CN117089580A - 干酪乳杆菌sf-l-12发酵物及其制备工艺、应用 - Google Patents
干酪乳杆菌sf-l-12发酵物及其制备工艺、应用 Download PDFInfo
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- CN117089580A CN117089580A CN202311354341.4A CN202311354341A CN117089580A CN 117089580 A CN117089580 A CN 117089580A CN 202311354341 A CN202311354341 A CN 202311354341A CN 117089580 A CN117089580 A CN 117089580A
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Abstract
本发明涉及微生物应用技术领域,具体涉及一种干酪乳杆菌SF‑L‑12发酵物及其制备工艺、应用。干酪乳杆菌(Lactobacillus casei)SF‑L‑12已于2020年10月21日保藏于中国微生物菌种保藏管理委员会普通微生物中心,保藏编号为CGMCC No.20927,被发酵底物包括净化水64~82份,党参提取物1~3份,当归提取物1~4份,枸杞子提取物1~4份,五味子提取物2~5份,蒲公英提取物2~3份,茯苓提取物2~3份,大枣粉2~5份,薏苡仁粉2~4份,苹果汁4~5份,均为药食同源的中药,合理配伍,安全无害。运用生物发酵工艺制备所得的干酪乳杆菌SF‑L‑12发酵物可显著提高血液中IgG、IgA、IgM等免疫球蛋白含量,有效提高机体免疫力。
Description
技术领域
本发明涉及微生物应用技术领域,具体涉及一种干酪乳杆菌SF-L-12发酵物及其制备工艺、应用。
背景技术
免疫球蛋白(immunoglobulin)指具有抗体活性的动物蛋白,主要存在于血浆中,也见于其他体液、组织和一些分泌液中。人血浆内的免疫球蛋白大多数存在于丙种球蛋白(γ-球蛋白)中。免疫球蛋白可以分为IgG、IgA、IgM、IgD、IgE五类。严重缺乏免疫球蛋白会引起免疫缺陷,在临床上能够诱发多种疾病如感染、肿瘤、***反应、自身免疫性疾病等。一般体内免疫球蛋白水平低的人群容易引发免疫缺陷类疾病,且这类人群的恶性肿瘤发作率要高于常人。
专利CN106999588A公开了具有增加的稳定性的包含免疫球蛋白的药物产品,其包含在含有除氧剂的第二包装内的预装填聚合物注射器中的多克隆免疫球蛋白溶液,通过静脉注射或皮下注射IgG以治疗免疫缺陷类疾病。但是该类疗法的成本较高,副作用较大。专利CN113598361A公开了一种提升人体免疫力的本草酵素及其制备方法,所述的本草酵素主要原料为药食同源的中药材,包括人参、茯苓、大枣、桑椹、黄精、枸杞、甘草、覆盆子、松花粉、蜂蜜等,上述中药材经过提取,加入蜂蜜、赤砂糖,依次进行乳酸发酵、酒精发酵和醋酸发酵,最终制得本草酵素。但是据其说明书记载,试食组的受试者服用上述本草酵素产品90天后,IgG、IgA、IgM含量较试食前有上升趋势,但无统计学差异(P>0.05);与对照组比较,试食组IgG、IgA、IgM含量有上升趋势,但无统计学差异(P>0.05),由此可知,上述本草酵素制备工艺较复杂,而增加免疫球蛋白分泌量的效果不明显。
发明内容
针对现有技术中增加免疫球蛋白的制剂存在生产成本高、副作用大或治疗效果不明显、见效慢的缺点问题,本发明提供一种干酪乳杆菌SF-L-12发酵物,原料均采用药食同源的的中药,运用生物发酵技术制备,安全无害且效果显著。
