Metabolites are small molecules involved in the metabolic processes within a horse's body. They are the intermediates and products of metabolism, encompassing a wide range of compounds, such as amino acids, lipids, carbohydrates, and nucleotides. These molecules play roles in energy production, growth, and cellular repair. The study of equine metabolites, often conducted through metabolomics, provides insights into the physiological and pathological states of horses. Changes in metabolite levels can indicate alterations in metabolic pathways, potentially reflecting health conditions or responses to environmental factors. This page compiles peer-reviewed research studies and scholarly articles that explore the identification, function, and diagnostic potential of metabolites in equine health.
Turner SE, Knych HK, Adams AA.To determine the pharmacokinetics, bioavailability, and pharmacological effects of cannabidiol (CBD) in senior horses. Methods: 8 university-owned senior horses. Methods: In this randomized, crossover study, horses were assigned to receive either a single oral dose of 2 mg/kg CBD in oil or a single IV dose of 0.1 mg/kg CBD in DMSO between August 10 and September 4, 2020. Blood samples were collected before and then 0.5, 1, 4, 8, 24, 48, 72, 96, 120, 144, 168, 192, 216, 240, and 264 hours after CBD administration. Serum biochemical analyses and CBCs were performed. Plasma concentrations of CBD ...
Göktaş EF, Kabil E, Yatanaslan L, Güneş E, Dirikolu L.Caffeine is a naturally occurring alkaloid and it is metabolized to paraxanthine, theophylline and theobromine. Analysis of caffeine and its metabolites is challenging since the metabolites theophylline and paraxanthine generate similar product and precursor ions. In this study, a new method was developed for the simultaneous analysis of caffeine, paraxanthine, theobromine and theophylline in horse urine using gas chromatography-mass spectrometry (GC-MS). Urine samples were treated using solid-phase extraction followed by the elution with dichloromethane-isopropanol (90:10) after the pH was ad...
Hu D, Tang Y, Wang C, Qi Y, Ente M, Li X, Zhang D, Li K, Chu H.The microbiota and its metabolites play an important role in regulating the host metabolism and immunity. However, the underlying mechanism is still not well studied. Thus, we conducted the LC-MS/MS analysis and RNA-seq analysis on with and without horse botfly infestation to determine the metabolites produced by intestinal microbiota in feces and differentially expressed genes (DEGs) related to the immune response in blood and attempted to link them together. The results showed that parasite infection could change the composition of microbial metabolites. These identified metabolites could b...
Philip M, Karakka Kal AK, Subhahar MB, Karatt TK, Mathew B, Koshy SA.It is important to remember that performance-enhancing agents such as non-peptide growth hormone secretagogues present a significant risk of abuse. Ibutamoren (MK-0677) is a potent, long-acting, selective non-peptide growth hormone secretagogue that can be taken orally. Methods: This study examines ibutamoren and its metabolites in thoroughbred horses after oral administration. Liquid chromatography/high-resolution mass spectrometry was used to determine the probable structures of the detected metabolites. Results: In this study, 22 metabolites of ibutamoren were identified (17 phase I and 5 p...
Li VL, He Y, Contrepois K, Liu H, Kim JT, Wiggenhorn AL, Tanzo JT, Tung AS, Lyu X, Zushin PH, Jansen RS, Michael B, Loh KY, Yang AC, Carl CS....Exercise confers protection against obesity, type 2 diabetes and other cardiometabolic diseases1-5. However, the molecular and cellular mechanisms that mediate the metabolic benefits of physical activity remain unclear6. Here we show that exercise stimulates the production of N-lactoyl-phenylalanine (Lac-Phe), a blood-borne signalling metabolite that suppresses feeding and obesity. The biosynthesis of Lac-Phe from lactate and phenylalanine occurs in CNDP2+ cells, including macrophages, monocytes and other immune and epithelial cells localized to diverse organs. In diet-induced obese mice,...
Pollard CL, Gibb Z, Clulow J, Ruiz A, Sheridan A, Bahrami M, Swegen A, Grupen CG.A deficiency in NAD has previously been linked with increased occurrences of congenital abnormalities and embryonic death in humans and mice. Early embryonic death is a major factor involved in pregnancy loss in mares, and very little is known regarding the NAD requirements for optimum reproductive function in horses. The aim of this study was to determine the effect of supplementing the diet of mares with nicotinic acid (NA) on the composition of NAD metabolites in the blood and follicular fluid. Vehicle alone or NA (3 g per os) were administered to seven mares over a minimum of 3 consecutive...
