质谱鸟枪法非靶向快速初筛青鱼中微囊藻毒素残留
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摘要
母离子扫描(PreIS)是一种功能强大的扫描模式,但较少受到质谱工作者关注。本实验利用PreIS的功能特性和微囊藻毒素(MCs)的分子结构特征,建立了一种鸟枪法用于非靶向快速初筛青鱼(Mylopharyngodon piceus)中MCs残留。通过优化固相萃取条件和质谱参数,选取m/z 135([C_9H_(11)O+H]~+)为特征碎片峰,快速初步定性筛查了所有含该基团的MCs。实验发现,MCs在质谱气相环境内更容易形成双电荷离子,且谱图较纯净,以[M+2H]~(2+)为主。通过方法学验证,该方法的线性关系良好,相关系数R~2介于0.992 5～0.997 1之间;灵敏度较高,检出限和定量限分别低于1.25μg/L和3.99μg/L;日内、日间精密度的相对标准偏差分别为6.8%～8.6%和7.2%～9.1%,重现性较好;方法回收率为68.2%～83.4%,RSD为4.5%～6.8%,能够满足一般分析测试的要求。采用该方法对市售青鱼进行MCs快速初筛,阳性检出率为15%,MCs总浓度为3.4～15.8μg/kg,主要检出的种类为微囊藻毒素RR(MC-RR)和LR(MC-LR)。该方法能够快速检测青鱼中MCs残留,可以为相关部门提供依据和参考,从而为消费者食品安全提供保障。
Precursor-ion scan(PreIS) is a versatile scanning mode but not commonly used by mass spectrometry analyzers. In this study, a shotgun method was developed for fast untargeted screening of MCs in Mylopharyngodon piceus(M. Piceus) on the basis of the specific functions of PreIS and the structural characteristics of MCs. The performance of this method was improved by optimizing the conditions of solid-phase extraction and parameters of mass spectrometry. Three types sorbent were compared, including florisil, HLB, and C18. The HLB(68.2%-83.4%) performed best, followed by C18(63.5%-78.5%) and florisil(ca.60%). 2% and 80% methonal were used as washing and eluting solvents, respectively. The MCs were easily ionized as doubly charged ions [M+2 H]~(2+). Under product-ion scan, all the MCs can generate fragment ions like m/z 375, 213 and 135, where m/z 135 is the most intensive one. Therefore, m/z 135 was selected for conducting PreIS of MCs. The standard solution of five MCs mixture was detected, and all the doubly charged ions of MCs were observed. Finally, the proposed shotgun method was validated in terms of linearity, sensitivity, precision and recovery. The results indicated that the calibration curves are linear with R~2 of 0.992 5-0.997 1. The LOD and LOQ are not more than 1.25 and 3.99 μg/L, respectively. The RSDs of intra-day and inter-day precision are 6.8%-8.6% and 7.2%-9.1%, respectively. All the MCs can be well recovered with recovery of 68.2%-83.4%, and RSDs of 4.5%-6.8%. This method was applied to the M. Piceus samples from local market, and 15% samples were detected to be positive. MC-RR and MC-LR are the two frequently detected MCs in these samples, with concentrations of 3.4-15.8 μg/kg. The shotgun method is robust and efficient in detecting MCs in M. Piceus. The data are important references for related functional department to ensure the food safety of domestic consumers.
