UPLC-QTOF-MS法研究甘油三酯作为水生环境中农药污染的潜在生物标志物

doi:10.7503/cjcu20190615

高等学校化学学报 ›› 2020, Vol. 41 ›› Issue (3): 431.doi: 10.7503/cjcu20190615

• 庆祝《高等学校化学学报》复刊40周年专栏 • 上一篇下一篇

UPLC-QTOF-MS法研究甘油三酯作为水生环境中农药污染的潜在生物标志物

何聿¹,夏倩¹,王文莉¹,罗芳²,陈金凤³,郭雅婷¹,王建¹,林振宇^1,^*,陈国南^1,^*

^1. 福州大学食品安全与生物分析教育部重点实验室
^2. 生物科学与工程学院, 福州 350116

收稿日期:2019-11-27 出版日期:2020-03-10 发布日期:2020-01-07
通讯作者: 林振宇,陈国南
作者简介:林振宇, 男, 博士, 教授, 博士生导师, 主要从事电化学方面的研究. E-mail: zylin@fzu.edu.cn|陈国南, 男, 博士, 教授, 博士生导师, 主要从事生物分析化学方面的研究. E-mail: gnchen@fzu.edu.cn
基金资助:
国家自然科学基金(Nos.21575025);国家自然科学基金(Nos.21575027);国家自然科学基金(Nos.21705025);国家自然科学基金(Nos.91843301);福建省自然科学基金资助(Nos.2018J01687);福建省自然科学基金资助.(Nos.2018J05039)

Triglycerides as Potential Biomarkers for Pesticides Pollution in Aquatic Environment Using UPLC-QTOF-MS ^†

HE Yu¹,XIA Qian¹,WANG Wenli¹,LUO Fang²,CHEN Jinfeng³,GUO Yating¹,WANG Jian¹,LIN Zhenyu^1,^*,CHEN Guonan^1,^*

^1. Key Laboratory for Analytical Science of Food Safety and Biology, Ministry of Education, 2. College of Biological Science and Engineering, Fuzhou University, Fuzhou 350116, China
^3. China Certification & Inspection Group Fujian Co. Ltd., Fuzhou 350116, China

Received:2019-11-27 Online:2020-03-10 Published:2020-01-07
Contact: Zhenyu LIN,Guonan CHEN
Supported by:
† Supported by the National Natural Science Foundation of China(Nos.21575025);† Supported by the National Natural Science Foundation of China(Nos.21575027);† Supported by the National Natural Science Foundation of China(Nos.21705025);† Supported by the National Natural Science Foundation of China(Nos.91843301);the Natural Science Foundation of Fujian Province, China(Nos.2018J01687);the Natural Science Foundation of Fujian Province, China(Nos.2018J05039)

摘要/Abstract

摘要：

以鲫鱼为代表性研究对象, 采用超高效液相色谱-四极杆飞行时间质谱(UPLC-QTOF-MS)技术, 分别研究了3种农药(敌百虫、高效氯氰菊酯和吡虫啉)对其体内代谢的影响. 通过Masslynx软件在MS ^E模式下采集数据, 使用UNIFI软件进行自动检测和数据过滤, 并将差异性物质与在线代谢组数据库进行比较, 鉴定出甘油三酯类(TGs)物质具有明显差异. 其中, 吡虫啉对TGs代谢的影响最大, 且肝脏的代谢差异比大脑更显著, 低浓度的急性暴露诱导肝脏中的TGs积累, 其含量与暴露浓度之间的关系符合Michaelis-Menten方程的增长趋势. 结果表明, TGs可在短时间内(≤2 h)灵敏地反映农药吡虫啉的低浓度(≤20 ng/mL)暴露, 可作为考察吡虫啉对水生生物毒性的潜在生物标记物, 有助于建立一种快速、灵敏的预警方法.