Deng L, Yang Y, Li Z, Li J, Zhu Y, Meng Q, Liu J, Wang X.Despite mare's milk being highly nutritious and beneficial to human health, the effects of different dietary regimens on the lipid profile of mare's milk remain to be completely understood. Herein we employed targeted multiple reaction monitoring-based lipidomics with ultra-high performance liquid chromatography-quadrupole time-of-flight-mass spectrometry to compare the milk lipid profiles of mares fed: pasture grass (P), corn stover and concentrate (H), or cornsilage and concentrate (S). Overall, 461 lipids belonging to 17 subclasses were identified; 261 significantly different lipids were id...
Haughan JE, Missanelli JR, You Y, Stefanovski D, Soma LR, Robinson MA.Glaucine, an aporphine alkaloid with anti-tussive, anti-inflammatory, and anti-nociceptive properties, has been identified in post-race samples from racehorses. To investigate pharmacokinetics of glaucine in horses, a three-way crossover study of intravenous and oral glaucine (0.1 mg/kg) and orally administered tulip poplar shavings (50 g shavings = 0.001 mg/kg glaucine) was performed in six horses. A two-compartment model best described IV administration with alpha ( ) and beta ( ) half-life lives of 0.3 (0.1-0.7) and 3.1 (2.4-7.8) h, respectively. The area under the curve ( ) was ...
Elbourne M, Cawley A, Stanley S, Bowen C, Fu S.Equine urine analysis has evolved over time to detect thousands of urinary compounds for doping control in the horse racing industry. The longitudinal assessment of 3-methoxytyramine to tyramine ratio (3-MT/T) values in equine urine by GC-MS profiling was investigated to support the Racing NSW Equine Biological Passport (EBP) for detection of dopaminergic manipulation in racehorses. This involved comparison of routine urine samples to administration studies of Sinemet, a common Parkinson's disease medication containing levodopa. Using an endogenous reference compound (ERC) in a urinary ratio e...
Cutler C, Viljanto M, Taylor P, Habershon-Butcher J, Van Eenoo P.Ibutamoren mesylate, or MK-0677, is an orally active, nonpeptide growth hormone secretagogue that has been developed to stimulate excretion of endogenous growth hormone. It has been evaluated for the treatment of a range of clinical conditions but is not available therapeutically. Nonetheless, MK-0677 is widely available to purchase online, sold as 'supplement' products. The mode of action and relative ease of purchase make MK-0677 a potential threat with regard to sports doping. The aim of this study was to investigate the metabolism of MK0677 in the horse following in vitro incubation and or...
Ishii H, Leung GN, Yamashita S, Nagata SI, Kushiro A, Sakai S, Toju K, Okada J, Kawasaki K, Kusano K, Kijima-Suda I.The use of nicotine stimulants in horses is generally banned in horse racing and equestrian sports-accidental consumption of tobacco products is one of the possible causes of nicotine exposure in horses. The authors recently reported a comprehensive metabolic study of nicotine in equines, differentiating between nicotine exposure and sample contamination by means of a nicotine biomarker trans-3'-hydroxycotinine. To identify potential biomarkers for the differentiation of genuine nicotine administration and consumption of tobacco products, tobacco leaves (equivalent to 250 mg of nicotine) wer...
Fitzgerald CCJ, Hedman R, Uduwela DR, Paszerbovics B, Carroll AJ, Neeman T, Cawley A, Brooker L, McLeod MD.The study of urinary phase II sulfate metabolites is central to understanding the role and fate of endogenous and exogenous compounds in biological systems. This study describes a new workflow for the untargeted metabolic profiling of sulfated metabolites in a urine matrix. Analysis was performed using ultra-high-performance liquid chromatography-high resolution tandem mass spectrometry (UHPLC-HRMS/MS) with data dependent acquisition (DDA) coupled to an automated script-based data processing pipeline and differential metabolite level analysis. Sulfates were identified through -means clustering...
Kozhakhmetov S, Babenko D, Kozhakhmetova S, Tuyakova A, Nurgaziyev M, Nurgozhina A, Muhanbetganov N, Chulenbayeva L, Sergazy S, Gulyayev A, Aljofan M....Ulcerative colitis is an inflammatory bowel disease that forms ulcerations in the mucous membrane of the colon and rectum, in which gut microbiota plays a pivotal role in its pathogenesis. Agents modulating microbial dysbiosis caused by colitis can help in the remission of this disease. The current study describes the potential therapeutic effects of active metabolites from Lactobacillus rhamnosus and mare's milk which have potential therapeutic values on the intestinal microbiota and proinflammatory cytokines. The analysis of the V1-V3 16S rDNA site revealed significant changes in the intesti...
