CN116948861A - 具有抗衰功能的菌株及其应用 - Google Patents
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Abstract
本发明公开了一种具有抗衰老功能的菌株,经鉴定为鼠李糖乳酪杆菌(Lacticaseibacillus rhamnosus)LC‑9,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,菌种保藏登记号为CGMCC No.26820,保藏时间为2023年3月17日。本发明提供的抗衰功能的菌株,该菌株的DPPH清除率较高,其在抗衰产品方面的应用有较大的前景。本发明将得到的菌株进行了实际应用,其发酵酸奶的效果较好,用其发酵得到的酸奶,在抗衰功能方面有较好的效果。同时,以该菌株为主要原料制备成其他食品,也有较好的抗衰效果。
Description
技术领域
本发明涉及生物工程技术领域,具体是一种具有抗衰老功能的菌株及其应用。
背景技术
近年来,国内外针对乳酸菌功能的研究不断增加,乳酸菌功能也不断细化,乳酸菌的新功能也被不断发掘;在研究中发现某些益生菌具有抗机体氧化应激的作用,被临床用于治疗与肠道相关的疾病。乳酸菌主要通过以下几种方式发挥抗氧化应激作用:清除体内自由基及活性氧、抑制脂质过氧化及螯合金属离子、抗氧化防御系统、调控与氧化应激相关的信号通路。乳酸菌菌体及其代谢产物都具有抗氧化的作用,而且不同菌株抗氧化的活性成分及能力均不同。
田园等采用蛋白质组学的方法深入分析了菌株的抗氧化机制。通过过氧化氢筛选,获得的植物乳杆菌KM1在5.0mmol/L的过氧化氢中存活率为78.26%,发现该菌株具有良好的肠道定植能力。蛋白质组学的结果显示共有112个差异表达蛋白,其中31个差异表达蛋白上调,81个下调。GO功能富集分析表明90%的差异表达蛋白质参与糖酵解等代谢过程,并且有14个差异表达蛋白与抗氧化活性有关,包括磷酸甘油激酶(PGK),α-甘油磷酸氧化酶(GlpO),丙酮酸氧化酶和NADH过氧化物酶等蛋白质。
益生菌中的乳酸菌在肠道中可产生一种四聚酸,可杀死大批有害的、具有抗药性的细菌。乳酸菌体抗原及其代谢物还可以通过刺激肠粘膜淋巴结,激发免疫活性细胞,产生特异性抗体和致敏淋巴细胞,激发机体的免疫应答,防止病原菌侵入和繁殖,还可以激活巨噬细胞,加强和促进吞噬作用,产生干扰素,加快细胞新陈代谢,增强机体免疫力。
益生菌能够提高血液中超氧化物歧化酶的含量和生物活性,而超氧化物歧化酶能够清除自由基,降低氧化还原电位,增强机体的抗氧化能力;益生菌代谢过程产生的酸性物质可大大提高肠道内的酸度,如甲酸、乙酸、乳酸等,阻碍了有害菌的生长和繁殖,抑制有毒有害物质的形成与吸收,减轻肝脏排毒负担,促进机体健康,延缓衰老。
目前国内的抗衰食品,主要以“妆食同源”概念为销售卖点,产品多为胶原蛋白、透明质酸、花青素和抗氧化成分,虽然从一定程度上能够缓解人体的衰老速度,但是产品整体价格偏高,功能单一,缺少对人体机能的反馈提升,不能有效提高人体自身对衰老的抵抗能力。所以抗衰的本质是提高人体对衰老的抵抗能力,因此开发一种功能全面的抗衰产品显得尤为重要,本产品具备提高人体的免疫能力,提高人体对环境风险的抵抗力;提供功能性成分,补充人体功能性成分的流失;清除体内自由基,减轻自由基对身体的伤害。
发明内容
