CN102625828A - 新型柠檬明串珠菌及将其用作发酵剂的发酵食品及组合物 - Google Patents
新型柠檬明串珠菌及将其用作发酵剂的发酵食品及组合物 Download PDFInfo
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- CN102625828A CN102625828A CN2009801604877A CN200980160487A CN102625828A CN 102625828 A CN102625828 A CN 102625828A CN 2009801604877 A CN2009801604877 A CN 2009801604877A CN 200980160487 A CN200980160487 A CN 200980160487A CN 102625828 A CN102625828 A CN 102625828A
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- lactic acid
- acid bacteria
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Abstract
本发明涉及柠檬明串珠菌CJGN34(Leuconostoc citreum CJGN34)KCTC 10974BP乳酸菌、包括将上述乳酸菌用作发酵剂而制成的泡菜在内的发酵食品以及含有上述乳酸菌的乳酸菌组合物。本发明的新型柠檬明串珠菌CJGN34提高了泡菜的整体味道品质,提高了采用冬季制备方式的传统制备方法而制得的泡菜固有的复杂的酸味和碳酸味道,从而在全年维持一定的发酵品质,特别是具有有效抑制味道品质会降低的夏季中发酵品质降低的效果,适用于包括泡菜的发酵食品,可用作发酵剂;或与载体或添加剂等进行混合,用作片剂、胶囊等药品以及食品益生菌;或以一定量混合到用于化妆品的原料中,从而能够应用于药品、食品、饲料、化妆品等多个领域中并可根据所属技术领域中通常的方法而使用。
Description
技术领域
本发明涉及新型柠檬明串珠菌菌株和将其用作发酵剂的发酵食品以及包括其的组合物。更具体地,涉及能够全年保持冬季泡菜的味道品质水平的发酵风味,并且特别能够有效抑制夏季发酵品质降低的新型柠檬明串珠菌菌株及将其用作发酵剂的发酵食品及包括其的组合物。
背景技术
泡菜是将各种蔬菜用盐腌渍并混合,自然发酵而制成的我国具有代表性的蔬菜发酵食品。其中,具有代表性的泡菜是以白菜作为主要材料的过冬泡菜(Kimjang Kimchi),其是将晚秋或初冬收获的白菜、萝卜用盐腌渍,并以大蒜、生姜、葱等作为配料混合后浸渍,然后埋于地下,可以食用一冬(Cheigh和Park,关于泡菜的生物化学、微生物及营养层面的研究,食品科学和营养学评论,卷134,1981,页码107~127(Cheigh and Park,Biochemical,microbiological and nutritional aspects of Kimchi,Crit.Rev.FoodSci.Nutr.Vol34,1981,pages107-127))。如上所述,过冬泡菜是利用食品保存法中的腌渍和发酵法,在不能够产蔬菜的整个冬季,将蔬菜在低温下保存,并且作为碳源、氨基酸、维生素的供给源来利用的低温发酵储藏食品,通过源自原料固有的乳酸菌进行发酵和熟成,从而具有丰富的发酵风味。
但是到了现代,由于品种改良和栽培技术的发达,在全年生产、储藏以泡菜为首的原料,从而使得在过冬季节之外也能够制备泡菜,并且由于冰箱和泡菜专用冰箱的普及,无论什么时候都能够在与冬天一样的低温下发酵储藏,这与传统的过冬白菜泡菜具有差异。
泡菜品质基本上受包括全年上市的白菜在内的多种原料品质的影响很大,从使泡菜商品化的企业的立场来看,这一点在给消费者提供均一品质方面成为巨大的绊脚石。特别是夏季(6月-8月)左右生产的泡菜,即使对其进行与冬季相同的发酵熟成管理,由于原料的品质问题和发酵熟成风味的降低,因此与其它时期的品质差异很大,发生来自消费者关于品质降低的多种投诉,这是相关业界众所周知的事实。
