CN113736713A - 食窦魏斯氏乳酸菌nc516·11及其胞外多糖和应用 - Google Patents
食窦魏斯氏乳酸菌nc516·11及其胞外多糖和应用 Download PDFInfo
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Abstract
本发明公开了食窦魏斯氏乳酸菌NC516.11及其胞外多糖和应用,保藏编号为:CGMCC No.23107。该菌株是在汾酒酒曲中分离纯化获得,经鉴定为食窦魏斯氏乳酸菌。经实验验证,该菌株能够提高荞麦面团中总酚、黄酮含量,促进营养物质的释放。同时可以改善荞麦面包的微观结构,减少烘焙损失。本发明所述食窦魏斯氏乳酸菌所制得的荞麦面包口感柔软、营养健康,市场前景广阔。
Description
技术领域
本发明涉及微生物及食品发酵,具体涉及食窦魏斯氏乳酸菌NC516.11及其胞外多糖和应用。
背景技术
近年来随着人们生活水平的提高,人们的饮食观念从吃得饱逐渐转变为吃得好。人们对粗粮、杂粮面包的需求逐渐上升。但是目前国内市场的面包制品多为纯小麦制成,粗粮、杂粮面包产量远远低于小麦面包。主要原因还是荞麦粉缺少麸质,制成的面团易塌陷,筋力弱,不易成型,制成的面包易碎屑,水分散失快,随着储藏时间增长面包变得干硬。
乳酸菌作为一种益生菌广泛应用于食品发酵行业,大部分乳酸菌在发酵过程中都会产生乳酸等次级代谢产物,将其应用于荞麦面团可以促进荞麦中总酚、黄酮析出、增加面包的抗氧化性。某些乳酸菌在发酵过程中还会产生胞外多糖,具有凝胶性,增稠性,稳定性等优良特性,可以改善荞麦面团的缺陷。
发明内容
发明目的:本发明的目的在于提供了一株食窦魏斯氏乳酸菌NC516.11以及其副产物胞外多糖,其中胞外多糖能够改善荞麦面团的流变特性。将NC516.11应用于荞麦面包制作可以增加营养物质释放,改善荞麦面包的质构的同时减少荞麦面包的烘焙损失,还能够减少荞麦面包在储藏时期的品质裂变。
技术方案:本发明所述的食窦魏斯氏乳酸菌NC516.11,已保藏于中国微生物菌种保藏管理委员会普通微生物中心,保藏时间为2021年8月2日,保藏编号为:CGMCCNo.23107;地址:北京市朝阳区北辰西路1号院3号,中国科学院微生物研究所,邮编:100101。
胞外多糖,由所述的食窦魏斯氏乳酸菌NC516.11所制。
所述胞外多糖的制备方法,包括以下步骤:
(1)将食窦魏斯氏乳酸菌NC516.11接种于培养基,30~40℃培养24~24h,培养结束后将三角瓶置于沸水浴中灭活乳酸菌,冷却至室温,离心除去乳酸菌;
(2)将上清与三氯乙酸溶液混合,2-6℃静置,2-6℃离心去除蛋白;
(3)在步骤(2)所得溶液中加入乙醇,2-6℃静置,2-6℃离心得到粗多糖。
荞麦面团,包括以下质量份的原料:90~110份荞麦粉、50~70份水、0.04~0.08份所述的胞外多糖;或者90~110份荞麦粉、50~70份菌液、4~6份蔗糖,其中所述菌液为含有权利要求1所述食窦魏斯氏乳酸菌NC516.11的水溶液,菌落数1*107~5*107CFU/mL。
所述荞麦面团的制作方法,包括以下步骤:
将荞麦粉、水、胞外多糖混合,制得荞麦面团,将荞麦面团置于30~40℃、80%~90%湿度条件下发酵6~10h得到;
或,将荞麦粉,菌液,蔗糖,制得荞麦面团,将荞麦面团置于25~35℃、80%~90%湿度条件下发酵6~10h得到。
荞麦面包,包括以下质量份的原料:10~30份权利要求4所述荞麦面团、10~30份小麦粉、40~50份水、0.5~1.2份酵母,25~35℃、80%~90%湿度条件下发酵1~1.5h,170~210℃烘焙15~30min得到。
