Analysis of doping control samples using supercritical fluid chromatography‐tandem mass spectrometry: Ready for routine use

Abstract

Supercritical fluid chromatography is proving to be a good separation and sample preparation tool for various analytical applications and, as such, has gained the attention of the anti‐doping community. Here, the applicability of supercritical fluid chromatography hyphenated to tandem mass spectrometry for routine doping control analysis was tested. A multi‐analyte method was developed to cover 197 drugs and metabolites that are prohibited in sport. More than 1000 samples were analyzed by applying a “dilute and inject” approach after hydrolysis of glucuronide metabolites. Additionally, a comparison with routinely used liquid chromatography‐mass spectrometry was performed with 250 of the 1000 samples and a number of past positive anti‐doping samples. It revealed some features where supercritical fluid chromatography‐tandem mass spectrometry was found to be complementary or advantageous to liquid chromatography‐mass spectrometry for anti‐doping purposes, such as better retention of analytes that are poorly retained in reversed‐phase liquid chromatography. Our results suggest that supercritical fluid chromatography‐tandem mass spectrometry is sensitive (limit of detection <50% relevant minimum required performance level required by the World Anti‐Doping Agency for anti‐doping analysis), reproducible, robust, precise (analytes of interest area coefficient of variation <5%; retention time difference coefficient of variation <1%) and complementary to existing techniques currently used for routine analysis in the World Anti‐Doping Agency accredited laboratories.

Annual banned‐substance review 16th edition—Analytical approaches in human sports drug testing 2022/2023

“…Wüst et al assessed the utility of supercritical fluid chromatography (SFC) tandem MS as an alternative and/or complementary tool for multi‐analyte ITP applications in sports drug testing 94 . Urine samples were enzymatically hydrolyzed and diluted with tetrahydrofuran prior to injection into the SFC‐ESI‐QqQ‐MS system, with the SFC being equipped with a 2‐EP analytical column (3.0 × 100 mm, 1.7 μm) and employing supercritical CO 2 and 5‐mM ammonium acetate in methanol/water (96.5/3.5, v / v ) as solvent and modifier, respectively.…”

Routine application of SFC‐MS in doping control: Analysis of 3 × 1000 urine samples using three different SFC‐MS instruments

“…Also, we undertook a semi‐quantitative calibration using four different concentrations from 25% MRPL (minimum required performance level) to 200% MRPL where we diluted with one blank urine all analytes in our quality controls (QCs; of 54 analytes). Generally, linearity was good (meeting WADA criteria) and seems to be a useful means of a WADA accredited laboratory providing an indicative estimate of concentration for assisting the results management authority's review of an anti‐doping violation 24 . A comprehensive review about the application of SFC‐MS for anti‐doping analysis has recently been provided by Parr and Botrè 26 …”

“…stimulants, glucocorticoids and β‐blockers) could easily be analysed by SFC‐MS. Furthermore, we comment on the ease of SFC‐LC switching, making the approach even more cost‐effective enabling the WADA accredited laboratory to invest in an LC and an SFC attached to the same mass spectrometer 24 …”

“…Furthermore, we comment on the ease of SFC-LC switching, making the approach even more cost-effective enabling the WADA accredited laboratory to invest in an LC and an SFC attached to the same mass spectrometer. 24 A SFC-MS screening method for the analysis of approximately 110 various drugs prohibited in sport including anabolic steroids, hormone modulators, stimulants, narcotics, synthetic cannabinoids and glucocorticoids has been published. [4][5][6][7] These authors optimised the choice of stationary phase as well as the composition of mobile phases and MS conditions to enable chromatographic separation and resolution and to ensure maximised MS response.…”

“…Generally, linearity was good (meeting WADA criteria) and seems to be a useful means of a WADA accredited laboratory providing an indicative estimate of concentration for assisting the results management authority's review of an anti-doping violation. 24 A comprehensive review about the application of SFC-MS for antidoping analysis has recently been provided by Parr and Botrè. 26 Our study shows practical reliability and readiness of SFC-MS to be used as a tool for routine sample analysis in anti-doping laboratories.…”