Preparation of Thermo-sensitive Microspheres by Premix Membrane Emulsification for Immobilization of Trypsin
快速膜乳化法制备温敏微球及其固定胰蛋白酶

Thermo-sensitive poly(N-isopropyl acrylamide-co-acrylic acid) P(NIPAM-co-AAc)] microspheres with narrow size distribution were successfully prepared by premix membrane emulsification at room temperature.The results show that the mean diameter of microspheres was 5.2 μm,their polydispersity index as low as 0.0323.The temperature-responsive behaviors of prepared microspheres were characterized.It was confirmed that the lower critical solution temperature(LCST) of P(NIPAM-co-AAc) microspheres decreased with addition of hydrophilic monmer.The higher the monmer amount,the lower the LCST of P(NIPAM-co-AAc) microspheres.The LCST of P(NIPAM-co-AAc) microspheres increased with more addition of N,N’-methylenebisacrylamide.The respond time increased with addition of hydrophilic monmer.Trypsin was covalently immobilized onto the microspheres.The result showed that when the concentrations of 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride,trypsin and PBS were 0.9 mg/mL,1.8 mg/mL and 70 mmol/L,the optimal value of trypsin loading and activity yield,276.10 mg/g and 75.07% respectively,was reached.The highest activity of immobilized trypsin was obtained at pH 8 and 37℃.