CN108823129B - 一种具有抵抗幽门螺旋杆菌作用的高活性益生菌固体饮料 - Google Patents
一种具有抵抗幽门螺旋杆菌作用的高活性益生菌固体饮料 Download PDFInfo
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Abstract
本发明的目的是提供一种具有抵抗幽门螺旋杆菌作用的高活性益生菌组合物,其按重量份计的包括:抗性糊精40‑60份、菊粉5‑15份、低聚果糖5‑15份、瑞士乳杆菌菌粉2‑8份、鼠李糖乳杆菌菌粉2‑8份、动物双岐杆菌菌粉2‑8份、乳双岐杆菌菌粉2‑8份、嗜酸乳杆菌菌粉2‑8份和两歧双岐杆菌菌粉2‑8份。本发明提供的高活性益生菌固体饮料各个组分相互作用,可以促使肠胃蠕动,促进肠胃菌群的平衡,增大了各组分单一作用的结果,可以更好的清除幽门螺旋杆菌,增加有益菌的含量。
Description
技术领域
本发明属于食品技术领域,具体涉及一种固体饮料及其制备方法,更具体地涉及一种具有抵抗幽门螺旋杆菌作用的高活性益生菌固体饮料。
背景技术
益生菌,英文probiotics一词由希腊语“for live”派生而来,译为“为了生命”,是含有生理活性的活菌,当被机体经过口服或其他给药方式摄入适当的数量后,能够定植于宿主并改善宿主微生态平衡,从而发挥有益作用。近年来的研究表明,人体内的细菌多达1000多种,特别是胃肠道内的细菌最为丰富,占全身微生态菌总数80%左右,是人体细胞的10倍,被誉为人类的“第二基因组”。这些细菌有助于人体消化食物,产生维生素以预防食物中细菌所诱发的疾病,同时刺激免疫***。随着研究的深入,发现肠道微生物在许多慢性疾病和症状中,如炎症、肥胖、糖尿病、还有癌症等也起了关键作用。微生物的数量和基因数量惊人,强烈影响人类的健康。这些发现为益生菌产品在食品、保健品、医药、养殖、环境、卫生用品等领域的应用带来了巨大的推动力。目前,益生菌被广泛用于酸奶、乳酸菌饮料、益生菌粉剂、动物饲料、环境治理等产品中,形成了相对完整和成熟的益生菌行业。
值得注意的是,益生菌只有达到一定的数量水平时,益生菌才能在宿主体内产生相应的效果。所以,以益生菌为功能成分的食品、保健品或药品需要保证所含活的益生菌数量保持在一定的水平。这一需求对益生菌的生产和制备提出了相应的要求。在益生菌的培养过程中,实现高密度培养,不但可以满足后续益生菌产品的功能需求,还可以提高益生菌生产环节中的生产效率,减少培养体积,缩短生产周期,降低生产成本。因此,益生菌的高密度培养是当前益生菌开发和生产中的关键技术问题。
液体发酵体系中,中国发明专利(公开号:CN 101230323A)在培养过程中添加碱性盐和脱氧剂以消除嗜酸乳杆菌或两歧双歧杆菌发酵时所受到的胁迫物质酸或氧,使最终产品中活菌数量提高2-4倍。中国发明专利(公开号:CN 101418270A)在干酪乳杆菌Zhang的培养过程中,通过加入NaOH、KOH或氨水等中和剂和通氮气的方法,得到1011CFU/mL以上活菌数的干酪乳杆菌的冻干菌粉。Hayakawa K等人(J Ferment Bioeng,1990,70,404)采用膜技术与发酵罐偶联,不断过滤除去乳酸等胁迫成分,实现最高菌体密度高达1011CFU/mL,该方法需耗费大量培养基,且仅限于实验室水平,很难扩大到生产应用规模。
半固体或固体发酵体系中,中国发明专利(公开号:CN 105132321A)采用由麦麸和豆粕等物质组成的培养基,接种屎肠球菌进行固态发酵,干燥产品中的活菌含量最高为3.8×1010CFU/g。
类似的还有中国专利(公开号:CN 101555510B;CN 101555511B;CN 103421710B;CN 102907563B;CN 103289937B;CN 1212391C)均采用豆粕、麸皮、玉米芯、米糠、花生壳、淀粉、玉米粉、草炭等农业废弃物或农产品为主要成分的固体培养基,接种微生物,进行高密度发酵,产品中的活菌数基本为1011CFU/g。这类产品主要用于动物饲料或进行药物等代谢产物生产。
