广西师范大学学报（自然科学版） ›› 2018, Vol. 36 ›› Issue (4): 108-118.doi: 10.16088/j.issn.1001-6600.2018.04.014

• • 上一篇    下一篇

使用国产环氧树脂LXEP-120固定化脂肪酶研究

徐珊1, 李任强1, 张继福2, 张云3,4, 孙爱君3,4, 胡云峰3,4*   

  1. 1.暨南大学生物工程学系,广东广州510632;
    2.广东省中医院,广东广州510120;
    3. 中国科学院南海海洋研究所中国科学院热带海洋生物资源与生态重点实验室,广东广州510301;
    4.中国科学院南海海洋研究所广东省海洋药物重点实验室,广东广州510301
  • 收稿日期:2018-05-08 发布日期:2018-10-20
  • 通讯作者: 胡云峰(1980—),男,山东日照人,中国科学院南海海洋研究所研究员,博士。E-mail:yunfeng.hu@scsio.ac.cn
  • 基金资助:
    中国科学院战略性先导科技专项(XDA11030404);广东省海洋渔业科技攻关与研发方向项目(A201701 C12);“科学”号高端用户项目(KEXUE2018G05);国家自然科学基金(21302199);广州市科技计划项目(201510010012)

Immobilization of Lipase Using Domestic Epoxy Resin LXEP-120

XU Shan1, LI Renqiang1, ZHANG Jifu2, ZHANG Yun3,4, SUN Aijun3,4, HU Yunfeng3,4*   

  1. 1. Department of Biotechnology, Jinan University, Guangzhou Guangdong 510632, China;
    2. Guangdong Provincial Hospital of Chinese Medicine, Guangzhou Guangdong 510120, China;
    3. CAS Key Laboratory of Tropical Marine Bio-resources and Ecology, South China SeaInstitute of Oceanology, Chinese Academy of Sciences, Guangzhou Guangdong 510301,China;
    4. Guangdong Key Laboratory of Marine Materia Medica, South China Sea Institute ofOceanology, Chinese Academy of Sciences, Guangzhou Guangdong 510301, China
  • Received:2018-05-08 Published:2018-10-20

PDF (PC)

484

摘要： 使用国产环氧树脂LXEP-120对猪胰脂肪酶进行固定化,研究其固定化工艺的最佳条件,并且对固定化酶和游离脂肪酶的酶学性质作初步的比较。结果显示,最佳固定化条件为:选用pH6.0、1.0 mol/L的磷酸钾缓冲液,载体投放量5.0 g,固定化温度22 ℃,固定化时间12 h。最终得到的固定化酶酶活400 U/g,酶活回收率92.79%,固定化酶重复使用10次后仍保持70%左右的酶活。与游离酶相比,固定化酶的最适反应pH(pH8.0)没有变化,最适反应温度(45 ℃)提高5 ℃,pH耐受性、热稳定性和重复使用性显著增强。同时,固定化酶具有较好的储存稳定性。研究结果证明国产环氧树脂LXEP-120在工业酶的固定化应用上具有很好的应用潜力。

关键词: 国产环氧树脂LXEP-120, 脂肪酶, 固定化, 酶学性质

Abstract: The immobilization of lipase from procine pancrestic was performed by using domestic epoxy resin LXEP-120. The immobilization conditions were optimized, and the enzymatic properties of immobilized lipase and free lipase were also compared. The optimal immobilization conditions were as follows: using 1.0 mol/L pH6.0 potassium phosphate buffer, the quantity of epoxy resin being 5.0 g, the appropriate temperature being 22 ℃, and the immobilized time being 12 h. Under above optimized conditions, the activity of immobilized lipase was about 400 U/g, and the recovery rate of the enzymatic activity of the immobilized lipase reached 92.79%. The immobilized lipase retained 70% of its initial enzymatic activity after repeated usage for 10 times. The optimal reaction pH (pH 8.0) was the same as that of free lipase, and the optimal reaction temperature(45 ℃) was 5 ℃ higher than that of free lipase. The thermal-stability and operating-stability reuse rate of immobilized lipase were significantly better than those of free lipase. Additionally, immobilized lipase exhibited very good storage stability. The study shows that domestic epoxy resins LXEP-120 possesses great potential in the immobilization of industrial enzymes.

