Doping in horses refers to the administration of prohibited substances or methods to enhance performance or alter behavior during competitions. This practice is a concern in equestrian sports due to its potential impact on animal welfare, fair play, and the integrity of the sport. Substances used in doping can range from stimulants and painkillers to sedatives and anti-inflammatory drugs. Detection methods include blood and urine tests, designed to identify the presence of banned substances or their metabolites. This page compiles peer-reviewed research studies and scholarly articles that explore the detection methods, regulatory frameworks, and ethical considerations associated with doping in equine sports.
Waraksa E, Wójtowicz-Zawadka M, Kwiatkowska D, Jarek A, Małkowska A, Wrzesień R, Namieśnik J.A novel assay for the simultaneous determination of ibuprofen (IBU) and its four probable metabolites, 1-hydroxyibuprofen (1-OH IBU), 2-hydroxyibuprofen (2-OH IBU), 3-hydroxyibuprofen (3-OH IBU) and carboxyibuprofen (CBX IBU) in equine urine samples with the application of Gas Chromatography-Electron Ionization-Mass Spectrometry (GC-EI-MS) has been developed and elaborated. The new approach for sample preparation including minimizing matrix effects by the application of weak cation exchange solid-phase extraction together with strong cation exchange solid-phase extraction has been applied. The...
Bailly-Chouriberry L, Garcia P, Cormant F, Loup B, Popot MA, Bonnaire Y.Cobra (Naja naja kaouthia) venom contains a toxin called α-cobratoxin (α-Cbtx) containing 71 amino acids (MW 7821 Da) with a reported analgesic power greater than morphine. In 2013, the first analytical method for the detection of α-Cbtx in equine plasma was developed by Bailly-Chouriberry et al, allowing the confirmation of the presence of α-Cbtx at low concentrations (1-5 ng/mL or 130-640 fmol/mL) in plasma samples. To increase the method sensitivity and therefore to improve the detection of α-Cbtx in post-administration plasma samples, a nano-liquid chromatography-mass spectrometry/hi...
Popot MA, Jacobs M, Garcia P, Loup B, Guyonnet J, Toutain PL, Bailly-Chouriberry L, Bonnaire Y.Tiludronate is a bisphosphonate drug marketed to treat different bone conditions in horses. Objective: The goal of this study was to measure the plasma concentrations of tiludronate in a population of race and sport horses under field conditions, and using pharmacokinetic population modelling, to estimate detection times for doping control. Methods: Prospective cohort. Methods: This study was conducted under field conditions on 39 race or sport horses diagnosed with bone conditions based on a lameness examination and treated with tiludronate. Each horse received 1 mg/kg of tiludronate (Tildren...
Hansson A, Knych H, Stanley S, Berndtson E, Jackson L, Bondesson U, Thevis M, Hedeland M.LGD-4033 has been found in human doping control samples and has the potential for illicit use in racehorses as well. It belongs to the pharmacological class of selective androgen receptor modulators (SARMs) and can stimulate muscle growth, much like anabolic steroids. However, SARMs have shown superior side effect profiles compared to anabolic steroids, which arguably makes them attractive for use by individuals seeking an unfair advantage over their competitors. The purpose of this study was to investigate the metabolites formed from LGD-4033 in the horse in order to find suitable analytical ...
Guan F, You Y, Li X, Robinson MA.α-Cobratoxin (CTX) is a large peptide (71 amino acids) with strong analgesic effect and may be misused in sports such as horse racing. To prevent such misuse, a sensitive method is required for detection and confirmation of the toxin in equine samples. CTX was extracted from equine plasma using an optimized mixed-mode solid-phase extraction (SPE) procedure. Extracted CTX was reduced with dithiothreitol and alkylated with iodoacetamide, and then was digested by trypsin at 56°C for 30min. The digest was analysed by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS), and trypt...
Hillyer LL, Ridd Z, Fenwick S, Hincks P, Paine SW.While cobalt is an essential micronutrient for vitamin B synthesis in the horse, at supraphysiological concentrations, it has been shown to enhance performance in human subjects and rats, and there is evidence that its administration in high doses to horses poses a welfare threat. Animal sport regulators currently control cobalt abuse via international race day thresholds, but this work was initiated to explore means of potentially adding to application of those thresholds since cobalt may be present in physiological concentrations. Objective: To devise a scientific basis for differentiation b...
