Steroids in horses refer to a class of compounds that include both naturally occurring and synthetic substances designed to mimic the effects of hormones such as cortisol and testosterone. These compounds can influence a wide range of physiological processes, including metabolism, immune response, and muscle growth. In equine medicine, steroids are sometimes used therapeutically to manage conditions such as inflammation, allergies, and certain musculoskeletal disorders. However, their use in competitive sports is often regulated due to potential performance-enhancing effects. This page compiles peer-reviewed research studies and scholarly articles that explore the pharmacology, therapeutic applications, and regulatory aspects of steroid use in equine health and athletic performance.
Hagedorn HW, Schulz R.Statistical analysis of normally occurring cortisol levels in serum and urine of horses served to recommend thresholds for this corticosteroid in these body fluids, as application of exogenous cortisol as well as ACTH may elevate the cortisol concentrations above the proposed threshold. The present study contributes to the general issue of how to establish thresholds for trotting horses upon sportive examination. 100 randomly selected post competition serum and urine samples, respectively, were submitted to cortisol analysis by means of HPLC. Concentrations of the endogenous corticosteroid in ...
Aguilera R, Becchi M, Mateus L, Popot MA, Bonnaire Y, Casabianca H, Hatton CK.A gas chromatography-combustion-isotope ratio mass spectrometry method for confirmation of hydrocortisone abuse in horseracing and equine sports is proposed. Urinary hydrocortisone was converted to a bismethylenedioxy derivative which presents good gas chromatographic properties and brings an extra carbon contribution of only two carbon atoms. Synthetic hydrocortisone has a different 13C abundance from that of natural urinary horse hydrocortisone and the difference is significant, therefore exogenous and endogenous hydrocortisone can be distinguished.
Raeside JI, Renaud RL, Christie HL.Dehydroepiandrosterone (DHEA) and 3 beta-hydroxyandrosta-5,7-dien-17-one (7-dehydro-DHEA) are secreted in large quantities by the remarkably hypertrophied fetal gonads of both sexes in the pregnant mare. Their secretion serves as the fetal component of a feto-placental unit for oestrogen production in equine pregnancies. They are secreted in large amounts but show a decline in late pregnancy when the fetal gonads regress and levels of oestrogens in the mare fall as a consequence. We have examined the levels of these precursor steroids in the newborn foal in the first days after birth. DHEA and...
Raeside JI, Christie HL.Large amounts of estrogens are secreted by the tests of the mature stallion. In a recent study by Claus et al. [Claus, Dimmick, T., Gimenez, T., Hudson, L.W., 1992. Estrogens and prostaglandin F2 alpha in the semen and blood plasma of stallions. Theriogenology 38, 687-693.], it was stated that high levels of estrogens were also present in semen. As a preliminary step to study possible implications for fertility in the stallion, we have measured estrone sulphate (E1S), the principal estrogen in blood, in both seminal plasma and spermatozoa. Semen was collected from four Standardbred stallions a...
Frisbie DD, Nixon AJ.To evaluate potential stimulatory or matrix-sparing effects of insulin-like growth factor 1 (IGF-1), alone or in combination with a corticosteroid, in an interleukin 1 (IL-1)-induced model of cartilage degradation. Methods: Cartilage from the weightbearing surfaces of trochlea and condyles of clinically normal 2-year-old male horses. Methods: Triamcinolone acetonide and IGF-1 effects were evaluated by assessing: matrix responses by sulfated glycosaminoglycan (GAG) assay and [35S]sulfated GAG synthesis; collagen content by hydroxyproline assay; and mitogenic response by [3H]thymidine incorporat...
Koskinen E, Marttila P, Katila T.The long-term effects of the anabolic steroid 19-norandrostenololylaurate on semen characteristics of Finnhorse colts were studied in 3 experiments. Semen was collected initially at 24 months of age and then twice a year. In experiment I, 500 mg or 100 mg of steroid per animal was given every 3rd week from 12 or 16 months to 24 months of age. In colts treated with 500 mg of anabolic steroid every 3rd week, azoospermia was observed in 3 out of 5 colts in the first semen collections, immediately after the end of treatment. The other 2 colts had low sperm numbers and a high percentage of proximal...
Koskinen E, Andersson M, Katila T.The effect of anabolic steroid on testicular growth was investigated in 3 experiments. In experiment I, 500 mg of the anabolic steroid was given to 4 colts and 100 mg to another 4 colts, every 3rd week, starting at age 16 months and ending at age 24 months. Six colts served as controls. Both treatments decreased total scrotal width (TSW) within 6 weeks. Seasonal testicular growth during spring partly overcame the effect of steroid treatment. Cessation of anabolic steroid treatment was followed by testicular growth at the same time as TSW in untreated colts was decreasing by virtue of the effec...
