CAMAG DBS Laboratory, Muttenz, Switzerland.
Institute of Forensic Medicine Bern, University of Bern, Bern, Switzerland.
Drug Test Anal. 2020 Aug;12(8):1126-1134. doi: 10.1002/dta.2819. Epub 2020 Jun 30.
The World Anti-Doping Agency (WADA) and the International Testing Agency (ITA) recently announced the development and implementation of dried blood spot (DBS) testing for routine analysis in time for the 2022 Winter Olympic and Paralympic Games in Beijing. Following the introduction of a ban on the use of tramadol in competition in March 2019, the Union Cycliste International (UCI) started a pilot study for the manual analysis of tramadol in DBS for antidoping purposes. In this context, we present a fully automated LC-MS/MS-based method with automated sample preparation using a CAMAG DBS-MS 500 for the analysis of tramadol and its metabolite O-desmethyltramadol in DBS. The presented approach reduces manual handling in the laboratory to an absolute minimum, only requiring the preparation of calibration and quality control DBS cards. The method was developed, optimized, and validated before performing cross-validation with a liquid blood-based analysis method using authentic samples from forensic cases. During the validation process, the method showed an extraction efficiency of 62%, linearity r > 0.99, accuracy and precision (within ± 15% and ± 20% at the LLOQ) for the determination of tramadol and O-desmethyltramadol. Method comparison in liquid blood with 26 samples showed good agreement (90 ± 19% for tramadol and 94 ± 14% for O-desmethyltramadol). In conclusion, automated analysis of tramadol and O-desmethyltramadol in DBS provides a fast and accurate solution for antidoping screening. It is suited for high-throughput analysis, having a run time of about 4 min per sample. Furthermore, with the automated approach, manual sample extraction becomes obsolete.
世界反兴奋剂机构（WADA）和国际检测机构（ITA）最近宣布开发和实施干血斑（DBS）检测，以便在北京 2022 年冬季奥运会和冬残奥会期间进行常规分析。在 2019 年 3 月禁止曲马多在比赛中使用后，国际自行车联盟（UCI）开始了一项针对 DBS 中曲马多及其代谢物 O-去甲曲马多进行抗兴奋剂目的的手动分析的试点研究。在这种情况下，我们提出了一种完全自动化的基于 LC-MS/MS 的方法，该方法使用 CAMAG DBS-MS 500 进行自动化样品制备，用于分析 DBS 中的曲马多及其代谢物 O-去甲曲马多。所提出的方法将实验室中的手动处理减少到最低限度，仅需要制备校准和质控 DBS 卡。该方法在使用来自法医案例的真实样品进行液体血基分析方法的交叉验证之前进行了开发、优化和验证。在验证过程中，该方法显示出 62%的提取效率、线性度 r>0.99、准确度和精密度（在LLOQ 处为±15%和±20%），用于曲马多和 O-去甲曲马多的测定。在 26 个样品的液体血中进行方法比较显示出良好的一致性（曲马多为 90±19%，O-去甲曲马多为 94±14%）。总之，DBS 中曲马多和 O-去甲曲马多的自动分析为兴奋剂筛查提供了快速准确的解决方案。它适合高通量分析，每个样品的运行时间约为 4 分钟。此外，采用自动化方法，手动样品提取变得多余。
