CN110982731A - 一株具有益生特性的太空诱变植物乳杆菌st20-71及其应用 - Google Patents
一株具有益生特性的太空诱变植物乳杆菌st20-71及其应用 Download PDFInfo
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Abstract
本申请涉及一株具有益生特性的太空诱变株植物乳杆菌ST20‑71及其应用,该菌被命名为Lactobacillus plantarum ST20‑71,已于2018年3月16日保藏于中国微生物菌种保藏管理委员会普通微生物菌种保藏中心保藏,保藏号为15456。本申请中的植物乳杆菌L.plantarum ST20‑71在胃肠液中较高的存活率,在人体内具有较强的存活能力。植物乳杆菌L.plantarum ST20‑71具有增强免疫抵抗作用、提高肠道菌群多样性、增加肠道菌群稳定性的益生特点。此外,L.plantarum ST20‑71可以发酵豆乳高产苯乳酸和4‑羟基苯乳酸。
Description
技术领域
本发明涉及微生物技术领域,特别涉及一株经过太空诱变后进行优良菌株选育得到、具有一定益生特性的植物乳杆菌(Lactobacillus plantarum)ST20-71及其作为功能性食品发酵剂和添加剂的应用。
背景技术
乳酸菌(Lactic acid bacteria,LAB)是一群革兰氏阳性、能发酵糖类产乳酸的细菌总称,其在自然界中广泛存在。乳酸菌是人和动物肠道内重要的生理菌群,参与人体的营养消化吸收、脂类和药物代谢、能量转换与供应和免疫调节等,其在人体健康中扮演着重要的角色。乳酸菌具有改善人体肠道菌群、调节机体免疫力等功效。益生菌是指通过摄取适当的量、对食用者的身体健康能发挥有效作用的活性微生物。乳酸菌中的Lactobacillus是最重要的益生菌菌属。根据卫生部印发的《可用于品的菌种名单》(2010版)规定,可以在食品中使用的乳杆菌属主要包括Lactobacillus casei、Lactobacillus plantarum、Lactobacillus acidophilus 和Lactobacillus johnsonii等。
Lactobacillus plantarum的最适生长温度为30-35℃,厌氧或兼性厌氧,呈直杆状或弯杆状,为兼性异型发酵乳酸菌。人类对Lactobacillus plantarum的应用有着悠久的历史。相关研究发现,一些Lactobacillus plantarum能通过胃并可定植于肠道,具有免疫调节、维持肠道平衡、预防心血管疾病等益生作用。植物乳杆菌能够代谢产生苯乳酸和4-羟基苯乳酸,上述物质具有广谱抗菌作用。与此同时,Lactobacillus plantarum对肠道菌群的调节也具有显著作用。近年来,太空诱变技术越来越多的应用于细菌育种研究,可快速产生新的亚群,对微生物生长动力、抗生素敏感、代谢产物的变化以及病原菌的毒力研究具有重大意义。太空诱变主要利用太空环境高辐射、微重力、高真空、弱磁场等特点,以高空气球、返回式卫星、航天飞机等为实验平台,完成对特定物种基因的诱变,具有诱变效率高、周期短,变异方向不确定等特点。我国益生菌菌种资源相对缺乏,经过太空诱变、具有优良益生特性的益生菌更是少之又少。
发明内容
本发明提供一株植物乳杆菌的新菌株(Lactobacillus plantarum)ST20-71及其应用,其特征在于具有增强免疫抵抗作用、提高肠道菌群多样性、增加肠道菌群稳定性等益生特点。
本发明提供一株植物乳杆菌的新菌株(Lactobacillus plantarum)ST20-71已于2018年3月16日在中国微生物菌种保藏管理委员会普通微生物菌种保藏中心保藏,保藏号为CGMCC No.15456,保藏单位地址:北京市朝阳区北辰西路1号院3号。
本发明的另一目的是具有优异的胃肠液耐受性的新型乳杆菌菌株Lactobacillus plantarum ST20-71,具备作为益生菌的基本条件。