Fernández-Hernández P, García-Marín LJ, Bragado MJ, Domingo A, González-Fernández L, Macías-García B.In the horse, a repeatable protocol for in vitro fertilization has not been developed, possibly due to incomplete sperm capacitation. We have previously identified the metabolites present in equine oviductal fluid (OF). We aimed to test the effects of different metabolites found in equine oviductal fluid on quality parameters of frozen-thawed spermatozoa. Different concentrations of myoinositol (5-25 mM), lactate (6-60 mM), glycine (0.1-5 mM), β-alanine (1-6 mM), and histamine (0.05-0.4 mM) were added independently to modified Whitten's medium (pH = 7.25). Thawed equine spermatozoa (three s...
Cutler C, Viljanto M, Taylor P, Hincks P, Biddle S, Van Eenoo P.Selective androgen receptor modulators, SARMs, are a large class of compounds developed to provide therapeutic anabolic effects with minimal androgenic side effects. A wide range of these compounds are available to purchase online and thus provide the potential for abuse in sports. Knowledge of the metabolism of these compounds is essential to aid their detection in doping control samples. In vitro models allow a quick, cost-effective response where administration studies are yet to be carried out. In this study, the equine phase I metabolism of the non-steroidal SARMs GSK2881078, LGD-2226, LG...
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...
Mathis D, Sass JO, Graubner C, Schoster A.Atypical myopathy (AM), an acquired multiple acyl-CoA dehydrogenase deficiency (MADD) in horses, induce changes in mitochondrial metabolism. Only few veterinary laboratories offer diagnostic testing for this disease. Inborn and acquired MADD exist in humans, therefore determination of organic acids (OA) in urine and acylcarnitines (AC) in blood by assays available in medical laboratories can serve as AM diagnostics. The evolution of OA and AC profiles in surviving horses is unreported. Methods: AC profiles using electrospray ionization tandem mass spectrometry (ESI-MS/MS) and OA in urine using...
Delcourt V, Garcia P, Pottier I, Mansoibou N, Bache N, Glavieux Y, Chabot B, Perot I, André F, Loup B, Barnabé A, Popot MA, Bailly-Chouriberry L.Synthetic androgenic anabolic steroids (AAS) are banned compounds and considered as major threats by both racing and sports international authorities. Hence, doping control laboratories are continually looking into analytical improvements to increase their detection capabilities, notably by means of emerging technologies. To enhance analytical performances for the detection of synthetic AAS such as stanozolol, specific chromatographic procedures have been developed using recent quaternary liquid chromatography technology originally designed for high-throughput standardized proteomics connected...
Waitt Wolker LH, Veltri CA, Pearman K, Lozoya M, Norris JW.This study aimed to investigate pharmacokinetics of fluoxetine in horses and validate a method for liquid chromatography mass spectrometry analysis of serum levels. Fluoxetine pharmacokinetics were determined using 10 healthy, adult horses. Fluoxetine pharmacokinetics following a single oral dose (0.25 mg/kg) were determined using blood samples collected prior to and at several time points over 7 days following administration. Serum concentrations of fluoxetine and its bioactive metabolite norfluoxetine were measured using liquid chromatography coupled to an accurate mass/high-resolution ma...
Paßlack N, van Bömmel-Wegmann S, Vahjen W, Zentek J.Zinc supplements are often used in equine nutrition to support skin and hoof quality or the immune function. However, no data on the effects of dietary zinc on the intestinal microbiota of horses and ponies are available so far. In the present study, varying dietary zinc concentrations (maintenance (4 mg/kg BW/day), 120 mg/kg dry matter (DM)/day and 240 mg/kg DM/day) were achieved by the supplementation of either zinc chloride hydroxide or zinc methionine (six treatment periods of 4 weeks each). Eight healthy adult ponies and two healthy adult horses were included, and faecal samples were coll...
Ishii H, Shibuya M, So YM, Wong JKY, Ho ENM, Kusano K, Sone Y, Kamiya T, Wakuno A, Ito H, Miyata K, Yamada M, Leung GN.IOX4 is a hypoxia-inducible factor prolyl hydroxylase (HIF-PHD) inhibitor, which was developed for the treatment of anemia by exerting hematopoietic effects. The administration of HIF-PHD inhibitors such as IOX4 to horses is strictly prohibited by the International Federation of Horseracing Authorities and the Fédération Équestre Internationale. To the best of our knowledge, this is the first comprehensive metabolic study of IOX4 in horse plasma and urine after a nasoesophageal administration of IOX4 (500 mg/day, 3 days). A total of four metabolites (three mono-hydroxylated IOX4 and one ...
Stanley S, Deng D, Van den Berg K, Foo HC.The abuse of performance-enhancing catecholamine-based stimulants, such as levodopa, is controlled in horse racing through the application of a regulatory threshold for the common major metabolite. However, catechol-O-methyltransferase (COMT) enzyme inhibitors can be used to restrict the catalysis of the stimulant, and so the concurrent administration of both substances would be a viable strategy to enhance racing performance while removing the risk of exceeding the threshold. A 200 mg dose of nitecapone, a COMT inhibitor, was administered to a Thoroughbred horse, and we have analysed the bl...
