脂肪酶固定化工艺优化及其酶学性质研究
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摘要
以D-101、AB-8和S-8三种大孔树脂为载体,进行黑曲霉脂肪酶的固定化研究。根据脂肪酶的固定化率及活力回收率,确定D-101为固定化载体。经响应曲面法优化得脂肪酶的固定化工艺:以5 g经预处理的D-101为载体,加入9.0 m L酶液(20 mg/m L),p H 7.6,39℃下吸附4.3 h,脂肪酶固定化率为95.11%,固定化脂肪酶活力回收率为101.36%。固定化酶最适反应温度升高2℃,最适p H不变,固定化脂肪酶的酸碱稳定性和热稳定性均优于游离酶。
Three macroporous resins(D-101, AB-8 and S-8) were invesgated as the carriers of the immobilized Aspergillus niger lipase, the rate of immobilized lipase and the activity recovery ratio were taken as indexes, we selected D-101 as the carrier for the immobilization of lipase, the immobilization conditions of and characterization of the immobilized lipase were studied. By response surface analysis, the optimum immobilization conditions were obtained as follows: 5 g pre-treated D-101, 9.0 m L lipase solution(20 mg/m L), p H 7.6, 39 ℃and 4.3 h. Under these conditions, the rate of immobilized lipase and the activity recovery ratio were 95.11 %and 101.36 % respectively. The optimum temperature of the immobilized lipase increased 2 ℃, and the temperature and p H stability of the immobilized enzyme were better than free emzyme.
Three macroporous resins(D-101, AB-8 and S-8) were invesgated as the carriers of the immobilized Aspergillus niger lipase, the rate of immobilized lipase and the activity recovery ratio were taken as indexes, we selected D-101 as the carrier for the immobilization of lipase, the immobilization conditions of and characterization of the immobilized lipase were studied. By response surface analysis, the optimum immobilization conditions were obtained as follows: 5 g pre-treated D-101, 9.0 m L lipase solution(20 mg/m L), p H 7.6, 39 ℃and 4.3 h. Under these conditions, the rate of immobilized lipase and the activity recovery ratio were 95.11 %and 101.36 % respectively. The optimum temperature of the immobilized lipase increased 2 ℃, and the temperature and p H stability of the immobilized enzyme were better than free emzyme.
引文
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[18]刘汝宽.脂肪酶固定化技术研究[D].郑州:河南工业大学,2007
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[20]贾佳,尹德忠,张宝亮,等.大孔树脂固定木瓜蛋白酶的研究[J].材料导报,2013,27(22):88-91
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[22]罗贵明.酶工程[M].北京:化学工业出版社,2003
[2]Rao P,Divakar S.Lipase catalyzed esterification of a-terpineol with various organic acids:application of the Plackett-Burman design[J].Process Biochem,2001,36(11):1125-1128
[3]Zhang L Q,Zhang Y D,Xu L,et al.Lipase catalyzed synthesis of RGD diamide in aqueous water-miscible organic solvents[J].Enzyme Microb Technol,2001,29(2):129-135
[4]Weber N,Klein E,Mukerjee K D.Long chain acyl thioesters prepared by solvent free thioesterification and transesterification catalyzed by microbial lipases[J].Appl Microbiol Biotechnol,1999,51(3):401-404
[5]刘光.黑曲霉脂肪酶的分离纯化及其性质研究[D].广州:华南理工大学,2013
[6]Sagiroglu A.Conversion of sunflower oil to biodiesel by alcoholysis using immobilized lipase[J].Artificial Cells Blood Substitutes and Biotechnology,2008,36(2):138-149
[7]罗文,谭天伟,袁振宏,等.多孔玻璃珠固定化脂肪酶及其催化合成生物柴油[J].现代化工,2007,21(11):40-42
[8]吴茜茜,吴克,刘斌,等.壳聚糖固定化德氏根霉脂肪酶的研究[J].工业微生物,2003,33(4):9-13
[9]López Serrano P,Cao L,van Rantwijk F,et al.Cross-linked enzyme aggregates with enhanced activity:application to lipases[J].Biotechnology Letters,2002,24(16):1379,1383
[10]Tsai S W,Shaw S S.Selection of hydrophobic membranes in the lipase-catalyzed hydrolysis of olive oil[J].Journal of Membrane Science,1998,146(1):l-8
[11]Sakaki K,Giorno L,Drioli E.Lipase-catalyzed optical resolution of racemic naproxen in biphasic enzyme membrane reactors[J].Journal of Membrane Science,2001,184(1):27-38
[12]Vaidya B K,Ingavle G C,Ponrathnam S,et al.Immobilization of Candida rugosa lipase on poly(allyl glycidyl ether-co-ethylene glycol dimethacrylate)macroporous polymer particles[J].Bioresource Technology,2008,99(9):3623-3629
[13]王华,王安明,周成,等.微波辐射高效共价固定青霉素酰化酶[J].化学学报,2008,66(2):271-275
[14]中华人民共和国国家质量监督检验检疫总局,中国国家标准化管理委员会.GB/T 23535-2009脂肪酶制剂[S].北京:中国标准出版社,2009
[15]波莱纳J,麦凯布A.P.工业酶——结构、功能与应用[M].北京:科学出版社,2010
[16]徐坚,王玉军,骆广生,等.膜材料的亲疏水性对固定化脂肪酶的影响[J].高校化学工程学报,2006,20(3):395-400
[17]万容兵.载体表面化学组成对蛋白质吸附行为的影响[D].浙江理工大学,2010
[18]刘汝宽.脂肪酶固定化技术研究[D].郑州:河南工业大学,2007
[19]徐向宏,何明珠.试验设计与Design-Expert、SPSS应用[M].北京:科学出版社,2010
[20]贾佳,尹德忠,张宝亮,等.大孔树脂固定木瓜蛋白酶的研究[J].材料导报,2013,27(22):88-91
[21]曹林秋.载体固定化酶-原理、应用和设计[M].北京:化学工业出版社,2008
[22]罗贵明.酶工程[M].北京:化学工业出版社,2003