本发明的另一目的是提供新型乳杆菌Lactobacillus plantarum ST20-71作为发酵剂或添加剂在酸奶或保健食品中的应用。
本发明的另一目的是提供新型乳杆菌Lactobacillus plantarumST20-71,具有高产苯乳酸以及4-羟基苯乳酸的特性。
本发明的目的是通过如下技术方案实现的:
本发明所涉及的植物乳杆菌(Lactobacillus plantarum)ST20-71、ST20-83 等103株植物乳杆菌是由植物乳杆菌(Lactobacillus plantarum)GT20搭载神州十号飞船(太空飞行15天)和天宫二号飞船(太空飞行60天)中经过太空诱变后进行优良菌株的选育得到,其中植物乳杆菌(Lactobacillus plantarum)ST20-71 具有增强免疫抵抗作用、提高肠道菌群多样性、增加肠道菌群稳定性的益生特点。
本发明涉及的植物乳杆菌ST20-71是通过如下方法分离筛选得到的。
a.菌株活化及单克隆挑取
本发明所涉及的植物乳杆菌(Lactobacillus plantarum)ST20-71、ST20-83等103株植物乳杆菌是由植物乳杆菌(Lactobacillus plantarum)GT20搭载神州十号飞船(太空飞行15天)和天宫二号飞船(太空飞行60天)中经过太空诱变后进行优良菌株的选育得。
将天空突变返回菌种接入到MRS液体培养基,37℃培养24h,继代培养3代,恢复菌株活力。对菌液进行10倍比稀释,得到10-1~10-7稀释梯度。分别吸取 200μL10-4、10-5、10-6、10-7梯度稀释液,均匀涂布于MRS固体培养基平皿内, 37℃厌氧培养48-72h。挑取形态、大小、颜色不同的单克隆接种于液体培养基中,于37℃恒温厌氧培养箱培养24h。待菌株生长良好后,进行革兰氏染色、镜检。
其中MRS培养基的组成是:大豆蛋白胨10g、牛肉膏5g、酵母粉4g、葡萄糖20g、Tween801ml、磷酸二氢钠2g、无水乙酸钠5g、柠檬酸三胺2g、硫酸锰0.02g、硫酸镁0.1g,蒸馏水1L,调节pH至6.2左右,琼脂15g,121℃、 15min灭菌。
b.分子生物学鉴定
将冷冻保存的供试菌株接种于TPY增菌液体培养基中,30℃恒温培养24h,经TPY传代培养2~3代后,取2mL对数生长末期的菌体培养物置于无菌EP管内离心,经8000×g离心3min(4℃)后收集菌体,弃上清,采用乳酸菌专用 CTAB冻融法提取菌株的基因组DNA。
其中,TPY增菌培养液的组成是:乳糖10g、牛肉膏5g、酵母粉5g、酪蛋白胨10g、大豆蛋白胨5g、磷酸氢二钾2.5g、磷酸二氢钾2.5g、硫酸镁0.1g、吐温 800.25g、L-半胱氨酸盐酸盐0.5g、15g琼脂、蒸馏水1L,121℃、15min灭菌。
利用IllumineHiSeq2000测序平台对Lactobacillus plantarum GT20原始菌株和ST20太空诱变复合体以及单克隆株等104株菌进行深度为500X的测序。
104株菌与模式菌株相比共获得20,465个SNP位点,除重复区以外,基因组上每Kb平均约有6.8个SNP位点。用SNP序列构建系统进化树,发现太空诱变菌株ST20-71和ST20-83与其他102株有较大的差异,其系统发育树如图1 所示。
c.形态学特性
本发明的植物乳杆菌L.plantarumST20-71具有如下形态学特征:细胞呈直杆状,有圆形末端,长3.0-8.0μm,宽0.9-1.2μm,呈单个、成对,或短链状出现 (见图2)。
d.菌落形态特性
本发明的植物乳杆菌L.plantarumST20-71在MRS培养基上生长形成乳白色菌落,不透明,圆形,表面光滑,中央凸起,直径约为1.2-1.5mm(见图3)。 e.植物乳杆菌上清液的制备方法
把冻存管中的保藏液接种到5ml的MRS培养基中,震荡混匀,37℃恒温厌氧培养24h,连续活化两代,用活化第二代的植物乳杆菌MRS培养液以(1×109 cfu/ml)以2%的比例(V/V)接种到全脂乳(11wt%全脂乳粉添加到89wt%无菌水中混匀后经巴氏杀菌)中,37℃恒温培养24h,取发酵液以4000×g离心10min,取上清,上清液经0.22μm微膜过滤,-20℃冰箱保存备用。
