氟虫腈亚砜荧光衍生化、HPLC-FLD检测方法的建立及应用

王梦 周香菊 聂嘉文 彭大勇 陈继光 张清峰 唐道邦 刘佳 尹忠平

王梦,周香菊,聂嘉文,等. 氟虫腈亚砜荧光衍生化、HPLC-FLD检测方法的建立及应用[J]. 食品工业科技,2021,42(20):286−293. doi:  10.13386/j.issn1002-0306.2021020236
引用本文: 王梦,周香菊,聂嘉文,等. 氟虫腈亚砜荧光衍生化、HPLC-FLD检测方法的建立及应用[J]. 食品工业科技,2021,42(20):286−293. doi:  10.13386/j.issn1002-0306.2021020236
WANG Meng, ZHOU Xiangju, NIE Jiawen, et al. Establishment and Application of HPLC-FLD Method for Fipronil Sulfoxide Detection by Pre-column Fluorescence Derivatization[J]. Science and Technology of Food Industry, 2021, 42(20): 286−293. (in Chinese with English abstract). doi:  10.13386/j.issn1002-0306.2021020236
Citation: WANG Meng, ZHOU Xiangju, NIE Jiawen, et al. Establishment and Application of HPLC-FLD Method for Fipronil Sulfoxide Detection by Pre-column Fluorescence Derivatization[J]. Science and Technology of Food Industry, 2021, 42(20): 286−293. (in Chinese with English abstract). doi:  10.13386/j.issn1002-0306.2021020236

氟虫腈亚砜荧光衍生化、HPLC-FLD检测方法的建立及应用

doi: 10.13386/j.issn1002-0306.2021020236
基金项目: 江西省食品药品监督管理局科技项目(2017SP05);国家自然科学基金项目(31460436);广州市科技计划项目(201909020001)
详细信息
    作者简介:

    王梦(1996−),女,硕士研究生,研究方向:食品安全检测及食品生物技术,E-mail:wmeng1019@163.com

    通讯作者:

    尹忠平(1971−),男,博士,教授,研究方向:食品安全检测及食品生物技术,E-mail:yin_zhongping@163.com

  • 中图分类号: TS207.5+3

Establishment and Application of HPLC-FLD Method for Fipronil Sulfoxide Detection by Pre-column Fluorescence Derivatization

  • 摘要: 建立了氟虫腈代谢物-氟虫腈亚砜的柱前荧光衍生化方法,并将其应用于鸡蛋中的残留检测。优化后的柱前荧光衍生化反应主要条件如下:催化缩合剂为EDC/DMAP;反应溶剂为二氯甲烷;氟虫腈亚砜与荧光衍生试剂的用量比为1:8;45 ℃水浴中反应75 min。衍生产物为N-(3-氰基-1-(2, 6-二氯-4-(三氟甲基)苯基)-4-((三氟甲基)硫代)-1H-吡唑-5-基)-4-(2-(9-乙基-9H-咔唑-3-基)-1H-菲并[9, 10-d]咪唑-1-基)丁酰胺(DX1),具有很高的荧光强度。衍生化产物DX1的高效液相色谱-荧光检测(HPLC-FLD)的主要条件如下:C18色谱柱;等度洗脱;流动相:乙腈/水(80/20,v/v);流速:1.0 mL/min;λex=310 nm,λem=404 nm;进样量20 μL;柱温40 ℃。在上述检测条件下,氟虫腈亚砜HPLC-FLD检测方法的检测限为0.033 μg/L,定量限为0.092 μg/L。在此基础上,建立了鸡蛋残留氟虫腈亚砜提取和检测的HPLC-FLD方法,检测限和定量限分别达到了0.052和0.132 µg/kg,精密度、稳定性和重现性在保留时间和峰面积上的RSD分别小于0.04%和2.7%,上机检测可在20 min内完成。该方法具有较好的灵敏度、精确性和可靠性,可应用于鸡蛋中残留氟虫腈亚砜的检测。
  • 图  1  氟虫腈亚砜与荧光衍生试剂L2柱前衍生化反应方程式

    Figure  1.  Reaction equation of precolumn fluorescence derivatization reaction between L2 and fipronil sulfide

    图  2  催化剂、反应时间、反应温度、衍生试剂与氟虫腈亚砜的用量比及反应溶剂对氟虫腈亚砜衍生化效率的影响

    Figure  2.  Influence of catalyzer, reaction time, reaction temperature, ratio of fipronil sulfoxide to fluorescence derivatization reagent, and reaction solvent on the derivatizing efficiency of fipronil

