同位素稀释技术结合超高效液相色谱-四极杆/线性离子阱质谱快速精准测定饲料及原料中16种霉菌毒素
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摘要
为优化饲料及原料中16种霉菌毒素的多残留提取,本试验结合稳定同位素稀释技术建立了饲料及原料中16种霉菌毒素的超高效液相色谱-四极杆/线性离子阱质谱检测法(Q-trap-UPLC-MS/MS)。样品经乙腈/水/乙酸(V/V/V∶70/29/1)溶液提取、稀释和上机检测。采用反相C-18色谱柱分离,以0.1%甲酸和1 mmoL NH4Ac溶液作为弱洗脱流动相与强洗脱流动相乙腈进行梯度洗脱,采用分段多反应监测(MRM)扫描模式进行定性定量分析。基质考察表明,饲料及原料中基质效应较强,故采用稳定同位素稀释以减少基质效应对定量分析的干扰。结果表明:16种霉菌毒素在一定浓度范围内均具有良好的线性关系,相关系数(R2)均大于0.998,5种常见配合饲料基质中的限量浓度添加水平的回收率(n=6)为70.2%～129.5%,相对标准偏差为0.4%～14.7%;4种常见原料基质中的限量浓度添加水平的回收率(n=6)为80.2%～116.1%,相对标准偏差为0.1%～7.7%,16种霉菌毒素的定量限为0.25～200 g/kg。该方法简单、快捷及准确,可满足批量饲料及原料中霉菌毒素的日常监测工作。
A UPLC-Quadrupole/Orbitrap Mass Spectrometric(Q-trap-UPLC-MS/MS) method was established for the determination of 16 mycotoxins in feed and feedstuff with stable isotope dilution technique.5 g sample(± 0.1 mg) was added in 50 mL centrifuge tube and extracted by acetonitrile/water/acetic acid(V/V/V:70/29/1).Chromatographic analyses were carried out on a reversed phase C18 column and using a gradient elution with 0.1% formic acid and 1 mmol/L NH4 Ac as weak elution mobile phase.The mass spectrometer was operated in a multiple reaction monitoring(MRM) mode for qualitative analysis.The most target compounds had different level of matrix effects and the stable isotope dilution was adopted to decrease it.The method showed good linearities( r2>0.998) in the certain concentration.The average recoveries and RSDs of mycotoxins from 5 kinds of feed were in the range of 70.2% ～ 129.5% and 0.41% ～ 14.7%,respectively.The average recoveries and RSDs of mycotoxins from in 4 kinds of feedstuffs were 80.2% ～ 116.1% and 0.1% ～ 7.7%,respectively.Limits of quantification ranged of the method was from 0.25 ～ 200 g/kg.The method was simple,quick and accurate,so it was suitable for the daily monitoring of mycotoxins in a large number of feed and feedstuffs.
A UPLC-Quadrupole/Orbitrap Mass Spectrometric(Q-trap-UPLC-MS/MS) method was established for the determination of 16 mycotoxins in feed and feedstuff with stable isotope dilution technique.5 g sample(± 0.1 mg) was added in 50 mL centrifuge tube and extracted by acetonitrile/water/acetic acid(V/V/V:70/29/1).Chromatographic analyses were carried out on a reversed phase C18 column and using a gradient elution with 0.1% formic acid and 1 mmol/L NH4 Ac as weak elution mobile phase.The mass spectrometer was operated in a multiple reaction monitoring(MRM) mode for qualitative analysis.The most target compounds had different level of matrix effects and the stable isotope dilution was adopted to decrease it.The method showed good linearities( r2>0.998) in the certain concentration.The average recoveries and RSDs of mycotoxins from 5 kinds of feed were in the range of 70.2% ～ 129.5% and 0.41% ～ 14.7%,respectively.The average recoveries and RSDs of mycotoxins from in 4 kinds of feedstuffs were 80.2% ～ 116.1% and 0.1% ～ 7.7%,respectively.Limits of quantification ranged of the method was from 0.25 ～ 200 g/kg.The method was simple,quick and accurate,so it was suitable for the daily monitoring of mycotoxins in a large number of feed and feedstuffs.
引文
[1]范志辰,韩铮,郭文博,等.超高效液相色谱-串联质谱法同时测定不同饲料中30种真菌毒素[J].色谱,2017,35(6),627~633.
[2]符金华,杨琳芬,董泽民,等.饲料中16种霉菌毒素液相色谱-三重四级杆/线性离子阱串联质谱法的条件优化[J].中国饲料,2017,15:5~10.