Joré C, Loup B, Garcia P, Paris AC, Popot MA, Audran M, Bonnaire Y, Varlet-Marie E, Bailly-Chouriberry L.Erythropoiesis Stimulating Agents (ESAs) were developed for therapeutic purposes to stimulate red blood cell (RBC) production. Consequently, tissue oxygenation is enhanced as athlete's endurance and ESAs misuse now benefits doping. Our hypothesis is that most of ESAs should have similar mechanisms and thus have the same effects on metabolism. Studying the metabolome variations could allow suspecting the use of any ESAs with a single method by targeting their effects. In this objective, a metabolomic study was carried out on 3 thoroughbred horses with a single administration of 4.2μg/kg of Mir...
Knych HK, Stanley SD, McKemie DS, Arthur RM, Bondesson U, Hedeland M, Thevis M, Kass PH.Although developed as a therapeutic medication, meldonium has found widespread use in human sports and was recently added to the World Anti-Doping Agency's list of prohibited substances. Its reported abuse potential in human sports has led to concern by regulatory authorities about the possible misuse of meldonium in equine athletics. The potential abuse in equine athletes along with the limited data available regarding the pharmacokinetics and pharmacodynamics of meldonium in horses necessitates further study. Eight exercised adult thoroughbred horses received a single oral dose of 3.5, 7.1, ...
Guan F, Robinson MA.The ability to analyze biological samples for multitudinous exogenous peptides with a single analytical method is desired for doping control in horse racing. The key to achieving this goal is the capability of extracting all target peptides from the sample matrix. In the present study, theory of mixed-mode solid-phase extraction (SPE) of peptides from plasma is described, and a generic mixed-mode SPE procedure has been developed for recovering multitudinous exogenous peptides with remarkable sequence diversity, from equine plasma and urine in a single procedure. Both the theory and the develop...
Popot MA, Ho ENM, Stojiljkovic N, Bagilet F, Remy P, Maciejewski P, Loup B, Chan GHM, Hargrave S, Arthur RM, Russo C, White J, Hincks P, Pearce C....Cobalt is an essential mineral micronutrient and is regularly present in equine nutritional and feed supplements. Therefore, cobalt is naturally present at low concentrations in biological samples. The administration of cobalt chloride is considered to be blood doping and is thus prohibited. To control the misuse of cobalt, it was mandatory to establish an international threshold for cobalt in plasma and/or in urine. To achieve this goal, an international collaboration, consisting of an interlaboratory comparison between 5 laboratories for the urine study and 8 laboratories for the plasma stud...
Shin HD, Suh JH, Kim J, Cho HD, Lee SD, Han KS, Wang Y, Han SB.A high throughput method for simultaneous screening of anabolic steroids and their metabolites (4-esterendione, trenbolone, boldenone, oxandrolone, nandrolone, methandrostenolone, testosterone, 1-androstendione, ethisterone, normethandrolone, methyltestosterone, 16β-Hydroxystanozolol, epitestosterone, bolasterone, norethandrolone, danazol, stanozolol and androstadienone) in equine urine by online turbulent flow extraction coupled with liquid chromatography-tandem mass spectrometry was developed. The use of turbulent flow chromatography could simplify pretreatment of horse urine, which has com...
Bailly-Chouriberry L, Loup B, Popot MA, Dreau ML, Garcia P, Bruyas JF, Bonnaire Y.Since the availability on the European market of the vaccine Improvac® dedicated to male pig immunological castration, the risk of misuse of this product in horses is now considered as a threat for the horseracing industry. Immunological castration is not allowed by the racing codes (immune system, Article 6). Indeed, this vaccination against the hypothalamic hormone luteinizing hormone-releasing hormone or gonadotropin-releasing hormone (GnRH) will prevent the release from the anterior pituitary of luteinizing hormone and follicle stimulating hormone, which are required for the development a...
