高效液相色谱检测赤霉素残留量的方法优化
摘要
农药在提高农作物产量及病虫害的综合防治中具有不可替代的作用,在农业生产中得到了广泛应用,但是,随之而来的是农药的不合理使用及滥用现象,导致农药在环境以及食物中残留量严重超标,对环境和人类健康不利。因此,国家正在逐步建立和完善可靠、灵敏、实用的农药残留分析技术,以达到合理规范的使用农药的目的。
赤霉素(GA3)是一种植物生长调节剂类农药,当大量使用时会产生一定毒性。本文建立了使用高效液相色谱(HPLC)对水果和蔬菜中的GA3残留进行分析检测的方法。优化了HPLC检测条件,使用键合固定相C18反相柱,紫外可见检测器,检测波长为254.00nm,流动相体系为pH4.0的40%甲醇水溶液。优化了样品的提取、分离和纯化步骤,使用80%甲醇水溶液超声波震荡法提取样品中的赤霉素,石油醚脱色,乙酸乙酯萃取,磷酸盐缓冲液反相萃取后,再以乙酸乙酯萃取,旋转蒸干后以甲醇溶解残渣,过滤膜后即为HPLC待测样品。
本文所建立的方法具有很高的灵敏度、精密度、准确度。利用这种检测方法检测各类水果和蔬菜中的GA3都能得到满意的结果。该方法符合灵敏、准确、易于推广使用的农药残留检测标准,是在国内现有条件下最具推广价值的适用于水果和蔬菜中GA3残留的检测方法。
Pesticides plays an irreplaceable role in improving agricultural production and in integrated pest control. It is widely applied in the field of agriculture. However, the irrational use of pesticides leads to seriously excessive residues in the environment and food and that is adverse to human health and environment. Therefore, our country is gradually establishing and perfecting a reliable, sensitive and practical pesticide residue analysis technology, to achieve a reasonable standard of the purpose of the reasonable use of pesticides.
Gibberellin (GA3) is a kind of plant growth regulator pesticides. When GA3 is used excessively, it would produce a certain toxicity. In this paper, we established an analysis method detecting the GA3 residue in fruit and vegetables with high-performance liquid chromatography (HPLC) . The HPLC testing conditions has been optimized: Stationary phase is C18; UV detector and the detection wavelength is 254.00 nm; The mobile phase is 40% methanol-water solution which pH is 4.0; The method of Sample extraction, separation and purification has also been optimized: GA3 is extracted with 80% methanol-water solution by ultrasonic shock from samples; The extract is decoloried with petroleum ether and GA3 extracted with ethyl acetate, and then, RP-extracted with PBS and extracted with ethyl acetate again. Volatile the ethyl acetate in the extract by rotary evaporation and then dissolve the residue with methanol followed by being filtered with 0.45 um membrane, and this solution is test sample for HPLC.
The method established in this paper has high sensitivity, precision and accuracy. The results is satisfying when it is used in detecting the GA3 of various fruits and vegetables. This method meets the pesticide residue testing standards that must be sensitive, accurate and easy to promote. So, this method has the most valuable promotion for detecting the GA3 Residues of various fruits and vegetables.
赤霉素(GA3)是一种植物生长调节剂类农药,当大量使用时会产生一定毒性。本文建立了使用高效液相色谱(HPLC)对水果和蔬菜中的GA3残留进行分析检测的方法。优化了HPLC检测条件,使用键合固定相C18反相柱,紫外可见检测器,检测波长为254.00nm,流动相体系为pH4.0的40%甲醇水溶液。优化了样品的提取、分离和纯化步骤,使用80%甲醇水溶液超声波震荡法提取样品中的赤霉素,石油醚脱色,乙酸乙酯萃取,磷酸盐缓冲液反相萃取后,再以乙酸乙酯萃取,旋转蒸干后以甲醇溶解残渣,过滤膜后即为HPLC待测样品。
本文所建立的方法具有很高的灵敏度、精密度、准确度。利用这种检测方法检测各类水果和蔬菜中的GA3都能得到满意的结果。该方法符合灵敏、准确、易于推广使用的农药残留检测标准,是在国内现有条件下最具推广价值的适用于水果和蔬菜中GA3残留的检测方法。
Pesticides plays an irreplaceable role in improving agricultural production and in integrated pest control. It is widely applied in the field of agriculture. However, the irrational use of pesticides leads to seriously excessive residues in the environment and food and that is adverse to human health and environment. Therefore, our country is gradually establishing and perfecting a reliable, sensitive and practical pesticide residue analysis technology, to achieve a reasonable standard of the purpose of the reasonable use of pesticides.