所述胞外多糖在提高荞麦面团流变特性中的应用。
所述食窦魏斯氏乳酸菌NC516.11在提高荞麦面团中总酚、黄酮含量的应用。
所述食窦魏斯氏乳酸菌NC516.11在减少荞麦面包烘焙损失中的应用。
所述食窦魏斯氏乳酸菌NC516.11在改善荞麦面包结构、质构、减少储藏过程中的品质裂变中的应用。
所述胞外多糖在在荞麦面团中的应用。
所述食窦魏斯氏乳酸菌NC516.11在荞麦面包中的应用。所述食窦魏斯氏乳酸菌NC516.11在荞麦面团中的应用。
有益效果:与现有技术相比,本发明具有以下显著优点:(1)本发明的食窦魏斯氏菌NC516.11在生长过程中产生的代谢产物胞外多糖可以作为一种凝胶剂、增稠剂应用于食品中。(2)食窦魏斯氏菌NC516.11所产生的胞外多糖可以增加了荞麦面团的流变学特性,改善了荞麦面团筋力弱的缺陷,使荞麦面团具有更好的加工品质。(3)由食窦魏斯氏菌NC516.11发酵的荞麦酸面团可以增加荞麦中营养物质总酚、黄酮含量的释放,其做成的面包质构好,烘焙损失小。(4)食窦魏斯氏菌NC516.11可以改善荞麦面包的微观结构,在储藏过程中减少水分流失,不易产生品质裂变。(5)食窦魏斯氏菌NC516.11可以使得荞麦面包更加柔软,硬度减小。
附图说明
图1是食窦魏斯氏菌胞外多糖的流变情况;
图2是食窦魏斯氏菌胞外多糖发酵荞麦面团的流变情况;
图3是荞麦面团总酚和黄酮含量;
图4是荞麦面包的烘焙损失;
图5是荞麦面包的微观结构;
图6是荞麦面包的在储藏期间硬度的变化情况;
图7是荞麦面包的在储藏期间低场核磁共振图谱(A:2h,B:24h,C:48h,D:72h)。
具体实施方式
实施例1菌株的筛选
本发明提供了一株食窦魏斯氏菌(Weissella cibaria)NC516.11.所述食窦魏斯氏菌NC516.11是在汾酒酒曲中分离纯化鉴定获得,以无菌操作称取10g酒曲样品,放入装有90ml无菌生理盐水的锥形瓶中充分震荡摇匀,用无菌生理盐水进行十倍梯度稀释,选取适宜稀释度的样品稀释液,涂布于MRS固体培养基上,置于37℃培养24~48h。挑取单菌落,在MRS琼脂平板上多次划线纯化,直至整个平板上的菌落形态一致,挑取单菌落到MRS液体培养基增菌培养。得到的菌株均在含40%甘油的MRS液体培养基中于-80℃冷冻保存。
菌种鉴定
利用16S rDNA对其测序得出食窦魏斯氏菌NC516.11的序列
在NCBI网站上进行BLAST序列比对,结果显示该序列与食窦魏斯氏菌的16SrDNA序列同源性超过99%,故判定菌株为食窦魏斯氏菌。
实施例2食窦魏斯氏菌NC516.11胞外多糖的制备及流变学特性测量
将食窦魏斯氏乳酸菌NC516.11以2%的体积比接种于mMRS培养基(每100mL MRS培养基中添加2%的蔗糖)35℃培养24-24h。培养结束后将三角瓶置于100℃水浴中煮10min灭活乳酸菌。冷却至室温后将培养液于4℃10000g离心20min以除去乳酸菌。将上清与等比例的10%三氯乙酸溶液混合,于4℃静置12h,4℃12000g 20min离心去除蛋白。在上清中加入4倍体积的乙醇,于4℃静置12h,4℃12000g 30min离心得到粗多糖。将粗多糖溶解于超纯水中,用截留分子量8000-14000的透析袋于4℃透析48h,每8h换水一次。将透析过的粗多糖溶液于-80℃冰箱冷冻,冷冻干燥后待用。
