CN113950247A - 组合物 - Google Patents
组合物 Download PDFInfo
- Publication number
- CN113950247A CN113950247A CN202080040762.8A CN202080040762A CN113950247A CN 113950247 A CN113950247 A CN 113950247A CN 202080040762 A CN202080040762 A CN 202080040762A CN 113950247 A CN113950247 A CN 113950247A
- Authority
- CN
- China
- Prior art keywords
- bifidobacterium
- lactoferrin
- composition
- bacteria
- present
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
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Abstract
本发明的课题在于,提供进一步促进双歧杆菌属细菌的增殖的技术。通过组合乳铁蛋白和/或乳铁蛋白水解物与人乳寡糖,从而协同地发挥双歧杆菌属细菌的增殖促进效果。优选前述人乳寡糖包含选自由2’‑岩藻糖基乳糖、3‑岩藻糖基乳糖和乳糖二岩藻四糖组成的组中的一种或二种以上。而且优选前述双歧杆菌属细菌为选自由短双岐杆菌、婴儿双歧杆菌和长双歧杆菌组成的组中的一种或二种以上。
Description
技术领域
本发明涉及含有乳铁蛋白和/或乳铁蛋白水解物、且含有人乳寡糖的组合物。
背景技术
已知婴幼儿的肠道菌群的双歧杆菌属细菌(以下,也记作双歧杆菌)最有优势,认为其与婴儿的健康的维持密切相关。因此,开发促进双歧杆菌的增殖的食品时,在维持婴幼儿的健康上是重要的。
以双歧杆菌的增殖为促进因子,乳蛋白中已知有乳铁蛋白(非专利文献1),牛乳铁蛋白被广泛用于育儿用粉乳、补充剂等。另外,对于乳铁蛋白分解物(源自乳铁蛋白的肽),也报道了具有双歧杆菌增殖促进作用(非专利文献2、专利文献1)。
另外,关于作为人的初乳中所含的多种寡糖的通称的人乳寡糖(以下,也记作HMO),也报道了促进双歧杆菌的增殖(专利文献2)。
现有技术文献
专利文献
专利文献1:美国专利申请公开第2015-139955号说明书
专利文献2:美国专利申请公开第2012-294840号说明书
非专利文献
非专利文献1:Rahman,M.M.et.al.,Int.J.Food Sci.Tech.45:453-458,2010
非专利文献2:H.Oda et.al.,Appl.Environ.Microbiol.,79(6)1843-1849,2013
发明内容
发明要解决的问题
本发明的课题在于,提供进一步促进双歧杆菌的增殖的技术。
用于解决问题的方案
本发明人等为了解决上述课题而进行了深入研究,结果发现:通过组合乳铁蛋白和/或乳铁蛋白水解物、与人乳寡糖,从而协同地发挥双歧杆菌属细菌的增殖促进效果,至此完成了本发明。
即,本发明的第一方式为一种组合物,其含有乳铁蛋白和/或乳铁蛋白水解物、且含有人乳寡糖。
本方式中,优选人乳寡糖相对于乳铁蛋白和/或乳铁蛋白水解物的质量比为1/100000以上且1/10以下。
本方式中,优选乳铁蛋白和/或乳铁蛋白水解物相对于组合物整体的含量为0.001质量%以上且低于100质量%。
本方式中,优选人乳寡糖相对于组合物整体的含量为0.001质量%以上且低于100质量%。
本方式中的人乳寡糖优选包含选自由2’-岩藻糖基乳糖、3-岩藻糖基乳糖和乳糖二岩藻四糖组成的组中的一种或二种以上。
本方式的组合物可以优选用于双歧杆菌属细菌增殖促进用途。此处,前述双歧杆菌属细菌优选为选自由短双岐杆菌、婴儿双歧杆菌和长双歧杆菌组成的组中的一种或二种以上。
另外,本发明的第二方式为一种经口组合物,其含有:乳铁蛋白和/或乳铁蛋白水解物、人乳寡糖、和双歧杆菌属细菌。
本方式中,优选前述双歧杆菌属细菌为选自由短双岐杆菌、婴儿双歧杆菌和长双歧杆菌组成的组中的一种或二种以上。
