以甘油为底物发酵生产新型黄原胶
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摘要
经过驯化得到了一株可以利用甘油发酵生产黄原胶的野油菜黄单胞菌突变株,Xanthomonas. campestris WXLB-006。RT-PCR结果显示:X. campestris WXLB-006中甘油代谢相关基因(glpF,glpK,glpD,fbp)的相对表达量均高于原始菌株,依次为glpD (8.53)> glpF(7.64)> glpK(6.61)> fbp(5.79)。初始甘油质量浓度40 g/L、变通气量和变搅拌转速(0~24 h,0.5 vvm和200 r/min;24~60 h,1.0 vvm和400 r/min)以及三阶段变速流加甘油策略(24~34 h,3g/L/h;34~44 h,2 g/L/h;44~54 h,1 g/L/h),使生物量由1.65 g/L增加到1.94 g/L,黄原胶产量由17.8 g/L提高到33.9 g/L,发酵周期由120 h缩短至60 h。重要的是,以甘油为底物发酵得到的黄原胶具有粘度低、透明性高、水化速率快和反复冻融处理粘度增大的特点。
A mutant strain,which was named X.campestris WXLB-006 and could use glycerol for xanthan production,was obtained by adaptive evolution.RT-PCR results showed that the transcriptional level of genes related to glycerol metabolism(glpF,glpK,glpD,and fbp) in X.campestris WXLB-006 was higher than those of the parent strain,and the order of these four genes was glpD(8.53) > glpF(7.64) > glpK(6.61) > fbp(5.79).With low glycerol concentration(40 g/L),adoption of variable aeration rate and stirring speed(0~24 h,0.5 vvm and 200 rpm;24~60 h,1.0 vvm and 400 rpm),and three-stage varied speed feeding glycerol strategy(24~34 h,3 g/L/h;34~44 h,2 g/L/h;44~54 h,1 g/L/h),biomass increased from 1.65 g/L to 1.94 g/L and xanthan yield from17.8 g/L to 33.9 g/L,but fermentation time decreased from 120 h to 60 h.Importantly,xanthan gum produced from glycerol has the properties of low viscosity,high transparency,fast hydration rate,and increased viscosity after treated with-20 ℃ freeze-thaw cycles.
A mutant strain,which was named X.campestris WXLB-006 and could use glycerol for xanthan production,was obtained by adaptive evolution.RT-PCR results showed that the transcriptional level of genes related to glycerol metabolism(glpF,glpK,glpD,and fbp) in X.campestris WXLB-006 was higher than those of the parent strain,and the order of these four genes was glpD(8.53) > glpF(7.64) > glpK(6.61) > fbp(5.79).With low glycerol concentration(40 g/L),adoption of variable aeration rate and stirring speed(0~24 h,0.5 vvm and 200 rpm;24~60 h,1.0 vvm and 400 rpm),and three-stage varied speed feeding glycerol strategy(24~34 h,3 g/L/h;34~44 h,2 g/L/h;44~54 h,1 g/L/h),biomass increased from 1.65 g/L to 1.94 g/L and xanthan yield from17.8 g/L to 33.9 g/L,but fermentation time decreased from 120 h to 60 h.Importantly,xanthan gum produced from glycerol has the properties of low viscosity,high transparency,fast hydration rate,and increased viscosity after treated with-20 ℃ freeze-thaw cycles.
引文
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[14]LI R,FEKE D L.Rheological and kinetic study of the ultrasonic degradation of xanthan gum in aqueous solutions[J].Food Chemistry,2015,172:808-813.
[15]LI J,LIU R,CHANG G,et al.A strategy for the highly efficient production of docosahexaenoic acid by Aurantiochytrium limacinum SR21 using glucose and glycerol as the mixed carbon sources[J].Bioresource Technology,2015,177:51-57.
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[17]BRETZ K.Succinic acid production in fed-batch fermentation of Anaerobiospirillum succiniciproducens using glycerol as carbon source[J].Chemical Engineering&Technology,2015,38(9):1659-1664.
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[19]CASAS J A,SANTOS V E,GARCIA-OCHOA F.Xanthan gum production under several operational conditions:molecular structure and rheological properties[J].Enzyme and Microbial Technology,2000,26(2-4):282-291.
[20]KRISHNA L J,SHARMA G.Studies on xanthan production from Xanthomonas campestris[J].Bioprocess Engineering,2000,23(6):687-689.
[21]CHEN S,CHU J,ZHUANG Y,et al.Enhancement of inosine production by Bacillus subtilis through suppression of carbon overflow by sodium citrate[J].Biotechnology Letters,2005,27(10):689-692.
[22]CHO H M,YOO B.Rheological characteristics of cold thickened beverages containing xanthan gum-based food thickeners used for dysphagia diets[J].Journal of the Academy of Nutrition and Dietetics,2015,115(1):106-111.