将胞外多糖以4%、6%、8%(m/v)溶解于蒸馏水中,利用流变仪测量多糖溶液的表观粘度曲线和扫描频率曲线。表观粘度采用50mm平板,测定距离1mm,25℃,剪切速率0.1~10l/s。扫描频率选择0.1~10Hz范围。图1A显示随着剪切速率的增加,各浓度EPS溶液的表观粘度均呈下降趋势,表现出剪切变稀的特点,在相同的剪切速率下,EPS的浓度越高,表观黏度越高。当EPS的浓度从4%增加到6%(w/v)时,EPS的剪切粘度增加了15倍左右。图1B和C显示不同浓度EPS的频率扫描。从图中可以看出,随着扫描频率的增加,各浓度EPS的G′和G″呈上升趋势。在扫描频率范围内,G″>G′,说明高浓度的EPS具有凝胶特性,可以很好地用作增稠剂。
实施例3利用食窦魏斯氏菌胞外多糖制备发酵荞麦面团
将胞外多糖以4%、6%、8%(m/v)溶解于蒸馏水中,将荞麦粉、包含食窦魏斯氏菌胞外多糖的水、混合,得到荞麦面团,将荞麦面团置于35℃、80%~90%湿度条件下发酵6~10h,得到发酵荞麦面团,利用流变仪测量面团的流变学特性,选用25mm平板,间隙2mm,频率0.1~10Hz。
图2显示荞麦面团中G′和G″都随频率的增加而增加。G′值在同一频率下随EPS含量的增加而增加,说明高浓度EPS能使荞麦面团具有更大的弹性。荞麦面团具有G′>G″的特点。所有面团都呈现出弹性大于粘度的特性,这在面团的切割和成型过程中起到了积极的作用。
实施例4利用食窦魏斯氏菌制备发酵荞麦面团
根据前期试验得出食窦魏斯氏菌NC516.11的最适生长温度为35℃,最佳产糖温度为25℃。故本发明设置了E35组、E25组以及对照组。
E35组:将食窦魏斯氏菌NC516.11以2%的体积比接种于MRS培养基37℃培养24-24h,以5000g离心力离心,用无菌水洗涤沉淀的菌体并调制成菌落数≥1*107CFU/mL。将100g荞麦粉,60mL菌液,5g蔗糖倒入和面机,搅拌速度100~150rpm,将搅拌好的面团置于35℃发酵箱。80%~90%湿度条件下发酵6~10h,从而制成E35组荞麦酸面团。
E25组:将食窦魏斯氏菌NC516.11以2%的体积比接种于MRS培养基37℃培养24-24h,以5000g离心力离心,用无菌水洗涤沉淀的菌体并调制成菌落数≥1*107CFU/mL。将100g荞麦粉,60mL菌液,5g蔗糖倒入和面机,搅拌速度100~150rpm,将搅拌好的面团置于25℃发酵箱。80%~90%湿度条件下发酵6~10h,从而制成E25组荞麦酸面团。
对照组:将100g荞麦粉,60mL水,5g蔗糖倒入和面机,搅拌速度100~150rpm,将搅拌好的面团置于35℃发酵箱80%~90%湿度条件下发酵6~10h,从而制成对照组荞麦酸面团。
测定各组荞麦面团总酚和黄酮含量测定结果见图3。由图可知,食窦魏斯氏菌NC516.11发酵后的荞麦面团总酚和黄酮含量都明显高于对照组(P<0.05),其中E35组发酵的荞麦面团总酚和黄酮含量最高。说明食窦魏斯氏菌NC516.11发酵可以增加荞麦中营养物质的析出。
实施例5利用食窦魏斯氏菌制备荞麦面包
将实施例4中发酵好的荞麦酸面团进行二次发酵并制成面包,具体方法如下:
E35组:将20g E35组酸面团与80g小麦粉,0.8g酵母,48mL水以100~150rpm的速度搅拌混合,将混合好的面团分割成90g的小面团置于35℃,80%~90%湿度条件下发酵1~1.5h。发酵完成后置于烤箱上火170℃,下火210℃,烘焙20min后取出。
E25组:将20g E25组酸面团与80g小麦粉,0.8g酵母,48mL水以100~150rpm的速度搅拌混合,将混合好的面团分割成90g的小面团置于35℃,80%~90%湿度条件下发酵1~1.5h。发酵完成后置于烤箱上火170℃,下火210℃,烘焙20min后取出。
对照组:将20g对照组酸面团与80g小麦粉,0.8g酵母,48mL水以100~150rpm的速度搅拌混合,将混合好的面团分割成90g的小面团置于35℃,80%~90%湿度条件下发酵1~1.5h。发酵完成后置于烤箱上火170℃,下火210℃,烘焙20min后取出。
