|本期目录/Table of Contents|

[1]马抒晗,张玮佳,吴茜,等.高产脂肪酶菌株的筛选鉴定及酶学、转酯特性[J].应用与环境生物学报,2014,20(04):602-608.[doi:10.3724/SP.J.1145.2014.01028]
　MA Shuhan,ZHANG Weijia,WU Qian,et al.Lipase-producing strain: isolation, identification, enzymology and transesterification[J].Chinese Journal of Applied & Environmental Biology,2014,20(04):602-608.[doi:10.3724/SP.J.1145.2014.01028]
点击复制

高产脂肪酶菌株的筛选鉴定及酶学、转酯特性()
分享到：

《应用与环境生物学报》[ISSN:1006-687X/CN:51-1482/Q]

卷:
20卷
期数:
2014年04期
页码:
602-608
栏目:
研究论文
出版日期:
2014-08-25

文章信息/Info

Title:
Lipase-producing strain: isolation, identification, enzymology and transesterification
作者:
马抒晗;  张玮佳;  吴茜;  姜腾飞;  乔代蓉;  曹毅
四川大学生命科学学院微生物与代谢工程四川省重点实验室 成都 610065
Author(s):
MA Shuhan;  ZHANG Weijia;  WU Qian;  JIANG Tengfei;  QIAO Dairong;  CAO Yi
Microbiology and Metabolic Engineering Key Laboratory of Sichuan Province, College of Life Sciences, Sichuan University, Chengdu 610065, China
关键词:
脂肪酶; 菌株筛选与鉴定; 酶学性质; 转酯; 伯克霍尔德氏菌; 生物柴油
Keywords:
lipase;  strain isolation and identification;  enzymology;  transesterification;  Burkholderia;  biodiesel
分类号:
Q936 : Q556
DOI:
10.3724/SP.J.1145.2014.01028
文献标志码:
A
摘要:
从自然环境中筛选水解酶活高且酶学、转酯特性优良的产脂肪酶菌株，对脂肪酶工业化发酵生产及生物柴油制备的研究具有重要意义. 采用罗丹明B平板初筛和摇瓶发酵复筛法，从70份含油脂丰富的样品中筛选产脂肪酶酶活较高的菌株进行16S rRNA鉴定，研究其酶学性质；用大孔树脂固定酶，在无溶剂体系中催化橄榄油制备生物柴油，研究其转酯特性. 结果筛选到一株高产脂肪酶的菌株WZ10-3，通过p-NPP法测得其初始酶活为78.68 U/mL，经16S rRNA鉴定属于伯克霍尔德氏菌（Burkholderia sp.），与B. stabilis同源性达到99%. 该菌在发酵48 h时达到产酶高峰，所产脂肪酶的最适作用温度为50 ℃，最适作用pH为7.0，70 ℃下的半衰期可达1 h，pH为7-9时稳定性良好. 以大孔树脂NKA-9和HPD600为载体制备的2种固定化脂肪酶，催化橄榄油生产生物柴油的转酯率均可达到97%. 综合表明，菌株WZ10-3脂肪酶的初始水解酶活高于大多数野生脂肪酶，热稳定性好且转酯特性优良，有很好的后续研究价值. 图7 表3 参25
Abstract:
For a better understanding of lipase industrial fermentation and biodiesel production, this study screened from natural environment lipase producing strains with high hydrolysis activity, stable enzymatic properties and excellent transesterification characteristics. The high lipase activity producing strain was isolated from 70 rich oil samples by Rhodamine B screening plates and shake flask fermentation, and was identified through 16S rRNA sequence. After studying of enzymatic properties, the lipase was immobilized on the macroporous resins NKA-9 and HPD600. The transesterification characteristics were studied by producing biodiesel from olive oil catalyzed by immobilized lipases in solvent-free system. The high lipase activity producing strain WZ10-3 was measured by p-NPP method to have an initial enzyme activity of 78.68 U/mL. The 16SrRNA sequence identified the strain as Burkholderia sp., of 99% homology with B. stabilis. Its lipase production reached a peak at 48 h of fermentation with the optimum reaction temperature of 50 oC and the optimum pH of 7.0. The lipase had a half-life of 1 h under 70 oC, and was stable under pH 7-9. The preparing biodiesel by lipases immobilized on the macroporous resins NKA-9 and HPD600 indicated that both immobilized lipases had 97% transesterification rate. The initial lipase activity of strain WZ10-3 is much higher than most other wild strains. Its good thermal stability and excellent transesterification characteristics prove the follow-up research value of the strain WZ10-3.

