The prevalence of doping in sport continues to be a concern in society

Wastewater-based Tracing of Doping Use

The prevalence of doping in sport and in the community at large continues to be a concern in many societies. Numerous test protocols are in place to test individuals but the increased sensitivity of modern analytical techniques and the purity of the ultrapure water available to use in testing have opened the scope for screening communal wastewater for drugs and their metabolites. 

Causanilles et al (1) have successfully developed and tested a procedure for 15 substances from the groups of anabolic steroids (used to promote muscle growth), weight loss products (used to increase metabolism by burning fat) and masking agents in wastewater using solid phase extraction to increase sensitivity and reduce matrix effects followed by liquid-chromatography coupled to high resolution mass spectrometry.

50 ml of sample were spiked with an isotope-labeled internal standard, filtered, and acidified before concentration on a mix-mode cationic polymer-based cartridge. The cartridge was washed repeatedly before elution with acetonitrile. This was evaporated, redissolved in water and finally reconstituted in 90:10 water/methanol to give a volume of 0.5 ml. This was analysed by UPLC (Ultra Performance Liquid Chromatography) coupled to a time-of-flight high-resolution mass spectrometer with a C18 column and a water/methanol mobile phase with the methanol content ranging from 20 to 100%.

Limits of detection ranged from 0.2 ngL-1 for anastrozole to 20 ngL-1 for trenbolone, nandrolone and mibolerone. 

Water from an ELGA PURELAB Chorus 1 was used throughout this work. The absence of trace organic contaminants in the water is critical as water is used repeatedly in the sample preparation: in the hydrochloric acid used to acidify the sample, to condition the SPE cartridge and to rinse it twice, to enhance the removal of acetonitrile
and, finally, to reconstitute the sample ready for UPLC. Water is also used as a mobile phase in the chromatography.

Samples were taken from the inlets to wastewater treatment plants in locations where sporting events of various types were taking place, before and during the events. 
24-hour composite samples were usually obtained. It was found to be essential to freeze the samples prior to analysis. A wide variety of analytes of interest were detected
particularly weight loss products. In many cases, significantly higher levels were detected during and just before the sporting events. 

The study was not exhaustive and was more intended to demonstrate the feasibility of the approach.

1) A. Causanilles, V. Nordmann, D. Vughs, E. Emke, O. De Hon, F. Hernandez and P. De Voogt, „wastewater-based tracing of doping use by the general population and 
amateur athletes“, Analytical and Bioanalytical Chemistry, 410, 1793-1803, 2018

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