全血中磷脂酰乙醇的UPLC-MS/MS检测  

Detection of Phosphatidylethanol in Whole Blood by UPLC-MS/MS

作　　者：胡双英 富显果 张明 曹联君 艾克拜尔·热合曼 HU Shuang-ying;FU Xian-guo;ZHANG Ming;CAO Lian-jun;AIKEBAIER Reheman(Department of Forensic Medicine,Basic Medical College,Xinjiang Medical University,Urumqi 830011,China;Ningde Municipal Hospital,Ningde Normal University,Ningde 352100,Fujian Province,China;Institute of Criminal Science and Technology of Ningde Public Security Bureau,Ningde 352100,Fujian Province,China;Fujian Key Laboratory of Toxicant and Drug Toxicology,Medical College,Ningde Nor-mal University,Ningde 352100,Fujian Province,China)

机构地区：[1]新疆医科大学基础医学院法医教研室,新疆乌鲁木齐830011 [2]宁德师范学院附属宁德市医院,福建宁德352100 [3]宁德市公安局刑事科学技术研究所,福建宁德352100 [4]宁德师范学院医学院福建省毒物与药物毒理学重点实验室,福建宁德352100

出　　处：《法医学杂志》2022年第3期367-373,共7页Journal of Forensic Medicine

基　　金：国家自然科学基金资助项目(2186010116)。

摘　　要：目的 建立全血中乙醇代谢物磷脂酰乙醇(phosphatidylethanol,PEth)的超高效液相色谱-串联质谱(ultra-performance liquid chromatography-tandem mass spectrometry,UPLC-MS/MS)检测方法。方法 取100μL全血加入适量含1%甲酸的水溶液,用丙酮沉淀蛋白,离心后取上清液过Bond Elut Certify小柱进行纯化富集,洗脱液经过氮吹后用V∶V=1∶1混合溶液复溶后进UPLC-MS/MS检测,负电离模式下进行选择反应监测扫描,采用外标法定量分析。结果 PEth在全血样本中1~160 ng/mL质量浓度范围内与峰面积呈线性关系(r=0.999 9),检出限为0.2 ng/mL,定量限为1 ng/mL。回收率为97.43%~103.61%,准确度为0.99%~1.77%,日内精密度为0.4%~2.4%,日间精密度为1.1%~3.3%,基质效应为91.00%~99.55%。在体外血样添加乙醇的实验中均未检出PEth。在3例酒后肇事案件中PEth检测均为阳性,检出量分别为195.49、83.67和876.12 ng/mL。结论 所建立方法具有较高的灵敏度和特异性,分析结果准确,适用于全血中PEth的定性定量分析。Objective To establish the ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) method to detect ethanol metabolites phosphatidylethanol(PEth) in whole blood.Methods An appropriate amount of aqueous solution including 1% formic acid was added to 100 μL whole blood, the protein was precipitated with acetone, centrifuged and the supernatant was purified and enriched by using Bond Elut Certify column. The eluent was redissolved with 1/1 isopropanol/acetonitrile(v/v) solution after nitrogen blowing and then tested by UPLC-MS/MS. Selective reaction monitoring scanning was carried out in negative ionization mode, and quantitative analysis was performed by external standard method. Results PEth showed a linear relationship over the concentration range of 1-160 ng/mL in whole blood(r=0.999 9) with peak area. The detection limit was 0.2 ng/mL,the quantification limit was 1 ng/mL, the recovery rate was 97.43%-103.61%, the accuracy was 0.99%-1.77%, the intra-day precision was 0.4%-2.4%, and the inter-day precision was 1.1%-3.3%, and the matrix effect was 91.00%-99.55%. PEth was not detected in the in vitro blood samples supplemented with ethanol. PEth was detected positive in three drunk driving cases, and the concentration were195.49, 83.67 and 876.12 ng/mL, respectively. Conclusion The established method has high sensitivity and specificity and the analysis results are accurate. It is suitable for the qualitative and quantitative analysis of PEth in whole blood.

关 键 词：法医毒物分析 乙醇生物标志物 磷脂酰乙醇 固相萃取 超高效液相色谱-串联质谱 

分 类 号：DF795.1[医药卫生—法医学]