本方式中,优选人乳寡糖相对于乳铁蛋白和/或乳铁蛋白水解物的质量比为1/100000以上且1/10以下。
本方式中的人乳寡糖优选包含选自由2’-岩藻糖基乳糖、3-岩藻糖基乳糖和乳糖二岩藻四糖组成的组中的一种或二种以上。
本方式的经口组合物优选为婴幼儿用营养组合物,特别优选为婴幼儿用配方乳。
发明的效果
根据本发明,提供能促进双歧杆菌增殖的组合物。上述组合物可以以添加物等方式包含于饮食品、药品。另外,通过将本发明的组合物进一步形成含有双歧杆菌的经口组合物的方式,可以得到肠道菌群的改善效果,期待有助于食用者、特别是婴幼儿的健康维持。
附图说明
图1为示出实施例1中的、2’-岩藻糖基乳糖和乳铁蛋白的短双岐杆菌的增殖促进作用的图。
图2为示出实施例2中的、2’-岩藻糖基乳糖和乳铁蛋白的婴儿双歧杆菌的增殖促进作用的图。
图3为示出实施例2中的、2’-岩藻糖基乳糖和乳铁蛋白的婴儿双歧杆菌的增殖促进作用的图。
具体实施方式
接着,对本发明详细地进行说明。但是,本发明不限定于以下的实施方式,在本发明的范围内可以自由变更。
本发明的第一方式的组合物含有乳铁蛋白和/或乳铁蛋白水解物、且含有人乳寡糖。
乳铁蛋白是哺乳动物、例如绵羊、山羊、猪、小鼠、水牛、骆驼(camel)、牦牛、马、驴、美洲驼(lama)、牛或人的乳、眼泪、唾液、血液等中所含的铁结合性的糖蛋白。
本发明中的乳铁蛋白可以源自任意哺乳动物,没有特别限定,从含量、获得容易性的方面出发,例如优选源自牛、人等的乳的乳铁蛋白。作为前述乳,可以为初乳、过渡乳、成熟乳、晚乳中的任一者。
另外,本发明中的乳铁蛋白可以是由作为前述乳的处理物的脱脂乳、乳清等通过常规方法(例如离子色谱法等)分离出的乳铁蛋白、通过基因操作由微生物、动物细胞、转基因动物等产生出的重组乳铁蛋白、合成乳铁蛋白、或它们的混合物。另外,乳铁蛋白可以为经非糖基化或糖基化而成者。作为这种乳铁蛋白,可以使用以工业规模制造的市售的乳铁蛋白(例如森永乳业株式会社制等)。
本发明中的乳铁蛋白中的金属含量没有特别限定,可以使用:选自由利用盐酸、柠檬酸等将乳铁蛋白脱铁而成的脱辅基型乳铁蛋白;使该脱辅基型乳铁蛋白以铁、铜、锌、锰等金属螯合而得到的饱和度100%以上的状态的金属饱和型乳铁蛋白;和金属以低于100%的各种饱和度结合的状态的金属部分饱和型乳铁蛋白组成的组中的任1种或2种以上的混合物。
此处,以下示出本技术中使用的乳铁蛋白的调制(乳铁蛋白自乳等原料的分离、纯化)方法的一例,但不限定于此。
使源自牛的乳原料在阳离子交换柱中通液,将该通过液回收,使该通过液适宜重复地在柱中通液。使去离子水在该柱中通液,使食盐水通液,得到吸附于该阳离子交换柱的碱性蛋白质的洗脱液。在该洗脱液中,以硫酸铵沉淀法,将其回收并适宜清洗。使回收后的沉淀物溶解于去离子水,将该溶解液用超滤膜过滤。进而,进行脱盐处理、冻干,从而得到粉末状的乳铁蛋白。
更详细地,首先,在柱中填充离子交换体,使盐酸通液,进行水洗,使离子交换体平衡化。然后,使冷却至4℃的pH6.9的脱脂乳在柱中通液,将透过液回收,再次同样地在柱中通液。接着,使去离子水在柱中通液,使食盐水通液,得到吸附于离子交换体的碱性蛋白质的洗脱液。在该洗脱液中添加饱和度80%的硫酸铵,使蛋白质沉淀,进行离心分离,将沉淀物回收。将回收后的沉淀物用饱和度80%的硫酸铵溶液清洗,添加去离子水以进行溶解,对于得到的溶液,用超滤膜组件进行脱盐、冻干,得到粉末状牛乳铁蛋白。如此得到了纯度为95质量%以上的牛乳铁蛋白。
乳铁蛋白中,根据种、属、个体等的差异,当然存在有1个或多个位置处的1个或多个碱基的置换、缺失、插入、附加、或逆位等基因变异,具有这种变异的基因编码的蛋白质的氨基酸中,也有时产生变异。本发明中的本技术中能使用的乳铁蛋白中,只要不有损本发明的效果就还含有包含这种变异者。
另外,本发明中的乳铁蛋白中,只要不有损本发明的效果就可以还包含进行了热处理、酸处理、或碱处理的乳铁蛋白处理物。
本发明中的乳铁蛋白水解物是指,对前述乳铁蛋白进行了水解处理而成者。
作为水解处理,例如可以举出日本特开2012-235768号公报中记载的方法。
具体而言,在对乳铁蛋白溶液进行酶反应处理之前,利用盐酸、柠檬酸、乙酸等酸,将pH调整为2~4、优选调整为2.5~3.5、特别优选调整为pH3。