测定各组荞麦面包的烘焙损失测定结果见图4。由图可知,由食窦魏斯氏菌NC516.11发酵制成的荞麦面包与对照组相比可以减少面包在烘焙过程中的损失(P<0.05)。其中E25组比E35组的烘焙损失略小。
拍摄各组荞麦面包的微观结构结果见图5。由图可知,在200倍放大倍数下,E25和E35组面包较对照组显示出更为细腻的结构。500倍下E25面包基本没有出现断层以及较大孔洞,1000倍下,E25面包光滑细腻,E35组略显粗糙并伴随少量的缝隙,对照组则显示出粗糙的特点并伴随许多大小不一的缝隙。在5000倍条件下。E25面包效果最好,没有出现E35面包的缝隙以及对照面包的大量孔洞。
测定各组荞麦面包的在储藏期间硬度的变化情况,选用质构仪,P/36探头。测前3mm/s,测中1mm/s,测后1mm/s。压缩程度40%,触发力5g,测定结果见图6。有图可知,在烘烤2h后,由食窦魏斯氏菌NC516.11制成的面包硬度低于对照组。随着储藏时间的延长,各组面包都呈现出硬度加大的趋势。在相同储藏时间内,E25组和E35组硬度都要小于对照组,E25组硬度都要小于E35组(P<0.05)。图7显示荞麦面包的在储藏期间低场核磁共振图谱(A:2h,B:24h,C:48h,D:72h)随着储藏时间的延长,各组水分含量都呈现降低趋势,其中E25组和E35组荞麦面包比对照组水分含量降低慢,说明食窦魏斯氏菌NC516.11可以帮助面包保持水分。
Claims (10)
1.食窦魏斯氏乳酸菌NC516.11,保藏编号为:CGMCC No.23107。
2.一种胞外多糖,其特征在于,由权利要求1所述的食窦魏斯氏乳酸菌NC516.11所制。
3.一种权利要求2所述胞外多糖的制备方法,其特征在于,包括以下步骤:
(1)将食窦魏斯氏乳酸菌NC516.11接种于培养基,30~40℃培养24~24h,培养结束后将三角瓶置于沸水浴中灭活乳酸菌,冷却至室温,离心除去乳酸菌;
(2)将上清与三氯乙酸溶液混合,2-6℃静置,2-6℃离心去除蛋白;
(3)在步骤(2)所得溶液中加入乙醇,2-6℃静置,2-6℃离心得到粗多糖。
4.一种荞麦面团,其特征在于,包括以下质量份的原料:90~110份荞麦粉、50~70份水、0.04~0.08份权利要求2所述的胞外多糖;或者90~110份荞麦粉、50~70份菌液、4~6份蔗糖,其中所述菌液为含有权利要求1所述食窦魏斯氏乳酸菌NC516.11的水溶液,菌落数1*107~5*107CFU/mL。
5.一种权利要求4所述荞麦面团的制作方法,其特征在于,包括以下步骤:
将荞麦粉、水、胞外多糖混合,制得荞麦面团,将荞麦面团置于30~40℃、80%~90%湿度条件下发酵6~10h得到;
或,将荞麦粉,菌液,蔗糖,制得荞麦面团,将荞麦面团置于25~35℃、80%~90%湿度条件下发酵6~10h得到。
6.一种荞麦面包,其特征在于,包括以下质量份的原料:10~30份权利要求4所述荞麦面团、10~30份小麦粉、40~50份水、0.5~1.2份酵母,25~35℃、80%~90%湿度条件下发酵1~1.5h,170~210℃烘焙15~30min得到。
7.权利要求2所述胞外多糖在提高荞麦面团流变特性中的应用。
8.权利要求1所述食窦魏斯氏乳酸菌NC516.11在提高荞麦面团中总酚、黄酮含量的应用。
9.权利要求1所述食窦魏斯氏乳酸菌NC516.11在减少荞麦面包烘焙损失中的应用。
10.权利要求1所述食窦魏斯氏乳酸菌NC516.11在改善荞麦面包结构、质构、减少储藏过程中的品质裂变中的应用。
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