几种新型电容生物传感器及复杂样品溶出伏安分析技术的研究与应用
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摘要
化学/生物传感器的研制已成为分析化学和生物化学领域中的一个非常重要和令人感兴趣的课题。其中自组装单层膜,分子印迹技术和纳米材料在这一领域中的应用尤为引人瞩目。本文将具有高选择性的分子印迹技术和可以提供实时响应信息的灵敏的电容传感方式相结合,研制了新型的分子印迹电容传感器; 将纳米材料固载抗体和电容传感相结合,研制了新型的电容免疫传感器; 分别将多孔的自组装膜和电化学聚合膜与电沉积纳米金阵列相结合,实现了在高浓度大分子污染物存在下,用溶出伏安法对痕量铜的灵敏测定; 最后对复杂样品中痕量重金属检测进行了探索。具体地说,本论文主要开展了以下几方面的创新研究工作:
1.将分子印迹技术和电容传感方式相结合,制备了一种新型分子印迹电容传感器,建立了测定谷胱甘肽含量的新方法。详细研究了传感器的制备和响应特性。其线性范围0.025~0.30 mmol L–1,检测限为1.25×10–3 mmol L-1。对人血清中谷胱甘肽的含量进行了直接测定,并取得了良好的结果。基于电容传感技术,建立了两步反应的动力学模型用于描述分析物与印迹位之间的特异性识别过程,并获取了相关的动力学参数。
2.以电合成的聚巯基苯并咪唑膜(PMBI)为介电层,结合分子印迹技术,制备一种新型分子印迹电容传感器用于苯噻草胺的检测。研究了传感器的制备和特性,并讨论了识别过程的动力学特征。此电容传感器制备方法经济简便,并且适用于不同结构的各种分子,有望应用于制备测定其他除草剂的传感器。
3.基于硫醇衍生物分子的头端基团和表面活性剂的相互反应,用电化学法定量评估了自组装单层膜在表面活性剂存在下的抑阻特性。结果表明,通过改变SAMs的表面结构,不同类型的表面活性剂可以用不同的方式来控制抑阻效应和电子传输效率。带有正电荷的表面活性剂降低了负电荷的氧化还原探针与单层膜上负电荷基团的静电斥力,通过增加双电层内的探针浓度而提高了电子传输的可逆性。中性的表面活性剂没有表现出明显的影响,而带负电荷的表面活性剂由于同号电荷的静电斥力而抑阻了探针的靠近和反应。表面活性剂与硫醇衍生物单层膜间的吸附模式,及建议的研究方法可进一步用于研究生物膜的穿透性能。
4.首次利用纳米羟基磷灰石的生物活性和生物兼容性,发展了一种新型的抗体固载方法。结合高灵敏的电容传感方式和共价键合方法,制备一种灵敏的非标记电容免疫传感器,并建立了人转铁蛋白的测定方法。线性范围: 1~100 ng
The fabrication of the chem/biosensor is one of the most promising and interesting areas in the field of analytical chemistry and biochemistry. Especially, the applications of self-assembled monolayer, molecular imprinted technology and nano-materials are distinguished. In this thesis, two new molecularly imprinted capacitive sensors were developed by combining high selectivity molecularly imprinted technology with high sensitivity capacitive transduction. Two new capacitive immunosensors were fabricated profiting from the biocompatibility of electrochemical deposited nano-gold array and capacitive transduction. A selectively penetrated SAM and a nano-gold array were sequentially modified on the glassy carbon electrode, by sensitive stripping voltammetry, trace copper in the presence of excess level of macromolecules was determinated. The analysis of heavy mental ions in complex environmental and biological samples was explored. The main work of this thesis, which was based on the techniques mentioned above, is summarized as follows:
1. A novel biosensor for glutathion direct detection in human serum was proposed by combination of capacitive transduction and molecularly imprinting technique. A two-step kinetic model was successfully derived to describe the recognition process between analyte and imprint sites by capacitive approach. The prospects of future development of this method include further used for the estimation of kinetic and thermodynamic parameters in the MIP recognition process.