在调整了pH的乳铁蛋白溶液中,以期望的量添加含有凝乳酶的酶组合物后,保持酶反应的温度为35~55℃、优选40~50℃、更优选42~48℃,边搅拌边使乳铁蛋白水解6小时~24小时、优选12~18小时。
接着,例如将反应溶液升温至80℃并维持10分钟,使酶加热失活。进而,优选添加氢氧化钠溶液等碱溶液,调整pH为5~7、例如为6。
需要说明的是,pH调整后的反应溶液(乳铁蛋白水解物)可以直接为溶液,优选进行冻干等而粉末化。另外,乳铁蛋白分解物也可以使用通过色谱法、或超滤等截流而成者。
本方式的组合物中,乳铁蛋白和/或乳铁蛋白水解物相对于组合物整体的含量优选0.001质量%以上且低于100质量%、更优选0.005~75质量%、进一步优选0.01~50质量%。
本发明中的人乳寡糖只要为人的乳中通常包含的寡糖就没有特别限定,优选可以举出2’-岩藻糖基乳糖、3-岩藻糖基乳糖、乳糖二岩藻四糖、2’,3-二岩藻糖基乳糖、乳糖-N-三糖II、乳糖-N-四糖、乳糖-N-新四糖、乳糖-N-岩藻五糖I、乳糖-N-新岩藻五糖、乳糖-N-岩藻五糖II、乳糖-N-岩藻五糖III、乳糖-N-岩藻五糖V、乳糖-N-新岩藻五糖V、乳糖-N-二岩藻六糖I、乳糖-N-二岩藻六糖II、6’-半乳糖基乳糖、3’-半乳糖基乳糖、乳糖-N-六糖和乳糖-N-新六糖等中性人乳寡糖、3’-唾液酸乳糖、6’-唾液酸乳糖、3-岩藻糖基-3’-唾液酸乳糖、二唾液酸-乳糖-N-四糖等酸性人乳寡糖。它们之中,从双歧杆菌属细菌的增殖促进效果的观点出发,优选2’-岩藻糖基乳糖、3-岩藻糖基乳糖、和乳糖二岩藻四糖,特别优选2’-岩藻糖基乳糖。
需要说明的是,本发明中的人乳寡糖只要不有损本发明的效果即可,可以为经纯化者,也可以为混合物。另外,可以包含前述人乳寡糖的一种或二种以上。
本方式的组合物中,人乳寡糖相对于组合物整体的含量优选0.001质量%以上且低于100质量%、更优选0.005~75质量%、进一步优选0.01~50质量%。
另外,本方式的组合物中,人乳寡糖相对于乳铁蛋白和/或乳铁蛋白水解物的质量比优选1/100000以上且1/10以下,更优选1/10000以上且1/10以下,进一步优选1/1000以上且1/10以下。
本发明的组合物可以促进双歧杆菌(Bifidobacterium)属细菌的增殖。
此处“促进增殖”是指,在体内或体外应用本发明的组合物时的双歧杆菌属细菌的菌数与不应用的情况相比增加。上述菌数的增加的程度没有特别限定,是指相对于不应用本发明的组合物时的双歧杆菌属细菌的菌数,优选大0.1%以上、更优选大1%以上、进一步优选大5%以上。
上述菌数的增加可以如下确认:除直接测量菌数之外,例如根据测定培养细菌的培养基的浊度(吸光度)或乙酸等短链脂肪酸量其值上升、测定前述培养基中的pH其值降低等,从而可以确认。
已知乳铁蛋白和/或乳铁蛋白水解物和人乳寡糖分别对双歧杆菌属细菌示出增殖促进作用,但如后述的实施例所示,通过组合两者,从而协同地发挥该作用,发挥优异的双歧杆菌属细菌的增殖促进效果。
作为由本发明的组合物促进增殖的双歧杆菌属细菌,没有特别限制,可以举出:短双歧杆菌(Bifidobacterium breve)、婴儿双歧杆菌(Bifidobacterium infantis、被再分类为长双歧杆菌婴儿亚种)、长双歧杆菌(Bifidobacterium longum)、两岐双岐杆菌(Bifidobacterium bifidum)、青春双岐杆菌(Bifidobacterium adolescentis)、链状双歧杆菌(Bifidobacterium catenulatum)、假小链双歧杆菌(Bifidobacteriumpseudocatenulatum)、动物双歧杆菌(Bifidobacterium animalis)、乳酸双歧杆菌(Bifidobacterium lactis)和假长双岐杆菌(Bifidobacterium pseudolongum)。它们之中,优选选自由短双歧杆菌、婴儿双歧杆菌和长双歧杆菌组成的组中的一种或二种以上。
本发明的第一方式的组合物可以将其本身制成饮食品、药品等形态,也可以制成作为添加物包含于饮食品、药品等的形态。
