碳酸酐酶在聚甲基戊烯中空纤维膜表面的固定及其性能分析

Immobilization and characterization of carbonic anhydrase on the surface of hollow fiber membrane of polymethyl pentene

摘要:

采用两步法将碳酸酐酶共价键合在聚甲基戊烯(Polymethyl-pentene, PMP)膜式氧合器表面以提高其清除血液中CO2的能力。首先采用等离子体处理法将羟基引入PMP表面, 然后用偶联剂溴化氰(CNBr)将碳酸酐酶(Carbonic anhydrase, CA)固定在PMP膜表面。采用XPS、表面接触角测定仪对等离子体处理后材料表面的物理化学性质进行了表征。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, 等离子体处理方法能将羟基成功引入PMP表面; CA能被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CNBr浓度的增加而增加, 最大可达理论单分子层接枝量的73%; CACI比物理吸附的CA(Physically adsorbed CA, CAPA)具更好的重复利用性; 37oC下, CACI比CA溶液表现出更好的储存稳定性。本方法有望应用在膜式氧合器上以提高其对血液中CO2的排除效率。

Abstract:

We immobilized carbonic anhydrase (CA) onto the surface of membrane oxygenator of polymethyl pentene (PMP) to enhance the removal of carbon dioxide in blood by two steps. We first introduced hydroxyl groups onto PMP surface by water plasma treatment, and then coupled CA onto PMP surface by using cyanate bromide (CNBr) as a crosslinker. After plasma treatment, the contact angle with water and chemical composition of PMP surface were characterized by analysis system of surface contact angle and XPS. 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 hydroxyl groups could be introduced on the surface of PMP by water plasma treatment, and CA with catalysis activity could be successfully introduced onto PMP surface in high immobilization efficiency. The activity of covalently immobilized CA increased with the increase of concentration of CNBr, and the maximum was 73% 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 37oC. The method would be used potentially in the membrane oxygenator to improve the capacity of removal of carbon dioxide in blood in the future.