Abstract
Stanozolol is still the most commonly used illicit anabolic-androgenic steroid (AAS) in professional sports. Therefore, accurate and fast analysis and long detection windows are of great interest in the field of antidoping analysis. In this work, a very simple, fast, and highly sensitive online solid-phase extraction method coupled with liquid chromatography-high-resolution tandem mass spectrometry (HPLC-HRMSMS) for the analysis of stanozolol-N-glucuronides was developed. This fully validated procedure is characterized by only a few manual steps (dilution and addition of internal standard) in the sample preparation. A limit of identification (LOI) of 75 pg/mL, high accuracy (87.1%-102.1%), precision (3.1%-7.8%), and sensitivity was achieved. Furthermore, good linearity (> 0.99) and robustness, as well as no carry-over effects, could be observed. In addition to excellent confirmation analysis performance, this method shows sufficient potential for the identification and characterization of unknown metabolites. Using this method, it was possible to unambiguously confirm the presence of 1'N- and 2'N-stanozolol-glucuronide in human urine for the first time due to the access to reference material.
Highlights
Since the beginning of doping analysis, knowledge about the window of opportunity for the detection of illicit substances has been of great interest
When the MS-range was set to m/z 505.2908 which corresponds to the theoretical mass of [stanozolol-mono-glucuronide + H] +, the results showed a perfect match for both retention times and fullHRMS spectra for both metabolites
We introduce a novel, highly functional analytical method for the analysis of stanozolol-glucuronides for doping control analysis
Summary
Since the beginning of doping analysis, knowledge about the window of opportunity for the detection of illicit substances has been of great interest. Conjugates using liquid chromatography–tandem mass spectrometry (LC–MS/MS), is a highly promising approach for the detection of unknown steroid metabolites, respectively LTMs.[7,8,9,10,11,12,13,14,15] In most cases, phase-II metabolites are excreted in urine as glucuronide- and/or sulfate-conjugates. Both conjugates lead to an increase in water solubility, allowing urinary excretion.[16,17,18,19]
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