第一方面,本发明提供一种干酪乳杆菌SF-L-12发酵物,由被发酵底物经干酪乳杆菌SF-L-12发酵后制得,干酪乳杆菌(Lactobacillus casei)SF-L-12已于2020年10月21日保藏于中国微生物菌种保藏管理委员会普通微生物中心,保藏编号为CGMCC No.20927,保藏地址为北京市朝阳区北辰西路1号院3号;被发酵底物包括如下重量份的组分:净化水64~82份,党参提取物1~3份,当归提取物1~4份,枸杞子提取物1~4份,五味子提取物2~5份,蒲公英提取物2~3份,茯苓提取物2~3份,大枣粉2~5份,薏苡仁粉2~4份,苹果汁4~5份。
进一步的,被发酵底物包括如下重量份的组分:净化水74份,党参提取物2份,当归提取物2份,枸杞子提取物2份,五味子提取物3份,蒲公英提取物3份,茯苓提取物3份,大枣粉3份,薏苡仁粉3份,苹果汁5份。
本发明发酵饮品的被发酵底物具备如下功效:
党参:健脾益肺,养血生津;
当归:补血活血,调经止痛;
枸杞子:滋补肝肾,益精明目。
五味子:收敛固涩,益气生津,补肾宁心;
蒲公英:清热解毒,消肿散结,利尿通淋;
茯苓:利水渗湿,健脾宁心;
大枣:补脾和胃,益气生津;
薏苡仁:利水渗湿,健脾止泻。
配方以党参为君药,健运中气,以当归为臣药,补血活血,以五味子为佐药,敛肺宁心,配枸杞子、蒲公英、茯苓、大枣、薏苡仁,清热解毒,补脾益肾。
第二方面,本发明提供上述干酪乳杆菌SF-L-12发酵物的制备工艺,具体包括以下步骤:
(1)首先将被发酵底物的各组分混合,得到混合浆料;
(2)将混合浆料输送至灭菌容器中,高压灭菌,冷却后得到灭菌液;
(3)将干酪乳杆菌SF-L-12接种到MRS液体培养基中,培养得到干酪乳杆菌SF-L-12接种剂;
(4)将干酪乳杆菌SF-L-12接种剂接种于灭菌液中,发酵得到发酵液;
(5)将发酵液输送至高压喷雾破壁机中进行破壁处理,得到破壁发酵液;
(6)破壁发酵液进行巴氏灭菌,得干酪乳杆菌SF-L-12发酵物。
进一步的,步骤(2)中,高压灭菌温度为120℃,时间为30min,然后冷却至35℃。
进一步的,步骤(3)中,干酪乳杆菌SF-L-12的接种量为1%,培养温度为37℃,培养时间为18h。
进一步的,步骤(4)中,接种剂的接种量为1%,发酵温度为37℃,发酵时间为24h。
进一步的,步骤(5)中,高压喷雾破壁机压力设置为60Mpa。
第三方面,本发明提供一种上述干酪乳杆菌SF-L-12发酵物在制备增加免疫球蛋白分泌量产品方面的应用。
本发明的有益效果在于:
本发明提供的被发酵底物基本采用药食同源性中药,合理配伍,安全无毒,同时使用干酪乳杆菌SF-L-12进行发酵,提高药物的吸收,协同增效,最终获得的干酪乳杆菌SF-L-12发酵物,经动物试验证明,可显著提高血液中IgG、IgA、IgM等免疫球蛋白含量,有效提高机体免疫力,无副作用,见效快。
具体实施方式
为了使本技术领域的人员更好地理解本发明中的技术方案,下面将对本发明实施例中的技术方案进行清楚、完整地描述,显然,所描述的实施例仅仅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做出创造性劳动前提下所获得的所有其他实施例,都应当属于本发明保护的范围。
本发明使用的MRS固体平板培养基包括以下组分:蛋白胨10g、牛肉粉5g、酵母粉4g、K2HPO4·7H2O 2g、柠檬酸三铵2g、葡萄糖20g、CH3COONa·3H2O 5g、MgSO4·7H2O 0.2g、MnSO4·4H2O 0.05g、吐温80 1mL、琼脂15.0g、蒸馏水1000mL。
本发明使用的MRS液体培养基包括以下组分:蛋白胨10g、牛肉粉5g、酵母粉4g、K2HPO4·7H2O 2g、柠檬酸三铵2g、葡萄糖20g、CH3COONa·3H2O 5g、MgSO4·7H2O 0.2g、MnSO4·4H2O 0.05g、吐温80 1mL、蒸馏水1000mL。
本发明使用的党参提取物,当归提取物,枸杞子提取物,五味子提取物,蒲公英提取物,茯苓提取物制备工艺为:将原料洗净后破碎成粗粒,用10倍量水煎煮2次,煎煮时间为3h和1h,合并两次滤液,静置2h后过滤得提取液。将提取液在真空度为-0.06到0.08Mpa浓缩至相对密度为1.10(70℃),然后进行喷雾干燥,最后粉碎成80目细粉即得提取物。
