A detailed description is given of the development and validation of a fully automated in-line solid-phase extraction-liquid chromatography-tandem mass spectrometry (SPE-LC–MS/MS) method capable of detecting 90 central-stimulating new psychoactive substances (NPS) and 5 conventional amphetamine-type stimulants (amphetamine, 3,4-methylenedioxy-methamphetamine (MDMA), 3,4-methylenedioxy-amphetamine (MDA), 3,4-methylenedioxy-N-ethyl-amphetamine (MDEA), methamphetamine) in serum. The aim was to apply the validated method to forensic samples. The preparation of 150 μL of serum was performed by an Instrument Top Sample Preparation (ITSP)-SPE with mixed mode cation exchanger cartridges. The extracts were directly injected into an LC–MS/MS system, using a biphenyl column and gradient elution with 2 mM ammonium formate/0.1% formic acid and acetonitrile/0.1% formic acid as mobile phases. The chromatographic run time amounts to 9.3 min (including re-equilibration). The total cycle time is 11 min, due to the interlacing between sample preparation and analysis. The method was fully validated using 69 NPS and five conventional amphetamine-type stimulants, according to the guidelines of the Society of Toxicological and Forensic Chemistry (GTFCh). The guidelines were fully achieved for 62 analytes (with a limit of detection (LOD) between 0.2 and 4 μg/L), whilst full validation was not feasible for the remaining 12 analytes. For the fully validated analytes, the method achieved linearity in the 5 μg/L (lower limit of quantification, LLOQ) to 250 μg/L range (coefficients of determination > 0.99). Recoveries for 69 of these compounds were greater than 50%, with relative standard deviations ≤ 15%. The validated method was then tested for its capability in detecting a further 21 NPS, thus totalling 95 tested substances. An LOD between 0.4 and 1.6 μg/L was obtained for these 21 additional qualitatively-measured substances. The method was subsequently successfully applied to 28 specimens from routine forensic case work, of which 7 samples were determined to be positive for NPS consumption.

详细描述了可检测90种中央刺激性新型精神活性物质（NPS）的全自动在线固相萃取-液相色谱-串联质谱（SPE-LC-MS / MS）方法的开发和验证）和5种常规的苯丙胺类兴奋剂（苯丙胺，3,4-亚甲二氧基-甲基苯丙胺（MDMA），3,4-亚甲二氧基-苯异丙胺（MDA），3,4-亚甲二氧基-N-乙基苯丙胺（甲基苯丙胺））在血清中。目的是将经过验证的方法应用于法医样品。通过带有混合模式阳离子交换剂小柱的仪器顶部样品制备（ITSP）-SPE进行150μL血清的制备。将萃取物直接注射到LC-MS / MS系统中，使用联苯柱并用2 mM甲酸铵/0.1%甲酸和乙腈/ 0进行梯度洗脱。1％甲酸作为流动相。色谱运行时间为9.3分钟（包括重新平衡）。由于样品制备和分析之间的交错，总的循环时间为11分钟。根据毒理学和法医化学学会（GTFCh）的指南，使用69 NPS和5种常规苯丙胺类兴奋剂对方法进行了充分验证。对于62种分析物（检测限（LOD）在0.2至4μg/ L之间）完全达到了指南，而对其余12种分析物进行完全验证是不可行的。对于经过充分验证的分析物，该方法在5μg/ L（定量下限，LLOQ）至250μg/ L范围（测定系数> 0.99）内实现了线性。这些化合物中有69种的回收率大于50％，相对标准偏差≤15％。然后对经过验证的方法测试其检测另外21种NPS的能力，从而总共测试了95种物质。对于这21种其他定性测量的物质，LOD在0.4至1.6μg/ L之间。该方法随后成功应用于例行法医案件中的28个标本，其中7个样品被确定为NPS消耗呈阳性。