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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.
Population Pharmacokinetics and Detection Times of Betamethasone After Intravenous, Intramuscular, and Intranasal Jet Administration of Betamethasone Sodium Phosphate in Horses. Betamethasone sodium phosphate (BETP) is a glucocorticosteroid used in humans. The primary objective was to estimate irrelevant plasma (IPC) and urine (IUC) concentrations using a pharmacokinetic/pharmacodynamic approach based on systemic clearance estimated from intravenous (IV) pharmacokinetic data. As a secondary objective, detection times (DTs) following IV, intramuscular (IM), and intranasal jet (INJ) administration were estimated for medication control. Plasma and urine concentrations were obtained from a total of 20 horses in Japan and Germany, with some horses contributing to more than...
Cycle length variation assists in differentiation of atrial, ventricular, and sinus arrhythmias in horses at rest. Criteria for arrhythmia classification in horses are insufficient, particularly when differentiating atrial premature complexes (APCs) from sinus arrhythmia (SA). This study investigated RR and PP interval changes between SA, APCs, and ventricular premature complexes (VPCs). Unassigned: 30 resting ECGs were reviewed: 10 each containing SA, APC, and VPC. Sequences of 10 RR and PP intervals were recorded for each arrhythmia (20 events/ECG). Percent RR and PP deviations were calculated (1) between sequential intervals (%ΔSEQPP and %ΔSEQRR, respectively) and (2) compared to averaged baseline int...
Detection of Cyclosporine in Horse Hair After Ocular Implant Administration. This study describes the detection and temporal distribution of cyclosporine in equine hair following ocular implant administration in a Thoroughbred gelding. Mane hair samples were collected at 172- and 217-day post-implantation and analysed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) after alkaline hydrolysis and liquid-liquid extraction. The method achieved an estimated limit of detection of 0.1 pg/mg. Cyclosporine was detected in hair at both sampling time points, with segmental analysis revealing distribution patterns consistent with systemic exposure and incorporation ...
Monitoring Estra-4,9-Diene-3,17-Dione Exposure in Equine Hair. This study investigated the incorporation of estra-4,9-diene-3,17-dione (dienedione) and its metabolites into equine hair following exogenous administration and evaluates the potential application of hair analysis for long-term doping surveillance. Mane hair samples were collected from three Thoroughbred castrated horses before and after oral administration of dienedione and from untreated colts for comparison. Dienedione was detected in post-administration hair from all treated horses but was absent in corresponding pre-administration samples. Segmental distribution patterns indicated contrib...
Detection and confirmation of hypoxia-inducible factor activating agents in equine plasma and urine by liquid chromatography coupled to mass spectrometry. Hypoxia-inducible factor (HIF) activating agents can promote the transcription of the erythropoietin gene, enhancing an athlete's performance, and are therefore prohibited in sports. A liquid chromatography coupled to high-resolution mass spectrometry (LCHRMS) method was developed for the detection and confirmation of 11 HIF-activating substances in equine plasma and urine. Ammonium formate added to the mobile phase system suppressed the responses of all the analytes in either positive or negative ion mode and thus was not used. The starting organic percentage in the mobile phase gradient and ...
Technical note on cervical epidural catheterization in two adult horses: Injectate distribution and epidural space effects. To determine the caudalmost extent of diffusion of contrast media administered through a cervical epidural catheter (CEC) and to describe pathologic changes associated with maintaining an indwelling CEC for up to 17 days. Methods: Pilot study. Methods: Two mature horses. Methods: CECs were placed under ultrasound guidance in two horses. Both horses were injected with 5 mL 50% Iohexol contrast solution, followed by radiographs of catheter insertion site to confirm placement. A total volume of 15 mL contrast solution was injected in 5 mL aliquots up to 25 min post-initial injection. La...
Approaches to Thoroughbred Horse Identification in Diverse Sample Types Using Multiplexed SNP-PCR Sequencing Panels. The ability to determine the identity of equine samples would be highly advantageous in cases of ambiguous sample source, as individual-specific controls for gene editing detection, or forensic examination of suspect biological materials in potential doping cases. To this end, a SNP-PCR sequencing-based panel comprising 93 markers in one reaction was designed and investigated for use in Thoroughbred horses. Marker amplification performance and discriminating power were calculated against a population survey of 264 Thoroughbred whole blood samples from routine doping testing. The sensitivity of...
