采用SFAD将碳酸酐酶偶联至无活性官能团聚合物表面的新方法

摘 要 采用异双官能团光偶联剂－Sulfosuccinimidyl (perfluoroazidobenzamido) ethyl-1,3-dithio propionate，SFAD)，将碳酸酐酶（Carbonic anhydrase，CA)接枝在无活性官能团聚合物－聚甲基戊烯（Polymethyl-pentene，PMP)膜式氧合器表面，以对硝基苯酚乙酸酯(p-nitrophenyl acetate,p-NPA)为底物，采用紫外分光光度计测定了接枝CA的活性、浓度、重复利用性、储存稳定性。结果表明，采用SFAD为交联剂能将酶成功地偶联在无活性官能团聚合物表面，在保持酶活性的同时获得较高的接枝效率；共价接枝CA（Covalently immobilized CA，CACI）的浓度随SFAD浓度的增加而增加，最大可达理论单分子层接枝量的85.86％；CACI比物理吸附的CA（Physically adsorbed CA，CAPA）具更好的重复利用性；37°C下, CACI 比CA溶液表现出更好的储存稳定性。

A New Method of Immobilization of Carbonic Anhydrase onto Polymer Surface without Active Functional Groups by Using SFAD as a Crosslinker

ABSTRACT Carbonic anhydrase(CA) was immobilized onto the surface of hollow fiber membrane of polymethyl pentene(PMP) without any active functional groups by using sulfosuccinimidyl perfluoroazidobenzamido] ethyl-1,3-dithiopropionate (SFAD), a photoactivatable heterobifunctional crosslinker. Using p-nitrophenyl acetate (p-NPA) as a substrate, the activity, concentration, storage stability and re-usability of immobilized CA on PMP hollow fibers were studied by ultraviolet spectrophotometer. The preliminary data showed that CA could be successfully introduced onto PMP surface with catalysis activity and immobilization efficiency.The activity of covalently immobilized CA increased with the increase of concentration of SFAD, and the maximum was 85.86% of the theoretical activity of CA spread on PMP surface in monolayer in studied range. Covalently immobilized CA showed higher reusability compared to physically adsorbed CA, and higher storage stability compared to free CA in solution at 37℃.