Topic:Metabolites
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.
Temporal relationships of a pulse of prolactin (PRL) to a pulse of a metabolite of PGF2α in mares. Hourly blood samples were collected from 10 mares during 24 h of each of the preluteolytic, luteolytic, and postluteolytic periods. The autocorrelation function of the R program was used to detect pulse rhythmicity, and the intra-assay CV was used to locate and characterize pulses of prolactin (PRL) and a metabolite of prostaglandin F2α (PGFM). Rhythmicity of PRL and PGFM concentrations was detected in 67% and 89% of mares, respectively. Combined for the three periods (no difference among periods), the PRL pulses were 5.2±0.4 h (mean±SEM) at the base, 7.5±1.5 h between nadirs of adjacent p...
Alterations in microbiota and fermentation products in equine large intestine in response to dietary variation and intestinal disease. We aimed to determine the effects of variations in dietary composition on equine gut microbiota and their fermentation products, and proposed that dietary modifications profoundly affect microbial ecosystems and their metabolites. Bacterial communities within the large intestine of three groups of horses were compared using oligonucleotide-RNA hybridisation methodology. Each group consisting of six horses was maintained on (1) a grass-only diet, (2) a concentrate diet (i.e. supplemented with hydrolysable carbohydrates) and (3) a concentrate diet but horses were affected by simple colonic obstr...
Implications of urine F2-isoprostane metabolite concentration in horses with colic and its potential use as a predictor for surgical intervention. F2-isoprostanes have been used extensively to quantify lipid peroxidation in association with risk factors in various diseases. Horses with colic may have intestinal ischaemia and/or inflammation characterised by oxidative stress and increased production of isoprostanes. Objective: To gather preliminary data regarding the feasibility of using urine F2-isoprostanes and isoprostane metabolites as early screening tools for the presence of gastrointestinal disease requiring surgical intervention in horses and ultimately develop a stall-side test capable of identifying these horses as early as poss...
A targeted lipidomics approach to the study of eicosanoid release in synovial joints. Articular tissues are capable of producing a range of eicosanoid mediators, each of which has individual biological effects and may be affected by anti-inflammatory treatment. We set out to develop and evaluate a high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) approach for the simultaneous analysis of multiple eicosanoid lipid mediators in equine synovial fluid (SF), and to illustrate its use for investigation of the in vivo effects of non-steroidal anti-inflammatory drug (NSAID) treatment. Methods: Synovial fluid samples were obtained from normal joints of 6 adult...
Pharmacokinetics of tramadol and metabolites O-desmethyltramadol and N-desmethyltramadol in adult horses. To determine the pharmacokinetics of tramadol and its metabolites O-desmethyltramadol (ODT) and N-desmethyltramadol (NDT) in adult horses. Methods: 12 mixed-breed horses. Methods: Horses received tramadol IV (5 mg/kg, over 3 minutes) and orally (10 mg/kg) with a 6-day washout period in a randomized crossover design. Serum samples were collected over 48 hours. Serum tramadol, ODT, and NDT concentrations were measured via high-performance liquid chromatography and analyzed via noncompartmental analysis. Results: Maximum mean ± SEM serum concentrations after IV administration for tramadol, ODT, ...
Comparison of different liquid chromatography stationary phases in LC-HRMS metabolomics for the detection of recombinant growth hormone doping control. Growth hormone (GH) is a polypeptide suspected of being used in horse racing to speed up physical performances. Despite scientific advances in the recent years, the control of its administration remains difficult. In order to improve it, a metabolomics study through LC-high resolution mass spectrometry measurements was recently initiated to assess the metabolic perturbations caused by recombinant equine growth hormone administration. Few tens of ions not identified structurally were highlighted as compounds responsible for the modification of metabolic profiling observed in treated animals. Th...
Pharmacokinetics of oral terbinafine in horses and Greyhound dogs. The objective of the study was to assess the pharmacokinetics of terbinafine administered orally to horses and Greyhound dogs. A secondary objective was to assess terbinafine metabolites. Six healthy horses and six healthy Greyhound dogs were included in the pharmacokinetic data. The targeted dose of terbinafine was 20 and 30 mg/kg for horses and dogs, respectively. Blood was collected at predetermined intervals for the quantification of terbinafine concentrations with liquid chromatography and mass spectrometry. The half-life (geometric mean) was 8.1 and 8.6 h for horses and Greyhounds, respe...