益生菌是通过改善宿主肠道微生物菌群的平衡而发挥作用的活性微生物,益生菌对健康的有益作用是俄国科学家Mctchnikoff首先提出的,他认为肠道乳酸菌能够通过防止腐败菌的生长而起到延长寿命的作用。到目前为止,人们已经发现益生菌可以改善肠道菌群结构,抑制病原菌,消除致癌因子,提高机体免疫力,降低胆固醇。益生菌与人体的健康有着密切的关系,在牛奶中添加益生菌可以改善消化功能,促进肠胃蠕动,减轻人体乳糖不耐受症,防止婴幼儿的腹泻和消化不良,在医学上,益生菌可以用于急性轮状病毒引起的腹泻,便秘,结肠功能失调以及放射性治疗引起的负作用,还可以用来抗肿瘤和抗癌。目前,常用的益生菌菌种有双歧杆菌,乳酸杆菌和一些链球杆菌。肠道内双歧杆菌、乳酸杆菌等有益菌具有多种生物拮抗功能,如乳酸杆菌在体内发酵乳糖,产生大量的乳酸、乙酸、可降低肠道局部pH值,改善微生态环境,抑菌或杀死肠道内的大肠杆菌、沙门菌、链球菌等。同时,低pH值有利于肠道蠕动,维持正常生理功能,阻止致病菌的定植。乳酸菌产生的组织酸对革兰氏阴性菌作用更明显,乳酸菌本身还有抑制病原体复制的作。双歧杆菌能产生一种未知的广谱抗菌物质,这种物质可以抑制沙门氏菌、志贺氏菌、霍乱弧菌等病原菌的活性。益生菌产品在增加营养的同时可起到预防和治疗疾病的作用。
益生菌作为一种新型功能性添加成分,再保健品或医药领域发挥着越来越重要的作用,然而由于菌体非常敏感,容易受到周围环境的影响而失去活性。益生菌常温保存是该产业一大技术问题。因此,益生菌产品逐渐获得大众的青睐,益生菌酸奶等也太产品因贮藏运输成本高,保质期短等限制了其销售范围,而固体类益生菌产品则储存方便。
固体饮料是指以糖、乳和乳制品、蛋或蛋制品、果汁或食用植物提取物等为主要原料,添加适量的辅料或食品添加剂制成的每100克成品水分不高于5克的固体制品,呈粉末状、颗粒状或块状等。随着人们生活水平的不断提高,消费升级趋势已越来越明显,从市场来看,为适应消费者对速溶固体饮料口味、营养保健等多方面的需要,开始遵循天然营养、回归自然的思想,固体饮料朝着组分营养化、品种多样化、功能多样化的良性方向发展。
中国专利201710385573.4公开了一种益生菌固体饮料的制作方法,该方法包括益生菌培养、益生菌包埋、冷冻干燥、调配、二次包埋、热风干燥和包装储藏步骤。该发明方法以乳酸菌粉和菊粉为主要原料,并添加功能性糖和果蔬粉混合制成益生菌固体饮料。该发明采用超声波辅助发酵技术、变温冷冻干燥技术和二次包埋技术,大大提高了乳酸菌在制作过程中及食用后在人体内的存活率、生存能力和生物活性。
中国专利20161038905306公开了一种草本酵母益生菌固体饮料及其制备方法和应用,所述草本酵母益生菌固体饮料由如下按照质量百分比计的组分组成:冻干益生菌粉20~40%、益生元5~20%、水溶性膳食纤维0.2~20%、中草药粉5~20%、甜味剂0~8%。所述草本酵母益生菌固体饮料具有调节肠道菌群平衡、提高机体免疫力等多种功效,可作为保健品,具有补脾润肺、消渴生津、理气和胃的效果,还可以调理失眠多梦、易疲劳等亚健康症状。
中国专利201710466714.5公开了一种益生菌固体饮料及其制备方法,包括原料、辅料和益生菌粉,所述原料按重量克包括:低聚果糖1.2克和水苏糖0.6克,所述辅料原料按重量克包括:酵母粉0.1克和草莓粉0.1克,所述益生菌粉包括:动物双岐杆菌20亿、乳双歧杆菌80亿和鼠李糖乳杆菌80亿;该种益生菌固体饮料能够维持肠道内的菌落平衡,既能够帮助消除腹泻,又能够杜绝便秘,对肠道起到良好的双向调节作用。
幽门螺旋杆菌是一种革兰氏阴性杆菌,是慢性活动性胃炎、消化性胃溃疡的只要病因,并与胃恶性肿瘤关系密切,世界卫生组织和国际抗癌联盟将其列为I类致癌物,现有技术报道较多的益生菌固体饮料多为可以维持肠道捏的菌落平衡,但是对抗幽门螺杆菌作用的益生菌固体饮料的研究较少,因此需要提供出一种具有抵抗幽门螺旋杆菌作用的益生菌固体饮料。
发明内容
为了解决背景技术中存在的问题,本发明的目的在于提供一种益生菌的方法。
本发明采用的技术方案如下:
一种益生菌的培养方法:
在含有促进益生菌生长繁殖的促生长因子的培养基中添加多孔材料,培养基与多孔材料的体积比为1:0.001–1:1000,再在培养基中接种益生菌并进行培养,实现益生菌高密度培养。
所述培养为厌氧培养,所接种的益生菌为厌氧或兼性厌氧的微生物。
厌氧培养是在30-42℃下培养12-48h。
培养结束后,所述益生菌聚集到多孔材料的表面或者内部或者表面和内部,得到干燥后的产品中总活菌数可达1011-1015CFU/g以上。活菌数受干燥方法等因素的影响,冷冻干燥的产品中活菌数更高。
所制备得到的益生菌具有良好的耐热性、耐酸性及耐药性等性能。