Kędzierski W, Sadok I, Kowalik S, Janczarek I, Staniszewska M.Tryptophan (Trp) is an essential amino acid which metabolises via the kynurenine pathway to generate a number of bioactive substances referred to as kynurenines. Among those are 3-hydroxykynurenine (3-HKyn) and quinolinic acid, which are neurotoxic, as well as kynurenic acid (Kyna) and xanthurenic acid (XA), which, similarly to nicotinamide (NAm), show neuroprotective and anti-depressive effects. Routine exercise is known to modulate Trp metabolism in skeletal muscle and is thus believed to reduce the risk of depressive states in humans and laboratory animals. Analogously, it was hypothesised ...
Paine SW, Aldurdunji M, Garth-Greeves A, Muir T, Hincks PR.Hydroxyzine and cetirizine are first- and second-generation oral antihistamine drugs, respectively, used to treat allergic reactions in horses. Cetirizine is also a metabolite of hydroxyzine, which may lead to complexities in regulating their use in equine sporting events. The aim of the research was to be able to provide detection times (DT) from pharmacokinetic studies in thoroughbred horses to better inform trainers, and their veterinary surgeons, prescribing these substances for treatment of Thoroughbred racehorses. Six and two horses were given 9 repeated administrations of hydroxyzine HC...
Wouters CP, Toquet MP, Renaud B, François AC, Fortier-Guillaume J, Marcillaud-Pitel C, Boemer F, De Tullio P, Richard EA, Votion DM.Atypical myopathy (AM) is a severe rhabdomyolysis syndrome that occurs in grazing horses. Despite the presence of toxins in their blood, all horses from the same pasture are not prone to display clinical signs of AM. The objective of this study was to compare the blood metabolomic profiles of horses with AM clinical signs with those of healthy co-grazing (Co-G) horses. To do so, plasma samples from 5 AM horses and 11 Co-G horses were investigated using untargeted metabolomics. Metabolomic data were evaluated using unsupervised, supervised, and pathway analyses. Unsupervised principal component...
Yılmaz İ, Maraş Z, Uğur Y, Erkan Özgür M, Durmaz M, İbrahim Ulusoy H, Erdoğan S.Metamizole sodium (MT) is an analgesic and antipyretic drug molecule used in humans, horses, cattle, swine, and dogs. Metamizole rapidly hydrolyzes and turns into methylamino antipyrine (MAA), an active primary metabolite of MT. The present study aims to determine the pharmacokinetic (PK) profiles of MT metabolites after intravenous (IV) and intramuscular (IM) administration into sex of Arabian horses (Equus ferus caballus) using a cross-over study design. The plasma samples were extracted by solid-phase extraction (SPE) method, and plasma concentrations of MT metabolites were analyzed by high...
Zhu Y, Wang X, Liu B, Yi Z, Zhao Y, Deng L, Holyoak R, Li J.Silage is fed to horses in China and other areas in the world, however, knowledge about the impact of feeding silage on horse health is still limited. In the current study, 12 horses were assigned into two groups and fed ryegrass silage and ryegrass hay, respectively, for 8 weeks. High-throughput sequencing was applied to analyze fecal microbiota, while liquid chromatography-tandem mass spectrometry (LC-MS/MS) based metabolomics technique was used for blood metabolite profile to investigate the influence of feeding ryegrass silage (group S) compared to feeding ryegrass hay (group H) on equine ...
Mycotoxin contaminated feed has been associated with colic of horses caused by intestinal disorders. Whether such disease conditions alter the intestinal toxin metabolism and transfer across a compromised mucosal barrier is unknown. A screening approach was used to relate blood residue levels of DON, ZEN and their metabolites to the status of the horses (sick vs. healthy). A total of 55 clinically healthy horses from 6 different farms with varying feeding background served as control for sick horses (N = 102) hospitalized due to colic. ZEN, alpha-zearalenol (ZEL), beta-ZEL and DON were detecta...
de Macêdo LB, Mouta AN, Araújo-Silva G, Urizar JTP, de Paula VV.Metamizole (MT), also known as dipyrone, is an analgesic and antipyretic drug labeled for use in humans and domestic animals in some countries. As with other drugs, the administration of MT in donkeys is based on studies carried out with horses. In the present report, we aimed to determine the pharmacokinetics of the two main metamizole active metabolites (N-methyl-4-aminoantipyrine [MAA] and 4-aminoantipyrine [AA]) following 10 (M ) and 25 mg/kg (M ) IV metamizole doses in Northeast Brazilian donkeys (n = 10). Blood was collected at predetermined times within over 48 h; MAA and AA plasma ...