Bailly-Chouriberry L, Baudoin F, Cormant F, Glavieux Y, Loup B, Garcia P, Popot MA, Bonnaire Y.The improvement of doping control is an ongoing race. Techniques to fight doping are usually based on the direct detection of drugs or their metabolites by analytical methods such as chromatography hyphenated to mass spectrometry after ad hoc sample preparation. Nowadays, omic methods constitute an attractive development and advances have been achieved particularly by application of molecular biology tools for detection of anabolic androgenic steroids (AAS), erythropoiesis-stimulating agent (ESA), or to control human growth hormone misuses. These interesting results across different animal spe...
Wong JKY, Kwok WH, Chan GHM, Choi TLS, Ho ENM, Jaubert M, Bailly-Chouriberry L, Bonnaire Y, Cawley A, Ming Williams H, Keledjian J, Brooks L....Acadesine, 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside, commonly known as AICAR, is a naturally occurring adenosine monophosphate-activated protein kinase (AMPK) activator in many mammals, including humans and horses. AICAR has attracted considerable attention recently in the field of doping control because of a study showing the enhancement of endurance performance in unexercised or untrained mice, resulting in the term 'exercise pill'. Its use has been classified as gene doping by the World Anti-Doping Agency (WADA), and since it is endogenous, it may only be possible to control del...
Wilkin T, Baoutina A, Hamilton N.A horse's success on the racetrack is determined by genetics, training and nutrition, and their translation into physical traits such as speed, endurance and muscle strength. Advances in genetic technologies are slowly explaining the roles of specific genes in equine performance, and offering new insights into the development of novel therapies for diseases and musculoskeletal injuries that cause early retirement of many racehorses. Gene therapy approaches may also soon provide new means to artificially enhance the physical performance of racehorses. Gene doping, the misuse of gene therapies f...
Liu Y, Uboh CE, Li X, Guan F, You Y, Maylin GA, Zhu F, Soma LR.A rapid and sensitive method for simultaneous screening, quantification and confirmation of 17 barbiturates in horse plasma using liquid chromatography-tandem mass spectrometry is described. Analytes were recovered from plasma by liquid-liquid extraction using methyl tert-butyl ether, separated on a C18 column, and analyzed in negative electrospray ionization mode. Multiple-reaction monitoring was employed for screening and quantification. Confirmation for the presence of the analytes was achieved by comparing ion intensity ratio. The ranges for limits of detection, quantification and confirma...
Herholz C, Zink N, Laska H, Gumpendobler M, Trolliet C, Probst S.Horse feed material of Swiss and foreign production available on the Swiss market was tested with regard to possible contamination with 7 different alkaloids, atropine and colchicine. Twenty-eight feed samples as well as 2 poppy seed samples were analyzed. Out of 28 feed samples 18 were positive for prohibited substances in Equestrian sports. The concentration of prohibited substances was in none of the samples high enough to cause an effect on the body of the horse. The poppy seed sample, which was obtained by conventional harvesting and cleaning methods, contained a very high Morphine concen...
Fragkaki AG, Kioukia-Fougia N, Kiousi P, Kioussi M, Tsivou M.The artificial increase of the physical capability of horses using drugs is well known in racing and other equine sports. Both illicit and therapeutic substances are regarded as prohibited substances in competition in most countries. Some countries make distinctions for a few, specific drugs which are, however, allowed for use in other countries. The primary objective in the case of doping control is the detection of any trace of drug exposure, either parent drug or any of its metabolites, using the most powerful analytical methods which are generally based on chromatographic/mass spectrometri...
Kwok KY, Chan GHM, Kwok WH, Wong JKY, Wan TSM.Selective estrogen receptor modulators (SERMs) are chemicals that possess the anti-oestrogenic activities that are banned 'in' and 'out' of competition by the World Anti-Doping Agency (WADA) in human sports, and by the International Federation of Horseracing Authorities (IFHA) in horseracing. SERMs can be used as performance-enhancing drugs to boost the level of androgens or to compensate for the adverse effects as a result of extensive use of androgenic anabolic steroids (AASs). SERMs have indeed been abused in human sports; hence, a similar threat can be envisaged in horseracing. Numerous an...
