Cy3标记农药核酸适配体表面增强拉曼光谱法特异性检测痕量啶虫脒

doi:10.3964/j.issn.1000-0593(2020)08-2462-06

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摘要：将具有拉曼信号的三氢-吲哚菁类（Cy3）染料分子标记农药核酸适配体（Aptamer）制备成拉曼检测试剂（Cy3-aptamer），对痕量啶虫脒进行了特异性的表面增强拉曼光谱法（SERS）检测研究。考虑到胶体的稳定和聚凝作用原理，采用聚丙烯酸钠作为分散剂，使作为SERS检测基底材料的银溶胶带负电荷，获得了良好的稳定性和分散性。由于聚丙烯酸钠分散的银溶胶为负电平衡体系，测试时需采用聚沉剂，使具有较高稳定性的银纳米颗粒团聚，形成SERS增强热点，从而提高SERS检测信号。以SERS信号较弱的啶虫脒为探针，考察了银溶胶中加入不同聚沉剂（NaCl，KCl，NaOH，HNO₃，H₃PO₄，H₂SO₄，HCl）对SERS信号的影响，实验结果表明，H⁺作为阳离子和PO^3-₄作为阴离子组成的电解质聚沉剂，对于带有一定负电荷σ^-基团分子，具有较好的拉曼增强效应。且通过紫外可见分光光谱，进一步说明了表面电荷性质对SERS的增强信号起决定作用。又由于Cy3-aptamer磷酸骨架上带有大量负电荷，其SERS信号较小。故选择带丰富正电荷的精胺分子以消除Cy3-aptamer磷酸骨架上的负电荷，使Cy3-aptemer更易吸附于银溶胶表面，使其产生较强的SERS光谱。此外，考察选择了精胺与Cy3-aptamer以及Cy3-aptamer与农药啶虫脒的最佳反应结合时间分别为5和20 min。最后，建立了定量检测农药啶虫脒的方法，并对检测机理进行了探讨。研究表明，农药啶虫脒在适配体银溶胶特效探针上于1 392 cm^-1处的SERS特征峰面积与水的OH伸缩振动峰面积组成相对拉曼峰面积强度，其相对强度与啶虫脒浓度的对数具有良好的负线性关系，浓度范围为1×10^-8～2.5×10^-7 mol·L^-1。将所建立的特效检测啶虫脒的方法用于实际水样的检测，回收率为97.4%～99.4%。结果表明，所提出的聚丙烯酸钠分散及精胺修饰的银溶胶有利于捕获Cy3-aptamer及其Cy3-aptamer与啶虫脒的反应物，提高了方法的灵敏度与可靠性。

关键词：表面增强拉曼光谱；核酸适配体；银溶胶；啶虫脒；聚丙烯酸钠；精胺

Abstract：In this work, aptamer modified by Cy3 (1,1’-bis(3-hydroxypropyl)-3,3,3’, 3’-tetramethylindocarbocyanine) dye which has a strong Raman signal, was applied for sensitive and specific detection of trace acetamiprid by Surface-enhanced Raman Scattering (SERS). The good stability and dispersity of negative silver colloid were obtained by adding the proper concentration of sodium polyacrylate according to the principle of colloid stabilization and coagulation. During the detection process, these high stable silver nanoparticles were needed to be agglomerated by using the agglomerating agent to form more SERS enhancement hotspots to improve the SERS intensity. The effects of different agglomerating agents (NaCl, KCl, NaOH, HNO₃, H₃PO₄, H₂SO₄, HCl) were investigated using acetamiprid as a probe. The results showed that the good SERS effect was showed when the electrolyte precipitator contains H⁺ and PO^3-₄. Furthermore, UV-Vis spectra show that the surface charge properties play a decisive role in the SERS effect. Furthermore, spermine with positive charges was selected to neutralize the negative charges on the phosphoric acid skeleton of Cy3-aptamer which could shorten the distance between Cy3-aptamer and silver colloid to enhance the Raman signal. The optimum reaction times of spermine with Cy3-aptamer and Cy3-aptamer with acetamiprid were 5 and 20 min, respectively. At last, quantitative detection of the acetamiprid method was established and the linearship was established between the logarithm concentration of acetamiprid and the relative intensity of the characteristic peak area at 1 392 cm^-1 divided by the OH stretching vibration of water. The linear range is from 1×10^-8 to 2.5×10^-7 mol·L^-1. The proposed method was applied to the determination of the spiked acetamiprid in water samples with the recovery of 97.4%～99.4%. The results show that the silver colloid dispersed by sodium polyacrylate and modified by spermine were helpful to capture the Cy3-aptamer and the reactants of Cy3-aptamer with acetamiprid, that the sensitivity and reliability of the method were improved.

Key words：Surface-enhanced Raman scattering; Aptamer; Silver colloid; Acetamiprid; Sodium polyacrylate; Spermine

收稿日期: 2019-07-17 修订日期: 2019-11-28

中图分类号:

O657.37

基金资助: 国家自然科学基金项目（21804119），浙江省自然科学基金项目（LQ17B050002）和浙江省公益性技术应用研究(分析测试)计划项目(2015C37068)资助

作者简介: 滕渊洁，1983年生，浙江工业大学化学工程学院讲师 e-mail: yuanjieteng@zjut.edu.cn

引用本文:

滕渊洁，韦其真，刘文涵，刘江美，聂永惠，李盼. Cy3标记农药核酸适配体表面增强拉曼光谱法特异性检测痕量啶虫脒[J]. 光谱学与光谱分析, 2020, 40(08): 2462-2467.
TENG Yuan-jie, WEI Qi-zhen, LIU Wen-han, LIU Jiang-mei, NIE Yong-hui, LI Pan. Cy3-Labeled Aptamer Combined with Surface-Enhanced Raman Scattering Using for Specific Detection of Trace Acetamiprid. SPECTROSCOPY AND SPECTRAL ANALYSIS, 2020, 40(08): 2462-2467.

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https://www.gpxygpfx.com/CN/10.3964/j.issn.1000-0593(2020)08-2462-06 或 https://www.gpxygpfx.com/CN/Y2020/V40/I08/2462