为了解决这样的问题,从现有的技术来看,添加多种微生物,主要添加乳酸菌作为发酵剂(starter),从而提高泡菜的品质(韩国授权专利第10-0536108号、第10-0181009号、第10-0330674号等)。但现有的方法不是通过明确研究全年发生的品质变化而改善问题,而是单纯为了改善品质而适用了发酵剂,并且在感官品质方面,也没有仅对未指定的非特定时期的整体味道品质差异进行确认等的、集中于夏季的品质改善并分析品质降低原因来改善问题的事例。
另一方面,一直以来将作为泡菜乳酸菌的泡菜明串珠菌(Leuconostoc kimchiii)、柠檬明串珠菌(Leuconostoc citreum)、肠膜明串珠菌(Leuconostoc mesenteroides)、卡西口米塔塔姆明串珠菌(Leuconostoc gasicomitatum)、乳明串珠菌(Leuconostoc lactis)、植物乳杆菌(Lactobacillus plantarum)、沙克乳杆菌(Lactobacillus sakei)等进行单独或组合使用,用于制备多种发酵食品。
其中,柠檬明串珠菌(Leuconostoc citreum)菌株与泡菜的熟成相关,特别是已知其具有利用白糖生产高分子粘质物的葡聚糖的能力(韩国授权专利第10-0718344号),已指出其对异位性皮炎具有治疗及预防的效果(韩国申请专利第10-2005-0041572号)。与此相关的还有:韩国授权专利第10-0814665号中公开了生产葡聚糖的柠檬明串珠菌S5菌株及利用其制备含葡聚糖发酵物的制备方法,韩国授权专利第10-080053号公开了6.具有生产甘露醇能力的柠檬明串珠菌及利用其生产甘露醇的方法。
由此,本发明人对全年白菜泡菜的发酵模式进行分析,对影响发酵品质变化的乳酸菌群落的变化和有机酸的变化进行了检验,通过这些知道了对于改善和稳定全年品质方面,冬季味道好的时期所分离出的乳酸菌对制备全年一定的发酵品质的泡菜具有有效的作用。能够全年保持冬季泡菜的味道品质水平的发酵风味,特别是对有效抑制夏季的发酵品质降低的新型明串珠菌属乳酸菌菌株进行分离、鉴定,从而完成了本发明。
发明内容
本发明要解决的技术问题
本发明的目的在于提供一种新型明串珠菌属乳酸菌菌株,将其用作泡菜的发酵剂,从而能够全年保持冬季泡菜的味道品质水平的发酵风味,特别是能够有效抑制夏季的发酵品质降低。
本发明的另一目的在于,提供将上述新型明串珠菌属乳酸菌菌株用作发酵剂的发酵食品。
本发明的再一目的在于,提供含有上述新型明串珠菌属乳酸菌菌株的菌体组合物。
技术方案
为了实现上述目的,本发明提供柠檬明串珠菌CJGN34(Leuconostoc citreum CJGN34)(保藏单位:韩国典型培养物保藏中心,保藏日期:2006.07.27,保藏编号:KCTC 10974BP)。
并且,本发明提供一种包含上述柠檬明串珠菌CJGN34作为有效成分的用于食品发酵的发酵剂组合物。
并且,本发明提供包括使用上述发酵剂组合物制备的泡菜的发酵食品。
并且,本发明提供一种包含上述柠檬明串珠菌CJGN34作为有效成分的乳酸菌组合物。
本发明中,有效成分是指包括有主要成分,其为根据内在的药理作用,有望直接或间接地表现该药品功效的物质或物质组合(包括没有示出药理学活性成分等的生药等)。
下面,更加详细地说明本发明。
本发明的柠檬明串珠菌CJGN34(Leuconostoc citreum CJGN34)的特征为,其是由泡菜中分离并鉴定的柠檬明串珠菌的新型菌株。上述菌株虽然是从冬季过冬季节时制备的白菜泡菜中分离得到,但相关菌株的分离范围并不限定于此。
为了对微生物进行鉴定和分类,对本发明的柠檬明串珠菌GJGN34进行了有关16S rRNA碱基序列分析,结果为:与柠檬明串珠菌标准菌株(Leuconostoc citreum KCTC 3526T,GenBank登录号AF111948)显示出了最高的同源性(99.7%),显示出了与柠檬明串珠菌最高的分子系统学上的亲缘关系。并且,将上述微生物鉴定为柠檬明串珠菌(Leuconostoc citreum),并命名为柠檬明串珠菌CJGN34,于2006年7月27日保藏于韩国典型培养物保藏中心(保藏编号为KCTC 10974BP)。
本发明的柠檬明串珠菌CJGN34为革兰氏阳性菌,是在有氧条件和厌氧条件下均能够生长的兼性厌氧菌(facultive anaerobe),并且不形成孢子,无运动性,细胞形态为单一或双球菌。采用本技术领域常规方法对柠檬明串珠菌CJGN34的更加具体的形态以及生理学特性进行分析的结果如下表1所示。
表1
形态及生理、生物化学特性 | 结果 |