参考文献/References:

1 Cho SS, Park DJ, Simkhada JR, Hong JH, Sohng JK, Lee OH, Yoo JC. A neutral lipase applicable in biodiesel production from a newly isolated Streptomyces sp. CS326 [J]. Bioprocess Biosyst Eng, 2012, 35 (1-2): 227-234
2 Tan T, Lu J, Nie K, Deng L, Wang F. Biodiesel production with immobilized lipase: a review [J]. Biotechnol Adv, 2010, 28 (5): 628-634
3 Defranceschi O, Anne C, Farion W, Felipe M, Coelho V, José V, Fernandes R, Maria L, Bellin M, André. Production of methyl oleate with a lipase from an endophytic yeast isolated from castor leaves [J]. Biocatal Agric Biotechnol, 2012, 1 (4): 295-300
4 Dheeman DS, Frias JM, Henehan G.T. Influence of cultivation conditions on the production of a thermostable extracellular lipase from Amycolatopsis mediterranei DSM 43304 [J]. J Ind Microbiol Biotechnol, 2010, 37 (1): 1-17
5 Kim BS, Hou CT. Production of lipase by high cell density fed-batch culture of Candida cylindracea [J]. Bioprocess Biosyst Eng, 2006, 29 (1): 59-64
6 A??kel ?, Er?an M, Sa? AY. Optimization of critical medium components using response surface methodology for lipase production by Rhizopus delemar [J]. Food Bioprod Process, 2010, 88 (1): 31-39
7 Ruchi G, Anshu G, Khare SK. Lipase from solvent tolerant Pseudomonas aeruginosa strain: production optimization by response surface methodology and application [J]. Bioresour Technol, 2008, 99 (11): 4796-4802
8 Kawakami K, Ueno M, Takei T, Oda Y, Ryo T. Application of a Burkholderia cepacia lipase-immobilized silica monolith micro-bioreactor to continuous-flow kinetic resolution for transesterification of (R, S)-1-phenylethanol [J]. Process Biochem, 2012, 47 (1): 147-150
9 Winkler UK. Glycogen hyaluronate and some other polysaccharides greatly enhance the formation of exolipase by Serratia marcescens [J]. Bacteriol, 1979, 138: 663-701
10 Lucas T, Francois J, Bohuon P. Factors influencing mass transfer during immersion cold storage of apples in NaCl/sucrose solutions [J]. Lebensm-Wissu-Technol, 1999, 32: 327-332
11 王艳华, 刘飞飞, 程仕伟. Bacillus subtilis CICC 20034产脂肪酶的培养条件研究[J]. 食品与药品, 2012, 14 (2): 84-87[Wang YH, Liu FF, Cheng SW. Study on culture condition for lipase-producing Bacillus subtilis CICC 20034 [J]. Food Drug, 2012, 14 (2): 84-87]
12 Liu CH, Huang CC, Wang YW, Lee DJ, Chang JS. Biodiesel production by enzymatic transesterification catalyzed by Burkholderia lipase immobilized on hydrophobic magnetic particles [J]. Appl Energy, 2012, 100: 41-46
13 Wyatt VT, Haas MJ. Production of fatty acid methyl esters via the in situ transesterification of soybean oil in carbon dioxide-expanded ethanol [J]. J Am Oil Chem Soc, 2009, 86 (10): 1009-1016
