聚电解质修饰的磁性纳米粒子和蛋白质相互作用的研究

发布时间：2018-01-27 12:45

本文关键词： 磁纳米粒子阳离子聚电解质离子强度蛋白质吸附　出处：《华东理工大学》2015年硕士论文　论文类型：学位论文

【摘要】：蛋白质在固体表面的吸附是当代生物技术、材料及化工领域的研究重点。由于蛋白质表面同时分布有正负电荷,理论上蛋白质可以吸附在任何带电的表面,聚电解质修饰的纳米粒子可以通过静电作用,疏水作用,氢键等选择性吸附蛋白质,因而在蛋白质的分离纯化、体内生物监测和体外生物传感器等领域有广阔的应用前景。本文分别采用原子转移自由基聚合和细乳液聚合的方法,制备了表面接枝聚(甲基丙烯酰氧乙基三甲基氯化铵)(PMATAC)的磁性球刷和包裹在苯乙烯微球里表面接枝聚(2-氨基乙基酯盐酸盐)的磁性聚电解质刷,研究了pH和盐浓度对蛋白质与上述磁刷相互作用的影响以及磁刷与不同蛋白质作用的差别,主要研究内容如下：(1)采用共沉淀法制得油酸修饰的磁性纳米粒子,然后以硅烷偶联剂作为ATRP引发剂,通过配体交换和缩聚法稳定地结合在磁性纳米粒子表面,在CuBr/bpy(2,2-联吡啶)催化体系下,成功制备了表面接枝阳离子聚电解质的磁性纳米粒子,TEM显示合成的磁性球刷具有较好的形貌和大小均一性,动态光散射表明接枝的聚电解质链的长度为20 nm左右,在水溶液中具有很好的分散性,通过磁分离,透析一系列纯化处理后,ICP表明残留的铜离子含量在0.92 ppb,去除量达到99%以上。(2)以p-乳球蛋白(BLG)作为模型蛋白研究了其与PMATAC-Fe3O4磁刷的相互作用。结合浊度滴定,动态光散射的研究方法,定性研究了蛋白质与PMATAC-Fe3O4磁刷作用随pH和盐浓度的变化,还采用等温滴定量热法定量研究了不同离子强度下两者相互作用的热力学参数和吸附量,研究表明其形成复合物的临界pH值(pHc)随盐浓度的增加呈现先增后减的非线性变化,从蛋白质电荷补丁和纳米粒子电荷密度的角度分析了盐浓度对其相互作用的影响。(3)以牛血清蛋白(BSA)为模型蛋白研究了其与PMATAC-Fe3O4磁刷的相互作用。并比较了其与BLG的作用区别,研究发现由于两种蛋白质的表面电荷分布不同,吸附效果受pH和盐浓度的影响程度差别显著,p-乳球蛋白的电荷分布较集中,所以在实验条件下(相同盐浓度)都要优先作用于牛血清蛋白。这使得利用PMATAC-Fe3O4磁刷分离不同蛋白质提供了可能性。(4)采用细乳液聚合和光乳液聚合相结合的方法制备了聚(2-氨基乙基酯盐酸盐)修饰的磁性球刷,由于磁纳米粒子以聚集体的形式包裹在苯乙烯微球里,与单个的磁纳米粒子相比,磁响应更明显,TGA结果显示磁含量为8%左右,聚电解质链长30 nm左右,以牛血清蛋白(BSA)为模型蛋白,通过调节pH与盐浓度能控制牛血清蛋白(BSA)在刷层间的吸附和脱附,高离子强度下不利于蛋白质的吸附和磁性阳离子聚电解质刷的稳定性。与不含磁纳米粒子的聚电解质刷相比,引入磁纳米粒子后的聚电解质刷具有了磁分离的功能。
[Abstract]:The adsorption of protein on solid surface is the research focus in the field of biotechnology, materials and chemical engineering. Because of the distribution of positive and negative charges on the surface of protein, protein can be adsorbed on any charged surface in theory. Polyelectrolyte modified nanoparticles can be selectively adsorbed by electrostatic action, hydrophobic action, hydrogen bond and so on, so they can be separated and purified. In vivo biosensor and in vitro biosensor have broad application prospects. Atom transfer radical polymerization and microemulsion polymerization are used in this paper. The surface grafted poly (methacryloxy ethyl trimethylammonium chloride) magnetic brushes and encapsulated poly (2-aminoethyl ester) hydrochloride on the surface of styrene microspheres were prepared. The magnetic polyelectrolyte brush. The effects of pH and salt concentration on the interaction of protein with the above mentioned magnetic brushes and the differences between the magnetic brushes and different proteins were studied. The main research contents are as follows: (1) oleic acid modified magnetic nanoparticles were prepared by coprecipitation method and then silane coupling agent was used as ATRP initiator. By ligand exchange and Polycondensation, the magnetic nanoparticles were stably bonded to the surface in CuBr-bpy-2-bipyridine catalyst system. The surface grafted cationic polyelectrolyte magnetic nanocrystalline particles (TEM) showed that the synthesized spheroidal brushes had good morphology and uniform size. Dynamic light scattering showed that the length of the graft polyelectrolyte chain was about 20 nm, and it had good dispersion in aqueous solution. After magnetic separation and dialysis, a series of purification treatment was carried out. ICP showed that the residual copper ion content was 0.92 ppb. The interaction with PMATAC-Fe3O4 magnetic brush was studied by using plactoglobulin (BLG) as the model protein, and the turbidity titration was combined. The dynamic light scattering (DLS) method was used to qualitatively study the interaction between protein and PMATAC-Fe3O4 with pH and salt concentration. The thermodynamic parameters and adsorption capacity of the two interactions under different ionic intensities were also quantitatively studied by isothermal drop quantitative calorimetry. The results show that the critical pH value of the complex increases firstly and then decreases with the increase of salt concentration. The effect of salt concentration on protein charge patch and charge density of nanoparticles was analyzed by using bovine serum protein (BSA). The interaction between the model protein and the PMATAC-Fe3O4 brush was studied and the difference between the model protein and BLG was compared. It was found that the surface charge distribution of the two proteins was different, the adsorption effect was influenced by pH and salt concentration, and the charge distribution of p-lactoglobulin was more concentrated. Therefore, under the experimental conditions (the same salt concentration), the bovine serum protein should be preferentially acted on. This makes it possible to separate different proteins by using PMATAC-Fe3O4 magnetic brushes. The magnetic sphere brush modified by poly (2-aminoethyl ester) hydrochloride was prepared by the combination of fine emulsion polymerization and photoemulsion polymerization. Because the magnetic nanoparticles are encapsulated in styrene microspheres in the form of aggregates, the magnetic response is more obvious than that of single magnetic nanoparticles. The results show that the magnetic content is about 8%. The adsorption and desorption of bovine serum protein (BSA) between brushes can be controlled by adjusting pH and salt concentration. High ionic strength is not conducive to protein adsorption and stability of magnetic cationic polyelectrolyte brushes compared with polyelectrolyte brushes without magnetic nanoparticles. The polyelectrolyte brush with magnetic nanoparticles has the function of magnetic separation.
【学位授予单位】：华东理工大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：TB383.1;O631

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