CN116731908A - 一种植物乳植杆菌mt213及其应用 - Google Patents
一种植物乳植杆菌mt213及其应用 Download PDFInfo
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Abstract
本发明涉及微生物技术领域，具体涉及一种植物乳植杆菌MT213及其应用。本发明自我国传统酸面团(老酵子)中分离筛选出植物乳植杆菌(Lactiplantibacillus plantarum)MT213，并且对其保藏，保藏编号为CGMCC No.26569。植物乳植杆菌MT213具有广谱的抑菌活性，可显著抑制多种常见致病菌以及腐败菌的生长，具有延长货架期的作用，在食品、保健品以及日化用品等领域具有广阔的应用前景。
Description
技术领域
本发明涉及微生物技术领域，具体涉及一种植物乳植杆菌MT213及其应用。
背景技术
微生物污染可导致食品等产品发生腐败变质，甚至会引发疾病，严重危害人类健康。目前，控制微生物生长的手段主要有物理防腐、化学防腐和生物防腐。其中，化学防腐剂最为常用，但是存在生产污染大、使用安全性不高等局限性。生物防腐具有天然、高效、安全等优势，受到消费者青睐，是目前防腐技术的主要开发方向。
乳酸菌(LAB)可通过菌体或产生的代谢产物发挥抑制致病菌以及腐败菌生长的作用，从而延长产品的货架期。此外，乳酸菌还可以改善食品品质和风味，产生功能性代谢产物提高食品营养价值。乳酸菌是我国传统发酵食品中的优势微生物，因此从我国传统发酵食品中挖掘具有抑菌特性的微生物，用于食品、日化用品等抑菌防腐，延长食品、日化用品等货架期/保质期，具有重要的开发价值。
发明内容
本发明自传统酸面团(老酵子)筛选并分离的一株植物乳植杆菌，具有抑制致病菌以及腐败菌生长的效果。本发明的植物乳植杆菌可应用于馒头、面包、蛋糕或饼干等发酵食品中，作为发酵剂使用，延长产品货架期。
第一方面，本发明提供一种植物乳植杆菌(Lactiplantibacillus plantarum)MT213，所述植物乳植杆菌的保藏编号为：CGMCC No.26569。
本发明分离筛选得到一种植物乳植杆菌，保藏后的相关信息如下：
保藏编号：CGMCC No.26569，分类命名为：植物乳植杆菌Lactiplantibacillusplantarum，保藏单位：中国微生物菌种保藏管理委员会普通微生物中心，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，保藏日期：2023年2月20日。
进一步地，所述植物乳植杆菌(Lactiplantibacillus plantarum)MT213的16SrDNA序列如SEQ ID No.1所示。
本发明进一步提供一种菌剂，包括所述的植物乳植杆菌(Lactiplantibacillusplantarum)MT213或其发酵液。
本发明进一步提供所述植物乳植杆菌(Lactiplantibacillus plantarum)MT213，或所述菌剂在抑制致病菌以及腐败菌中的应用；
所述腐败与致病微生物包括革兰氏阳性菌、革兰氏阴性菌或真菌中的一种或多种。
进一步地，所述革兰氏阳性菌包括金黄色葡萄球菌、单细胞增生李斯特菌、枯草芽孢杆菌、蜡样芽孢杆菌、藤黄微球菌等中的一种或多种；和/或，所述革兰氏阴性菌包括大肠杆菌、沙门氏菌、荧光假单胞菌、铜绿假单胞菌等中的一种或多种；和/或，所述真菌包括黑曲霉、产黄青霉、扩展青霉、桔青霉、黄曲霉等中的一种或多种。
本发明进一步提供所述植物乳植杆菌(Lactiplantibacillus plantarum)MT213，或所述菌剂在制备功能性产品中的应用；所述功能性产品包括：食品、保健品、或日化用品。
第二方面，本发明提供一种提高面制品防腐保鲜的方法，在面制品的制备过程中加入所述的植物乳植杆菌(Lactiplantibacillus plantarum)MT213，或所述的菌剂。
进一步地，所述植物乳植杆菌(Lactiplantibacillus plantarum)MT213的添加量为106-109CFU/g。
第三方面，本发明提供一种面制品，制备所述面制品时，使用了植物乳植杆菌(Lactiplantibacillus plantarum)MT213。
进一步地，所述面制品为发酵面制品，制备所述发酵面制品时，所用发酵剂包括植物乳植杆菌(Lactiplantibacillus plantarum)MT213；优选地，所述发酵面制品为馒头、面包、蛋糕或饼干等中的一种或多种。