所述的培养基为液态、半固态或者固态,培养体系可为液体培养、半固体培养和固体培养三种培养方式中的任一种,所述的多孔材料为固态。
所述的培养基为YPD(酵母菌)、双歧杆菌培养基(青春双歧杆菌、动物双歧杆菌、乳双歧杆菌、两歧双歧杆菌、短双歧杆菌、婴儿双歧杆菌、长双歧杆菌)、MRS培养基(嗜酸乳杆菌、干酪乳杆菌、德氏乳杆菌保加利亚亚种、瑞氏乳杆菌、约氏乳杆菌、植物乳杆菌、罗伊氏乳杆菌、鼠李糖乳杆菌、唾液乳杆菌、副干酪乳杆菌、乳酸乳球菌乳酸亚种、乳酸乳球菌双乙酰亚种、格氏乳杆菌、片球菌属、肠球菌属、魏斯氏菌属、蜜蜂球属、卷曲乳杆菌)、SIIM0012培养基(发酵乳杆菌、鼠李糖乳杆菌)、M17培养基(嗜热链球菌)、乳酸菌培养基(乳酸乳球菌乳脂亚种)、肠膜明串珠菌培养基(肠膜明串珠菌肠膜亚种)、丙酸细菌培养基(丙酸杆菌属)、麦芽汁琼脂(德巴利酵母属、假丝酵母属、球拟酵母属)、血琼脂培养基(拟杆菌属、梭杆菌属)、番茄汁培养基(酒球菌属)、PDA马铃薯、葡萄糖琼脂培养基(青霉属)、SICC0070丁酸菌培养基(梭菌属)、TSA胰蛋白胨大豆琼脂(气球菌属)、肉汤培养基(芽孢杆菌属)、LB培养基(微球菌属)等。
所述的培养基中含有促进益生菌生长繁殖的促生长因子。含有促进益生菌生长繁殖的促生长因子如乳清发酵液、胶原蛋白发酵液、马铃薯发酵液、海藻提取物、菌菇类发酵液或提取物、豆乳、番茄汁、胡萝卜汁、玉米汁、冬虫夏草提取物、灵芝提取物、铁皮石斛提取物、当归提取物、银杏叶提取物、麦芽提取物、参类提取物、黄芪提取物、罗汉果提取物、菊粉、酵母葡聚糖、低聚木糖、维生素、氨基酸、多巴胺、植酸、黄腐酸、腐殖酸、镁盐、锌盐、锰盐、钙盐、铁盐、硒盐等的一种或多种。
所述的培养基中的多孔材料的平均孔径大小为1纳米-500微米,体积孔隙率为5%-99.9%(v/v)。5%-99.9%(v/v)指的是多孔材料中的孔隙占整个材料的体积比为5%-99.9%。
所述的多孔材料为凝胶材料、气凝胶材料、泡沫材料、纤维材料、微孔材料、介孔材料或者大孔材料等。
所述的凝胶材料、气凝胶材料、泡沫材料和纤维材料的平均孔径从纳米级尺寸到微米级尺寸不等;所述的微孔材料的平均孔径小于2纳米,介孔材料的平均孔径为2-50纳米,大孔材料的平均孔径大于50纳米。
所述的多孔材料为天然多糖类及其衍生物、天然蛋白类及其衍生物、合成高分子的均聚物或者共聚物、无机材料等材料中的一种或多种。
天然多糖类如甲壳素、壳聚糖、纤维素、木质素、葡聚糖、琼脂糖、低聚果糖、大豆低聚糖、魔芋葡甘聚糖、低聚异麦芽糖、低聚木糖、低聚麦芽糖、淀粉、糊精、菊粉、海藻酸及其盐、右旋糖酐、透明质酸、海藻多糖、褐藻多糖、螺旋藻多糖、地衣多糖、香菇多糖、灵芝多糖、银耳多糖、***胶、果胶、瓜尔胶、黄原胶角叉菜胶等来源于植物、动物、海藻和微生物的多糖及其衍生物。
天然蛋白类如大豆蛋白、绿豆蛋白、小麦蛋白、玉米蛋白、谷蛋白、蚕丝蛋白、蜘蛛丝蛋白、胶原蛋白、乳清蛋白、明胶、酪蛋白酸钠、纤维蛋白、角蛋白等来源于植物、动物和微生物的蛋白及其衍生物。
合合成高分子的均聚物和共聚物如聚氨基酸、聚乙二醇、聚乳酸、聚乳酸-聚羟基乙酸共聚物、聚乙烯基吡咯烷酮、聚己酸内酯、聚β羟基丁酸酯、聚β羟基戊酸等。
无机材料如珍珠岩、蒙脱土、炭黑、白炭黑、陶瓷等。
所述的多孔材料为以上所述材料的一种或多种,但又不局限于以上材料。
所用的益生菌为对人和(或)动物有益的微生物,如双歧杆菌属、青春双歧杆菌、动物双歧杆菌、乳双歧杆菌、两歧双歧杆菌、短双歧杆菌、婴儿双歧杆菌、长双歧杆菌、乳杆菌属、嗜酸乳杆菌、干酪乳杆菌、卷曲乳杆菌、德氏乳杆菌保加利亚亚种(保加利亚乳杆菌)、德氏乳杆菌亚种、发酵乳杆菌、格氏乳杆菌、瑞士乳杆菌、约氏乳杆菌、副干酪乳杆菌、植物乳杆菌、罗伊氏乳杆菌、鼠李糖乳杆菌、唾液乳杆菌、链球菌属、嗜热链球菌、乳球菌属、乳酸球菌乳酸亚种、乳酸乳球菌乳脂亚种、乳酸乳球菌双乙酰亚种、肠膜明串珠菌肠膜亚种、丙酸杆菌属、酵母属、德巴利酵母数、假丝酵母属、球拟酵母属、拟杆菌属、梭杆菌属、酒球菌 属、消化链球菌属、青霉属、链球属、梭菌属、气球菌属、片球菌属、芽孢杆菌属、蜜蜂球菌属、肠球菌属、微球菌属、魏斯氏菌属等的一种或多种,但又不局限于以上菌种。
所述培养基中,多孔材料上接种的益生菌菌液量为0.1%-50%(v/v)。0.1%-50%(v/v)指的是益生菌菌液加入的体积量为加入后整个培养基体积的0.1%-50%。
