高产D-塔格糖植物乳杆菌WU14的L-阿拉伯糖异构酶发酵培养基优化
|
摘要
D-塔格糖(D-tagatose)是一种新型甜味剂,具有降低血糖、改善肠道菌群等功能,被广泛用于食品及药品等领域。植物乳杆菌WU14(Lactobacillus plantarum WU14)可通过L-阿拉伯糖异构酶转化D-塔格糖,为提高其转化能力,以植物乳杆菌WU14的生物量及L-阿拉伯糖异构酶酶活为指标,通过单因素实验与响应面实验对高产D-塔格糖植物乳杆菌WU14发酵培养基及培养条件进行优化。结果表明,发酵培养基组成为(g/L):葡萄糖4.35 g、L-阿拉伯糖2.71 g、酵母浸粉17.6g、大豆蛋白胨17.6 g、无水乙酸钠10 g、MgSO_4·7H_2O 0.4 g、MnSO_4·2H_2O 0.05 g、K_2HPO_4 0.4g;发酵条件为:发酵初始pH值6.24、发酵温度28.6℃、接种量2%、发酵20 h,在此条件下,植物乳杆菌WU14的菌体量比优化前提高了近48%,L-阿拉伯糖异构酶酶活达42.23 U/mL,D-塔格糖的转化率达到69.6%。培养基的优化显著提高植物乳杆菌WU14的菌体量及L-阿拉伯糖异构酶活力,为D-塔格糖进一步工业化发酵生产提供理论基础。
D-tagatose is a new sweetener that has the functions of lowering blood sugar and improving intestinal flora, and is widely used in food and medicine. Lactobacillus plantarum WU14 has the ability to transform D-tagatose by L-arabinose isomerase. In order to improve the ability of tranformation, the biomass and the activity of L-arabinose isomerase of L. plantarum WU14 were used as the indicators, the single factor and response surface analysis were employed to optimize the fermentation medium and conditions of L. plantarum WU14 with high D-tagatose yield. The results showed that the optimal composition of the fermentation medium was(g/L): glucose 4.35 g, L-arabinose 2.71 g, yeast extract powder 17.6 g, soy peptone 17.6 g, anhydrous sodium acetate 10 g, MgSO_4·7 H_2O 0.4 g, MnSO_4·2 H_2O 0.05 g, K_2HPO_4 0.4 g; fermentation conditions: initial pH6.24, fermentation temperature 28.6 ℃, inoculum 2%, fermentation 20 h. With these optimizations, the biomass of L. plantarum WU14 increased by nearly 48%, the L-arabinose isomerase activity reached 42.23 U/mL and the conversion rate of D-tagatose reached 69.6%. The optimization of the medium significantly increased the biomass and L-arabinose isomerase activity of L. plantarum WU14, which provided a theoretical basis for further industrial fermentation production of D-tagatose.
D-tagatose is a new sweetener that has the functions of lowering blood sugar and improving intestinal flora, and is widely used in food and medicine. Lactobacillus plantarum WU14 has the ability to transform D-tagatose by L-arabinose isomerase. In order to improve the ability of tranformation, the biomass and the activity of L-arabinose isomerase of L. plantarum WU14 were used as the indicators, the single factor and response surface analysis were employed to optimize the fermentation medium and conditions of L. plantarum WU14 with high D-tagatose yield. The results showed that the optimal composition of the fermentation medium was(g/L): glucose 4.35 g, L-arabinose 2.71 g, yeast extract powder 17.6 g, soy peptone 17.6 g, anhydrous sodium acetate 10 g, MgSO_4·7 H_2O 0.4 g, MnSO_4·2 H_2O 0.05 g, K_2HPO_4 0.4 g; fermentation conditions: initial pH6.24, fermentation temperature 28.6 ℃, inoculum 2%, fermentation 20 h. With these optimizations, the biomass of L. plantarum WU14 increased by nearly 48%, the L-arabinose isomerase activity reached 42.23 U/mL and the conversion rate of D-tagatose reached 69.6%. The optimization of the medium significantly increased the biomass and L-arabinose isomerase activity of L. plantarum WU14, which provided a theoretical basis for further industrial fermentation production of D-tagatose.
