.
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