所述益生菌接种菌液中的菌液的浓度是108-109CFU/mL。
依据本发明的培养方法培养,菌种在培养基中多组分的促进下,其生长繁殖速率大幅提升,培养结束后,干燥即可制得益生菌产品。产品中的总活菌数可达1011-1015CFU/g以上,可用作益生菌发酵剂或益生菌制剂。这种益生菌具有良好的耐热性、耐酸碱性及耐药性等性能。
本发明可以概述为:
本发明的目的是提供一种高活性益生菌固体饮料,其按重量份计的包括:抗性糊精40-60份、菊粉5-15份、低聚果糖5-15份、瑞士乳杆菌菌粉2-8份、鼠李糖乳杆菌菌粉2-8份、动物双岐杆菌菌粉2-8份、乳双岐杆菌菌粉2-8份、嗜酸乳杆菌菌粉2-8份和两歧双岐杆菌菌粉2-8份。本发明提供的高活性益生菌固体饮料各个组分相互作用,可以促使肠胃蠕动,促进肠胃菌群的平衡,增大了各组分单一作用的结果,可以更好的清除幽门螺旋杆菌,增加有益菌的含量。本发明属于食品技术领域,具体涉及一种固体饮料及其制备方法,更具体地涉及一种高活性益生菌固体饮料及其制备方法。
针对现有技术的不足,本发明提供了一种高活性益生菌固体饮料及其制备方法。
一方面,本发明提供了一种高活性益生菌固体饮料,其按重量份计的包括:抗性糊精40-60份、菊粉5-15份、低聚果糖5-15份、瑞士乳杆菌菌粉2-8份、鼠李糖乳杆菌菌粉2-8份、动物双岐杆菌菌粉2-8份、乳双岐杆菌菌粉2-8份、嗜酸乳杆菌菌粉2-8份和两歧双岐杆菌菌粉2-8份。
优选地,所述的抗性糊精、鼠李糖乳杆菌菌粉和两歧双岐杆菌菌粉的质量比为8:1:1;
优选地,所述的低聚果糖、瑞士乳杆杆菌菌粉和动物双岐杆菌菌粉的质量比为3:1:2;
优选地,所述的菊粉、乳双岐杆菌菌粉、嗜酸乳杆菌菌粉的质量比为2:1:1。
在一些更有选的实施方案中,所述的高活性益生菌固体饮料,还包括葛根粉和黑豆粉,葛根内含12%的黄酮类化合物,如葛根素、大豆黄酮苷、花生酥等营养成分,还含有蛋白质、氨基酸、糖和人体必须的铁、钙、铜、硒等矿物质,是老少皆宜的名贵补品,有“千年人参”之美誉。黑豆又名乌豆,味甘性平,具有高蛋白、低热量的特性,具有抗氧化,延缓衰老的作用。
因此,作为一种更优选的实施方法,上述的高活性益生菌固体饮料,其按重量份计的还包括:葛根粉5份、黑豆粉8份。
在本发明中的实施例中,所述的瑞士乳杆菌菌粉为瑞士杆菌R52菌粉;所述的鼠李糖乳杆菌菌粉为鼠李糖乳杆菌R1l菌粉;所述的动物双岐杆菌菌粉为动物双岐杆菌BB-12菌粉;所述的乳双岐杆菌菌粉为乳双岐杆菌Bi-07菌粉;所述的嗜酸乳杆菌菌粉为嗜酸乳杆菌NCFM菌粉;所述的两歧双岐杆菌菌粉为两岐双岐杆菌Bb-06菌粉。
另一方面,本发明提供了一种高活性益生菌固体饮料的制备方法,具体包括以下步骤:
(1)将配方用量的瑞士乳杆菌菌粉、鼠李糖乳杆菌菌粉、动物双岐杆菌菌粉、乳双岐杆菌菌粉、嗜酸乳杆菌菌粉和两歧双岐杆菌菌粉搅拌均匀后,得到混合物A;
(2)将菊粉、抗性糊精和混合物A混合,搅拌均匀后,得到混合物B;
(3)将得到的混合物B与配方中其余的原料混合,搅拌均匀后,得到混合物D;
(4)将步骤(3)得到的混合物D冷冻干燥,即得到所述的益生菌固体饮料。
所述步骤(1)中搅拌的转速为5-15r/min,搅拌的时间为30-50min。
所述步骤(2)中搅拌的转速为20-30r/min,搅拌的时间为20-40min。
所述步骤(3)中搅拌的转速为35-45r/min,搅拌的时间为10-30min。
所述步骤(4)中冷冻干燥的条件为:真空度0.095-0.01Mpa,在5-10oC下保持3-4h后,在0-5oC下保持2-3h后,在-5-0oC下保持1-2h后,在-10-0o下保持1-2h后,在-20--10oC下保持1-2h后,在-30--20oC下保持1-2h。
无特别说明,本发明中的瑞士乳杆菌菌粉、鼠李糖乳杆菌菌粉、动物双岐杆菌菌粉、乳双岐杆菌菌粉、嗜酸乳杆菌菌粉和两歧双岐杆菌菌粉中的活菌数均为2x1011cfu/g。
本发明的有益效果为:
(1)本发明提供的高活性益生菌固体饮料,包括:抗性糊精、菊粉、低聚果糖、瑞士乳杆菌菌粉、鼠李糖乳杆菌菌粉、动物双岐杆菌菌粉、乳双岐杆菌菌粉、嗜酸乳杆菌菌粉和两歧双岐杆菌菌粉,彼此之间相互作用,可以促使肠胃蠕动,促进肠胃菌群的平衡,增大了各组分单一作用的结果。
(2)本发明提供的高活性益生菌固体饮料,可以更好的清除幽门螺旋杆菌,促进肠道中有机酸和短链脂肪酸的增加,降低胃的pH值,导致酸度降低而抑制幽门螺旋杆菌,增加有益菌的含量。