关键词: 超高效液相色谱-四极杆飞行时间质谱, 甘油三酯, 生物标记物, 农药污染

Abstract:

Pesticides exert a long-term negative influence on the aquatic organism and even the aquatic environment. It is necessary to find some potential biomarkers to give insight into the pesticides pollution and provide a rapid, sensitive, early-warning method. In this study, ultra-performance liquid chromatography with quadrupole-time-of-flight mass spectrometry(UPLC-QTOF-MS) was applied to analyze the effect of pesticides on the metabolomics in liver and brain of Crucian carp. Three pesticides, namely trichlorfon, β-cypermethrin and imidacloprid were investigated, respectively. Data was collected by masslynx software in MS ^E mode, analyzed by UNIFI software for automatic detection and data filtering, compared to the online metabolome database. The metabolic difference of liver was more significant than that of the brain. The triglycerides(TGs) species were identified as the different metabolites. Among these three pesticides, imidacloprid had the most significant influence on the metabolism of TGs. The low concentration of acute exposure induced the accumulation of TGs in the liver, and the relationship between the content and the exposure concentration was in line with the growth trend of Michaelis-Menten equation. The results showed that TGs can sensitively reflect the low concentration exposure of imidacloprid pesticides(≤ 20 ng/mL) in a short period of time(≤ 2 h), which can be used as a potential biomarker to investigate the toxicity of imidacloprid to aquatic organisms, and help to establish a new method for estimation the pesticides contamination in aquatic environment.

Key words: Ultra-performance liquid chromatography with quadrupole-time-of-flight mass spectrometry(UPLC-QTOF-MS), Triglycerides, Biomarker, Pesticides pollution

中图分类号:

O657.63

TrendMD:

何聿, 夏倩, 王文莉, 罗芳, 陈金凤, 郭雅婷, 王建, 林振宇, 陈国南. UPLC-QTOF-MS法研究甘油三酯作为水生环境中农药污染的潜在生物标志物. 高等学校化学学报, 2020, 41(3): 431.

HE Yu, XIA Qian, WANG Wenli, LUO Fang, CHEN Jinfeng, GUO Yating, WANG Jian, LIN Zhenyu, CHEN Guonan. Triglycerides as Potential Biomarkers for Pesticides Pollution in Aquatic Environment Using UPLC-QTOF-MS ^†. Chem. J. Chinese Universities, 2020, 41(3): 431.

图/表 8

Fig.1 Total ion chromatograms(TIC) of brain(A) and liver tissues(B) (A1, B1) Blank; (A2, B2) exposure in trichlorfon; (A3, B3) exposure in β-cypermethrin; (A4, B4) exposure in imidacloprid.

Fig.2 Volcano plots of endogenous metabolites(n=12, *P<0.05) (A1) Trichlorfon-treated brain; (A2) trichlorfon-treated liver; (B1) β-cypermethrin-treated brain; (B2) β-cypermethrin-treated liver; (C1) imidacloprid-treated brain; (C2) imidacloprid-treated liver.

Fig.3 Binary contrast spectra of brain(A) and liver(B) tissues (A1) Base peak intensity(BPI) of brain exposed for 0 and 2 h; (A2) chromatogram of difference in brain; (A3) mass spectrum of difference in brain; (B1) BPI of liver exposed for 0 and 2 h; (B2) chromatogram of difference in liver; (B3) mass spectrum of difference in liver.