Turner JW, Liu IK, Kirkpatrick JF.Regulation of local overpopulations of free-roaming feral equids is in demand worldwide for ecological balance and habitat preservation. Contraceptive vaccines have proven effective in feral horses, which breed seasonally, but no data are available for equids such as the burro, which is reproductively active all year round. In the present study, 27 individually identified female feral burros (Equus asinus) roaming free in Virgin Islands National Park (St John, US Virgin Islands; Lesser Antilles) were remotely treated with pig zonae pellucidae (PZP) vaccine. Between January and May, 16 burros w...
Schutzer WE, Holtan DW.The mare possesses unique steroid hormone metabolic activity during pregnancy in that peripheral 4-pregnene-3,20-dione (progesterone; P4) is undetectable by 220 days gestation. This study examines in vivo metabolism of progestins by the pregnant mare and in vitro metabolic activity of maternal and fetal tissues. Pregnant mares (n = 3) received intravenous infusions of 3 beta-hydroxy-5-pregnen-20-one (pregnenolone; P5), P4, 5 alpha-pregnane-3,20-dione (5 alpha-DHP), 3 beta-hydroxy-5 alpha-pregnan-20-one (3 beta-5 alpha), deuterium labeled (D4)-P5, D4-3 beta-5 alpha and vehicle. Anestrous mares ...
Lillich JD, Bertone AL, Schmall LM, Ruggles AJ, Sams RA.OBJECTIVE--To document plasma, urine, and synovial fluid disposition of 2 common intra-articularly administered steroid preparations, methylprednisolone acetate (MPA) and isoflupredone acetate (IPA). DESIGN--Descriptive investigation. SAMPLE POPULATION--100 mg of MPA or 4 mg of IPA was administered to 2 groups of 4 healthy sound radiographically normal female horses. PROCEDURE--Blood samples were collected at time 0 (before) and 2, 4, 6, 8, 10, 12, 24, 36, 48, 72, and 96 hours after administration of the designated steroid. Complete urine collection for measurement of designated steroid was ac...
Chen CL, Zhu D, Gillis KD, Meleka-Boules M.To develop a simple and sensitive ELISA for detection of dexamethasone in horse serum and urine. Methods: Blood and urine samples from 3 thoroughbred mares. Methods: A dexamethasone oxime was prepared and conjugated to hemocyanin, bovine serum albumin and to horseradish peroxidase. One- and two-step double-antibody ELISA methods, as well as a radioimmunoassay method, were performed. The one-step ELISA was used to test urine from 3 Thoroughbred mares injected with 5 mg of dexamethasone, IV. Results: The ELISA could detect dexamethasone in the range of 0.01 to 50 ng/ml, with intra- and interassa...
Gu X, Meleka-Boules M, Chen CL.A capillary electrophoresis technique was developed for the separation of synthetic glucocorticoids and the determination of dexamethasone and flumethasone in horse urine. Pretreatment of the sample using a dexamethasone affinity column resulted in low background that enabled the authors to detect levels as low as 1.1 ng/mL and 2.7 ng/mL for dexamethasone and flumethasone in horse urine, respectively. The developed method was used to detect dexamethasone in horse urine samples after the injection of a therapeutic dose of dexamethasone for up to 12 hr postinjection. The optimum conditions for c...
Geor R, Hope E, Lauper L, Piela S, Klassen J, King V, Murphy M.The effects of dexamethasone (0.2 mg/kg of body weight; IV, IM, and PO) and methylprednisolone acetate (120 mg given intra-articularly) on serum osteocalcin and cortisol concentrations were studied in 6 horses. Serum osteocalcin and cortisol concentrations were serially monitored after each treatment. A significant (P < 0.05) decrease in serum osteocalcin and cortisol concentrations was observed from 12 to 24 and 2 to 48 hours, respectively, after IV and IM administrations of dexamethasone. Serum osteocalcin and cortisol concentrations were significantly decreased from 6 to 48 and 3 to 72 h...
Thomas PG, Ignotz GG, Ball BA, Miller PG, Brinsko SP, Currie B.Oviduct epithelial cells (OEC) increasingly are used to support embryonic development and to study gamete interactions with the female reproductive tract in vitro. This series of experiments was designed to characterize monolayers derived from oviduct epithelium. Epithelial cells harvested from the isthmus and ampulla of the oviducts of five estrous mares were cultured with or without the basal lamina extract, Matrigel. Within each group OEC were cultured in the presence of either estradiol-17 beta or a carrier control. All groups were subcultured three times. Epithelial cell morphology and fu...