引文
[1]王静.甜味剂D-塔格糖的生产及其在食品中的应用[J].中国调味品,2016,41(01):140-143.
[2]Kwak J H,Kim M S,Lee J H,et al.Beneficial effect of tagatose consumption on postprandial hyperglycemia in Koreans:a double-blind crossover designed study[J].Food&Function,2013,8(4):1223-1228.
[3]M Rouhi,R Mohammadi,A M Mortazavian,et al.Combined effects of replacement of sucrose with D-tagatose and addition of different probiotic strains on quality characteristics of chocolate milk[J].Dairy Science&Technology,2015,95(2).
[4]梁真.塔格糖酸乳发酵条件及稳定性的研究[D].济南:齐鲁工业大学,2015.
[5]徐铮,李贵祥,李莎,等.生物法生产D-塔格糖的研究进展[J].生物加工过程,2013,11(04):64-71.
[6]Cheetham P S J,Wootton A N.Bioconversion of D-galactose into D-tagatose[J].Enzyme Microb Techol,1993,15:105-108.
[7]Babu A,Manjasetty,Mark R.Chance Crystal structure of E.coli L-arabinose isomerase,the putative target of biological tagatose production[J].J MolBiol,2006,360:297-309.
[8]Rhimi M,Juy M,Aghajari N,et al.Probing the essential catalytic residues and the substrate affinity in the thermoactive Bacillus stearothermophilus US100L-arabinose isomerase by site-directed mutagenesis[J].JBacteriol,2007,189:3556-3563.
[9]Oh HJ,Kim HJ,Oh DK.Increase in D-tagatose production rate by sitedirected mutagenesis of L-arabinose isomerase from Geobacillus thermodenitrificans[J].Biotechnol Lett,2006,28:145-149.
[10]Kim BC,Lee YH,Lee HS,et al.Cloning,expression and characterization of L-arabinose isomerase from Thermotoga neapolitana:bioconversion of D-galactose to D-tagatose using the enzyme[J].FEMS Microbiol Lett,2002,212:121-126.
[11]Wending Mei,Lu Wang,Ying Zang,et al.Characterization of an L-arabinose isomerase from Bacillus coagulans NL01 and its application for D-tagatose production[J].BMC Biotechnology,2016,16(1).
[12]Wei Xu,Chen Fan,Tao Zhang,et al.Cloning,Expression,and Characterization of a Novel L-Arabinose Isomerase from the Psychrotolerant Bacterium Pseudoalteromonas haloplanktis[J].Molecular Biotechnology,2016,58(11).
[13]Salonen N,Salonen K,Leisola M,et al.D-Tagatose production in the presence of borate by resting Lactococcus lactis cells harboring Bifidobacterium longum L-arabinose isomerase[J].Bioproc Biosyst Eng,2013,36(4):489-497.
[14]Fina Amreta Laksmi,Shigeki Arai,Hirohito Tsurumaru,et al.Improved substrate specificity for D-galactose of L-arabinose isomerase for industrial application[J].BBA-Proteins and Proteomics,2018.
[15]梁敏.Thermoanaerobacter mathranii来源的L-阿拉伯糖异构酶(TaMAI)及其转化D-塔格糖的研究[D].济南:山东大学,2012.
[16]Jebors S,Tauran Y,Aghajari N,et al.Supramolecular stabilization of acid tolerant L-arabinose isomerase from Lactobacillus sakei[J].Chem Commun,2011,47:12307-12309.
[17]J Jayamuthunagai,G Srisowmeya,M Chakravarthy,et al.D-Tagatose production by permeabilized and immobilized Lactobacillus plantarum using whey permeate[J].Bioresource Technology,2017,235.
[18]刘新颖.大肠杆菌工程菌发酵乳糖制备D-塔格糖的研究[D].济南:山东大学,2014.
[19]应碧.高产D-塔格糖乳酸菌L-AI基因的分析、食品级诱导表达及其酶学性质的研究[D].南昌:江西农业大学,2015.
[20]张华.塔格糖的生物制备及L-阿拉伯糖异构酶性质研究[D].无锡:江南大学,2007.