本方式的组合物的摄取(给予)路径可以为经口或非经口,均可,通常为经口。另外,作为非经口摄取(给予),可以举出直肠给予等。
本发明的第一方式的组合物经经口摄取(给予)的情况下,优选还含有双歧杆菌属细菌。
即,本发明的第二方式为一种经口组合物,其含有:乳铁蛋白和/或乳铁蛋白水解物、人乳寡糖和双歧杆菌属细菌。
本方式中的涉及乳铁蛋白和/或乳铁蛋白水解物、以及人乳寡糖的说明依据前述第一方式中的说明。
第二方式的组合物中,乳铁蛋白和/或乳铁蛋白水解物相对于组合物整体的含量优选0.001~5.0质量%、更优选0.005~1.0质量%、进一步优选0.01~0.5质量%。
另外,第二方式的组合物中,人乳寡糖相对于组合物整体的含量优选0.001~5.0质量%、更优选0.005~1.0质量%、进一步优选0.01~0.5质量%。
这些可以为通常作为经口组合物流通时的含量的范围。
作为经口摄取(给予)第二方式的组合物时的含量,可以设为上述范围,也可以适宜稀释等。例如,经口摄取(给予)时的乳铁蛋白和/或乳铁蛋白水解物相对于组合物整体的含量可以设为0.0001~5.0质量%。另外,经口摄取(给予)时的人乳寡糖相对于组合物整体的含量可以设为0.0001~5.0质量%。
本方式中,至少包含活菌作为双歧杆菌属细菌,每1mL经口组合物,含有优选1.0×106cfu以上、更优选1.0×107cfu以上、进一步优选2.0×107cfu以上的双歧杆菌属细菌的活菌。需要说明的是,只要含有活菌即可,也可以包含死菌。
需要说明的是,cfu是指菌落形成单元(Colony forming unit)。本说明书中,例如,可以设为包含还原脱脂乳粉10质量%的固体培养基中以38℃进行培养时的值。
这些可以为通常作为经口组合物流通时的含量的范围。
作为本方式中经口组合物中含有的双歧杆菌属细菌,可以举出前述第一方式中说明的种类者,优选为选自由短双岐杆菌、婴儿双歧杆菌和长双歧杆菌组成的组中的一种或二种以上。
作为短双歧杆菌,更具体而言,可以举出短双歧杆菌M-16V。短双歧杆菌M-16V于2018年1月26日、在国家技术评估学会,专利微生物保藏中心(NPMD)(〒292-0818日本国千叶县木更津市上总镰足2-5-8 122号室)以保藏编号NITE BP-02622进行了基于布达佩斯条约的国际保藏。可以使用作为市售品能获得的、例如森永乳业株式会社制的“短双歧杆菌M-16V”。
另外,作为短双歧杆菌,更具体而言,还可以举出短双歧杆菌MCC1274。短双歧杆菌MCC1274于2009年8月25日、在独立行政法人产业技术综合研究所专利生物保藏中心(现·国家高级工业科学技术学院,国际专利生物保藏中心(IPOD,NITE)(〒292-0818日本国千叶县木更津市上总镰足2-5-8 120号室)以保藏编号FERM BP-11175进行了基于布达佩斯条约的国际保藏。
作为婴儿双歧杆菌,更具体而言,还可以举出婴儿双歧杆菌M-63。婴儿双歧杆菌M-63于2018年1月26日、在NPMD以保藏编号NITE BP-02623进行了基于布达佩斯条约的国际保藏。
作为长双歧杆菌,更具体而言,可以使用长双歧杆菌NITE BP-02621、(别名:BB536或长双歧杆菌长亚种(Bifidobacterium longum subsp.longum)ATCC BAA-999)。长双歧杆菌BB536在NPMD、于2018年1月26日以NITE BP-02621的保藏编号进行了基于布达佩斯条约的国际保藏。作为与其相同的细菌的长双歧杆菌长亚种(Bifidobacterium longumsubsp.longum)ATCC BAA-999(编号:ATCC BAA-999)可以自American Type CultureCollection(ATCC:美国、20110、弗吉尼亚州、马纳萨斯大学大道10801(10801 UniverSityBoulevard,Manassas,VA 20110,United States of America))作为ATCC BAA-999获得(例如参照日本特开2012-223134号公报等)。