Kwok WH, Choi TLS, Tsoi YYK, Leung GNW, Wan TSM.A fast method for the direct analysis of enzyme-hydrolysed horse urine using an automated on-line solid-phase extraction (SPE) coupled to a liquid-chromatography/high resolution mass spectrometer was developed. Over 100 drugs of diverse drug classes could be simultaneously detected in horse urine at sub to low parts per billion levels. Urine sample was first hydrolysed by β-glucuronidase to release conjugated drugs, followed by centrifugal filtration. The filtrate (1mL) was directly injected into an on-line SPE system consisting of a pre-column filter and a SPE cartridge column for the separa...
Choi TLS, Wong JKY, Ho ENM, Kwok WH, Leung GNW, Curl P, Wan TSM.Lithium salts are commonly prescribed to treat bipolar disorder in humans. They are effective for the treatment of acute mania and the prophylaxis of manic relapses through long-term use. Although there is no reported legitimate therapeutic use of lithium in horses, its potential mood-stabilizing effect, low cost, and ready availability make lithium salt a potential agent of abuse in equine sports, especially for equestrian competition horses. Lithium can be found in soil, plants, and water, as such it is naturally present in the equine body, thus a threshold is necessary to control its misuse...
Wong KS, Chan GHM, Ho ENM, Wan TSM.Growth hormone (GH), or somatotropin, is a protein that may enhance physical performance and facilitate growth and wound healing. For this reason, growth hormones and their recombinant analogues are prohibited in human sports by the World Anti-Doping Agency (WADA) and in horseracing under Article 6 of the International Agreement on Breeding, Racing and Wagering published by the International Federation of Horse Racing Authorities (IFHA). Identifying the illicit use of GHs in both human athletes and racehorses is challenging, especially when differentiating between endogenous and exogenous GHs,...
Waller CC, McLeod MD.In recent years, the potential for anabolic steroid abuse in equine sports has increased due to the growing availability of designer steroids. These compounds are readily accessible online in 'dietary' or 'nutritional' supplements and contain steroidal compounds which have never been tested or approved as veterinary agents. They typically have unusual structures or substitution and as a result may pass undetected through current anti-doping screening protocols, making them a significant concern for the integrity of the industry. Despite considerable focus in human sports, until recently there ...
Machnik M, Kaiser S, Koppe S, Kietzmann M, Schenk I, Düe M, Thevis M, Schänzer W, Toutain PL.Methylxanthines positives in competition samples have challenged doping control laboratories and racing jurisdictions since methylxanthines are naturally occurring prohibited substances and often constituents of feed. For theobromine, an international threshold (renamed in International Residue Limit, IRL) of 2 µg/mL in urine has been established. On the basis of the data presented herein, a threshold or rather an IRL for theobromine in plasma of 0.3 µg/mL was proposed and was thereupon approved by the International Federation of Horseracing Authorities (IFHA). Official recommendations f...
Salomonsson ML, Bondesson U, Hedeland M.This paper describes quantitative methods for the determination of dimethylsulfoxide (DMSO) in equine plasma and urine based on simple precipitation and dilution followed by hydrophilic interaction liquid chromatography coupled to tandem mass spectrometry (HILIC-MS/MS). DMSO is a polar solvent with analgesic and anti-inflammatory properties. Its pharmacological features make it prohibited in horse racing. However, since DMSO is naturally present in the horses' environment, international threshold values have been implemented for plasma and urine (1 and 15 µg/mL, respectively). Previously pr...
Wong ASY, Leung GNW, Leung DKK, Wan TSM.Anabolic steroids are banned substances in equine sports. Gas chromatography-mass spectrometry (GC-MS) has been the traditional technique for doping control analysis of anabolic steroids in biological samples. Although liquid chromatography-mass spectrometry (LC/MS) has become an important technique in doping control, the detection of saturated hydroxysteroids by LC-MS remains a problem due to their low ionization efficiency under electrospray. The recent development in fast-scanning gas-chromatography-triple-quadrupole mass spectrometry (GC-MS/MS) has provided a better alternative with a sign...