(3)本发明提供的高活性益生菌固体饮料可以减轻炎症,促进消化,增强免疫力。
具体实施方式
说明:本发明所有表格中的“-”表示该项内容不存在。
一种高活性益生菌固体饮料,其按重量份计的包括:抗性糊精40-60份、菊粉5-15份、低聚果糖5-15份、瑞士乳杆菌菌粉2-8份、鼠李糖乳杆菌菌粉2-8份、动物双岐杆菌菌粉2-8份、乳双岐杆菌菌粉2-8份、嗜酸乳杆菌菌粉2-8份和两歧双岐杆菌菌粉2-8份。
在本发明中的实施例中,瑞士乳杆菌菌粉为瑞士杆菌R52菌粉;所述的鼠李糖乳杆菌菌粉为鼠李糖乳杆菌R1l菌粉;所述的动物双岐杆菌菌粉为动物双岐杆菌BB-12菌粉;所述的乳双岐杆菌菌粉为乳双岐杆菌Bi-07菌粉;所述的嗜酸乳杆菌菌粉为嗜酸乳杆菌NCFM菌粉;所述的两歧双岐杆菌菌粉为两岐双岐杆菌Bb-06菌粉。
在本发明中一些实施例中,上述的高活性高益生菌固体饮料还包括葛根粉和黑豆粉。
该高活性益生菌固体饮料的制备方法为:
(1)将配方用量的瑞士乳杆菌菌粉、鼠李糖乳杆菌菌粉、动物双岐杆菌菌粉、乳双岐杆菌菌粉、嗜酸乳杆菌菌粉和两歧双岐杆菌菌粉搅拌均匀后,得到混合物A;
(2)将菊粉、抗性糊精和混合物A混合,搅拌均匀后,得到混合物B;
(3)将得到的混合物B与配方中其余的原料混合,搅拌均匀后,得到混合物D;
(4)将步骤(3)得到的混合物冷冻干燥,即得到所述的益生菌固体饮料。
步骤(1)中搅拌的转速为5-15r/min,搅拌的时间为30-50min。
步骤(2)中搅拌的转速为20-30r/min,搅拌的时间为20-40min。
步骤(3)中搅拌的转速为35-45r/min,搅拌的时间 为10-30min。
步骤(4)中冷冻干燥的条件为:真空度0.095-0.1 Mpa,在5-10oC下保持3-4h后,在0-5oC下保持2-3h后,在-5-0oC下保持1-2h后,在-10-0o下保持1-2h后,在-20--10oC下保持1-2h后,在-30--20oC下保持1-2h。
通过调整基础实施例中的具体实验参数,得到了如表1所示的实施例1-10。
实施例1 | 实施例2 | 实施例3 | |
抗性糊精(份) | 40 | 60 | 50 |
菊粉(份) | 5 | 15 | 10 |
低聚果糖(份) | 15 | 5 | 10 |
瑞士乳杆菌R52菌粉(份) | 2 | 8 | 5 |
鼠李糖乳杆菌R1l菌粉(份) | 8 | 2 | 4 |
动物双岐杆菌BB-12菌粉(份) | 8 | 2 | 4 |
乳双岐杆菌Bi-07菌粉(份) | 2 | 8 | 5 |
嗜酸乳杆菌NCFM菌粉(份) | 2 | 8 | 4 |
两岐双岐杆菌Bb-06菌粉(份) | 8 | 2 | 5 |
葛根粉(份) | — | — | — |
黑豆粉(份) | — | — | — |
抗性糊精、鼠李糖乳杆菌R1l菌粉和两岐双岐杆菌Bb-06菌粉的质量比 | 5:1:1 | 30:1:1 | 12.5:1:1.25 |
低聚果糖、瑞士乳杆菌R52菌粉和动物双岐杆菌BB-12菌粉的质量比 | 7.5:1:4 | 2.5:4:1 | 2.5:1.25:1 |
菊粉、乳双岐杆菌Bi-07菌粉和嗜酸乳杆菌NCFM菌粉的质量比 | 2.5:1:1 | 1.875:1:1 | 2.5:1.25:1 |
步骤(1)中的搅拌转速(r/min) | 5 | 15 | 10 |
步骤(1)中的搅拌时间(min) | 30 | 50 | 40 |
步骤(2)中的搅拌转速(r/min) | 20 | 30 | 25 |
步骤(2)中的搅拌时间(min) | 20 | 40 | 30 |
步骤(3)中搅拌转速(r/min) | 35 | 45 | 40 |
步骤(3)中搅拌时间(min) | 10 | 30 | 20 |
步骤(4)中真空度(Mpa) | 0.095 | 0.1 | 0.095 |
第1次冷冻的温度(<sup>o</sup>C) | 5 | 10 | 8 |
第1次冷冻的时间(h) | 3 | 4 | 3.5 |
第2次冷冻的温度(<sup>o</sup>C) | 0 | 5 | 3 |
第2次冷冻的时间(h) | 2 | 3 | 2.5 |
第3次冷冻的温度(<sup>o</sup>C) | -5 | 0 | -3 |
第3次冷冻的时间(h) | 1 | 2 | 1.5 |