McDowell KJ, Adams MH, Franklin KM, Baker CB.A cDNA library was constructed from poly(A) RNA obtained from Day 14 nonbred equine endometrium. A cDNA probe for porcine retinol-binding protein (RBP) was used to screen the library, and a complete cDNA sequence (1133 bp, excluding the poly(A) tail) was obtained. Endometrial biopsies were obtained from cycling, nonbred mares at Days 0, 1, 4, 8, 10, 11, 13, and 15 and from pregnant mares at Days 11, 13, 15, and 17 after ovulation (n = 2 mares each day). Endometrial biopsies were also taken from 18 noncycling anestrous mares after the following treatments: C (vehicle control for 1 day, n = 3), ...
Chen CL, Goldberg J, Gronwall RR.A radioimmunoassay was developed for prednisolone using IgG purified from rabbit antiserum. The assay was employed to determine the pharmacokinetics of prednisolone following intravenous administration of 450 mg of prednisolone sodium succinate (Solu Delta Cortef) to five adult Thoroughbred horses. The RIA had a sensitivity of 2 ng/ml and was relatively specific. It had cross-reactivity with 21-deoxycortisol (83.3%) cortisol (27.8%), 11-beta-hydroxyprogesterone (39.2%) and 17-hydroxyprogesterone (50%). However, it did not cross-react with naturally occurring steroids (cholesterol, testosterone...
Rossdale PD, Ousey JC, McGladdery AJ, Prandi S, Holdstock N, Grainger L, Houghton E.Plasma progestagen concentrations were measured daily by radioimmunoassay (RIA) in 35 sick foals for the duration of their illness. The foals were divided into three groups on the basis of time to stand after birth. Foals were given intensive care treatment according to the severity of their illness. Plasma and urine concentrations of pregnenolone (P5) and pregnenediol (P5 beta beta) were measured by gas chromatography--mass spectrometry; plasma cortisol concentrations were measured by RIA and the foals' renal and respiratory status were assessed by creatinine clearance ratios and arterial oxy...
Eisenhauer KM, Roser JF.A stallion testicular cell incubation system was developed and used to investigate the regulation of steroidogenesis in stallion testes. Cells isolated from testes of 2- to 4-year-old stallions (n = 6) were cultured for 12 hours in a defined medium with and without varying doses of lipoprotein, equine luteinizing hormone (eLH), human chorionic gonadotropin (hCG), equine follicle-stimulating hormone (eFSH), and/or equine prolactin (ePRL). Estrogen conjugate (EC), testosterone (T), and estradiol-17 beta (E2) production were determined by RIA. Increasing doses of lipoprotein significantly (P <...
Foland JW, McIlwraith CW, Trotter GW, Powers BE, Lamar CH.Osteochondral fragments were created arthroscopically on the distal aspect of both radial carpal bones in 12 horses. On day 14 after surgery, one middle carpal joint of each horse was injected with 2.5 mL Betavet Soluspan (3.9 mg betamethasone sodium phosphate and 12 mg betamethasone acetate per milliliter) and the contralateral joint was injected with 2.5 mL saline as a control. Intra-articular treatments were repeated on day 35. On day 17, six horses began exercising 5 days per week on a high-speed treadmill. The other six horses were kept in box stalls throughout the study as nonexercised c...
Harkins JD, Carney JM, Tobin T.Corticosteroids possess potent anti-inflammatory activity and are commonly injected intra-articularly for local relief of inflammatory lesions in performance horses. However, the suppression of anabolic activity in the joint may lead to an increased rate of joint breakdown. Complications associated with intra-articular corticosteroid therapy include septic arthritis, which is usually due to inadvertent joint contamination at the time of corticosteroid injection, and steroid arthropathy, which is characterized by an accelerated rate of joint destruction and radiographic evidence of severe degen...
Muyan M, Roser JF, Dybdal N, Baldwin DM.The objective of the present study was to determine whether the testicular steroids, i.e., testosterone (T), dihydrotestosterone (DHT), estradiol (E2), estrone (E1), and estrone sulfate (E1SO4), play a physiological role in regulating LH release in the male horse by direct actions at the anterior pituitary gland. Enzymatically dispersed anterior pituitary cells from stallions (n = 4) or geldings (n = 3) were cultured for 48 h in alpha-modified Eagle's medium containing 10% steroid-free horse medium. To determine the effects of the steroids on the LH response to GnRH, the cells were incubated f...
