克里布所类芽孢杆菌6hRe76产木聚糖酶条件的优化
|
摘要
为了提高木聚糖酶产量,采用单因素试验和响应面分析法,对木聚糖酶产生菌克里布所类芽孢杆菌6hRe76产木聚糖酶的发酵培养基和发酵条件进行优化。利用单因素试验确定最佳碳源和最佳氮源分别为麸皮、酵母膏。然后利用PB试验筛选出影响木聚糖酶产量的主要因素为装液量、麸皮、K_2HPO_4,进一步利用Box-Behnken软件进行试验设计,并对试验数据处理得到响应模型。预测得到菌株6hRe76的最佳培养条件为:麸皮5.92%,K_2HPO_40.71%,酵母膏0.75%,MgSO_40.05%,CaCl_20.05%,吐温80 0.15%,装液量11mL/50mL三角瓶,接种量10%,种龄24h,34℃,初始pH9.0,培养时间36h。在最佳培养条件下,菌株6hRe76产酶量达92.11U/mL,比原始水平57.15U/mL提高了61.17%。
In order to obtain high yield of xylanase,the fermentation medium and fermentation conditions on xylanase production by Paenibacillus kribbensis 6 hRe76 were optimized by single-factor experiments and response surface methodology.The single-factor experiment was used to screen the best carbon source and nitrogen source.The results showed that wheat bran and yeast extract was as the optimum carbon source and nitrogen source,respectively.Wheat bran,K_2HPO_4 and liquid volume were certified as the major factors influencing xylanase production by Plackett-Burman experiment.Then,Box-Behnken was used in experimental design,experimental data were analyzed and the response surface mode constructed indicated that the optimal conditions for xylanase production by strain 6 hRe76 were as following:wheat bran 5.92%,K_2HPO_40.71%,yeast extract 0.75%,MgSO_40.05%,CaCl_20.04%,Tween80 0.15%,liquid volume 11/50 mL,inoculum 10%,seed age 24 h,34 ℃,initial pH9.0,cultured for 36 h.Under the optimal conditions,the maximum activity of xylanase was92.11 U/mL.The xylanase production of 6 hRe76 increased 61.17% than the original level of 57.15 U/mL.
In order to obtain high yield of xylanase,the fermentation medium and fermentation conditions on xylanase production by Paenibacillus kribbensis 6 hRe76 were optimized by single-factor experiments and response surface methodology.The single-factor experiment was used to screen the best carbon source and nitrogen source.The results showed that wheat bran and yeast extract was as the optimum carbon source and nitrogen source,respectively.Wheat bran,K_2HPO_4 and liquid volume were certified as the major factors influencing xylanase production by Plackett-Burman experiment.Then,Box-Behnken was used in experimental design,experimental data were analyzed and the response surface mode constructed indicated that the optimal conditions for xylanase production by strain 6 hRe76 were as following:wheat bran 5.92%,K_2HPO_40.71%,yeast extract 0.75%,MgSO_40.05%,CaCl_20.04%,Tween80 0.15%,liquid volume 11/50 mL,inoculum 10%,seed age 24 h,34 ℃,initial pH9.0,cultured for 36 h.Under the optimal conditions,the maximum activity of xylanase was92.11 U/mL.The xylanase production of 6 hRe76 increased 61.17% than the original level of 57.15 U/mL.
引文
[1]Saha B C.Hemicellulose bioconversion[J].Journal of Industrial Microbiology and Biotechnology,2003,30(5):279-291.
[2]Li X,Li E,Zhu Y,et al.A typical endo-xylanase fromStreptomyces rameus L2001and its unique characteristics in xylooligosaccharide production[J].Carbohydrate Research,2012,359(5):30-36.
[3]陈磊,赵祥颖,刘建军.木聚糖酶研究进展[J].山东食品发酵,2014,175:7-10.
[4]方微,单玉萍,李峰,等.木聚糖酶的作用机理及其在饲料中的应用[J].中国饲料,2011,21:21-24.
[5]陈阳雷,吉兴香,许凤.纸浆木聚糖酶生物助漂技术进展[J].中国造纸,2017,36(12):74-78.
[6]Lin C,Shen Z,and Qin W.Characterization of xylanase and cellulase produced by a newly isolated Aspergillus fumigatus N2and its efficient saccharification of barley straw[J].Appl Biochem Biotech.2017,82,559–569.
[7]Miao Y Z,Kong Y Q,Li P,et al.Efect of CBM1and linker region on enzymatic properties of a novel thermostable dimeric GH10xylanase(Xyn10A)from flamentous fungus Aspergillus fumigatus Z5[J].AMB Expr 2018(8):44-53.
[8]Deesukon W,Nishimura Y,Sakamoto T,et al.Purification,characterization of GH11endo-β-1,4-xylanase fromthermotolerant Streptomyces sp.SWU10and overexpression in Pichia pastoris KM71H[J].Molecular Biotechnology,2013,54(1):37-46.
[9]Oliveira J V,Borges T A,Corrêa R A,et al.Pseudozyma brasiliensis sp.nov,a xylanolytic,ustilaginomycetous yeast species isolated from an insect pest of sugarcane roots[J].International Journal of Systematic and Evolutionary Microbiology,2014,64(6):2159-2168.
[10]Basit A,Liu J,Miao T,et al.Characterization of two endo-b-1,4-xylanases from Myceliophthora thermophila and their saccharification efficiencies,synergistic with commercial cellulase[J].Frontiers in Microbiology,2018,9:1-11.
[11]朱辉,丁延芹,田方,等.多粘类芽孢杆菌SC2xynD和gluB的克隆及序列分析[J].生物技术通报,2008(4):171-174.