Characterization of in vivo plasma metabolites of tepoxalin in horses using LC-MS-MS. Tepoxalin is a veterinary drug registered for use in the dog as a dual inhibitor (cyclooxygenase-5 lipoxygenase). In the horse, it predominantly triggers a strong cyclooxygenase inhibition; this bias seems to be due to the action of its metabolite(s). Among these, only the RWJ-20142 is well known, while to the best of our knowledge no information is available on the other metabolites produced in vivo. Hence, the identification of its main metabolic pathway is pivotal to better understand its clinical activity. A suitable high performance liquid chromatography method has been applied to liquid ...
Easy stereoselective synthesis of 5α-estrane-3β,17α-diol, the major metabolite of nandrolone in the horse. 5α-Estrane-3β,17α-diol is the major metabolite of nandrolone in horse urine. The presence of 5α-estrane-3β,17α-diol in female and gelding urines is prohibited by Racing Rules and its natural presence in male urine led regulation authorities to establish a concentration threshold of 45 ng/mL. This paper describes a rapid, simple and stereoselective synthesis of 5α-estrane-3β,17α-diol, providing horseracing laboratories with an essential reference material for their antidoping performance.
Nuclear magnetic resonance-based metabonomic study of early time point laminitis in an oligofructose-overload model. NMR-metabonomics is an unbiased evaluation method, which allows to comprehensively study changes of the equine metabolic profile in early time point laminitis. This might give insight into the early stages of disease development. Objective: To detect hitherto unknown changes in blood metabolites during the development of oligofructose-induced laminitis by comparing pre- and post induction blood samples. Methods: Prior to laminitis induction blood was sampled to establish control values. Post oligofructose administration (POA) blood was collected every 3 h for 24 h. One-dimensional (1) H-NMR sp...
In vitro metabolic studies using homogenized horse liver in place of horse liver microsomes. The study of the metabolism of drugs, in particular steroids, by both in vitro and in vivo methods has been carried out in the authors' laboratory for many years. For in vitro metabolic studies, the microsomal fraction isolated from horse liver is often used. However, the process of isolating liver microsomes is cumbersome and tedious. In addition, centrifugation at high speeds (over 100 000 g) may lead to loss of enzymes involved in phase I metabolism, which may account for the difference often observed between in vivo and in vitro results. We have therefore investigated the feasibility of us...
The use of in vitro technologies and high-resolution/accurate-mass LC-MS to screen for metabolites of ‘designer’ steroids in the equine. Detection of androgenic-anabolic steroid abuse in equine sports requires knowledge of the drug's metabolism in order to target appropriate metabolites, especially where urine is the matrix of choice. Studying 'designer' steroid metabolism is problematic since it is difficult to obtain ethical approval for in vivo metabolism studies due to a lack of toxicological data. In this study, the equine in vitro metabolism of eight steroids available for purchase on the Internet is reported; including androsta-1,4,6-triene-3,17-dione, 4-chloro,17α-methyl-androsta-1,4-diene-3,17β-diol, estra-4,9-diene-...
Analysis of bioactive eicosanoids in equine plasma by stable isotope dilution reversed-phase liquid chromatography/multiple reaction monitoring mass spectrometry. Oxidative metabolites of arachidonic acid (AA) are implicated in inflammation. Thus, we evaluated cycloxygenases (COXs) and lipoxygenases (LOs) mediated metabolism of AA to eicosanoids in equine plasma. Eicosanoids were extracted from plasma by two liquid-liquid extraction (LLE) steps; first was by chloroform/isopropanol and second by methyl-tert-butyl ether. For identification and quantification of 25 eicosanoids, a highly specific, selective and sensitive stable isotope dilution liquid chromatography (LC) multiple reaction monitoring (MRM) mass spectrometric (MS) method was developed. To avo...