Peña FJ, Ortiz-Rodríguez JM, Gaitskell-Phillips GL, Gil MC, Ortega-Ferrusola C, Martín-Cano FE.An overview of the sperm metabolism is presented; using the stallion as a model we review glycolysis, Krebs Cycle and oxidative phosphorylation, paying special attention to the interactions among them. In addition, metabolism implies a series of coordinated oxidation-reduction reactions and in the course of these reactions reactive oxygen species (ROS) and reactive oxoaldehydes are produced ; the electron transport chain (ETC) in the mitochondria is the main source of the anion superoxide and hydrogen peroxide, while glycolysis produces 2-oxoaldehydes such as methylglyoxal as byproducts; due t...
Gollnick PD, Bertocci LA, Kelso TB, Witt EH, Hodgson DR.The effect of high-intensity exercise on the respiratory capacity of skeletal muscle was studied in horses which ran five 600-m bouts on a track with 2 min of rest between exercise bouts, or once to fatigue on a treadmill at an intensity that elicited the maximal oxygen uptake. Venous blood and biopsy samples of the middle gluteal muscle were collected at rest, after each exercise bout, and 30 and 60 min post-exercise. Blood samples were analyzed for lactate concentration and pH and muscle samples for metabolites, pH, and respiratory capacity. Venous blood and muscle pH declined to 6.91 +/- 0....
Scarth JP, Teale P, Kuuranne T.A detailed understanding of equine drug metabolism is important for detection of drug abuse in horseracing and also in veterinary drug development and practice. To date, however, no comprehensive review of equine drug metabolism has been published. The majority of literature regarding equine drug metabolite profiles is derived from sports drug detection research and is generally targeted at detecting marker metabolites of drug abuse. However, the bulk of the literature on equine drug metabolism enzymology is derived from veterinary studies aimed at determining the molecular basis of metabolism...
Letendre LT, Tessman RK, McClure SR, Kvaternick VJ, Fischer JB, Hanson PD.To determine pharmacokinetic parameters and variables, firocoxib concentrations in urine and plasma, urine-to-plasma ratios, and the urine depletion profile of firocoxib and to evaluate whether the pharmacokinetic behavior of firocoxib was governed by linear processes after multiple doses of firocoxib were administered IV and orally. Methods: 6 healthy female horses (5 Paint horses and 1 Quarter Horse) in experiment 1 and 12 healthy male and female horses in experiment 2. Methods: In experiment 1, 6 horses were orally administered firocoxib paste once daily for 12 consecutive days, and plasma ...
Silver M, Fowden AL, Knox J, Ousey JC, Franco R, Rossdale PD.The effects of insulin-induced hypoglycaemia on plasma catecholamines, cortisol and metabolites have been examined in newborn and 7-14-day-old foals. The fall in plasma glucose elicited by the highest dose of insulin (1.0 i.u./kg) given to the neonates was slower in onset and less severe in effect than 0.5 i.u./kg in the older foals. There was a significant inverse correlation between the concentrations of glucose and adrenaline (but not noradrenaline) in plasma once the glucose level had fallen below 2 mmol/l; the adrenergic response to hypoglycaemia was greater in the 7-14-day-old foals than...
Shilo Y, Britzi M, Eytan B, Lifschitz T, Soback S, Steinman A.Tramadol is a centrally acting analgesic drug that has been used clinically for the last two decades to treat moderate to moderately severe pain in humans. The present study investigated tramadol administration in horses by intravenous, intramuscular, oral as immediate-release and oral as sustained-release dosage-form routes. Seven horses were used in a four-way crossover study design in which racemic tramadol was administered at 2 mg/kg by each route of administration. Altogether, 23 blood samples were collected between 0 and 2880 min. The concentration of tramadol and its M1 metabolite were ...
Ho EN, Leung DK, Leung GN, Wan TS, Wong AS, Wong CH, Soma LR, Rudy JA, Uboh C, Sams R.Administration studies of levamisole in horses were carried out using two different levamisole preparations, namely, levamisole hydrochloride oral bolus and levamisole phosphate injectable solution. These preparations were analysed in detail for the presence of aminorex-like impurities. Both levamisole preparations were found to contain 1-(2-mercaptoethyl)-4-phenyl-2-imidazolidinone (I) and 4-phenyl-2-imidazolidinone (II) as degradation impurities, but neither aminorex nor rexamino was detected in these preparations. After the administration of these preparations to horses, aminorex, rexamino,...