Goff AK, Leduc S, Poitras P, Vaillancourt D.The objective of this study was to determine if changes in steroid synthesis occurred in the horse blastocyst about the time of maternal recognition of pregnancy. Embryos collected between days 7.5 and 14.5 were incubated for 8 hr in vitro in HAM's F10 containing radiolabelled pregnenolone. The steroid metabolites in the incubation medium were separated by reverse phase HPLC and the major peaks expressed as a percentage of total metabolites. It was found that there were no major changes in the profile of metabolites throughout the period of study, although there was increased conversion as the...
Hamir AN, Moser G, Galligan DT, Davis SW, Granstrom DE, Dubey JP.A 5-year (1985-1989) retrospective immunohistochemical study was conducted using an avidin-biotin complex (ABC) immunoperoxidase method to demonstrate Sarcocystis neurona in histologically suspect cases of equine protozoal myeloencephalitis (EPM). Primary antibodies against S. neurona and S. cruzi were utilized for the ABC technique. The findings were compared with those from cases in which the organisms were detected by examination of hematoxylin and eosin (HE)-stained neuronal sections. HE-stained sections detected the presence of the organisms in 20% of the suspect cases; whereas the ABC te...
Oladosu LA, Olufemi BE.An investigation was carried out to study the haematology of steroid immunosuppressed horses experimentally infected with Babesia equi and Ehrlichia equi, separately or simultaneously. Horses infected with both pathogens showed less marked changes in their haematology than those inoculated with either pathogen separately. This appeared to result from early elimination of the more pathogenic Babesia as Ehrlichia spread through the granulocytes. The apparent suppression of Babesia by Ehrlichia is of field clinical importance and merits further investigation for its apparent useful potentials in ...
Steffenrud S, Maylin G.A high-performance liquid chromatographic method was developed for thermospray mass spectrometric analysis of steroidal hormones. Using a Nova-Pak C18 reversed-phase column and isocratic elution with a solvent comprised of 25 mM ammonium formate in 30% acetonitrile, corticosteroids were separated within 10 min. This solvent also permitted ultraviolet absorbance detection down to 220 nm with low-nanogram sensitivity. The use of acetonitrile was favourable for thermospray mass spectrometric analysis because mass spectra were obtained with a pseudomolecular ion as the base peak. A combination of ...
Cohen ND, Carter GK.Steroid hepatopathy was diagnosed in a horse with glucorticoid-induced hyperadrenocorticism on the basis of anamnesis, serum biochemical data, and histologic findings of hepatic biopsy. Initially, clinical signs of polyuria, polydypsia, and muscular degeneration were seen. The horse developed laminitis during hospitalization.
Weber JA, Freeman DA, Vanderwall DK, Woods GL.This study was conducted to identify embryonic products whose secretion was temporally associated with the oviductal transport period of the mare. Chemicals secreted by oviductal-transport-stage equine embryos were identified by incubating Day 6 or Day 7 early uterine embryos with 35S-methionine/cysteine, 3H-progesterone, or 3H-arachidonic acid for 24 h, and subsequently identifying radioactively labeled proteins (SDS-PAGE; n = 3 embryos), steroids (HPLC; n = 3 embryos), or prostaglandins (HPLC; n = 3 embryos) in the culture medium. Early uterine embryos secreted 116.1 +/- 45.5 pg of prostagla...
Watson ED, Sertich PL.This study investigated the sequence of hormonal changes within the preovulatory follicles of mares. Mares were injected i.v. with 2500 IU human chorionic gonadotrophin (hCG) when a preovulatory follicle of 35 mm in diameter was detected. Fluid was aspirated from preovulatory follicles before (0 h), and 12, 24 and 36 h after administration of hCG. Concentrations of progesterone, prostaglandin (PG) E2, PGF, 6-keto-PGF1 alpha and thromboxane B2 in follicular fluid increased significantly (P less than 0.01) between 0 and 36 h. At 36 h, PGE2 was present in highest concentrations, followed by PGF a...
Teale P, Houghton E.A screening procedure for anabolic steroid residues in horse urine has been developed based upon solid-phase extraction and gas chromatographic/mass spectrometric analysis in the selected ion mode. For moderate sample throughput the method provides a viable alternative to radioimmunoassay screening and has advantages over the latter technique due to its flexibility, specificity and ability to detect a number of steroids in a single analysis. Full automation of the gas chromatographic/mass spectrometric analysis is an additional feature of the methodology.
