Use of dried blood spots in doping control analysis of anabolic steroid esters

• Whole blood dried on filter paper – a minimal invasive sampling technique.
• Screening of anabolic steroid esters in dried blood spots for doping control analysis.
• Direct detection of exogenous testosterone via the intact esters.
• Enhancement of the assay sensitivity by preparation of the methyloxime derivatives.
• Liquid chromatography coupled to high sensitivity mass spectrometry.

Dried blood spot (DBS) sampling, a technique for whole blood sampling on a piece of filter paper, has more than 50-years tradition, particularly in the diagnostic analysis of metabolic disorders in neonatal screening. Due to the minimal invasiveness, straightforwardness, robustness against manipulation and fastness DBS sampling recommends itself as an advantageous technique in doping control analysis. The present approach highlights the development of a screening assay for the analysis of eight anabolic steroid esters (nandrolone phenylpropionate, trenbolone enanthate, testosterone acetate, testosterone cypionate, testosterone isocaproate, testosterone phenylpropionate, testosterone decanoate and testosterone undecanoate) and nandrolone in DBS. The detection of the intact esters allows an unequivocal proof of the administration of conjugates of exogenous testosterone and its derivatives. Precise, specific and linear conditions were obtained by means of liquid chromatography high resolution/high accuracy mass spectrometry. Sensitivity in the low ppb range was accomplished by the preparation of the methyloxime derivatives of the target compounds. Labeled internal standards (d3-nandrolone, d3-nandrolone caproate and d3-nandrolone undecanoate) were applied to compensate for the broad range in chain length of the esters.The assay presented here outlines the application of DBS for the analysis of anabolic steroid esters in doping controls for the first time providing great potential to simplify the proof of exogenous administration of testosterone.

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