第4次冷冻的温度(<sup>o</sup>C) | -10 | 0 | -5 |
第4次冷冻的时间(h) | 2 | 1 | 1.5 |
第5次冷冻的温度(<sup>o</sup>C) | -20 | -10 | -15 |
第5次冷冻的时间(h) | 1 | 2 | 1.5 |
第6次冷冻的温度(<sup>o</sup>C) | -30 | -20 | -25 |
第6次冷冻的时间(h) | 1 | 2 | 1.5 |
实施例4 | 实施例5 | 实施例6 | |
抗性糊精(份) | 48 | 50 | 50 |
菊粉(份) | 10 | 10 | 10 |
低聚果糖(份) | 10 | 9 | 10 |
瑞士乳杆菌R52菌粉(份) | 5 | 3 | 5 |
鼠李糖乳杆菌R1l菌粉(份) | 6 | 4 | 4 |
动物双岐杆菌BB-12菌粉(份) | 4 | 6 | 4 |
乳双岐杆菌Bi-07菌粉(份) | 5 | 5 | 5 |
嗜酸乳杆菌NCFM菌粉(份) | 4 | 4 | 5 |
两岐双岐杆菌Bb-06菌粉(份) | 6 | 5 | 5 |
葛根粉(份) | — | — | — |
黑豆粉(份) | — | — | — |
抗性糊精、鼠李糖乳杆菌R1l菌粉和两岐双岐杆菌Bb-06菌粉的质量比 | 8:1:1 | 12.5:1:1.25 | 12.5:1:1.25 |
低聚果糖、瑞士乳杆菌R52菌粉和动物双岐杆菌BB-12菌粉的质量比 | 2.5:1.25:1 | 3:1:2 | 2.5:1.25:1 |
菊粉、乳双岐杆菌Bi-07菌粉和嗜酸乳杆菌NCFM菌粉的质量比 | 2.5:1.25:1 | 2.5:1.25:1 | 2:1:1 |
步骤(1)中的搅拌转速(r/min) | 10 | 10 | 10 |
步骤(1)中的搅拌时间(min) | 40 | 40 | 40 |
步骤(2)中的搅拌转速(r/min) | 25 | 25 | 25 |
步骤(2)中的搅拌时间(min) | 30 | 30 | 30 |
步骤(3)中搅拌转速(r/min) | 40 | 40 | 40 |
步骤(3)中搅拌时间(min) | 20 | 20 | 20 |
步骤(4)中真空度(Mpa) | 0.095 | 0.095 | 0.095 |
第1次冷冻的温度(<sup>o</sup>C) | 8 | 8 | 8 |
第1次冷冻的时间(h) | 3.5 | 3.5 | 3.5 |
第2次冷冻的温度(<sup>o</sup>C) | 3 | 3 | 3 |
第2次冷冻的时间(h) | 2.5 | 2.5 | 2.5 |
第3次冷冻的温度(<sup>o</sup>C) | -3 | -3 | -3 |
第3次冷冻的时间(h) | 1.5 | 1.5 | 1.5 |
第4次冷冻的温度(<sup>o</sup>C) | -5 | -5 | -5 |
第4次冷冻的时间(h) | 1.5 | 1.5 | 1.5 |
第5次冷冻的温度(<sup>o</sup>C) | -15 | -15 | -15 |
第5次冷冻的时间(h) | 1.5 | 1.5 | 1.5 |
第6次冷冻的温度(<sup>o</sup>C) | -25 | -25 | -25 |
第6次冷冻的时间(h) | 1.5 | 1.5 | 1.5 |
实施例7 | 实施例8 | 实施例9 | 实施例10 | |
抗性糊精(份) | 48 | 48 | 48 | 48 |
菊粉(份) | 10 | 10 | 10 | 10 |
低聚果糖(份) | 9 | 9 | 9 | 9 |
瑞士乳杆菌R52菌粉(份) | 3 | 3 | 3 | 3 |
鼠李糖乳杆菌R1l菌粉(份) | 6 | 6 | 6 | 6 |
动物双岐杆菌BB-12菌粉(份) | 6 | 6 | 6 | 6 |
乳双岐杆菌Bi-07菌粉(份) | 5 | 5 | 5 | 5 |
嗜酸乳杆菌NCFM菌粉(份) | 5 | 5 | 5 | 5 |
两岐双岐杆菌Bb-06菌粉(份) | 6 | 6 | 6 | 6 |
葛根粉(份) | — | 5 | — | 5 |
黑豆粉(份) | — | — | 8 | 8 |
抗性糊精、鼠李糖乳杆菌R1l菌粉和两岐双岐杆菌Bb-06菌粉的质量比 | 8:1:1 | 8:1:1 | 8:1:1 | 8:1:1 |
