The 23rd International Conference of Racing Analysts and Veterinarians

The 23rd International Conference of Racing Analysts and Veterinarians (ICRAV) was held in Hong Kong from September 17–23, 2023, under the theme of “Sustaining the Integrity of Racing.” ICRAV is a biennial event that brings together analysts, veterinarians, and administrators in the racing industry to share expertise and address crucial veterinary, scientific, regulatory, and welfare issues essential for upholding the integrity of racing and ensuring the welfare of animal athletes. The conference attracted over 200 participants from 30 nations, featuring a record-breaking total of more than 150 presentations.

For the first time, ICRAV has curated a selection of articles in a special issue of Drug Testing and Analysis, highlighting the key research topics presented at the 23rd conference. This issue also honors the memory of esteemed experts who passed away between 2018 and 2024, including Dr. David Lloyd Crone [1], Dr. Walter Hyde [2], Dr. Peter Haywood [3], Dr. John Vine [4], Dr. Dennis Hill [5], and Dr. Alan Malcolm Duffield [6]. Their significant contributions to equine anti-doping have laid a strong foundation for future developments. I extend my heartfelt gratitude to Editor-in-Chief Professor Mario Thevis for the opportunity to guest edit this special issue dedicated to the conference.

This issue opens with a comprehensive review by Gray et al. of the analytical advances in horseracing medication and doping control since the 22nd ICRAV in 2018 [7]. These encompass advancements in the detection of both the “small” and “large” molecule drugs, sample preparation methodologies, utilization of alternative matrices, advances in instrumentation, studies on drug metabolism and pharmacokinetics, the detection and prevalence of endogenous compounds, as well as the exploration of biomarkers and OMICs approaches. Research on equine gene doping control is also featured. Toutain contributed a mini-review on population pharmacokinetics (POP PK), a valuable tool for measuring and explaining the variability in drug exposure among individuals, with thorough discussions on its applications within the horseracing and equestrian industry [8].

Following this, this special issue presents an impressive collection of nine original research articles, five short communications, one application note, two case reports, two tutorials, and two perspectives, showcasing recent technological advancements, discoveries, and emerging challenges within the equine anti-doping industry. Recent progress includes advancements in detection capabilities for small molecules. Dorakumbura et al. reported a simple LC–MS method for screening over 150 compounds, including pregabalin and metformin, in equine and canine urine [9], monitoring the prevalence of these substances in race-day urine samples in Western Australia. Steel et al. contributed a tutorial on enhancing the specificity of sandwich enzyme-linked immunosorbent assay (ELISA)-based drug screening [10]. With respect to bisphosphonates (BPs), which will face a total ban in racehorses from January 1, 2027, as mandated by the International Federation of Horseracing Authorities (IFHA), several developments have emerged in their control. Klingberg et al. evaluated the performance of two types of SPE cartridges for recovering BPs from equine plasma [11]. Their findings included visual comparisons of signal-to-noise ratios in extracted chromatograms and recovery measurements to identify the best routine screening approach. Furthermore, Wong et al. reported an efficient ion chromatography–high resolution mass spectrometry (IC–HRMS) method for the simultaneous detection of myo-inositol trispyrophosphate (ITPP) and 10 BPs at sub-parts-per-billion (ppb) to low-ppb levels in equine plasma after SPE purification [12]. Another challenge in detecting BPs in horses is their erratic biological eliminations. Tou et al. developed an alternative approach to detect the misuse of BPs in horses by monitoring identified lipid and corticosteroid potential biomarkers [13].

