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Topic:Analytical Methods
Analytical methods in equine research encompass a variety of scientific techniques and tools used to study and evaluate different aspects of horse health, performance, and physiology. These methods help advance our understanding of equine biology, diagnosing conditions, and improving management practices. Common analytical methods include molecular techniques like PCR and ELISA for detecting pathogens and measuring biomarkers, imaging technologies such as ultrasound and MRI for assessing musculoskeletal health, and statistical models for analyzing genetic data and performance metrics. This page compiles peer-reviewed research studies and scholarly articles that explore the development, application, and impact of various analytical methods in equine science.
Doping Control Analysis of Perfluorocarbons in Equine Plasma by Headspace Gas Chromatography-Tandem Mass Spectrometry. In order to control the potential misuse of perfluorocarbons as an oxygen carrier in equine sports, a simple and sensitive method for detecting perfluorocarbons in equine plasma by gas chromatography/tandem mass spectrometry using negative chemical ionization with methane as reagent gas has been developed and fully validated. The method covers seven perfluorocarbons, which are the active components in blood substitute products, and shows good sensitivity and robustness. Limits of detection as low as 0.01 ng/mL could be achieved by the method. To the best of our knowledge, this is the first r...
Evaluating the Effects of Solid-Phase Cartridge Chemistry on Extraction of Bisphosphonates and ITPP From Equine Plasma. Bisphosphonate drugs and myo-inositol trispyrophosphate are of concern to the racing industry and have been listed as prohibited substances in equine sports. The current bisphosphonate plasma screening analysis employed at the Australian Racing Forensic Laboratory involves the use of sequential solid-phase extraction procedures, passing the samples through a mixed mode cartridge, followed by a weak anion exchange cartridge. The eluates collected following the second extraction are then methylated and analysed by liquid chromatography-mass spectrometry. Under these extraction conditions, some b...
Pharmacokinetics of Ethyl Glucuronide and Ethyl Sulfate and Pharmacodynamic Effects Following Intravenous and Oral Administration of Ethanol to Exercised Horses. Ethanol, a central nervous system depressant and banned substance in horseracing, has reportedly been administered to horses prior to competition to "calm a horse's nerves." In this study, the pharmacokinetics of two metabolites of ethanol were studied to better understand the behavior of this compound in the horse and provide a scientific basis for regulation of its administration. Six horses received a single intravenous (30 mL; 1200 mg) and oral (90 mL; 3600 mg) administration of ethanol (vodka, 40% ABV) in a balanced cross-over design. Blood and urine samples were collected at vari...
Horseshoe effects on equine gait-A systematic scoping review. To provide an overview of available research about effects of horseshoes on equine kinetics and kinematics. Methods: The terms, "horse/equine," "hoof," "shoes/horseshoes," "kinetics," and "kinematics" were searched in PubMed, Web of Science, Center for Agriculture and Bioscience International, and United States Department of Agriculture National Agricultural Library for manuscripts from first availability to 2024. Independent reviewers used preferred reporting items for systematic reviews and meta-analyses guidelines to map and extract evidence-based data from sources. Metrics included partici...
In vitro evaluation of the automated hematology analyzer XN-31 for rapid diagnosis of equine piroplasmosis. Equine piroplasmosis (EP) is a protozoal disease affecting equids, caused by and . EP is conventionally diagnosed using microscopic, molecular, and/or serological methods, which are time-consuming. Consequently, there is a need for faster testing methods. In this study, we evaluated the application of the Sysmex XN-31 automated hematology analyzer, originally a rapid test for detecting malaria in humans, for the diagnosis of EP. The cultured parasites were measured using the XN-31 that had been customized for horse blood samples (XN-31m). The following parameters were evaluated: limit of dete...
Detection of levamisole and its metabolites in horses after oral levamisole administration over seven days. Levamisole is a regulated substance sometimes administered to racehorses to treat equine protozoal myelitis. Metabolites include compound II, aminorex, and pemoline. Aminorex and pemoline are Horseracing Integrity and Safety Authority-banned substances. Previous studies have examined single doses of the drug. This study examined the disposition of levamisole after 7 days of dosing. Methods: 6 healthy Thoroughbred geldings. Methods: Horses were treated with 500 mg (approx 0.91 to 1 mg/kg) of compounded levamisole hydrochloride paste PO every 12 hours for a total of 13 doses over 7 days. Serum a...
Extracted Plasma Cell-Free DNA Concentrations Are Elevated in Colic Patients with Systemic Inflammation. Colic is a common and potentially life-threatening condition in horses; in many cases, it remains challenging for clinicians to determine the cause, appropriate treatment, and prognosis. One approach that could improve patient care and outcomes is identification of novel diagnostic and prognostic biomarkers. Plasma cell-free DNA (cfDNA) is a biomarker that shows promise for characterizing disease severity and predicting survival in humans with acute abdominal pain or requiring emergency abdominal surgery. In horses, we recently determined that extracted plasma cfDNA concentrations are elevated...