Doping control analysis of insulin and its analogues in equine urine by liquid chromatography-tandem mass spectrometry. Insulin and its analogues have been banned in both human and equine sports owing to their potential for misuse. Insulin administration can increase muscle glycogen by utilising hyperinsulinaemic clamps prior to sports events or during the recovery phases, and increase muscle size by its chalonic action to inhibit protein breakdown. In order to control insulin abuse in equine sports, a method to effectively detect the use of insulins in horses is required. Besides the readily available human insulin and its synthetic analogues, structurally similar insulins from other species can also be used a...
Use of in vitro technologies to study phase II conjugation in equine sports drug surveillance. Within equine drug surveillance, there is significant interest in analyzing intact phase II conjugates of drugs in urine, but progress has been limited by a lack of reference material. Methods: In this study, in vitro techniques using equine liver fractions were employed to produce glucuronide and sulfate conjugates of stanozolol, 16β-hydroxystanozolol and nandrolone, the glucuronide conjugate of morphine and the glutathione metabolite of chlordinitrobenzene for the first time in equine sports drug surveillance. Results: The glucuronide conjugate of the synthetic progestagen altrenogest was a...
Pharmacokinetics of ceftiofur sodium and ceftiofur crystalline free acid in neonatal foals. Ceftiofur, a third generation cephalosporin, demonstrates in vitro efficacy against microorganisms isolated from septicemic neonatal foals. This pharmacokinetic study evaluated the intravenous and subcutaneous administration of ceftiofur sodium (5 mg/kg body weight; n = 6 per group) and subcutaneous administration of ceftiofur crystalline free acid (6.6 mg/kg body weight; n = 6) in healthy foals. Plasma ceftiofur- and desfuroylceftiofur-related metabolite concentrations were measured using high performance liquid chromatography following drug administration. Mean (±SD) noncompartmental pharma...
Drug metabolism in the horse: a review. 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...
Enantiomeric composition analysis of pranoprofen in equine plasma and urine by chiral liquid chromatography-tandem mass spectrometry in selected reaction monitoring mode. 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 ...
The use of in vitro technologies coupled with high resolution accurate mass LC-MS for studying drug metabolism in equine drug surveillance. 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...
Association of season and pasture grazing with blood hormone and metabolite concentrations in horses with presumed pituitary pars intermedia dysfunction. Pituitary pars intermedia dysfunction (PPID) is a risk factor for pasture-associated laminitis, which follows a seasonal pattern. Objective: Hormonal responses to season differ between PPID and unaffected horses. Methods: Seventeen horses aged 8-30 years (14 horses ≥ 20 years of age). Methods: Longitudinal observational study. Blood was collected monthly from August 2007 until July 2008 after pasture grazing and again after overnight stall confinement. Blood hormone and metabolite concentrations were measured and pasture grass samples were analyzed to determine carbohydrate content. Analysis...
Identification of etamiphylline and metabolites in equine plasma and urine by accurate mass and liquid chromatography/tandem mass spectrometry. Etamiphylline camsylate (Millophylline V) was administered intravenously to two horses at a dose of 2.8 mg/kg. Urine and blood samples were taken up to 32 h post administration. Unhydrolyzed plasma and urine was extracted using solid phase extraction (SPE). The identity of the parent drug and metabolites was confirmed using a linear ion trap mass spectrometer and accurate mass analysis on an orbitrap mass spectrometer. Desethyletamiphylline (molecular weight 251) was the main metabolite observed in the urine and plasma samples and resulted from the N-deethylation of etamiphylline. The second m...
Quantitative analysis of lignocaine and metabolites in equine urine and plasma by liquid chromatography-tandem mass spectrometry. In this paper, a method for the sensitive and reproducible analysis of lignocaine and its four principal metabolites, monoethylxylidide (MEGX), glycylxylidide (GX), 3-hydroxylignocaine (3-HO-LIG), 4-hydroxylignocaine (4-HO-LIG) in equine urine and plasma samples is presented. The method uses liquid chromatography coupled to tandem mass spectrometry operating in electrospray ionisation positive ion mode (+ESI) via multiple reaction monitoring (MRM). Sample preparation involved solid-phase extraction using a mixed-mode phase. The internal standard adopted was lignocaine-d(10). Lignocaine and its...
