Doping of horses is a fraud that has to be combatted to protect the welfare of the animals, the integrity of competition, and the breeding industry. Doping control of blood and urine from competition horses by doping control laboratories plays a crucial role in this. An increased sensitivity enabled by sophisticated detection methods, e.g. LC-HRMS and LC-MS2 potentially makes it attractive to analyse alternative matrices such as hair.
The matrix hair can provide a longer detection time window for e.g. steroid esters than blood and urine and can also provide doping control laboratories and racing authorities with complementary information about the ‘drug history’ of a horse.
In this paper, a method for the combined screening of anabolic agents and sedatives in hair is presented. After cleaning and pulverisation of the hair, liquid-liquid extraction was used to extract steroid esters, selected androgen receptor modulators (SARMs), and stanozolol, which were subsequently analyzed by LC-HRMS and LC-MS2. The organic layer was also used for the detection of a selection of sedatives e.g. acepromazine, butorphanol, ketamine and xylazine.
After liquid-liquid extraction, the remaining aqueous phase was subjected to a mixed-mode cation exchange solid-phase extraction, facilitating the analysis of β2-agonists. Method sensitivities were estimated at 0.5 - 10 pg per mg hair, depending on the individual analyte.
The here described method has been successfully used to analyse hair samples from AORC PT programs and also for the detection of sedatives in authentic hair samples.
Additionally, the method was validated in accordance with ISO 17025 requirements.