形态(Morphology) | 球菌(Coccus) |
运动性(Motility) | - |
孢子(Spore) | - |
过氧化氢酶 | - |
同型与异型发酵 | 兼性异型发酵 |
15℃下的繁殖 | + |
45℃下的繁殖 | - |
通过葡萄糖代谢的葡聚糖生成 | + |
在3%的NaCl中的繁殖 | + |
厌氧性繁殖 | + |
利用葡萄糖生成CO2 | + |
糖发酵特性 | |
丙三醇 | - |
赤藓糖醇 | - |
D-阿拉伯糖 | - |
L-阿拉伯糖 | + |
核糖 | - |
D-木糖 | - |
L-木糖 | - |
戊五醇 | - |
木糖苷 | - |
半乳糖 | + |
D-葡萄糖 | + |
D-果糖 | + |
D-甘露糖 | + |
L-山梨糖 | - |
鼠李糖 | - |
半乳糖醇 | - |
肌醇 | - |
甘露醇 | + |
山梨醇 | - |
D-甘露糖苷 | - |
D-葡萄糖苷 | + |
葡萄糖胺 | + |
苦杏仁苷 | + |
熊果苷 | + |
七叶苷 | + |
水杨苷 | + |
纤维二糖 | + |
麦芽糖 | + |
乳糖 | - |
蜜二糖 | - |
蔗糖 | + |
海藻糖 | + |
菊糖 | - |
松三糖(melizitose) | - |
D-蜜三糖 | - |
美沙酮(amidon) | - |
糖原 | - |
木糖醇 | - |
龙胆二糖 | - |
D-松二糖 | + |
D-来苏糖 | - |
D-塔格糖 | - |
D-岩藻糖 | - |
L-岩藻糖 | - |
D-阿拉伯糖醇 | - |
L-阿拉伯糖醇 | - |
葡萄糖酸盐 | - |
2-葡萄糖酸盐 | - |
5-葡萄糖酸盐 | - |
+:阳性反应
-:阴性反应
为了长时间稳定地保存本发明的柠檬明串珠菌CJGN34,优选在混合有一定量的丙三醇的水溶液的保存溶液中,使菌体散开,于-70℃条件下保存,或者将其悬浮在灭菌的10%脱脂乳中进行冷冻干燥。
此外,本发明的柠檬明串珠菌CJGN34作为由天然发酵的泡菜中分离的乳酸菌,分类为对人无害的GRAS(一般认为安全,Generally recognized as safe)菌株,独自地进行了与安全性相关的明胶液化反应检测、苯丙氨酸脱氨生成检测、氨生成检测、溶血性实验检测等,结果为:在明胶液化反应检测、苯丙氨酸脱氨生成检测、氨生成检测、溶血性实验检测中均被确认为阴性,从而被确认为能够用于食品的安全的菌。
本发明的柠檬明串珠菌CJGN34在包括泡菜在内的发酵食品中用作发酵剂时,有助于提高整体的味道品质,提高采用冬季制备方式的传统制备方法而制得的泡菜固有的复杂酸味和碳酸味道,从而在全年稳定地维持泡菜的品质,特别有助于有效抑制味道品质会降低的夏季中发酵品质降低。并且,对含有上述菌株的菌体进行培养后,进行回收能够制备菌体组合物,这样的组合物能够与饲料中作为原料的谷物粉进行混合,用作饲料益生菌,或混合载体或添加剂等而用作片剂、胶囊等药品以及食品益生菌,或以一定量混合到用于化妆品的原料中,从而能够应用于药品、食品、饲料、化妆品等多个领域中并可根据所属技术领域中通常的方法而使用。
此外,本发明的柠檬明串珠菌CJGN34能够用于乳酸菌药物组合物。这样的情况下,可以制备成本技术领域中公知的常规药剂学剂型。上述药品优选为口服剂型。例如可制成液体剂、悬浮剂、散剂、颗粒剂、片剂、胶囊剂、丸剂或浸膏剂等口服给药剂型。
制备成上述各个剂型时,可以加入制备各个剂型所需的并在药剂学上可接受的载体或添加剂。制备成具有典型的口服给药剂型时,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一种以上,作为添加剂可以使用选自香料、维生素类以及抗氧化剂中一种以上。
上述载体以及添加剂只要是药剂学上可接受的均可,优选地,作为稀释剂具体有乳糖、玉米淀粉、大豆油、微晶纤维素或甘露醇,作为润滑剂有硬脂酸镁或滑石粉,作为粘合剂有聚乙烯吡咯烷酮或羟丙基纤维素。此外,优选地,作为崩解剂有羧甲基纤维素钙、羧基乙酸淀粉钠、波拉克林钾或交联聚维酮,作为甜味剂有白糖、果糖、山梨醇或阿斯巴甜,作为稳定剂有羧甲基纤维素钠、β-环糊精、白蜡或黄原胶,作为防腐剂有对羟基苯甲酸甲酯、对羟基苯甲酸丙酯或山梨酸钾。
