Supramolecular Solvent Extraction for Doping Control Analysis of Prohibited Substances in Horse Urine

Despite the recent success in introducing supramolecular solvents (SUPRAS)-based extraction to drug analysis, its application and robustness in day-to-day regular urine testing have yet to be demonstrated. Moreover, the applicability of SUPRAS in equine doping control testing remains unexplored. In this work, we have successfully developed for the first time a simple, rapid, inexpensive, and environmentally friendly SUPRAS extraction method for analyzing 76 prohibited substances of different classes (selective androgen receptor modulators, hypoxia-inducible factor prolyl hydroxylase inhibitors, angiotensin II receptor antagonists, benzodiazepines, etc.) in hydrolyzed horse urine with liquid chromatography-mass spectrometry (LC/MS) for detection. The developed 1,2-hexanediol-based SUPRAS-LC/MS method has been fully validated, and its applicability and robustness in day-to-day testing of horse urine have also been demonstrated. This work marks a significant milestone in the advancement of green and sustainable drug testing methodology in equine sports, offering a novel approach to address one of the complexities inherent in equine doping control.