Determination of Amitraz and Its Metabolites in Honey by LC-MS/MS
摘 要
称取蜂蜜样品2.00 g,加入同位素内标40 ng,用pH 7的磷酸二氢钾-磷酸氢二钠缓冲溶液溶解并稀释至10.0 mL。充分混匀后,加入乙腈10.0 mL和氯化钠2.0 g,离心涡旋提取5 min。取上层乙腈相保留待用,于下层水相中再加入乙腈10 mL重复提取1次。合并两次提取液,加乙腈定容至20.0 mL。分取此提取液1.0 mL,加入于己预置N-丙基乙二胺(PSA)25 mg、十八烷基硅烷(C18)10 mg和MgSO4 30 mg的离心管中,充分混匀后,高速离心,进行分散固相萃取(dSPE)净化。转移全部上清液,加水定容至2.0 mL,此溶液供液相色谱-串联质谱法(LC-MS/MS)分析。色谱分离中采用XDB-C18色谱柱为固定相,用不同比例的φ 0.15%甲酸溶液(A)和乙腈(B)的混合液作为流动相进行梯度淋洗。所得各洗脱液按工作条件进行MS/MS测定双甲脒及其代谢物单甲脒、2,4-二甲基苯基甲酰胺和2,4-二甲基苯胺的残留量。由于采用空白蜂蜜作基体加入混合标准溶液制作工作曲线,并加入同位素内标参与定量,有效地克服了基质效应。上述4种化合物工作曲线的线性范围为0~100 μg·kg-1,测定下限(10S/N)依次为0.10,0.20,5.0,2.0 μg·kg-1。在实际样品基体中加入3个浓度水平的标准溶液(5.0,10,20 μg·kg-1)进行回收试验,测得回收率均大于80%,测定值的相对标准偏差(n=6)均小于10%。所提出方法具有简便、快速的优点,其测定下限能满足目前国内外的规定要求。
Abstract
2.00 g of honey sample were taken and 40 ng of the isotopic internal standard were added. The mixture was dissolved and diluted to 10.0 mL with potassium dihydrogen phosphate-sodium hydrogen phasphate buffer solution of pH 7. The solution was extracted twice with acetonitrile (10.0 mL for each extraction and 2.0 g of NaCl were added in the 1st extraction) by centrifuging and swirling for 5 min. The 2 supernatants were combined and its volume was made up to 20.0 mL with acetonitrile. An aliquot of 1.0 mL of the solution was transferred to a centrifuging tube in which 25 mg of PSA, 10 mg of C18 and 30 mg of MgSO4 were added preliminarily, and the mixture was centrifuged at high speed to perform purification by dispersive SPE. Whole of the supernatant was taken and its volume was made up to 2.0 mL with water. This solution was used for LC-MS/MS analysis. In LC separation, XDB-C18 chromatographic column was used as stationary phase, and mixtures of (A) φ 0.15% formic acid and (B) acetonitrile in various proportions were used as mobile phase in gradient elution. Eluates obtained at various time-intervals were used for MS/MS determination of residual amounts of Amitraz and its 3 main metabolites, i.e., N-2,4-dimethylphenyl-N-methylformamidine, N-(2,4-dimethylphenyl)formamide and 2,4-dimethylaniline in the honey sample. In preparation of working standard curves, mixed standard solutions of the 4 analytes were added to a blank honey sample as matrix, and internal standard was used for quantification simultaneously, interference due to matrix effect was effectively overcome. Linearity ranges for the 4 compounds were in the same range of 0-100 μg·kg-1, with their lower limits of determination (10S/N) were found as follows 0.10, 0.20, 5.0 2.0 μg·kg-1. Tests for recovery and precision were performed by addition of standard solutions at 3 concentration levels (5.0, 10 and 20 μg·kg-1) to a blank honey sample as matrix. Values of recovery found were all greater than 80%, and values of RSD (n=6) found were all less than 10%. It was shown that the proposed method has specialties of easy and rapid in manipulations, and furthermore, values of lower limits of determination for the 4 analytes were found to meet with the requirements of regulations given by our country or abroad.
