通过优化葡萄糖醛酸苷共轭物的酶水解,建立测定死后尿液中五种精神活性药物的 LC-MS/MS 方法。

IF 2.8 4区 医学 Q2 TOXICOLOGY
Forensic Toxicology Pub Date : 2024-07-01 Epub Date: 2024-04-01 DOI:10.1007/s11419-024-00685-1
Tomohito Matsuo, Tadashi Ogawa, Masae Iwai, Katsutoshi Kubo, Fumio Kondo, Hiroshi Seno
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引用次数: 0

摘要

目的:生物样本的毒理学分析在法医和临床调查中发挥着重要作用。摄入的药物会以与葡萄糖醛酸等内源性物质共轭的形式排泄到尿液中;水解这些共轭物可提高液相色谱-串联质谱法(LC-MS/MS)对目标药物的测定。在本研究中,我们试图改进五种精神活性药物(11-去甲-9-羧基-Δ9-四氢大麻酚、奥沙西泮、劳拉西泮、替马西泮和阿米替林)葡萄糖醛酸轭合物的酶水解:通过改变温度、酶量和反应时间来优化尿液中葡萄糖醛酸共轭物的酶水解效率。水解直接在提取柱上进行。经过充分验证后,将这种使用 LC-MS/MS 的分析方法应用于法医尸检样本:结果:我们发现,重组β-葡糖醛酸酶 B-One® 可在室温下直接在提取柱上 3 分钟内定量水解这些共轭物。这种柱上方法节省了时间,并避免了宝贵样品在转移到提取柱过程中的损失。用这种方法处理的基于 LC-MS/MS 的校准曲线显示出良好的线性关系,r2 值超过 0.998。该方法的日内和日间准确度和精密度分别为 93.0-109.7% 和 0.8-8.8%。回收率为 56.1-104.5%。基质效应为 78.9% 至 126.9%:我们建立了一种直接在萃取柱上酶水解尿液中葡萄糖醛酸苷结合物的 LC-MS/MS 方法。该方法已成功应用于法医尸检样本。该方法应用广泛,包括法医和临床毒理学研究。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Development of an LC-MS/MS method for the determination of five psychoactive drugs in postmortem urine by optimization of enzymatic hydrolysis of glucuronide conjugates.

Purpose: Toxicological analyses of biological samples play important roles in forensic and clinical investigations. Ingested drugs are excreted in urine as conjugates with endogenous substances such as glucuronic acid; hydrolyzing these conjugates improves the determination of target drugs by liquid chromatography-tandem mass spectrometry (LC-MS/MS). In this study, we sought to improve the enzymatic hydrolysis of glucuronide conjugates of five psychoactive drugs (11-nor-9-carboxy-Δ9-tetrahydrocannabinol, oxazepam, lorazepam, temazepam, and amitriptyline).

Methods: The efficiency of enzymatic hydrolysis of glucuronide conjugates in urine was optimized by varying temperature, enzyme volume, and reaction time. The hydrolysis was performed directly on extraction columns. This analysis method using LC-MS/MS was applied to forensic autopsy samples after thorough validation.

Results: We found that the recombinant β-glucuronidase B-One® quantitatively hydrolyzed these conjugates within 3 min at room temperature directly on extraction columns. This on-column method saved time and eliminated the loss of valuable samples during transfer to the extraction column. LC-MS/MS-based calibration curves processed with this method showed good linearity, with r2 values exceeding 0.998. The intra- and inter-day accuracies and precisions of the method were 93.0-109.7% and 0.8-8.8%, respectively. The recovery efficiencies were in the range of 56.1-104.5%. Matrix effects were between 78.9 and 126.9%.

Conclusions: We have established an LC-MS/MS method for five psychoactive drugs in urine after enzymatic hydrolysis of glucuronide conjugates directly on extraction columns. The method was successfully applied to forensic autopsy samples. The established method will have broad applications, including forensic and clinical toxicological investigations.

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来源期刊
Forensic Toxicology
Forensic Toxicology TOXICOLOGY-
CiteScore
5.80
自引率
9.10%
发文量
40
审稿时长
3 months
期刊介绍: The journal Forensic Toxicology provides an international forum for publication of studies on toxic substances, drugs of abuse, doping agents, chemical warfare agents, and their metabolisms and analyses, which are related to laws and ethics. It includes original articles, reviews, mini-reviews, short communications, and case reports. Although a major focus of the journal is on the development or improvement of analytical methods for the above-mentioned chemicals in human matrices, appropriate studies with animal experiments are also published. Forensic Toxicology is the official publication of the Japanese Association of Forensic Toxicology (JAFT) and is the continuation of the Japanese Journal of Forensic Toxicology (ISSN 0915-9606).
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