需要说明的是,以上述示例的菌株名所特定的菌株中不限定于以该菌株名保藏、注册在规定的机关的菌株其本身(以下为了方便说明,也称为“保藏菌株”),还包括与其实质上同等的菌株(也称为“派生菌株”或“衍生株”)。即,不限定于以上述保藏编号保藏于上述保藏机构的菌株本身,也包含与其实质上等同的菌株。对于各菌株,“与上述保藏菌株实质上同等的菌株”是指如下菌株:属于与上述保藏菌株相同的种,可以得到肠道菌群改善效果,进一步其16SrRNA基因的碱基序列相对于上述保藏菌株的16SrRNA基因的碱基序列、具有优选99.86%以上、更优选99.93%以上、进一步优选100%的同一性,且优选具有与上述保藏菌株相同的菌学性质。对于各菌株,与上述保藏菌株实质上同等的菌株例如也可以是将该保藏菌株作为亲本菌株的派生菌株。作为派生菌株,可以举出:由保藏菌株育种的菌株、自保藏菌株自然生长的菌株。作为育种方法,可以举出基于基因工程学方法的修饰、基于突变处理的修饰。作为突变处理,可以举出X射线的照射、紫外线的照射、以及利用N-甲基-N’-硝基-N-亚硝基胍、甲磺酸乙酯和甲磺酸甲酯等诱变剂的处理。作为自保藏菌株自然生长的菌株,可以举出使用保藏菌株时自然生长的菌株。作为这种菌株,可以举出通过保藏菌株的培养(例如传代培养)而自然生长的突变株。派生菌株可以通过一种修饰来构建,也可以通过二种或其以上的修饰来构建。
作为本方式的经口组合物中能含有的双歧杆菌属细菌的菌体,可以使用市售品,也可以使用适宜制造而取得者。
另外,本方式的经口组合物中能含有的双歧杆菌属细菌的菌体可以通过对前述双歧杆菌属细菌进行培养而容易取得。培养方法只要双歧杆菌属细菌可以增殖就没有特别限制。作为培养方法,例如可以直接使用双歧杆菌属细菌的培养中通常使用的方法,或适宜修正而使用。培养温度例如可以为25~50℃,优选35~42℃。培养优选可以在厌氧条件下实施,例如可以边使二氧化碳气体等厌氧气体通气边实施。另外,培养也可以在液体静置培养等微好氧条件下实施。例如可以实施培养,直至双歧杆菌属细菌增殖至期望程度。
培养中使用的培养基只要双歧杆菌属细菌可以增殖就没有特别限制。作为培养基,例如可以直接使用双歧杆菌属细菌的培养中通常使用的培养基、或适宜修正而使用。即,作为碳源,可以根据同化性而使用例如半乳糖、葡萄糖、果糖、甘露糖、纤维二糖、麦芽糖、乳糖、蔗糖、海藻糖、淀粉、淀粉水解物、赤糖糊等糖类。作为氮源,例如可以使用氨、硫酸铵、氯化铵、硝酸铵等铵盐类、硝酸盐类。另外,作为无机盐类,例如可以使用氯化钠、氯化钾、磷酸钾、硫酸镁、氯化钙、硝酸钙、氯化锰、硫酸亚铁等。另外,也可以使用蛋白胨、大豆粉、脱脂大豆粕、肉提取物、酵母提取物等有机成分。另外,作为双歧杆菌属细菌的培养中通常使用的培养基,具体而言,可以举出:梭菌强化培养基(Reinforced Clostridialmedium)、MRS培养基(de Man,Rogosa,and Sharpe medium)、mMRS培养基(modified MRSmedium)、TOSP培养基(TOS propionate medium)、TOSP Mup培养基(TOS propionatemupirocin medium)。
作为本方式的经口组合物中能含有的双歧杆菌属细菌,可以没有特别限制地使用其菌体或含有其的级分。即,作为双歧杆菌属细菌,例如可以直接使用通过培养而得到的培养物,也可以将培养物稀释或浓缩而使用,还可以使用从培养物回收的菌体。另外,只要不有损肠道菌群改善效果即可,就可以在培养后进行加热、冻干等各种追加操作。追加操作优选菌体的生存性高者。即,作为本方式的经口中能含有的双歧杆菌属细菌,具体而言,可以举出双歧杆菌属细菌的培养物、或从前述培养物回收的菌体、它们的处理物,作为前述处理物,可以举出稀释物、浓缩物、或干燥物等。需要说明的是,菌体通常优选以含有活菌体的形态使用。菌体例如可以由活菌体构成,也可以为活菌体与死菌体的混合物。
本方式的经口组合物如第一方式中说明可以促进双歧杆菌属细菌的增殖。此处,增殖对象的双歧杆菌属细菌不限定于经口组合物中含有的双歧杆菌属细菌,也包含存在于摄取的人等动物的消化道内的双歧杆菌属细菌。
