摘要
芥子气可以与血红蛋白的组氨酸侧链发生反应,生成芥子气暴露染毒的生物标志物。本研究建立了定性、定量检测痕量芥子气暴露染毒人血红蛋白加合物的方法。采用2-氯乙基乙基硫醚(2-CEES)染毒的血红蛋白溶液作为内标,用链霉蛋白酶对芥子气-珠蛋白加合物进行酶解,经PPL柱固相萃取纯化以及苄氧羰基氯(Cbz-Cl)衍生,使用超高效液相色谱-串联质谱(UHPLC-MS/MS)进行分析检测。经过优化实验条件,最终选择链霉蛋白酶的酶解温度55℃,酶解时间10.0 h,使用3.0 mL 5%甲醇水溶液淋洗,1.0 mL 50%甲醇水溶液洗脱,Cbz-Cl衍生时间30 min。结果表明,在10.0~1 000μg/L线性范围内,线性关系良好(R^2>0.997),准确度为89.8%~113%,精密度(RSD)不高于12.4%,检出限(LOD)为10.0μg/L(S/N>5)。该方法采用2对离子对进行确证,有助于痕量芥子气暴露染毒的溯源性分析,与传统酸解法相比,该方法样品处理时间短,反应条件温和、操作简单,且灵敏度可提高约150倍。
Sulfur mustard(2, 2’-dichloroethyl-sulfide, HD) is an alkylating agent and a powerful vesicant, which was widely imposed and caused the bulk of injuries and deaths in World War Ⅰ and Iran-Iraq War. Since World War Ⅱ, many casualties have been caused by the chemical weapons containing HD abandoned by Japanese in China. It still represents potential threat to military and civilian population. It’s very important to study reliable bioanalytical methods for detecting HD exposure. HD reacts with histidine residues of hemoglobin producing the HD adduct which has been used as a biomarker for retrospective detection of HD exposure because of its long half-life(about 120 days). A novel method based on the HD-hemoglobin adduct for qualitative and quantitative detection of trace HD exposure was developed. The HD-hemoglobin was isolated from blood through centrifugation and hemolyzation in water. Then the heme was removed with acidic acetone to obtain the HD-globin. HD-globin adduct was digested with pronase, purified by PPL solid phase extraction following benzyl chloroformate(Cbz-Cl) derivatization, and analyzed with ultra-high performance liquid chromatography-tandem mass spectrometry(UHPLC-MS/MS). Upon optimization, the digestion was performed at 55 ℃ for 10.0 h. The PPL loading samples was washed with 3.0 mL 5% methanol solution and eluted with 1.0 mL 50% methanol solution. The derivatization time and temperature were 30 min and 4 ℃, respectively. With 2-chloroethyl ethyl sulfide spiked hemoglobin solution serving as internal standard(ISTD), the method was found to be linear between 10.0 and 1 000 μg/L HD exposure(R^2>0.997) with precision of ≤12.4% and accuracy ranged between 89.8% and 113%. In spite of a shorter sample treatment time(compared with the acid hydrolysis, which needed reflux in hydrochloric acid at 110 ℃ for 24 h), the limit of detection(LOD) was 10.0 μg/L(S/N>5) which was 150 times lower than that by the acid hydrolysis in a previous article. Furthermore, two transitions were more benef
作者
陈博
于惠兰
刘石磊
刘昌财
梁龙辉
杨旸
吴姬娜
李晓森
CHEN Bo;YU Hui-lan;LIU Shi-lei;LIU Changcai;LIANG Long-hui;YANG Yang;WU Ji-na;LI Xiao-sen(State Key Laboratory of NBC Protection for Civilian,Beijing 102205,China;Laboratrry of Analytical Chemistry,Research Institute of Chemical Defenee,Beijing 102205,China)
出处
《质谱学报》
EI
CAS
CSCD
北大核心
2019年第4期305-313,I0001,共10页
Journal of Chinese Mass Spectrometry Society