本发明所要解决的技术问题是:提供一株具有抗衰老功能的菌株,并鉴定其菌落形态学特征和生理与生化特性,以及所筛选得到的抗衰功能菌株的应用。
为解决上述技术问题,本发明采用的技术方案如下:
(一)抗衰功能菌株的筛选。
本发明所提供的筛选菌株的方法就是进行高通量筛选,以耐受过氧化氢并能清除自由基为目标。通过对不同生境的菌种进行分离,获得乳酸菌,然后对乳酸菌的耐过氧化氢能力,生长能力、产酸能力、清除自由基能力等进行评价,获得具有抗衰能力的乳酸菌。
本发明使用的第一步筛选培养基为改良MRS培养基。在灭菌结束后,在MRS培养基中加入无菌过氧化氢,使其在培养基中的浓度为0.5%-1%,无菌接入天然发酵产物后,进行厌氧培养,之后,对存活的微生物菌群进行单菌落平板分离。
平板分离培养基采用MRS固体培养基:蛋白胨5g,牛肉膏5g,酵母粉5g,胰蛋白胨5g,葡萄糖20g,Tween 80 1g,硫酸锰 0.25g,乙酸钠5g,柠檬酸氢二铵2g,硫酸镁0.58g,3水磷酸氢二钾2g,碳酸钙20g,琼脂20g。纯化水定至1000ml,pH7.0。
挑取单菌落后,使用MRS培养基进行纯培养,之后检测发酵液的DPPH清除率。
(二)对所得到的具有抗衰功能的菌株进行生理特性的研究
通过本方法,得到了一株抗衰功能较强的乳酸菌,在实验室进行菌株的基本鉴定为乳酸菌,再经过中国工业微生物菌种保藏管理中心(CICC)进一步鉴定为鼠李糖乳酪杆菌(Lacticaseibacillus rhamnosus)LC-9,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,菌种保藏登记号为CGMCC No. 26820,保藏时间为2023年3月17日,保藏地址为北京市朝阳区北辰西路1号院3号。
鼠李糖乳酪杆菌( Lacticaseibacillus rhamnosus)LC-9菌株具有下述性质:
1、菌落形态学特征:
本发明的鼠李糖乳酪杆菌( Lacticaseibacillus rhamnosus)LC-9,在MRS培养基中,37±1℃厌氧培养24小时,菌体呈短杆状,无鞭毛,不运动,革兰氏染色阳性。在营养琼脂培养基上,37±1℃℃厌氧培养24小时,菌落白色,圆形,表面湿润,不透明。
2、生理与生化特性:
a.培养温度: 37±1℃,最适温度为37℃;
b.在pH 7.0±1范围内生长;
3、营养特征:
鼠李糖乳酪杆菌( Lacticaseibacillus rhamnosus)LC-9可在多种培养基上生长,培养基成分明确且价格低廉。该培养基碳源一般为乳糖或葡萄糖或麦芽糖,氮源可选用蛋白胨,酵母粉,玉米浆等,培养基中还包括钠盐、钙盐、镁盐和盐酸盐等常用的无机盐类。
(三)所筛选到的鼠李糖乳酪杆菌( Lacticaseibacillus rhamnosus)LC-9的应用
将鼠李糖乳酪杆菌( Lacticaseibacillus rhamnosus)LC-9进行扩大培养后,接种入灭菌的纯牛奶中,37±1℃静置发酵10±6小时,此明呈凝乳状态,符合发酵乳饮料的特征,4℃冷藏后饮用,口感较好。对发酵乳进行DPPH清除率检测,检测其DPPH清除率,其DPPH清除率较高。
DPPH清除率的检测方法:
用乙醇配制不同浓度的样品稀释液和0.04mg/mL的DPPH溶液,向2.0mL样品溶液中加入2.0mL的DPPH溶液,充分混合,室温避光反应30min后测定该混合溶液在517nm处的吸光值A1;以等体积的无水乙醇代替样品稀释液,同法操作,测定吸光值为A0;以等体积的无水乙醇代替DPPH乙醇溶液,加入样品稀释液同样操作。测定吸光值为A2。以VC作阳性对照,做三次平行实验,按下面公式计算清除率。
清除率(%)=(1- (A1- A2)/ A0)*100。
(四)一种抗衰酸奶的制备方法和应用
本发明利用上述抗衰老菌株鼠李糖乳酪杆菌LC-9,制备了一种抗衰老酸奶,提供的抗衰酸奶包括以下重量份原料:全脂奶粉8~15份、蔗糖5~10份、灰树花多糖0.1~0.5份、透明质酸0.05~0.1份、蓝莓花青素0.03~0.1份、低聚果糖0.3~0.8份、胶原蛋白肽 0.05~0.1份、分离乳清蛋白粉0.3~0.8份、γ-氨基丁酸0.3~0.8份、酸奶发酵剂3~10份、其余为水。
所述酸奶发酵剂为副干酪乳酪杆菌LC-6,鼠李糖乳酪杆菌LC-9,植物乳植物杆菌LP-28,鼠李糖乳酪杆菌 LP-45,嗜热链球菌ST-23。
抗衰酸奶的制备方法及应用,包括以下步骤:
(1)原料称取
称取全脂乳粉、蔗糖、灰树花多糖、透明质酸、蓝莓花青素、低聚果糖、胶原蛋白肽、分离乳清蛋白粉、γ-氨基丁酸、稳定剂、甜味剂等。
(2)物料水溶
准备55-60℃的配料水,在搅拌状态下,将物料投入并完全溶解后,静置恒温45-55℃水合30分钟。
(3)乳液均质
将水合奶粉定容到设定体积,将其升温到55-60℃,将其全部打入均质机,均质压力为18-20Mpa。
(4)杀菌
杀菌条件选择93-98℃,320s;杀菌完成后迅速降温到培养温度43℃。
(5)接种、发酵
将活化好的菌种按照5%的接种量进行接种,先在搅拌状态下发酵1h,后进行静置发酵。
(6)发酵产品
发酵中止后,迅速降温到20℃,后灌装,之后进入冷库冷藏。
本发明的有益效果:
本发明的主要目的是获得具有清除自由基的功能乳酸菌,产品搭配胶原蛋白、透明质酸、多糖等功能成分,以乳酸菌饮品为媒介,具有提高免疫力、清除自由基,延缓衰老等功能,以理化指标检测、抗衰细胞模型等手段,确定最佳配方,实现产品功效最大化,综上所述,本研究可为功能性抗衰食品提供一种可行的方法依据。
本发明采用酸奶进行抗衰产品制作,成本较低,经实验证明具有明显的抗衰老效果,能够增强免疫力。动物实验证明,通过小鼠试验,发现液体药物对衰老小鼠脾脏中的NK细胞活性有一定的增强效果,从而增强衰老小鼠免疫系统活性,同时液体药物一定程度上能使脑组织中SOD、GSH-PX含量升高,以及脑组织中MDA含量降低,对脑组织的衰老具有一定延缓的作用。
附图说明
图1是47个菌株DPPH清除率。
图2是菌株9和菌株45的生长情况。
图3鼠李糖乳酪杆菌( Lacticaseibacillus rhamnosus)LC-9的显微观察图片。
图4鼠李糖乳酪杆菌( Lacticaseibacillus rhamnosus)LC-9的平板菌落图片。
图5是雌雄各组小鼠脾脏指数,其中 A:不分雌雄各组小鼠脾脏指数;B:分雌雄各组小鼠脾脏指数。
图6是各组脑组织中SOD、GSH-Px、MDA含量水平检测与NK细胞活性检测(与对照组比较,*P<0.05)
实施方式
根据下述实施例,可以更好地理解本发明。然而,本领域的技术人员容易理解,实施例所描述的仅用于说明本发明,而不应当也不会限制权利要求书中所详细描述的本发明。
实施例
抗衰功能菌株的筛选方法