[2]Kwak J H,Kim M S,Lee J H,et al.Beneficial effect of tagatose consumption on postprandial hyperglycemia in Koreans:a double-blind crossover designed study[J].Food&Function,2013,8(4):1223-1228.
[3]M Rouhi,R Mohammadi,A M Mortazavian,et al.Combined effects of replacement of sucrose with D-tagatose and addition of different probiotic strains on quality characteristics of chocolate milk[J].Dairy Science&Technology,2015,95(2).
[4]梁真.塔格糖酸乳发酵条件及稳定性的研究[D].济南:齐鲁工业大学,2015.
[5]徐铮,李贵祥,李莎,等.生物法生产D-塔格糖的研究进展[J].生物加工过程,2013,11(04):64-71.
[6]Cheetham P S J,Wootton A N.Bioconversion of D-galactose into D-tagatose[J].Enzyme Microb Techol,1993,15:105-108.
[7]Babu A,Manjasetty,Mark R.Chance Crystal structure of E.coli L-arabinose isomerase,the putative target of biological tagatose production[J].J MolBiol,2006,360:297-309.
[8]Rhimi M,Juy M,Aghajari N,et al.Probing the essential catalytic residues and the substrate affinity in the thermoactive Bacillus stearothermophilus US100L-arabinose isomerase by site-directed mutagenesis[J].JBacteriol,2007,189:3556-3563.
[9]Oh HJ,Kim HJ,Oh DK.Increase in D-tagatose production rate by sitedirected mutagenesis of L-arabinose isomerase from Geobacillus thermodenitrificans[J].Biotechnol Lett,2006,28:145-149.
[10]Kim BC,Lee YH,Lee HS,et al.Cloning,expression and characterization of L-arabinose isomerase from Thermotoga neapolitana:bioconversion of D-galactose to D-tagatose using the enzyme[J].FEMS Microbiol Lett,2002,212:121-126.
[11]Wending Mei,Lu Wang,Ying Zang,et al.Characterization of an L-arabinose isomerase from Bacillus coagulans NL01 and its application for D-tagatose production[J].BMC Biotechnology,2016,16(1).
[12]Wei Xu,Chen Fan,Tao Zhang,et al.Cloning,Expression,and Characterization of a Novel L-Arabinose Isomerase from the Psychrotolerant Bacterium Pseudoalteromonas haloplanktis[J].Molecular Biotechnology,2016,58(11).
[13]Salonen N,Salonen K,Leisola M,et al.D-Tagatose production in the presence of borate by resting Lactococcus lactis cells harboring Bifidobacterium longum L-arabinose isomerase[J].Bioproc Biosyst Eng,2013,36(4):489-497.
[14]Fina Amreta Laksmi,Shigeki Arai,Hirohito Tsurumaru,et al.Improved substrate specificity for D-galactose of L-arabinose isomerase for industrial application[J].BBA-Proteins and Proteomics,2018.
[15]梁敏.Thermoanaerobacter mathranii来源的L-阿拉伯糖异构酶(TaMAI)及其转化D-塔格糖的研究[D].济南:山东大学,2012.
[16]Jebors S,Tauran Y,Aghajari N,et al.Supramolecular stabilization of acid tolerant L-arabinose isomerase from Lactobacillus sakei[J].Chem Commun,2011,47:12307-12309.
[17]J Jayamuthunagai,G Srisowmeya,M Chakravarthy,et al.D-Tagatose production by permeabilized and immobilized Lactobacillus plantarum using whey permeate[J].Bioresource Technology,2017,235.
[18]刘新颖.大肠杆菌工程菌发酵乳糖制备D-塔格糖的研究[D].济南:山东大学,2014.
[19]应碧.高产D-塔格糖乳酸菌L-AI基因的分析、食品级诱导表达及其酶学性质的研究[D].南昌:江西农业大学,2015.
[20]张华.塔格糖的生物制备及L-阿拉伯糖异构酶性质研究[D].无锡:江南大学,2007.
© 2004-2018 中国地质图书馆版权所有 京ICP备05064691号 京公网安备11010802017129号 地址：北京市海淀区学院路29号 邮编：100083 电话：办公室：(+86 10)66554848；文献借阅、咨询服务、科技查新：66554700 |