Gibberellin (GA3) is a kind of plant growth regulator pesticides. When GA3 is used excessively, it would produce a certain toxicity. In this paper, we established an analysis method detecting the GA3 residue in fruit and vegetables with high-performance liquid chromatography (HPLC) . The HPLC testing conditions has been optimized: Stationary phase is C18; UV detector and the detection wavelength is 254.00 nm; The mobile phase is 40% methanol-water solution which pH is 4.0; The method of Sample extraction, separation and purification has also been optimized: GA3 is extracted with 80% methanol-water solution by ultrasonic shock from samples; The extract is decoloried with petroleum ether and GA3 extracted with ethyl acetate, and then, RP-extracted with PBS and extracted with ethyl acetate again. Volatile the ethyl acetate in the extract by rotary evaporation and then dissolve the residue with methanol followed by being filtered with 0.45 um membrane, and this solution is test sample for HPLC.
The method established in this paper has high sensitivity, precision and accuracy. The results is satisfying when it is used in detecting the GA3 of various fruits and vegetables. This method meets the pesticide residue testing standards that must be sensitive, accurate and easy to promote. So, this method has the most valuable promotion for detecting the GA3 Residues of various fruits and vegetables.
引文
[1]潘瑞炽,董愚得.植物生理学(第三版)[M].高教出版社,1995.
[2]周宇,佟兆国,张开春,等.赤霉素在落叶果树生产中的应用[J].中国农业科技导报,2006,8(2):27-31.
[3]王军荣,雷虹.浅谈赤霉素在制麦中的应用[J].大麦科学,2005,1:47-48.
[4]王贵元,夏仁学,曾祥国,等.外源脱落酸和赤霉素对红肉脐橙果肉糖含量的影响[J].应用生态学报,2007,18(11):2451-2455.
[5] 张玉宗,陈振玲,吕杏者,等.高效液相色谱法测定谷子幼苗中的 GA1和 GA3[J].色谱,1999,17(5):469-472.
[6]辛丰.蔬菜巧用赤霉素,优质丰产增效益[J] .山东农药信息,2006,(5):32.
[7]刘海波.赤霉素在农业生产上的应用[J] .现代农业,2007,3:15.
[8]杨斌.赤霉素对葡萄无核化的影响[J] .安徽农业通报,2007,13(9):168.
[9]陈家华.食品安全让你吃得更健康[N].上海科技报,2004-5-11(3).
[10]曹坳程.我国农产品的农药污染问题及对策[Z] .中国农业科学院植物保护研究所
[11]袁祖华,粟建文,蔡雁平.湖南蔬菜产业现状及发展对策[J] .现代农业科技,2007,4:32-35.
[12]张中一,施正香,周清.农用化学品对生态环境和人类健康的影响及其对策[J] .中国农业大学学报 2003 ,8 (2) :73～77.
[13]贾蕊,陆迁,何学松.我国农业污染现状、原因及对策研究[J] .中国农业科技导报,2006,8(1):59～63.
[14]孙向东.蔬菜农药残留的危害、种类、超标原因及应对措施[J] .贵州农业科学,2005,33(6):99～100.
