基于聚乙烯醇微球为载体固定化脂肪酶的研究
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摘要
以具有广泛工业应用性和生物相容性的聚乙烯醇(PVA)为原料,利用PVA分子结构特点对其进行改性,进一步采用高速搅拌制备聚乙烯醇微球,采用扫描电镜表征其形貌。采用1,1′-羰基二(1,2,3-三氮唑)将聚乙烯醇微球活化,以活化后的聚乙烯醇微球为载体固定化脂肪酶,并采用聚乙烯醇物理吸附脂肪酶为实验对照组。结果表明:聚乙烯醇微球粒径分布在2～3μm之间,微球表面形态较为光滑。聚乙烯醇微球固定化脂肪酶可重复使用,重复10次酶催化水解反应,相对酶活仍维持在70%以上。固定化脂肪酶的最优催化水解反应温度为30℃,与游离酶相比降低了10℃,最佳催化水解反应pH为8,与游离酶最佳催化pH相比,催化反应条件更温和。
Polyvinyl alcohol(PVA)with good biocompatibility is widely used in industry.PVA was used as raw material to make PVA microsphere.PVA microsphere functionalized with ester imidazole groups was prepared with treatment by carbonyl diazole(CDI).SEM pictures showed that the surface of PVA microsphere was smooth,and had particle size of 2～3μm.The lipase 03(Leveking)immobilized on the actived PVA microsphere exhibited excellent stability and reusability.The enzymatic hydrolysis reaction was repeated for 10 times,and the relative enzyme activity remained above 70%.The optimal temperature of hydrolysis reaction catalyzed by immobilized lipase was 30℃,compared with the free enzyme decreased 10℃.The optimal pH changed from pH 9 of free enzyme to pH 8 of immobilized lipase.The results indicated that PVA microsphere immobilized lipase with more wild reaction conditions could be widely used in insudtry for the lower energy cost.
Polyvinyl alcohol(PVA)with good biocompatibility is widely used in industry.PVA was used as raw material to make PVA microsphere.PVA microsphere functionalized with ester imidazole groups was prepared with treatment by carbonyl diazole(CDI).SEM pictures showed that the surface of PVA microsphere was smooth,and had particle size of 2～3μm.The lipase 03(Leveking)immobilized on the actived PVA microsphere exhibited excellent stability and reusability.The enzymatic hydrolysis reaction was repeated for 10 times,and the relative enzyme activity remained above 70%.The optimal temperature of hydrolysis reaction catalyzed by immobilized lipase was 30℃,compared with the free enzyme decreased 10℃.The optimal pH changed from pH 9 of free enzyme to pH 8 of immobilized lipase.The results indicated that PVA microsphere immobilized lipase with more wild reaction conditions could be widely used in insudtry for the lower energy cost.
引文
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[15]Abrami'ca M,Le2ˇci'cI,Korica T,et al.Purification and properties of extracellular lipase from streptomyces rimosus[J].Enzym Microb Tech,1999,25(6):522-529.
[2]Cardenas J A E,De Castro Alvarrez M S.Screening and catalytic activity in organic synthesis of novel fungal and yeast lipases[J].J Mol Catal B:Enzym,2001,14:111-123.
[3]Osada K T K,Hatano M.Polyunsaturated fatty acid glyceride syntheses by microbial lipase[J].J Am Oil Chem Soc,1990,67:921-922.
[4]Undurraga D M A,Erazo S.Cocoa butter equivalent through enzymic interesterfication of palm oil mid fraction[J].Process Biochem,2001,26:933-939.
[5]Zaborsky O.Immmobilised enzymes[M].Cleveland:CRCPress,1973.
[6]Tan T,Lu J,Nie K,et al.Biodiesel production with immobilized lipase:a review[J].Biotechnology Advances,2010,28(5):628-634.
[7]Lu J,Deng L,Wang F,et al.Stability of immobilized candida sp.99-125lipase for biodiesel production[J].Chem Eng Technol,2012,35(12):2120-2124.
[8]Idris A,Bukhari A.Immobilized candida antarctica lipase b:hydration,stripping off and application in ring opening polyester synthesis[J].Biotechnology Advances,2012,30(3):550-563.
[9]Orive G,Tam S K,Pedraz J L,et al.Biocompatibility of alginate-poly-L-iysine microcapsules for cell therapy[J].Biomaterials,2006,27(20):3691-3700.
[10]Kaity S,Isaac J,Ghosh A.Interpenetrating polymer network of locust bean gum-poly(vinyl alcohol)for controlled release drug delivery[J].Carbohydrate Polymers,2013,94(1):456-467.
[11]Kaity S,Ghosh A.Facile preparation of acrylamide grafted locust bean gum-poly(vinyl alcohol)interpenetrating polymer network microspheres for controlled oral drug delivery[J].Journal of Drug Delivery Science and Technology,2016,33:1-12.
[12]Jeon Y S,Rhim J W.Performance study on membrane capacitive deionization(MCDI)processes using the composite carbon electrodes coated by cation and anion exchange polymers based on PVA[J].Polymer-Korea,2017,41(2):351-360.
[13]Gao L,Kong T F,Huo Y P.Dual thermoresponsive and pH-responsive poly(vinyl alcohol)derivatives:synthesis,phase transition study,and functional applications[J].Macromolecules,2016,49(19):7478-7489.
[14]Sun X X,Xin Y R,Wang X H,et al.Functionalized acetoacetylated poly(vinyl alcohol)monoliths for enzyme immobilization:aphase separation method[J].Colloid and Polymer Science,2017,295(10):1827-1881.
[15]Abrami'ca M,Le2ˇci'cI,Korica T,et al.Purification and properties of extracellular lipase from streptomyces rimosus[J].Enzym Microb Tech,1999,25(6):522-529.