Measurements of hydrocortisone and cortisone for longitudinal profiling of equine plasma by liquid chromatography–tandem mass spectrometry

The conventional detection of exogenous drugs in equine doping samples has been used for confirmation and subsequent prosecution of participants responsible. In recent years, alternative methods using indirect detection have been investigated due to the expanding number of pharmaceutical agents available with the potential of misuse. The monitoring of endogenous biomarkers such as hydrocortisone (HC) has been studied in equine urine with an international threshold of 1 μg/ml established; however, there is no current threshold for equine plasma. The aim of this research was to investigate plasma concentrations of HC and cortisone (C) in race day samples compared to an administration of Triamcinolone Acetonide (TACA). The reference population (n = 1150) provided HC (6 to 145 ng/ml) and C (0.7 to 13 ng/ml) levels to derive the HC to C ratio (HC/C). Population reference limits (PRLs) were proposed for HC/C values at 0.2 (lower) and 61 (upper). Administration of TACA resulted in down‐regulation of HC/C values below the estimated PRLs for up to 96 h post‐administration. This indirect detection period was longer than the detection of TACA for 72 h. The use of individual reference limits (IRLs) for HC/C values was investigated to support the Equine Biological Passport (EBP), an intelligence model developed by Racing NSW for longitudinal monitoring of biomarkers.