A Study On Optimizing Of Lipase Immobilization Process

Lipase is a green,efficient biocatalyst that used widely in industry including the food,medicine,leather,environmental protection,fabric processing,biodiesel production and so on.However,free lipase is unstable,difficult to recycle and to continue operation,difficult to perform the large-scale industrial production.These disadvantages could be better overcome by immobilizing the lipase.Therefore,the study on the enzymatic immobilization has become a focus.The immobilization technology of lipase and the properties of immobilized lipase were studied in this paper.The main points were as follows:?1?Sodium alginate and CMC used as carriers,EGDE as cross-linking agent to immobilize the lipase were studied.The optimum conditions for the preparation of immobilization lipase were as follows:sodium alginate concentration 2.5%,CMC concentration 1.5%,lipase concentration 10 mg/mL,solution pH=7.0,CaCl₂ concentration 5%,EGDE concentration 0.02%,and immobilization time 30 min.Under the optimal conditions,the activity of immobilized lipase was about 379.34 U/g,and the recovery of the enzymatic activity of the immobilized lipase reached to about 50.09%.The optimal reaction pH of immobilized lipase was at 8.5 and the optimal temperature was at 45?that were higher compared with the free lipase.The thermal stability of immobilized lipase was better than that of the free lipase.The immobilized lipase could retain still about 60%activities after reused for seven times.?2?The properties of glutaraldehyde cross-linked immobilized lipase,EGDE cross-linked immobilized lipase and uncross-linked immobilized lipase were studied.Based on the activity of uncross-linked immobilized lipase was as 100%,the activity of the glutaraldehyde cross-linked immobilized lipase and the EGDE cross-linked immobilized lipase was respectively as 118.46%and 134.00%.The activity of glutaraldehyde cross-linked,EGDE cross-linked and uncross-linked immobilized lipase was respectively 47.24%,59.66%and39.42%of the initial activity after reused for seven times.?3?The domestic epoxy resin LXEP-120 was used to immobilize the lipase.The optimal immobilization technology was as follows:the potassium phosphate buffer with 1 mol/L,pH 6.0,2.0 mg/mL of the concentration of lipase,the quantity of epoxy resin was 5.0 g?4 m L lipase solution?,the temperature was 22?,and the immobilization time was 12 h.Under these conditions,the activity of immobilized lipase was about 398.46 U/g,the recovery rate of the enzymatic activity reached to 92.79%.The optimal reaction pH of immobilized lipase was at 8.0,and the optimal reaction temperature was at 45?with 5?higher than that of the free lipase.The acid-alkali tolerance,the thermal and operating stability of immobilized lipase were significantly increased than that of the free lipase.?4?After the immobilization,glycine was used to block the epoxy resin groups that were not covalently linked to the enzyme molecules.Glycine incubation was found to significantly improve the thermal stability of the immobilized lipase,but not any effects appeared to the acid-alkali tolerance and re-use rate of the immobilized lipase.The optimal glycine incubation technology was that the glycine with 2.5 mol/L,pH 7.0,was incubated with the resin for 24h at 25?.