1)改良MRS培养基的制备。
准确称取MRS培养基(海博生物)66.2g,加热溶解于 1000ml 纯化水中,115℃高压灭菌 15 分钟,灭菌结束后,冷却备用。
使用无菌滤器过滤30%过氧化氢至无菌容器中,按照0.8%的终浓度计算,1000mlMRS培养基中加入26.7ml无菌35%过氧化氢。
2)从泡菜中筛选菌种。
①取多组生长旺盛的泡菜原液,分别编号为a,b,c,d,e。用移液枪吸取1ml转移至改良MRS培养基中,43℃,震荡厌氧培养24小时。
②制备平板筛选培养基,准确称取蛋白胨5g,牛肉膏5g,酵母粉5g,胰蛋白胨5g,葡萄糖20g,Tween 80 1g,硫酸锰 0.25g,乙酸钠5g,柠檬酸氢二铵2g,硫酸镁0.58g,3水磷酸氢二钾2g,碳酸钙20g,琼脂20g于烧杯中,加水1000ml混匀,调pH至中性,115℃灭菌20min。灭菌结束后,倒平板冷却备用。
③取各组发酵液进行梯度稀释,涂平板筛选培养基,37℃恒温培养24小时,挑取其中有透明圈的单菌落,进行保存,按顺序进行编号1-47。
④将初步筛选获得的47个菌株,接种入MRS培养基,37℃恒温培养24小时,检测其DPPH清除率。其结果见图1。
⑤对清除率>80%的菌株进行初步的鉴定,其中4,17号非乳酸菌,舍弃。根据生长情况(OD600),产酸情况,综合比较,本发明选择了菌株9作为以后的生产菌株,结果见图2。
实施例
①菌株的显微观察
从附图3,鼠李糖乳酪杆菌( Lacticaseibacillus rhamnosus)LC-9的显微图片上可以看出,鼠李糖乳酪杆菌( Lacticaseibacillus rhamnosus)LC-9的菌体呈短杆状,无鞭毛,不运动。
②菌株的平板观察
从附图4,鼠李糖乳酪杆菌( Lacticaseibacillus rhamnosus)LC-9,在营养琼脂培养基上,37±1℃℃厌氧培养24小时,菌落白色,圆形,表面湿润,不透明。
③菌株的革兰氏染色
取少量菌体,进行革兰氏染色,鉴别鼠李糖乳酪杆菌( Lacticaseibacillusrhamnosus)LC-9为革兰氏阳性菌。
实施例
鼠李糖乳酪杆菌( Lacticaseibacillus rhamnosus)LC-9的应用
①菌体培养
将菌株鼠李糖乳酪杆菌( Lacticaseibacillus rhamnosus)LC-9接种于灭菌后的MRS培养基中,37℃,摇床震荡培养12小时,之后将种子液进行无菌离心,6000g离心5min,得到菌泥。用无菌生理盐水洗涤2次,得到纯菌体。纯菌体用无菌纯化水进行重悬,此菌悬液为培养菌种。
②酸奶培养
将纯牛乳与稳定剂,甜味剂等进行混合后灭菌,接入菌悬液,在37℃恒温放置8-12小时后,等牛奶凝乳后,将酸奶于冰箱中静置4-8小时,进行后熟,将此方法得到的发酵乳进行DPPH清除率的检测,其清除率达80%以上。
本实施例中的抗衰酸奶,按下述配方称取各重量份原料:
为实现上述目的,所述抗衰酸奶包括以下重量份原料:全脂奶粉10份、蔗糖7份、灰树花多糖0.3份、透明质酸0.05份、蓝莓花青素0.05份、低聚果糖0.5份、胶原蛋白肽 0.08份、分离乳清蛋白粉0.5份、γ-氨基丁酸0.5份、酸奶发酵剂7份、其余为水。
所述酸奶发酵剂为干酪乳杆菌LC-6,干酪乳杆菌LC-9,植物乳杆菌 LP-28,植物乳杆菌 LP-45,嗜热链球菌ST-23。
