黄曲霉毒素氧化酶/壳聚糖-单壁碳纳米管/聚邻苯二胺修饰电极对杂色曲霉素的检测

Detection of sterigmatocystin based on the novel Aflatoxin- Oxidase/chitosan-single-walled carbon nanotubes/poly-o- phenylenediamine modified electrode

摘要:

将黄曲霉毒素氧化酶(AFO)固定在壳聚糖(CS)-单壁碳纳米管(SWCNTs)杂交膜中, 组装在聚邻苯二胺(POPD)修饰的金电极(Au)表面, 制备了对杂色曲霉素(ST)敏感的生物传感器(AFO/CS-SWCNTs/POPD/Au)。运用原子力显微镜(AFM)、傅立叶变换红外光谱(FT-IR)和交流阻抗技术(EIS)对电极组装过程进行了表征。循环伏安法研究表明, AFO在修饰电极上发生了准可逆的氧化还原反应, 是表面控制过程, 其式量电位为-0.436 V (vs.Ag/AgCl), 说明包埋在CS-SWCNTs中的AFO和电极之间发生了直接电子传递。AFO修饰电极对ST具有明显的电催化作用, 其表观米氏常数为7.13 mmol/L, 催化电流与ST浓度在10~310 ng/mL范围内呈线性关系, 相关系数为0.997, 检出限为3 ng/mL (S/N=3), 响应时间小于10 s。组装的生物传感器具有较好的稳定性与重现性, 连续检测20 ng/mL的ST标准溶液11次, 电流值RSD为3.9%; 放置一个月后, 其电流响应值仍为初始值的85.6%。该方法具有较高的选择性和灵敏度, 应用于实际样品检测时, 其回收率在87.6%~105.5%之间。建立的ST含量的电化学测定方法简便快捷, 测定结果令人满意。

Abstract:

A sensitive electrochemical biosensor based on Aflatoxin-Oxidase (AFO) was developed for detection of sterigmatocystin (ST). The enzyme was immobilized on chitosan-single-walled carbon nanotubes (CS-SWCNTs) hybrid film, which attached to the poly-o-phenylenediamine (POPD)-modified Au electrode. The fabricated procedures of the biosensor were characterized with atomic force microscopy (AFM), fourier transform-infrared spectroscopy (FT-IR), and electrochemical impedance spectroscopy (EIS). The cyclic voltammetric results of the biosensor indicated that AFO exhibited a surface-controlled and quasi-reversible electrochemical redox behavior with a formal potential of -0.436 V (vs. Ag/AgCl) in 0.1 mol/L PBS (pH 7.0), which resulted from the direct electron transfer between entrapped AFO and the underlying electrode. The enzymatic electrode exhibited an excellent electrocatalytic response to ST. The linear range of ST determination was from 10 ng/mL to 310 ng/mL with correlation coefficient of 0.997, the detection limit was 3 ng/mL (S/N=3), and the response time was less than 10 seconds. The apparent Michaelis-Menten constant () was estimated to be 7.13 mmol/L. The biosensor had the advantages of good repeatability and stability, remaining 85.6% of its original current value after storage at 4oC for a month, and the RSD for 11 replicate determination of 20 ng/mL ST was 3.9%. This AFO/CS-SWCNTs/POPD/Au modified electrode showed high selectivity and sensitivity in real sample analysis, giving values of recovery in the range of 87.6%-105.5%. The proposed method can be applied to the determination of ST in real samples with satisfactory results.