f.苯乳酸产量
将筛选后得到的2株太空诱变Lactobacillus plantarum ST20-71和Lactobacillus plantarum ST20-83与原始菌株Lactobacillus plantarum GT20在 MRS培养基、豆乳、脱脂乳和全脂乳中发酵24h后测定苯乳酸产生量。选择苯乳酸产量较高菌株为复筛菌株。
g.4-羟基苯乳酸产量
将筛选后得到的2株太空诱变Lactobacillus plantarum ST20-71和Lactobacillus plantarum ST20-83与原始菌株Lactobacillus plantarumGT20在MRS 培养基、豆乳、脱脂乳和全脂乳中发酵24h后测定4-羟基苯乳酸产生量。选择 4-羟基苯乳酸产量较高的菌株为复筛菌株。
h.菌株胃肠液耐受性
模拟胃肠液的制备方法:将PBS灭菌后,用1mol/L HCL调节pH值至2.5,加入3.0mg/ml胃蛋白酶,用0.22μm微孔滤膜过滤除菌,制成模拟人工胃液;将PBS灭菌后,用0.1mol/LNaOH调节pH值至8.0,加入0.1%胰蛋白酶和1.8%牛胆盐,用0.22μm微孔滤膜过滤除菌,制成人工模拟胰液。
胃肠液耐受性:将g中筛选出的复筛菌株活化培养两代,离心洗菌两次,收集菌体,取0.5ml复筛菌体悬液加入到4.5ml pH 2.5模拟人工胃液中,37℃消化 3h,同时分别于0h和3h用MRS琼脂培养基倾注法计数测定活菌数。之后,取 0.5ml已消化3h的人工含菌胃液加入到4.5ml的人工肠液中,继续于37℃水浴培养,分别于4h和8h用MRS琼脂培养基倾注法计数测定活菌数,每个样品做 4个平行。
菌株存活率计算公式
N1――菌株处理后活菌数;
N0――菌株初始活菌数。
本发明的太空诱变植物乳杆菌L.plantarum ST20-71具有优良的胃肠液耐受性,能存活于包括人在内的动物的胃肠器官内,上述特性是作为益生菌的基础。本发明的植物乳杆菌L.plantarum ST20-71是具有益生菌活性的益生菌,当以干燥的细胞形态或发酵产物形态用于人或动物的情况下,能够对宿主的肠道菌群产生有利的影响,具有增强免疫抵抗作用、提高肠道菌群多样性、增加肠道菌群稳定性的益生特点。本发明的植物乳杆菌L.plantarum ST20-71具有产生大量苯乳酸及4-羟基苯乳酸,苯乳酸及4-羟基苯乳酸具有良好的抑菌作用和调节肠道稳态的作用。
本发明提供含有本发明太空诱变植物乳杆菌L.plantarum ST20-71的用于预防和治疗肠道疾病的组合物及其用途,能够用于预防或治疗包括人在内的哺乳动物的肠道疾病,包括牛、马、猪等家畜。肠道危害细菌感染、以及炎症性肠道疾病等都包括在上述“肠道疾病”中,例如,包括由病原性微生物(大肠杆菌、沙门氏菌、梭菌等)引起的感染性腹泻、肠胃炎、炎症性肠道疾病、神经性肠炎综合症、小肠细菌过度生长、急性腹泻等,但是并不限于这些。
上述用于预防和治疗肠道疾病的组合物中含有的植物乳杆菌L.plantarum ST20-71能够以活菌体或死菌体方式存在,但是优选为以活菌体方式存在。一般活菌体具有增强免疫抵抗作用、提高肠道菌群多样性、增加肠道菌群稳定性的效果,用于人以及动物时,能够密集、停留在肠内的消化管道壁上,从而起到使有害菌不能够停留的作用,并且产生乳酸来降低肠内的pH值,抑制有害细菌的繁殖。并且,所用的活菌体生成细菌素和过氧化物,从而能够抑制病原菌的繁殖,对负责吸收营养成分的肠绒毛的活动起到帮助作用。
对于上述本发明的用于预防或治疗肠道疾病的组合物的给药方式没有特别限定,但是优选为经口给药。虽然给药量根据肠道疾病的种类、疾病程度、年龄、性别、人种、治疗或预防目的等有所不同,但是一般情况下以成人为基础。
制备成上述各个剂型时,可以加入各个剂型的制备所需的并在药剂学可接受的载体或添加剂来制备。制备成具有典型的用于经口给药的剂型时,上述载体可以使用选自稀释剂、润滑剂、粘结剂、崩解剂、甜味剂、稳定剂以及防腐剂中的一种或多种,作为添加剂可以使用选自香料、维生素类以及抗氧化剂