Quantification of osilodrostat in horse urine using LC/ESI-HRMS to establish an elimination profile for doping control. The use of osilodrostat, developed as a medication for Cushing's disease but categorized as an anabolic agent, is banned in horses by both the International Federation of Horseracing Authorities and the Fédération Equestre Internationale. For doping control purposes, elimination profiles of hydrolyzed osilodrostat in horse urine were established and the detectability of free forms of osilodrostat and its major metabolite, mono-hydroxylated osilodrostat (M1c), was investigated. Post-administration urine samples obtained from a gelding and three mares were analyzed to establish the eliminatio...
Metabolic Pathway of Osilodrostat in Equine Urine Established through High-resolution Mass Spectrometric Characterization for Doping Control. Osilodrostat, used to treat Cushing's disease, exhibits an anabolic effect, leading to its classification as a prohibited substance in horseracing and equestrian sports. This study reports the characterization of osilodrostat metabolites in horse urine and elucidates its metabolic pathways for the first time for doping control purposes. Methods: Osilodrostat was administered nasoesophageally to four thoroughbreds (one gelding and three mares) at a dose of 50 mg each. Potential metabolites were extensively screened via our developed generic approach employing differential analysis to identify m...
Effects of storage-handling methods on nutrient analysis of fresh-forage samples. Forage low in nonstructural carbohydrates (NSC) is recommended for insulin dysregulated (ID) horses, indicating the importance of an accurate forage analysis. However, it remains to be fully understood how handling forage samples pre-analysis impacts nutrient values. The objective of this study was to compare the effect of fresh-forage storage-handling methods: microwave-oven (MO; 9kw; 70s then stored at -20℃), room temperature (20℃), 3℃, -20℃, and -80℃. Subsamples collected from a mixed-grass pasture and a ryegrass plot were analyzed for crude protein (CP), water soluble carbohydrat...
Comparison of a hand-held high-end resolution infrared thermography (FLIR P640) and a smartphone infrared thermographic device (FLIR One) for the assessment of skin surface temperature after anaesthetising the median nerve in Healthy horses. Accuracy of a median nerve block is normally assessed by testing skin sensitivity on the medial and dorsal aspects of fetlock and pastern. The present study evaluated subjective and objective analysis of skin surface temperature obtained with two different infrared (IR) thermography cameras (a high-end [FLIR P640] and a smartphone IR thermography device [FLIR One®]) before and after anaesthesia of the median nerve. Thermographic images were obtained at 0, 30, 60 and 90 minutes after performing a median nerve block with 2% mepivacaine hydrochloride. The subjective analysis of thermographic ima...
Einthoven’s triangle adapted for horses: Proposal for the Delta configuration. Reliable ECGs are crucial for diagnosing arrhythmias, yet a lack of standardization impedes arrhythmia diagnosis and treatment in horses. Objective: To objectively determine an optimal position of Einthoven's triangle for ECG recordings in horses at rest, which can form the basis for standardized ECG recording and improve diagnosis and treatment of arrhythmias. Methods: The study involved 72 healthy, warmblood horses aged between 3 and 20 years. Methods: In view of future 12-lead studies and vectorcardiography, requiring an orthogonal system, Einthoven's triangle was positioned around the he...
Exploring the Dissolution, Solid-state Properties, and Long-term Storage Stability of Cryoprotectant-free Fenbendazole Nanoparticles. Fenbendazole is an antiparasitic drug widely used in veterinary medicine to treat parasitic infections caused in animals like cattle, horses, sheep, and dogs. Recently, it has been repositioned as a potential alternative for cancer treatment. However, it is a highly hydrophobic molecule (0.9 ug/mL), which can compromise its dissolution rate and absorption. Thus, this work aimed to apply a nanotechnological approach to improve drug solubility and dissolution performance. Fenbendazole nanoparticles stabilized by different poloxamers were obtained by lyophilization without cryoprotectants. The be...
Quality Assessment of Reconstructed Cow, Camel and Mare Milk Powders by Near-Infrared Spectroscopy and Chemometrics. Milk powders are becoming a major attraction for many industrial applications due to their nutritional and functional properties. Different types of powdered milk, each with their own distinct chemical compositions, can have different functionalities. Consequently, the development of rapid monitoring methods is becoming an urgent task to explore and expand their applicability. Lately, there is growing emphasis on the potential of near-infrared spectroscopy (NIRS) as a rapid technique for the quality assessment of dairy products. In the present work, we explored the potential of NIRS coupled wi...
