Department of Forensic Medicine, Wroclaw Medical University, Wroclaw, Poland.
Institute of Toxicology Research, Borowa, Poland.
J Forensic Sci. 2021 May;66(3):1003-1012. doi: 10.1111/1556-4029.14674. Epub 2021 Jan 29.
The paper presents a method for the determination of methadone, EDDP, and EMDP in postmortem biological materials using liquid-liquid extraction with ethyl acetate (pH9) and UHPLC-MS/MS technique. Methadone-d and EDDP-d were used as the internal standards. The method validation results for blood and urine were as follows: linearity: 0.5-1000 ng/ml; R > 0.9993 for methadone, EDDP and R > 0.9944 for EMDP. Intra- and inter-day precision: 0.1%-7.5% and 0.3%-8.6%, respectively; intra- and inter-day accuracy: -11.8% to 13.9% and -9.3 to 14.8%, respectively; recovery: 91.5%-123.0%; matrix effect: 83.5%-123.9%. This study also describes 18 postmortem cases, where methadone concentrations ranged 2.3-1180 ng/ml in blood (n = 17), from 11.0 to >10,000 ng/ml in urine (n = 13) and 135.2-409.0 in vitreous humor (VH, n = 3). EDDP concentrations ranged from not detectable to 180 ng/mL in blood, from 42.4 to >10,000 ng/ml in urine and 18.3-36.5 in VH. EMDP concentrations were found in four cases in blood from below LLOQ to 1.8 ng/ml and in seven cases in urine, ranged 2.1-243.0 ng/ml. EMDP was not detected in VH samples. The EDDP/methadone ratios and blood/urine ratios for methadone and EDDP in EMDP-positive and negative cases were performed. The paper presents mass spectra of other methadone metabolites, than EDDP and EMDP (ring hydroxylated methadone, ring hydroxylated EDDP, ring hydroxylated EMDP, methadol, and DDP). Simultaneous determination of methadone and its metabolites in order to unequivocally interpret the results of toxicological tests seems to be useful in cases related to prescription/illicit use of methadone.
本文提出了一种使用乙酸乙酯（pH9）液液萃取和 UHPLC-MS/MS 技术测定死后生物材料中甲氨蝶呤、EDDP 和 EMDP 的方法。甲氨蝶呤-d 和 EDDP-d 被用作内标。血液和尿液的方法验证结果如下：线性：0.5-1000ng/ml；甲氨蝶呤、EDDP 的 R>0.9993，而 EMDP 的 R>0.9944。日内和日间精密度：分别为 0.1%-7.5%和 0.3%-8.6%；日内和日间准确度：分别为-11.8%至 13.9%和-9.3%至 14.8%；回收率：91.5%-123.0%；基质效应：83.5%-123.9%。本研究还描述了 18 例死后案例，其中血液中甲氨蝶呤浓度范围为 2.3-1180ng/ml（n=17），尿液中甲氨蝶呤浓度范围为 11.0 至>10,000ng/ml（n=13），玻璃体液（VH，n=3）中浓度为 135.2-409.0。EDDP 浓度在血液中从不可检测到 180ng/mL，在尿液中从 42.4 至>10,000ng/ml，在 VH 中从 18.3 至 36.5。在血液中，有 4 例低于LLOQ 至 1.8ng/ml，有 7 例尿液中甲氨蝶啶阳性。在 VH 样本中未检测到 EMDP。对 EMDP 阳性和阴性病例中甲氨蝶呤和 EDDP 的 EDDP/甲氨蝶呤比值和血液/尿液比值进行了分析。本文还介绍了甲氨蝶呤的其他代谢物，如 EDDP 和 EMDP（环羟化甲氨蝶呤、环羟化 EDDP、环羟化 EMDP、甲多醇和 DDP）的质谱图。为了在与处方/非法使用甲氨蝶呤有关的病例中明确解释毒理学测试的结果，似乎有必要同时测定甲氨蝶呤及其代谢物。