Table 1 Information of representative components with high responses in brain and liver samples exposed to imidacloprid(0 and 2 h)

Sample	Component, m/z	Unknown intensity/ counts	Reference intensity/ counts	Unknown/ reference	Observed retention time/min
In brain	806.5728	3092718	3191399	0.9691	1.37
	896.7709	2516408	2318847	1.0852	1.24
	870.7539	1748903	1984113	0.8815	1.23
	898.7881	1546268	1502813	1.0289	1.54
	832.5871	1376215	1468047	0.9374	1.33
	872.7726	1266039	1342851	0.9428	1.54
In liver	896.7709	4236852	287846	14.7192	1.22
	898.7881	3388133	181398	18.6779	1.52
	872.7726	2467030	158469	15.5679	1.52
	900.8013	2209446	227011	9.7328	1.79
	874.7853	2057944	266732	7.7153	1.79
	806.5728	3340418	3378909	0.9886	1.37
	832.5871	2278588	2178444	1.046	1.33
	146.1175	119687	124507	1.056	2.99

Fig.4 Effect of exposure time on endogenous metabolite(m/z 896.7709) (A) BPI of endogenous metabolite(m/z 896.7709). Exposure time/h: a. 0; b. 2; c. 4; d. 6; e. 8; f. 10; (B) mass spectrum of endogenous metabolite(m/z 896.7709); (C) effect of exposure time on peak area of endogenous metabolite(m/z 896.7709)(n=12, *P<0.05).

Table 2 Summary of information on different endogenous metabolites mainly lipids detected in ESI positive ion mode of UPLC-QTOF-MS mass spectrometry

Experimental accurate m/z	Theoretical m/z	10⁶δ	Chemical formula	Lipid name	Ion
806.5728	806.5694	4	C₄₆H₈₀NO₈P	PC(38:6)	[M+H]⁺
832.5871	832.5851	2	C₄₈H₈₂NO₈P	PC(40:7)	[M+H]⁺
872.7726	872.7702	3	C₅₅H₉₈O₆	TG(52:4)	[M+NH₄]⁺
874.7867	874.7858	1	C₅₅H₁₀₀O₆	TG(52:3)	[M+NH₄]⁺
896.7709	896.7702	1	C₅₇H₉₈O₆	TG(54:6)	[M+NH₄]⁺
898.7881	898.7858	3	C₅₇H₁₀₀O₆	TG(54:5)	[M+NH₄]⁺
900.8013	900.8015	0	C₅₇H₁₀₂O₆	TG(54:4)	[M+NH₄]⁺

Fig.5 Comparison in detected MS/MS spectra and predicted MS/MS spectra in HMDB (A) Detected MS/MS spectrum of m/z 806.5728; (B) detected MS/MS spectrum of m/z 896.7709; (C) predicted MS/MS spectrum of m/z 806.5728; (D) predicted MS/MS spectrum of m/z 896.7709.

Fig.6 Effect of exposure concentration(n=12, *P<0.05) (A) Heatmap of the metabolites in fish liver exposed to different concentrations(0, 1, 2, 5, 10, 20 ng/mL) of imidacloprid for 2 h. Each concentration is represented in columns, and each metabolites in lines. Blue indicates low content, whereas red indicates high content; (B) Michaelis Menten equation fitting curve of m/z 896.7709 change with exposure concentration.

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UPLC-QTOF-MS法研究甘油三酯作为水生环境中农药污染的潜在生物标志物

Triglycerides as Potential Biomarkers for Pesticides Pollution in Aquatic Environment Using UPLC-QTOF-MS ^†

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[1]	黄玉, 谷彩云, 吴翰钟, 夏晓爽, 李新. 基于超高效液相色谱-四极杆飞行时间质谱的缺血性脑卒中的代谢组学研究[J]. 高等学校化学学报, 2017, 38(10): 1742.
[2]	余艺华, 孙君坦, 何炳林, 顾汉卿. 一种新型甘油三酯吸附剂的制备及其对血清中甘油三酯的吸附性能研究[J]. 高等学校化学学报, 1997, 18(10): 1729.

UPLC-QTOF-MS法研究甘油三酯作为水生环境中农药污染的潜在生物标志物

Triglycerides as Potential Biomarkers for Pesticides Pollution in Aquatic Environment Using UPLC-QTOF-MS †

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Triglycerides as Potential Biomarkers for Pesticides Pollution in Aquatic Environment Using UPLC-QTOF-MS ^†