Detection of β-methylphenethylamine, a novel doping substance, by means of UPLC/MS/MS

Self Cite

Higenamine (Norcoclaurine) is a very popular substance in Chinese medicine and is present in many plants. The substance may be also found in supplements or nutrients, consumption of which may result in violation of anti-doping rules. Higenamine is prohibited in sport at all times and included in Class S3 (β-2-agonists) of the World Anti-Doping Agency (WADA) 2017 Prohibited List. The presence of higenamine in urine samples at concentrations greater than or equal to 10 ng/mL constitutes an adverse analytical finding (AAF). This work presents a new metabolite of higenamine in urine sample which was identified by means of ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Samples were prepared according to 2 protocols - a Dilute and Shoot (DaS) approach and a method involving acid hydrolysis and double liquid-liquid extraction (LLE). To meet the requirements typical for a confirmatory analysis, the screening procedure was further developed. In samples prepared by the DaS method, 2 peaks were observed; the earlier one was specific for higenamine and the later one unknown. MS scan analysis showed mass about 80 Da higher than that of higenamine. In turn, in samples prepared in accordance with the protocol involving hydrolysis, an increase in the area under peak for higenamine was observed, while the second peak was absent. It seems that the described strategy of detection of higenamine in urine avoids false negative results.

“…Urine samples were prepared by 2 methods: dilute and shoot (DaS) and a method involving acid hydrolysis and double extraction described previously, with modification.…”

Section: Methodsmentioning

confidence: 99%

Analysis for higenamine in urine by means of ultra‐high‐performance liquid chromatography–tandem mass spectrometry: Interpretation of results

Grucza

Kwiatkowska

Kowalczyk

et al. 2017

Self Cite

“…13,14 2.3 | Dilute-and-shoot approach Briefly, for DaS, 100 μL of urine was spiked with 5 μL of Amphetamine-D11 (internal standard) at final concentration of 500 ng/mL and diluted with 900 μL of water. 13,14 2.3 | Dilute-and-shoot approach Briefly, for DaS, 100 μL of urine was spiked with 5 μL of Amphetamine-D11 (internal standard) at final concentration of 500 ng/mL and diluted with 900 μL of water.…”

Section: Sample Preparationmentioning

confidence: 99%

Detection of bemitil and its metabolite in urine by means of LC–MS/MS in view of doping control analysis

Kwiatkowska

Kowalczyk

Grucza

et al. 2018

Self Cite

2-(Ethylthio) benzimidazole is an active ingredient of Antihot, a dietary supplement sold in Ukraine. The substance, available also under names of Bemitil, Metaprot, and Bemaktor, was developed in the USSR in 1970s, and after tests on Soviet cosmonauts and soldiers, several studies on its influence on athletes' performances were conducted. The research showed that bemitil is a synthetic adaptogen which is capable to significantly increase physical performance and reduce the time of regeneration.Moreover, according to supplement's distributor, the substance improves both physical performance and resistance to stress. Taking into account these properties, it appears plausible that the World Anti-Doping Agency (WADA) decided to include bemitil in its 2018 monitoring program. To select markers of bemitil use, six doses of the supplement (two per day, on three consecutive days) were administrated to six healthy volunteers (three men, three women, 26-49 years). Urine samples were collected before, during and up to 30 days after the first ingestion. Samples were analyzed by means of ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The study revealed that bemitil can be traced in urine as either a parent compound or its glucuronide conjugate, which is more abundant and has a wider detection window. KEYWORDS 2-ethylsulfanyl-1H-benzimidazole (WADA), 2-(Ethylthio)benzimidazol (ANTIHOT), anti-doping analysis, Bemitil, excretion study

“…[50] There have been reports of several cases of violations by athletes in various sporting events. [101] Four urine samples found positive for β-MePEA were used to evaluate the impact of glucuronide and sulfate deconjugation on method performance. [101] Four urine samples found positive for β-MePEA were used to evaluate the impact of glucuronide and sulfate deconjugation on method performance.…”

Section: Synthetic Phenethylamines In Dietary Supplementsmentioning

confidence: 99%

“…[99,100] Cholbinski et al reported the development and application of a simple and rapid analytical procedure for urine samples that enables discrimination between β-MePEA and amphetamine, its positional isomer. [101] This work is expected to be of significance in the area of forensic analysis of anti-doping samples because it provides a validated procedure for discriminating between amphetamine and β-MePEA. [101] This work is expected to be of significance in the area of forensic analysis of anti-doping samples because it provides a validated procedure for discriminating between amphetamine and β-MePEA.…”

Section: Synthetic Phenethylamines In Dietary Supplementsmentioning

confidence: 99%

Overview of regulation of dietary supplements in the USA and issues of adulteration with phenethylamines (PEAs)

Pawar

Grundel

2016

The multi-billion dollar dietary supplement industry is global in reach. The industry has been criticized for problems related to poor quality control, safety, misbranding, and adulteration. In this review, we describe how the US Food and Drug Administration (FDA) regulates dietary supplements within the framework of the Federal Food, Drug, and Cosmetic Act (FD&C Act). The Dietary Supplement Health and Education Act of 1994 (DSHEA), which amended the FD&C Act, gave the FDA the authority to promulgate Good Manufacturing Practices for dietary supplements and required that manufacturers provide the FDA information supporting a conclusion that the ingredients are reasonably expected to be safe if the dietary ingredients were not marketed in the USA before 15 October 1994. Recent amendments to the FD&C Act require that serious dietary-supplement-related adverse events be reported to the FDA and provide the agency with mandatory recall authority. We discuss the presence of naturally occurring (e.g. Ephedra, Citrus aurantium, Acacia) and synthetic (e.g. β-methylphenethylamines, methylsynephrine, α-ethyl-phenethylamine) biologically active phenethylamines (PEAs) in dietary supplements and of PEA drugs (e.g. clenbuterol, fenfluramine, sibutramine, lorcaserin) in weight-loss products. Regulatory actions against manufacturers of products labelled as dietary supplements that contain the aliphatic amines 1,3-dimethylamine and 1,3-dimethylbutylamine, and PEAs such as β-methylphenethylamine, aegeline, and Dendrobium illustrate the FDA's use of its authority under the FD&C Act to promote dietary supplement safety. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.