Preliminary data on the potential for unintentional antidoping rule violations by permitted cannabidiol (CBD) use.

According to the World Anti-Doping Agency (WADA) regulations, cannabinoids use is prohibited in competition except for cannabidiol (CBD) use. For an adverse analytical finding (AAF) in doping control, cannabinoid misuse is based on identification of the pharmacologically inactive metabolite 11-nor-delta-9-carboxy-tetrahydrocannabinol-9-carboxylic acid (carboxy-THC) in urine at a concentration greater than 180 ng/ml. All other (minor) cannabinoids are reported as AAF when identified, except for CBD that has been explicitly excluded from the class of cannabinoids on WADA\u27s Prohibited List since 2018. However, due to the fact that CBD isolated from cannabis plants may contain additional minor cannabinoids, the permissible use of CBD can lead to unintentional violations of antidoping regulations. An assay for the detection of 16 cannabinoids in human urine was established. The sample preparation consisted of enzymatic hydrolysis of glucuronide conjugates, liquid-liquid extraction, trimethylsilylation, and analysis by gas chromatography/tandem mass spectrometry (GC-MS/MS). Spot urine samples from CBD users, as well as specimens obtained from CBD administration studies conducted with 15 commercially available CBD products, were analyzed, and assay characteristics such as selectivity, reproducibility of detection at the minimum required performance level, limit of detection, and limit of identification were determined. An ethical committee approved controlled single dose commercially available CBD products administration study was conducted to identify 16 cannabinoids in urine samples collected after ingestion or application of the CBD products as well as their presence in spot urine samples of habitual CBD users. Variable patterns of cannabinoids or their metabolites were observed in the urine samples, especially when full spectrum CBD products were consumed. The presence of minor cannabinoids or their metabolites in an athlete\u27s in-competition urine sample represents a substantial risk of an antidoping rule violation

Investigation of Testosterone, Androstenone, and Estradiol Metabolism in HepG2 Cells and Primary Culture Pig Hepatocytes and Their Effects on 17βHSD7 Gene Expression

Steroid metabolism is important in various species. The accumulation of androgen metabolite, androstenone, in pig adipose tissue is negatively associated with pork flavor, odour and makes the meat unfit for human consumption. The 17β-hydroxysteroid dehydrogenase type 7 (17βHSD7) expressed abundantly in porcine liver, and it was previously suggested to be associated with androstenone levels. Understanding the enzymes and metabolic pathways responsible for androstenone as well as other steroids metabolism is important for improving the meat quality. At the same time, metabolism of steroids is known to be species- and tissue-specific. Therefore it is important to investigate between-species variations in the hepatic steroid metabolism and to elucidate the role of 17βHSD7 in this process. Here we used an effective methodological approach, liquid chromatography coupled with mass spectrometry, to investigate species-specific metabolism of androstenone, testosterone and beta-estradiol in HepG2 cell line, and pig cultured hepatocytes. Species- and concentration-depended effect of steroids on 17βHSD7 gene expression was also investigated. It was demonstrated that the investigated steroids can regulate the 17βHSD7 gene expression in HepG2 and primary cultured porcine hepatocytes in a concentration-dependent and species-dependent pattern. Investigation of steroid metabolites demonstrated that androstenone formed a 3′-hydroxy compound 3β-hydroxy-5α-androst-16-ene. Testosterone was metabolized to 4-androstene-3,17-dione. Estrone was found as the metabolite for β-estradiol. Inhibition study with 17βHSD inhibitor apigenin showed that apigenin didn't affect androstenone metabolism. Apigenin at high concentration (50 μM) tends to inhibit testosterone metabolism but this inhibition effect was negligible. Beta-estradiol metabolism was notably inhibited with apigenin at high concentration. The study also established that the level of testosterone and β-estradiol metabolites was markedly increased after co-incubation with high concentration of apigenin. This study established that 17βHSD7 is not the key enzyme responsible for androstenone and testosterone metabolism in porcine liver cells. © 2012 Chen et al

Doping practices in international weightlifting: analysis of sanctioned athletes/support personnel from 2008 to 2019 and retesting of samples from the 2008 and 2012 Olympic Games.

Background The pervasiveness of doping and findings of anti-doping corruption threaten weightlifting's position at the 2024 Olympic Games. Analysing the practices of doping in weightlifters could identify patterns in doping that assist in future detection. Methods We analysed publicly available data on sanctioned athletes/support personnel from the International Weightlifting Federation between 2008 and 2019 and announced retrospective Anti-Doping Rule Violations (ADRVs) from the 2008 and 2012 Olympic Games. Results There were 565 sanctions between 2008 and 2019 of which 82% related to the detection of exogenous Anabolic Androgenic Steroid (AAS) metabolites and markers indicating endogenous AAS usage. The detection of exogenous AAS metabolites, markers of endogenous AAS usage and other substance metabolites varied by IWF Continental Federation (p ≤ 0.05) with Europe (74%, 11%, 15%) and Asia (70%, 15%, 15%) showing a higher detection of exogenous AAS compared to Pan America (37%, 30%, 33%) and Africa (50%, 17%, 33%). When looking at the 10 most detected substances, the nations with the highest number of sanctions (range 17-35) all had at least one overrepresented substance that accounted for 38-60% of all detected substances. The targeted re-analysis of samples from the 2008 and 2012 Olympic Games due to the discovery of long-term metabolites for exogenous AAS resulted in 61 weightlifters producing retrospective ADRVs. This includes 34 original medallists (9 gold, 10 silver and 15 bronze), the highest of any sport identified by Olympic Games sample re-testing. The exogenous AAS dehydrochloromethyltestosterone and stanozolol accounted for 83% of detected substances and were present in 95% of these samples. Conclusion Based on these findings of regional differences in doping practices, weightlifting would benefit from the targeted testing of certain regions and continuing investment in long-term sample storage as the sensitivity and specificity of detection continues to improve

Detection and localization of pectin methylesterase isoforms in pollen tubes of Nicotiana tabacum L.