此外,为了增加味觉,除了上述成分之外,作为公知的添加剂,还可以混合使用梅花香、柠檬香、菠萝香、香草香等天然香料;天然果汁、叶绿酸、类黄酮等天然色素;果糖、蜂蜜、糖醇、白糖等甜味成分;或柠檬酸、柠檬酸钠等酸味剂。
关于这种制剂化方法以及制剂化时所需的载体以及添加剂在《雷明顿的制药科学》(Remington′s Pharmaceutical Sciences)(第19版,1995)中有详细记载。
并且,本发明的柠檬明串珠菌CJGN34可以作为乳酸菌食品组合物使用。上述食品组合物不仅包括保健功能性食品,还包括人们每日经常摄取的一般食品。用作保健功能性食品的情况下,可以和食品学上可接受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、浸膏剂、茶、果冻或饮料等。关于上述食品学上可接受的载体或添加剂,根据需要制备的剂型可以选择使用在本技术领域可接受的公知的任一载体或添加剂。
并且,由于本发明的柠檬明串珠菌CJGN34可以含有叫作葡聚糖(dextran)的高含湿性的聚合物,因此可以用作乳酸菌化妆品组合物。当用于本发明涉及的化妆品组合物的情况下,可以制备成该化妆品技术领域公知的常规剂型的多种化妆品。制备成上述各种剂型时,可以添加制备各个剂型所需的制备化妆品时可接受的载体或添加剂来制备。
并且,本发明的柠檬明串珠菌CJGN34还可以作为饲料添加剂或饲料组合物来使用。
作为饲料添加剂使用的情况下,可以将上述组合物制备成20-90%的高浓缩液或粉末或颗粒形态。上述饲料添加剂还可以包括选自柠檬酸、富马酸、己二酸、乳酸、苹果酸等有机酸;或磷酸钠、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,可以一起包括常规饲料成分。
所述饲料添加剂以及饲料还可以包括谷类,例如有粉碎或破碎的小麦、燕麦、大麦、玉米以及米;植物性蛋白饲料,例如有以油菜、大豆以及葵花籽为主要成分的饲料;动物性蛋白饲料,例如有血粉、肉粉、骨粉以及海鲜粉;糖分以及乳制品,例如有由各种奶粉以及乳清粉构成的干燥成分等,除此之外,还可以包括营养补充剂、消化和吸收提高剂以及生长促进剂等。
上述饲料添加剂可以单独喂养动物,或者是和食用载体中的其它饲料添加剂组合喂养。并且,上述饲料添加剂可以作为追肥(topdressing)和这些动物饲料直接混合,或者是不与饲料一起而另外制备成口服剂型的方式容易地对动物进行喂养。在上述饲料添加剂不与动物饲料一起喂养的情况下,可以和本技术领域已知的药剂学上允许的食用载体组合使用,制备成能够立刻释放的剂型或缓释的剂型。这些食用载体可以是固体或液体,例如有玉米淀粉、乳糖、蔗糖、大豆碎片、花生油、橄榄油、芝麻油以及丙二醇。当使用固体载体的情况下,作为饲料添加剂可以是片剂、胶囊剂、散剂、糖锭剂或含糖片剂或微分散性形态的追肥。当使用液体载体的情况下,饲料添加剂可以是明胶软胶囊剂、糖浆剂、悬浮液、乳液剂或溶液剂等剂型。
上述饲料中可以包括任意一种满足动物食欲的通常使用的含有蛋白质的有机谷粉。这种含有蛋白质的谷粉通常主要由玉米、大豆谷粉、或玉米/大豆谷粉混合物构成。
此外,上述饲料添加剂以及饲料还可以含有辅助剂,例如有保存剂、稳定剂、润湿剂或乳化剂、溶液促进剂等。上述饲料添加剂可以以浸透、喷雾或混合的方式来添加到动物饲料中使用。
本发明的饲料或饲料添加剂可以应用在包括哺乳类、家禽、以及鱼类的多数动物的食物中。作为上述哺乳类,不仅可以用于猪、牛、羊、山羊、实验用啮齿动物以及实验用啮齿动物,还可以用于宠物(例如狗、猫)等。作为上述家禽类,可以用于鸡、火鸡、鸭、鹅、雉、以及鹌鹑等,作为上述鱼类可以用于鳟鱼等,但并不限于这些。
有益效果
如上面所述,本发明的新型柠檬明串珠菌CJGN34在包括泡菜的发酵食品中用作发酵剂时,提高了泡菜的整体味道品质,提高了采用冬季制备方式的传统制备方法而制得的泡菜固有的复杂酸味和碳酸味道,从而在全年稳定地维持泡菜的品质,特别是具有有效抑制味道品质会降低的夏季中发酵品质降低的效果,并且,本发明的新型柠檬明串珠菌CJGN34制备成菌体组合物,从而适用于包括上述泡菜的发酵食品,可用作发酵剂;或混合载体或添加剂等而用作片剂、胶囊等药品以及食品益生菌;或以一定量混合到用于化妆品的原料中,从而能够应用于药品、食品、饲料、化妆品等多个领域中并可根据所属技术领域中通常的方法而使用。
附图说明
图1为示出制备泡菜不同时期的整体味道平均分数的图表。
图2为示出制备泡菜不同时期的发酵菌落的变化比率的图表。