Plasma Homocysteine Concentrations in Horses with Left-Sided Valvular Heart Disease. In human medicine, homocysteine, a sulfur-containing amino acid, is used as a biomarker reflecting endothelial dysfunction, inflammation, thrombogenesis, oxidative stress and cardiovascular disease. An automated enzyme-cycling assay for homocysteine, established for humans, has been validated and reference ranges have been created for horses. In this study the homocysteine assay was further evaluated by assessing short-term stability (whole blood and plasma samples stored at 4 °C and room temperature for 24 h), long-term stability (two years), the effect of different anticoagulants (lithium h...
High-Throughput Equine Plasma Profiling Using Laser-Assisted Rapid Evaporative Ionisation Mass Spectrometry. Remnants of erythrocyte removal from blood constitute therapeutic plasma-a water-based solution containing mainly platelets, extracellular vesicles (EVs), protein and lipid complexes and ions. Plasma-based therapies, including their use to accelerate wound healing in human and animal patients, are increasingly gaining prominence in regenerative medicine; however, methods for the optimal preparation of individual plasma are still under development. Rapid and accurate analysis of plasma composition is essential for the development of therapeutic applications and for ensuring the quality of plasm...
Effect of lingual gauze swab placement and light exclusion on pulse oximeter agreement with arterial haemoglobin-oxygen saturation derived from blood-gas analysis in healthy anaesthetised horses. To compare peripheral arterial haemoglobin-oxygen saturation values obtained using pulse oximetry (SpO) with and without a lingual gauze swab under ambient light and light-exclusion conditions, with arterial haemoglobin-oxygen saturation derived from arterial blood-gas analysis (SaO). Methods: Prospective method-comparison study. Methods: A group of 60 healthy adult horses undergoing elective general anaesthesia. Methods: In American Society of Anesthesiologists (ASA) physical status classification I-II horses, SpO readings were obtained under six conditions during anaesthesia: no swab, no swa...
Evaluation of an Immunoturbidometric Assay for Measurement of Total Adiponectin Concentration in Equine Serum Samples. Total adiponectin (TA) is a marker of metabolic health in equids, with values below a critical decision point of < 7.9 μg/mL conferring increased risk of laminitis. Objective: To evaluate an IT assay to measure serum [TA] in equids and to assess its correlation with [high molecular weight adiponectin] (HMWA) measured by ELISA. Methods: Total serum adiponectin concentrations from 207 adult horses were measured using both an IT assay for TA and HMWA using a previously validated ELISA assay. Data were analyzed using Spearman correlation and Passing-Bablok regression. Intra- and inter-assa...
Plasma and Urinary Concentrations of Synephrine in Horses Following Controlled Oral Administration. Synephrine is a sympathomimetic alkaloid of regulatory interest in both horse racing and equestrian sports, where feed‑related exposure may lead to its detection in equine blood and urine. This study characterized plasma and urinary concentrations of synephrine in horses following controlled oral administration. Six Thoroughbred horses received single oral doses of 100 mg or 800 mg synephrine, and blood and urine samples were collected sequentially. Each sample was analyzed both before and after β‑glucuronidase hydrolysis, demonstrating that synephrine was present predominantly as conjuga...
Simple and efficient HPLC-MS/MS method for simultaneous extraction and quantification of anthelmintics in equine manure. Anthelmintic treatment is a common practice in horses. The presence of pharmaceutical residues in fields and pastures is well-documented affecting various organisms, like nematodes. This study describes the development and validation of a high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method using multiple reaction monitoring (MRM) for extracting and determining the concentrations of four commonly used anthelmintics in horse manure. Manure was initially dried, then spiked with various concentrations (10-10,000 ng) of anthelmintics and extracted using an acetonit...
Detection of ionisable lipids in equine plasma by supported liquid extraction and liquid chromatography-high-resolution tandem mass spectrometry for doping control of mRNA agents. Messenger ribonucleic acid (mRNA) has emerged as a new class of therapeutic agent with unprecedented potential. In particular, the lipid nanoparticle (LNP)-mRNA formulations enabled safe and effective delivery of mRNA in vivo, leading to the success of mRNA vaccines during the COVID-19 pandemic. To control the potential misuse of LNP-mRNA agents in equine sports, a simple and sensitive method has been developed for the detection of ionisable lipids in equine plasma using supported liquid extraction (SLE) followed by liquid chromatography-high-resolution tandem mass spectrometry. To overcome th...