Precursor-ion scan(PreIS) is a versatile scanning mode but not commonly used by mass spectrometry analyzers. In this study, a shotgun method was developed for fast untargeted screening of MCs in Mylopharyngodon piceus(M. Piceus) on the basis of the specific functions of PreIS and the structural characteristics of MCs. The performance of this method was improved by optimizing the conditions of solid-phase extraction and parameters of mass spectrometry. Three types sorbent were compared, including florisil, HLB, and C18. The HLB(68.2%-83.4%) performed best, followed by C18(63.5%-78.5%) and florisil(ca.60%). 2% and 80% methonal were used as washing and eluting solvents, respectively. The MCs were easily ionized as doubly charged ions [M+2 H]~(2+). Under product-ion scan, all the MCs can generate fragment ions like m/z 375, 213 and 135, where m/z 135 is the most intensive one. Therefore, m/z 135 was selected for conducting PreIS of MCs. The standard solution of five MCs mixture was detected, and all the doubly charged ions of MCs were observed. Finally, the proposed shotgun method was validated in terms of linearity, sensitivity, precision and recovery. The results indicated that the calibration curves are linear with R~2 of 0.992 5-0.997 1. The LOD and LOQ are not more than 1.25 and 3.99 μg/L, respectively. The RSDs of intra-day and inter-day precision are 6.8%-8.6% and 7.2%-9.1%, respectively. All the MCs can be well recovered with recovery of 68.2%-83.4%, and RSDs of 4.5%-6.8%. This method was applied to the M. Piceus samples from local market, and 15% samples were detected to be positive. MC-RR and MC-LR are the two frequently detected MCs in these samples, with concentrations of 3.4-15.8 μg/kg. The shotgun method is robust and efficient in detecting MCs in M. Piceus. The data are important references for related functional department to ensure the food safety of domestic consumers.
引文
[1] SUN S, YE J, CHEN J, et al. Effect of dietary fish oil replacement by rapeseed oil on the growth, fatty acid composition and serum non-specific immunity response of fingerling black carp, Mylopharyngodon piceus[J]. Aquaculture Nutrition, 2011, 17(4): 441-450.
[2] FAN H, LUO Y, YIN X, et al. Biogenic amine and quality changes in lightly salt-and sugar-salted black carp (Mylopharyngodon piceus) fillets stored at 4 C[J]. Food Chemistry, 2014, (159): 20-28.
[3] 苏雅玲,邓一荣. 富营养化湖泊中微囊藻毒素及其控制去除技术[J]. 环境科学与技术,2013, (6): 62-66. SU Yaling, DENG Yirong. Microcystins in eutrophic lakes and their controlling and removing methods[J]. Environmental Science & Technology, 2013, (6): 62-66(in Chinese).
[4] CHRISTIANSEN G, FASTNER J, ERHARD M, et al. Microcystin biosynthesis in Planktothrix: genes, evolution, and manipulation[J]. Journal of Bacteriology, 2003, 185(2): 564-572.
[5] SABLINA A A, CHEN W, ARROYO J D, et al. The tumor suppressor PP2A abeta regulates the ralA GTPase[J]. Cell, 2007, 129(5): 969-982.
[6] NISHIWAKI-MATSUSHIMA R, OHTA T, NISHIWAKI S, et al. Liver tumor promotion by the cyanobacterial cyclic peptide toxin microcystin-LR[J]. Journal of Cancer Research and Clinical Oncology, 1992, 118(6): 420-424.
[7] CHRISTOFFERSEN K, KAAS H. Toxic cyanobacteria in water. A guide to their public health consequences, monitoring, and management[J]. Limnology & Oceanography, 2000, 45(5): 255-258.
[8] 盛建武,何苗,施汉昌,等. 水环境中微囊藻毒素检测技术研究进展[J]. 环境污染与防治,2006,28(2):132-136. SHENG Jianwu, HE Miao, SHI Hanchang, et al. Review on detection technologies of microcystins in aquatic environment[J]. Environmental Pollution & Control, 2006, 28(2): 132-136(in Chinese).
[9] 李诗言,王扬,王鼎南,等. 通过型固相萃取-液相色谱-串联质谱法同时快速测定鱼肉中7种微囊藻毒素[J]. 色谱,2017,35(8):794-800. LI Shiyan, WANG Yang, WANG Dingnan, et al. Rapid and simultaneous determination of seven microcystins in fish meat by liquid chromatography-tandem mass spectrometry coupled with pass-through solid phase extraction[J]. Chinese Journal of Chromatography, 2017, 35(8): 794-800(in Chinese).
