Over the years, illicit doping practices in sports have changed, from wide use of stimulants in the first half of the 20th century to the rise of anabolic steroids, which make up the majority of anti-doping rule violations nowadays. These changes in doping prevalences also forced sport federations, anti-doping agencies, and of course anti-doping laboratories to adapt their strategies to detect doping. Similarly, the analytical methods to detect doping changed, following technological advances. Not only do anabolic steroids make up an important part of all reported anti-doping rule violations, but their detection can be challenging, depending on the substance that was administered The work performed in this thesis was aimed at expanding the detection of anabolic steroids by using alternative sample matrices and applying analytical techniques that are currently not widely used in the anti-doping field.
While the detection of exogenous steroids in urine is relatively straightforward, doping with endogenous steroids like testosterone is much more complex. As endogenous steroids are produced by the body as well, their origin has to be determined which is the differentiation between endogenous production or exogenous administration. Currently the only technique capable of distinguishing between natural and synthetic molecules is isotope ratio mass spectrometry (IRMS). However, IRMS requires meticulous sample purification and large sample volumes, with urine as the routinely used matrix of choice. Therefore, it is not feasible to analyse every sample with this technique, and suspicious samples have to be preselected carefully. At the moment, the most important criterion for selection is the urinary steroid profile, which is based on the concentrations of six endogenous steroids and their ratios. In the past, population derived thresholds were used to evaluate steroid concentrations, but large inter-individual variations were observed, making the population derived thresholds not sensitive enough. To develop individual thresholds, the athlete biological passport was created, including the steroidal module which contains longitudinal and individual urinary steroid profiles of every monitored athlete. Using this tool, each athlete is monitored as their own reference, and the targeting of samples for IRMS analysis was improved, aiding in the detection of doping with endogenous steroids. However, this approach has some limitations, as the detection of transdermal applications remains difficult and the urinary steroid profiles require the excretion of testosterone, which can be reduced in certain athletes.
In the first part of the thesis, blood matrices were explored for the detection of anabolic steroids to supplement regularly used urine samples. In the last years, serum steroid profiling was introduced into the athlete biological passport, adding another selection criterion for IRMS analysis. The benefit of analysing serum samples is to allow for the detection of testosterone esters, which are used as pro-hormones.