大枣粉、薏苡仁粉的制备方法为:将去核大枣、薏苡仁烘干至含水量5%以下,磨碎成80目的粉,待用。
苹果汁制备方法为:将苹果预煮5min后打浆,用卧螺机提取浆液中的汁液,同时用两效真空蒸发器将物料浓缩至40%,高温杀菌,待物料冷却,即得苹果汁。
本发明使用的干酪乳杆菌(Lactobacillus casei)SF-L-12为自行采集所得,已于2020年10月21日保藏于中国微生物菌种保藏管理委员会普通微生物中心,保藏编号为CGMCC No.20927,保藏地址为北京市朝阳区北辰西路1号院3号,其它相关信息已公开于中国专利CN113444675A。
实施例1一种干酪乳杆菌SF-L-12发酵物
首先准备被发酵底物,以重量份数计,净化水74份,党参提取物2份,当归提取物2份,枸杞子提取物2份,五味子提取物3份,蒲公英提取物3份,茯苓提取物3份,大枣粉3份,薏苡仁粉3份,苹果汁5份。
按照以下工艺制备:(1)首先将被发酵底物的各组分混合,得到混合浆料;(2)将混合浆料输送至灭菌容器中,120℃高压灭菌30min,然后冷却至35℃,得到灭菌液;(3)将干酪乳杆菌SF-L-12接种到MRS液体培养基中,接种量为1%,37℃下培养18h得干酪乳杆菌SF-L-12接种剂;(4)将干酪乳杆菌SF-L-12接种剂接种于灭菌液中,接种量为1%,37℃下发酵24h,得到发酵液;(5)将发酵液输送至高压喷雾破壁机中,压力设置60MPa,进行破壁处理,得到破壁发酵液;(6)破壁发酵液进行巴氏灭菌,得发酵物。
实施例2一种干酪乳杆菌SF-L-12发酵物
首先准备被发酵底物,以重量份数计,净化水64份,党参提取物3份,当归提取物4份,枸杞子提取物4份,五味子提取物5份,蒲公英提取物3份,茯苓提取物3份,大枣粉5份,薏苡仁粉4份,苹果汁4份。
按照实施例1的工艺制备发酵物。
实施例3一种干酪乳杆菌SF-L-12发酵物
首先准备被发酵底物,以重量份数计,净化水82份,党参提取物1份,当归提取物1份,枸杞子提取物1份,五味子提取物2份,蒲公英提取物2份,茯苓提取物2份,大枣粉2份,薏苡仁粉2份,苹果汁5份。
按照实施例1的工艺制备发酵物。
对比例1一种类干酪乳杆菌CICC 20251发酵物
首先准备与实施例1相同的被发酵底物。
按照以下工艺制备:(1)首先将被发酵底物的各组分混合,得到混合浆料;(2)将混合浆料输送至灭菌容器中,120℃高压灭菌30min,然后冷却至35℃,得到灭菌液;(3)将类干酪乳杆菌CICC 20251接种到MRS液体培养基中,接种量为1%,37℃下培养18h得类干酪乳杆菌CICC 20251接种剂;(4)将类干酪乳杆菌CICC 20251接种剂接种于灭菌液中,接种量为1%,37℃下发酵24h,得到发酵液;(5)将发酵液输送至高压喷雾破壁机中,压力设置60MPa,进行破壁处理,得到破壁发酵液;(6)破壁发酵液进行巴氏灭菌,得发酵物。
对比例2一种干酪乳杆菌ACCC 10640发酵物
首先准备与实施例1相同的被发酵底物。
按照以下工艺制备:(1)首先将被发酵底物的各组分混合,得到混合浆料;(2)将混合浆料输送至灭菌容器中,120℃高压灭菌30min,然后冷却至35℃,得到灭菌液;(3)将干酪乳杆菌ACCC 10640接种到MRS液体培养基中,接种量为1%,37℃下培养18h得干酪乳杆菌ACCC 10640接种剂;(4)将干酪乳杆菌ACCC 10640接种剂;接种于灭菌液中,接种量为1%,37℃下发酵24h,得到发酵液;(5)将发酵液输送至高压喷雾破壁机中,压力设置60MPa,进行破壁处理,得到破壁发酵液;(6)破壁发酵液进行巴氏灭菌,得发酵物。
测试例干酪乳杆菌SF-L-12发酵物对小鼠血清免疫球蛋白的影响