Bijault C, Dehennin L.Steroid 21-hydroxylase activity of the microsome-enriched fraction of follicular linings from equine ovaries has been demonstrated by gas chromatography-mass spectrometry. The 21-hydroxylated metabolites were quantified by isotope dilution with deuterated analogues. The two most abundant potential substrates for follicular steroid 21-hydroxylase, progesterone (P) and 17-hydroxyprogesterone (17OHP), were converted respectively to 11-deoxycorticosterone (DOC) and 11-deoxycortisol with corresponding apparent specific activities of 308 and 24 pmol/mg protein/h and apparent Km values of 1.1 and 6.4...
Stucki M, Voegel CD, Binz TM, Kraemer T, Lavaud A, Voelter K.Equine sport agencies list steroids as prohibited substances for competing horses. Objective: The objective of this study was to investigate if the controlled substances dexamethasone and prednisolone are detectable in equine serum and urine samples during and after treatment with eye drops and if this can generate a positive doping test. Methods: Prospective cohort study. Methods: The study cohort included 11 horses. One eye of the horses was treated with either dexamethasone (Maxitrol 0.1%, n = 5 eyes) or prednisolone (Pred forte 1%, n = 6 eyes) eye drops 3 times daily for 14 days. Dexa...
Oladosu LA, Olufemi BE.An investigation was carried out to study the haematology of steroid immunosuppressed horses experimentally infected with Babesia equi and Ehrlichia equi, separately or simultaneously. Horses infected with both pathogens showed less marked changes in their haematology than those inoculated with either pathogen separately. This appeared to result from early elimination of the more pathogenic Babesia as Ehrlichia spread through the granulocytes. The apparent suppression of Babesia by Ehrlichia is of field clinical importance and merits further investigation for its apparent useful potentials in ...
Karatt TK, Sathiq MA, Laya S, Kal AKK, Subhahar MB, Muhammed Ajeebsanu MP, Philip M, Caveney MR, Graiban FM.Nonsteroidal selective androgen receptor modulators (SARMs) are a novel class of compounds that have not yet been clinically approved; however, they appear to have a better anabolic/androgenic ratio than steroids and cause slighter side effects. Sports drug testing laboratories are required to maintain continuously updated doping control analytical methods in light of the widespread misuse of SARMs in elite and amateur sports. This paper describes the metabolic conversion of SARM GSK2881078 in thoroughbred horses following oral administration and in vitro with equine liver microsomes. A liquid...
Stanley SM, Kent S, Rodgers JP.In this study the equine metabolism of fluoxymesterone (9alpha-fluoro-11beta-17beta-dihydroxy-17alpha-meth ylandrost-4-ene-3-one) given orally has been investigated. The parent material was not detected, but two major 16-hydroxy metabolites which corresponded to a mono- and a di-hydroxylation product were evident. One of the hydroxylation positions was identified as C-16. Phase II metabolism in the form of glucuronide formation was also common. These steroids will provide target compounds for confirming abuse of this drug in the horse.
Marshall DE, Gower DB, Silver M, Fowden A, Houghton E.Equine umbilicus was cannulated in utero and a series of cord plasma samples removed for analysis. After steroid extraction and derivatisation, gas chromatographic-mass spectrometric (GC-MS) analysis demonstrated large differences in steroid content between the plasma samples obtained from the umbilical artery and vein, the blood supplies leading to and from the placental surface, respectively. 3Beta-hydroxy-5,7-androstadien-17-one, dehydroepiandrosterone, pregnenolone, 3beta-hydroxy-5alpha-pregnan-20-one, 5-pregnene-3beta,20beta-diol and 5beta-pregnane-3beta,20beta-diol were identified as maj...
Roser JF, Tarleton M, Belanger JM.Recent studies in our laboratory investigating plasma gonadotrophin concentrations before and after castration indicate that the hypothalamic-pituitary axis is not the original site of dysfunction in stallions with idiopathic infertility. In the present study, fertile, subfertile and infertile stallions were subjected to replacement therapy with oestradiol and testosterone 1 year after castration to investigate hypothalamic-pituitary axis function further. Fertile (n=5), subfertile (n=3) and infertile (n=2) castrated stallions of Light horse breeds, aged 11-23 years, were given either oestradi...
Viljanto M, Scarth J, Hincks P, Hillyer L, Cawley A, Suann C, Noble G, Walker CJ, Kicman AT, Parkin MC.Detection of testosterone and/or its pro-drugs in the gelding is currently regulated by the application of an international threshold for urinary testosterone of 20 ng/mL. The use of steroid ratios may provide a useful supplementary approach to aid in differentiating between the administration of these steroids and unusual physiological conditions that may result in atypically high testosterone concentrations. In the current study, an ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method was developed to quantify testosterone (T) and epitestosterone (E). ...