因此,本方式的经口组合物可以用于改善肠道菌群。此处,“菌群改善”是指,包括增加双歧杆菌属细菌的存在于肠道菌群的细菌数、其存在比例。另外,“菌群改善”不限定于双歧杆菌属细菌的肠道菌群中的存在比例增加,也可以包括增加其他好细菌在肠道菌群中的存在比例、减少坏细菌在肠道菌群中的存在比例。作为其他好细菌,可以举出乳酸菌等。作为坏细菌,可以举出产气荚膜梭菌(Clostridium perfringens)、沙门氏属细菌、金黄色葡萄球菌、致病性大肠杆菌等。
前述“存在比例”也可以换言之相对于在肠道菌群中检测到的菌群整体的“占有率”。
作为本发明的组合物通过改善肠内菌群来能预防或改善的疾病、病态的对象者、或源自肠内菌群恶化的疾病、病态的对象者,可以变得有用。例如可以作为肠道调节用、免疫调节用、抗变态反应用、细菌/病毒感染防御用、氧化应激降低用、腹泻的预防/改善用、便秘的预防/改善用、炎症性肠疾病用、大肠癌的预防用等。
本发明的另一方式为一种乳铁蛋白和/或乳铁蛋白水解物、以及人乳寡糖在制造双歧杆菌属细菌的增殖促进用组合物中的应用。
本发明的另一方式为一种乳铁蛋白和/或乳铁蛋白水解物、以及人乳寡糖在双歧杆菌属细菌的增殖促进中的应用。
本发明的另一方式为一种用于促进双歧杆菌属细菌的增殖的、乳铁蛋白和/或乳铁蛋白水解物、以及人乳寡糖。
本发明的另一方式一种促进双歧杆菌属细菌的增殖的方法,其包括将乳铁蛋白和/或乳铁蛋白水解物、以及人乳寡糖给予至动物。此处,动物没有特别限定,通常为人。
本发明的组合物的摄取(给予)时期没有特别限定,可以根据给予对象的状态而适宜选择。
本发明的组合物的摄取(给予)量可以根据摄取(给予)对象的年龄、性别、状态、其他条件等而适宜选择。
需要说明的是,无论摄取(给予)的量、时间,药剂均1日1次给予或分多次给予。
本发明的组合物优选形成饮食品的方式。
作为饮食品,只要不有损本发明的效果就对形态、性状没有特别限制,可以利用通常饮食品中使用的原料根据通常的方法而制造。
需要说明的是,饮食品中添加的添加物的方式也包含于本发明的组合物中。作为上述方式,例如可以举出挤奶后的人乳、配方乳中添加的添加物,设想添加后的乳被新生儿、婴儿摄取。
饮食品通常经经口摄取,但不限定于此,例如可以经鼻摄取、可以由胃瘘、肠瘘摄取。例如,对于新生儿、婴儿,设想由经鼻胃营养管等摄取作为后述的本发明的组合物的婴幼儿用配方乳、添加了本发明的组合物的人乳。
作为饮食品,液态、糊剂状、凝胶状固体、粉末等的形态均可,例如可以举出片剂糖果;流食(经管摄取用营养食品);面包、通心粉、意大利面、面类、蛋糕混合料、油炸粉、面包粉等小麦粉制品;方便面、杯面、蒸煮/熟食食品、熟食罐头、微波炉用食品、速溶汤/日式炖菜、速溶大酱汤/汤物、罐装浓汤、冻干食物、其它速食品等速食品类;农作物罐头、水果罐头、果酱(jam)/橘子酱(marmalade)类、酱菜、煮豆类、农产品干物类、谷物(谷物加工品)等农产加工品;水产罐头、鱼肉火腿/香肠、水产糜类制品、水产鱼贝类、佃煮类等水产加工品;畜产罐头/糊类、畜肉火腿/香肠等畜产加工品;加工乳、乳饮料、酸奶类、乳酸菌饮料类、乳酪、奶油冰淇淋类、奶油、其它乳制品等乳/乳制品;黄油、人造奶油类、植物油等油脂类;酱油、大酱、沙司(sauce)类、西红柿加工调味剂、味醂类、食醋类等基础调味剂;烹调混合料、咖喱的原料类、酱汁类、沙拉(dressing)类、面汤类、香辛料类、其它复合调味剂等复合调味剂/食品类;原材料冷冻食品、半熟食冷冻食品、经烹调的冷冻食品等冷冻食品;焦糖、糖果、口香糖、巧克力、曲奇(cookie)、饼干(biscuits)、蛋糕(cake)、馅饼(pie)、小吃(snack)、薄脆饼(cracker)、日本点心、米点心、豆点心、西式甜点、果冻、其他点心等点心类;碳酸饮料、天然果汁、果汁饮料、带有果汁的清凉饮料、果肉饮料、带有果粒的果实饮料、蔬菜系饮料、豆奶、豆奶饮料、咖啡饮料、茶饮料、粉末饮料、浓缩饮料、运动饮料、营养饮料、酒精饮料、其他嗜好饮料等嗜好饮料类、婴儿食品、米饭伴侣、茶泡饭海带等其他市售食品等;补充剂、配方乳(包含乳粉、液态乳等)等营养组合物;育儿用补充剂、育儿用配方乳(包含乳粉、液态乳等)等育儿用(婴幼儿用)营养组合物;经肠营养食品;功能性食品(特定保健用食品、营养功能食品)等。
