基于新型碳纳米复合材料构建电致化学发光酶生物传感器的研究

发布时间：2018-01-16 04:10

本文关键词：基于新型碳纳米复合材料构建电致化学发光酶生物传感器的研究　出处：《西南大学》2015年硕士论文　论文类型：学位论文

【摘要】：电致化学发光(Electrochemiluminescence,ECL)是在电极表面施加一定的电压发生化学反应从而产生一些特殊的电极产物,这些电极产物之间或者电极产物与体系中某组分之间通过电子传递形成激发态,由激发态跃迁返回基态过程中产生的一种光辐射。它是电化学方法和化学发光方法相结合形成的一种新型检测技术,具有灵敏度高、选择性好、过程易于控制、操作简单等优点。酶生物传感器是以固定化酶膜为生物识别元件、修饰电极为信号转换器件,因此其不仅具有酶的分子识别功能和选择催化功能,还具有电极响应迅速、易于操作等特点。ECL酶生物传感器将ECL检测技术与酶的高度专一性和高效催化性融合在一起,具有高灵敏度、高选择性等特点。随着纳米技术的快速发展,各种纳米材料已被广泛应用于生物传感器领域,在表现纳米材料各种优势性能(如大的比表面积、独特的电化学性质、好的生物相容性等)的基础上为生物传感器的发展开辟了广阔的应用前景并实现了生物传感器的高灵敏检测。其中,碳纳米管和石墨烯均具有特殊的物理和化学性质,具有良好的导电性和电催化特性,能够使电子在固定化酶与电极之间进行快速而有效地传递。同时,将基于碳纳米材料研制的新型纳米复合材料应用于酶生物传感器的构建中可为酶的固定提供良好的微环境,从而保持酶的生物活性。基于以上考虑,本文研制了基于碳纳米材料的新型纳米复合材料并构建了一系列灵敏的ECL酶生物传感器,且性能都较以往的酶生物传感器有很大程度上的提高。本论文主要研究工作如下：1.基于Hemin功能化多壁碳纳米管纳米复合材料的电致化学发光胆固醇生物传感器该工作结合多壁碳纳米管(MWCNTs)对过氧化氢(H202)具有电催化氧化活性以及Hemin对H202电化学氧化从而增强Luminol-H2O2体系的ECL信号的特性,制备了新型的Hemin-MWCNTs纳米复合材料,并将其应用于胆固醇的检测。首先,合成Hemin-MWCNTs纳米复合材料,将其滴涂于预先处理好的裸玻碳电极表面,待电极在室温下干燥后再在其表面滴涂胆固醇氧化酶并于4℃孵育若干小时即可。由于Hemin-MWCNTs对H2O2有很好的电催化性能,可增强体系的ECL信号。在最优实验条件下,该传感器的ECL响应与胆固醇浓度在3×10-7～1.2×10-3 mol·L-1范围内呈现良好的线性关系,检测限为1×10-7 mol·L-1。此外,该传感器具有灵敏度高\准确度及稳定性好等优点。2.基于碳纳米管-氧化石墨烯.硫堇-纳米金的电致化学发光胆固醇生物传感器首先,碳纳米管与氧化石墨烯(GO)以质量比为1：1结合成纳米复合物可展现出良好的电催化活性。其次,某些还原剂(如肼)有毒,因此必须控制用量。由于硫堇(Thionine,Thi)结构中有大量氨基以及GO结构中的大量羟基,可作为一些物质(例如氯金酸)的还原剂,还原得到的纳米金(AuNPs)对于Luminol - H2O2体系的ECL有极大的增强作用,并且GO作为还原剂还原氯金酸的同时,还可以作为AuNPs的生长模板。基于以上两点,在未加入其它还原剂的情况下,利用GO和Thi自身的还原性,原位还原氯金酸制备了MWCNTs-GO-Thi-Au纳米复合材料,并将该纳米复合材料应用于胆固醇的检测。由于该纳米复合材料对Luminol - H2O2体系的ECL信号具有放大作用,使得检测限大大降低。在最优实验条件下,该传感器呈现了较宽的线性范围(0.15～828μmol·L-1)和相对较低的检测限(50nmol·L-1)。结合纳米复合物以及ECL检测方法的优势,将本实验所构建的生物传感器应用于临床生物检测具有巨大的潜在价值。3.基于生物功能化的AMs-ChO生物复合材料的电致化学发光胆碱生物传感器由于石墨烯(GR)和AuNPs对于Luminol - H2O2体系的ECL具有极大的促进作用,并且壳聚糖(CS)带正电荷,可通过静电作用与带负电荷的酶或者含酶的生物复合材料结合,因此该工作首先将GO、氯金酸和CS通过电化学还原沉积修饰到电极表面(dpGR-AuNPs-CS)。其次,由于Fe304具有过氧化物模拟酶的特性,可催化氧化H2O2成为各种活性氧分子,因为传统的酶易失活,采用Fe304可以很好的避免酶失活的不足。另外,Ti02能增强Luminol - H2O2体系ECL信号,因此制备了带负电荷的Fe3O4-TiO2-ChO (AMs-ChO)生物复合材料,并能成功固载到带正电荷的电极表面,构建了一种新型的ECL胆碱生物传感器。该传感器显示出较高的灵敏度,线性范围是0.003～1120 μmol·L-1,检出限低至1 nmol·L-1。该传感器具有良好的稳定性,选择性和很高的灵敏度,在临床诊断方面有较好的应用前景。
[Abstract]:Electrochemiluminescence (Electrochemiluminescence, ECL) is a certain voltage is applied on the electrode surface chemical reaction to produce some special product of electrode, between these electrodes or electrode product product and system between a component formed by electron transfer from excited state, excited state transition back to produce ground state in the process of a light radiation. It is the electrochemical method and chemiluminescence method combined with the formation of a new detection technique, which has high sensitivity, good selectivity, easy control, simple operation and other advantages. Enzyme biosensors with immobilized enzyme membrane as biological recognition element, modified electrode for signal converter, so the function of molecular recognition and selection not only has the catalytic function of the enzyme electrode, has quick response, easy operation,.ECL biosensor will be highly specific and ECL detection technology with enzyme Catalytic performance together with high sensitivity, high selectivity and so on. With the rapid development of nanotechnology, nano materials have been widely used in the field of biosensors, in the performance of nano materials of various advantages (such as a large surface area, unique electrical and chemical properties, good biocompatibility etc.). The basis for the development of biosensor has opened up a broad application prospect and high sensitive detection of the biosensor. Among them, carbon nanotubes and graphene have particular physical and chemical properties, good electrical conductivity and catalytic properties, which enables electrons between the immobilized enzyme and the electrode transfer quickly and effectively. At the same time, the construction of development based on carbon nano materials nano composite materials used in enzyme biosensor can provide good microenvironment for enzyme immobilization and maintain the enzyme from Biological activity. Based on the above considerations, this paper developed a new type of nano composite materials based on carbon nano materials and