Guan F, Robinson MA, Soma LR.Etanercept is a protein-based medication for the treatment of human patients with rheumatoid arthritis and other autoimmune-based diseases; its pharmacological action is to inhibit and antagonize tumour necrosis factor alpha. Etanercept was rumoured to be used in horse racing in North America. To detect such use, the aim of this study was to develop a liquid chromatography-mass spectrometry (LC-MS) method for confirmation of etanercept in equine plasma. Etanercept was extracted from plasma by anti-human IgG antibody linked to magnetic beads. The analyte was reduced and alkylated, and then dige...
Bachmann V, von Salis B, Fürst A.The goal of this study was to describe the development of equine drug testing in horses in Switzerland. This was achieved through evaluation of a film made by the Institute of Forensic Medicine at the University of Basel entitled 'Doping von Rennpferden' [Doping of Race Horses], toxicological detection, 1962', the analysis of doping test results of the Swiss Equestrian Federation and by interviewing individuals of various professions who were involved in equine drug testing at the time. The study compares early and modern methods of drug testing and highlights the changes in the attitude of eq...
Knych HK, Arthur RM, Kanarr KL, McKemie DS, Kass PH.Methamphetamine is a central and peripheral nervous system stimulant. There is only a single study that describes exposure to and disposition of this compound in horses. The potential for abuse and inadvertent exposure in equine athletes along with the limited data available necessitates further study. The objectives of the current study were to describe drug and metabolite concentrations, develop an analytical method that could be used to regulate its use, and describe selected pharmacodynamic effects. In phase 1, six horses were randomized into three transmucosal dose groups (n = 2/group; ...
Moeller BC, Yang Z.Controlling the abuse of prohibited substances such as anabolic steroids, selective androgen receptor modulators, β-adrenoceptor agonists, and blood doping agents is of great interest to racing authorities. The use of dried blood spots (DBS) as an alternative sampling approach may be a feasible approach for controlling the use of these agents. To assess the feasibility of using DBS in equine blood, an 11-min liquid chromatography-mass spectrometry method was developed on a triple quadrupole mass spectrometer following extraction from Whatman 903 DBS cards. A total of 50 compounds across multi...
Choi TLS, Wong JKY, Ho ENM, Kwok WH, Leung GNW, Curl P, Wan TSM.Lithium salts are commonly prescribed to treat bipolar disorder in humans. They are effective for the treatment of acute mania and the prophylaxis of manic relapses through long-term use. Although there is no reported legitimate therapeutic use of lithium in horses, its potential mood-stabilizing effect, low cost, and ready availability make lithium salt a potential agent of abuse in equine sports, especially for equestrian competition horses. Lithium can be found in soil, plants, and water, as such it is naturally present in the equine body, thus a threshold is necessary to control its misuse...
Dirikolu L, Lehner AF, Karpiesiuk W, Harkins JD, Woods WE, Carter WG, Boyles J, Fisher M, Tobin T.Lidocaine is a local anesthetic drug that is widely used in equine medicine. It has the advantage of giving good local anesthesia and a longer duration of action than procaine. Although approved for use in horses in training by the American Association of Equine Practitioners (AAEP), lidocaine is also an Association of Racing Commissioners International (ARCI) Class 2 drug and its detection in forensic samples can result in significant penalties. Lidocaine was observed as a monoprotonated ion at m/z 235 by ESI+ MS/MS (electrospray ionization-positive ion mode) analysis. The base peak ion at m/...
Keen B, Cawley A, Fouracre C, Pyke J, Fu S.The emergence of novel doping agents is a continuous issue for analysts who aim to maintain the integrity of horseracing together with the well-being and safety of the animals and riders involved. Untargeted mass spectrometric analysis presents a potential improvement for antidoping as it enables the detection of compounds being indirectly affected by an administered drug. In this study, liquid chromatography-high-resolution mass spectrometry was used to investigate a 12-horse administration study of the synthetic opioid, butorphanol. A mass spectrometric workflow capable of detecting metaboli...
Lasne F, Popot MA, Varlet-Marie E, Martin L, Martin JA, Bonnaire Y, Audran M, de Ceaurriz J.A direct detection method for anti-doping control of recombinant human erythropoietin (rHuEPO) abuse in racehorses is proposed. This method involves screening of plasma (or serum) by an enzyme-linked immunosorbent assay specific for human EPO and confirmation in urine samples by characterization of the urinary EPO isoelectric profile. This method was tested on horses that were administered epoetin alpha (rHuEPO) and the hyper-glycosylated form of this drug (darbepoetin alpha).
