Doping control container for urine stabilization: a pilot study

Abstract

Urine collection containers used in the doping control collection procedure do not provide a protective environment for urine, against degradation by microorganisms and proteolytic enzymes. An in-house chemical stabilization mixture was developed to tackle urine degradation problems encountered in human sport samples, in cases of microbial contamination or proteolytic activity. The mixture consists of antimicrobial substances and protease inhibitors for the simultaneous inactivation of a wide range of proteolytic enzymes. It has already been tested in lab-scale, as part of World Anti-Doping Agency's (WADA) funded research project, in terms of efficiency against microbial and proteolytic activity. The present work, funded also by WADA, is a follow-up study on the improvement of chemical stabilization mixture composition, application mode and limitation of interferences, using pilot urine collection containers, spray-coated in their internal surface with the chemical stabilization mixture. Urine in plastic stabilized collection containers have been gone through various incubation cycles to test for stabilization efficiency and analytical matrix interferences by three WADA accredited Laboratories (Athens, Ghent, and Rome). The spray-coated chemical stabilization mixture was tested against microorganism elimination and steroid glucuronide degradation, as well as enzymatic breakdown of proteins, such as intact hCG, recombinant erythropoietin and small peptides (GHRPs, ipamorelin), induced by proteolytic enzymes. Potential analytical interferences, observed in the presence of spray-coated chemical stabilization mixture, were recorded using routine screening procedures. The results of the current study support the application of the spray-coated plastic urine container, in the doping control collection procedure. Copyright © 2016 John Wiley & Sons, Ltd.

Improved detection methods significantly increase the detection window for EPO microdoses

“…Once in the laboratory, the sample or at least the aliquot for EPO analysis should be kept refrigerated or frozen until EPO analysis is performed and protease inhibitors are added to the urine in the first step of EPO preparation. 27 Alternatively the use of a stabilizer (combination of antibiotics, antimycotic substances and protease inhibitors) in the urine collection container used for the doping control has also been suggested 28,29 with good results on EPO detection but an effect on urine specific gravity was observed and remains to be defined. It might impact the results from initial testing procedures for some other substances.…”

Horseradish‐peroxidase‐conjugated anti‐erythropoietin antibodies for direct recombinant human erythropoietin detection: Proof of concept

“…Samples on lanes 8, 9 and 11 did not show any EPO bands, which could be related to an extremely low/no EPO excretion or more likely to a bacterial or proteolytic degradation of the protein during the shipment to the laboratory. 16 Although the Santa Cruz antibody was able to bind to the target analytes, it was less sensitive. Even when the exposure time was increased by more than threefold (8 to 25 s), some bands that had weak intensities on the routine gel (lanes 6 and 7) were still almost invisible using the modified method.…”

Les défis du passeport biologique: du prélèvement à l’interprétation

“…Il existe des études qui démontrent que les échantillons urinaires peuvent être stabilisés par l'ajout de bactériostatique [15]. Cependant, les substances utilisées ont un impact sur certaines variables telles que la gravité spécifique, le pH… Une autre stratégie suggérée est la congélation ou la réfrigération des urines immédiatement après la récolte afin de stopper une quelconque dégradation.…”