【作者】 王振刚；

【导师】 徐志康；

【作者基本信息】 浙江大学 ， 高分子化学与物理， 2008， 博士

【摘要】 鉴于它们在生物反应器和生物传感器等领域的良好应用前景,高分子分离膜的制备及其酶固定化研究在近几十年来备受关注。膜材料的形态结构和化学组成对于改善固定化酶的性能具有重要的意义。本论文旨在探索丙烯腈共聚物分离膜形态结构和化学组成的调控方法,以期得到理想的固定化酶膜,并研究影响固定化酶性能的内在因素。分别使用浸没沉淀相转化和静电纺丝法制备了丙烯腈/丙烯酸共聚物不对称膜和纤维膜,采用场发射扫描电镜(FESEM)研究膜的形态结构,纯水通量和牛血清白蛋白截留实验研究膜的渗透、分离性能。结果表明:就不对称膜而言,以水作为制膜液添加剂可显著改善膜的形态结构和渗透性能,并维持其分离特性;水和PEG或丙三醇组合则具有协同效应;就纤维膜而言,纺丝液浓度和组成显著影响纤维的形态和直径。选取纤维膜作为酶固定化载体,分别通过掺杂碳纳米管和蛋白质仿生修饰对膜进行改性,并用FESEM、透射电镜、激光共聚焦显微镜和紫外-可见光谱进行表征。研究发现掺杂碳纳米管和仿生修饰都可提高固定化酶的活性。此外,碳纳米管掺杂可提高酶的储存稳定性。仿生修饰可提高酶的热稳定性,牛血清清蛋白与胶原蛋白的修饰对酶热稳定性和储存稳定性影响迥异。结合原子力显微镜、水接触角、荧光光谱、圆二色谱和紫外-可见光谱分析,发现酶-蛋白质和酶-碳纳米管相互作用可在很大程度上影响固定化酶的构象和活性中心。将纤维膜应用于葡萄糖传感器的制备,使用计时电流法研究传感器响应时间、响应强度、灵敏度、线性检测范围、动力学参数和使用稳定性,发现掺杂碳纳米管可显著提高传感器的响应强度和灵敏度。本论文为高分子分离膜形态结构和化学性质的调控提供了可行的方案,并为固定化酶膜在生物传感器中的应用打下了基础;载体与酶相互作用的研究在一定程度上促进了酶固定化研究的深入。更多还原

【Abstract】 Polymer membranes for enzyme immobilization have attracted much attention in recent years, especially in the fields of bioreactor and biosensor. The morphology and chemical structure of membranes play a significant role in the performance of enzyme-immobilized membranes. Considering these, in this thesis, novel strategies were explored to tailor the membrane structures for improving the properties of enzyme-immobilized membranes. Furthermore, the intercorrelation between enzymes and the supports were studied.First, immersion precipitation phase inversion and electrospining were used to fabricate polyacrylonitrile-based asymmetric and fibrous membranes, respectively. The morphologies of the membranes were characterized by field emission electron microscopy (FESEM). Pure water flux and BSA rejection experiment were applied to study the properties of the asymmetric membranes. It was found that the nonsolvent additive, water, could obviously improve the permeation performance of the asymmetric membranes, while retaining the BSA rejection. The mixtures of water with PEG or glycerol show cooperative effect on the permeation performance and morphology of the membranes. For the fibrous membrane, on the other hand, the concentration and composition of spinning solution also have a significant impact on the diameter and morphology of the fibers.Second, fibrous membranes were selected for enzyme immobilization. Influences of pre-modified through Multiwalled Carbon Nanotubes (MWCNTs) filling or protein-involved biomimetic tethering were explored. The membranes were characterized by FESEM, transmission electron microscopy, confocal laser scanning microscopy and UV-vis spectra. It was found that the both modifications increase the activities of the immobilized enzymes. In addition, MWCNTs filling improves the storage stabilities and biomimetic tethering enhances the operational stabilities. However, different proteins tethering have distinct effects on the thermal and storage stabilities of the enzymes. The enzyme-support interactions were studied by AFM, water contact angle measurement, circular dichroism, fluorescence and UV-vis spectra. The results indicate that the interactions largely decide the conformations and active sites of the enzymes, which affect their performance in turn. Subsequently, the enzyme-immobilized fibrous membranes were applied in glucose sensor and chronoamperometry was used to study the parameters of the biosensor. It was found that MWCNTs filling improves the current and sensitivity.To sum up, this thesis has established several effective methods to tailor the morphological and chemical properties of the membranes for enzyme immobilization. It has also explored a foundation for the application of fibrous membranes in biosensor. Furthermore, the study of the enzyme-support interactions will promote the design of supports for enzyme immobilization.更多还原