Development of Fields for Enzymatic Reaction Using Grafted Polymer Chains Immobilizing Enzyme at a High Density

使用高密度固定酶的接枝聚合物链开发酶反应领域

基本信息

批准号：
07555253
负责人：
SAITO Kyoichi
金额：
$ 3.84万
依托单位：
Chiba University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1997
项目状态：
已结题

项目摘要

Extensive research has been carried out on immobilized enzymes because recovery of expensive enzymes after reactions with substrates is not required. Entrapping enzymes in a gel matrix, for example, in a caragenan gel, is a representative immobilization method where the substrate is required to diffuse into the gel to reach the active site of the immobilized enzyme. When an intrinsic reaction rate is fast, the overall reaction rate is governed by the diffusion rate of the substrate into the gel. A convection-aided reaction system using an enzyme immobilized on a porous membrane will realize a higher throughput, because the diffusional path of the substrate to the enzyme can be minimized.The objective of our study was to evaluate a high performance enzyme reaction system using a porous hollow-fiber membrane containing a polymer chain grafted on the pore surface. Here, alpha-amylase was bound by a phenyl group introduced onto the graft chain, and starch was used as a substrate.A phenyl group was introduced onto the polymer chain grafted onto a porous hollow-fiber membrane to bind alpha-amylase. The resulting phenyl-group-containing porous membrane (Ph fiber) had inner and outer diameters of 0.73 and 1.45 mm, respectively. An alpha-amylase solution was forced to permeate radially outward through the pores of the Ph fiber. The amount of alpha-amylase immobilized was 12 mg per g of the Ph fiber. Quantitative hydrolysis of starch during permeation of a 20 g/L starch solution in buffer (0.010 M Tris-HCl + 0.10 M CaCl_2, pH 7) was attained up to a space velocity of 2,200 h^<-1> ; this was achieved at an operating pressure of 0.15 MPa because of negligible diffusional mass-transfer resistance of the starch to the alpha-amylase due to convective flow. Although the activity of alpha-amylase was lowered to 10% by immobilization, the stability of alpha-amylase captured by the graft chain was higher than that of soluble alpha-amylase.

已经对固定酶进行了广泛的研究，因为不需要与底物的反应后恢复昂贵的酶。在凝胶基质中捕获的酶，例如，在卡拉格凝胶中，是一种代表性的固定方法，其中需要底物扩散到凝胶中以到达固定酶的活性位点。当固有反应速率迅速时，总体反应速率受底物扩散速率的影响。使用固定在多孔膜上的酶的对流辅助反应系统将实现更高的吞吐量，因为可以最大程度地减少底物向酶的扩散路径。我们的研究的目的是评估使用含有多孔的空心含量的高性能酶反应系统，其中包含Polymer链链链孔，并在表面上进行了孔。在这里，α-淀粉酶由引入移植链引入的苯基结合，并将淀粉用作底物。将苯基组引入移植到多孔的空心纤维膜上的聚合物链上，以结合α-淀粉酶。所得的含苯基的多孔膜（pH纤维）的内径和外径分别为0.73和1.45 mm。 α-淀粉酶溶液被迫通过pH纤维的孔向外渗透。固定的α-淀粉酶的量为每克pH纤维12毫克。在缓冲液（0.010 m Tris-HCl + 0.10 M CaCl_2，pH 7）中渗透到20 g/l淀粉溶液期间的淀粉的定量水解被达到2,200 H^<-1>的空间速度；由于对流流动，淀粉对α-淀粉酶的扩散质量转移性可忽略不计，因此在0.15 MPa的工作压力下实现了这一点。尽管通过固定化将α-淀粉酶的活性降低至10％，但通过移植链捕获的α-淀粉酶的稳定性高于可溶性α-淀粉酶的稳定性。