另外,作为饮食品,可以举出面向妊娠期/哺乳期的母亲的妈妈用牛奶(均衡性良好地配合有妊娠/哺乳期所需的营养的配方乳)、学童期及以后的配方乳、面向成人的配方乳、面向老年人的流食等营养调整食品、营养辅助食品、流食等营养功能食品、患者用食品(特殊用途食品)等
其中,更优选婴幼儿用营养组合物、特别优选婴幼儿用配方乳。
另外,作为饮食品的一方式,也可以制成饲料。作为饲料,可以举出宠物食品、家畜饲料、养鱼饲料等。
作为饲料的形态,没有特别限制,例如可以含有玉米、小麦、大麦、黑麦、芦粟(milo)等谷类;大豆油渣、菜籽油渣、棕榈油渣、亚麻籽油渣等植物性油渣类;麦麸、麦糠、米糠、脱脂米糠等糠类;玉米蛋白粗粉、玉米浆粗粉等制造糟粕类;鱼粉、脱脂奶粉、乳清、黄色油脂、牛脂等动物性饲料类;圆酵母、啤酒酵母等酵母类;磷酸三钙、碳酸钙等矿物质饲料;油脂类;单体氨基酸;糖类等。
本发明的组合物为饮食品(包含饲料)的方式的情况下,可以作为标示了使肠内的双歧杆菌增殖的用途的饮食品提供/销售。
上述“标示”行为中包括用于向需求者通知上述用途的全部行为,只要是能唤起/类推前述用途的表现,无论标示的目的、标示的内容、标示的对象物/介质等如何都属于本发明的“标示”行为。
另外,“标示”优选的是通过需求者能直接识别上述用途的表现来进行。具体而言,可列举出:为了转让、交付、用于转让或交付而展示在饮食品的商品或商品的包装上记载了前述用途的物品,在进口行为、涉及商品的广告、价格表或说明书类上将上述用途记载并展示、或进行发布,或在将这些作为内容的信息中记载上述用途并利用电磁(网络等)方法来提供的行为等。
另一方面,作为标示内容,优选为根据行政等许可的标示(例如基于行政规定的各种制度而获得许可,在基于这样的许可的状态下进行的标示等)。另外,优选将这样的标示内容附于包装、容器、目录、小册子、POP(购买点广告,Point of purchase advertising)等销售现场的宣传材料、其它资料等上。
另外,“标示”还可列举出作为健康食品、功能性食品、经肠营养食品、特殊用途食品、保健功能食品、特定保健用食品、营养功能食品、功能性标示食品、药物用准药物等的标示。其中特别是被消费者厅许可的标示、例如可列举出涉及特定保健用食品、营养功能食品、或功能性标示食品的制度、或与这些类似的制度认可的标示等。具体而言,可以列举出作为特定保健用食品的标示、作为带有条件的特定保健用食品的标示、对身体的构造、功能产生影响内容的标示、疾病风险降低标示、基于科学根据的功能性的标示等等,更具体而言,涉及健康增进法中规定的特殊用途标示的许可等的日本内阁府令(日本平成二十一年八月三十一年内阁府令第五十七号)中规定的作为特定保健用食品的标示(特别是保健的用途的标示)和与其类似的标示是典型例。
作为上述标示,例如可以举出标示为“想要增加双歧杆菌”、“用于改善肠道菌群”、“用于婴幼儿的胃健康”等。
本发明的组合物为药品的形态的情况下,其给予路径可以为经口或非经口,均可,优选经口。另外,作为非经口摄取(给予),可以举出直肠给予等。
作为药品的形态,可以根据给予方法而适宜制剂化为期望的剂型。例如,经口给予的情况下,可以制剂化为散剂、颗粒剂、片剂、胶囊剂等固体制剂;溶液剂、糖浆剂、悬浮剂、乳剂等液体制剂等。另外,非经口给予的情况下,可以制剂化为栓剂、软膏剂、注射剂等。
制剂化时,可以使用通常制剂化中使用的赋形剂、pH调节剂、着色剂、矫味剂等成分。另外,也可以组合使用其他药效成分、公知的或对将来会发现的双歧杆菌属细菌的益生元等。
此外,制剂化可以根据剂型通过公知的方法适宜实施。制剂化时,可以适宜配合制剂载体而制剂化。
作为赋形剂,例如可以举出乳糖、白糖、葡萄糖、甘露糖醇、山梨糖醇等糖衍生物;玉米淀粉、马铃薯淀粉、α-淀粉、糊精、羧甲基淀粉等淀粉衍生物;结晶纤维素、羟丙基纤维素、羟丙基甲基纤维素、羧甲基纤维素、羧甲基纤维素钙等纤维素衍生物;阿拉伯胶;葡聚糖;普鲁兰多糖;轻质无水硅酸、合成硅酸铝、偏硅酸铝酸镁等硅酸盐衍生物;磷酸钙等磷酸盐衍生物;碳酸钙等碳酸盐衍生物;硫酸钙等硫酸盐衍生物等。
作为结合剂,除了上述赋形剂之外可列举出例如:明胶;聚乙烯基吡咯烷酮;聚乙二醇等。