Pectin methylesterases (PMEs) were detected in tobacco (Nicotiana tabacum) pollen tubes grown in vitro. Seven PME isoforms exhibiting a wide isoelectric-point (pI) range (5.3-9.1) were found in crude extracts of pollen tubes. These isoforms were mainly retrieved in supernatants after low- and high-speed separation of the crude extract. Two isoforms, with pIs 5.5 and 7.3 and molecular weight about 158 kDa, were detected by immunoblotting with anti-flax PME antiserum. Localization of pectins and PME isoforms in pollen tubes was investigated by immunogold labelling with JIM5 monoclonal antibodies and anti-flax PME antiserum, respectively. In germinated pollen grains, two PME isoforms were mainly detected in the exine, Golgi apparatus and secretory vesicles. In pollen tubes the same two PME isoforms were distributed along the outer face of the plasma membrane in the vicinity of the inner layer of the cell wall, in the Golgi and around secretory vesicles. In pollen grains, PME isoforms were, in some cases, mixed with acidic pectins m proximity to the outer surface of the plasma membrane. In pollen tubes the presence of PMEs inside secretory vesicles carrying esterified pectins supports the hypothesis that, during pollen tube growth, PMEs could be transferred by secretory vesicles in a precursor form and be activated at the tip where exocytosis takes place

Risk of unintentional antidoping rule violations by consumption of hemp products

Consumption of hemp products is continuously growing, with an expanding scope of applications. Suppliers operate through different distribution channels, but the Internet is a major retail platform. Hemp products are prepared from cannabis plants and, therefore, might contain a variety of different natural cannabinoids. According to the regulations of the World Anti-Doping Agency, all natural and synthetic cannabinoids are prohibited in-competition, with the explicit exemption of cannabidiol. Therefore, an investigation of 23 hemp products for the presence of cannabinoids was performed to determine the likelihood of unintentional violations of anti-doping regulations. An assay for the detection of 16 cannabinoids in nutritional supplements was developed and validated. The sample preparation consisted of QuEChERS extraction, trimethylsilylation, and analysis by gas chromatography/tandem mass spectrometry. All 23 commercially available hemp products were analyzed, and assay characteristics such as selectivity, limit of detection, limit of identification, limit of quantification, linearity, imprecision, recovery, and accuracy were determined. Twenty of 23 hemp products included a variety of cannabinoids at, occasionally, substantial concentrations, with four products covering the entire spectrum of tested cannabinoids. An ethics committee-approved single-dose administration study was conducted with the commercially available hemp products, investigating the presence of 16 cannabinoids in urine collected pre- and post-consumption. Variable patterns of cannabinoids or their metabolites in urine were observed. In 30% of the urine samples collected 8 h after ingestion, the presence of a prohibited cannabinoid would have resulted in an unintentional violation of anti-doping regulations

A biochemical model for neurite outgrowth during brain development

International audienc

Preliminary data on the potential for unintentional antidoping rule violations by permitted cannabidiol (CBD) use

According to the World Anti-Doping Agency (WADA) regulations, cannabinoids use is prohibited in competition except for cannabidiol (CBD) use. For an adverse analytical finding (AAF) in doping control, cannabinoid misuse is based on identification of the pharmacologically inactive metabolite 11-nor-delta-9-carboxy-tetrahydrocannabinol-9-carboxylic acid (carboxy-THC) in urine at a concentration greater than 180 ng/ml. All other (minor) cannabinoids are reported as AAF when identified, except for CBD that has been explicitly excluded from the class of cannabinoids on WADA's Prohibited List since 2018. However, due to the fact that CBD isolated from cannabis plants may contain additional minor cannabinoids, the permissible use of CBD can lead to unintentional violations of antidoping regulations. An assay for the detection of 16 cannabinoids in human urine was established. The sample preparation consisted of enzymatic hydrolysis of glucuronide conjugates, liquid-liquid extraction, trimethylsilylation, and analysis by gas chromatography/tandem mass spectrometry (GC-MS/MS). Spot urine samples from CBD users, as well as specimens obtained from CBD administration studies conducted with 15 commercially available CBD products, were analyzed, and assay characteristics such as selectivity, reproducibility of detection at the minimum required performance level, limit of detection, and limit of identification were determined. An ethical committee approved controlled single dose commercially available CBD products administration study was conducted to identify 16 cannabinoids in urine samples collected after ingestion or application of the CBD products as well as their presence in spot urine samples of habitual CBD users. Variable patterns of cannabinoids or their metabolites were observed in the urine samples, especially when full spectrum CBD products were consumed. The presence of minor cannabinoids or their metabolites in an athlete's in-competition urine sample represents a substantial risk of an antidoping rule violation