CN116210836A - 一种含有益生菌的固体饮料组合物及其应用 - Google Patents
一种含有益生菌的固体饮料组合物及其应用 Download PDFInfo
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Abstract
本发明公开了一种含有益生菌的固体饮料组合物及其应用。所述固体饮料组合物中包含乳酸片球菌菌粉1‑5份、乳果糖3‑20份和膳食纤维3‑17份,其中膳食纤维为红豆粉、圆苞车前子壳粉、黑芝麻粉;所述乳果糖为低聚果糖、赤藓糖醇、山梨糖醇。所述乳酸片球菌菌粉是采用保藏编号为CCTCC NO:M 20221643的乳酸片球菌NSS0302制备而成的,其菌含量不低于7×1011CFU/g。将所述固体饮料组合物中的原料通过混合的方法制得固体饮料,其通过促进肠道消化、加强代谢,对改善便秘有显著的效果。
Description
技术领域
本发明属于微生物领域,具体涉及一种含有益生菌的固体饮料组合物及其应用。
背景技术
便秘是指大便次数减少,一般每周少于3次,伴排便困难、粪便干结。便秘是临床上常见的症状,多长期持续存在,会严重影响生活质量。
益生菌是对于人体非常有益的营养成分,能够缓和胃肠的不适,因为益生菌能够调节人体肠道功能,对于一些腹泻、便秘、恶心等胃肠不适的症状有非常好的缓和作用,也能够促进机体对于营养的吸收,还能够提高人体的免疫力,抵抗有害细菌的侵袭。
现有治疗便秘的产品主要是通过药物或者增加膳食纤维的方式促进肠胃蠕动,长此以往容易破坏肠道菌群的正常水平,因此,研究一种能够改善便秘的益生菌制品非常重要。
发明内容
本发明目的在于提供一种含有益生菌的固体饮料组合物及其应用。所述固体饮料组合物以乳酸片球菌NSS0302作为主要原料,辅助添加乳果糖和多种膳食纤维,在促进肠道蠕动的同时增加肠道有益菌群的数量。
为实现上述发明目的,本发明采用以下技术方案予以实现:
本发明提供了一种改善便秘的固体饮料组合物,按重量份数计,所述固体饮料组合物中包含乳酸片球菌菌粉1-5份、乳果糖3-20份和膳食纤维3-17份。
进一步的,所述膳食纤维为红豆粉、圆苞车前子壳粉、黑芝麻粉中的一种或多种。
进一步的,其特征在于:所述乳果糖为低聚果糖、赤藓糖醇、山梨糖醇中的一种或多种。
进一步的,按重量份数计,所述固体饮料组合物的组分包含:
进一步的,所述乳酸片球菌菌粉是采用保藏编号为CCTCC NO:M 20221643的乳酸片球菌NSS0302制备而成的,其菌含量不低于7×1011CFU/g。
进一步的,所述乳酸片球菌菌粉的制备步骤为:
(1)将所述乳酸片球菌NSS0302活化后,以2%-5%(v/v)的接种量接种于发酵培养基中,37℃培养12-18h,得到发酵液经低温离心、清洗3-5次后,离心得到菌泥;
(2)将所述菌泥与冻干保护剂以1:1~2的质量体积比混匀,充分乳化后,得到乳化液;
(3)将所述乳化液经真空冷冻干燥,过后,得到乳酸片球菌NSS0302菌粉。
进一步的,所述真空冷冻干燥的条件步骤为:所述乳化液-45℃预冻,保持-45℃2h;2h内升温至-10℃,控制真空度0.20mbar,保持5h;2h升温至5℃,控制真空度0.20mbar,保持10h;2h升温至30℃,控制真空度0.20mbar,保持6h;抽极限真空,持续2h;反填洁净空气恢复至常压,菌粉出仓。
进一步的,所述冻干保护剂的配方为:海藻糖20g/L-30g/L,脱脂乳粉5g/L-10g/L,菊粉4g/L-7g/L、低聚异麦芽糖0.2g/L-0.5g/L,甘氨酸0.1g/L-0.2g/L,吐温80 0.5g/L-1g/L。
本发明还提供了所述的固体饮料组合物在制备改善便秘的固体饮料中的应用。
进一步的,所述固体饮料的制备方法为:制备乳酸片球菌菌粉,然后向其中加入乳果糖和膳食纤维,混合均匀后,无菌环境下包装即可。