[12]Ghio S,Insani E,Pinninny F,et al.GH10XynA is the main xylanase identified in the crude enzymatic extractof Paenibacillus sp.A59when grown on xylan or lignocellulosic biomass[J].Microbiological Research,2016,186:16-26
[13]王中月,滕超,汤回花,等.产高分子量木聚糖酶细菌的筛选及产酶条件优化[J].食品与发酵工业,2017,43(10):88-95.
[14]马春玲,李思含,赵东芳,等.可产木聚糖酶的蕙兰根内生细菌的筛选与鉴定[J].河北大学学报,2017,37(5):518-522.
[15]董媛媛,李亚贤,沈艺红,等.响应面法优化木聚糖酶和α-葡萄糖醛酸酶联合水解桦木木聚糖工艺[J].食品科学,2018,39(4):125-131.
[16]卢海强,谷新晰,李晨,等.响应面法优化棒状乳杆菌HS4廉价增殖培养基研究[J].河北农业大学学报,2017,40(2):51-55.
[17]王晓宇,刘伟娜,谢响明,等.青霉L1来源具有生产木寡糖应用潜力的高比活GH11木聚糖酶[J].生物工程学报,2018,34(1):68-77.
[18]易旭东,潘虎,田云,等.一株碱性木聚糖酶产生菌的筛选及发酵条件的优化[J].中国农学通报2017,33(17):12-18.
[19]葛慧华,林锦霞,张光亚.以响应面法优化短小芽孢杆菌木聚糖酶产酶条件[J].华侨大学学报:自然科学版,2012(2):172-175.
[20]黄国勇,吴振强.枯草芽孢杆菌产木聚糖酶发酵条件的研究[J].河南工业大学学报:自然科学版,2006(1):32-35.
[2]Li X,Li E,Zhu Y,et al.A typical endo-xylanase fromStreptomyces rameus L2001and its unique characteristics in xylooligosaccharide production[J].Carbohydrate Research,2012,359(5):30-36.
[3]陈磊,赵祥颖,刘建军.木聚糖酶研究进展[J].山东食品发酵,2014,175:7-10.
[4]方微,单玉萍,李峰,等.木聚糖酶的作用机理及其在饲料中的应用[J].中国饲料,2011,21:21-24.
[5]陈阳雷,吉兴香,许凤.纸浆木聚糖酶生物助漂技术进展[J].中国造纸,2017,36(12):74-78.
[6]Lin C,Shen Z,and Qin W.Characterization of xylanase and cellulase produced by a newly isolated Aspergillus fumigatus N2and its efficient saccharification of barley straw[J].Appl Biochem Biotech.2017,82,559–569.
[7]Miao Y Z,Kong Y Q,Li P,et al.Efect of CBM1and linker region on enzymatic properties of a novel thermostable dimeric GH10xylanase(Xyn10A)from flamentous fungus Aspergillus fumigatus Z5[J].AMB Expr 2018(8):44-53.
[8]Deesukon W,Nishimura Y,Sakamoto T,et al.Purification,characterization of GH11endo-β-1,4-xylanase fromthermotolerant Streptomyces sp.SWU10and overexpression in Pichia pastoris KM71H[J].Molecular Biotechnology,2013,54(1):37-46.
[9]Oliveira J V,Borges T A,Corrêa R A,et al.Pseudozyma brasiliensis sp.nov,a xylanolytic,ustilaginomycetous yeast species isolated from an insect pest of sugarcane roots[J].International Journal of Systematic and Evolutionary Microbiology,2014,64(6):2159-2168.
[10]Basit A,Liu J,Miao T,et al.Characterization of two endo-b-1,4-xylanases from Myceliophthora thermophila and their saccharification efficiencies,synergistic with commercial cellulase[J].Frontiers in Microbiology,2018,9:1-11.
[11]朱辉,丁延芹,田方,等.多粘类芽孢杆菌SC2xynD和gluB的克隆及序列分析[J].生物技术通报,2008(4):171-174.
[12]Ghio S,Insani E,Pinninny F,et al.GH10XynA is the main xylanase identified in the crude enzymatic extractof Paenibacillus sp.A59when grown on xylan or lignocellulosic biomass[J].Microbiological Research,2016,186:16-26
[13]王中月,滕超,汤回花,等.产高分子量木聚糖酶细菌的筛选及产酶条件优化[J].食品与发酵工业,2017,43(10):88-95.
[14]马春玲,李思含,赵东芳,等.可产木聚糖酶的蕙兰根内生细菌的筛选与鉴定[J].河北大学学报,2017,37(5):518-522.
[15]董媛媛,李亚贤,沈艺红,等.响应面法优化木聚糖酶和α-葡萄糖醛酸酶联合水解桦木木聚糖工艺[J].食品科学,2018,39(4):125-131.
[16]卢海强,谷新晰,李晨,等.响应面法优化棒状乳杆菌HS4廉价增殖培养基研究[J].河北农业大学学报,2017,40(2):51-55.
[17]王晓宇,刘伟娜,谢响明,等.青霉L1来源具有生产木寡糖应用潜力的高比活GH11木聚糖酶[J].生物工程学报,2018,34(1):68-77.
[18]易旭东,潘虎,田云,等.一株碱性木聚糖酶产生菌的筛选及发酵条件的优化[J].中国农学通报2017,33(17):12-18.
[19]葛慧华,林锦霞,张光亚.以响应面法优化短小芽孢杆菌木聚糖酶产酶条件[J].华侨大学学报:自然科学版,2012(2):172-175.
[20]黄国勇,吴振强.枯草芽孢杆菌产木聚糖酶发酵条件的研究[J].河南工业大学学报:自然科学版,2006(1):32-35.