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Teale P.,HFL Sport Science An LGC Company | Scarth J.,HFL Sport Science An LGC Company | Hudson S.,HFL Sport Science An LGC Company
Bioanalysis | Year: 2012

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. © 2012 Future Science Ltd.

Scarth J.P.,HFL Sport Science An LGC Company | Kay J.,Veterinary Medicines Directorate | Teale P.,HFL Sport Science An LGC Company | Akre C.,Canadian Food Inspection Agency | And 5 more authors.
Drug Testing and Analysis | Year: 2012

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. © Her Majesty the Queen in Right of Canada.

Clarke A.,HFL Sport Science An LGC Company | Scarth J.P.,HFL Sport Science An LGC Company | Hands J.,HFL Sport Science An LGC Company | Teale P.,HFL Sport Science An LGC Company | And 3 more authors.
Chromatographia | Year: 2011

Under European legislation, the use of anabolic steroids as growth promoters in meat production is prohibited. Currently, there is no internationally accepted method used for the detection of the potentially endogenous steroids nandrolone and boldenone in the ovine. In the current study, a multi-residue GC-MS-MS-based urinary assay has been validated for boldenone as well as the nandrolone metabolites 5α-estrane-3β,17α-diol and epinandrolone. Using a standard addition calibration line approach in pooled bovine urine, the method was linear between the endogenous concentrations and those augmented with 6,000 pg mL-1. The method was then applied to populations of wether (n = 242) and ewe (n = 237) ovine animals in order to establish urinary thresholds for detecting nandrolone and boldenone abuse. A statistical model (the Chebyshev inequality) was used to produce threshold concentrations for each analyte. Adjustment of the nandrolone metabolite data for specific gravity, a measure of the hydration status of the animal, allowed the effective thresholds to be reduced; potentially leading to a lower number of false positives. Furthermore, the proposed epinandrolone confirmatory thresholds (38,628 and 57,950 pg mL-1 in wethers and ewes, respectively) were found to be effective in detecting abuse of nandrolone for at least 1 month post-dose of this steroid. However, further studies would be required to assess the efficacy of the proposed boldenone confirmatory thresholds (19,857 and 56,080 pg mL-1 in wethers and ewes, respectively) since data on its excretion following administration to the ovine are lacking. © 2011 Springer-Verlag.

Scarth J.P.,HFL Sport Science An LGC Company | Seibert C.,HFL Sport Science An LGC Company | Brown P.R.,HFL Sport Science An LGC Company | Teale P.,HFL Sport Science An LGC Company | And 3 more authors.
Chromatographia | Year: 2011

Recombinant human erythropoietin (rhEPO) analogues are known to have been used in horse sports for their assumed performance enhancing properties. Recently, several authors have published liquid chromatographic-tandem mass spectrometric (LC-MS/MS) methods for confirming the presence of rhEPO analogues in horse plasma. In the current study, an improved LC-MS/MS confirmatory procedure for rhEPO, darbepoetin (DPO) and continuous erythropoietin receptor activator (CERA) in horse plasma was developed and validated. The method was also adapted for and applied to urine samples for the first time. Similar to previously published plasma assays, the methods utilise size exclusion and immunoaffinity extraction prior to tryptic cleavage, enzymatic deglycosylation and LC-MS/MS analysis of the resulting signature tryptic peptides (rhEPO/CERA T5, rhEPO/CERA/DPO T6 and DPO T9). However, the novel application of UPLC chromatography significantly improves the run time of the method compared to nano- or micro-LC and its robustness compared to nano-LC. Furthermore, recombinant canine EPO was found to serve as an effective internal standard, thus allowing confidence in interpretation of the success/failure of every step in the procedure. Limits of detection for confirming the presence of rhEPO, CERA and DPO in plasma were 0.1, 0.25 and 0.05 ng mL -1, respectively, which were equal to or lower than limits achieved using previously published LC-MS/MS based methods. Limits of detection for confirming the presence of rhEPO, CERA and DPO in urine were 0.05, 0.15 and 0.025 ng mL -1 and the analysis of urine samples collected from horses administered rhEPO (Eprex™) or DPO (Aranesp™) demonstrated the use of this matrix as a suitable alternative in situations where plasma is not available. © 2011 Springer-Verlag.

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