基于多酚氧化酶-金纳米粒子-壳聚糖杂化膜-石墨烯掺杂碳糊电极的敏感双酶生物传感器用于检测氨基甲酸酯。

Sensitive bi-enzymatic biosensor based on polyphenoloxidases-gold nanoparticles-chitosan hybrid film-graphene doped carbon paste electrode for carbamates detection.

机构信息

REQUIMTE, Instituto Superior de Engenharia do Porto, Instituto Politécnico do Porto, Rua Dr. Bernardino de Almeida 431, 4200-072 Porto, Portugal; GELCORR, Dep. de Química Analítica e Físico-Química, Centro de Ciências, Universidade Federal do Ceará, Bloco 940, Campus do Pici, 60455-970 Fortaleza, CE, Brazil.

REQUIMTE, Instituto Superior de Engenharia do Porto, Instituto Politécnico do Porto, Rua Dr. Bernardino de Almeida 431, 4200-072 Porto, Portugal; REQUIMTE, Dep. de Ciências Químicas, Faculdade de Farmácia, Universidade do Porto, Rua de Jorge Viterbo Ferreira 228, 4050-313 Porto, Portugal.

出版信息

Bioelectrochemistry. 2014 Aug;98:20-9. doi: 10.1016/j.bioelechem.2014.02.003. Epub 2014 Mar 5.

Abstract

A bi-enzymatic biosensor (LACC-TYR-AuNPs-CS/GPE) for carbamates was prepared in a single step by electrodeposition of a hybrid film onto a graphene doped carbon paste electrode (GPE). Graphene and the gold nanoparticles (AuNPs) were morphologically characterized by transmission electron microscopy, X-ray photoelectron spectroscopy, dynamic light scattering and laser Doppler velocimetry. The electrodeposited hybrid film was composed of laccase (LACC), tyrosinase (TYR) and AuNPs entrapped in a chitosan (CS) polymeric matrix. Experimental parameters, namely graphene redox state, AuNPs:CS ratio, enzymes concentration, pH and inhibition time were evaluated. LACC-TYR-AuNPs-CS/GPE exhibited an improved Michaelis-Menten kinetic constant (26.9±0.5M) when compared with LACC-AuNPs-CS/GPE (37.8±0.2M) and TYR-AuNPs-CS/GPE (52.3±0.4M). Using 4-aminophenol as substrate at pH5.5, the device presented wide linear ranges, low detection limits (1.68×10(-9)±1.18×10(-10)-2.15×10(-7)±3.41×10(-9)M), high accuracy, sensitivity (1.13×10(6)±8.11×10(4)-2.19×10(8)±2.51×10(7)%inhibitionM(-1)), repeatability (1.2-5.8% RSD), reproducibility (3.2-6.5% RSD) and stability (ca. twenty days) to determine carbaryl, formetanate hydrochloride, propoxur and ziram in citrus fruits based on their inhibitory capacity on the polyphenoloxidases activity. Recoveries at two fortified levels ranged from 93.8±0.3% (lemon) to 97.8±0.3% (orange). Glucose, citric acid and ascorbic acid do not interfere significantly in the electroanalysis. The proposed electroanalytical procedure can be a promising tool for food safety control.

摘要

一种双酶生物传感器(LACC-TYR-AuNPs-CS/GPE)通过在掺杂石墨烯的碳糊电极(GPE)上电沉积混合膜一步制备而成。通过透射电子显微镜、X 射线光电子能谱、动态光散射和激光多普勒测速法对石墨烯和金纳米粒子(AuNPs)进行形态学表征。电沉积的混合膜由漆酶(LACC)、酪氨酸酶(TYR)和金纳米粒子(AuNPs)包埋在壳聚糖(CS)聚合物基质中组成。评估了实验参数,即石墨烯的氧化还原状态、AuNPs:CS 比、酶浓度、pH 值和抑制时间。与 LACC-AuNPs-CS/GPE(37.8±0.2M)和 TYR-AuNPs-CS/GPE(52.3±0.4M)相比,LACC-TYR-AuNPs-CS/GPE 的米氏常数(26.9±0.5M)得到了提高。在 pH5.5 下使用 4-氨基酚作为底物,该装置呈现出宽的线性范围、低检测限(1.68×10(-9)±1.18×10(-10)-2.15×10(-7)±3.41×10(-9)M)、高准确性、灵敏度(1.13×10(6)±8.11×10(4)-2.19×10(8)±2.51×10(7)%inhibitionM(-1))、重复性(1.2-5.8%RSD)、重现性(3.2-6.5%RSD)和稳定性(约 20 天),可根据其对多酚氧化酶活性的抑制能力来测定柑橘类水果中的甲萘威、甲酰胺盐酸盐、丙虫磷和福美锌。在两个加标水平的回收率范围为 93.8±0.3%(柠檬)至 97.8±0.3%(橙子)。葡萄糖、柠檬酸和抗坏血酸在电分析中不会产生显著干扰。所提出的电化学生物分析程序可以成为食品安全控制的一种有前途的工具。

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