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GAS CHROMATOGRAPHY MASS SPECTROMETRY
Screening Techniques in Doping Analysis by GC/MS
Sports doping is not only contradictory to the concept to-volatilize drugs (Screening Method No.2), diuretics of fair play, but it has a negative impact on the health (No.5), and β-blocker agents (No.7).
of athletes as well as society in general. For these This Application News introduces an example of the reasons, drug doping testing is conducted based on analysis of a difficult-to-volatilize drug (Screening regulations imposed by the World Anti-Doping Agency Method No.2) obtained with the cooperation of MITSUBISHI KAGAKU BIO-CLINICAL LABORATORIES, Table 1 lists the sports doping screening techniques. INC., officially recognized as a WADA testing agency.
The quadrupole GC/MS is used for analysis of difficult- Table 1 Classification of Screening Methods in Sport Doping Analysis
Analytical Procedures
The pretreatment flow chart and GC/MS analytical
conditions for Screening Method No.2 are shown in In the pretreatment procedure, 6 M of hydrochloric Add internal standardAdjust pH (9.6±0.1) acid was added to 5 mL of urine, and this was heated Salting-out with anhydrous sodium nitrate for 30 minutes at 105 ºC to conduct hydrolysis. After washing with diethyl ether, 2-methyl-2-propanol and internal standards were added to the liquid phase, and Add nethyl orange / acetonitrile / TFA solution after adjusting the pH to 9.6+/-0.1, extraction was Add MSTFA until solution turns yellow, heat at conducted with diethyl ether. The extract was dried under a stream of nitrogen gas, and after adding Add MBTFA, heat at 80 ˚C for 10 min.
methyl orange/acetonitrile/TFA solution, MSTFA was added until the solution turned yellow, after which the solution was heated for 5 minutes at 80 ºC. Then, MBTFA was added, and the solution was heated for Fig.1 Pretreatment Flow for Screening Method No.2
10 minutes at 80 ºC to conduct N-TFA-O-TMS derivatization.
Table 2 Analytical Conditions
: DB-5 (15 m × 0.25 mm I.D. df=0.25 µm) : 100 ˚C (1 min)-16 ˚C/min-300 ˚C (2 min) Sports Doping Test Report Format
In order to present test results in the most effective
manner, the results of each analyte must be arranged GCMSsolution allows the reporting items to be pasted in an easy-to-view format. For instance, the report to the screen and freely positioned to easily generate must be as compact as possible, displaying highly effective doping test reports. (Fig.2) chromatograms of the drug and its metabolites side- Amphetamine and p-OH-amphetamine, and methamphetamine and p-OH-methamphetamine are drug and metabolite pairs. Placing their chromatograms side-by-side enables easy confirmation in the report.
Fig.2 Report Creation Screen
To ensure data reliability, WADA requires various pretreatment procedure and system blank.
confirmation tests. In the case of Screening Method Fig.3 shows these testing results in a report formatted No. 2, a Minimum Required Performance Limit (MRPL) using GCMSsolution. The chromatograms of the of 0.5 µg/mL (strychnine only, 0.2 µg/mL) is set to analyte target ions and their identifying ions are verify the GC/MS sensitivity.✽1 In addition, analysis of positioned one above the other, enabling convenient a control sample, consisting of drug-free urine, and a judgment of the presence or absence of the blank sample is required to ensure the reliability of the Fig.3 Example of Report Format for Sports Doping Test
Reference✽1 : MINIMUM REQUIRED PERFORMANCE LIMITS FOR DETECTION OF PROHIBITED SUBSTANCES - WADA Technical Document TD2004MRPL NOTES:
✽This Application News has been produced and edited using information that was available when the
data was acquired for each article. This Application News is subject to revision without prior notice.
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