具体实施方式
下面根据下述实施例更具体说明本发明。但是,这些实施例只是为了有助于本发明的理解,本发明的范围并不能以任何方式由这些实施例来限定。
实施例1:通过对全年泡菜品质变化的检验得到改善点及CJGN34菌株的分离
为了掌握品质变化原因并进行改善,通过对全年泡菜的品质变化进行检验,从而确认了全年泡菜品质的变化。
泡菜使用了根据CJ第一制糖的制备方法制备的泡菜,将泡菜进行区分。将3月至5月15日制备的泡菜称为春季泡菜,将5月15日至8月制备的泡菜称为夏季泡菜,将9月至11月15日制备的泡菜称为秋季泡菜,将11月16日至2月底制备的泡菜称为冬季泡菜。
采用5分评分法对各个时期泡菜的感官品质进行分析确认,得到的味道品质的结果为:确认了下述特征,夏季泡菜的整体味道的得分为3.5水平,这与其他季节制得的泡菜的整体味道平均维持在3.8-4.0左右相比要低。上述结果如图1所示。
为了分析发酵微生物对如上所述的全年味道品质变化的影响,采用崔等(崔等人.,泡菜的微生物种群动态,发酵的卷心菜,FEMS微生物通讯.,卷.257,2006,第262-267页)(Choi et.al.,Microobial population dynamics of kimchi,a fermented cabbage,FEMS microbial.Lett.,Vol.257,2006,pages 262-267)的方法,通过对16SrRNA的分析进行了鉴定。其结果为:采用传统方法腌渍了过冬泡菜,味道品质最为优异的冬季制备的泡菜进行绝对异型发酵,从而检测出利用糖生成除乳酸之外的多种酸并产生气体的明串珠菌属为80%以上,确认了其对冬季泡菜的复杂的发酵风味和碳酸味起到主要作用。相反,发生发酵品质降低的夏季制备的泡菜进行偏异型发酵或同型发酵,确认了在一般的环境下利用糖仅生成乳酸而不产生气体的作为乳杆菌属(Lactobacillus)的沙克乳杆菌(Lactobacillus sakei)、弯曲乳杆菌(Lactobacillus curvatus)、植物乳杆菌(Lactobacillus plantarum)等菌的比率为50%以上,这与其他季节制备的泡菜相比升高了,能够预料到这种发酵菌落的变化会对夏季泡菜中单纯的酸味和碳酸味道变弱造成影响。上述结果如图2所示。
为了更加明确这样的发酵菌落的变化对泡菜发酵风味的影响,采用HPLC对与微生物分析中使用的样品相同的样品进行了有机酸分析。所述微生物分析为采用上述崔等人的方法而开始发酵的总酸度为0.4%以上的微生物分析。有机酸作为乳酸菌消耗糖而生成的主要代谢产物,在偏异型发酵,同型发酵的情况下,主要产生乳酸,在绝对异型发酵的情况下,产生乳酸,同时还产生醋酸、二氧化碳等发酵产物,因此有机酸是可判断发生了什么种类的发酵的主要指标物质。其结果如表2所示。
如表2所示,对乳酸和醋酸的摩尔浓度的比率进行计算的结果,可确认乳酸与醋酸的比率在全年维持在1.3-1.6左右,与此相反,夏季制备的泡菜为2.2-2.8,乳酸的比率升高了。这是由于发酵乳酸菌的改变,从而在发酵特性上显示出差异,具体表现为在夏季时的同型发酵的比率升高,能够确认上述特征为夏季品质降低的原因。
表2
制备时期 | 样品编号 | 乳酸/醋酸浓度比率(mM) |
春季 | 春季泡菜1 | 1.5±0.08 |
春季 | 春季泡菜2 | 1.6±0.11 |
夏季 | 夏季泡菜1 | 2.8±0.13 |
夏季 | 夏季泡菜2 | 2.2±0.09 |
夏季 | 夏季泡菜3 | 2.4±0.25 |
夏季 | 夏季泡菜4 | 2.8±0.13 |
秋季 | 秋季泡菜1 | 1.4±0.09 |
冬季 | 冬季泡菜1 | 1.4±0.25 |
冬季 | 冬季泡菜2 | 1.5±0.39 |
冬季 | 冬季泡菜3 | 1.3±0.02 |
冬季 | 冬季泡菜4 | 1.3±0.09 |
通过上述结果,本发明人将冬季味道好的时期所分离出的乳酸菌用作发酵剂,从而在全年制备了具有一定发酵品质的泡菜,特别是为了有效抑制夏季泡菜品质的降低,从冬季制备的泡菜中分离了约500来个菌株,从其中选择是绝对异型发酵菌且从冬季泡菜的优势种的菌株中分离出符合目标的新型乳酸菌。
实施例2微生物柠檬明串珠菌CJGN34菌株的分离及鉴定