Determination of (13)C/(12)C ratios of urinary excreted boldenone and its main metabolite 5beta-androst-1-en-17beta-ol-3-one. Boldenone (androsta-1,4-dien-17beta-ol-3-one, Bo) is an anabolic steroid known to have been used in cattle breeding or equine sport as a doping agent for many years. Although not clinically approved for human application, Bo or its main metabolite 5beta-androst-1-en-17beta-ol-3-one (BM1) were detected in several doping control samples. For more than 15 years the possibility of endogenous Bo production in human beings has been discussed. This is a challenging issue for doping control laboratories as Bo belongs to the list of prohibited substances of the World Anti-Doping Agency and therefore th...
Determination of non-steroidal anti-inflammatory drugs residues in animal muscles by liquid chromatography-tandem mass spectrometry. A confirmatory method for the determination of residues of nine non-steroidal anti-inflammatory drugs and one metabolite in animal muscles has been developed. After enzymatic hydrolysis samples were extracted with acetonitrile and cleaned up using alumina and C(18) SPE cartridges. Liquid chromatography-tandem mass spectrometry was used for the separation and determination of analytes. The method was validated in bovine muscles, according to the Commission Decision 2002/657/EC criteria. Applicability of the method in the analysis of swine, horse and chicken muscles was checked by precision and ...
Comparative in vitro metabolism of the ‘designer’ steroid estra-4,9-diene-3,17-dione between the equine, canine and human: identification of target metabolites for use in sports doping control. Effective detection of the abuse of androgenic-anabolic steroids in human and animal sports often requires knowledge of the drug's metabolism in order to target appropriate urinary metabolites. 'Designer' steroids are problematic since it is difficult to obtain ethical approval for in vivo metabolism studies due to a lack of a toxicological profile. In this study, the in vitro metabolism of estra-4,9-diene-3,17-dione is reported for the first time. This is also the first study comparing the metabolism of a designer steroid in the three major species subject to sport's doping control; namely th...
Depletion of urinary zilpaterol residues in horses as measured by ELISA and UPLC-MS/MS. Three horses were dosed with dietary zilpaterol and the urine concentrations measured from withdrawal day 0 to withdrawal day 21. The analyses were carried out using both enzyme-linked immunosorbent assay (ELISA) and an ultraperformance liquid chromatography with triple-quadrupole-tandem mass spectrometric detection (UPLC-MS/MS). The UPLC-MS/MS method was developed to provide rapid analysis with positive analyte identification by following three product ions and computing the two independent ion ratios. When urinary zilpaterol concentrations were between 0.2 and 2 ng/mL, the ELISA had interday...
Determination of oral tramadol pharmacokinetics in horses. The determination of the pharmacokinetic parameters of tramadol in plasma and a better characterization of its metabolites after oral administration to horses is necessary to design dosage regimens to achieve target plasma concentrations that are associated with analgesia. The purpose of this study was to determine the pharmacokinetics and elimination pattern in urine of tramadol and its metabolites after oral administration to horses. Tramadol was administered orally to six horses and its half-life, T(max) and C(max) in plasma were 10.1, 0.59 h, and 132.7 ng/mL, respectively. The half-life, T...
Species comparison of oral bioavailability, first-pass metabolism and pharmacokinetics of acetaminophen. Species differences in oral bioavailability, first-pass metabolism and pharmacokinetics of biopharmaceutics classification system (BCS) class I compound acetaminophen were studied. The absolute bioavailability was 42.2%, 39.0%, 44.5%, 75.5% and 91.0% in chickens, turkeys, dogs, pigs and horses, respectively. After hydrolysis of metabolites by beta-glucuronidase/sulfatase, apparent bioavailability increased significantly in all species (turkeys: 72.4%, dogs: 100.5%, pigs: 102.2%), except horses (91.6%). Mean metabolic ratios of [acetaminophen glucuronide]/[acetaminophen] between 0 and 1h were s...
Simplified method to measure glucocorticoid metabolites in faeces of horses. 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...