Fast and Sensitive Screening of Oxandrolone and Its Major Metabolite 17-Epi-Oxandrolone in Human Urine by UHPLC-MS/MS with On-Line SPE Sample Pretreatment.

Oxandrolone, a synthetic testosterone analog, is used for the treatment of several diseases associated with weight loss. Unfortunately, oxandrolone is abused by many athletes and bodybuilders due to its strong anabolic effect. We have developed and validated a highly sensitive and rapid on-line SPE-UHPLC-MS/MS method for the determination of oxandrolone and simultaneous identification of its major metabolite 17-epi-oxandrolone in urine matrices. Enrichment of the analytes via an integrated solid-phase extraction was achieved using an Acquity UPLC BEH C18 Column. Subsequently, the chromatographic separation of the on-line preconcentrated sample fraction was achieved using an Acquity HSS T3 C18 Column. For the structural identification of these analytes, a high-resolution mass spectrometer Synapt-G2Si coupled to the Acquity M-class nano-LC system with ionKey source was used. A highly sensitive determination of oxandrolone was achieved using a tandem quadrupole mass spectrometer XEVO TQD. The method was successfully validated in the linear range of oxandrolone from 81.63 pg·mL-1 (limit of quantification, LOQ) to 5000 pg·mL-1 in the human urine matrix. It was applied to the analysis of real urine samples obtained from a healthy volunteer after the oral administration of one dose (10 mg) of oxandrolone. Concentration vs. time dependence was tested in the time interval of 4 h-12 days (after oral administration) to demonstrate the ability of the method to detect the renal elimination of oxandrolone from the human body. Favorable performance parameters along with successful application indicate the usefulness of the proposed method for its routine use in antidoping control labs.

Development of an ultra-high performance liquid chromatography-tandem mass spectrometry method for the determination of anabolic steroids currently available on the black market in the Czech Republic and Slovakia.

Over the past decade, the use of anabolic androgenic steroids (AAS) by recreational bodybuilders and regular gym members has been increasing exponentially. However, there is a lack of knowledge on AAS products sold in the Czech Republic and Slovakia. The aim of this study was to assess the quantity of active substances in AAS products obtained from AAS users in the Czech Republic and Slovakia. In addition, the study also examines the current trends in counterfeit AAS products used by recreational bodybuilders and regular gym members. For this purpose, the authors developed and validated a universal ultra-high performance liquid chromatography with tandem mass spectrometry method to determine the most popular doping substances in different pharmaceutical formulations (oil-based injectables, tablets, and capsules). This method was successfully utilized for the analysis of 358 AAS products. Our results showed that 58.9% AAS products analyzed contained the declared active substances at the declared concentrations, 15.9% contained no active substance, 16.8% were under-concentrated, 4.5% contained active substances different from those declared on the label, and 3.6% products were over-concentrated. Alarmingly, the results demonstrated that over 40% of the AAS analyzed failed to meet label claims and therefore may pose serious health risks to consumers. This study also highlighted that the availability of AAS should be more rigorously restricted and their quality closely monitored in order to protect AAS users. In conclusion, the authors have developed a precise, accurate, sensitive, selective, and robust method for the routine screening of products containing anabolic androgenic steroids.

Development of the fast, simple and fully validated high performance liquid chromatographic method with diode array detector for quantification of testosterone esters in an oil-based injectable dosage form.

Counterfeit steroids are available on the black market, ultimately to consumers who believe they are buying a legitimate pharmaceutical item from the labeled company. In many cases, counterfeit steroids can contain lower doses or some products can be overdosed. This can unwittingly expose users to a significant health risks. The mixture of testosterone propionate, phenylpropionate, isocaproate and decanoate in an oil-based injectable dosage form belongs to the one of the most misused illicit drugs by a variety of athletes. This study developed a new, fast, simple and reliable HPLC method combined with a simple sample preparation step to determine testosterone propionate, phenylpropionate, isocaproate and decanoate in an oil-based injectable dosage form without the use of sophisticated and expensive instrumentation. The developed analytical procedure provides high throughput of samples where LC analysis takes only 6min and sample preparation of oil matrix in one step takes approximately 10min with precision ranging from 1.03 to 3.38% (RSD), and accuracy (relative error %) within ±2.01%. This method was found to be precise, linear, accurate, sensitive, selective and robust for routine application in screening of commercial pharmaceutical products based on content of mentioned testosterone esters in their oil-based injectable dosage form for counterfeit drugs. This method was successfully applied to the analysis of nine samples of commercial testosterone mixtures purchased from various sources and will be further used as an effective screening method for determination of previously mentioned testosterone esters in samples confiscated by Institute of Forensic Science (Slovakia) during the illegal trade.