14 Dandavate V, Jinjala J, Keharia H, Madamwar D. Production, partial purification and characterization of organic solvent tolerant lipase from Burkholderia multivorans V2 and its application for ester synthesis [J]. Bioresour Technol, 2009, 100 (13): 3374-3381
15 Uttatree S, Winayanuwattikun P, Charoenpanich J. Isolation and characterization of a novel thermophilic-organic solvent stable lipase from Acinetobacter baylyi [J]. Appl Biochem Biotechnol, 2010, 162 (5): 1362-1376
16 夏宇, 周文, 邓学良, 伍金娥. 脂肪酶高产菌株的筛选及产酶条件优化[J]. 中南林业科技大学学报, 2013, 33 (9): 116-120 [Xia Y, Zhou WH, Deng XL, Wu JE. Isolation of high lipase-producing strains and optimization of lipase producing conditions [J]. J Central South Univ For Technol, 2013, 33 (9):116-120]
17 Sorokin DY, Jones BE. Improved method for direct screening of true lipase-producing microorganisms with particular emphasis on alkaline conditions [J]. Microbiology, 2009, 78 (1): 125-130
18 Shu Z, Lin R, Jiang H, Zhang Y, Wang M, Huang J. A rapid and efficient method for directed screening of lipase-producing Burkholderia cepacia complex strains with organic solvent tolerance from rhizosphere [J]. J Biosci Bioeng, 2009, 107 (6): 658-661
19 Mahenihiralingam E, Urban TA, Goldberg JB. The multifarious, multireplieon Burkholderia cepacia complex [J]. Nat Rev Microbiol, 2005, 3 (2): 144-156
20 陈贵元, 魏云林. 低温脂肪酶的研究现状与应用前景[J]. 生物技术通报, 2006, 12: 29-32 [Chen GY, Wei YL. The research status and application prospect of cold-adapted lipases [J]. Biotechnol Bull, 2006, 12: 29-32]
21 薛静, 陶树兴, 田泽英, 苏蕊, 丛寅. 脂肪酶产生菌的筛选、产酶条件及酶特性研究[J]. 安徽农业科学, 2011, 39 (15): 8826-8830 [Xue J, Tao SX, Tian ZY, Su R, Cong Y. Screen of lipase production strains and study on condition for lipase production and properties of lipase [J]. J Anhui Agric.Sci, 2011, 39 (15) : 8826-8830]
22 蔡少丽, 邹有土, 黄建忠, 林琳. 定点突变对扩展青霉脂肪酶热稳定性的改善[J]. 应用与环境生物学报, 2013, 19 (1): 43-47 [Cai SL, Zou YT, Huang JZ, Lin L. Improvement of thermostability of Penicillium expansum lipase by site-directed mutagenesis [J]. Chin J Appl Environ Biol, 2013, 19 (1): 43-47]
23 Horchani H, Cha?bouni M, Gargouri Y, Sayari A. Solvent-free lipase-catalyzed synthesis of long-chain starch esters using microwave heating: Optimization by response surface methodology [J]. Carbohyd Polym, 2010, 79 (2): 466-474
24 Baron AM, Barouh N, Barea B, Villeneuve P, Mitchell DA, Krieger N. Transesterification of castor oil in a solvent-free medium using the lipase from Burkholderia cepacia LTEB11 immobilized on a hydrophobic support [J]. Fuel, 2014, 117: 458-462
25 张学林, 唐湘华, 李俊俊, 宋拓, 慕跃林, 许波, 杨云娟, 黄遵锡. 低温脂肪酶酶促酯交换制备生物柴油[J]. 中国油脂, 2013, 38 (2): 66-68 [Zhang XL, Tang XH, Li JH, Song T, Mu YL, Xu B, Yang YJ, Huang ZX. Biodiesel production by low temperature lipase catalysed transesterification [J]. China Oils Fats, 2013, 38 (2): 66-68]

相似文献/References:

[1]张晓梅,窦文芳,许泓瑜,等.一株产脂肪酶嗜盐菌的鉴定及耐盐机理[J].应用与环境生物学报,2010,16(01):100.[doi:10.3724/SP.J.1145.2010.00100]
　ZHANG Xiaomei,DOU Wenfang,XU Hongyu,et al.Identification of a Lipase Producing Halophilic Strain and Its Halophilic Mechanism[J].Chinese Journal of Applied & Environmental Biology,2010,16(04):100.[doi:10.3724/SP.J.1145.2010.00100]
[2]张银波,江木兰,万霞,等.环境基因组DNA脂肪酶基因克隆与分析[J].应用与环境生物学报,2010,16(02):269.[doi:10.3724/SP.J.1145.2010.00269]
　ZHANG Yinbo,JIANG Mulan,WAN Xia,et al.Cloning and Analysis of a Lipase Gene from Environmental Genome DNA[J].Chinese Journal of Applied & Environmental Biology,2010,16(04):269.[doi:10.3724/SP.J.1145.2010.00269]
[3]吴尧,万霞,张银波,等.白地霉（lip42）脂肪酶的纯化及酶学性质[J].应用与环境生物学报,2010,16(05):710.[doi:10.3724/SP.J.1145.2010.00710]
　WU Yao,WAN Xia,ZHANG Yingbo,et al.Purification and Properties of a Lipase from Geotrichum candidum lip42[J].Chinese Journal of Applied & Environmental Biology,2010,16(04):710.[doi:10.3724/SP.J.1145.2010.00710]
[4]蔡少丽,邹有土,黄建忠,等.定点突变对扩展青霉脂肪酶热稳定性的改善[J].应用与环境生物学报,2013,19(01):43.[doi:10.3724/SP.J.1145.2013.00043]
　CAI Shaoli,ZOU Youtu,HUANG Jianzhong,et al.Improvement of Thermostability of Penicillium expansum Lipase by Site-directed Mutagenesis[J].Chinese Journal of Applied & Environmental Biology,2013,19(04):43.[doi:10.3724/SP.J.1145.2013.00043]
[5]付玮,李友国,李一星.基于黄单胞菌冰核蛋白N端的表面展示体系建立[J].应用与环境生物学报,2014,20(03):351.[doi:10.3724/SP.J.1145.2014.10001]
　FU Wei,LI Youguo,LI Yixing.Establishment of cell surface display system based on N-domain of ice nucleation protein of Xanthomonas[J].Chinese Journal of Applied & Environmental Biology,2014,20(04):351.[doi:10.3724/SP.J.1145.2014.10001]
[6]嵇少泽 勾长龙 张喜庆 王羽 田佳琪 李晨 刘继军 高云航**.病死猪堆肥高效降解复合菌系的构建及应用效果评价*[J].应用与环境生物学报,2020,26(03):1.[doi:10.19675/j.cnki.1006-687x.2019.07007]
　JI Shaoze,GOU Changlong,ZHANG Xiqing,et al.Construction and application of a high efficient complex microbial system to degrade dead-pig carcass in compost and assessment of its effect*[J].Chinese Journal of Applied & Environmental Biology,2020,26(04):1.[doi:10.19675/j.cnki.1006-687x.2019.07007]

备注/Memo

备注/Memo:
“十二五”国家科技支撑计划项目（2011BAD14B05、2013BAD10B01、2014BAD02B02）、国家自然科学基金项目（31171447）和四川省科技厅项目（2014GZX0005、2013HH0007、2013GZX0161-3、2013GZ0058、2013JCPT003）资助 Supported by the Sci-tech Pillar Project of the Twelfth Five-year Plan of China (2011BAD14B05, 2013BAD10B01, 2014BAD02B02), the National Natural Science Foundation of China (31171447) , and the Project of the Science and Technology Department of Sichuan Province (2014GZX0005, 2013HH0007, 2013GZX0161-3, 2013GZ0058, 2013JCPT003)
更新日期/Last Update: 2014-08-26