Liu Y, Siard M, Adams A, Keowen ML, Miller TK, Garza F, Andrews FM, Seeram NP.The human health benefits attributed to turmeric/curcumin spice has resulted in its wide utilization as a dietary supplement for companion pets and other animals including horses. While the quantification of free curcuminoids (curcumin, demethoxycurcumin, bisdemethoxycurcumin) and their phase-2 metabolites (curcumin-O-sulfate, curcumin-O-glucuronide) have been extensively investigated in human and rodent biological samples (primarily plasma and serum), there is lack of similar data for horses. Herein, we report a validated LC-ESI-MS/MS method for the simultaneous quantification of the aforemen...
Laus F, Gialletti R, Bazzano M, Laghi L, Dini F, Marchegiani A.Osteoarthritis (OA) is a common cause of lameness in sport horses with a significant economic impact. The prevention of OA is crucial since no effective treatment is available. This study aimed to apply untargeted metabolomic analysis to investigate the differences in synovial fluid (SF) composition between healthy and OA-affected joints in horses. SF collected from healthy (n.8) and OA (n.11) horses was analyzed using H-NMR analysis. Metabolomic analysis allowed 55 different metabolites to be identified and quantified in SF samples. Nineteen metabolites were found to be differently concentrat...
Jacob SI, Murray KJ, Rendahl AK, Geor RJ, Schultz NE, McCue ME.Metabolomics, the study of small-molecule metabolites, has increased understanding of human metabolic diseases, but has not been used to study equine metabolic syndrome (EMS). Objective: (1) To examine the serum metabolome of Welsh Ponies with and without insulin dysregulation before and during an oral sugar test (OST). (2) To identify differences in metabolites in ponies with insulin dysregulation, obesity, or history of laminitis. Methods: Twenty Welsh Ponies (mean ± SD; 13.8 ± 9.0 years) classified as non-insulin dysregulated [CON] (n = 10, insulin 60 mU/L) at 75 minutes ...
Walter J, Huwiler F, Fortes C, Grossmann J, Roschitzki B, Hu J, Naegeli H, Laczko E, Bleul U.Maturation of oocytes under in vitro conditions (IVM) results in impaired developmental competence compared to oocytes matured in vivo. As oocytes are closely coupled to their cumulus complex, elucidating aberrations in cumulus metabolism in vitro is important to bridge the gap towards more physiological maturation conditions. The aim of this study was to analyze the equine "cumulome" in a novel combination of proteomic (nano-HPLC MS/MS) and metabolomic (UPLC-nanoESI-MS) profiling of single cumulus complexes of metaphase II oocytes matured either in vivo (n = 8) or in vitro (n = 7). Re...
González-Fernández L, Sánchez-Calabuig MJ, Calle-Guisado V, García-Marín LJ, Bragado MJ, Fernández-Hernández P, Gutiérrez-Adán A....A repeatable protocol for equine in vitro fertilization (IVF) has remained elusive. This is likely, in part, due to suboptimal composition of capacitation or IVF media that are currently in use. Hence, we aimed to analyse the metabolome of equine oviductal fluid (OF) at the pre- (PRE) and immediate post-ovulatory (PST) stages using proton magnetic resonance spectroscopy (H NMR). Oviductal fluid from eight PRE and six PST mares were used to prepare a total of five samples per group. A total of 18 metabolites were identified. The five metabolites with the highest concentrations in the OF sample...
Weidolf LO, Lee ED, Henion JD.Sulfoconjugated anabolic steroids were separated by micro-bore high-performance liquid chromatography. The eluent was introduced into the atmospheric pressure ion source of the triple-quadrupole mass spectrometer via an ion spray liquid chromatograph/mass spectrometer interface operated in the negative ion mode. The limit of detection was 10 pg on-column by selected ion monitoring of the molecular ion and the response increased linearly over a concentration range of 2.4 orders of magnitude. Following work-up by a liquid-solid extraction procedure of equine urine samples, full-scan daughter ion...
Mozūraitis R, Būda V, Kutra J, Borg-Karlson AK.Urine samples from 14 mares, belonging to five breeds, were collected at estrus and diestrus to search for estrous specific volatile compounds which could be used to determine ovulation time. Around 150 volatiles were collected from urine head-space samples by solid phase micro extraction technique, and analyses were conducted by gas chromatography-mass spectrometry methods. Comparison of chromatographic profiles of volatile substances revealed that concentrations of m- and p-cresols were significantly greater during estrus while diethylphthalate was more abundant at diestrus. Monitoring of m-...
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...
Flauger B, Krueger K, Gerhards H, Möstl E.Glucocorticoids or their metabolites can be measured in several body fluids or excreta, including plasma, saliva, urine and faeces. In recent years the measurement of glucocorticoid metabolites (GCMs) in faeces has gained increasing attention, because of its suitability for wild populations. In horses, however, the group-specific enzyme immunoassay described so far has a limited practicability due to its complex extraction procedure. Therefore, we tested the applicability of other enzyme immunoassays for glucocorticoid metabolites. The present study clearly proved that an enzyme immunoassay (E...