进一步的,所述固体饮料中还包含添加剂;所述添加剂为低聚异麦芽糖、抗性糊精、菊粉、低聚异半乳糖、水苏糖、甘氨酸、茶多酚、富马酸、硅酸钙、抗坏血酸、普鲁兰多糖、阿斯巴甜中的一种或多种。
进一步的,所述固体饮料的使用为3.30g/kg·BW-6.6g/kg·BW。
与现有技术相比,本发明具有以下优点和技术效果:
本发明采用具有改善便秘功能的乳酸片球菌NSS0302作为主要的原料,辅助添加乳果糖和多种膳食纤维,在促进肠道蠕动的同时增加肠道有益菌群的数量,从而达到促进肠道消化、加强代谢,缓解便秘的效果。
膳食纤维中圆苞车前子壳粉具有清热解毒,消炎杀菌作用,其含有葡萄糖甙、蛋白质、维生素B1、胆碱、木糖、半乳胶醛酸、膳食纤维、树胶醛糖等营养物质。黑芝麻粉具有延缓衰老、补肝肾、润五脏、润肠,缓解大便干燥,调节内分泌的作用。红豆粉含有丰富的蛋白质、锌、钾,可以有效缓解便秘。因此,本发明的固体饮料兼具食补方法和微生态方法,营养全面,容易吸收,微生物的作用保证各项活性物质更好的被人体吸收,保护肠道菌群至平衡状态,乳果糖用于调味,口感好,且制成的固体饮料便于携带,适合便秘人群的使用。
附图说明
图1为乳酸片球菌NSS0302的平板培养照片。
图2为小肠推进率。
具体实施方式
结合以下具体实例对本发明的技术方案作进一步详细的说明。
下述实施例中,如无特殊说明,所使用的实验方法均为常规方法,所用材料、试剂等均可从生物或化学试剂公司购买。
实施例1:乳酸片球菌NSS0302的分离鉴定
1、分离筛选
以来源于3份四川三台县传统农家发酵豆豉为样品。将多份豆豉按5%的体积分别接种到MRS液体培养基中,37℃培养16h,无菌试管中加入1.0mL菌液和9.0mL 0.85%无菌生理盐水,稀释成浓度为10-1的菌液,重复上述操作依次将菌液稀释成浓度为10-2、10-3、10-4、10-5四个梯度,用无菌吸管各吸取0.1mL的10-3、10-4、10-5三个浓度梯度的菌液涂于MRS固体培养基上,每个浓度梯度做3个平行试验,37℃静置培养24h。进行菌落形态观察,挑选不同形态的单菌落再次于MRS固体培养基上划线分离,37℃下培养24h,重复划线分离3代,得到纯化后菌株,进行革兰氏染色并显微镜观测,挑选出革兰氏染色阳性,显微镜观测呈球状、杆状、多形状的菌株。在筛选得到的多种菌株中进一步筛选得到一株编号为NSS0302的菌株。
将菌株NSS0302接种于MRS固体平板上,37℃恒温培养24h,观察菌落形态特征。结果见图1,菌株NSS0302的菌落呈圆形,边缘整齐,表面光滑,中间突出,颜色呈乳白色,质地均匀。
2、16S rRNA鉴定
提取菌株NSS0302的DNA作为模板,利用细菌16S rRNA通用引物进行扩增,扩增片段送至生物公司进行序列测定,测序结果如SEQ ID No.1所示。BLAST比对结果表明,菌株NSS0302与Pediococcus acidilactici的序列相似性为99.72%,因此结合上述实验结果,判断菌株NSS0302为乳酸片球菌。
将筛选到的菌株NSS0302进行菌种保藏,所述乳酸片球菌NSS0302的保藏单位:中国典型培养物保藏中心(CCTCC);地址:中国.武汉.武汉大学;保藏日期:2022年10月24日;乳酸片球菌Pediococcus acidilactici的保藏编号为CCTCC NO:M 20221643。
实施例2:乳酸片球菌NSS0302菌粉的获得
将保存的乳酸片球菌NSS0302在改良MRS固体平板上划线,37℃培养活化,然后挑取单菌落接种于改良MRS液体培养基中,37℃培养16h,再以3%的接种量再接种至改良MRS培养基,37℃培养16h,再连续培养2代扩大培养;得到的发酵液在4℃下,12000r/min,离心10min,弃上清液,收集菌泥,使用超纯水洗涤3次后,将菌泥与冻干保护剂(配方为30g海藻糖,10g脱脂乳粉,7g菊粉、0.5g低聚异麦芽糖,0.2g甘氨酸,1g吐温80,1000mL纯水)以1:1.7的质量比混合,充分乳化后,经真空冷冻干燥(乳化液-45℃预冻,保持-45℃2h;2h内升温至-10℃,控制真空度0.20mbar,保持此过程5h;2h升温至5℃,控制真空度0.20mbar,保持此过程10h;2h升温至30℃,控制真空度0.20mbar,保持此过程6h;抽极限真空,持续2h;反填洁净空气恢复至常压,制品出仓),得到乳酸片球菌NSS0302的冻干活菌粉。