[10] 王蕾,李小艳,薛文通,等. 微囊藻毒素检测方法研究进展[J]. 食品科学,2005,26(增刊1):135-138. WANG Lei, LI Xiaoyan, XUE Wentong, et al. Review on research progress of microcystins detection[J]. Food Science, 2005, 26(Suppl 1): 135-138(in Chinese).
[11] KAYA K, SANO T. Total microcystin determination using erythro-2-methyl-3-(methoxy-d 3)-4-phenylbutyric acid (MMPB-d 3) as the internal standard[J]. Analytica Chimica Acta, 1999, 386(1): 107-112.
[12] 郭坚,杨新磊,叶明立. 全自动在线固相萃取-高效液相色谱法测定水体中痕量微囊藻毒素[J]. 分析化学,2011,39(8):1 256-1 260. GUO Jian, YANG Xinlei, YE Mingli. Determination of trace level microcystins in water by fully automated online solid phase extraction coupled with high performance liquid chromatography[J]. Chinese Journal of Analytical Chemistry, 2011, 39(8): 1 256-1 260(in Chinese).
[13] JIN R, LI L, GUO L, et al. A graphene tip coupled with liquid chromatography tandem mass spectrometry for the determination of four synthetic adulterants in slimming supplements[J]. Food Chemistry, 2017, (224): 329-334.
[14] SHEN Q, JIN R, XUE J, et al. Analysis of trace levels of sulfonamides in fish tissue using micro-scale pipette tip-matrix solid-phase dispersion and fast liquid chromatography tandem mass spectrometry[J]. Food Chemistry, 2016, (194): 508-515.
[15] ORTELLI D, EDDER P, COGNARD E, et al. Fast screening and quantitation of microcystins in microalgae dietary supplement products and water by liquid chromatography coupled to time of flight mass spectrometry[J]. Analytica Chimica Acta, 2008, 617(1): 230-237.
[16] 李彦文,黄献培,吴小莲,等. 固相萃取-高效液相色谱串联质谱法同时测定土壤中3种微囊藻毒素[J]. 分析化学,2013,41(1):88-92. LI Yanwen, HUANG Xianpei, WU Xiaolian, et al. Simultaneous extraction and determination of three microcystins from soil using solid phase extraction and liquid chromatography-tandem mass spectrometry[J]. Chinese Journal of Analytical Chemistry, 2013, 41(1): 88-92(in Chinese).
[17] SHEN Q, GONG L, BAIBADO J T, et al. Graphene based pipette tip solid phase extraction of marine toxins in shellfish muscle followed by UPLC-MS/MS analysis[J]. Talanta, 2013, 116(22): 770-775.
[2] FAN H, LUO Y, YIN X, et al. Biogenic amine and quality changes in lightly salt-and sugar-salted black carp (Mylopharyngodon piceus) fillets stored at 4 C[J]. Food Chemistry, 2014, (159): 20-28.
[3] 苏雅玲,邓一荣. 富营养化湖泊中微囊藻毒素及其控制去除技术[J]. 环境科学与技术,2013, (6): 62-66. SU Yaling, DENG Yirong. Microcystins in eutrophic lakes and their controlling and removing methods[J]. Environmental Science & Technology, 2013, (6): 62-66(in Chinese).
[4] CHRISTIANSEN G, FASTNER J, ERHARD M, et al. Microcystin biosynthesis in Planktothrix: genes, evolution, and manipulation[J]. Journal of Bacteriology, 2003, 185(2): 564-572.
[5] SABLINA A A, CHEN W, ARROYO J D, et al. The tumor suppressor PP2A abeta regulates the ralA GTPase[J]. Cell, 2007, 129(5): 969-982.
[6] NISHIWAKI-MATSUSHIMA R, OHTA T, NISHIWAKI S, et al. Liver tumor promotion by the cyanobacterial cyclic peptide toxin microcystin-LR[J]. Journal of Cancer Research and Clinical Oncology, 1992, 118(6): 420-424.
