In vitro metabolic studies of REV-ERB agonists SR9009 and SR9011

SR9009 and SR9011 are attractive as performance-enhancing substances due to their REV-ERB agonist effects and thus circadian rhythm modulation activity. Although no pharmaceutical preparations are available yet, illicit use of SR9009 and SR9011 for doping purposes can be anticipated, especially since SR9009 is marketed in illicit products. Therefore, the aim was to identify potential diagnostic metabolites via in vitro metabolic studies to ensure effective (doping) control. The presence of SR9009 could be demonstrated in a black market product purchased over the Internet. Via human liver microsomal metabolic assays, eight metabolites were detected for SR9009 and fourteen metabolites for SR9011 by liquid chromatography-high resolution mass spectrometry (LC-HRMS). Structure elucidation was performed for all metabolites by LC-HRMS product ion scans in both positive and negative ionization mode. Retrospective data analysis was applied to 1511 doping control samples previously analyzed by a full-scan LC-HRMS screening method to verify the presence of SR9009, SR9011 and their metabolites. So far, the presence of neither the parent compound nor the metabolites could be detected in routine urine samples. However, to further discourage use of these potentially harmful compounds, incorporation of SR9009 and SR9011 into screening methods is highly recommended

In Vitro Metabolic Studies of REV-ERB Agonists SR9009 and SR9011

SR9009 and SR9011 are attractive as performance-enhancing substances due to their REV-ERB agonist effects and thus circadian rhythm modulation activity. Although no pharmaceutical preparations are available yet, illicit use of SR9009 and SR9011 for doping purposes can be anticipated, especially since SR9009 is marketed in illicit products. Therefore, the aim was to identify potential diagnostic metabolites via in vitro metabolic studies to ensure effective (doping) control. The presence of SR9009 could be demonstrated in a black market product purchased over the Internet. Via human liver microsomal metabolic assays, eight metabolites were detected for SR9009 and fourteen metabolites for SR9011 by liquid chromatography-high resolution mass spectrometry (LC–HRMS). Structure elucidation was performed for all metabolites by LC–HRMS product ion scans in both positive and negative ionization mode. Retrospective data analysis was applied to 1511 doping control samples previously analyzed by a full-scan LC–HRMS screening method to verify the presence of SR9009, SR9011 and their metabolites. So far, the presence of neither the parent compound nor the metabolites could be detected in routine urine samples. However, to further discourage use of these potentially harmful compounds, incorporation of SR9009 and SR9011 into screening methods is highly recommended

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Improvements in GC-QqQ-MS screening, 1 year of experience

The previously presented screening method for doping substances using GC-MS/MS by Van Eenoo et al. has been one year in operation in our laboratory. This experience allowed us to make improvements to this method in terms of detection limits and quality control. Now the method is able to detect over 150 compounds from different classes (steroids, narcotics, stimulants, beta-2-agonists and hormone antagonists) in a qualitative way. In the quantitative part, the traditional steroid profile with most important endogenous steroids is expanded in order to improve the detection and identification of endogenous steroid misuse. Besides these also norandrosterone, salbutamol, morfine and the major metabolite of cannabis are quantified. Methods developed for anti-doping purposes should be subjected to the highest level of quality. Here, parameters to control each step in the methodology were added: hydrolysis efficiency, derivatisation efficiency and microbiological degradation are monitored in every single sample. Additionally, special attention is paid to the relation between parameters indicating degradation by micro-organisms and the reliability of the steroid profile. In this presentation, the modifications, improvements and ‘tricks’ will be presented