选取清洁级雄性昆明小鼠50只,体重为18~22g,经过3天的适应性饲养后,取体重相近的小鼠48只,随机分为6组,每组8只,分别将实施例1、实施例2、实施例3、对比例1、对比例2制备的发酵物灌胃,对照组用净化水,每次2ml,每天一次,正常饮食饮水,连续灌胃10天,再禁食1天,禁食后第二天摘眼球取血置于低温冰箱过夜后,3000r/min离心15min,分离上清液,采用间接酶联免疫吸附分析法测定IgA、IgM、IgG的含量。采用SPSS17.0进行统计分析,结果如表1所示。
表1 各组发酵物对小鼠血清免疫球蛋白的影响
注明:*表明p<0.05。
免疫球蛋白是体液免疫的主要抗体,血清抗体含量的高低在一定程度上反映了机体对疾病的抵抗能力。血清免疫球蛋白的测定是检查体液免疫水平常用的方法,通常检测IgG、IgA、IgM这3 类免疫球蛋白可以反映血清免疫球蛋白的总体水平。由上表可知,服用干酪乳杆菌SF-L-12发酵物能提高小鼠血清中的IgG、IgA含量、IgM含量,说明干酪乳杆菌SF-L-12发酵物能提高机体的免疫能力,其中实施例1组小鼠的血清中IgG、IgA的含量最高,与对照组相比有显著提高,IgM含量有提高但未达到统计学意义上的显著差异。
尽管通过优选实施例的方式对本发明进行了详细描述,但本发明并不限于此。在不脱离本发明的精神和实质的前提下,本领域普通技术人员可以对本发明的实施例进行各种等效的修改或替换,而这些修改或替换都应在本发明的涵盖范围内/任何熟悉本技术领域的技术人员在本发明揭露的技术范围内,可轻易想到变化或替换,都应涵盖在本发明的保护范围之内。
Claims (8)
1. 一种干酪乳杆菌SF-L-12发酵物,其特征在于,由被发酵底物经干酪乳杆菌SF-L-12发酵制得,干酪乳杆菌(Lactobacillus casei)SF-L-12已于2020年10月21日保藏于中国微生物菌种保藏管理委员会普通微生物中心,保藏编号为CGMCC No.20927,保藏地址为北京市朝阳区北辰西路1号院3号;被发酵底物包括如下重量份的组分:净化水64~82份,党参提取物1~3份,当归提取物1~4份,枸杞子提取物1~4份,五味子提取物2~5份,蒲公英提取物2~3份,茯苓提取物2~3份,大枣粉2~5份,薏苡仁粉2~4份,苹果汁4~5份。
2.如权利要求1所述的干酪乳杆菌SF-L-12发酵物,其特征在于,被发酵底物包括如下重量份的组分:净化水74份,党参提取物2份,当归提取物2份,枸杞子提取物2份,五味子提取物3份,蒲公英提取物3份,茯苓提取物3份,大枣粉3份,薏苡仁粉3份,苹果汁5份。
3.一种如权利要求1所述干酪乳杆菌SF-L-12发酵物的制备工艺,其特征在于,具体包括以下步骤:
(1)首先将被发酵底物的各组分混合,得到混合浆料;
(2)将混合浆料输送至灭菌容器中,高压灭菌,冷却后得到灭菌液;
(3)将干酪乳杆菌SF-L-12接种到MRS液体培养基中,培养得到干酪乳杆菌SF-L-12接种剂;
(4)将干酪乳杆菌SF-L-12接种剂接种于灭菌液中,发酵得到发酵液;
(5)将发酵液输送至高压喷雾破壁机中进行破壁处理,得到破壁发酵液;
(6)破壁发酵液进行巴氏灭菌,得干酪乳杆菌SF-L-12发酵物。
4.如权利要求3所述的制备工艺,其特征在于,步骤(2)中,高压灭菌温度为120℃,时间为30min,然后冷却至35℃。
5.如权利要求3所述的制备工艺,其特征在于,步骤(3)中干酪乳杆菌SF-L-12的接种量为1%,培养温度为37℃,培养时间为18h。
6.如权利要求3所述的制备工艺,其特征在于,步骤(4)中接种剂的接种量为1%,发酵温度为37℃,发酵时间为24h。
7.如权利要求3所述的制备工艺,其特征在于,步骤(5)中,高压喷雾破壁机压力设置为60Mpa。
8.一种如权利要求1所述的干酪乳杆菌SF-L-12发酵物在制备增加免疫球蛋白分泌量的产品方面的应用。
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