Philip M, Karakka Kal AK, Subhahar MB, Karatt TK, Mathew B, Perwad Z.Performance-enhancing substances and methods have become a serious problem in competitive sports. The hypoxia-inducible factor (HIF) stabilizers can enhance the organism's capacity for molecular oxygen transport and are likely to be abused as performance-enhancing agents in sports. This paper describes the metabolic conversion of the popular hypoxia inducible factor-prolyl hydroxylase (HIF-PH) inhibitors, namely, daprodustat, desidustat, and vadadustat using equine liver microsomes, determined on a QExactive high-resolution mass spectrometer. During this study, a total of 10 metabolites for da...
Wynne PM, Vine JH, Amiet RG.The influence of sampling variables on the concentration of the dopamine metabolites 3-methoxytyramine (3MT), dihydroxyphenylacetic acid (DOPAC) and homovanilic acid (HVA) was examined in equine urine. A logarithmic transformation of the data for all horses gave distribution which approximated the normal distributions for each metabolite. The mean urinary concentration of 3 MT in horses was 214 ng/mL and the application of a threshold with a probability of 1 in 10,000 gave an actionable level of 4 microg/mL. Environmental variables were not forensically significant in determining the populatio...
Jaussaud P, Guieu D, Courtot D, Barbier B, Bonnaire Y.A tolfenamic acid metabolite, a hydroxylated product, has been identified in equine plasma and urine samples using gas chromatography-mass spectrometry in the electron-impact and chemical-ionization modes. The method also allows the qualitative monitoring of the elimination of the drug and its metabolites from plasma. The two compounds are detected up to 48 and 24 h, respectively, after a single oral administration of a 30 mg/kg dose. The simultaneous detection of the two products increases the reliability of anti-doping control analysis.
Wong ASY, Leung GNW, Leung DKK, Wan TSM.Anabolic steroids are banned substances in equine sports. Gas chromatography-mass spectrometry (GC-MS) has been the traditional technique for doping control analysis of anabolic steroids in biological samples. Although liquid chromatography-mass spectrometry (LC/MS) has become an important technique in doping control, the detection of saturated hydroxysteroids by LC-MS remains a problem due to their low ionization efficiency under electrospray. The recent development in fast-scanning gas-chromatography-triple-quadrupole mass spectrometry (GC-MS/MS) has provided a better alternative with a sign...
Hagedorn HW, Böck M, Schulz R.Equine blood may contain salicylic acid (SA) taken up as free acid or represents the metabolite of acetylsalicylic acid (ASA). To obtain information of SA in race horses we screened blood samples of trotting-horses routinely drawn to be analyzed for doping substances. The individual values determined followed a Gaussian distribution displaying a geometric mean of 19 ng SA per ml serum. A probit analysis revealed linear relationship (r = 0.995). Additional studies examined the antithrombotic efficacy of ASA in the horse. An oral dose of 300 mg ASA considerably elevated the bleeding time for mor...
The Journal of nutritionDecember 1, 1994
Volume 124, Issue 12 Suppl 2730S-2735S doi: 10.1093/jn/124.suppl_12.2730S
Snow DH.Since the domestication of the horse and its use in various types of athletic competition, its diet has developed from that associated with grazing to feeding of additional energy in the form of grains up to the present situation when numerous supplements are fed in hopes of enhancing performance. Many if not all of these may be considered as ergogenic aids, and under the rules of racing in many countries should be considered as a prohibited substance. Until recently, a blind eye has been turned to whether the rules should be enforced against many nutritional supplements, for example, vitamins...
Göktaş EF, Kabil E, Arıöz F.Nonsteroidal anti-inflammatory drugs (NSAIDs) are commonly used in therapeutic doses in human and veterinary medicine for the treatment of inflammation, pain, and fever. A method for the simultaneous determination of nine NSAIDs, known as therapeutic prohibited substances, in equine urine was developed and fully validated according to the European Commission Decision 2002/657/EC and Association of Official Racing Chemists criteria. The validation was performed for naproxen, flunixin, ketoprofen, diclofenac, eltenac, meclofenamic acid, phenylbutazone, vedaprofen, and carprofen in equine urine i...