[3]王瑞国,苏晓鸥,程芳芳,等.液相色谱-串联质谱法测定饲料原料中26种霉菌毒素[J].分析化学研究报告,2015,43(2):264~270.
[4]薛毅,张玥,吴银良.液相色谱-串联质谱法同时测定饲料中17种霉菌毒素[J].饲料加工与检测[J].2016,52(19):90~94.
[5]应永飞,朱聪英,韦敏珏,等.液相色谱-串联质谱法测定饲料中14种霉菌毒素及其类似物[J].分析化学,2010,38(12):1759~1764.
[6]赵孔祥,葛宝坤,陈旭艳,等.在线免疫亲和柱-液相色谱-串联质谱快速测定中药及中药中10中真菌毒素[J].分析化学,2011,39(9):1341~1346.
[7]郑翠梅,张艳,王松雪,等.液相色谱-串联质谱法测定小麦中T-2、ZEN及DON3种镰刀菌毒素[J].分析测试学报,2012,31(4):383~389.
[8]郑翠梅,张艳,王雪松,等.液相色谱-质谱联用同时检测粮食中多种真菌毒素的应用进展[J].粮食科技与经济,2012,37(1):45~49.
[9]中华人民共和国国家质量监督检验检疫总局,中国国家标准化管理委员会.GB 13078.2-2006[S].饲料卫生标准-饲料中赭曲霉毒素A和玉米赤霉烯酮的允许量.北京:中国标准出版社,2006-07-01.
[10]中华人民共和国国家质量监督检验检疫总局,中国国家标准化管理委员会.GB 13078.3-2007[S].配合饲料中脱氧雪腐镰刀菌烯醇的允许量.北京:中国标准出版社,2007-03-01.
[11]中华人民共和国国家质量监督检验检疫总局,中国国家标准化管理委员会.GB 21693-2008[S].配合饲料中T-2毒素的允许量.北京:中国标准出版社,2008-07-01.
[12]中华人民共和国国家质量监督检验检疫总局.GB 13078-2001[S].饲料卫生标准.北京:中国标准出版社,2001-10-01.
[13]中华人民共和国卫生部.GB 2761-2011[S].食品中真菌毒素限量.北京:中国标准出版社,2011-10-20.
[14]朱聪英,应永飞,韦敏钰,等.液相色谱-串联质谱法测定饲料中黄曲霉毒素的研究[J].质谱学报,2010,31(4):240~246.
[15]Binder E M,Tan L M,Chin L J,et al.Worldwide occurrence of mycotoxins in commodities,feeds and feed ingredients[J].J Anim.Feed Sci.Technol.2007,137(3):265~282.
[16]Cavaliere C,Foglia P,Guarino C,et al.Anal Chim Acta,2007,596(24),141-152.
[17]Chiar A C,Patrizia F,et al.Determination of aflatoxins in olive oil by liquid chromatography-tandem mass spectrometry[J].J Analytica Chimica Acta,2007,596(1):141~148.
[18]Johannes M W,Van D O,Ido P K.The role of liquid chromatography-tandem mass spectrometry in the clinical laboratory[J].JChromatogr B.2012,883(884):18~32.
[19]Marin S,Ramos A,Cano-Sancho G,et al.Mycotoxins:occurrence,toxicology,and exposure assessment[J].J Food Chem Toxicol,2013,60:218~237.
[20]Markus H.10 years of MS instrumental developments-impact on LC-MS/MS in clinical chemistry[J].J Chromatogr B,2012,3:883(884):3~17.
[21]Micheal J,Riordan O,Martin G,et al.Comparison of analyticalmethods for aflatoxin determination in commercial chilli spicepreparations and subsequent development of an improved method[J].Food Control,2009,20(8):700~705.
[22]Mira P,Marinella F,Miren A L,et al.Recent trends in the liquid chromatography mass spectrometry analysis of?organic contaminants in environmental samples[J].J Chromatogr A,2010,1217(25):4004~4017.
[23]Monbaliu S,van Poucke C,Detavernier C,et al.Occurrence of mycotoxins in feed as analyzed by a multi-mycotoxin LC-MS/MSMethod[J].J Agr Food Chem,2009,58(1):66~71.
[24]Shephard G S,Berthiller F,Burdaspal P A,et al.Developments in mycotoxin analysis:an update for 2011-2012[J].J World Mycotoxin,2012,5(1):3~30.