Muyan M, Roser JF, Dybdal N, Baldwin DM.The objective of the present study was to determine whether the testicular steroids, i.e., testosterone (T), dihydrotestosterone (DHT), estradiol (E2), estrone (E1), and estrone sulfate (E1SO4), play a physiological role in regulating LH release in the male horse by direct actions at the anterior pituitary gland. Enzymatically dispersed anterior pituitary cells from stallions (n = 4) or geldings (n = 3) were cultured for 48 h in alpha-modified Eagle's medium containing 10% steroid-free horse medium. To determine the effects of the steroids on the LH response to GnRH, the cells were incubated f...
Dedar RK, Kumar N, Narnaware SD, Tripathi BN.Wound healing in horses is complicated by the excessive growth of granulation tissue, commonly known as proud flesh and is similar to keloids in human beings. At present, there is no satisfactory treatment for proud flesh in horses. In this study, we, for the first time, demonstrated that leaf extract of Aerva javanica suppresses excessive growth of granulation tissue in horses. Many plant flavonoids are claimed to have antiproliferative properties. Kaempferol is a natural flavonoid containing 3-hydroxy flavone backbone found in many plants in its aglycone form and attached with various sugars...
Chen CL, Goldberg J, Gronwall RR.A radioimmunoassay was developed for prednisolone using IgG purified from rabbit antiserum. The assay was employed to determine the pharmacokinetics of prednisolone following intravenous administration of 450 mg of prednisolone sodium succinate (Solu Delta Cortef) to five adult Thoroughbred horses. The RIA had a sensitivity of 2 ng/ml and was relatively specific. It had cross-reactivity with 21-deoxycortisol (83.3%) cortisol (27.8%), 11-beta-hydroxyprogesterone (39.2%) and 17-hydroxyprogesterone (50%). However, it did not cross-react with naturally occurring steroids (cholesterol, testosterone...
Skrabalak DS, Covey TR, Henion JD.Several important corticosteroids were qualitatively determined in the plasma and urine of horses by micro-liquid chromatography-mass spectrometry (micro-LC-MS). The sensitivity and specificity of micro-LC-MS are demonstrated as is the ability of micro-LC-MS to deal with endogenous interferences. In turn, the relative amount of dexamethasone and its major unconjugated metabolite were determined in equine urine by micro-LC-MS; the conclusions drawn are reported.
Curcio BR, Canisso IF, Pazinato FM, Borba LA, Feijó LS, Muller V, Finger IS, Toribio RE, Nogueira CEW.The overall goal of this study was to assess the efficacy of various therapeutic combinations of estradiol cypionate (ECP, a long-acting estrogen) and altrenogest (ALT, a long-acting progestin) in addition to basic treatment for placentitis with trimethoprim-sulfamethoxazole (TMS) and flunixin meglumine (FM). Specific outcomes measured in this experiment were (i) time from induction of bacterial placentitis to delivery, and foal parameters (high-risk, survival, and birth weight); and (ii) serum steroid concentrations (progesterone, 17α-hydroxyprogesterone, 17β-estradiol, and cortisol) in res...
Shin HD, Suh JH, Kim J, Cho HD, Lee SD, Han KS, Wang Y, Han SB.A high throughput method for simultaneous screening of anabolic steroids and their metabolites (4-esterendione, trenbolone, boldenone, oxandrolone, nandrolone, methandrostenolone, testosterone, 1-androstendione, ethisterone, normethandrolone, methyltestosterone, 16β-Hydroxystanozolol, epitestosterone, bolasterone, norethandrolone, danazol, stanozolol and androstadienone) in equine urine by online turbulent flow extraction coupled with liquid chromatography-tandem mass spectrometry was developed. The use of turbulent flow chromatography could simplify pretreatment of horse urine, which has com...
Teale P, Houghton E.A screening procedure for anabolic steroid residues in horse urine has been developed based upon solid-phase extraction and gas chromatographic/mass spectrometric analysis in the selected ion mode. For moderate sample throughput the method provides a viable alternative to radioimmunoassay screening and has advantages over the latter technique due to its flexibility, specificity and ability to detect a number of steroids in a single analysis. Full automation of the gas chromatographic/mass spectrometric analysis is an additional feature of the methodology.
Fidani M, Pompa G, Mungiguerra F, Casati A, Fracchiolla ML, Arioli F.After the detection of low concentrations of prednisolone in racehorse urine samples collected at Italian racetracks, a study was initiated to investigate the accuracy of the analytical protocol used and the possible endogenous origin of detected prednisolone. Methods: Multiple reaction monitoring (MRM) MS(2) acquisition with a triple quadrupole (n = 780) and full scan MS(2) and MS(3) (n = 180) acquisition with a linear ion trap were checked. As a further confirmation, ten urine samples were analysed by high-resolution mass spectrometry (HRMS). Results: The study showed the difficulty of ident...