[7] CHRISTOFFERSEN K, KAAS H. Toxic cyanobacteria in water. A guide to their public health consequences, monitoring, and management[J]. Limnology & Oceanography, 2000, 45(5): 255-258.
[8] 盛建武,何苗,施汉昌,等. 水环境中微囊藻毒素检测技术研究进展[J]. 环境污染与防治,2006,28(2):132-136. SHENG Jianwu, HE Miao, SHI Hanchang, et al. Review on detection technologies of microcystins in aquatic environment[J]. Environmental Pollution & Control, 2006, 28(2): 132-136(in Chinese).
[9] 李诗言,王扬,王鼎南,等. 通过型固相萃取-液相色谱-串联质谱法同时快速测定鱼肉中7种微囊藻毒素[J]. 色谱,2017,35(8):794-800. LI Shiyan, WANG Yang, WANG Dingnan, et al. Rapid and simultaneous determination of seven microcystins in fish meat by liquid chromatography-tandem mass spectrometry coupled with pass-through solid phase extraction[J]. Chinese Journal of Chromatography, 2017, 35(8): 794-800(in Chinese).
[10] 王蕾,李小艳,薛文通,等. 微囊藻毒素检测方法研究进展[J]. 食品科学,2005,26(增刊1):135-138. WANG Lei, LI Xiaoyan, XUE Wentong, et al. Review on research progress of microcystins detection[J]. Food Science, 2005, 26(Suppl 1): 135-138(in Chinese).
[11] KAYA K, SANO T. Total microcystin determination using erythro-2-methyl-3-(methoxy-d 3)-4-phenylbutyric acid (MMPB-d 3) as the internal standard[J]. Analytica Chimica Acta, 1999, 386(1): 107-112.
[12] 郭坚,杨新磊,叶明立. 全自动在线固相萃取-高效液相色谱法测定水体中痕量微囊藻毒素[J]. 分析化学,2011,39(8):1 256-1 260. GUO Jian, YANG Xinlei, YE Mingli. Determination of trace level microcystins in water by fully automated online solid phase extraction coupled with high performance liquid chromatography[J]. Chinese Journal of Analytical Chemistry, 2011, 39(8): 1 256-1 260(in Chinese).
[13] JIN R, LI L, GUO L, et al. A graphene tip coupled with liquid chromatography tandem mass spectrometry for the determination of four synthetic adulterants in slimming supplements[J]. Food Chemistry, 2017, (224): 329-334.
[14] SHEN Q, JIN R, XUE J, et al. Analysis of trace levels of sulfonamides in fish tissue using micro-scale pipette tip-matrix solid-phase dispersion and fast liquid chromatography tandem mass spectrometry[J]. Food Chemistry, 2016, (194): 508-515.
[15] ORTELLI D, EDDER P, COGNARD E, et al. Fast screening and quantitation of microcystins in microalgae dietary supplement products and water by liquid chromatography coupled to time of flight mass spectrometry[J]. Analytica Chimica Acta, 2008, 617(1): 230-237.
[16] 李彦文,黄献培,吴小莲,等. 固相萃取-高效液相色谱串联质谱法同时测定土壤中3种微囊藻毒素[J]. 分析化学,2013,41(1):88-92. LI Yanwen, HUANG Xianpei, WU Xiaolian, et al. Simultaneous extraction and determination of three microcystins from soil using solid phase extraction and liquid chromatography-tandem mass spectrometry[J]. Chinese Journal of Analytical Chemistry, 2013, 41(1): 88-92(in Chinese).
[17] SHEN Q, GONG L, BAIBADO J T, et al. Graphene based pipette tip solid phase extraction of marine toxins in shellfish muscle followed by UPLC-MS/MS analysis[J]. Talanta, 2013, 116(22): 770-775.
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