低聚果糖、瑞士乳杆菌R52菌粉和动物双岐杆菌BB-12菌粉的质量比 | 3:1:2 | 3:1:2 | 3:1:2 | 3:1:2 |
菊粉、乳双岐杆菌Bi-07菌粉和嗜酸乳杆菌NCFM菌粉的质量比 | 2:1:1 | 2:1:1 | 2:1:1 | 2:1:1 |
步骤(1)中的搅拌转速(r/min) | 10 | 10 | 10 | 10 |
步骤(1)中的搅拌时间(min) | 40 | 40 | 40 | 40 |
步骤(2)中的搅拌转速(r/min) | 25 | 25 | 25 | 25 |
步骤(2)中的搅拌时间(min) | 30 | 30 | 30 | 30 |
步骤(3)中搅拌转速(r/min) | 40 | 40 | 40 | 40 |
步骤(3)中搅拌时间(min) | 20 | 20 | 20 | 20 |
步骤(4)中真空度(Mpa) | 0.095 | 0.095 | 0.095 | 0.095 |
第1次冷冻的温度(<sup>o</sup>C) | 8 | 8 | 8 | 8 |
第1次冷冻的时间(h) | 3.5 | 3.5 | 3.5 | 3.5 |
第2次冷冻的温度(<sup>o</sup>C) | 3 | 3 | 3 | 3 |
第2次冷冻的时间(h) | 2.5 | 2.5 | 2.5 | 2.5 |
第3次冷冻的温度(<sup>o</sup>C) | -3 | -3 | -3 | -3 |
第3次冷冻的时间(h) | 1.5 | 1.5 | 1.5 | 1.5 |
第4次冷冻的温度(<sup>o</sup>C) | -5 | -5 | -5 | -5 |
第4次冷冻的时间(h) | 1.5 | 1.5 | 1.5 | 1.5 |
第5次冷冻的温度(<sup>o</sup>C) | -15 | -15 | -15 | -15 |
第5次冷冻的时间(h) | 1.5 | 1.5 | 1.5 | 1.5 |
第6次冷冻的温度(<sup>o</sup>C) | -25 | -25 | -25 | -25 |
第6次冷冻的时间(h) | 1.5 | 1.5 | 1.5 | 1.5 |
对比例1一种高活性益生菌固体饮料及其制备方法
配方基本同实施例3,与实施例3的差别在于配方中不含瑞士乳杆菌R52菌粉,制备方法同实施例3。
对比例2一种高活性益生菌固体饮料及其制备方法
配方基本同实施例3,与实施例3的差别在于配方中不含鼠李糖乳杆菌R1l菌粉,制备方法同实施例3。
对比例3一种高活性益生菌固体饮料及其制备方法
配方基本同实施例3,与实施例3的差别在于配方中不含动物双岐杆菌BB-12菌粉,制备方法同实施例3。
对比例4一种高活性益生菌固体饮料及其制备方法
配方基本同实施例3,与实施例3的差别在于配方中不含乳双岐杆菌Bi-07菌粉,制备方法同实施例3。
对比例5一种高活性益生菌固体饮料及其制备方法
配方基本同实施例3,与实施例3的差别在于配方中不含嗜酸乳杆菌NCFM菌粉,制备方法同实施例3。
对比例6一种高活性益生菌固体饮料及其制备方法
配方基本同实施例3,与实施例3的差别在于配方中不含两岐双岐杆菌Bb-06菌粉,制备方法同实施例3。
对比例7一种高活性益生菌固体饮料及其制备方法
配方基本同实施例3,与实施例3的差别在于步骤(4)中只进行一次冷冻干燥,冷冻干燥的时间为-25oC,冷冻时间为11小时。
实验例1
将实施例1-10与对比例1-7制备的高活性益生菌固体饮料进行体外抑制幽门螺旋杆菌试验,具体步骤如下:
1、材料
菌株H.CagA(+)pylori为国际标准菌株NCTC11637,为CagA(+)和VacA(+)。
2、方法
在无菌环境下,称取实施例1-10和对比例1-7的5g益生菌固体饮料,分别溶于200ml无菌水中,用盐酸调节溶液pH为2.0,并40℃恒温搅拌2h,过滤,滤液旋转蒸发至体积小于50ml,然后用无菌水定容至100ml。然后按倍比稀释为1:40。用滤纸打孔器打成若干直径为6mm的圆形滤纸片,120℃高压蒸汽灭菌。无菌环境下,用无菌镊子分别夹取灭菌的滤纸片,每张滤纸片上加10μl组合物溶液,水分挥发后备用。
(2)称取空肠弯曲琼脂培养基450g,加入蒸馏水1000ml,加热搅拌溶解,高温高压灭菌,冷却至50-60℃。加入100ml脱纤维羊血,搅拌均匀,倾倒无菌培养皿。将H.pylori于三气培养箱中培养3d,用布氏肉汤将其洗脱配制成悬液。通过酶标仪检测幽门螺旋杆菌浓度,将其制备为108cfu/ml的布氏肉汤菌液,涂于平板上。