Explainable machine learning for assessing upper respiratory tract of racehorses from endoscopy videos. Laryngeal hemiplegia (LH) is a major upper respiratory tract (URT) complication in racehorses. Endoscopy imaging of horse throat is a gold standard for URT assessment. However, current manual assessment faces several challenges, stemming from the poor quality of endoscopy videos and subjectivity of manual grading. To overcome such limitations, we propose an explainable machine learning (ML)-based solution for efficient URT assessment. Specifically, a cascaded YOLOv8 architecture is utilized to segment the key semantic regions and landmarks per frame. Several spatiotemporal features are then ex...
Parasympathetic tone activity, heart rate, and grimace scale in conscious horses of 3 breeds before, during, and after nociceptive mechanical stimulation. Parasympathetic tone activity (PTA) in response to nociceptive stimulus in conscious non-sedated horses is unknown. Objective: Study PTA, heart rate (HR), and horse grimace scale (HGS) at rest and during mechanical nociceptive stimulation. Methods: Ninety healthy young adult horses (females, males): 30 each of Friesians, Quarter Horses, and Warmbloods. Methods: Prospective control study. The study consisted of habituation to equipment (Day 1), baseline recordings (Days 2 and 3), and nociceptive testing applying mild pressure to the metacarpus (Day 4). Parasympathetic tone, HR, and HGS were rec...
Flowmetry and spectrophotometry can detect reduced intestinal microperfusion in nonsurvivors during equine colic surgery for large intestinal strangulation. To evaluate the use of laser Doppler flowmetry and spectrophotometry (LDFS) for large intestinal viability assessment in horses with naturally occurring large intestinal strangulations. Methods: By use of LDFS, intestinal microperfusion was quantified as tissue oxygen saturation (tSo2), hemoglobin (tHB), and blood flow (tBF) in cases with large colon volvulus and small colon strangulations undergoing colic surgery (n = 17). Intestinal biopsies were taken from the pelvic flexure in all large colon cases and in small colon cases that underwent intraoperative euthanasia. Measurements were compare...
In vivo comparative study of hemp straw exposure and cannabidiol oil administration in horse urine. The non-psychoactive cannabinoids cannabidiol (CBD) and cannabidiolic acid (CBDA) are available on the market in different forms, mostly for their anti-inflammatory and potential analgesic properties. These substances are prohibited during equine competitions. CBD and CBDA are naturally present in hemp straw, commonly used as a bedding substitute for wheat straw. Unfortunately, horses can eat it, which therefore could lead to a possible risk of positive findings for CBD/CBDA in biological samples after doping control tests. The goals of this study were, first, to provide recommendations on the...
Saliva and plasma metabolome changes during anoestrus, the oestrous cycle and early gestation in the mare: A pilot study. Successful reproductive management of domestic mammals depends primarily upon timely identification of oestrous cycle stages. There is a need to develop an alternative non-invasive, welfare-friendly, accurate and reliable method to identify reproductive cycle stages. This is of particular interest for horse breeders, because horses are high-value farm animals that require careful management and individual monitoring. Saliva sampling is non-invasive, painless and welfare-friendly. Thus, we performed a metabolomic analysis of equine saliva during different reproductive stages to identify changes...
Rapid Detection of Getah Virus Antibodies in Horses Using a Recombinant E2 Protein-Based Immunochromatographic Strip. The prevalence and impact of Getah virus (GETV) are significant concerns in China. GETV can infect a wide range of animals, including horses, pigs, sheep, cattle, birds, and humans, resulting in substantial losses in the livestock and agricultural industries. GETV infection can cause the development of ulcers and inflammation in the mouth and gums of horses, which result in pain and discomfort and lead to symptoms such as reduced appetite, drooling, and difficulty chewing. As a result, there is a pressing need for efficient and rapid disease diagnosis methods. However, the currently available ...
Pharmacokinetics and Alterations in Glucose and Insulin Levels After a Single Dose of Canagliflozin in Healthy Icelandic Horses. Canagliflozin (CFZ) is a sodium-glucose cotransporter-2 inhibitor that has shown promising results as a drug for the treatment of insulin dysregulation in horses. Even though CFZ is used clinically, no pharmacokinetic data has previously been published. In this study, the pharmacokinetics of CFZ after administration of a single oral dose of 1.8 mg/kg in eight healthy Icelandic horses was examined. Additionally, the effect of treatment on glucose and insulin levels in response to a graded glucose infusion was investigated. Plasma samples for CFZ quantification were taken at 0, 0.33, 0.66, 1, ...
Enhanced analysis of equine plasma for the presence of recombinant human erythropoietin – Implementation of an improved workflow. An improved screening workflow and a robust capillary flow LC-MS confirmatory method for the detection of recombinant human erythropoietin (rHuEPO) has been implemented to increase the sensitivity of rHuEPO detection and to reduce the number of suspect samples committed to confirmatory testing. The influence of repeated dosing of epoetin-β on the detection window of rHuEPO in equine plasma was assessed using the optimised method. Samples were initially assessed using an economical R&D Human EPO Duo-Set ELISA Development System. Samples indicating a result greater than the batch baseline were ...