Stucki M, Voegel CD, Binz TM, Kraemer T, Lavaud A, Voelter K.Equine sport agencies list steroids as prohibited substances for competing horses. Objective: The objective of this study was to investigate if the controlled substances dexamethasone and prednisolone are detectable in equine serum and urine samples during and after treatment with eye drops and if this can generate a positive doping test. Methods: Prospective cohort study. Methods: The study cohort included 11 horses. One eye of the horses was treated with either dexamethasone (Maxitrol 0.1%, n = 5 eyes) or prednisolone (Pred forte 1%, n = 6 eyes) eye drops 3 times daily for 14 days. Dexa...
Díaz S, Kienast ME, Villegas-Castagnasso EE, Pena NL, Manganare MM, Posik D, Peral-García P, Giovambattista G.In order to detect switching and/or manipulation of samples, the owner of a stallion asked our lab to perform a DNA test on a positive doping urine sample. The objective was to compare the urine DNA profile versus blood and hair DNA profiles from the same stallion. At first, 10 microsatellite markers were investigated to determine the horse identity. No results were obtained when horse specific markers were typed in the urine sample. In order to confirm the species origin of this sample we analyzed the mitochondrial cytochrome b gene. This analysis from blood and hair samples produced reproduc...
Frincke JM, Henderson GL.Fentanyl, a potent, synthetic narcotic analgesic, has reportedly been used to "dope" racehorses. Urine was collected from a horse dosed with 70 mg of [3H]fentanyl, and the primary metabolite, a water-soluble, amphoteric compound, was isolated by high-pressure liquid chromatography and identified by spectroscopic analysis. This metabolite was found to be N-[1-(2-phenethyl-4-piperidinyl)] malonanilinic acid.
Karatt TK, Sathiq MA, Laya S, Kal AKK, Subhahar MB, Muhammed Ajeebsanu MP, Philip M, Caveney MR, Graiban FM.Nonsteroidal selective androgen receptor modulators (SARMs) are a novel class of compounds that have not yet been clinically approved; however, they appear to have a better anabolic/androgenic ratio than steroids and cause slighter side effects. Sports drug testing laboratories are required to maintain continuously updated doping control analytical methods in light of the widespread misuse of SARMs in elite and amateur sports. This paper describes the metabolic conversion of SARM GSK2881078 in thoroughbred horses following oral administration and in vitro with equine liver microsomes. A liquid...
Momose A, Tsuji T.When sulpyrine and aminopyrine are administered to the horse, unchanged aminopyrine and its metabolites, 4-methylaminoantipyrine and 4-aminoantipyrine, are detected in the urine by means of thin-layer chromatography (TLC) and gas liquid chromatography. Further identification of aminopyrine and these metabolites was carried out by the gas chromatography-mass spectrometry (GC-MS) method. The procedures for separation and identification are as follows : The excretions were adjusted to pH 9 with ammonium hydroxide and extracted with chloroform. The extract was separated by TLC. The spots were loca...
Waraksa E, Owczarek K, Kubica P, Kłodzińska E, Ozimek M, Wrzesień R, Bobrowska-Korczak B, Namieśnik J.Bromhexine (BH), expectorant used in the treatment of respiratory disorders associated with viscid or excessive mucus, is not permitted for use in the competing horse by many authorities in horseracing and Olympic disciplines. Metabolic studies are of the great importance in anti-doping field because they allow for updating the selection of the most appropriate markers for prohibited substances, such as metabolites present at higher concentration levels and/or lasted for a longer period of time in biological samples than a parent drug. This study describes LC-MS/MS-based method for simultaneou...
Harkins JD, Lehner A, Karpiesiuk W, Woods WE, Dirikolu L, Boyles J, Carter WG, Tobin T.Bupivacaine is a potent local anaesthetic used in equine medicine. It is also classified as a Class 2 foreign substance by the Association of Racing Commissioners International (ARCI). The identification of residues in postrace urine samples may cause regulators to impose significant penalties. Therefore, an analytical/pharmacological database was developed for this medication. The highest no-effect dose (HNED) for the local anaesthetic effect of bupivacaine was determined to be 0.25 mg by using an abaxial sesamoid local anaesthetic model. Administration of the HNED of bupivacaine to eight hor...