对冻干活菌粉进行倾注计数,菌粉真空保存在-18℃的冰箱中以备后用。乳酸片球菌NSS0302冻干活菌粉的菌含量为7×1011CFU/g。
实施例3:小鼠推进实验
一种固体饮料,以重量份数计,包括以下组分:
将上述组分混合均匀后,直接无菌环境下装入铝箔复合膜袋中即可。
依据卫生部颁发的《保健食品检验与评价技术规范》(2003版)中的通便功能检测方法建立小鼠便秘模型。按照每组6只小鼠随机分组,分为正常组、模型组、乳果糖组、固体饮料低剂量组、固体饮料中剂量组、固体饮料高剂量组(乳果糖组为3.5g/kg·BW,低剂量组为0.33g/kg·BW,中剂量组为3.30g/kg·BW,高剂量组为6.60g/kg·BW)共6组。
小鼠限食不限水12h后,于18d上午8:00进行灌胃,正常组小鼠灌胃0.25mL蒸馏水,其余五组均灌胃0.25mL盐酸洛哌丁胺溶液。30min后正常组和模型组小鼠用0.25mL 0.04%的酚红溶液灌胃,乳果糖组和低剂量、中剂量、高剂量组分别灌胃0.25mL含有乳果糖、低剂量、中剂量和高剂量菌粉的0.04%酚红溶液。30min后立即处死小鼠,取出小肠,小肠近端置于刻度板零处,测量肠管长度记为“小肠总长度”,溶液前沿处刻度值为“酚红推进长度”。
小肠推进率(%)=酚红推进长度(cm)/小肠总长度(cm)×100。
实验结果如图2所示,模型组小鼠的小肠推进率显著降低,说明便秘小鼠的小肠蠕动明显减缓。与模型组相比,乳果糖组小鼠的小肠推进率明显有所提高,进行显著性分析可知,低剂量固体饮料组小鼠的小肠推进率和便秘模型组相比没有差异,中剂量、高剂量组均与和模型组之间存在显著差异,这表明了中高剂量的固体饮料对肠道蠕动的改善作用。
实施例4:人群实验
实验共招募60例18岁以上的功能性便秘患者,符合罗马Ⅳ标准中功能性便秘诊断标准,并排除了IBS或肠道器质性病变。便秘患者以随机抽签的方式进行分组,实验分为2组,即固体饮料组、安慰剂组(安慰剂为抗性糊精,2g/d)。以4周为1个试验周期。第1周为导入阶段,期间受试者不能吃任何导泻药或者益生菌产品,第1周结束后受试者填写便秘量表并采集粪便样本;第2周~第4周为实验阶段,受试者每天服用2g固体饮料或安慰剂,在第2周~第3周期间、第4结束后受试者分别填写便秘量表并采集粪便样本。
(1)便秘量表:采用《基本信息调查表》、《膳食结构与生活状况调查表》调查便秘患者的基本信息,采用《便秘症状评估表》、便秘患者生存质量量表(PAC-QOL)》、《随访记录表》记录便秘患者在项目期间便秘症状和精神状态的变化。
①《基本信息调查表》:包含姓名、年龄、性别、身高、体重、疾病史以及近1个月以来的益生菌服用状况和药物服用状况等内容。
②《膳食结构与生活状况调查表》:包含常用主食、膳食口味、日常饮水量、益生菌或药物使用频率、运动情况、睡眠情况、心理压力、是否吸烟饮酒等项目。
③《便秘症状评估表》:采用评分的方式评价便秘症状的严重程度。总共5个项目,分别为排便困难/过度用力排便、粪便性状、排便不尽/下坠胀感、排便有肛门/直肠阻塞感、排便频率(d/次),每项0分~3分,总分15分为严重便秘,0分为无便秘症状。
④《便秘患者生存质量量表(PAC-QOL)》:包含便秘患者生理状态、心理状态、社交状态及满意度等方面的28项内容,采用评分的方法评价便秘患者的生存质量。根据生存质量的优劣程度共分为1分~5分的5个等级,总分越高患者的生存状态越差。
⑤《随访记录表》:为研究人员自用表格,包含饮食方式、食欲、饮食量、排便频率、粪便性状、是否腹痛/腹胀、是否服用泻剂等内容,用于实验期间对便秘患者的跟踪访问。
