Detection and pharmacokinetics of salmeterol in thoroughbred horses following inhaled administration.

Salmeterol is a man-made beta-2-adrenergic receptor agonist used to relieve bronchospasm associated with inflammatory airway disease in horses. Whilst judicious use is appropriate in horses in training, they cannot race with clinically effective concentrations of medications under the British Horseracing Authority's Rules of Racing. Salmeterol must therefore be withdrawn prior to race day and pharmacokinetic (PK) studies used to establish formal detection time advice. Salmeterol xinafoate (Serevent Evohaler® ) was administered (0.1 mg twice daily for 4.5 days) via inhalation to six horses. Urine and blood samples were taken up to 103 h postadministration. Hydrolysed samples were extracted using solid phase extraction. A sensitive Ultra high performance tandem mass spectrometry (UPLC-MS/MS) method was developed, with a Lower limit of quantification (LLOQ) for salmeterol of 10 pg/mL in both matrices. The majority of salmeterol plasma concentrations, postlast administration, were below the method LLOQ and so unusable for PK analysis. Urine PK analysis suggested a half-life consistent with duration of pharmacological effect. Average estimated urine concentration at steady-state was obtained via PK modelling and used to estimate a urine concentration of 59 ± 34 pg/mL as a marker of effective lung concentration. From this, potential detection times were calculated using a range of safety factors.

Detection and pharmacokinetics of salbutamol in thoroughbred racehorses following inhaled administration.

Salbutamol sulphate (Ventolin Evohaler) was administrated via the inhalation route to six horses at a dose of 0.5 mg every 4 h during the day for 2 days (total dose 4 mg). Urine and blood samples were taken up to 92 h postadministration. Hydrolyzed plasma and urine were extracted using solid phase extraction (SPE). A sensitive tandem mass spectrometric method was developed in this study, achieving a lower limit of quantification (LLOQ) for salbutamol of 10 pg/mL in plasma and urine. The parent drug was identified using UPLC-MS/MS. Most of the determined salbutamol plasma concentrations, post last administration, lie below the LLOQ of the method and so cannot be used for plasma PK analysis. Urine PK analysis suggests a half-life consistent with the pharmacological effect duration. An estimate of the urine average concentration at steady-state was collected by averaging the concentration measurements in the dosing period from -12 to 0 h relative to the last administered dose. The value was averaged across the six horses and used to estimate an effective urine concentration as a marker of effective lung concentration. The value estimated was 9.6 ng/mL and from this a number of detection times were calculated using a range of safety factors.

A review of analytical strategies for the detection of 'endogenous' steroid abuse in food production.

Detection of the abuse of synthetic steroids in food production is nowadays relatively straightforward using modern techniques such as gas or liquid chromatography coupled to mass spectrometry (GC-MS/MS or LC-MS/MS, respectively). However, proving the abuse of 'endogenous' (or naturally occurring) steroids is more difficult. Despite these difficulties, significant progress in this area has recently been made and a number of methods are now available. The aim of the current review was to systematically review the available analytical approaches, which include threshold concentrations, qualitative 'marker' metabolites, intact steroid esters, gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS), longitudinal testing and 'omics' biomarker profiling. The advantages/disadvantages of these methods are considered in detail, but the choice of which to adopt is dictated by a number of practical, political, and economic factors, which vary in different parts of the world. These include the steroid/species combination requiring analysis, the matrix tested, whether samples are collected from live or slaughtered animals, available analytical instrumentation, sample throughput/cost, and the relevant legal/regulatory frameworks. Furthermore, these approaches could be combined in a range of different parallel and/or sequential screening/confirmatory testing streams, with the final choice being determined by the aforementioned considerations. Despite these advances, more work is required to refine the different techniques and to respond to the ever increasing list of compounds classified as 'endogenous'. At this advanced stage, however, it is now more important than ever for scientists and regulators from across the world to communicate and collaborate in order to harmonize and streamline research efforts.

Impact of the emergence of designer drugs upon sports doping testing.

Historically, dope-testing methods have been developed to target specific and known threats to the integrity of sport. Traditionally, the source of new analytical targets for which testing was required were derived almost exclusively from the pharmaceutical industry. More recently, the emergence of designer drugs, such as tetrahydrogestrinone that are specifically intended to evade detection, or novel chemicals intended to circumvent laws controlling the sale and distribution of recreational drugs, such as anabolic steroids, stimulants and cannabinoids, have become a significant issue. In this review, we shall consider the emergence of designer drugs and the response of dope-testing laboratories to these new threats, in particular developments in analytical methods, instrumentation and research intended to detect their abuse, and we consider the likely future impact of these approaches.

Detection of endogenous steroid abuse in cattle: results from population studies in the UK.