[2]符金华,杨琳芬,董泽民,等.饲料中16种霉菌毒素液相色谱-三重四级杆/线性离子阱串联质谱法的条件优化[J].中国饲料,2017,15:5~10.
[3]王瑞国,苏晓鸥,程芳芳,等.液相色谱-串联质谱法测定饲料原料中26种霉菌毒素[J].分析化学研究报告,2015,43(2):264~270.
[4]薛毅,张玥,吴银良.液相色谱-串联质谱法同时测定饲料中17种霉菌毒素[J].饲料加工与检测[J].2016,52(19):90~94.
[5]应永飞,朱聪英,韦敏珏,等.液相色谱-串联质谱法测定饲料中14种霉菌毒素及其类似物[J].分析化学,2010,38(12):1759~1764.
[6]赵孔祥,葛宝坤,陈旭艳,等.在线免疫亲和柱-液相色谱-串联质谱快速测定中药及中药中10中真菌毒素[J].分析化学,2011,39(9):1341~1346.
[7]郑翠梅,张艳,王松雪,等.液相色谱-串联质谱法测定小麦中T-2、ZEN及DON3种镰刀菌毒素[J].分析测试学报,2012,31(4):383~389.
[8]郑翠梅,张艳,王雪松,等.液相色谱-质谱联用同时检测粮食中多种真菌毒素的应用进展[J].粮食科技与经济,2012,37(1):45~49.
[9]中华人民共和国国家质量监督检验检疫总局,中国国家标准化管理委员会.GB 13078.2-2006[S].饲料卫生标准-饲料中赭曲霉毒素A和玉米赤霉烯酮的允许量.北京:中国标准出版社,2006-07-01.
[10]中华人民共和国国家质量监督检验检疫总局,中国国家标准化管理委员会.GB 13078.3-2007[S].配合饲料中脱氧雪腐镰刀菌烯醇的允许量.北京:中国标准出版社,2007-03-01.
[11]中华人民共和国国家质量监督检验检疫总局,中国国家标准化管理委员会.GB 21693-2008[S].配合饲料中T-2毒素的允许量.北京:中国标准出版社,2008-07-01.
[12]中华人民共和国国家质量监督检验检疫总局.GB 13078-2001[S].饲料卫生标准.北京:中国标准出版社,2001-10-01.
[13]中华人民共和国卫生部.GB 2761-2011[S].食品中真菌毒素限量.北京:中国标准出版社,2011-10-20.
[14]朱聪英,应永飞,韦敏钰,等.液相色谱-串联质谱法测定饲料中黄曲霉毒素的研究[J].质谱学报,2010,31(4):240~246.
[15]Binder E M,Tan L M,Chin L J,et al.Worldwide occurrence of mycotoxins in commodities,feeds and feed ingredients[J].J Anim.Feed Sci.Technol.2007,137(3):265~282.
[16]Cavaliere C,Foglia P,Guarino C,et al.Anal Chim Acta,2007,596(24),141-152.
[17]Chiar A C,Patrizia F,et al.Determination of aflatoxins in olive oil by liquid chromatography-tandem mass spectrometry[J].J Analytica Chimica Acta,2007,596(1):141~148.
[18]Johannes M W,Van D O,Ido P K.The role of liquid chromatography-tandem mass spectrometry in the clinical laboratory[J].JChromatogr B.2012,883(884):18~32.
[19]Marin S,Ramos A,Cano-Sancho G,et al.Mycotoxins:occurrence,toxicology,and exposure assessment[J].J Food Chem Toxicol,2013,60:218~237.
[20]Markus H.10 years of MS instrumental developments-impact on LC-MS/MS in clinical chemistry[J].J Chromatogr B,2012,3:883(884):3~17.
[21]Micheal J,Riordan O,Martin G,et al.Comparison of analyticalmethods for aflatoxin determination in commercial chilli spicepreparations and subsequent development of an improved method[J].Food Control,2009,20(8):700~705.
[22]Mira P,Marinella F,Miren A L,et al.Recent trends in the liquid chromatography mass spectrometry analysis of?organic contaminants in environmental samples[J].J Chromatogr A,2010,1217(25):4004~4017.
[23]Monbaliu S,van Poucke C,Detavernier C,et al.Occurrence of mycotoxins in feed as analyzed by a multi-mycotoxin LC-MS/MSMethod[J].J Agr Food Chem,2009,58(1):66~71.
[24]Shephard G S,Berthiller F,Burdaspal P A,et al.Developments in mycotoxin analysis:an update for 2011-2012[J].J World Mycotoxin,2012,5(1):3~30.