[23]LAN H J,ZHAO L,SU L,et al.Food cold chain equilibrium based on collaborative replenishment[J].Journal of Applied Research and Technology,2014,12(2):201-211.
[2]GALLARDO R,FARIA C,RODRIGUES L R,et al.Anaerobic granular sludge as a biocatalyst for 1,3-propanediol production from glycerol in continuous bioreactors[J].Bioresource Technology,2014,155:28-33.
[3]ANASTACIO G S,SANTOS K O,SUAREZ P A Z,et al.Utilization of glycerin byproduct derived from soybean oil biodiesel as a carbon source for heterologous protein production in Pichia pastoris[J].Bioresource Technology,2014,152:505-510.
[4]ZHU H,YI X,LIU Y,et al.Production of acetol from glycerol using engineered Escherichia coli[J].Bioresource Technology,2013,149:238-243.
[5]ROSALAM S,ENGLAND R.Review of xanthan gum production from unmodified starches by Xanthomonas camprestris sp[J].Enzyme and Microbial Technology,2006,39(2):197-207.
[6]TANG Minmin,HONG Yan,GU Zhengbiao,et al.Effect of xanthan on long-term retrogradation of rice starch[J].Journal of Food Science and Biotechnology,2013,32(7).(in Chinese)
[7]PALANIRAJ A,JAYARAMAN V.Production,recovery and applications of xanthan gum by Xanthomonas campestris[J].Journal of Food Engineering,2011,106(1):1-12.
[8]WANG Z,WU J,ZHU L,et al.Activation of glycerol metabolism in Xanthomonas campestris by adaptive evolution to produce a high-transparency and low-viscosity xanthan gum from glycerol[J].Bioresource Technology,2016,211:390-397.
[9]SUN X M,REN L J,JI X J,et al.Adaptive evolution of Schizochytrium sp.by continuous high oxygen stimulations to enhance docosahexaenoic acid synthesis[J].Bioresource Technology,2016,211:374-381.
[10]SHAW R G,SHAW F H.Quantitative genetic study of the adaptive process[J].Heredity,2014,112(1):13-20.
[11]KALOGIANNIS S,IAKOVIDOU G,LIAKOPOULOU-KYRIAKIDES M,et al.Optimization of xanthan gum production by Xanthomonas campestris grown in molasses[J].Process Biochemistry,2003,39(2):249-256.
[12]WANG D,JU X,ZHOU D,et al.Efficient production of pullulan using rice hull hydrolysate by adaptive laboratory evolution of Aureobasidium pullulans[J].Bioresource Technology,2014,164:12-19.
[13]SUTHERLAND I W.Microbial polysaccharides from gram-negative bacteria[J].International Dairy Journal,2001,11(9):663-674.
[14]LI R,FEKE D L.Rheological and kinetic study of the ultrasonic degradation of xanthan gum in aqueous solutions[J].Food Chemistry,2015,172:808-813.
[15]LI J,LIU R,CHANG G,et al.A strategy for the highly efficient production of docosahexaenoic acid by Aurantiochytrium limacinum SR21 using glucose and glycerol as the mixed carbon sources[J].Bioresource Technology,2015,177:51-57.
[16]SWINNEN S,KLEIN M,CARRILLO M,et al.Re-evaluation of glycerol utilization in Saccharomyces cerevisiae:characterization of an isolate that grows on glycerol without supporting supplements[J].Biotechnolgy for Biofuels,2013,6(1):1-25.
[17]BRETZ K.Succinic acid production in fed-batch fermentation of Anaerobiospirillum succiniciproducens using glycerol as carbon source[J].Chemical Engineering&Technology,2015,38(9):1659-1664.
[18]BANDAIPHET C,PRASERTSAN P.Effect of aeration and agitation rates and scale-up on oxygen transfer coefficient,k La in exopolysaccharide production from Enterobacter cloacae WD7[J].Carbohydrate Polymers,2006,66(2):216-228.
[19]CASAS J A,SANTOS V E,GARCIA-OCHOA F.Xanthan gum production under several operational conditions:molecular structure and rheological properties[J].Enzyme and Microbial Technology,2000,26(2-4):282-291.
[20]KRISHNA L J,SHARMA G.Studies on xanthan production from Xanthomonas campestris[J].Bioprocess Engineering,2000,23(6):687-689.
[21]CHEN S,CHU J,ZHUANG Y,et al.Enhancement of inosine production by Bacillus subtilis through suppression of carbon overflow by sodium citrate[J].Biotechnology Letters,2005,27(10):689-692.
[22]CHO H M,YOO B.Rheological characteristics of cold thickened beverages containing xanthan gum-based food thickeners used for dysphagia diets[J].Journal of the Academy of Nutrition and Dietetics,2015,115(1):106-111.
[23]LAN H J,ZHAO L,SU L,et al.Food cold chain equilibrium based on collaborative replenishment[J].Journal of Applied Research and Technology,2014,12(2):201-211.