Diuretics can be misused to force diuresis to achieve weight loss or to mask the intake of a prohibited substance and are therefore prohibited by the World Anti-Doping Agency (WADA). For similar reasons other masking agents (vaptans, probenecid, etc.) are also prohibited by the WADA. The currently employed methods to detect diuretics in urine use extraction or dilute-and-shoot, combined with 1D- liquid chromatography (LC) high resolution mass spectrometry (MS) or LC-triple quadrupole MS. Dilute-and-shoot methods save time and work, but these methods encounter some problems (e.g., peak drift and matrix effect). Therefore, a 2D-LC-MS/MS application was developed, validated and evaluated as an alternative. The effect of a turbulent flow rate was studied by loading samples under different conditions and the turbulent flow rate was found to be more effective in removing matrix interferences. A correlation with the specific gravity was observed. A turbulent flow online solid phase extraction (SPE) method combined with LC-MS/MS for the detection of 50 diuretics and masking agents was developed and validated for identification purposes. This method combines the advantages of dilute-and-shoot while solving the issues of matrix effect and retention time shift. Furthermore, the presented method is compliant with WADA's identification criteria and can hence be used for screening and/or confirmation.
Keywords: Diuretics; Doping; Mass spectrometry; Turbulent flow online solid phase extraction; Two-dimensional liquid chromatography; Urine.
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