Administration Study of Bedinvetmab, an Antinerve Growth Factor Monoclonal Antibody, in Horses for Doping Control Purposes. Monoclonal antibodies (mAbs) targeting nerve growth factor (NGF) represent a novel therapeutic approach for managing acute and chronic pain in humans and animals. However, their potential misuse as pain-masking agents in competitive equine sports raises significant welfare and integrity concerns. To enhance understanding of the elimination profile of anti-NGF mAb if misused in horses, this study describes the first administration of bedinvetmab (a canine anti-NGF mAb) in three geldings, establishing its plasma elimination profile following subcutaneous administration. Bedinvetmab plasma concen...
Surface electromyography and joint angle kinematics of horses at walk and trot on a treadmill. This study aimed to evaluate electromyographic and kinematic parameters of the thoracic and pelvic limbs of horses at a walk and trot on a treadmill. Eleven Brazilian Sport Horses trained for Eventing were assessed in a completely randomized design under two conditions: walk (1.8 m/s) and trot (3.6 m/s). Horses were evaluated on a high-speed treadmill using surface electromyography (sEMG) and kinematic analysis. sEMG was performed bilaterally on the mm. extensor carpi radialis and mm. flexor carpi ulnaris; bilateral muscles of the pelvic limb: mm. tensor fascia latae and mm. biceps femoris u...
Repeatability and test-retest reliability of thermal and pressure pain threshold testing in healthy thoroughbred horses. Quantitative sensory testing (QST) has been used in horses to assess nociceptive responsiveness but its broader clinical application is limited by variability across anatomical sites, environmental conditions, operator technique, and behavioral endpoints. This study aimed to evaluate the feasibility, within-session repeatability, and test-retest reliability of thermal threshold (TT) and pressure pain threshold (PPT) testing in healthy Thoroughbred horses. Unassigned: Eleven adult Thoroughbred horses (4 mares, 7 geldings; median [IQR] age 6 [5-6] years; body weight 558 ± 47 kg) underwent...
Correction: Phenylbutazone concentrations in synovial fluid following administration via intravenous regional limb perfusion in the forelimbs of six adult horses. [This corrects the article DOI: 10.3389/fmed.2026.1810077.].
Detecting and measuring respiratory events in horses during exercise with a microphone: deep learning vs. standard signal processing. Monitoring respiration parameters such as respiratory rate could be beneficial to understand the impact of training on equine health and performance and ultimately improve equine welfare. In this work, we compare deep learning-based methods to an adapted signal processing method to automatically detect cyclic respiratory events and extract the dynamic respiratory rate from microphone recordings during high-intensity exercise in Standardbred trotters. Results: Our deep learning models are able to detect exhalation sounds (median F1 score of 0.94) in noisy microphone signals and show promising r...
Forage nutritive analysis in practical equine feeding: challenges and opportunities. Previous research has shown that forage nutritive analyses are not used to any large extent in equine nutrition. As forages differ widely in nutritional quality, knowledge of the nutritive value is essential to avoid under- or overfeeding of horses. Objective: The aim of this study was to investigate how forage nutritive analyses were used among horse owners in Sweden, and to identify challenges horse owners encounter with such analyses. Methods: Data was collected through an online survey with questions about the respondent, access to forage analytical reports, if and how the respondent used ...
A Comprehensive Analysis of the Agreement and Performance of Variant Annotation Programs in Equine Genomes. Advances in whole-genome sequencing (WGS) technology have led to the widespread adoption of WGS for investigating genetic diseases and traits in domestic animals. This has created a need for improved methods for prioritizing candidate causal variants. One way variants are prioritized is using variant annotators that predict variant effects based on their proximity to genomic features and effect on amino acid sequence. However, validation of variant annotators for domestic animal genomes is lacking. In this study, we calculated the agreement of three popular variant annotators, Ensembl Varian...
Evaluation of a canine Coll2-1 ELISA kit for use in equine serum and its correlation with radiographic joint disease. This study evaluated an enzyme-linked immunosorbent assay (ELISA) for the detection of Coll2-1, a peptide fragment of type II collagen released during cartilage degradation in horses using a kit originally developed for dogs. Objective: The objectives of this study were to: 1) assess the analytical applicability of the canine Coll2-1 ELISA in equine serum and 2) explore the correlations between Coll2-1 concentrations and radiographic scores of osteoarthritis (OA) in horses. Methods: Twelve Lusitano stallions, of similar age and workload but presenting different radiographic scores were include...