如上述实施例1所述,对全年泡菜品质变化的原因进行分析,通过这一分析,将从冬季制备的熟成的味道好的泡菜中分离出的乳酸菌菌株命名为CJGN34,将上述菌株涂布在含有1.5%琼脂(agar)的MRS固体培养基(Difco,USA)上,于30℃下培养24小时后,用接种环(loop)挑取已经被确认为是纯分离的菌落,将其在MRS液体培养基(Difco,USA)中,于30℃下培养18-24小时。
然后,对CJGN34菌株的形态以及生理学特性采用金等(金等,仁海明串珠菌新种,一种由泡菜中分离出的乳酸菌,国际系统进化微生物学杂志,第53卷,2003年7月,第1123-1126页)(Kim et.al.,Leuconostoc inhae sp.nov.,a lactic acid bacterium isolated from kimchi,International Journal of Systematic and Evolutional Microbiology,Volume53,July 2003,pages 1123-1126)的方法,并且使用API50CH以及API50CHL试剂盒(生物梅里埃公司产品)来进行了确定。将最后确认的CJGN34菌株的形态及生理学特性整理在上述表1中。
为了对乳酸菌进行鉴定及分类,分析了16S rRNA基因的碱基序列。采用金等(金等,泡菜乳酸菌新品种,来源于泡菜的新品种,国际系统与进化微生物学杂志,第50卷,2000年9月,第1915-1919页)(Kim et.al.,Leuconostoc kimchii sp.nov.,a new species from kimchi.International Journal of Systematic and Evolutional Microbiology,Volume50,September 2000,pages 1915-1919)的方法对16S rRNA基因的碱基序列进行确定及分析。
对于16S rRNA碱基序列进行分析的结果,本发明的柠檬明串珠菌CJGN34菌株与柠檬明串珠菌标准菌株(Leuconostoc citreum KCTC 3526T,GenBank登录号AF111948)显示出了最高的同源性(99.7%),从而被鉴定为柠檬明串珠菌(Leuconostoc citreum),并且将其命名为柠檬明串珠菌CJGN34,于2006年7月27日保藏于韩国典型培养物保藏中心(保藏编号为KCTC 10974BP)。
实施例3:柠檬明串珠菌CJGN34菌株的安全性评价
为了对上述实施例1中分离得到的菌株进行安全性评价,根据韩国生物企业协会的组合标准中提出的安全性评价试验方法进行了溶血现象检测、明胶液化反应检测、有害代谢产物(氨)生成确认、以及苯丙氨酸脱氨检测。结果如下表3所示。
表3柠檬明串珠菌CJGN34的安全性评价结果
根据上述结果,确认了柠檬明串珠菌CJGN34(Leuconostoc citreum CJGN34)对于明胶液化反应、有害代谢产物(氨)生成、苯丙氨酸脱氨的生成、溶血现象检测中均确认为阴性。并且还确认了柠檬明串珠菌CJGN34是能够用于人体的安全的菌株。
实施例4:柠檬明串珠菌CJGN34作为发酵剂而制得的泡菜的特征
为了使本发明的柠檬明串珠菌CJGN34适用于发酵剂,制备了菌株的培养液或菌体组合物,培养液和菌体组合物采用下述方法制得。
培养液是将10g葡萄糖、5g酵母提取物和5g柠檬酸二铵(diammonium citrate)溶于水中,经过灭菌后接种菌,在25-37℃的温度下培养8-24小时而制得。菌体组合物按照如下方法制备:为了生产菌,用氨气气体将MRS液体培养基(Difco)的pH调节至5.5-7.0,并于30℃下培养约18小时,然后进行离心分离回收得到菌体。使用作为冷冻保护剂的5%的糊精和10%脱脂牛奶,将回收的菌体于-40℃以下进行冷冻后,在30-37℃下进行干燥,然后将菌体用搅拌器研磨后将其打碎,并且将打碎的活菌配成作为目标的菌数,为了保管,将其和适量的葡萄糖、乳糖、脱脂牛奶等赋形剂进行混合放入密封的铝袋中进行包装。
在制备泡菜时,将按照上述方法制备的培养液或菌体组合物以泡菜整体重量的0.001%-3%的比例进行添加使用。泡菜采用CJ第一制糖的制备方法进行制备发酵,采用相同的材料和配比制备,对将CJGN34作为发酵剂添加而制备的发酵泡菜和没有添加发酵剂的自然发酵的泡菜的感官品质及理化特性进行了分析。由于泡菜的感官品质及理化特性根据熟成度的不同会发生差异,因此以总酸度为基准,将熟成程度控制在0-0.05%内并进行了调查。如上制备的泡菜的感官品质的分析结果如下表4所示。
