【作者】 陆杨；

【导师】 姜忠义；

【作者基本信息】 天津大学 ， 化学工艺， 2006， 硕士

【摘要】 海藻酸(ALG)钙凝胶具有生物相容性好、价格低廉、材料易得、制备简便等优点,已成为使用最为广泛的酶固定化载体之一。但其突出缺点是结构松散、孔径大,酶泄漏严重,且易发生溶胀。这些缺点严重限制了海藻酸凝胶固定化酶的使用寿命和进一步的应用。为克服上述缺点,本研究根据酶固定化载体设计中“亲疏平衡”和“刚柔相济”的思想,在海藻酸凝胶中添加不同亲疏水性、结构形态的纳米管(碳纳米管,CNTs;二氧化硅纳米管,SiNTs)和纳米粒子(二氧化硅胶体,SG;石墨)制备出新型的无机-有机杂化凝胶:海藻酸-碳纳米管杂化凝胶(ALG-CNTs gel),海藻酸-二氧化硅纳米管杂化凝胶(ALG-SiNTs gel),海藻酸-硅胶颗粒杂化凝胶(ALG-SG gel)和海藻酸-石墨杂化凝胶(ALG-graphite gel),并通过SEM,EDX,FTIR、SEM,DMA和BET等多种表征手段对空白及杂化海藻酸凝胶的结构形态、物化性质进行了研究。并通过非稳态扩散模型研究底物NADH在空白及杂化海藻酸凝胶的扩散行为。研究结果表明,杂化后,凝胶的机械强度显著增强,底物在凝胶中的有效扩散系数也大大提高。首先采用先吸附、后包埋的方法将模型蛋白(牛血清白蛋白,BSA)包埋于空白及杂化海藻酸凝胶中,通过对各种凝胶在固定化过程中BSA泄漏率的研究发现,由于杂化介质对BSA分子和水分子的吸附,杂化海藻酸凝胶中的BSA泄漏率显著下降,且凝胶中BSA泄漏率从高到低依次为:ALG > ALG-graphite > ALG-SG > ALG-CNTs > ALG-SiNTs。采用筛选出的ALG-SiNTs和ALG-SG杂化海藻酸凝胶进一步包埋酵母醇脱氢酶,并对空白及杂化海藻酸凝胶中酶泄漏率、酶促反应最适温度和pH值、酶催化活性、动力学性质、循环使用和储存稳定性进行了研究。研究结果显示,与空白海藻酸凝胶相比,ALG-SiNTs杂化凝胶中的酶泄漏率降低了50%。此外,ALG-SiNTs杂化凝胶固定化酶与底物的亲和力、酶活保持率、贮存和循环使用稳定性均显著高于空白海藻酸凝胶和ALG-SG杂化凝胶两种载体。更多还原

【Abstract】 Calcium alginate (ALG) gel is one of the most commonly used carriers in the entrapment immobilization of enzyme owing to their significant advantages such as good biocompatibility, low cost, easy availability, and simplicity of preparation. However, some disadvantages are often associated with this carrier, including high enzyme leakage, low mechanical strength, and serious swelling due to their open structure, large pore size. These disadvantages significantly restrict the lifetime of these enzymes immobilized in alginate gels and limit its further application.To overcome these disadvantages, in this paper, novel and efficient Organic-inorganic hybrid gels consisting of alginate gels doped with nanotubes or nanoparticles were prepared by incorporating nanotubes (carbon nanotubes, CNTs; silica nanotubes, SiNTs) and nanoparticles (silica gel, SG; graphite) into alginate gels. The hybrid alginate gel prepared in the present study included ALG-CNTs gel, ALG-SiNTs gel, ALG-SG gel and ALG-graphite gel. The structures and morphologies, chemical and physical properties of pure and hybrid alginate gels were evaluated by scanning electron microscopy (SEM) equipped with energy dispersive X-ray spectroscopy (EDX), Fourier Transform IR spectrophotometer (FTIR), dynamic mechanical analysis (DMA) and BET nitrogen adsorption-desorption method. In addition, the diffusion characterisitics of NADH in pure and hybrid alginate gels were determined according an unsteady-state model. And characterization results showed that the mechanical properties of these hybrid gels were remarkably reinforced and effective diffusion coefficient for substrates were significantly increased.To deeply understand the effect of doping on the enzyme leakage, pure and hybrid alginate gels were firstly employed to encapsulate bovine serum albumin (BSA). During the encapsulation, first BSA was adsorbed on the dopants and then BSA and dopants were suspended in ALG solution, followed by Ca2+ cross-linking to form biocomposites. It was found that BSA leakage decreased significantly after immobilized in these hybrid alginate gels compared to that from ALG gel, which was mainly due to, firstly, the protein molecule was adsorbed on the dopants which could not be washed out easily, secondly, during the gel formation, water loss in hybrid alginate gels became less than that in pure alginate gel. And the BSA leakages in these gels occurred in the following order: ALG > ALG-graphite > ALG-SG >更多还原