Leung GN, Kwok WH, Wan TS, Lam KK, Schiff PJ.Formestane (4-hydroxyandrost-4-ene-3,17-dione) is an irreversible steroidal aromatase inhibitor with reported abuse in human sports. In 2011, our laboratory identified the presence of formestane in a horse urine sample from an overseas jurisdiction. This was the first reported case of formestane in a racehorse. The metabolism of formestane in humans has been reported previously; however, little is known about its metabolic fate in horses. This paper describes the in vitro and in vivo metabolic studies of formestane in horses, with the objective of identifying the target metabolite with the lon...
Lehner AF, Hughes CG, Karpiesiuk W, Harkins JD, Dirikolu L, Bosken J, Camargo F, Boyles J, Troppmann A, Woods WE, Tobin T.Amitraz (N'-(2,4-dimethylphenyl)-N-[[(2,4-dimethylphenyl)imino]methyl]-N-methyl-methanimidamide) is an alpha-2 adrenergic agonist used in veterinary medicine primarily as a scabicide- or acaricide-type insecticide. As an alpha-2 adrenergic agonist, it also has sedative/tranquilizing properties and is, therefore, listed as an Association of Racing Commissioners International Class 3 Foreign Substance, indicating its potential to influence the outcome of horse races. We identified the principal equine metabolite of amitraz as N-2,4-dimethylphenyl-N'-methylformamidine by electrospray ionization(+...
Yu J, Han KS, Lee G, Paik MJ, Kim KR.The enantioseparation of pranoprofen after its addition in racemic form into equine plasma and urine was conducted by chiral liquid chromatography-tandem mass spectrometry in selected reaction monitoring mode. The methods for the assay of both enantiomers were linear (r≥0.9943) in the low range from 0.001 to 0.1μg/mL and high range from 0.01 to 1.0μg/mL with good precision (% RSD≤5.6) and accuracy (% RE=-5.3 to 1.9). When racemic pranoprofen was orally administered to four horses at a single dose of 3.1mg/kg, the median plasma concentrations of (R)-pranoprofen were lower than the levels ...
Matassa LC, Woodard D, Leavitt RK, Firby P, Beaumier P.Glycopyrrolate (Robinul) is a quaternary ammonium salt which serves as a respiratory enhancing drug. It is reportedly used in horse racing to improve breathing. Extraction of glycopyrrolate from equine urine employing unique solid-phase extraction techniques gave a residue suitable for liquid chromatography-tandem mass spectrometry (LC-MS-MS) and gas chromatography-mass spectrometry (GC-MS). LC-MS-MS analysis employed an extract derived from 5 ml of urine subjected to cation-exchange chromatography. The daughter ion of m/z 318 monitored in the positive-ion mode was m/z 116. Recovery of glycopy...
You Y, Proctor RM, Haughan J, Missanelli JR, Robinson MA.Fentanyl, a powerful synthetic mu opioid receptor agonist, is banned in equine sports by the Association of Racing Commissioners International and the Fédération Équestre Internationale. The presence of fentanyl in equine blood has been confirmed during routine post-race screening for doping substances in the authors' laboratory. While fentanyl can be detected and confirmed in blood, it is rapidly metabolized, and screening for the metabolite N-[1-(2-phenethy-4-piperidinyl)] maloanilinic acid (PMA) in equine urine is expected to allow for a longer detection time. In this study, a quantitati...
Tozaki T, Ohnuma A, Kikuchi M, Ishige T, Kakoi H, Hirota KI, Takahashi Y, Nagata SI.Gene doping, which is prohibited in horseracing and equestrian sports, can be performed by introducing exogenous genes, known as transgenes, into the bodies of postnatal animals. To detect exogenous genes, a method utilizing quantitative polymerase chain reaction (qPCR) with a hydrolysis probe was developed to test whole blood and plasma samples, thereby protecting the fairness of competition and the rights of stakeholders in horseracing and equestrian sports. Therefore, we aimed to develop sample storage methods suitable for A and B samples in gene doping tests using blood. For sample A, suff...
