芥子气三肽加合物合成新路线开发及在确证分析中的应用  

Development of a New Synthetic Route of Tripeptide-adduct of Sulfur Mustard Exposure and Application in Confirmatory Analysis

作　　者：杨展 刘玉龙 陈佳 吴剑峰 何跃忠 谢剑炜 YANG Zhan;LIU Yu-Long;CHEN Jia;WU Jian-Feng;HE Yue-Zhong;XIE Jian-Wei(Academy of Military Medical Sciences,Academy of Military Sciences,Beijing 100850,China;State Key Laboratory of Toxicology and Medical Countermeasures and Laboratory of Toxicant Analysis,Institute of Pharmacology and Toxicology,Academy of Military Medical Sciences,Beijing 100850,China)

机构地区：[1]军事科学院军事医学研究院,北京100850 [2]军事科学院军事医学研究院毒物药物研究所,抗毒药物与毒理学国家重点实验室,北京100850

出　　处：《分析化学》2022年第1期136-144,I0006,I0007,共11页Chinese Journal of Analytical Chemistry

基　　金：国家重点研发计划(Nos.2018YFC1602600,2020YFF0305000)资助。

摘　　要：芥子气(Sulfur mustard,SM)是一种典型的难防难治化学战剂,生物体SM暴露后,除自身发生氧化水解代谢反应外,还会与体内的蛋白质、谷胱甘肽及DNA等生成一系列的加合物。白蛋白在血浆中丰度较高,芥子气与其34位半胱氨酸(Cys-34)结合形成加合物,再经蛋白酶K水解后生成三肽加合物(HETE-Cys-Pro-Phe,HETE-CPF)。HETE-CPF半衰期长、稳定性好,是溯源SM暴露最有效,也是最重要的生物标志物。本研究以易获取的商品化试剂氯乙醇为起始原料,开发了一条新的HETE-CPF合成方法。相比于原有以2-氯乙基巯基乙醇为起始原料的合成方法,新方法副反应更少、纯化更简便。在此基础上建立了可用于全血样本中HETE-CPF的高灵敏分析检测方法,检测灵敏度可达2 ng/mL。同时,还鉴定出大鼠经尾静脉注射SM暴露后,其与血浆白蛋白形成的暴露标志物与人源血浆差异显著,为HETE-Cys-Pro-Tyr(HETE-CPY),并非HETECPF,两者白蛋白一级序列差异是导致标志物不同的根本原因。Sulfurmustard(SM)is a typical representative of highly-toxic chemical warfare agent that is difficult to be prevented and treated.SM could produce various biomarkers with protein,glutathione and DNA,etc,besides oxidative and hydrolytic metabolism.The tripeptide-adduct hydroxyethylthioethyl-Cys-Pro-Pro(HETE-CPF)originating from Cys-34 on the human serum albumin(HSA)is a very important retrospective biomarker,which is usually used to identify SM exposure due to its long half-life and good stability.In this work,a novel synthetic route of HETE-CPF was developed with chloroethanol as the starting material.Compared with the original synthetic route using 2-chloroethyl mercaptoethanol as the starting material,the new synthetic method had less side reactions,higher yield and easier purification.Furthermore,a highly sensitive analysis method for HETE-CPF in whole blood samples was established,and the detection sensitivity was up to 2 ng/mL,which was enough to identify human plasma sample after exposure to SM.Meanwhile,it was also identified that after SM exposure via caudal vein in rats,the exposure biomarkers formed with plasma albumin were significantly different from those of human plasma,which was HETE-Cys-Pro-Tyr(HETE-CPY),not HETE-CPF.The fundamental reason might be due to their different primary sequence of albumin.

关 键 词：芥子气 固相肽合成 白蛋白加合物 高分辨质谱 

分 类 号：TJ92[兵器科学与技术—武器系统与运用工程]