Key words: domestic epoxy resin LXEP-120, lipase, immobilization, enzymatic property

中图分类号: 

  • Q814
[1] 谢雪凤. 嗜热子囊菌产过氧化氢酶的生产、固定化及应用研究[D]. 杭州:浙江工业大学, 2009.
[2] KUMARI A, GUPTA R. Functional characterization of a novel aspartic acid rich lipase, talipc, from trichosporon asahii msr54: solvent-dependent enantio inversion during esterification of 1-phenylethanol[J]. Biotechnol Lett, 2015, 37(1): 121-130.
[3] CAO Y, ZHUANG Y, YAO C, et al. Purification and characterization of an organic solvent-stable lipase from pseudomonas stutzeri lc2-8 and its application for efficient resolution of (r, s)-1-phenylethanol[J]. Biochemical Engineering Journal, 2012, 64: 55-60.
[4] ROMDHANE I B, ROMDHANE Z B, BOUZID M, et al. Application of a chitosan-immobilized talaromyces thermophilus lipase to a batch biodiesel production from waste frying oils[J]. Appl Biochem Biotechnol, 2013, 171(8): 1986-2002.
[5] JAMIE A, ALSHAMI A S, MALIABARI Z O, et al. Immobilization and enhanced catalytic activity of lipase on modified MWCNT for oily wastewater treatment[J]. Environmental Progress and Sustainable Energy, 2016, 35(5): 1441-1449.
[6] HA S H, LAN M N, LEE S H, et al. Lipase-catalyzed biodiesel production from soybean oil in ionic liquids[J]. Enzyme and Microbial Technology, 2007, 41(4): 480-483.
[7] VAHEDI H. Evaluating the effects of different levels of lipase enzyme on the quality of baguette bread[J]. Acta Medica Mediterranea, 2015, 31: 1359-1363.
[8] LAUPRASERT P, CHANSIRIRATTANA J,PAENGJAN J. Effect of selected bacteria as bioremediation on the degradation of fats oils and greases in wastewater from cafeteria grease traps[J]. European Journal of Sustainable Development, 2017, 6(2): 180-185.
[9] 曾淑华, 杨江科, 闫云军. 固定化脂肪酶性质及其应用研究[J]. 生物加工过程, 2007, 5(1): 45-49.
[10] KIM H, CHOI N, OH S W, et al. Synthesis of alpha-linolenic acid-rich triacylglycerol using a newly prepared immobilized lipase[J]. Food Chemistry, 2017, 237: 654-658.
[11] MENDES A A, DE CASTRO H F, RODRIGUES D D, et al. Multipoint covalent immobilization of lipase on chitosan hybrid hydrogels: influence of the polyelectrolyte complex type and chemical modification on the catalytic properties of the biocatalysts[J]. Journal of Industrial Microbiology and Biotechnology, 2011, 38(8): 1055-1066.
[12] LEI M, HU D, YANG H, et al. Preparation and characterization of hollow magnetic composite nanoparticles for immobilized pectinase[J]. Surface and Coatings Technology, 2015, 271: 2-7.
[13] NUNES M A P, VILA-REAL H, FERNANDES P C B, et al. Immobilization of naringinase in pva-alginate matrix using an innovative technique[J]. Applied Biochemistry and Biotechnology, 2010, 160(7): 2129-2147.
[14] LOPEZ-GALLEGO F, BETANCOR L, MATEO C, et al. Enzyme stabilization by glutaraldehyde crosslinking of adsorbed proteins on aminated supports[J]. Journal of Biotechnology, 2005, 119(1): 70-75.
[15] TORRES P, BATISTA-VIERA F. Immobilization of β-galactosidase from bacillus circulans onto epoxy-activated acrylic supports[J]. Journal of Molecular Catalysis B: Enzymatic, 2012, 74(3/4): 230-235.
[16] 刘文涛, 段洪东, 王兴建, 等. 环氧基固定化酶载体的研究进展[J]. 山东轻工业学院学报, 2012, 26(3): 40-44.
[17] AGHABABAIE M, BEHESHTI M, RAZMJOU A, et al. Covalent immobilization of candida rugosa lipase on a novel functionalized Fe3O4@SiO2 dip-coated nanocomposite membrane[J]. Food and Bioproducts Processing, 2016, 100: 351-360.