i.菌株发酵特性研究
将筛选后得到的2株太空诱变Lactobacillus plantarum ST20-71和Lactobacillus plantarum ST20-83与原始菌株Lactobacillus plantarum GT20分别在豆乳、脱脂乳和全脂乳三种培养基中发酵24h后,进行菌株活菌计数。
j.菌株发酵特性研究
用活化第二代的植物乳杆菌MRS培养液(1×109cfu/ml)0.03wt‰的比例混合后接种到100g的全脂乳(12wt%全脂乳粉、7wt%白砂糖和81wt%无菌水混匀后经匀质和巴氏灭菌)中,并且以未添加植物乳杆菌的普通发酵酸奶为对照,37℃静置培养6h,取发酵液以4000×g离心10min,取上清,上清液经0.22μm微膜过滤,-20℃冰箱保存备用。
把上述植物乳杆菌进行发酵特性的研究,分别测定在脱脂乳中发酵0-24h 时的pH值、滴定酸度(TA),以此来评价和筛选对发酵酸奶的发酵特性有较小影响作用的菌株,从而筛选出适于酸奶发酵的植物乳杆菌。
本发明的植物乳杆菌L.plantarumST 20-71的组合物可以作为食品使用。例如发酵乳制品及其他健康保健食品中,还包括人们每日经常摄取的一般的食品。用于保健功能性食品的情况下,可以和食品学上允许的载体或添加剂一起制备成本技术领域公知的常规保健功能性食品的剂型,作为所述保健功能性食品例如可以制备成散剂、颗粒剂、片剂、胶囊剂、悬浮剂、乳剂、糖浆剂、液体剂、浸膏、茶、果冻或饮料等。上述食品学允许使用的载体或添加剂,根据需要制备的剂型可以选择使用在本技术领域允许使用的公知的任一载体或添加剂。
酸奶或者发酵乳制品是以乳为主要原料,通过添加白砂糖、增稠剂、稳定剂、发酵剂等配料,再经均质、杀菌、冷却、发酵、灌装、冷藏后熟等工艺加工而成的一种发酵型乳制品。常规酸奶的制备过程中,为了使酸奶具有口感细腻、稠厚感适宜、风味优良等特性,通常在制备中添加稳定剂和增稠剂。纵观市场中的酸奶,在其配料表中都可找到稳定剂和增稠剂的身影,典型的稳定剂和增稠剂包括:明胶、果胶、羟丙基二淀粉磷酸酯、琼脂等。
根据本发明的具体实施方式,本发明的发酵乳制品及其他健康保健食品中含有植物乳杆菌ST20-71。本发明的发酵乳制品及其他健康保健食品中含有还包括适量(常规用量)的发酵剂。优选地,所述发酵剂包括保加利亚乳杆菌、嗜热链球菌、嗜酸乳杆菌、双歧杆菌、干酪乳杆菌和鼠李糖乳杆菌等中的一种或几种的组合。当所述发酵剂为上述几种菌种的组合时,在能够顺利进行反应的前提下,本领域一般技术人员可以对各菌种的用量比例进行调配和选择。
根据本发明的具体实施方式,本发明植物乳杆菌L.plantarumST20-71可以与适量其他常规发酵剂复配,用于乳制品及其他健康保健食品的发酵。优选地,所述发酵剂包括保加利亚乳杆菌、嗜热链球菌、嗜酸乳杆菌、双歧杆菌、干酪乳杆菌和鼠李糖乳杆菌等中的一种或几种的组合。当所述发酵剂为上述几种菌种的组合时,在能够顺利进行反应的前提下,本领域一般技术人员可以对各菌种的用量比例进行调配和选择。
上述本发明的含有植物乳杆菌L.plantarumST20-71的组合物还可以作为饲料添加剂或饲料来使用。上述饲料添加剂还可以使用包括选自柠檬酸、富马酸、己二酸、乳酸、苹果酸等有机酸;或膦酸钠、磷酸钾、酸性焦磷酸盐、聚磷酸盐 (聚合磷酸盐)等磷酸盐;或多元酚、儿茶酸、α-生育酚、迷迭香提取物、维生素C、绿茶提取物、甘草提取物、壳聚糖、鞣酸、植酸等天然抗氧化剂中的一种或多种。作为饲料使用的情况下,可以将上述组合物制备成常规饲料形态,可以包括常规饲料成分。
本发明的植物乳杆菌L.plantarum ST20-71及与之相关的技术方案带来如下有益效果:
(1)植物乳杆菌L.plantarumST20-71在模拟胃肠液中有较高的存活率;
(2)植物乳杆菌L.plantarumST20-71具有增强免疫抵抗作用、提高肠道菌群多样性、增加肠道菌群稳定性的益生特点;
(3)植物乳杆菌L.plantarumST20-71在豆乳中有较高活菌数;