Fitzgerald CCJ, Hedman R, Uduwela DR, Paszerbovics B, Carroll AJ, Neeman T, Cawley A, Brooker L, McLeod MD.The study of urinary phase II sulfate metabolites is central to understanding the role and fate of endogenous and exogenous compounds in biological systems. This study describes a new workflow for the untargeted metabolic profiling of sulfated metabolites in a urine matrix. Analysis was performed using ultra-high-performance liquid chromatography-high resolution tandem mass spectrometry (UHPLC-HRMS/MS) with data dependent acquisition (DDA) coupled to an automated script-based data processing pipeline and differential metabolite level analysis. Sulfates were identified through -means clustering...
Kieken F, Pinel G, Antignac JP, Monteau F, Christelle Paris A, Popot MA, Bonnaire Y, Le Bizec B.Despite the worldwide existing regulation banning the use of the recombinant equine growth hormone (reGH) as growth promoter, it is suspected to be used in horseracing to improve performances. Various analytical methods previously developed to screen for its misuse have encountered some limitations in terms of detection timeframes, in particular during the first days following reGH administration. A novel strategy involving the characterization of global metabolomic fingerprints in urine samples of non-treated and reGH-treated horses by liquid chromatography-electrospray-high-resolution mass s...
Theurillat R, Knobloch M, Schmitz A, Lassahn PG, Mevissen M, Thormann W.CE with multiple isomer sulfated beta-CD as the chiral selector was assessed for the simultaneous analysis of the enantiomers of ketamine and metabolites in extracts of equine plasma and urine. Different lots of the commercial chiral selector provided significant changes in enantiomeric ketamine separability, a fact that can be related to the manufacturing variability. A mixture of two lots was found to provide high-resolution separations and interference-free detection of the enantiomers of ketamine, norketamine, dehydronorketamine, and an incompletely identified hydroxylated metabolite of no...
Karlíková R, Široká J, Jahn P, Friedecký D, Gardlo A, Janečková H, Hrdinová F, Drábková Z, Adam T.Atypical myopathy (AM) is a potentially fatal disease of grazing horses. It is reportedly caused by the ingestion of sycamore seeds containing toxic hypoglycin A. In order to study metabolic changes, serum and urine samples from nine horses with atypical myopathy and 12 control samples from clinically healthy horses were collected and then analysed using a high-performance liquid chromatography coupled with tandem mass spectrometry; serum metabolic profiles as the disease progressed were also studied. Metabolic data were evaluated using unsupervised and supervised multivariate analyses. Signif...
Bazzano M, Laghi L, Zhu C, Lotito E, Sgariglia S, Tesei B, Laus F.In the present study, data related to the metabolomics of saliva and serum in trained standardbred horses are provided for the first time. Metabolomic analysis allows to analyze all the metabolites within selected biofluids, providing a better understanding of biochemistry modifications related to exercise. On the basis of the current advances observed in metabolomic research on human athletes, we aimed to investigate the metabolites' profile of serum and saliva samples collected from healthy standardbred horses and the relationship with physical exercise. Twelve trained standardbred horses we...
Dunnett M, Harris RC, Orme CE.A simple, robust and reproducible analytical method for the determination of phosphocreatine (PCr), creatine (Cr) and creatinine (Cn) in equine skeletal muscle is presented. The technique used isocratic reverse-phase ion-pairing high-performance liquid chromatography. Neutralized perchloric acid extracts of equine muscle biopsies were analysed and the values obtained were compared with determinations from an established enzymic procedure. Good resolution of all three metabolites was achieved within a retention time of less than 11 min. Linearity for each metabolite within the concentration ran...
Lankveld DP, Driessen B, Soma LR, Moate PJ, Rudy J, Uboh CE, van Dijk P, Hellebrekers LJ.Ketamine (KET) possesses analgesic and anti-inflammatory activity at sub-anesthetic doses, suggesting a benefit of long-term KET treatment in horses suffering from pain, inflammatory tissue injury and/or endotoxemia. However, data describing the pharmacodynamic effects and safety of constant rate infusion (CRI) of KET and its pharmacokinetic profile in nonpremedicated horses are missing. Therefore, we administered to six healthy horses a CRI of 1.5 mg/kg/h KET over 320 min following initial drug loading. Cardiopulmonary parameters, arterial blood gases, glucose, lactate, cortisol, insulin, non...
