TB500

TB500/TB1000 and SGF1000: A scientific approach for a better understanding of misbranded and adulterated drugs

Nowadays, numerous websites attempt to commercialize over the internet various products, regardless of the lack of approval by the EMA or the FDA either for human or veterinary use. These products are often produced after aborted drug development due to insufficient or deleterious biological effects, synthesized based on natural products, or only based on scientific literature. However, the administration of such products is dangerous, considering the lack of official control over the production of these substances and the absence of approval by health authorities. In this short communication, we provide an extensive analysis of three misbranded and adulterated products sold over the internet named TB500, TB1000, and SGF1000. We confirm that the content of TB500/TB1000 products is not systematically consistent with it's former descriptions, but also that SGF1000 is mainly composed of sheep extracellular matrix (ECM) and blood proteins, and the signal corresponding to the purported growth promoters is excessively diluted.

Multi-analyte screening of small peptides by alkaline pre-activated solid phase extraction coupled with liquid chromatography-high resolution mass spectrometry in doping controls

Peptidic drugs with wide spectrum of physiological activity are of interest for cheating athletes and can be misused as doping in sports. A growing number of small peptide drugs capable of enhancing performance are included in the prohibited list issued by World Anti-Doping Agency (WADA), therefore the improvement of the detection methods is constantly needed. In the present study, a screening assay was developed comprising 54 prohibited small peptides and the related substances in urine by means of the alkaline pre-activated weak cation exchange-solid phase extraction (WCX-SPE) with liquid chromatography-high resolution mass spectrometry (LCHRMS). This method performed good enrichment and purification effect of traditional WCX for basic peptides, and also improve the purification power of acidic peptides, which significantly expanded the coverage of detection substances. The method was validated in accordance with WADA relevant criteria and validated with a main focus on qualitative parameters including selectivity, limits of detection (0.02-0.2 ng/mL), linearity (0.1-20 ng/mL for 46 analytes and 0.2-20 ng/mL for 9 analytes), accuracy and precision (RE% and RSD% < 20% at 1, 5 and 10 ng/mL), recovery (39.2%-100.1% except for the TB500(1-2) free acid: 9.2%), matrix effects (ion suppression effect: 0 to 49.4% and ion enhancement effect: 100% and 264.6%), carryover, reliability and sample extract stability. As a proof-of-principle, urine samples from two patients received a single injection of leuprorelin acetate microspheres (3.75 mg) 30 days before were analyzed and the results proved the applicability of the method.

Analytical approaches for the detection of emerging therapeutics and non-approved drugs in human doping controls

The number and diversity of potentially performance-enhancing substances is continuously growing, fueled by new pharmaceutical developments but also by the inventiveness and, at the same time, unscrupulousness of black-market (designer) drug producers and providers. In terms of sports drug testing, this situation necessitates reactive as well as proactive research and expansion of the analytical armamentarium to ensure timely, adequate, and comprehensive doping controls. This review summarizes literature published over the past 5 years on new drug entities, discontinued therapeutics, and 'tailored' compounds classified as doping agents according to the regulations of the World Anti-Doping Agency, with particular attention to analytical strategies enabling their detection in human blood or urine. Among these compounds, low- and high-molecular mass substances of peptidic (e.g. modified insulin-like growth factor-1, TB-500, hematide/peginesatide, growth hormone releasing peptides, AOD-9604, etc.) and non-peptidic (selective androgen receptor modulators, hypoxia-inducible factor stabilizers, siRNA, S-107 and ARM036/aladorian, etc.) as well as inorganic (cobalt) nature are considered and discussed in terms of specific requirements originating from physicochemical properties, concentration levels, metabolism, and their amenability for chromatographic-mass spectrometric or alternative detection methods.

Adsorption effects of the doping relevant peptides Insulin Lispro, Synachten, TB-500 and GHRP 5

The tendency of peptides to adsorb to surfaces can raise a concern in variety of analytical fields where the qualitative/quantitative measurement of low concentration analytes (ng/mL-pg/mL) is required. To demonstrate the importance of using the optimal glassware/plasticware, four doping relevant model peptides (GHRP 5, TB-500, Insulin Lispro, Synachten) were chosen and their recovery from various surfaces were evaluated. Our experiments showed that choosing expensive consumables with low-bind characteristics is not beneficial in all cases. A careful selection of the consumables based on the evaluation of the physico/chemical features of the peptide is recommended.

In vitro models for metabolic studies of small peptide hormones in sport drug testing

Peptide hormones represent an emerging class of potential doping agents. Detection of their misuse is difficult due to their short half-life in plasma and rapid elimination. Therefore, investigating their metabolism can improve detectability. Unfortunately, pharmacokinetic studies with human volunteers are often not allowed because of ethical constraints, and therefore alternative models are needed. This study was performed in order to evaluate in vitro models (human liver microsomes and S9 fraction) for the prediction of the metabolism of peptidic doping agents and to compare them with the established models. The peptides that were investigated include desmopressin, TB-500, GHRP-2, GHRP-6, hexarelin, LHRH and leuprolide. Several metabolites were detected for each peptide after incubation with human liver microsomes, S9 fraction, and serum, which all showed endopeptidase and exopeptidase activity. In vitro models from different organs (liver vs. kidney) were compared, but no significant differences were recorded. Deamidation was not observed in any of the models and was therefore evaluated by incubation with α-chymotrypsin. In conclusion, in vitro models are useful tools for forensic and clinical analysts to detect peptidic metabolic markers in biological fluids.