抗衰酸奶的制备方法及应用,包括以下步骤:
(1)原料称取
称取全脂乳粉、蔗糖、灰树花多糖、透明质酸、蓝莓花青素、低聚果糖、胶原蛋白肽、分离乳清蛋白粉、γ-氨基丁酸、稳定剂、甜味剂。
(2)物料水溶
准备55-60℃的配料水,在搅拌状态下,将物料投入并完全溶解后,静置恒温45-55℃水合30分钟。
(3)乳液均质
将水合奶粉定容到设定体积,将其升温到55-60℃,将其全部打入均质机,均质压力为18-20Mpa。
(4)杀菌
杀菌条件选择93-98℃,320s;杀菌完成后迅速降温到培养温度43℃。
(5)接种、发酵
将活化好的菌种按照5%的接种量进行接种,先在搅拌状态下发酵1h,后进行静置发酵。
(6)发酵产品
发酵中止后,迅速降温到20℃,后灌装,之后进入冷库冷藏。
抗衰酸奶,配方包括,按下述配方称取各重量份原料:全脂奶粉8份、蔗糖8份、灰树花多糖0.5份、透明质酸0.06份、蓝莓花青素0.04份、低聚果糖0.6份、胶原蛋白肽 0.05份、分离乳清蛋白粉0.3份、γ-氨基丁酸0.3份、酸奶发酵剂5份、其余为水。
抗衰酸奶,配方包括,按下述配方称取各重量份原料:全脂奶粉12份、蔗糖8份、灰树花多糖0.8份、透明质酸0.08份、蓝莓花青素0.06份、低聚果糖0.8份、胶原蛋白肽 0.03份、分离乳清蛋白粉0.6份、γ-氨基丁酸0.5份、酸奶发酵剂8份、其余为水。
抗衰酸奶,配方包括,按下述配方称取各重量份原料:全脂奶粉9份、蔗糖6份、灰树花多糖0.5份、透明质酸0.05份、蓝莓花青素0.03份、低聚果糖0.5份、胶原蛋白肽 0.04份、分离乳清蛋白粉0.8份、γ-氨基丁酸0.4份、酸奶发酵剂5份、其余为水。
抗衰酸奶,,配方包括,按下述配方称取各重量份原料:全脂奶粉15份、蔗糖10份、灰树花多糖0.6份、透明质酸0.03份、蓝莓花青素0.05份、低聚果糖0.5份、胶原蛋白肽 0.04份、分离乳清蛋白粉0.4份、γ-氨基丁酸0.3份、酸奶发酵剂10份、其余为水。
为了评估抗衰酸奶的应用效果,将18只18~20g的KM小鼠随机分成3组,每组6只;3组分别为:
正常对照组;
衰老模型组;
衰老模型和抗衰酸奶,其采用实施例4-3制备的抗衰酸奶;
1)动物分组及造模 老鼠随机分为3组,正常对照组(n=6),雌雄各半,模型组(n=6),雌雄各半,模型+给药组(n=6),雌雄各半,动物适应性饲养7天后,对模型组和模型+药物组,每天颈背部皮下注射D-半乳糖溶液,0.1ml/10g,对对照组注射同等生理盐水,连续42天。在造模的同时给予模型+药物组小鼠灌胃给药处理,每天颈背部皮下注射D-半乳糖溶液0.1ml/10g,同时对模型+药物组进行灌胃液体药物0.52ml/10g/d,对对照组注射同等生理盐水。
(2)观察指标 在末次灌胃结束24小时后将各组小鼠称重后处死,取大脑组织,进行ELISA检测,检测各组脑组织的超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)、丙二醛(MDA);取脾脏称重,计算脏器比(脏器比=脏器重量/动物体重*100);分离脾脏淋巴细胞,做NK细胞活性检测。
(3) ELISA检测
1)所有试剂和组分都先恢复到室温,标准品和样品,做复孔。
2)配制好试剂盒各种组分的工作液,可按试剂盒说明书中试剂准备中描述的方法操作。
3)从铝箔袋中取出所需板条,剩余的板条用自封袋密封放回冰箱保存。