中图分类号 O657.63 DOI 10.11973/lhjy-hx201907007
所属栏目 工作简报
基金项目 浙江省科技计划项目(2017C37011);浙江出入境检验检疫局科技计划项目(ZK201710X)
收稿日期 2018/10/11
修改稿日期
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备注侯建波,高级工程师,博士,研究方向为食品中药物残留分析,houjb@ziq.gov.cn
引用该论文: HOU Jianbo,XIE Wen,QIAN Yan,YAO Binbin,HU Xiaoli,MAO Renyi,HAN Chao. Determination of Amitraz and Its Metabolites in Honey by LC-MS/MS[J]. Physical Testing and Chemical Analysis part B:Chemical Analysis, 2019, 55(7): 784~790
侯建波,谢文,钱艳,姚滨滨,胡晓莉,毛壬熠,韩超. 液相色谱-串联质谱法测定蜂蜜中双甲脒及其代谢产物残留量[J]. 理化检验-化学分册, 2019, 55(7): 784~790
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参考文献
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【21】薛晓锋,赵静,邱静,等.气相色谱-质谱法同时测定蜂蜜中的双甲脒及其代谢物残留[J].现代科学仪器, 2005(1):65-67.
【22】RYOO J J, KIM S H, JEONG Y H, et al. Simultaneous determination of amitraz, bromopropylate, coumaphos, cymiazole and 2,4-dimethylaniline in korean honey samples by high-performance liquid chromatography[J]. Bulletin of the Korean Chemical Society, 2008,29(5):1043-1047.
【23】GB/T 21169-2007国家标准蜂蜜中双甲脒及其代谢物残留量测定液相色谱法[S].
【24】赵增运,吴斌,沈崇钰,等.高效液相色谱法测定蜂蜜中的双甲脒残留[J].中国养蜂, 2005,56(5):4-6.
【25】MARTEL A C, ZEGGANE S. Determination of acaricides in honey by high-performance liquid chromatography with photodiode array detection[J]. Journal of Chromatography A, 2002,954(1/2):173-180.
【26】ZHENG W J, PARK J A, ABD E A M, et al. Development and validation of modified QuEChERS method coupled with LC-MS/MS for simultaneous determination of cymiazole, fipronil, coumaphos, fluvalinate, amitraz, and its metabolite in various types of honey and royal jelly[J]. Journal of Chromatography B, 2018,1072:60-69.
【27】FU Y, YANG T, ZHAO J, et al. Determination of eight pesticides in lycium barbarum by LC-MS/MS and dietary risk assessment[J]. Food Chemistry, 2017,218:192-198.
【28】NAKAJIMA T, TSURUOKA Y, KANDA M, et al. Determination and surveillance of nine acaricides and one metabolite in honey by liquid chromatography-tandem mass spectrometry[J]. Food Additives & Contaminants:Part A, 2015,32(7):1099-1104.
【29】TOMASINI D, SAMPAIO M R F, CALDAS S S, et al. Simultaneous determination of pesticides and 5-hydroxymethylfurfural in honey by the modified QuEChERS method and liquid chromatography coupled to tandem mass spectrometry[J]. Talanta, 2012,99:380-386.
【30】XU J Z, MIAO J J, LIN H, et al. Determination of amitraz and 2,4-dimethylaniline residues in honey by using LC with UV detection and MS/MS[J]. Journal of Separation Science, 2009,32(23/24):4020-4024.
【31】侯建波,谢文,曾淦宁,等.液相色谱-串联质谱法测定蜂王浆中双甲脒及其代谢物的残留量[J].质谱学报, 2019(2):131-138.
【32】GAO X, TAN Y L, GUO H. Simultaneous determination of amitraz, chlordimeform, formetanate and their main metabolites in human urine by high performance liquid chromatography-tandem mass spectrometry[J]. Journal of Chromatography B, 2017,1052:27-33.