(4)植物乳杆菌L.plantarumST20-71可以产生大量苯乳酸和4-羟基苯乳酸。
保藏信息:
本发明提供的植物乳杆菌(Lactobacillus plantarum)菌株ST20-71,其生物学名称为Lactobacillus plantarum,于2018年3月16日在中国微生物菌种保藏管理委员会普通微生物菌种保藏中心保藏,保藏号为15456。
附图说明
图1:L.plantarumST20复合体系统发育树
图2:L.plantarumST20-71菌落形态图片
图3:L.plantarumST20-71菌体显微镜图片(X1000)
图4:L.plantarumST20-71对人体抗炎因子和免疫球蛋白的影响
图5:L.plantarumST20-71对肠道菌群α多样性的影响
图6:L.plantarum ST20-71对肠道菌群β多样性的影响
图7:L.plantarum ST20-71的苯乳酸产量测定
图8:L.plantarum ST20-71的4-羟基苯乳酸产量测定
具体实施方式
下面结合附图和实施例对本发明作进一步说明,以下实施例仅为说明性的,本发明并不受这些实例的限制。
本发明中的试验菌株:所用植物乳杆菌L.plantarum ST20-71和L.plantarumST20-83等103株植物乳杆菌,是由植物乳杆菌GT20搭载神州十号飞船(太空飞行15天)和天宫二号飞船(太空飞行60天)经过太空诱变后进行优良菌株选育后得到。
实施例1:L.plantarumST20-71胃肠液耐受性
将PBS灭菌后,用1mol/L HCL调节pH值至2.5,加入3.0mg/ml胃蛋白酶,用0.22μm微孔滤膜过滤除菌,制成模拟人工胃液;将PBS灭菌后,用0.1mol/L NaOH调节pH值至8.0,加入0.1%胰蛋白酶和1.8%牛胆汁,用0.22μm微孔滤膜过滤除菌,制成人工模拟胰液。
将复筛菌株活化培养两代,离心洗菌两次,收集菌体,取0.5ml复筛菌液加入到4.5ml的pH 2.5模拟人工胃液中,37℃消化3h,同时分别于0h和3h用 MRS琼脂培养基倾注法计数测定活菌数。之后,取0.5ml已消化3h的人工含菌胃液加入到4.5ml的人工肠液中,继续于37℃水浴培养,分别于4h和8h用 MRS琼脂培养基倾注法计数测定活菌数。
菌株存活率计算公式
N1――菌株处理后活菌数;
N0――菌株初始活菌数。
L.plantarum ST20-71、L.plantarum ST20-83和GT20对胃肠液中的存活率见表1,在模拟胃液(pH2.5)条件下存活3h后的活菌数呈现下降的趋势,对模拟胃液有一定的耐受性。L.plantarumST20-71在pH8.0的人工肠液中该菌株存活率下降缓慢,4h时的存活率为91.64±0.02%,保持至8h时该菌株存活率为 92.87±0.71%,表明太空诱变菌株L.plantarum ST20-71有较好的胃肠液耐受性。
表1 人工胃肠液中菌株存活率(%)
实施例2:L.plantarumST20-71的益生作用
1.研究对象
选取符合诊断和排除标准的65例讲课志愿者作为研究对象,性别不限。其中青年组(年龄20-50岁)35例,老年组(年龄大于等于50岁)。排除标准:伴有消化道病变、心肝肾功能不全、恶性肿瘤、糖尿病、既往有腹部手术史者、妊娠或哺乳期妇女、试验前4周内曾服用影响本试验观察的药物(抗生素、微生态制剂)、自身免疫性疾病及不能按期复诊及随访者。
2.实验方法
将65例健康志愿者随机分为对照组和试验组,实验组将“益适优”菌粉溶于温水或牛奶中(1袋口服1/日饭后),而对照组将安慰剂溶于温水或牛奶中(1 袋口服1/日饭后),连续服用28天。
3.观察指标
(1)血液指标测定:对志愿者的IgG和IL4的含量进行测定。
(2)肠道菌群测定:对65例健康志愿者的粪便样品进行PacBio SMRT三代测序,分析其菌群构成并计算其α和β多样性。
4.观测方法
比较两组不同方法在血液指标和肠道菌群方面差异。
5.治疗结果
对照组和实验组在人体抗炎因子和免疫球蛋白的差异如图所示。