4)标准品的加样:设置标准品孔和样本孔,标准品孔各加不同浓度的标准品 50μL。
5)加样:分别设空白孔(空白对照孔不加样品及酶标试剂,其余各步操作相同)、待测样品孔。在酶标包被板上待测样品孔中加样品50μL,加样将样品加于酶标板孔底部,尽量不触及孔壁,轻轻晃动混匀。
6)加酶:每孔加入酶标试剂 100μL,空白孔除外。
7)温育:用封板膜封板后置 37 ℃温育 60 min。
8)配液:将 20 倍浓缩洗涤液用蒸馏水 20 倍稀释后备用。
9)洗涤:小心揭掉封板膜,弃去液体,甩干,每孔加满洗涤液,静置 30 s后弃去,如此重复 5 次,拍干。
10)显色:每孔先加入显色剂A 50μL,再加入显色剂B 50μL,轻轻震荡混匀,37 ℃避光显色 15 min。
11)终止:每孔加终止液 50μL,终止反应(此时蓝色立转黄色)。
12) 测定:以空白孔调零,450 nm 波长依序测量各孔的吸光度(OD 值)。测定应在加终止液后 15 min以内进行。
(4)脾脏淋巴细胞悬液制备
取下的脾脏组织用PBS清洗1~3次,清洗过后,将脾脏组织置于60mm培养皿中,加入适量的PBS,用10mL无菌注射器内芯轻轻研磨脾脏(力度适中),用70uM滤网过滤,收集细胞悬液于10mL离心管中,1500rpm 5min离心弃上清;在细胞中加入适量的红细胞裂解液,450×g离心10分钟,弃上清,细胞加入1640完全培养基进行重悬,细胞进行流式计数,将细胞浓度调整为5×106 个/mL备用。
(5) NK细胞活性检测
实验前 24h 先将 K562细胞(靶细胞)传代培养,收集细胞后用不含胎牛血清的RPM1640 培养液洗涤三次,计数后将细胞浓度调整为1x105个/ml,取靶细胞(K562细胞)和效应细胞(脾细胞)各100μL (效应细胞:靶细胞=50:1),加入96孔培养板中,同时设靶细胞最大释放孔即每孔加入靶细胞 K562 和1%NP40各100ul,靶细胞自然释放孔即每孔各加入靶细胞K562 和RPMI1640培养液100ul,所有样本均设三个复孔,置细胞培养板于37℃、5%CO2培养箱中孵育22h后,每孔各吸取上清100pl,按乳酸脱氢酶 ELISA 试剂盒说明书操作,检测结果经酶联仪450nm 波长测定反应孔、自然释放孔、最大释放孔的OD值,按以下公式计算 NK 细胞的杀伤活性:NK细胞活性=(反应孔OD-自然释放孔OD) /(最大释放孔 OD-自然释放孔OD)×100%
统计方法 应用SPSS 20.0软件进行统计分析。所有实验重复3次,定量结果采用均数±标准差(X±S)表示。两组之间定量数值比较采用独立样本T检验,多组之间定量数值比较采用单因素方差分析,两两比较采用S-N-K法。检验水准α=0.05。
实验结果
与对照组相比,模型组和模型药物组的小鼠脾脏指数有所增加;与模型组相比,模型加药组小鼠脾脏比无明显变化。与模型组相比,模型药物组小鼠脑组织中的SOD、GSH-Px含量呈升高趋势,MDA含量呈下降趋势,NK细胞活性呈升高趋势。
1)液体药物对小鼠免疫器官的影响
如图5左侧的A所示,与对照组相比,脾脏指数都略微增加,无显著差异;与模型
组相比,模型药物组小鼠脾脏指数无明显变化;如图5右侧的B所示,雄性小鼠与雌性小鼠的脾脏指数也有所不同,与对照组相比,模型组与模型药物组的小鼠脾脏指数略微增加,无显著差异。
ELISA检测结果:
图6:各组脑组织中SOD、GSH-Px、MDA含量水平检测与NK细胞活性检测(与对照组比较,*P<0.05)
如图6所示,与对照组相比,脑组织中的SOD含量呈下降趋势,无显著性差异,MDA含量明显增加,差异具有显著性(p<0.05),模型组中GSH-Px含量显著下降,差异具有显著性(p<0.05),模型组中小鼠NK细胞活性呈下降趋势,无显著性差异。与模型组相比,模型药物组脑组织中的SOD、GSH-Px含量呈升高趋势,MDA含量呈下降趋势,NK细胞活性呈升高趋势。
当然,上述说明并非是对本发明的限制,本发明也并不限于上述举例,本技术领域的普通技术人员,在本发明的实质范围内,作出的变化、改型、添加或替换,都应属于本发明的保护范围。
Claims (10)
1. 一株具有抗衰老功能的菌株,经鉴定为鼠李糖乳酪杆菌( Lacticaseibacillus rhamnosus)LC-9,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,菌种保藏登记号为CGMCC No. 26820,保藏时间为2023年3月17日。
2.权利要求1所述的鼠李糖乳酪杆菌在制备抗衰老产品上的应用。
3.如权利要求2所述的应用,其特征在于,所述抗衰老产品为医药食品。
4.一种利用权利要求1所述的具有抗衰老功能的菌株制备的酸奶。
5. 权利要求4所述的酸奶,其特征在于,包括以下重量份原料:全脂奶粉8~15份、蔗糖5~10份、灰树花多糖0.1~0.5份、透明质酸0.05~0.1份、蓝莓花青素0.03~0.1份、低聚果糖0.3~0.8份、胶原蛋白肽 0.05~0.1份、分离乳清蛋白粉0.3~0.8份、γ-氨基丁酸0.3~0.8份、酸奶发酵剂3~10份、其余为水。
6. 权利要求5所述的酸奶,其特征在于,所述酸奶发酵剂为副干酪乳酪杆菌LC-6,鼠李糖乳酪杆菌LC-9,植物乳植物杆菌 LP-28,鼠李糖乳酪杆菌 LP-45,嗜热链球菌ST-23。
7. 权利要求5或6所述的抗衰老酸奶的制备方法,其特征在于,包括以下步骤:
(1)原料称取
称取全脂乳粉、蔗糖、灰树花多糖、透明质酸、蓝莓花青素、低聚果糖、胶原蛋白肽、分离乳清蛋白粉、γ-氨基丁酸、稳定剂、甜味剂;
(2)物料水溶
准备55-60℃的配料水,在搅拌状态下,将物料投入并完全溶解后,静置恒温45-55℃水合30分钟;
(3)乳液均质
将水合奶粉定容到设定体积,将其升温到55-60℃,将其全部打入均质机,均质压力为18-20Mpa;
(4)杀菌
杀菌条件选择93-98℃,320s;杀菌完成后迅速降温到培养温度43℃;
(5)接种、发酵
将活化好的菌种按照5%的接种量进行接种,先在搅拌状态下发酵1h,后进行静置发酵;
(6)发酵产品
发酵中止后,迅速降温到20℃,后灌装,之后进入冷库冷藏。
8.根据权利要求7所述的抗衰老酸奶的制备方法,其特征在于:在步骤(2)中物料水溶,准备60℃的配料水,在搅拌状态下,将物料投入并完全溶解后,静置恒温55℃水合30分钟。
9.根据权利要求7所述的抗衰老酸奶的制备方法,其特征在于:在步骤(4)中杀菌条件选择95℃,300s。
10.根据权利要求7所述的抗衰老酸奶的制备方法,其特征在于:在步骤(5)中培养温度为30℃。
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