    注:a:催化剂的影响;b:反应时间的影响;c:反应温度的影响;d:衍生试剂的影响;e:反应溶剂的影响。图a、e中大写字母表示差异极显著(P<0.01);小写字母表示差异显著(P<0.05)。

    图  3  荧光衍生试剂L2和氟虫腈亚砜衍生化产物DX1的荧光光谱扫描图

    Figure  3.  Spectrograms of fluorescence scanning of fluorescent derivatization reagent L2 and its derivative DX1

    注:a:L2荧光光谱扫描图;b:DX1荧光光谱扫描图。

    图  4  鸡蛋样品中残留氟虫腈亚砜HPLC-FLD检测色谱图

    Figure  4.  Chromatograms of HPLC-FLD determination of fipronil sulfide in sample eggs

    注:峰P1:氟虫腈亚砜和荧光试剂L2的荧光衍生产物DX1。

    表  1  氟虫腈亚砜HPLC-FLD检测方法的线性回归方程、相关系数、检测限和定量限

    Table  1.   Linear regression equation, determination coefficient, LOD and LOQ of fipronil sulfide determination by HPLC-FLD

    待测物名称线性回归方程线性相关系数
    r
    检测限
    (µg/L)
    定量限
    (µg/L)
    DX1Y=194.95X−0.06090.99990.0330.092
    下载: 导出CSV

    表  2  氟虫腈亚砜HPLC-FLD检测方法的精密度、稳定性和重现性

    Table  2.   Precision, stability and reproducibility of fipronil sulfide determination by HPLC-FLD

    评价指标平均值±标准偏差(Mean±SD)相对标准偏差(RSD,%)
    精密度保留时间(min)8.568±0.00340.040
    峰面积106.93±2.2382.093
    稳定性保留时间(min)8.454±0.00190.023
    峰面积107.57±0.9510.884
    重现性保留时间(min)8.474±0.00550.065
    峰面积78.003±1.3991.793
    下载: 导出CSV

    表  3  鸡蛋中残留氟虫腈亚砜柱前衍生化HPLC-FLD检测的加标回收率

    Table  3.   Recovery rate of HPLC-FLD determination of fipronil sulfide in eggs by pre-column derivatization

    待测物名称添加量(mg/kg)平均加标回收率(%)相对标准偏差RSD(%)重复性RSD(%)
    氟虫腈亚砜0.0184.683.650.82
    0.187.851.050.75
    0.594.502.980.91
    下载: 导出CSV

    表  4  鸡蛋中残留氟虫腈亚砜HPLC-FLD检测方法的检测限、定量限和灵敏度

    Table  4.   LOD and LOQ of HPLC-FLD determination of fipronil sulfide in eggs by pre-column derivatization

    待测物名称检测限LOD
    (µg/kg)
    定量限LOQ
    (µg/kg)
    灵敏度
    氟虫腈亚砜0.0520.13265.5
    下载: 导出CSV

    表  5  鸡蛋中残留氟虫腈亚砜HPLC-FLD检测方法的精密度、稳定性和重现性

    Table  5.   Precision, stability and reproducibility of HPLC-FLD determination of fipronil sulfide in eggs by pre-column derivatization

    评价指标平均值±标准偏差(Mean±SD)相对标准偏差(RSD,%)
    精密度保留时间(min)8.340±0.00280.034
    峰面积89.403±1.6411.836
    稳定性保留时间(min)8.46±0.00180.021
    峰面积109.693±1.8781.712
    重现性保留时间(min)8.42±0.00330.039
    峰面积116.718±3.1162.670
    下载: 导出CSV

    表  6  五省二十五份鸡蛋样品残留氟虫腈亚砜HPLC-FLD检测结果

    Table  6.   Determination results of residual fipronil sulfide in 25 sample eggs collected from 5 provinces by HPLC-FLD

    样品地区取样数氟虫腈亚砜平均检测值(µg/kg)检测结果
    江西50未检出
    湖北50未检出
    山东50未检出
    河南50未检出
    安徽50未检出
    下载: 导出CSV
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  • 收稿日期:  2021-03-01
  • 网络出版日期:  2021-08-31
  • 刊出日期:  2021-10-11

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