Regarding macromolecules, Richards et al. reported an improved ELISA screening workflow alongside a robust capillary flow liquid chromatography–mass spectrometry (LC–MS) confirmatory method for detecting recombinant human erythropoietin (rHuEPO) in equine plasma [14]. These optimized methods enhanced the sensitivity and specificity of rhEPO detection and were employed to study the impact of repeated dosing of epoetin-β on its detection window. Steel et al. developed a high-throughput LC–MS assay incorporating size exclusion solid-phase extraction (SPE), peptide derivatization, and tryptic digestion for detecting adrenocorticotropic hormone (ACTH) and insulin variants [15], demonstrating applicability in detecting endogenous ACTH and insulin in plasma from horses suffering from pituitary pars intermedia dysfunction (PPID). Delcourt et al. evaluated three data-independent acquisition (DIA) strategies, namely, diaPASEF, slicePASEF, and prmPASEF, on a trapped ion mobility spectrometry quadrupole-time-of-flight mass spectrometer for high-throughput equine doping control analyses [16]. Although slicePASEF and prmPASEF outperformed diaPASEF in detecting humanized monoclonal antibodies in post-administration equine plasma, diaPASEF still demonstrated superior performance in untargeted proteomics studies.

Research focusing on endogenous substances includes Ho et al.'s study confirming the endogenous nature of estra-4,9-diene-3,17-dione (dienedione) in entire male horses as well as its metabolism and elimination in castrated horses [17]. Dienedione and its hydroxylated metabolites were proposed as markers for controlling the misuse in geldings. Cawley et al. developed a reliable analytical method capable of detecting recent ethanol exposure in greyhounds [18]. Urinary metabolites of ethanol, ethyl glucuronide, and ethyl sulfate were quantified in greyhound urine using a simple isotope dilution liquid–liquid extraction followed by LC–MS detection. A urinary threshold was proposed based on a population study and has been successfully applied to prosecute ethanol misuse in greyhounds. Barnabé explored Bayesian Individual Limits, customized for each horse, to complement a single limit for monitoring the endogenous IGF-1 profile of horses [19]. The method has been implemented in the French equine biological passport for IGF-1 monitoring.

In the area of equine pharmacokinetics, Knych contributed a tutorial describing the critical considerations for administration studies in equine anti-doping research and the design of scientific investigations aimed at effectively guiding medication control in racehorses [20]. Loy et al. examined the pharmacokinetics of two formulations of altrenogest administered orally and intramuscularly to mares [21]. The study provided valuable insights into female horses, assisting veterinarians and analysts in determining more accurate dosing and detection intervals.

Recent discoveries are also noteworthy for case investigations. Brooks et al. further explored the environmental source of aminorex [22]. Their findings indicate that resedine serves as an intermediary compound between barbarin and aminorex in horses and can act as a biomarker to differentiate aminorex positives linked to the ingestion of plants from the Brassicaceae family. Brooks et al. also conducted oral administration studies of hemp oil products in horses to investigate the metabolism and elimination of cannabidiolic acid (CBDA), cannabidiol (CBD), and its metabolites in horse plasma and urine [23]. Trevisiol et al. conducted an in vivo comparative study of hemp straw exposure versus CBD oil administration in horse urine [24], aiming to provide recommendations regarding the use of hemp straw before horse competitions and to distinguish between exposure to hemp bedding and the administration of CBD oil. Karamatic et al. discussed a thoughtful approach for establishing harmonized screening limits and detection times in greyhound racing [25].

In addition to the biological samples typically used in doping control testing, Yamada et al. presented a case report detailing the workflow of investigative testing of miscellaneous materials in Racing Analytical Services Limited (RASL) [26], illustrated with flowcharts to assist in handling of non-routine samples. Steel published an application note describing a simple method for identifying hyaluronic acid in seized samples through chemical hydrolysis, followed by Hypercarb chromatography and mass spectrometry analysis [27]. Chambers reported a pooled sampling technique to monitor medication in the racehorse industry, offering timely insights into medications being administered to horses and substances present at racetracks and training centers [28].

Maintaining the integrity of racing remains a significant challenge for many regulatory authorities in light of the rapidly evolving doping landscape. Kwok et al. detailed the development and implementation of an intelligence-based anti-doping approach aimed at enhancing enforcement capabilities through improved drug testing analytics and effective management of intelligence and integrity information [29].

Taken together, this special issue encapsulates the key advancements in the equine doping control industry since the 22nd ICRAV and highlights efforts made during the 23rd ICRAV for “Sustaining the Integrity of Racing.” Sincere thanks to all authors and reviewers who contributed to this special issue, which will serve as a valuable resource for equine anti-doping in the future.

The authors declare no conflicts of interest.