固载化木聚糖酶交联酶聚集体的制备及性质
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摘要
目的提高木聚糖酶YS1069的稳定性、重复利用率以及便于从反应体系中分离。方法本文将交联酶聚集体CLEAs技术与载体固定化技术相结合,用LKZ-128氨基型树脂对木聚糖酶CLEAs进行固定化,并对其性质进行初步研究。结果及结论采用单因素法和响应面法对影响固定化酶活性的因素进行分析和优化,获得最佳固定化条件为:pH 9.0的酶液,以2mL的异丙醇为沉淀剂,沉淀1h,以1.59g/L的京尼平为交联剂,交联2h,20℃的环境中固定化9.25h,加酶量为8mg时回收率最高,达到65.63%,加酶量为60mg时酶活性最高,达到190U/g。重复使用10次后,其相对酶活仍为42.3%,说明具有良好的批次操作稳定性。该酶经过固定化后热稳定性和pH稳定性均得到了提高。
Objective To enhance the stability,repeated utilization of Xylanase and easily separate from the reaction system.Methods In this study,the cross-linked enzyme aggregates(CLEAs)technology was combined with carrier immobilization technique.Using amino resin LKZ-128,Xylanase CLEAs were immobolized and its properties were studied.Results and ConclusionImmobilized conditions of Xylanase CLEAs were studied by using single factor experiment and the response surface design.The study indicated that the best immobilized conditions were:pH 9.0enzyme liquid precipitated by 2mL isopropyl,precipitation 1h,and cross-linked 2hwith the carrier LKZ-128 by 1.59g/L genipin,immobilized 9.25 hin the environment of 20℃.The recovery rate was up to 65.63% when the enzyme dosage was 8mg,and activity of enzyme was up to 190U/g when the enzyme dosage was 60 mg.After repeated 10 times,the relative activity of the enzyme was still 42.3%,showing good reusability.Thermal stability and pH stability of the enzyme were improved after immobilization.
Objective To enhance the stability,repeated utilization of Xylanase and easily separate from the reaction system.Methods In this study,the cross-linked enzyme aggregates(CLEAs)technology was combined with carrier immobilization technique.Using amino resin LKZ-128,Xylanase CLEAs were immobolized and its properties were studied.Results and ConclusionImmobilized conditions of Xylanase CLEAs were studied by using single factor experiment and the response surface design.The study indicated that the best immobilized conditions were:pH 9.0enzyme liquid precipitated by 2mL isopropyl,precipitation 1h,and cross-linked 2hwith the carrier LKZ-128 by 1.59g/L genipin,immobilized 9.25 hin the environment of 20℃.The recovery rate was up to 65.63% when the enzyme dosage was 8mg,and activity of enzyme was up to 190U/g when the enzyme dosage was 60 mg.After repeated 10 times,the relative activity of the enzyme was still 42.3%,showing good reusability.Thermal stability and pH stability of the enzyme were improved after immobilization.
引文
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[15]任延刚,朱启忠,王娜,等.壳聚糖/海藻酸钠固定木聚糖酶的研究[J].安徽农业科学,2008,36(35):15298-15301.
[16]京尼平交联壳聚糖磁性微球的制备及脂肪酶的固定化[J].应用化学,2013,30(8):922-926.
[17]艾志录,江正强,李里特.大孔树脂D380固定化橄榄绿链霉菌E-86来源木聚糖酶的研究[J].食品与发酵工业,2004,30(2):10-14.
[18]崔建涛.树脂固定化木聚糖酶及其酶解动力学研究[D],郑州:河南工业大学,2007.
[2]吴学敏,宣振华,刘毅.低聚木糖的研究进展[J].贵阳中医学院学报,2013,35(3):25-28.
[3]王伟,王飞,胡国华.稻草低聚木糖的制备及其抗氧化活性研究[J].中国食品添加剂,2012,6:78-82.
[4]朱路甲,贾克军,王宝军.我国功能糖产业的现状与发展前景[J].中国食品添加剂,2010,1:52-56.
[5]武秋颖,陈复生,时冬梅,等.酶法制备低聚木糖研究进展[J].粮油加工,2010,5:118-121.
[6]Cao L,van Langen L,Sheldon R A.Immobilised enzymes:carrier-bound or carrier-free[J].Curr Opin Biotechnol,2003,14(4):387-394.
[7]Cao L,Van Langen L M,Van Rantwijk F.Cross-linked aggregates of penicillin acylase:robust catalysts for the synthesis ofβ-lactam antibiotics[J].J Molecular Catalysis B:Enzymatic,2001,11(4/6):665-670.
[8]李小冬,吴嘉,贾东晨,等.固定化酶的研究方法概述[J].中国酿造,2011,11:5-9.
[9]孙丰慧,李安明.耐碱性木聚糖酶产生菌的筛选及发酵条件研究[J].应用与环境生物学报,2008,14(03):436-439.
[10]王镜岩.生物化学[M].北京:高等教育出版社,2002:88-90.
[11]Cao L,van Langen L,Sheldon R A.Effect of crosslinked condition on characteristics of chitosan/tripolyphosphate/genipin beads and their application in the selective adsorption of phytic acid from[J].Carbohydr Polym,2011,14(4):387-394.
[12]Chiono V,Pulieri E,Vozzi G,et al.Genipin-crosslinked chitosan/gelatin blends for biomedical applications[J].J Mater Sci Mater Med,2008,19(2):889-898.
[13]李笃信,贾德民,周萍.聚苯乙烯阴离子交换树脂吸附交联法固定α-淀粉酶[J].华南理工大学学报(自然科学版),1998,26(8):63-67.
[14]Sarbu A,de Pinho M N,Freixo Md R,et al.New method for the covalent immobilization of a xylanase by radical grafting of acrylamide on cellulose acetate membranes[J].Enzyme Microb Technol,2006,36(39):125-130.
[15]任延刚,朱启忠,王娜,等.壳聚糖/海藻酸钠固定木聚糖酶的研究[J].安徽农业科学,2008,36(35):15298-15301.
[16]京尼平交联壳聚糖磁性微球的制备及脂肪酶的固定化[J].应用化学,2013,30(8):922-926.
[17]艾志录,江正强,李里特.大孔树脂D380固定化橄榄绿链霉菌E-86来源木聚糖酶的研究[J].食品与发酵工业,2004,30(2):10-14.
[18]崔建涛.树脂固定化木聚糖酶及其酶解动力学研究[D],郑州:河南工业大学,2007.