Baldwin DM, Roser JF, Muyan M, Lasley B, Dybdal N.Enzymatically dispersed anterior pituitary cells from donor mares were cultured for 48 h in alpha-modified Eagles' medium containing 10% steroid-free horse serum. The cells were then incubated for 24 h in fresh medium oestrogen followed by a 4-h incubation with or without GnRH. Media and cell extracts were analyzed for LH by radioimmunoassay. In the first series of experiments, pituitary cells from Day-3 dioestrous mares were preincubated with ethanol (control) or different concentrations of E2 (10(-11) to 10(-7) M) for 24 h prior to a 4-h incubation without (basal) or with 1.0 nM GnRH. E2 inc...
Viljanto MJ, Kicman AT, Walker CJ, Parkin MC, Wolff K, Pearce CM, Scarth J.Boldenone is an anabolic-androgenic steroid that is prohibited in equine sports. Urine from the uncastrated male horse contains boldenone that is thought to be of endogenous origin and thus a threshold ('cut-off') concentration has been adopted internationally for free and conjugated boldenone to help distinguish cases of doping from its natural production. The testis is likely to be a source of boldenone. Qualitative analysis was performed on extracts of equine testicular homogenates (n = 3 horses) incubated non-spiked and in the presence of its potential precursors using liquid chromatog...
Weiser G, Kohn C, Vachon A.Immune-mediated hemolytic anemia was diagnosed in two horses on the basis of regenerative anemia, increased erythrocyte fragility in hypotonic saline, autoagglutination, and a positive direct antiglobulin (Coomb's) test. During steroid therapy partial resolution of the anemia was indicated by rising packed cell volume, macrocytosis, and bone marrow erythroid hyperplasia. Using erythrocyte volume distribution histograms (erythrograms), the regenerative response was characterized by analysis of macrocytic and normocytic erythrocyte subpopulations. In both horses, a gradual net increase of about ...
Bhavnani BR, Woolever CA.[3H]Equilin [3H-labeled 3-hydroxy-1,3,5(10), 7-estratetraen-17-one] was administered iv to a pregnant mare in the 10th month of gestation. Maternal urine was collected for 3 days, and blood samples were taken 35 min and 3, 6, 12, and 24 h after the injection. The half-life of the disappearance of radioactivity from the blood was approximately 2.5 h. Over 90% of the administered dose was excreted in the first 24 h. The urine was extracted, hydrolyzed, and fractionated. The bulk of the radioactive material (75%) was present in the phenolic sulfate fraction from which radiochemically pure equilin...
Houghton E, Dumasia MC, Wellby JK.Negative ion chemical ionization mass spectra of some corticosteroids have been obtained by direct syringe introduction on to the Finnigan moving belt high-performance liquid chromatography-mass spectrometer interface. Proprietary preparations based upon dexamethasone, betamethasone and prednisolone were administered to horses at therapeutic dose level. Urine samples were extracted, the extracts purified by Sephadex LH-20 chromatography and the presence of the parent steroids in the eluates was confirmed by combined high-performance liquid chromatography negative ion chemical ionization mass s...
Liu Y, Uboh CE, Soma LR, Li X, Guan F, You Y, Rudy JA, Chen JW.In 2008, Pennsylvania (PA) became the first State in the USA to ban and enforce the ban on the use of anabolic and androgenic steroids (AAS) in equine athletes by using plasma for analysis. To enforce the ban, a rapid and high-throughput method for analysis of 60 AAS in equine plasma was developed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Analytes were recovered from plasma by liquid-liquid extraction (LLE) using methyl tert-butyl ether, separated on a reversed-phase C₁₈ column and analyzed by electrospray ionization mass spectrometry. Multiple-reaction monitoring (MRM)...
Watson ED, Thomson SR, Howie AF.A steroidogenic acute regulatory (StAR) protein has been identified in several species as a probable important rate-limiting step in steroidogenesis. This protein is believed to be responsible for transporting cholesterol from the outer to the inner mitochondrial membrane. It is known that equine chorionic gonadotrophin (eCG) stimulates steroidogenesis in the corpora lutea of early pregnant mares and that eCG also upregulates StAR mRNA in bovine ovaries. In the present study, ovarian tissue from cyclic and early pregnant mares was immunostained to detect the distribution of the StAR protein. W...