IgG是血液和细胞液中的主要抗体类型,由B细胞产生和分泌,与多种致病菌结合发挥抗感染的作用。ST20-71干预显著增加了中青年组血液中IgG水平 (p<0.01),表明ST20-71可以增加中青年志愿者的免疫抵抗作用。IL-4是属于抗炎因子,介导CD4T辅助2(Th2)细胞的活化,进而调节Th1/Th2细胞的免疫平衡。ST20-71干预显著增加了血液中抗炎因子IL-4水平(p<0.05)。
菌群α多样性显示,安慰剂组菌群丰度和多样性没有明显变化,而服用 ST20-71组的菌群丰度和多样性持续升高。丰富的肠道菌群丰度和多样性是人体健康的重要标志。显示出ST20-71具有提高肠道菌群多样性的益生功效。
如图所示为健康志愿者肠道菌群β多样性差异比较。摄入ST20后,志愿者肠道菌群结构没有发生根本性的改变,个体差异是影响肠道菌群的重要因素之一。在28天时间内,摄入ST20后志愿者肠道菌群点更为接近,而对照组没有发生大的变化。上述结果表明摄入ST20可在一定程度上增加肠道菌群稳定性。
综上所述,植物乳杆菌ST20-71(Lactobacillus plantarumST20-71)具有增强免疫抵抗作用、提高肠道菌群多样性、增加肠道菌群稳定性的益生特点。
实施例3:L.plantarumST20-71在豆乳、脱脂乳和全脂乳中发酵计数
将筛选后得到的Lactobacillus plantarumST20-71、ST20-83与诱变原始菌株GT20分别在豆乳、脱脂乳和全脂乳三种培养基中发酵24h后,进行菌株活菌计数。结果如表2所示,结果显示原始菌株GT20和太空诱变株ST20-83在脱脂乳中的活菌数显著高于全脂乳和豆乳中活菌数,豆乳中的活菌数最低。而太空诱变株ST20-71在豆乳中活菌数显著高于脱脂乳和全脂乳,表明突变株ST20-71更适合豆乳环境中生长。
表2 菌株活菌计数结果(log10cfu/mL)
实施例4:L.plantarumST20-71发酵实验
将筛选后得到的太空诱变Lactobacillus plantarum ST20-71、ST20-83与原始菌株GT20分别进行发酵实验,3株Lactobacillus plantarum在发酵10h后的pH 值和滴定酸度存在差异,突变株ST20-71产酸速率较快(3.56Δ°T/h),但差异均不显著。
表3 L.plantarum发酵的pH和滴定酸度的测定
实施例5:L.plantarumST20-71筛选株苯乳酸产量测定
将筛选后得到的Lactobacillus plantarumST20-71、ST20-83与诱变原始菌株GT20在MRS培养基、豆乳、脱脂乳和全脂乳中发酵24h后测定苯乳酸产生量,结果发现突变株ST20-71发酵四种培养基产生苯乳酸量显著高于GT20和 ST20-83。除此之外,GT20和ST20-83发酵豆乳的苯乳酸产生量高于发酵全脂乳和脱脂乳,ST20-71发酵豆乳中产苯乳酸的量少于发酵全脂乳和脱脂乳。如图7 所示,3株Lactobacillus plantarum在MRS培养基中的苯乳酸产生量最大,其中突变株ST20-71发酵产生苯乳酸量最高为16.46mg/L。
实施例6:L.plantarum筛选株4-羟基苯乳酸产量测定
3株Lactobacillus plantarum在四种同样培养环境下4-羟基苯乳酸产生量分析发现,发酵豆乳4-羟基苯乳酸产生量在0.05-0.08mg/L范围内,菌株间无显著性差异。如图8所示GT20和ST20-83发酵四种培养基4-羟基苯乳酸产生量在 0.06-0.10mg/L范围内,菌株间无显著性差异。除发酵豆乳外,突变株ST20-71 发酵全脂乳、脱脂乳以及MRS中4-羟基苯乳酸产生量显著高于GT20和ST20-83 菌株,最高产量为脱脂乳中的1.17mg/L。
Claims (3)
1.一株太空诱变的植物乳杆菌(Lactobacillusplantarum)ST20-71,该菌株于2018年3月16日保藏于中国微生物菌种保藏管理委员会普通微生物菌种保藏中心保藏,保藏号为15456。
2.权利要求1所述的植物乳杆菌,其特征在于该菌株应用于功能性食品中的应用,其特征在于具有增强免疫抵抗作用、提高肠道菌群多样性、增加肠道菌群稳定性的益生特点。
