The Metabolism of Anabolic Agents in the Racing Greyhound

Williams, Keith Robert (1999) The Metabolism of Anabolic Agents in the Racing Greyhound. PhD thesis, University of Glasgow.

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Abstract

The work presented in this thesis was concerned with the detection of anabolic drugs in the racing greyhound. The use of these substances in racing animals is prohibited by the National Greyhound Racing Club in the rules governing the sport. Prior to this study, little information existed regarding the metabolism of these anabolic agents in the racing greyhound or the detection of their administration. This study initially established a greyhound metabolic unit and developed an experimental protocol based on fully trained racing greyhounds as the animal model. The unit and protocol were licenced by the Home Office for this purpose. Because of the lack of information in the literature, work was carried out to establish the nornial urinary steroid profile in the racing greyhound and some basic information regarding steroid metabolism in the racing greyhound. Subsequently, the metabolism of selected anabolic steroids (nandrolone and boldenone) in the greyhound was studied, together with the effects of these exogenous steroids on the normal steriodal profile. Finally, the metabolism in the greyhound of another anabolic agent, clenbuterol, was examined. Methodology was developed for the analysis of steroids in urine and plasma based on solid phase extraction, enzymatic hydrolysis, Sephadex LH-20 fractionation and qualitative and quantitative analysis by gas chromatography-mass spectrometry (GC-MS). The extraction protocols were assessed using radiolabelled steroids and found to be acceptable. Hydrolysis procedures were assessed by HPLC and found to be satisfactory. A variety of derivatisation reagents were evaluated to optimise the GC-MS procedure . It was not possible to detect the presence of steroids in the urine of male greyhounds usmg full scan mass spectrometry. In female greyhounds, however, it was possible to identify the presence of a pregnane, an estrogen and two corticosteroids. Additional constituents were present which may also have been steroids but their identification could not be confirmed. Testosterone was shown to be present in greyhound urine using radioimmunoassay. Maximum concentrations of testosterone measured were approximately 25ng/mg creatinine and 10 ng/mg creatinine for two male greyhounds. The metabolism and turnover of testosterone in the greyhound were investigated using labelled forms of the drug. Radiolabelled testosterone was administered to a male greyhound at a low dose, designed to increase the metabolic pool size of testosterone in the animal as little as possible. Plasma samples and complete urine collections were obtained. Elimination of the radiolabelled material from the plasma was rapid and approximately 40% of the dose was recovered in the urine by 7 days post dose. An apparent volume of distribution of 5.8 litres was calculated for testosterone. A quantitatively significant dose of deuterium labelled testosterone (2 milligrams) was administered intravenously to a greyhound. The testosterone pool size for the animal was estimated by isotopic dilution at approximately 190mug. Two exogenous anabolic steroids (nandrolone and boldenone) were administered intramuscularly to two male racing greyhounds. Nandrolone was found to be present in predose urine and plasma samples indicating that nandrolone is in fact an endogenous steroid in the racing greyhound. Following administration of nandrolone to the two male dogs, levels of nandrolone in plasma rose from 30 and 60 ng/ml to over 100 ng/ml in both dogs. Elimination of nandrolone after a few days brought the plasma concentration back to the initial starting levels. Levels of endogenous steroids were depressed in plasma samples along with the concentration of creatinine excreted in the urine. Boldenone was not detected in either plasma or urme samples following mtramuscular administration. While depression of the endogenous profile did occur, it was less than that which occurred following dosing with nandrolone. Clenbuterol is a beta-agonist drug that has been abused as an anabolic agent both in human sport and in livestock farming. Methodology was developed for the detection of clenbuterol in plasma and urine samples from the racing greyhound. Although the greatest sensitivity could be achieved using negative ion chemical ionisation-mass spectrometry in association with a heptafluorobutyrate derivative, the use of a simpler trimethylsilyl derivative and electron impact ionisation was found to be adequate and was considered more appropriate as a routine method. Clenbuterol was administered orally to one male and one female racing greyhound. Severe side effects were noted within 2 hours post dose and continued past 24 hours post dose. Peak plasma concentrations of clenbuterol occurred between 5-24 hours post dose and it was possible to determine the presence of clenbuterol in plasma and urine of the dogs for at least 8 days post dose. It was concluded that the principal aims of the project had been achieved.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Keywords: Analytical chemistry, Animal sciences
Date of Award: 1999
Depositing User: Enlighten Team
Unique ID: glathesis:1999-76208
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 19 Nov 2019 16:28
Last Modified: 19 Nov 2019 16:28
URI: http://theses.gla.ac.uk/id/eprint/76208

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