2. A new artificial receptor layer was electrosynthesized on the gold electrode to fabricate a capacitive chemosensor. Based on molecular imprinting technique and capacitive transduction, the sensor was applied for herbicide detection. The MIP-modified electrode was selective to mefenacet among other anilide herbicide, and it is applicable to direct mefenacet sensing. Non-MIP-modified electrodes did not show selective response to mefenacet. MIPs proved to be effective as a molecular-recognition element of a chemosensor based on capacitive transduction of the recognized analyte. The preparation of MIPs is simple and low-cost and applicable to various molecules with different structure. The combination of molecular imprinting technique and capacitive
transduction will provide simple, specific, and inexpensive sensing systems. This method can be expected for further applications to other herbicides. 3. The barrier properties and electron transfer of self-assembled thiol derivative monolayer modified electrodes in the presence of different surfactants were quantitatively investigated by seriate electrochemical experiments. It was found that by changing the surface structure of SAMs, different surfactants could regulate the barrier properties and electron-transfer efficiency in different ways. A positively charged surfactant lowers the electrostatic repulsion between the negative redox probes and negatively charged surface groups of the monolayer, enhancing the reversibility of electron transfer by virtue of increasing the redox probe concentration within the electric double-layer region. A neutral surfactant shows no significant effect, while a negative surfactant hinders the access and reaction of a redox probe by an electrostatic repulsion of same-sign charges. The adsorption model between the surfactant and the thiol derivative monolayers and the suggested methods can be further used to study the penetration property of biomembranes. 4. A new antibody immobilization strategy was successfully developed for the fabrication of a label-free capacitive immunosensor. Based on electrodeposition of nanometer-sized bioactive hydroxyapatite, covalently coupled with DS and capacitive transduction, the sensor was applied for human transferrin detection. The fabrication and the specificity of the immunosensor were discussed in detail. Compared with traditional immunoassays, the merit of this approach is that it achieved the real-time monitoring the antibody-antigen interaction and omitted the labeling procedure to simplify the analysis process. The immobilization strategy developed in the present work can be applied for the fabrication of other specific immunosensor. 5. An alumina sol-gel-derived ultrathin film was modified on the gold electrode surface followed by an electrochemical deposited nano-gold array. Anti-human transferrin was further immobilized on the nano-gold array to fabricate a novel capacitive immunosensor. 6. A sensitive and convenient voltammetric approach was developed for trace copper determination free from macromolecule interferences using a doubly
modified GCE. After successively modified with a direct electrochemically deposited Au nanocrystals array and a spontaneously adsorbed MES monolayer film, the resulting GCE achieved a high sensitivity and a large linear response range in the determination of trace copper, even the concentration of coexisting model contaminants (Tween 80 and BSA) reached 500 ppm. Satisfied results were obtained in the detection of copper in the metallurgic wastewater sample. 7. A sensitive and convenient voltammetric approach was developed for trace copper (II) determination freeing from macromolecular interferences at PMBI film/Au nanocrystals modified GCEs. After successively modified with a electrodeposited Au nanocrystals array and a porous PMBI film, the resulting GCE achieved a high sensitivity and a large linear response range in the determination of trace copper, even the concentration of coexisting model contaminants (BSA and Tween 80) reached 500 ppm. The described method provided an alternative choice for the determination of trace copper in the analysis of complex environmental and biological samples by stripping voltammetry.
1.将分子印迹技术和电容传感方式相结合,制备了一种新型分子印迹电容传感器,建立了测定谷胱甘肽含量的新方法。详细研究了传感器的制备和响应特性。其线性范围0.025~0.30 mmol L–1,检测限为1.25×10–3 mmol L-1。对人血清中谷胱甘肽的含量进行了直接测定,并取得了良好的结果。基于电容传感技术,建立了两步反应的动力学模型用于描述分析物与印迹位之间的特异性识别过程,并获取了相关的动力学参数。
2.以电合成的聚巯基苯并咪唑膜(PMBI)为介电层,结合分子印迹技术,制备一种新型分子印迹电容传感器用于苯噻草胺的检测。研究了传感器的制备和特性,并讨论了识别过程的动力学特征。此电容传感器制备方法经济简便,并且适用于不同结构的各种分子,有望应用于制备测定其他除草剂的传感器。
3.基于硫醇衍生物分子的头端基团和表面活性剂的相互反应,用电化学法定量评估了自组装单层膜在表面活性剂存在下的抑阻特性。结果表明,通过改变SAMs的表面结构,不同类型的表面活性剂可以用不同的方式来控制抑阻效应和电子传输效率。带有正电荷的表面活性剂降低了负电荷的氧化还原探针与单层膜上负电荷基团的静电斥力,通过增加双电层内的探针浓度而提高了电子传输的可逆性。中性的表面活性剂没有表现出明显的影响,而带负电荷的表面活性剂由于同号电荷的静电斥力而抑阻了探针的靠近和反应。表面活性剂与硫醇衍生物单层膜间的吸附模式,及建议的研究方法可进一步用于研究生物膜的穿透性能。
4.首次利用纳米羟基磷灰石的生物活性和生物兼容性,发展了一种新型的抗体固载方法。结合高灵敏的电容传感方式和共价键合方法,制备一种灵敏的非标记电容免疫传感器,并建立了人转铁蛋白的测定方法。线性范围: 1~100 ng
The fabrication of the chem/biosensor is one of the most promising and interesting areas in the field of analytical chemistry and biochemistry. Especially, the applications of self-assembled monolayer, molecular imprinted technology and nano-materials are distinguished. In this thesis, two new molecularly imprinted capacitive sensors were developed by combining high selectivity molecularly imprinted technology with high sensitivity capacitive transduction. Two new capacitive immunosensors were fabricated profiting from the biocompatibility of electrochemical deposited nano-gold array and capacitive transduction. A selectively penetrated SAM and a nano-gold array were sequentially modified on the glassy carbon electrode, by sensitive stripping voltammetry, trace copper in the presence of excess level of macromolecules was determinated. The analysis of heavy mental ions in complex environmental and biological samples was explored. The main work of this thesis, which was based on the techniques mentioned above, is summarized as follows:
1. A novel biosensor for glutathion direct detection in human serum was proposed by combination of capacitive transduction and molecularly imprinting technique. A two-step kinetic model was successfully derived to describe the recognition process between analyte and imprint sites by capacitive approach. The prospects of future development of this method include further used for the estimation of kinetic and thermodynamic parameters in the MIP recognition process.