[15]藤威,柳琪,郭栋梁.当前我国农产品中农药、兽药残留污染原因的探讨与治理展望[J] .食品研究与开发,2003,8(24):11-12.
[16]黄示忠.好年冬在土壤和苹果中残留测定方法的研究[J] .农业环境保护,1998,1(11) :33-34.
[17]liu Z , Phillips J B. Comprehensive two - dimensional gas chromatography using an on column thermal modulator interface[J]. Chromatogr. Sci. ,1991 ,29 :227～231
[18]Zrostkou J , Hajlov J , Ajka T. Evaluation of two -dimensional gas chromatography - time of flight - mass spectrometry for thedetermination of multiple pesticide residues in fruit [J] . Chromatogr. A. , 2003 ,19(10):173～186.
[19]张政,张强,王转花. HPLC 测定赤霉素 GA3的含量[J].色谱, 1993,11(4):254.
[20]PicóY , Font G , MoltoJ C , Manes J . Pesticide residue determination in fruit and vegetables by liquid chromatography -mass spectrometry[J]. Chromatogr. A ,2000 ,882 :153～173.
[21]Fernández M , PicóY , Manes J . Determination of carbamate residues in fruits and vegetables by matrix solid - phase dispersion and liquid chromatography - mass spectrometry[J]. Chromatogr. A ,2000 ,871 :43～56.
[22]张静,寇登民.农药残留的现代仪器分析方法[J].现代仪器分析,2005,1:8-10.
[23]王朝瑾,蔡琦.农产品中农药残留的检测趋势[J].现代科学仪器,2006,1:106-108.
[24]郭永泽.当前农药残留分析技术及发展趋势[J].天津农业科学,2002(8):22-25.
[25] 黄宝美,郑妍鹏,李学谦.毛细管电泳法测定青菜中敌百虫的残留量[J].分析试验室,2004,23(3):1-3.
[26]费新平,王立世,张水锋.毛细管电泳-安培检测法对甲基对硫磷、对硫磷、西维因和速灭威农药残留的测定研究[J].分析测试学报,2004,23(5):70-73.
[27]何永红,高志贤,晁福寰.基于生物学原理的农药残留检测技术研究进展[J].卫生研究,2004,33(1):112-114.
[28]仲维科,郝戬,樊耀波,等.食品农药残留分析进展[J].分析化学,2000,28(7):904-910.
[29] 高景红,肖志勇.我国农药残留限量标准存在问题探讨[J].农业环境与发展,2002,4:32-33.
[30]吴娟,王明林,李艳霞,等. 我国食品中农药残留限量标准现状分析[J]. 食品安全,2006,2;55-57
[31]周艳明.仪器分析[M].吉林:吉林科学技术出版社,2002
[32]葛雅琨,张元新,胡睿,等.蔬菜中赤霉素残留量的高效液相色谱检测方法研究[J].吉林化工学院学报,2005,22(2):13-16
[33] 北京清华大学.赤霉素发酵滤液的溶剂萃取法浓缩工艺 [P] .中国专利,91100074.7.1991—01—11.
[2]周宇,佟兆国,张开春,等.赤霉素在落叶果树生产中的应用[J].中国农业科技导报,2006,8(2):27-31.
[3]王军荣,雷虹.浅谈赤霉素在制麦中的应用[J].大麦科学,2005,1:47-48.
[4]王贵元,夏仁学,曾祥国,等.外源脱落酸和赤霉素对红肉脐橙果肉糖含量的影响[J].应用生态学报,2007,18(11):2451-2455.
[5] 张玉宗,陈振玲,吕杏者,等.高效液相色谱法测定谷子幼苗中的 GA1和 GA3[J].色谱,1999,17(5):469-472.
[6]辛丰.蔬菜巧用赤霉素,优质丰产增效益[J] .山东农药信息,2006,(5):32.
[7]刘海波.赤霉素在农业生产上的应用[J] .现代农业,2007,3:15.
[8]杨斌.赤霉素对葡萄无核化的影响[J] .安徽农业通报,2007,13(9):168.