本发明具备如下有益效果：
本发明分离筛选得到一种植物乳植杆菌，将其命名为植物乳植杆菌(Lactiplantibacillus plantarum)MT213，该菌株具有广谱抑菌性，可以抑制多种致病菌和腐败菌的生长。
本发明将植物乳植杆菌MT213用于馒头的制备中，可以使得馒头贮藏期间微生物数量显著降低，从而延长馒头货架期。
本发明提供了一种具有广谱抑菌性的植物乳植杆菌MT213，可以显著抑制多种致病菌以及腐败菌的生长，为开发新型营养安全化食品、保健品、或日化用品等产品提供了新的菌株资源。
附图说明
为了更清楚地说明本发明或现有技术中的技术方案，下面将对实施例或现有技术描述中所需要使用的附图作一简单地介绍，显而易见地，下面描述中的附图是本发明的一些实施例，对于本领域普通技术人员来讲，在不付出创造性劳动的前提下，还可以根据这些附图获得其他的附图。
图1为本发明实施例1中植物乳植杆菌(Lactiplantibacillus plantarum)MT213的菌落形态图。
图2为本发明实施例1中植物乳植杆菌(Lactiplantibacillus plantarum)MT213的革兰氏染色图。
图3为本发明实施例1中植物乳植杆菌(Lactiplantibacillus plantarum)MT213***发育树示意图。
具体实施方式
为使本发明的目的、技术方案和优点更加清楚，下面将结合本发明中的附图，对本发明中的技术方案进行清楚、完整地描述，显然，所描述的实施例是本发明一部分实施例，而不是全部的实施例。基于本发明中的实施例，本领域普通技术人员在没有作出创造性劳动前提下所获得的所有其他实施例，都属于本发明保护的范围。
实施例1抗微生物植物乳植杆菌MT213的筛选与鉴定
1、菌株MT213的分离纯化
挑选传统老酵子为样品稀释后，选取合适的稀释液涂布于含有CaCO3的MRS固体培养基平板，静置，于30-37℃培养36-48h，挑选出有溶钙圈的菌株，将分离菌株于MRS固体培养基培养24-48h后，观察发现菌落形态多呈乳白色圆形、中间略微***、不透明或半透明、边缘整齐(图1)。革兰氏染色呈阳性，菌体形态呈短杆(图2)。将那些在周围产生透明圈的单菌落进行划线分离培养，然后挑取单菌落于MRS液体培养基中30-37℃下培养至对数期后，将发酵液放入制作好的甘油管中放入-40℃保藏备用。
2、具有广谱抑菌活性的菌株筛选
指示菌株包括细菌(金黄色葡萄球菌ATCC 25923、大肠杆菌ATCC 44752、蜡样芽孢杆菌ATCC 11778、荧光假单胞菌ATCC 13525、枯草芽孢杆菌CGMCC 1.3358、沙门氏菌ATCC14028、单细胞增生李斯特菌ATCC 10403s、藤黄微球菌ATCC 10240)和真菌(黑曲霉ATCC16404、产黄青霉ATCC 10106、扩展青霉CICC 40658、桔青霉、黄曲霉ATCC 9643)。
指示菌株及LAB保存于河北科技大学食品与生物技术实验室。细菌和真菌分别在营养肉汤(NB)和马铃薯葡萄糖琼脂(PDA)中培养。上述细菌摇床培养至对数期后，用紫外分光光度计调至菌悬液浓度为108CFU/mL，备用。上述真菌在PDA平板上培养4-7d后，用0.85％生理盐水洗涤孢子，血球计数板调整孢子数为106CFU/mL，4℃储存备用。
菌株抑菌特性评价：将上述指示菌接种到NA和PDA培养基后倒平板，细菌平板终浓度为106CFU/mL，真菌平板终浓度为104CFU/mL。将保藏的乳酸菌菌种于液体MRS培养基中30-37℃活化18-24h，之后按体积百分比1-3％接种量加入到液体MRS培养基，于30-37℃培养24-48h后，将发酵液离心过膜后取上清注入细菌平板牛津杯中；将上清液冷冻干燥后将浓缩上清液注入真菌平板牛津杯中，平板放于4℃冰箱中预扩散后于30-37℃(真菌25-28℃)培养箱中培养18-24h(真菌24-48h)，观察抑菌效果，测量并记录抑菌圈的直径大小，见表1。
表1植物乳植杆菌MT213的抑菌谱
3、乳酸菌菌株的鉴定
对上述分离得到的乳酸菌菌株MT213进行16S rDNA鉴定，结果显示该乳酸菌16SrDNA序列长度为1492bp如SEQ ID No.1所示。将测序结果与GenBank中的16S rDNA序列进行同源性比对，然后构建***发育树(图3)，以确定菌株的种属关系。同源性分析结果表明，乳酸菌MT213与Lactiplantibacillus plantarum YLL-03和CHOL-200相似度100％，鉴定为植物乳植杆菌，分类命名为植物乳植杆菌MT213。
