The advent of hundreds of new compounds aimed at the substance misuse market has posed new analytical challenges. A semi-quantitative liquid chromatography–high-resolution mass spectrometry (LC–HRMS) method has been developed to detect exposure to two novel stimulants, mephedrone and ethylphenidate, and selected metabolites. Centrifuged urine (50 µL) was diluted with LC eluent containing internal standards (mephedrone-d₃, methylphenidate-d₉ and ritalinic acid-d₁₀; all 0.02 mg/L) (450 µL). Intra- and inter-assay accuracy and precision were within ±15% and <6%, respectively, for all analytes. The limit of detection was 0.01 mg/L for all analytes. Urine samples from mephedrone and ethylphenidate users were analyzed using immunoassay (amphetamine-group cloned enzyme donor immunoassay [CEDIA]) and LC–HRMS. Ethylphenidate, mephedrone and selected metabolites all had low cross-reactivity (<1%) with the immunoassay. The median (range) amphetamine-group CEDIA concentration in urine samples from mephedrone users (n = 11) was 0.30 (<0.041–3.04) mg/L, with only 1 sample giving a positive CEDIA result. The amphetamine-group CEDIA concentration in the urine sample from an ethylphenidate user was <0.041 mg/L. Improving the detection of novel compounds is of increasing importance to enable accurate diagnosis and treatment. Immunoassay methods used for drug screening may be inappropriate and lead to false-negative results. Conversely, detection of these compounds is possible through use of LC–HRMS and can provide information on the metabolites present after exposure to these drugs.

中文翻译：

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免疫分析和液相色谱-高分辨率质谱分析法检测新型兴奋剂：哌醋甲酯和甲氧麻黄酮的案例研究

针对物质滥用市场的数百种新化合物的出现提出了新的分析挑战。已开发出一种半定量液相色谱-高分辨率质谱（LC-HRMS）方法，以检测与两种新型兴奋剂，甲氧麻黄酮和哌醋甲酯以及某些代谢物的接触情况。离心尿液（50 µL）用含有内标（甲氧麻黄酮-d ₃，哌醋甲酯-d ₉和利尿酸-d ₁₀）的LC洗脱液稀释。; 全部0.02 mg / L）（450 µL）。所有分析物的批内和批间准确性和精密度分别在±15％和<6％之内。所有分析物的检出限为0.01 mg / L。使用免疫测定法（安非他明组克隆的酶供体免疫测定法[CEDIA]）和LC-HRMS分析了甲氧麻黄酮和哌醋甲酯使用者的尿液样本。免疫分析结果显示，哌醋甲酯，甲氧麻黄酮和某些代谢物的交叉反应性均较低（<1％）。甲氧麻黄酮使用者尿液样本中苯丙胺类CEDIA浓度的中位数（范围）（n = 11）为0.30（<0.041–3.04）mg / L，只有1个样品的CEDIA结果为阳性。哌醋乙酯使用者尿液样本中的苯丙胺类CEDIA浓度<0.041 mg / L。改善新化合物的检测对于实现准确的诊断和治疗越来越重要。用于药物筛选的免疫测定方法可能不合适，并会导致假阴性结果。相反，通过使用LC-HRMS可以检测这些化合物，并且可以提供接触这些药物后存在的代谢物的信息。