Qualitative Screening and Quantitative Analysis of 110 Pesticides in Blood by Ultra-High Performance Liquid Chromatography- Quadrupole-Time of Flight Mass Spectrometry
摘 要
建立了超高效液相色谱-四极杆-飞行时间质谱法(UPLC-Q-TOF/MS)同时分析血液中110种农药的方法。在2 mL离心管中,加入200 μL血液样品和1 000 μL甲醇,混合均匀,涡旋振荡2 min,离心10 min,取上清液在优化的仪器工作条件下进行测定。采用Kinetex Biphenyl色谱柱为固定相,以5 mmol·L-1甲酸铵溶液(A)和含5 mmol·L-1甲酸铵的甲醇溶液(B)的混合溶液为流动相进行梯度洗脱。在电喷雾离子源正离子(ESI+)模式和信息依赖采集(IDA)模式下,依次采集一级和二级质谱信息。采用Master View软件通过质量误差、保留时间误差、差异同位素比、谱库相似程度等4个因子获得的综合得分来进行定性筛查;采用Multi Quant软件通过提取离子流(XIC)色谱图中的前体离子的峰面积进行定量分析。结果表明:110种目标农药化合物的综合得分为81.8~97.0分,所得结果均大于定性筛查的最低限(80分)。110种目标农药化合物的质量浓度在5~200 μg·L-1内与其对应的色谱峰面积呈线性关系,相关系数在0.99以上,检出限(3S/N)为2 μg·L-1。以空白血为基质进行3个浓度水平的加标回收试验,目标农药的回收率为80.3%~120%,测定值的相对标准偏差(n=6)为1.9%~20%。以混合标准溶液和用空白基质配制的同浓度的混合标准溶液中各农药的峰面积的比值评估基质效应,110种农药的基质效应为80.65%~119.86%。采用此方法分析了2起农药中毒案件中的血液样品,分别检出了敌敌畏、毒死蜱、氯氰菊酯(案件1)和阿特拉津、氯氟氰菊酯、克百威(案件2),其定性筛查中4个因子的综合得分均大于80分,质量浓度分别为183.98,156.79,104.67 μg·L-1和144.57,165.32,138.91 μg·L-1。
Abstract
A method of ultra-high performance liquid chromatography-quadrupole-time of flight mass spectrometry (UPLC-Q-TOF/MS) for simultaneous analysis of 110 pesticides in blood was established. 200 μL blood sample and 1 000 μL methanol were mixed in a 2 mL centrifuge tube, and the mixture was vortexed for 2 min to have the target extracted. After centrifuging for 10 min, the supernatant was analyzed under the optimized instrumental conditions. A Kinetex Biphenyl chromatographic column was used as the stationary phase, and a mixed solution containing of 5 mmol·L-1 ammonium formate solution (A) and methanol solution containing 5 mmol·L-1 ammonium formate (B) was used as the mobile phase for gradient elution. Under the mode of ESI+ and IDA, primary and secondary mass spectrometry information were collected in sequence. Qualitative screening was performed using Master View software through a combined score obtained from 4 factors including mass error, retention time error, isotope ratio difference and library similarity; Multi Quant software was used for quantitative analysis by the peak area of the precursor ion in XIC chromatogram. The results showed that the combined scores of 110 target pesticides were ranged from 81.8 to 97.0 points, all above the minimum score of qualitative screening (80 point). The mass concentrations of 110 target pesticides were linearly related to the corresponding chromatographic peak areas in the range of 5-200 μg·L-1, and the correlation coefficients were all above 0.99. The limits of detection (3S/N) found were same of 2 μg·L-1. The spiked recovery test was made by using the blank blood as matrix at 3 concentration levels, giving the recoveries of the target pesticides in the range of 80.3%-120%, and RSDs (n=6) of the determined values were ranged from 1.9% to 20%. The matrix effects were evaluated by the ratio of the peak area of the same compound with the same mass concentration between the standard solution and the blank matrix matching solution, and matrix effects of 110 pesticides were found in the range of 80.65%-119.86%. Blood samples from two pesticide poisoning cases were analyzed by the present method, and dichlorvos, chlorpyrifos, cypermethrin (for case 1) and atrazine, cyfluthrin, carbofuran (for case 2) were detected, with combined scores all above 80 point, and their mass concentrations were 183.98, 156.79, 104.67 μg·L-1 and 144.57, 165.32, 138.91 μg·L-1, respectively.
中图分类号 O657.63 DOI 10.11973/lhjy-hx202008002
所属栏目 工作简报
基金项目 国家重点研发计划项目(2017YFC0803504)
收稿日期 2019/10/24
修改稿日期
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备注杜秋瑶,硕士研究生,研究方向为毒物毒品分析
引用该论文: DU Qiuyao,ZHANG Yunfeng,WANG Jifen,WANG Lu,CHANG Jing,DONG Linpei,WU Xiaojun,LIU Bingjie. Qualitative Screening and Quantitative Analysis of 110 Pesticides in Blood by Ultra-High Performance Liquid Chromatography- Quadrupole-Time of Flight Mass Spectrometry[J]. Physical Testing and Chemical Analysis part B:Chemical Analysis, 2020, 56(8): 851~864
杜秋瑶,张云峰,王继芬,王璐,常靖,董林沛,吴小军,刘冰洁. 超高效液相色谱-四极杆-飞行时间质谱法定性筛查和定量分析血液中110种农药[J]. 理化检验-化学分册, 2020, 56(8): 851~864
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【2】赵森,张云峰,王炯,等.超高效液相色谱质谱联用法检测季铵盐类农药的研究[J].湖北警官学院学报, 2012(12):173-174.