实施例2植物乳植杆菌MT213在馒头生产中的应用
本实施例用于说明植物乳植杆菌MT213在发酵制备馒头中的应用研究。
1、面团的制备
酵母菌发酵组：小麦粉(100％)、酵母(0.5％)、水(50％)。
乳酸菌+酵母菌发酵组：小麦粉(100％)、酵母(0.5％)、水(50％)、乳酸菌MT213接种量106-109CFU/g。
将上述两组中的各组分混匀，于30℃恒温恒湿培养箱中醒发6h，取两组面团待分析。
2、馒头的制备
酵母菌发酵组：小麦粉(100％)、酵母(0.5％)、水(50％)。
乳酸菌+酵母菌发酵组：小麦粉(100％)、酵母(0.5％)、水(50％)、乳酸菌MT213接种量106-109CFU/g。
将上述两组于30℃恒温恒湿培养箱中醒发6h后取出排气，分割等分面团，搓圆整形。沸水蒸制25min。自然冷却1h后装于自封袋中，放置在25℃培养箱中贮藏。
3、植物乳植杆菌MT213对馒头贮藏期间菌落总数的影响
每2天随机选取3个馒头，用无菌生理盐水(0.85％，w/v)稀释，均质混匀，连续稀释，在30-37℃培养48h后在平板计数培养基(PCA)平板上对细菌计数，28℃培养48h后在孟加拉红板上对真菌计数，见表2。
表2不同处理组馒头贮藏期间菌落总数
结果(表2)显示加入乳酸菌的馒头在贮藏期间菌落总数显著低于酵母组，贮藏第10d时细菌和真菌菌落总数分别为6.79Lg CFU/g、4.63Lg CFU/g，显著低于酵母组，表明植物乳植杆菌MT213能够抑制多种微生物的生长，延长馒头的货架期。
最后应说明的是：以上实施例仅用以说明本发明的技术方案，而非对其限制；尽管参照前述实施例对本发明进行了详细的说明，本领域的普通技术人员应当理解：其依然可以对前述各实施例所记载的技术方案进行修改，或者对其中部分技术特征进行等同替换；而这些修改或者替换，并不使相应技术方案的本质脱离本发明各实施例技术方案的精神和范围。
Claims (10)
1.植物乳植杆菌(Lactiplantibacillus plantarum)MT213，其特征在于，其保藏编号为：CGMCC No.26569。
2.根据权利要求1所述的植物乳植杆菌(Lactiplantibacillus plantarum)MT213，其特征在于，所述植物乳植杆菌(Lactiplantibacillus plantarum)MT213 16S rDNA序列如SEQ ID No.1所示。
3.一种菌剂，其特征在于，含有权利要求1或2所述的植物乳植杆菌(Lactiplantibacillus plantarum)MT213或其发酵液。
4.权利要求1或2所述的植物乳植杆菌(Lactiplantibacillus plantarum)MT213或权利要求3所述的菌剂在抑制致病菌以及腐败菌中的应用；
所描述致病菌以及腐败菌包括革兰氏阳性菌、革兰氏阴性菌或真菌中的一种或多种。
5.根据权利要求4所述的应用，其特征在于，所述革兰氏阳性菌包括金黄色葡萄球菌、单细胞增生李斯特菌、枯草芽孢杆菌、蜡样芽孢杆菌、藤黄微球菌中的一种或多种；和/或，所述革兰氏阴性菌包括大肠杆菌、沙门氏菌、荧光假单胞菌、铜绿假单胞菌中的一种或多种；和/或，所述真菌包括黑曲霉、产黄青霉、扩展青霉、桔青霉、黄曲霉中的一种或多种。
6.权利要求1或2所述的植物乳植杆菌(Lactiplantibacillus plantarum)MT213，或权利要求3所述菌剂在制备功能性产品中的应用；所述功能性产品包括：食品、保健品、或日化用品。
7.一种提高面制品防腐保鲜的方法，其特征在于，包括：在面制品的制备过程中加入权利要求1或2所述的植物乳植杆菌(Lactiplantibacillus plantarum)MT213，或权利要求3所述的菌剂。
8.根据权利要求7所述的方法，其特征在于，所述植物乳植杆菌(Lactiplantibacillusplantarum)MT213的添加量为106-109CFU/g。
9.一种面制品，其特征在于，制备所述面制品时，使用了植物乳植杆菌(Lactiplantibacillus plantarum)MT213。
10.根据权利要求9所述的面制品，其特征在于，所述面制品为发酵面制品，制备所述发酵面制品时，所用发酵剂包括植物乳植杆菌(Lactiplantibacillus plantarum)MT213；
优选地，所述面制品为馒头、面包、蛋糕或饼干。
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