(2)评价标准:根据《便秘症状评估表》的积分计算固体饮料缓解便秘的有效率:有效率(%)=治疗后积分-治疗前积分/治疗前积分×100%。
有效率的评价如下:
≥75%为疗效很好;≥50%且<75%为疗效好;≥25%且<50%为疗效一般;<25%为疗效差。
显效率=疗效很好+疗效好;有效率=疗效一般;总有效率=显效率+有效率。
实验中分别收集便秘患者服用固体饮料前后的便秘症状量表,通过有效率计算公式获得便秘患者症状的变化,结果如表1所示。
表1:固体饮料缓解便秘有效率
固体饮料缓解便秘的显效率为:疗效很好+疗效好=13.3%+23.3%=36.6%;有效率为:疗效一般=43.3%;总有效率为:显效率+有效率=36.6%+43.3%=79.9%。安慰剂缓解便秘的显效率:效果很好+效果好=0%+3.3%=3.3%;有效率:疗效一般=33.0%;总有效率为:显效率+有效率=3.3%+33.0%=36.3%。可以看出,固体饮料缓解便秘的效果明显优于安慰剂,说明本发明制备的含有益生菌乳酸片球菌NSS0302的固体饮料具有很好缓解便秘的作用。
以上实施例仅用于说明本发明的技术方案,而非对其进行限制;尽管参照前述实施例对本发明进行了详细的说明,对于本领域的普通技术人员来说,依然可以对前述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;而这些修改或替换,并不使相应技术方案的本质脱离本发明所要求保护的技术方案的精神和范围。
Claims (10)
1.一种含有益生菌的固体饮料组合物,其特征在于:按重量份数计,所述固体饮料组合物中包含乳酸片球菌菌粉1-5份、乳果糖3-20份和膳食纤维3-17份。
2.根据权利要求1所述的固体饮料组合物,其特征在于:所述膳食纤维为红豆粉、圆苞车前子壳粉、黑芝麻粉中的一种或多种。
3.根据权利要求1所述的固体饮料组合物,其特征在于:所述乳果糖为低聚果糖、赤藓糖醇、山梨糖醇中的一种或多种。
4.根据权利要求1所述的固体饮料组合物,其特征在于:所述乳酸片球菌菌粉是采用保藏编号为CCTCC NO:M 20221643的乳酸片球菌NSS0302制备而成的,其菌含量不低于7×1011CFU/g。
5.根据权利要求4所述的固体饮料组合物,其特征在于:所述乳酸片球菌菌粉的制备步骤为:
(1)将所述乳酸片球菌NSS0302活化后,以2%-5%(v/v)的接种量接种于发酵培养基中,37℃培养12-18h,得到发酵液经低温离心、清洗3-5次后,离心得到菌泥;
(2)将所述菌泥与冻干保护剂以1:1~2的质量体积比混匀,充分乳化后,得到乳化液;
(3)将所述乳化液经真空冷冻干燥,过后,得到乳酸片球菌NSS0302菌粉。
6.根据权利要求5所述的固体饮料组合物,其特征在于:所述真空冷冻干燥的条件步骤为:所述乳化液-45℃预冻,保持-45℃2h;2h内升温至-10℃,控制真空度0.20mbar,保持5h;2h升温至5℃,控制真空度0.20mbar,保持10h;2h升温至30℃,控制真空度0.20mbar,保持6h;抽极限真空,持续2h;反填洁净空气恢复至常压,菌粉出仓。
7.权利要求1-6任一项所述的固体饮料组合物在制备改善便秘的固体饮料中的应用。
8.根据权利要求7所述的应用,其特征在于:所述固体饮料的制备方法为:制备乳酸片球菌菌粉,然后向其中加入乳果糖和膳食纤维,混合均匀后,无菌环境下包装即可。
9.根据权利要求7所述的应用,其特征在于:所述固体饮料中还包含添加剂;所述添加剂为低聚异麦芽糖、抗性糊精、菊粉、低聚异半乳糖、水苏糖、甘氨酸、茶多酚、富马酸、硅酸钙、抗坏血酸、普鲁兰多糖、阿斯巴甜中的一种或多种。
10.根据权利要求7所述的应用,其特征在于,所述固体饮料的使用量为3.30g/kg·BW-6.6g/kg·BW。
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