Shape and joint angle data for seven European horse breeds and their repeatability. Conformation traits are important in the selection and distinction between horse breeds, but tend to be evaluated subjectively within a breed and cannot be compared between them. The horse shape space model, using a combination of 253 landmarks and semi-landmarks, provides objective information on the shape of a horse photographed from the side that can be compared between breeds. In this dataset, we are providing the full set of 253 landmarks for 1241 horses from seven breeds, including an R code file to extract joint angle information and transform the raw data into csv files for further ana...
Artificial mitochondrial transplantation (AMT) reverses aging of mesenchymal stem cells and improves their immunomodulatory properties in LPS-induced synoviocytes inflammation. Nowadays, regenerative medicine techniques are usually based on the application of mesenchymal stem cells (MSCs) for the repair or restoration of injured damaged tissues. However, the effectiveness of autologous therapy is limited as therapeutic potential of MSCs declines due to patient's age, health condition and prolonged in vitro cultivation as a result of decreased growth rate. For that reason, there is an urgent need to develop strategies enabling the in vitro rejuvenation of MSCs prior transplantation in order to enhance their in vivo therapeutic efficiency. In presented study, we attemp...
The Effects of Intra-Articular Triamcinolone and Autologous Protein Solution on Metabolic Parameters in Horses. Intra-articular corticosteroids are a popular treatment choice for joint-associated pain and inflammation in horses despite recent work on the metabolic effects of these drugs. The goal of this project was to compare metabolic effects between intra-articular (IA) triamcinolone acetonide (TA) and an autologous protein solution (APS). Five mixed-breed geldings (4-9 years) were utilized for this project. Three identical and consecutive 28-day treatment blocks were used, with metacarpophalangeal IA treatments consisting of equal volumes of saline, a commercially available APS, or 9 mg of TA. Regul...
Development and validation of general plasma screening method for performance enhancing drugs in racehorses utilizing liquid chromatography-high-resolution mass spectrometry (LC-HRMS). The screening of drugs in plasma and urine often requires initial extraction (such as liquid-liquid extraction and solid-phase extraction) before the samples are submitted to instrumental analyses. These extraction procedures are often laborious and time-consuming. In this manuscript, a high-throughput automated assay based on liquid chromatography-high-resolution mass spectrometry (LC-HRMS) suitable for use as an initial testing procedure covering multiple classes of compounds prohibited in horse racing is described. The assay requires a 600-μL plasma aliquot, which is subjected to solid pha...
Selenium deposition in an atypically disintegrated hoof wall in a Thoroughbred racehorse with alkali disease: Proof by energy-dispersive X-ray fluorescence analysis. From summer 2018 to summer 2019, several Thoroughbred racehorses held at the Miho Training Centre of the Japan Racing Association inadvertently ingested excessive amounts of sodium selenite, resulting in typical chronic selenium (Se) poisoning - the so-called alkali disease. The typical abnormality was a hoof wall disorder with a circumferentially deep ring and/or transverse hoof wall cracks parallel to the coronet on all feet and appearing after excessive ingestion. One affected Thoroughbred male was unique in that all the hooves had a rough surface with a very fragile hoof wall, but no wall ...
Analytical advances in horseracing medication and doping control from 2018 to 2023. The analytical approaches taken by laboratories to implement robust and efficient regulation of horseracing medication and doping control are complex and constantly evolving. Each laboratory's approach will be dictated by differences in regulatory, economic and scientific drivers specific to their local environment. However, in general, laboratories will all be undertaking developments and improvements to their screening strategies in order to meet new and emerging threats as well as provide improved service to their customers. In this paper, the published analytical advances in horseracing me...
Detection of nonsteroidal and steroidal selective androgen receptor modulators in equine hair after oral administrations. This paper describes the detections of nonsteroidal and steroidal selective androgen receptor modulators (SARMs), namely, RAD140 and YK-11, in mane hair collected from horses having been orally administered with the respective drugs. SARMs are potent anabolic agents with a high potential of misuse in horseracing and equestrian sports, and the misuses of RAD140 and YK-11 in human sports have been reported. To better control the misuse of RAD140 and YK-11 in horses, two separate oral administration studies of RAD140 (0.3 mg/kg daily for 3 days) and YK-11 (0.2 mg/kg daily for 3 days) were pre...
Intelligence-based anti-doping via an Intelligence and Drug Testing Management (IDTM) system. The Intelligence and Drug Testing Management (IDTM), a system that can enhance drug testing analytics with related horse information and intelligence in a single platform, can help identify and mitigate potential doping and other threats.