3.权利要求1所述的植物乳杆菌,其特征在于作为发酵剂或食品添加剂在普通食品中的应用。
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Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN111996146A (zh) * | 2020-08-31 | 2020-11-27 | 山东宝来利来生物工程股份有限公司 | 一株高产苯乳酸的植物乳杆菌及其应用 |
CN116694503A (zh) * | 2023-03-27 | 2023-09-05 | 上海华聿康生物科技有限公司 | 一种具有润肠通便和提高免疫功能的植物乳杆菌Lp-HZ55 |
Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN102732447A (zh) * | 2012-04-18 | 2012-10-17 | 北京和美科盛生物技术有限公司 | 一株抑制面包中霉菌和酵母菌的植物乳杆菌及其使用方法 |
CN102864096A (zh) * | 2012-04-18 | 2013-01-09 | 北京和美科盛生物技术有限公司 | 一株植物乳杆菌及其高密度培养和冻干菌粉制备的方法 |
CN104830731A (zh) * | 2015-05-08 | 2015-08-12 | 内蒙古农业大学 | 一株具有广谱抑菌特性的植物乳杆菌ab-1及其应用 |
CN108004189A (zh) * | 2018-01-18 | 2018-05-08 | 北京科拓恒通生物技术股份有限公司 | 一种复合益生乳酸菌粉剂及其制备方法与应用 |
-
2019
- 2019-10-28 CN CN201911027909.5A patent/CN110982731A/zh active Pending
Patent Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN102732447A (zh) * | 2012-04-18 | 2012-10-17 | 北京和美科盛生物技术有限公司 | 一株抑制面包中霉菌和酵母菌的植物乳杆菌及其使用方法 |
CN102864096A (zh) * | 2012-04-18 | 2013-01-09 | 北京和美科盛生物技术有限公司 | 一株植物乳杆菌及其高密度培养和冻干菌粉制备的方法 |
CN104830731A (zh) * | 2015-05-08 | 2015-08-12 | 内蒙古农业大学 | 一株具有广谱抑菌特性的植物乳杆菌ab-1及其应用 |
CN108004189A (zh) * | 2018-01-18 | 2018-05-08 | 北京科拓恒通生物技术股份有限公司 | 一种复合益生乳酸菌粉剂及其制备方法与应用 |
Non-Patent Citations (1)
Title |
---|
任发政 吴剑涛主编: "《健康好搭档:酸奶与益生菌》", 31 May 2008, 中国商业出版社 * |
Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN111996146A (zh) * | 2020-08-31 | 2020-11-27 | 山东宝来利来生物工程股份有限公司 | 一株高产苯乳酸的植物乳杆菌及其应用 |
CN116694503A (zh) * | 2023-03-27 | 2023-09-05 | 上海华聿康生物科技有限公司 | 一种具有润肠通便和提高免疫功能的植物乳杆菌Lp-HZ55 |
CN116694503B (zh) * | 2023-03-27 | 2024-01-05 | 上海华聿康生物科技有限公司 | 一种具有润肠通便和提高免疫功能的植物乳杆菌Lp-HZ55 |
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