Kikuchi M, Nagata SI, Ishige T, Minamijima Y, Hirota KI, Tozaki T, Kakoi H, Kizaki K.Glucocorticoid preparations have anti-inflammatory effects, and are commonly used in the equine clinical setting; however, such treatments can cause a number of side effects. Adrenal insufficiency is an adverse effect induced by the suppression of adrenal function following drug administration. This study aimed to investigate the influence of two glucocorticoid preparations, dexamethasone and hydrocortisone, on adrenocortical function in horses. The usual doses of dexamethasone and hydrocortisone preparations in equine practice were administered intramuscularly to six horses, and peripheral bl...
Eiler H, Goble D, Oliver J.The plasma concentration of hydrocortisone was determined in mares given either cosyntropin (100 IU, given IV) or corticotropin (200 IU, given IM). Plasma hydrocortisone concentrations of the mares treated with cosyntropin increased by 46%, 57% and 80% at 30, 60, and 120 minutes, respectively, when compared with base-line values; these values returned to base line at 240 minutes. In mares treated with corticotropin, mean plasma hydrocortisone concentrations increased by 42%, 143%, 101% and 155% at 30, 60, 120, and 240 minutes, respectively, when compared with base-line values. Differences in t...
Dintinger T, Gaillard JL, Zwain I, Bouhamidi R, Silberzahn P.The results of the measurement of 19-nortestosterone in the testiscular artery and vein of the stallion, the very low levels of this steroid in the peripheral blood of geldings and the similar patterns of increase in the peripheral levels of 19-nortestosterone and testosterone after hCG stimulation, show that 19-nortestosterone, like testosterone, is essentially synthesized in the testis. This testicular origin was confirmed by the ability of testicular tissue to synthesize 19-norandrogens from [4-14C]androgens in vitro. 19-Nortestosterone was 50% conjugated in the peripheral blood and almost ...
Steffenrud S, Maylin G.A high-performance liquid chromatographic method was developed for thermospray mass spectrometric analysis of steroidal hormones. Using a Nova-Pak C18 reversed-phase column and isocratic elution with a solvent comprised of 25 mM ammonium formate in 30% acetonitrile, corticosteroids were separated within 10 min. This solvent also permitted ultraviolet absorbance detection down to 220 nm with low-nanogram sensitivity. The use of acetonitrile was favourable for thermospray mass spectrometric analysis because mass spectra were obtained with a pseudomolecular ion as the base peak. A combination of ...
Dumasia MC, Houghton E, Jackiw M.After homogenization of testicular tissue from stallions aged 1, 2 and 5 years, the unconjugated and conjugated steroids were isolated by a combined solvent-solid extraction procedure. The conjugates were further separated into glucuronides and sulphates by chromatography using Sephadex LH-20. After enzyme hydrolysis and solvolysis of the respective conjugate classes, the three extracts, unconjugated steroids, aglycones and solvolysed sulphates, were purified by chromatography using Kieselgel 60H columns. Five fractions were resolved from each extract; an aliquot of each fraction was derivatiz...
Bijault C, Dehennin L.Steroid 21-hydroxylase activity of the microsome-enriched fraction of follicular linings from equine ovaries has been demonstrated by gas chromatography-mass spectrometry. The 21-hydroxylated metabolites were quantified by isotope dilution with deuterated analogues. The two most abundant potential substrates for follicular steroid 21-hydroxylase, progesterone (P) and 17-hydroxyprogesterone (17OHP), were converted respectively to 11-deoxycorticosterone (DOC) and 11-deoxycortisol with corresponding apparent specific activities of 308 and 24 pmol/mg protein/h and apparent Km values of 1.1 and 6.4...
Waller CC, Cawley AT, Suann CJ, Ma P, McLeod MD.Furazadrol ([1',2']isoxazolo[4',5':2,3]-5α-androstan-17β-ol) is a designer anabolic androgenic steroid that is readily available via the internet. It contains an isoxazole fused to the steroid A-ring which offers metabolic stability and noteworthy anabolic activity raising concerns over the potential for abuse of this compound in equine sports. The metabolism of furazadrol was studied by in vivo and in vitro methods for the first time. Urinary furazadrol 17-sulfate and furazadrol 17-glucuronide metabolites were detected in vivo after a controlled administration and compared with syntheticall...
Her GR, Watson JT.Sensitive and specific methodology based on capillary column gas chromatography-electron capture-negative chemical ionization-mass spectrometry has been developed for the quantitative analysis of corticosteroids from biological fluids. The feasibility of this method is demonstrated in the quantitative analysis of dexamethasone in horse urine following administration of the drug. A structurally similar compound, 6 alpha-methylprednisolone, is added to the urine as an internal standard. The free dexamethasone and the internal standard are extracted and oxidized to high-electron-affinity 1,4-andr...