表4
如表4中所示,根据制备时期的不同,自然发酵泡菜的整体味道的喜好度由夏季泡菜3.59到冬季泡菜4.1,味道品质的偏差很大,与此相比,使用CJGN34作为发酵剂的情况下,由夏季泡菜的3.85到冬季泡菜的4.05,可以确认味道品质的偏差缩小了,由此能够看出在制备期望的全年具有均一的发酵品质的泡菜时,起到有效的作用。特别是夏季制备的自然发酵泡菜的酸味喜好度、碳酸感喜好度、余味喜好度降低,使整体味道喜好度降至3.5的分值带,与此相反CJGN34发酵剂泡菜的情况下,酸味喜好度、余味喜好度显示出了95%的显著性差异,碳酸感喜好度显示出了90%的显著性差异,并且整体味道喜好度也显示出了95%的显著性差异,因此可以确认作为目的的能够有效地抑制夏季发酵品质降低。
此外,在异型发酵和同型发酵中,对能够确认发生了什么种类的发酵的主要指标物质,即对乳酸和醋酸的摩尔浓度比率确认的结果,可确认夏季中升高的乳酸生成比率降低了,CJGN34发酵剂能够正常优化,从而抑制降低夏季味道品质的同型发酵或偏异型发酵乳酸菌生长,由此可以再次确认有效抑制泡菜的发酵品质。
上述各个制备时期泡菜的有机酸分析结果如下表5所示。
表5
制备时期 | 样品 | 乳酸/醋酸浓度比率(mM) |
夏季 | 自然发酵泡菜 | 2.4±0.3 |
夏季 | 发酵剂泡菜 | 1.5±0.1 |
冬季 | 自然发酵泡菜 | 1.4±0.2 |
冬季 | 发酵剂泡菜 | 1.3±0.3 |
春季 | 自然发酵泡菜 | 1.5±0.1 |
春季 | 发酵剂泡菜 | 1.5±0.1 |
秋季 | 自然发酵泡菜 | 1.4±0.1 |
秋季 | 发酵剂泡菜 | 1.4±0.1 |
实施例5:将柠檬明串珠菌CJGN34菌株作为发酵剂而制得的发酵食品及腌制食品
为了使本发明的柠檬明串珠菌CJGN34菌株适用于除泡菜以外的发酵食品发酵剂,采用与实施例4相同的方法制备了菌株培养液或菌体组合物,将其作为发酵剂制备了发酵食品。发酵食品可以使用黄瓜、萝卜、胡萝卜、洋葱、大蒜、辣椒、白菜等多种原料制备,可以应用于腌制萝卜等腌制食品,从而可以制备发酵腌制萝卜。
实施例6:含有柠檬明串珠菌CJGN34(Leuconostoc citreu m CJGN34)的菌体组合物的制备及应用
为了将上述实施例1中鉴定的柠檬明串珠菌CJGN34(Leuconostoc citreum CJGN34)作为药品、食品、饲料、饲料添加剂或化妆品的原料使用,进行了大量生产,并且采用与实施例4相同的方法制备了菌体组合物。
这样制得的菌体组合物能够与作为饲料原料的谷粉进行混合用作饲料用益生菌;或与载体或添加剂等进行混合而用作片剂、胶囊等药品以及食品益生菌;或以一定量混合到用于化妆品的原料中,从而能够应用于药品、食品、饲料、化妆品等多个领域中并可根据所属技术领域中通常的方法而使用。
Claims (7)
1.柠檬明串珠菌CJGN34(Leuconostoc citreum CJGN34)KCTC 10974BP。
2.包含柠檬明串珠菌CJGN34(Leuconostoc citreum CJGN34)KCTC 10974BP作为有效成分的用于食品发酵的发酵剂组合物。
3.由权利要求2所述的用于食品发酵的发酵剂组合物制备的发酵食品。
4.由权利要求2所述的用于食品发酵的发酵剂组合物制备的腌制食品。
5.由权利要求2所述的用于食品发酵的发酵剂组合物制备的泡菜。
6.包含柠檬明串珠菌CJGN34(Leuconostoc citreum CJGN34)KCTC 10974BP作为有效成分的乳酸菌药物组合物。
7.包含柠檬明串珠菌CJGN34(Leuconostoc citreum CJGN34)KCTC 10974BP作为有效成分的乳酸菌食品组合物。
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JP2009-190443 | 2009-08-19 | ||
JP2009190443A JP5028458B2 (ja) | 2009-07-16 | 2009-08-19 | 新規のロイコノストックシトレウム及びこれをスタータで使った醗酵食品及び組成物{NovelLeuconostoccitreum,fermentationfoodsandcompositionscomprisingthesame} |