【33】GUO H, ZHANG P, WANG J W, et al. Determination of amitraz and its metabolites in whole blood using solid-phase extraction and liquid chromatography-tandem mass spectrometry[J]. Journal of Chromatography B, 2014,951/952:89-95.
【34】KORTA E, BAKKALI A, BERRUETA L A, et al. Study of acaricide stability in honey. Characterization of amitraz degradation products in honey and beeswax[J]. Journal of Agricultural and Food Chemistry, 2001,49(12):5835-5842.
【2】王少成,姜元振.双甲脒(螨克)防治果、菜、棉植食性螨、蚜、木虱等的药效评价[J].农药科学与管理, 1992,13(2):6-7.
【3】代平礼,王强,孙继虎,等.4种农药对意大利蜜蜂的毒力测定[J].农药, 2007,46(8):546-547.
【4】SANTIAGO G P, OTERO C G, SÁNCHEZ D M, et al. Comparing effects of three acaricides on varroa jacobsoni (acari:varroidae) and apis mellifera (hymenoptera:apidae) using two application techniques[J]. The Florida Entomologist, 2000,83(4):468-476.
【5】陈锐,戴珍科,蔡道基.甲基异柳磷等四种农药对蜜蜂的安全评价研究[J].生态科学, 1992,11(2):100-103.
【6】OSANO O, ADMIRAAL W, KLAMER H J C, et al. Comparative toxic and genotoxic effects of chloroacetanilides, formamidines and their degradation products on vibrio fischeri and chirono-mus riparius[J]. Environmental Pollution, 2002,119(2):195-202.
【7】RIALOTERO R, GASPAR E, MOURA I, et al. Chromatographic-based methods for pesticide determination in honey:An overview[J]. Talanta, 2007,71(2):503-514.
【8】European Union. Council Regulation(EU) No. 2377/90/EEC/1990[S].
【9】Japan. Positive list system for agricultural chemical residues in foods[EB/OL]. http://www.mhlw.go.jp/english/topics/foodsafety/positivelist060228/.
【10】United States of America. Code of federal regulations (40 CFR Part 180,180.287)[S]. 2006.
【11】JIMÉNEZ J, NOZAL M, BERNAL J, et al. Factors affecting the extraction, hydrolysis and deri-vatization steps for the quantitation of total residues of amitraz in honey by gas chromatography with electron capture detection[J]. Analytical and Bioanalytical Chemistry, 2002,374(2):300-304.
【12】LENÍĈEK J, SEKYRA M, NOVOTNÁ A R, et al. Solid phase microextraction and gas chroma-tography with ion trap detector (GC-ITD) analysis of amitraz residues in beeswax after hydrolysis to 2,4-dimethylaniline[J]. Analytica Chimica Acta, 2006,571(1):40-44.
【13】SHAMSIPUR M, HASSAN J, SALAR-AMOLI J, et al. Headspace solvent microextraction-gas chromatographic thermionic specific detector determination of amitraz in honey after hydrolysis to 2,4-dimethylaniline[J]. Journal of Food Composition and Analysis, 2008,21(3):264-270.
【14】AMOLI J S, HASAN J, HEJAZY M. Determination of amitraz residue by headspace gas chromatography in honey and beeswax samples from iran[J]. American Journal of Food Technology, 2009,4(1):56-59.
【15】JIMÉNEZ J J, BERNAL J L, DEL NOZAL M J, et al. Extraction and clean-up methods for the determination of amitraz total residues in beeswax by gas chromatography with electron capture detection[J]. Analytica Chimica Acta, 2004,524(1/2):271-278.
【16】郑用权,姚建仁,邵向东,等.蜂蜜中双甲脒残留量检测方法[J].农药科学与管理, 2000,21(3):14-16.
【17】李阿丹,王晶,崔宗岩,等.固相微萃取-气相色谱-串联质谱法测定蜂蜜中双甲脒及其代谢产物残留[J].燕山大学学报, 2016,40(3):236-240.