2. A new artificial receptor layer was electrosynthesized on the gold electrode to fabricate a capacitive chemosensor. Based on molecular imprinting technique and capacitive transduction, the sensor was applied for herbicide detection. The MIP-modified electrode was selective to mefenacet among other anilide herbicide, and it is applicable to direct mefenacet sensing. Non-MIP-modified electrodes did not show selective response to mefenacet. MIPs proved to be effective as a molecular-recognition element of a chemosensor based on capacitive transduction of the recognized analyte. The preparation of MIPs is simple and low-cost and applicable to various molecules with different structure. The combination of molecular imprinting technique and capacitive
transduction will provide simple, specific, and inexpensive sensing systems. This method can be expected for further applications to other herbicides. 3. The barrier properties and electron transfer of self-assembled thiol derivative monolayer modified electrodes in the presence of different surfactants were quantitatively investigated by seriate electrochemical experiments. It was found that by changing the surface structure of SAMs, different surfactants could regulate the barrier properties and electron-transfer efficiency in different ways. A positively charged surfactant lowers the electrostatic repulsion between the negative redox probes and negatively charged surface groups of the monolayer, enhancing the reversibility of electron transfer by virtue of increasing the redox probe concentration within the electric double-layer region. A neutral surfactant shows no significant effect, while a negative surfactant hinders the access and reaction of a redox probe by an electrostatic repulsion of same-sign charges. The adsorption model between the surfactant and the thiol derivative monolayers and the suggested methods can be further used to study the penetration property of biomembranes. 4. A new antibody immobilization strategy was successfully developed for the fabrication of a label-free capacitive immunosensor. Based on electrodeposition of nanometer-sized bioactive hydroxyapatite, covalently coupled with DS and capacitive transduction, the sensor was applied for human transferrin detection. The fabrication and the specificity of the immunosensor were discussed in detail. Compared with traditional immunoassays, the merit of this approach is that it achieved the real-time monitoring the antibody-antigen interaction and omitted the labeling procedure to simplify the analysis process. The immobilization strategy developed in the present work can be applied for the fabrication of other specific immunosensor. 5. An alumina sol-gel-derived ultrathin film was modified on the gold electrode surface followed by an electrochemical deposited nano-gold array. Anti-human transferrin was further immobilized on the nano-gold array to fabricate a novel capacitive immunosensor. 6. A sensitive and convenient voltammetric approach was developed for trace copper determination free from macromolecule interferences using a doubly
modified GCE. After successively modified with a direct electrochemically deposited Au nanocrystals array and a spontaneously adsorbed MES monolayer film, the resulting GCE achieved a high sensitivity and a large linear response range in the determination of trace copper, even the concentration of coexisting model contaminants (Tween 80 and BSA) reached 500 ppm. Satisfied results were obtained in the detection of copper in the metallurgic wastewater sample. 7. A sensitive and convenient voltammetric approach was developed for trace copper (II) determination freeing from macromolecular interferences at PMBI film/Au nanocrystals modified GCEs. After successively modified with a electrodeposited Au nanocrystals array and a porous PMBI film, the resulting GCE achieved a high sensitivity and a large linear response range in the determination of trace copper, even the concentration of coexisting model contaminants (BSA and Tween 80) reached 500 ppm. The described method provided an alternative choice for the determination of trace copper in the analysis of complex environmental and biological samples by stripping voltammetry.
引文
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