[9]陈家华.食品安全让你吃得更健康[N].上海科技报,2004-5-11(3).
[10]曹坳程.我国农产品的农药污染问题及对策[Z] .中国农业科学院植物保护研究所
[11]袁祖华,粟建文,蔡雁平.湖南蔬菜产业现状及发展对策[J] .现代农业科技,2007,4:32-35.
[12]张中一,施正香,周清.农用化学品对生态环境和人类健康的影响及其对策[J] .中国农业大学学报 2003 ,8 (2) :73～77.
[13]贾蕊,陆迁,何学松.我国农业污染现状、原因及对策研究[J] .中国农业科技导报,2006,8(1):59～63.
[14]孙向东.蔬菜农药残留的危害、种类、超标原因及应对措施[J] .贵州农业科学,2005,33(6):99～100.
[15]藤威,柳琪,郭栋梁.当前我国农产品中农药、兽药残留污染原因的探讨与治理展望[J] .食品研究与开发,2003,8(24):11-12.
[16]黄示忠.好年冬在土壤和苹果中残留测定方法的研究[J] .农业环境保护,1998,1(11) :33-34.
[17]liu Z , Phillips J B. Comprehensive two - dimensional gas chromatography using an on column thermal modulator interface[J]. Chromatogr. Sci. ,1991 ,29 :227～231
[18]Zrostkou J , Hajlov J , Ajka T. Evaluation of two -dimensional gas chromatography - time of flight - mass spectrometry for thedetermination of multiple pesticide residues in fruit [J] . Chromatogr. A. , 2003 ,19(10):173～186.
[19]张政,张强,王转花. HPLC 测定赤霉素 GA3的含量[J].色谱, 1993,11(4):254.
[20]PicóY , Font G , MoltoJ C , Manes J . Pesticide residue determination in fruit and vegetables by liquid chromatography -mass spectrometry[J]. Chromatogr. A ,2000 ,882 :153～173.
[21]Fernández M , PicóY , Manes J . Determination of carbamate residues in fruits and vegetables by matrix solid - phase dispersion and liquid chromatography - mass spectrometry[J]. Chromatogr. A ,2000 ,871 :43～56.
[22]张静,寇登民.农药残留的现代仪器分析方法[J].现代仪器分析,2005,1:8-10.
[23]王朝瑾,蔡琦.农产品中农药残留的检测趋势[J].现代科学仪器,2006,1:106-108.
[24]郭永泽.当前农药残留分析技术及发展趋势[J].天津农业科学,2002(8):22-25.
[25] 黄宝美,郑妍鹏,李学谦.毛细管电泳法测定青菜中敌百虫的残留量[J].分析试验室,2004,23(3):1-3.
[26]费新平,王立世,张水锋.毛细管电泳-安培检测法对甲基对硫磷、对硫磷、西维因和速灭威农药残留的测定研究[J].分析测试学报,2004,23(5):70-73.
[27]何永红,高志贤,晁福寰.基于生物学原理的农药残留检测技术研究进展[J].卫生研究,2004,33(1):112-114.
[28]仲维科,郝戬,樊耀波,等.食品农药残留分析进展[J].分析化学,2000,28(7):904-910.
[29] 高景红,肖志勇.我国农药残留限量标准存在问题探讨[J].农业环境与发展,2002,4:32-33.
[30]吴娟,王明林,李艳霞,等. 我国食品中农药残留限量标准现状分析[J]. 食品安全,2006,2;55-57
[31]周艳明.仪器分析[M].吉林:吉林科学技术出版社,2002
[32]葛雅琨,张元新,胡睿,等.蔬菜中赤霉素残留量的高效液相色谱检测方法研究[J].吉林化工学院学报,2005,22(2):13-16
[33] 北京清华大学.赤霉素发酵滤液的溶剂萃取法浓缩工艺 [P] .中国专利,91100074.7.1991—01—11.