PCT/KR2009/005139 WO2011007924A1 (ko) | 2009-07-16 | 2009-09-10 | 신규한 류코노스톡 시트리움 및 이를 스타터로 사용한 발효식품 및 조성물 |
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CN103070378A (zh) * | 2013-02-17 | 2013-05-01 | 哈尔滨伟平科技开发有限公司 | 混合菌种发酵制作酸黄瓜的方法 |
CN104256459A (zh) * | 2014-10-20 | 2015-01-07 | 哈尔滨派特纳生物科技开发有限公司 | 一种发酵橄榄的制作方法 |
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CN110177866A (zh) * | 2016-07-15 | 2019-08-27 | Cj第一制糖株式会社 | 气体产生量低的肠膜明串珠菌cjlm627菌株以及使用该菌株制作泡菜的方法 |
CN110249042A (zh) * | 2016-07-15 | 2019-09-17 | Cj第一制糖株式会社 | 气体产生量低的肠膜明串珠菌cjlm181菌株以及使用该菌株制作泡菜的方法 |
CN110249042B (zh) * | 2016-07-15 | 2023-06-20 | Cj第一制糖株式会社 | 气体产生量低的肠膜明串珠菌cjlm181菌株以及使用该菌株制作泡菜的方法 |
CN110177866B (zh) * | 2016-07-15 | 2023-06-20 | Cj第一制糖株式会社 | 气体产生量低的肠膜明串珠菌cjlm627菌株以及使用该菌株制作泡菜的方法 |
CN108285878A (zh) * | 2017-11-24 | 2018-07-17 | 延边大学 | 高产甘露醇的柠檬明串株菌hm1菌株及发酵制备甘露醇的方法 |
CN108285878B (zh) * | 2017-11-24 | 2022-10-28 | 延边大学 | 高产甘露醇的柠檬明串株菌hm1菌株及发酵制备甘露醇的方法 |
WO2021218130A1 (zh) * | 2020-04-29 | 2021-11-04 | 中国农业大学 | 柠檬明串珠菌及其在沉淀淀粉浆液中的应用 |
CN112715890A (zh) * | 2020-12-25 | 2021-04-30 | 贵州统之源食品有限公司 | 一种固定化泡菜发酵剂及其应用 |
CN112715890B (zh) * | 2020-12-25 | 2024-01-09 | 贵州统之源食品有限公司 | 一种固定化泡菜发酵剂及其应用 |
Also Published As
Publication number | Publication date |
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US10123558B2 (en) | 2018-11-13 |
WO2011007924A1 (ko) | 2011-01-20 |
DK2455451T3 (da) | 2017-11-27 |
EP2455451A4 (en) | 2014-05-07 |
US20170006905A1 (en) | 2017-01-12 |
JP2011019504A (ja) | 2011-02-03 |
EP2455451B1 (en) | 2017-08-30 |
ES2649338T3 (es) | 2018-01-11 |
HK1173744A1 (en) | 2013-05-24 |
CN102625828B (zh) | 2013-11-06 |
US20120100256A1 (en) | 2012-04-26 |
JP5028458B2 (ja) | 2012-09-19 |
EP2455451A1 (en) | 2012-05-23 |
KR101099924B1 (ko) | 2011-12-28 |
KR20110007478A (ko) | 2011-01-24 |
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