【18】YAMINI Y, FARAJI M, GHAMBARIAN M. Hollow-fiber liquid-phase microextraction followed by gas chromatography flame ionization detection for the determination of amitraz in honey and water samples[J]. Food Analytical Methods, 2015,8(3):758-766.
【19】CALDOW M, FUSSELL R J, SMITH F, et al. Development and validation of an analytical method for total amitraz in fruit and honey with quantification by gas chromatography-mass spectrometry[J]. Food Additives and Contaminants, 2007,24(3):280-284.
【20】BASHIRI J M, MEHDINIA A, JABBARI A, et al. Determination of amitraz in the honey samples by dispersive liquid-liquid microextraction followed by gas chromatography:Flame ionization detection[J]. American Journal of Analytical Chemistry, 2011,2(5):632-637.
【21】薛晓锋,赵静,邱静,等.气相色谱-质谱法同时测定蜂蜜中的双甲脒及其代谢物残留[J].现代科学仪器, 2005(1):65-67.
【22】RYOO J J, KIM S H, JEONG Y H, et al. Simultaneous determination of amitraz, bromopropylate, coumaphos, cymiazole and 2,4-dimethylaniline in korean honey samples by high-performance liquid chromatography[J]. Bulletin of the Korean Chemical Society, 2008,29(5):1043-1047.
【23】GB/T 21169-2007国家标准蜂蜜中双甲脒及其代谢物残留量测定液相色谱法[S].
【24】赵增运,吴斌,沈崇钰,等.高效液相色谱法测定蜂蜜中的双甲脒残留[J].中国养蜂, 2005,56(5):4-6.
【25】MARTEL A C, ZEGGANE S. Determination of acaricides in honey by high-performance liquid chromatography with photodiode array detection[J]. Journal of Chromatography A, 2002,954(1/2):173-180.
【26】ZHENG W J, PARK J A, ABD E A M, et al. Development and validation of modified QuEChERS method coupled with LC-MS/MS for simultaneous determination of cymiazole, fipronil, coumaphos, fluvalinate, amitraz, and its metabolite in various types of honey and royal jelly[J]. Journal of Chromatography B, 2018,1072:60-69.
【27】FU Y, YANG T, ZHAO J, et al. Determination of eight pesticides in lycium barbarum by LC-MS/MS and dietary risk assessment[J]. Food Chemistry, 2017,218:192-198.
【28】NAKAJIMA T, TSURUOKA Y, KANDA M, et al. Determination and surveillance of nine acaricides and one metabolite in honey by liquid chromatography-tandem mass spectrometry[J]. Food Additives & Contaminants:Part A, 2015,32(7):1099-1104.
【29】TOMASINI D, SAMPAIO M R F, CALDAS S S, et al. Simultaneous determination of pesticides and 5-hydroxymethylfurfural in honey by the modified QuEChERS method and liquid chromatography coupled to tandem mass spectrometry[J]. Talanta, 2012,99:380-386.
【30】XU J Z, MIAO J J, LIN H, et al. Determination of amitraz and 2,4-dimethylaniline residues in honey by using LC with UV detection and MS/MS[J]. Journal of Separation Science, 2009,32(23/24):4020-4024.
【31】侯建波,谢文,曾淦宁,等.液相色谱-串联质谱法测定蜂王浆中双甲脒及其代谢物的残留量[J].质谱学报, 2019(2):131-138.
【32】GAO X, TAN Y L, GUO H. Simultaneous determination of amitraz, chlordimeform, formetanate and their main metabolites in human urine by high performance liquid chromatography-tandem mass spectrometry[J]. Journal of Chromatography B, 2017,1052:27-33.
【33】GUO H, ZHANG P, WANG J W, et al. Determination of amitraz and its metabolites in whole blood using solid-phase extraction and liquid chromatography-tandem mass spectrometry[J]. Journal of Chromatography B, 2014,951/952:89-95.
【34】KORTA E, BAKKALI A, BERRUETA L A, et al. Study of acaricide stability in honey. Characterization of amitraz degradation products in honey and beeswax[J]. Journal of Agricultural and Food Chemistry, 2001,49(12):5835-5842.
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