constructed a series of sensitive ECL biosensor, and the performance is compared to previous enzyme biosensor has largely improved. The main research work of this thesis are as follows: 1. multi walled carbon nanotubes composites based on the Hemin function of the electrochemiluminescence of the cholesterol biosensor work combined with multi walled carbon nanotubes (MWCNTs) on hydrogen peroxide (H202) with electro catalytic oxidation activity and Hemin on the electrochemical oxidation of H202 enhanced ECL signal characteristics of Luminol-H2O2 system, Hemin-MWCNTs new type nanocomposites were prepared and applied to detect cholesterol at first, the synthesis of Hemin-MWCNTs nano composite materials, it will drop onto the glassy carbon electrode surface treated in advance, the electrode was dried at room temperature after The surface coating of cholesterol oxidase and in 4 hours of incubation. C because Hemin-MWCNTs has good electrocatalytic properties of H2O2 enhanced ECL signal system. Under the optimum conditions, the ECL response of the sensor and the concentration of cholesterol in 3 * 10-7 to 1.2 * 10-3 mol - L-1 range show a good linear relationship, detection limit of 1 * 10-7 mol L-1.. In addition, the sensor has high sensitivity, good accuracy and stability of the advantages of.2. such as carbon nanotubes and graphene oxide based on thionine gold nanoparticles electrochemiluminescence cholesterol biosensor first, carbon nanotubes and graphene oxide (GO) in the mass ratio of 1:1 with nano composites can exhibit good electrocatalytic activity. Secondly, some reducing agent (such as toxic hydrazine), so we must control the amount. The thionine (Thionine, Thi) has a large number of hydroxyl amino and GO structure in large structure The base, can be used as some substances (e.g. chloroauric acid) of the reducing agent, reducing gold nanoparticles (AuNPs) have obtained the enormous enhancement effect for the Luminol - H2O2 system ECL, and GO as the reducing agent and reduction of chloroauric acid, can also be used as growth template AuNPs. Based on the above two points, without adding the other reducing agent, by reducing GO and Thi, MWCNTs-GO-Thi-Au nanocomposites were prepared by in situ reduction of chloroauric acid system, and the application of the nanometer composite material to detect cholesterol. Because the ECL signal of the nano composite materials of Luminol - H2O2 system has the function of amplification, the detection limit is greatly reduced under optimal conditions, the sensor showed a wide linear range (0.15 ~ 828 mol - L-1) and low detection limit (50nmol - L-1). The combination of nano composites and ECL method for detection of advantage, will this experiment. The application of biosensor in clinical biological detection has the potential value of.3. huge AMs-ChO biological composite biological functions of the ECL choline biosensor based on graphene (GR) and AuNPs for Luminol - H2O2 system ECL has great role in promoting, and chitosan (CS) with positive charge can be. Through the combination of biological composite electrostatic interaction with the negatively charged enzyme or enzyme, so the work of the GO, chloroauric acid and CS by electrochemical deposition of modified electrodes (dpGR-AuNPs-CS). Secondly, due to the nature of the Fe304 mimetic peroxidase, which can catalyze the oxidation of H2O2 into reactive oxygen molecules. Because the traditional enzyme deactivation, Fe304 can well avoid the lack of enzyme inactivation. In addition, Ti02 can enhance the Luminol - H2O2 system ECL signal, thus the preparation of the negatively charged Fe3O4-TiO2-Ch O (AMs-ChO) composite material, and can be successfully immobilized onto the electrode surface with positive charge, construction of a new type of ECL choline biosensor. The sensor showed high sensitivity, linear range is 0.003 ~ 1120 mol - L-1, the detection limit as low as 1 nmol. The sensor has a good L-1. the stability, selectivity and high sensitivity, and has a good application prospect in clinical diagnosis.

【学位授予单位】：西南大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：TB33;TP212.3

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