The use of steroids as growth-promoting agents in food production is banned under European Union legislation. Detecting the abuse of testosterone, nandrolone, boldenone, oestradiol and progesterone is complicated by the fact that these steroids are known to be endogenous in certain situations. In this study, the concentrations of characteristic metabolites of each of these steroids were quantified in populations of untreated steers and heifers. Steroid concentration population data were then used by a statistical model (the Chebyshev inequality) to produce threshold concentrations for screening and confirming the abuse of these steroids in steer and non-pregnant heifer urine. In addition to thresholds based on testing one animal (a '1 out of 1' approach), new methods based on testing multiple animals from a herd (a 'y out of n' approach) allowed threshold concentrations to be significantly reduced and hence false compliances to be minimised. In the majority of cases, the suggested thresholds were found to be capable of confirming the abuse of endogenous steroids in steers and heifers. In the case of oestradiol abuse in the female, however, confirmation based on a threshold is not possible and alternative methods such as gas chromatography-combustion-isotope ratio mass spectrometry are required. In addition to the steer and heifer populations, a small number of pregnant animals were also tested, yielding insights into the biosynthetic pathways of some of the steroids.

Presence and metabolism of endogenous androgenic-anabolic steroid hormones in meat-producing animals: a review.

The presence and metabolism of endogenous steroid hormones in meat-producing animals has been the subject of much research over the past 40 years. While significant data are available, no comprehensive review has yet been performed. Species considered in this review are bovine, porcine, ovine, equine, caprine and cervine, while steroid hormones include the androgenic-anabolic steroids testosterone, nandrolone and boldenone, as well as their precursors and metabolites. Information on endogenous steroid hormone concentrations is primarily useful in two ways: (1) in relation to pathological versus 'normal' physiology and (2) in relation to the detection of the illegal abuse of these hormones in residue surveillance programmes. Since the major focus of this review is on the detection of steroids abuse in animal production, the information gathered to date is used to guide future research. A major deficiency in much of the existing published literature is the lack of standardization and formal validation of experimental approach. Key articles are cited that highlight the huge variation in reported steroid concentrations that can result when samples are analysed by different laboratories under different conditions. These deficiencies are in most cases so fundamental that it is difficult to make reliable comparisons between data sets and hence it is currently impossible to recommend definitive detection strategies. Standardization of the experimental approach would need to involve common experimental protocols and collaboratively validated analytical methods. In particular, standardization would need to cover everything from the demographic of the animal population studied, the method of sample collection and storage (especially the need to sample live versus slaughter sampling since the two methods of surveillance have very different requirements, particularly temporally), sample preparation technique (including mode of extraction, hydrolysis and derivatization), the end-point analytical detection technique, validation protocols, and the statistical methods applied to the resulting data. Although efforts are already underway (at HFL and LABERCA) to produce more definitive data and promote communication among the scientific community on this issue, the convening of a formal European Union working party is recommended.

Modulation of the growth hormone-insulin-like growth factor (GH-IGF) axis by pharmaceutical, nutraceutical and environmental xenobiotics: an emerging role for xenobiotic-metabolizing enzymes and the transcription factors regulating their expression. A review.

The growth hormone-insulin-like growth factor (GH-IGF) axis has gained considerable focus over recent years. One cause of this increased interest is due to a correlation of age-related decline in plasma GH/IGF levels with age-related degenerative processes, and it has led to the prescribing of GH replacement therapy by some practitioners. On the other hand, however, research has also focused on the pro-carcinogenic effects of high GH-IGF levels, providing strong impetus for finding regimes that reduce its activity. Whereas the effects of GH/IGF activity on the action of xenobiotic-metabolizing enzyme systems is reasonably well appreciated, the effects of xenobiotic exposure on the GH-IGF axis has not received substantial review. Relevant xenobiotics are derived from pharmaceutical, nutraceutical and environmental exposure, and many of the mechanisms involved are highly complex in nature, not easily predictable from existing in vitro tests and do not always predict well from in vivo animal models. After a review of the human and animal in vivo and in vitro literature, a framework for considering the different levels of direct and indirect modulation by xenobiotics is developed herein, and areas that still require further investigation are highlighted, i.e. the actions of common endocrine disruptors such as pesticides and phytoestrogens, as well as the role of xenobiotic-metabolizing enzymes and the transcription factors regulating their expression. It is anticipated that a fuller appreciation of the existing human paradigms for GH-IGF axis modulation gained through this review may help explain some of the variation in levels of plasma IGF-1 and its binding proteins in the population, aid in the prescription of particular dietary regimens to certain individuals such as those with particular medical conditions, guide the direction of long-term drug/nutraceutical safety trials, and stimulate ideas for future research. It also serves to warn athletes that using compounds touted as performance enhancing because they promote short-term GH release could in fact be detrimental to performance in the long-run.