【3】侯睿,张云峰,王炯,等.常见季铵盐类农药分析检测方法研究进展[J].刑事技术, 2010(6):36-39.
【4】HERNÁNDEZ F, SANCHO J V, IBÁÑEZ M, et al. Investigation of pesticide metabolites in food and water by LC-TOF-MS[J]. TrAC Trends in Analytical Chemistry, 2008,27(10):862-872.
【5】FERRER I, THURMAN E M, FERNÁNDEZ-ALBAA R. Quantitation and accurate mass analysis of pesticides in vegetables by LC/TOF-MS[J]. Analytical Chemistry, 2005,77(9):2818-2825.
【6】李春苗,胡雪艳,范春林,等.GC-Q-TOF/MS快速筛查测定半夏中322种农药及化学污染物残留[J].分析测试学报, 2017,36(4):431-448.
【7】周闰,吴林林,刘文卫.基于QuEChERS方法的血液中30种农药的气相色谱-质谱快速检测法[J].现代预防医学, 2019,46(13):2448-2454.
【8】ROSANO T G, OHOUO P Y, WOOD M. Application of high-resolution UPLC-MSE/TOF confirmation in forensic urine drug screening by UPLC-MS/MS[J]. Journal of Analytical Toxicology, 2019,43(5):353-363.
【9】ROSANO T G, WOOD M, IHENETU K, et al. Drug screening in medical examiner casework by high-resolution mass spectrometry (UPLC-MSE-TOF)[J]. Journal of Analytical Toxicology, 2013,37(8):580-593.
【10】常靖,王芳琳,张泽楠,等.法医毒物检验中毒物的快速筛查与检测研究进展[J].中国法医学杂志, 2015,30(1):49-51.
【11】郭杰,许秋实,石辉丽,等.利用Q-TOFLC/MS建立二级谱图数据库快速检测90种农药的初步研究[J].刑事技术, 2013(4):41-45.
【12】GUVVALA V, CHIDAMBARAM S V, ANIREDDY J S. A study on structural characterization of degradation products of cangrelor using LC/QTOF/MS/MS and NMR[J]. Journal of Pharmaceutical and Biomedical Analysis, 2019,170:327-334.
【13】SHI M G, GAO T T, ZHANG T, et al. Characterization of curcumin metabolites in rats by ultra-high-performance liquid chromatography with electrospray ionization quadrupole time-of-flight tandem mass spectrometry[J]. Rapid Communications in Mass Spectrometry, 2019,33(13):1114-1121.
【14】WARD P, CHADHA M, CONNOLLY C, et al. A comparative assessment of water-soluble selenium metabolites in commercial selenised yeast supplements by liquid chromatography-electrospray ionisation QTOF-MS[J]. International Journal of Mass Spectrometry, 2019,439:42-52.
【15】XIA Y G, GONG F Q, GUO X D, et al. Rapid screening and characterization of triterpene saponins in Acanthopanax senticosus leaves via untargeted MSAll and SWATH techniques on a quadrupole time of flight mass spectrometry[J]. Journal of Pharmaceutical and Biomedical Analysis, 2019,170:68-82.
【16】AZRAD M, FREIDUS V, KASSEM R, et al. Identification of dermatophytes by MALDI-TOF MS technology in the clinical laboratory[J]. International Journal of Mass Spectrometry, 2019,440:32-36.
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【18】陈威,吴军辉,万娟,等.QuEChERS-LC-ESI-Q-TOF快速筛查稻米中59种农药残留[J].食品工业, 2018,39(1):312-317.
【19】GUAN W B, LI C S, LIU X, et al. Graphene as dispersive solidphase extraction materials for pesticides LC-MS/MS multi-residue analysis in leek, onion and garlic[J]. Food Additives & Contaminants:Part A, 2014,31(2):250-261.
【20】LI X H, WANG M M, ZHAO J, et al. Ultrasound-assisted emulsification liquid phase microextraction method based on deep eutectic solvent as extraction solvent for determination of five pesticides in traditional Chinese medicine[J]. Journal of Pharmaceutical and Biomedical Analysis, 2019,166:213-221.
【21】邓慧芬,张建莹,黄科,等.QuEChERS-液相色谱-串联质谱法测定蔬菜中105种农药残留[J].色谱, 2018,36(12):1211-1222.
【22】WANG X B, ZHENG J, LI J J. et al. Simultaneous analysis of 23 illegal adulterated aphrodisiac chemical ingredients in health foods and Chinese traditional patent medicines by ultrahigh performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry[J]. Journal of Food and Drug Analysis, 2018,26(3):1138-1153.
【23】HE Z Y, XU Y P, ZHANG Y W, et al. On the use of in-source fragmentation in ultrahigh-performance liquid chromatography-electrospray ionization-high-resolution mass spectrometry for pesticide residue analysis[J]. Journal of Agricultural and Food Chemistry, 2019,67(38):10800-10812.
【24】LOWRY J, TRUVER M T, SWORTWOOD M J. Quantification of seven novel synthetic opioids in blood using LC-MS/MS[J]. Forensic Toxicology, 2019,37(1):215-223.
【25】MATUSZEWSKI B K, CONSTANZER M L, CHAVEZ-ENG C M. Strategies for the assessment of matrix effect in quantitative bioanalytical methods based on HPLC-MS/MS[J]. Analytical Chemistry, 2003,75(13):3019-3030.
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