(3)采用制片扩散法,通过游标卡尺测量抑菌圈直径。
2、实验结果如表2所示
从表2中可以看出,本发明实施例1-10中提供的益生菌组合物对幽门螺旋杆菌有良好的抑菌效果,远高于对比例1-10中提供的益生菌组合物。其中实施例4中通过调节抗性糊精、鼠李糖乳杆菌R1l菌粉和两岐双岐杆菌Bb-06菌粉的质量比为8:1:1,可以提高益生菌组合物对幽门螺旋杆菌的抑菌活性。
实验例2
1、材料:将雄性小鼠称重,每组10只,按照本发明实施例1-10提供的高活性益生菌固体饮料,对小鼠称重,分别按照1.67g/kg.bw对小鼠练血灌胃10天,空白组用无菌稀释液罐胃。在给与受试物之前,取小鼠粪便0.1g,加无菌生理盐水10倍稀释,计算每克湿便中的菌数,取对数后进行数据统计,最后一次给予受试物24之后,无菌采取粪便,检测肠道菌群,方法同上,结果与表3所示:
从表3中可以看出,本发明实施例1-10提供的高活性益生菌固体饮料,经小鼠灌服后,双岐杆菌和乳酸杆菌的含量高于灌服前,而对比例1-7中提供的高活性益生菌固体饮料的中的双岐杆菌和乳酸杆菌的数量没有明显增加,甚至降低,说明本配方提供的益生菌固体饮料具有改善肠道菌落的功能。其中实施例5与实施例3的差别在于,调整了低聚果糖、瑞士乳杆菌R52菌粉和动物双岐杆菌BB-12菌粉的质量比为3:1:2,可以显著地增加小鼠肠道中乳酸杆菌和双歧杆菌的含量,具有统计学差异(P<0.05)。其中实施例6与实施例3的差别在于调整菊粉、乳双岐杆菌Bi-07菌粉和嗜酸乳杆菌NCFM菌粉的质量比为2:1:1,实施例6提供的高活性益生菌固体饮料可以显著降低小鼠肠道的肠球菌的含量,具有统计学差异(P<0.05)。实施例10与实施例7的差别在于含有黑豆粉和葛根粉,实施例10提供的高活性益生菌固体饮料可以显著降低小鼠肠道的产气荚膜梭菌的含量,具有统计学差异(P<0.05)。
惟以上所述者,仅为本发明之较佳实施例而已,当不能以此限定本发明实施之范围,即大凡依本发明权利要求及发明说明书所记载的内容所作出简单的等效变化与修饰,皆仍属本发明权利要求所涵盖范围之内。此外,摘要部分和标题仅是用来辅助专利文件搜寻之用,并非用来限制本发明之权利范围。
Claims (4)
1.一种高活性益生菌固体饮料,其按重量份计的包括:抗性糊精40-60份、菊粉5-15份、低聚果糖5-15份、瑞士乳杆菌菌粉2-8份、鼠李糖乳杆菌菌粉2-8份、动物双歧 杆菌菌粉2-8份、乳双歧 杆菌菌粉2-8份、嗜酸乳杆菌菌粉2-8份和两歧双歧 杆菌菌粉2-8份;
所述的抗性糊精、鼠李糖乳杆菌菌粉和两歧双歧 杆菌菌粉的质量比为8:1:1;
所述的低聚果糖、瑞士乳杆杆菌菌粉和动物双歧 杆菌菌粉的质量比为3:1:2;
所述的菊粉、乳双歧 杆菌菌粉和嗜酸乳杆菌菌粉的质量比为2:1:1;
所述的固体饮料,其按重量份计的还包括:葛根粉5份和黑豆粉8份。
2.如权利要求1所述的高活性益生菌固体饮料的制备方法,其特征在于:具体包括以下步骤:
(1)将配方用量的瑞士乳杆菌菌粉,鼠李糖乳杆菌菌粉,动物双歧 杆菌菌粉,乳双歧杆菌菌粉,嗜酸乳杆菌菌粉,两歧双歧 杆菌菌粉搅拌均匀后,得到混合物A;
(2)将菊粉,抗性糊精和混合物A混合,搅拌均匀后,得到混合物B;
(3)将得到的混合物B与配方中其余的原料混合,搅拌均匀后,得到混合物D;
(4)将步骤(3)得到的混合物D冷冻干燥,即得到所述的益生菌固体饮料。
3.如权利要求2所述的高活性益生菌固体饮料的制备方法,其特征在于:所述步骤(1)中搅拌的转速为5-15r/min,搅拌的时间为30-50min;所述步骤(2)中搅拌的转速为20-30r/min,搅拌的时间为20-40min;所述步骤(3)中搅拌的转速为35-45r/min,搅拌的时间为10-30min。
4.如权利要求2所述的高活性益生菌固体饮料的制备方法,其特征在于:所述步骤(4)中冷冻干燥的条件为:真空度0.095-0.01Mpa,在5-10oC下保持3-4h后,在0-5oC下保持2-3h后,在-5-0oC下保持1-2h后,在-10-0oC下保持1-2h后,在-20--10oC下保持1-2h后,在-30--20oC下保持1-2h。
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