Grippa E, Santini L, Castellano G, Gatto MT, Leone MG, Saso L.Ethyl acetate extracts of equine serum, containing 0-5 microg/ml of hydrocortisone (HYD), dexamethasone (DEX), oxyphenbutazone (OPB), indomethacin (IND), phenylbutazone (PB) and probenecid as internal standard, were evaporated with nitrogen, resuspended in methanol and analyzed by HPLC, using a C-18 column equilibrated with 51:49 acetonitrile-water, 0.1% trifluoroacetic acid, at 1 ml/min. The eluate was monitored at 254 nm. The selectivity (inter-assay C.V.<4%), sensitivity (limits of quantitation of 0.25 microg/ml for HYD, DEX and IND, 0.5 microg/ml for PB and 1 microg/ml for OPB, despite ...
Steelman SM, Johnson P, Jackson A, Schulze J, Chowdhary BP.Acute laminitis is an inflammatory disease of the equine foot that often occurs secondarily to sepsis or systemic inflammation associated with gastrointestinal disease. It has been suggested that laminitis is similar to multiple organ dysfunction syndrome in humans, although in horses the weight-bearing laminar epithelium of the foot appears to be the tissue most sensitive to insult and the first "organ" to fail. Metabolomics performed on serum samples collected before (Con) and after (Lmn) experimental induction of gastrointestinal-associated sepsis in six horses detected 1,177 metabolites of...
Le Moyec L, Robert C, Triba MN, Bouchemal N, Mach N, Rivière J, Zalachas-Rebours E, Barrey E.Endurance racing places high demands on energy metabolism pathways. Metabolomics can be used to investigate biochemical responses to endurance exercise in humans, laboratory animals, and horses. Although endurance horses have previously been assessed in the field (i.e., during races) using broad-window Nuclear Magnetic Resonance metabolomics, these studies included several different race locations, race distances, age classes, and race statuses (finisher or elimination). The present NMR metabolomics study focused on 40 endurance horses racing in three race categories over 90, 120, or 160 km. T...
Shepherd ML, Ponder MA, Burk AO, Milton SC, Swecker WS.The purpose of the present study was to compare digestibility of grass hay, faecal and plasma volatile fatty acid (VFA) concentrations, and faecal bacterial abundance in overweight and moderate-condition mares. Five overweight adult mixed-breed mares and five adult mixed-breed mares in moderate condition were housed individually and limit-fed orchard grass (Dactylis glomerata) hay at 20 g/kg body weight (as fed) daily for 14 d. Forage DM and fibre digestibility were determined using AOAC methods; digestible energy was measured using bomb calorimetry; plasma and faecal VFA concentrations were ...
Wang T, Zeng Y, Ma C, Meng J, Wang J, Ren W, Wang C, Yuan X, Yang X, Yao X.In this study, the plasma non-targeted metabolomics of Yili horses were characterized before and after exercise on tracks that differed in surface hardness to better understand exercise-related biochemical changes. Blood samples were obtained from eight trained Yili horses before and immediately after exercise. Samples were used for metabolomic analysis by ultra-performance liquid chromatography-Q-EXACTIVE mass spectrometry. In total, 938 significantly different metabolites involving sugar, lipid, and amino acid metabolism were detected in the plasma, with significant increases in glucose, glu...
Higgins AJ, Lees P, Wright JA.In a series of experiments to examine equine inflammatory exudates for the presence of metabolites of arachidonic acid, including prostaglandin E2 (PGE2), a model for the induction and collection of exudates in ponies has been developed. Multiperforated polypropylene practice golf balls implanted subcutaneously in the mid-neck region were well tolerated and proved to be the most successful model. One such cage was implanted in the neck of each of seven ponies. Inflammatory exudates were induced by injecting 3.0 or 0.5 ml carrageenin into the cages and aspirates collected between three and 48 h...
Knych HK, Arthur RM, McKemie DS, Seminoff K, Hamamoto-Hardman B, Kass PH.Phenylbutazone (PBZ) is a potent mon-steroidal anti-inflammatory drug used commonly in performance horses. The objectives of the current study were to describe blood and urine concentrations and the pharmacokinetics of PBZ and its metabolites following intravenous (IV) and oral administration and to describe the duration of pharmacodynamic effect. To that end, 17 horses received an IV administration and 18 horses an oral administration of 2 g of PBZ. Blood and urine samples were collected prior to and for up to 96 hours post drug administration. Whole blood samples were collected at various t...
Scarth JP, Spencer HA, Timbers SE, Hudson SC, Hillyer LL.The detection of drug abuse in horseracing often requires knowledge of drug metabolism, especially if urine is the matrix of choice. In this study, equine liver/lung microsomes/S9 tissue fractions were used to study the phase I metabolism of eight drugs of relevance to equine drug surveillance (acepromazine, azaperone, celecoxib, fentanyl, fluphenazine, mepivacaine, methylphenidate and tripelennamine). In vitro samples were analyzed qualitatively alongside samples originating from in vivo administrations using LC-MS on a high resolution accurate mass Thermo Orbitrap Discovery instrument and by...