Single-dose intravenous α2-agonists do not acutely alter anti-inflammatory bioactive protein profiles but reduce interleukin-8 in equine autologous protein solution and platelet-rich plasma. Determine the acute effects of xylazine and detomidine on concentrations of selected growth factors, cytokines, and α2-macroglobulin (A2M) in platelet-rich plasma (PRP) and autologous protein solution (APS) and the long-term effects of these sedatives on serum A2M. We hypothesized that sedative administration would decrease concentrations of all bioactive proteins in PRP and APS acutely and A2M in serum for 6 days following sedative administration. Unassigned: 6 horses were randomized to receive IV xylazine (200 mg) or detomidine (10 mg). Blood was collected before and 5 minutes after sedativ...
Validation of measurable radiographic parameters for quantitative assessment of carpal and metacarpal bone morphology in foals. Radiographic morphometry provides a quantitative means to evaluate bone conformation in horses, yet validation of these parameters in foals has not previously been undertaken. This study aimed to determine the intra- and inter-rater repeatability and reproducibility of established carpal and third metacarpal (MC3) radiographic parameters in Thoroughbred foals and to establish baseline conformational data. Twenty Thoroughbred foals were radiographed without sedation from birth to 14 months of age at two-month intervals. Dorsopalmar carpal and lateromedial MC3 radiographs were obtained using sta...
OPTIMIZING EXTRACTION AND CONCENTRATION METHODS FOR LC-MS/MS ANALYSIS OF VETERINARY PHARMACEUTICALS IN EQUINE-ASSOCIATED ENVIRONMENTAL MATRICES. This study developed a method for the simultaneous quantification of six potentially environmentally toxic veterinary drugs: benzylpenicillin, acepromazine, flunixin, dexamethasone, ketamine and romifidine, in soils, sediments, equine faeces and natural waters. Three methods were compared for their extraction from solid matrices: Ultrasonic extraction (USE), QuEChERS and Accelerated Solvent Extraction (ASE). ASE method was optimized according to temperature and extraction cycles. Quantification was performed by LC-MS/MS. LC and MS parameters were optimized. The method was validated according t...
Comparative Analysis of Meat Quality and Flavor Among Four Categories of Mongolian Horses. This study aims to conduct a comparative analysis of the quality and flavor of meat from four categories of Mongolian horses (Wushen, Baicha, Barhu, and Ujimqin). Physicochemical indicators, electronic nose, electronic tongue, and lipidomics were used to characterize meat quality and flavor and to screen for differential markers. Results showed that Wushen Horses had the highest pH, serine, glutamic acid, total free amino acids (∑FAA), total non-essential amino acids (∑NEAA), total amino acids (∑TAA), NEAA/TAA, W2S sensor response, umami and richness values, and had the lowest cooking lo...
Integrative Metabolomic and Echocardiographic Profiling Reveals Metabolic-Cardiac Structural Coupling in Yili Horses During Incremental Exercise. This study integrated echocardiography with widely targeted metabolomics to decipher how plasma metabolic dynamics couple with cardiac geometry in Yili horses during incremental treadmill exercise. Nine speed-type horses underwent a graded exercise test (6% incline; 0 to 9 m/s). Jugular venous blood samples collected at rest (0 m/s) and at 3, 5, 7, and 9 m/s were profiled by LC-MS, and Pearson correlation analysis was applied to relate differentially expressed metabolites (DEMs) to twenty echocardiographic structural indices. A core set of 314 shared DEMs (124 upregulated, 190 downregulated) w...
Feasibility of a hand-held myotonometry device for measuring biomechanical muscle parameters in horses. To evaluate the feasibility of a hand-held myotometry device for measuring the biomechanical parameters (frequency, stiffness, and decrement) of the longissimus dorsi muscle in the thoracolumbar region of healthy horses. Unassigned: Data were obtained from 50 adult horses. The biomechanical parameters were measured using a MyotonPRO device in triple-scan mode. Measurements were recorded on both sides, with the horses standing square. The analysis focused on whether there were differences between the sides and correlations between the variables. Unassigned: The mean muscle frequency (hertz) was...
Derivatisation-free FIA-MS/MS assay for rapid simultaneous screening of atypical myopathy biomarkers and acylcarnitine profiles from dried blood spots. Atypical myopathy (AM) is a frequently fatal, toxin-induced disease in horses, characterised by acute progression and mortality rates exceeding 70% within 72 h. The disease is caused by ingestion of seedlings or seeds of certain maple species that contain the toxins hypoglycin A (HGA) and methylenecyclopropylglycine (MCPrG). Early diagnosis depends on reliable detection of these compounds and their metabolites. However, most available LC-MS/MS assays are limited by derivatisation requirements and dependence on serum and urine matrices. Here, a rapid derivatisation-free flow injection analysis...
