‘Wrong-way-round ionization’ and screening for doping substances in human urine by high-performance liquid chromatography/orbitrap mass spectrometry

Moscow Antidoping Center, 105005, Moscow, Elizavetynsky10.
Journal of Mass Spectrometry (Impact Factor: 2.38). 03/2012; 47(3):381-91. DOI: 10.1002/jms.2055
Source: PubMed


To free analytical resources for new classes of doping substances, such as banned proteins, maximization of the number of compounds that can be determined with high sensitivity in a single run is highly urgent. This study demonstrates an application of 'wrong-way-round ionization' for the simultaneous detection of multiple classes of doping substances without the need to switch the polarity. A screening method for the detection of 137 compounds from various classes of prohibited substances (stimulants, diuretics, β(2)-agonists, β-blockers, antiestrogens, glucocorticosteroids and anabolic agents) has been developed. The method involves an enzymatic hydrolysis, liquid-liquid extraction and detection by liquid chromatography/orbitrap mass spectrometry with wrong-way-round ionization. Up to 64% of compounds had a 10-fold lower limit of detection (LOD) than the minimum required performance limit. To compare the efficiency of conventional ionization relative to wrong-way-round ionization of doping substances in + ESI, a fortified blank urine sample at the minimum required performance limit was analyzed using two ESI approaches. All compounds were detected with markedly better S/N in a high-pH mobile phase, with the exception of acetazolamide (minimal change in S/N, < 20%).The method was validated by spiking 10 different blank urine samples at five different concentrations. Validation parameters included the LOD, selectivity, ion suppression, extraction recovery and repeatability.

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