[18] XIANG X R, SUO H B, XU C, et al. Covalent immobilization of lipase onto chitosan-mesoporous silica hybrid nanomaterials by carboxyl functionalized ionic liquids as the coupling agent[J]. Colloids and Surfaces B, Biointerface, 2018, 165: 262-269.
[19] ZHAO J Q, WANG Y J, LUO G S, et al. Covalent immobilization of penicillin g acylase on aminopropyl-functionalized mesostructured cellular foams[J]. Bioresour Technol, 2010, 101(19): 7211-7217.
[20] 顾恺, 邹树平, 王志才, 等. 环氧树脂固定化卤醇脱卤酶的研究[J]. 现代化工, 2016, 36(11): 69-74.
[21] EPHRAIM K K, KRAEMER D M. Eupergit (r) c, a carrier for immobilization of enzymes of industrial potential[J]. Journal of Molecular Catalysis B-Enzymatic, 2000, 10(1-3): 157-176.
[22] 刘向前, 李道明, 王卫飞, 等. Lipozyme~CALB L固定化及催化合成EPA/DHA甘油酯的研究[J]. 中国油脂, 2016(11): 21-26.
[23] GONZALEZ-CORONEL L A, COBAS M, Rostro-Alanis M D, et al. Immobilization of laccase of pycnoporus sanguineus cs43[J]. New Biotechnology, 2017, 39: 141-149.
[24] KNEZEVIC-JUGOVIC Z D, GRBAVCIC S Z, JOVANOVIC J R, et al. Covalent immobilization of enzymes on Eupergit() supports: effect of the immobilization protocol[J]. Methods Mol Biol, 2017, 1504: 75-91.
[25] WANG J, LI W, NIU D D, et al. Improved synthesis of isomaltooligosaccharides using immobilized alpha-glucosidase in organic-aqueous media[J]. Food Science and Biotechnology, 2017, 26(3): 731-738.
[26] 侯爱军,徐冰斌,梁亮,等.改进铜皂-分光光度法测定脂肪酶活力[J].皮革科学与工程,2011,21(1):22-27.
[27] 嵇东情, 郑仁朝, 郑裕国. 环氧树脂固定化拟南芥腈水解酶NIT的研究[J]. 精细与专用化学品, 2015, 23(5): 12-16.
[28] 崔培梧, 黎继烈, 肖作为, 等. 环氧基树脂固定化青霉菌柚苷酶的工艺条件[J]. 食品与发酵工业, 2014, 40(5): 87-92.
[29] 侯丽云. 脂肪酶固定化及其在催化合成乙酸香茅酯中的应用[D]. 扬州: 扬州大学, 2013: 11.
[30] 刘自琴. 脂肪酶和胰蛋白酶的固定化及共固定化研究[D]. 广州: 华南理工大学, 2012: 4.
[31] MATEO C, GRAZú V, PESSELA B C, et al. Advances in the design of new epoxy supports for enzyme immobilization-stabilization[J]. Biochemical Society Transactions, 2007, 35(6):1593-1601.
[32] 雷生姣, 王可兴, 吕晓燕, 等. 聚乙烯醇-海藻酸钙固定化柚苷酶[J]. 食品科学,2011,32(3): 138-143.
[33] FERNANDES A I, GREGORIADIS G. Synthesic, characterization and properties of sialylated catalase[J]. Biochimica et Biophysica Acta (BBA) -Protein Structure and Molecular Enzymology, 1996, 1293(1): 90-96.
[34] AMIN H A, SECUNDO F, AMER H, et al. Improvement of aspergillus flavus saponin hydrolase thermal stability and productivity via immobilization on a novel carrier based on sugarcane bagasse[J]. Biotechnol Rep (Amst), 2018, 17: 55-62.
[35] DILEK A, BARIS B, DENIZ Y, et al. Improving stability of formate dehydrogenase from Candida methylica by immobilization onto Eupergit C 250 L[J]. Applied Biochemistry and Microbiology, 2016(1): 37-42.
[36] DRISS D, DRISS Z, CHAARI F, et al. Immobilized his-tagged recombinant xylanase from penicillium occitanis on nickel-chelate Eupergit C[J]. Bioengineered, 2014, 5(4): 274-279.
[1] 林海蛟, 张继福, 张云, 胡云峰. 基于大孔吸附树脂先交联后吸附法固定化脂肪酶[J]. 广西师范大学学报（自然科学版）, 2020, 38(4): 100-108.
[2] 钟成华, 张文东, 刘鹏, 陈建. 包埋固定化复合菌低温下处理养猪废水研究[J]. 广西师范大学学报（自然科学版）, 2011, 29(3): 52-56.
Viewed
Full text


Abstract

Cited
  Shared   
  Discussed   
No Suggested Reading articles found!
版权所有 © 广西师范大学学报（自然科学版）编辑部
地址：广西桂林市三里店育才路15号　邮编：541004
电话：0773-5857325　E-mail: gxsdzkb@mailbox.gxnu.edu.cn
本系统由北京玛格泰克科技发展有限公司设计开发