作为崩解剂,除了上述赋形剂之外可列举出例如:交联羧甲基纤维素钠、羧甲基淀粉钠、交联聚乙烯基吡咯烷酮等经化学修饰的淀粉或纤维素衍生物等。
作为润滑剂,可列举出例如:滑石;硬脂酸;硬脂酸钙、硬脂酸镁等硬脂酸金属盐;胶体二氧化硅;硅酸铝镁盐(Veegum)、鲸蜡等蜡类;硼酸;二元醇;富马酸、己二酸等羧酸类;苯甲酸钠等羧酸钠盐;硫酸钠等硫酸盐类;亮氨酸;月桂基硫酸钠、月桂基硫酸镁等月桂基硫酸盐;硅酸酐、硅酸水合物等硅酸类;淀粉衍生物等。
作为稳定剂,可列举出例如:对羟基苯甲酸甲酯、对羟基苯甲酸丙酯等对羟基苯甲酸酯类;氯丁醇、苄醇、苯基乙醇等醇类;苯扎氯铵;乙酸酐;山梨酸等。
作为矫味矫臭剂,例如可列举出:甜味剂、酸味剂、香料等。
需要说明的是,作为经口给予用的液体制剂的情况下使用的载体,可以举出水等溶剂等。
摄取本发明的药品的时机例如为饭前、饭后、饭间、睡觉前等,没有特别限定。
实施例
以下,利用实施例对本发明进一步具体地进行说明,但本发明不限定于这些实施例。
<实施例1>人乳寡糖和乳铁蛋白对短双岐杆菌的增殖活性的影响的研究
(1)培养基和试剂的制备
(i)MRS培养基的制备
使DifcoMRS(BD)如所附说明书记载溶解于纯化水,在115℃下进行15分钟灭菌。使L-半胱氨酸盐酸盐(L-Cysteine hydrochloride)(Wako)溶解于纯化水,进行过滤灭菌,在灭菌后冷却至常温的MRS培养基中以最终浓度成为0.05质量%的方式以无菌的方式添加。
(ii)不含有葡萄糖的MRS培养基的制备
以表1中记载的组成使试剂溶解于纯化水,在115℃下,进行15分钟灭菌。使L-半胱氨酸盐酸盐(L-Cysteine hydrochloride)(Wako)溶解于纯化水,进行过滤灭菌,在灭菌后冷却至常温的MRS培养基中以最终浓度成为0.05质量%的方式以无菌的方式添加。
[表1]
组成 | 量 |
聚氧乙烯山梨糖醇酐单油酸酯(Nacalai Tesque) | 1.25g |
Bacto Beef Extract(BD) | 12.5g |
BBL Polypeptone(BD) | 12.5g |
Bacto Yeast Extract(BD) | 6.25g |
磷酸二氢钾(国产化学) | 2.5g |
乙酸钠(Wako) | 6.25g |
柠檬酸氢二铵(Wako) | 2.5g |
硫酸镁·七水合物(国产化学) | 0.5g |
硫酸锰(II)五水合物(Wako) | 0.0625g |
纯化水 | 900mL |
(iii)人乳寡糖溶液和乳铁蛋白溶液的制备
使人乳寡糖(2’-岩藻糖基乳糖(2’-FL)(Carbosynth))、和乳铁蛋白(森永乳业株式会社制、铁含量15.8mg/100g)分别溶解于纯化水。溶解后进行过滤灭菌。
(2)前培养液的制备
(1)在(i)中制备的MRS培养基3mL中,添加短双岐杆菌M-16V的前培养液150μL,以厌氧的方式以37℃培养16小时。培养后,以1500×g进行5分钟离心分离,将菌体和培养液分离。去除培养液,再次悬浮于(1)(ii)中制备的不含有葡萄糖的MRS培养基3mL。
(3)增殖促进效果的评价
(1)在(ii)中制备的不含葡萄糖的MRS培养基中,添加(2)中制备的前培养液使得终浓度成为1v/v%。进一步,添加(1)(iii)中制备的2’-岩藻糖基乳糖(2’-FL)和乳铁蛋白溶液使其分别成为表2所示的终浓度,在96孔板中每200μL/孔进行分注。
需要说明的是,1.0mg/mL换算为质量百分率时为0.1质量%。
之后,以厌氧的方式以37℃培养24小时。培养后,用酶标仪(电晕电气),在600nm的波长下测定浊度,评价双歧杆菌的增殖。
将结果示于表2和图1。在2’-岩藻糖基乳糖和乳铁蛋白共存下培养的情况下,相对于对照(未添加2’-岩藻糖基乳糖和乳铁蛋白),确认到短双岐杆菌M-16V的显著的增殖。
[表2]
<实施例2>人乳寡糖和乳铁蛋白对婴儿双歧杆菌的增殖活性的影响的研究
将双歧杆菌替换为婴儿双歧杆菌M-63,除此之外,进行与实施例1同样的操作,进行增殖的评价。
将结果示于表3以及图2和图3。在2’-岩藻糖基乳糖和乳铁蛋白共存下培养的情况下,相对于对照(未添加2’-岩藻糖基乳糖和乳铁蛋白),确认到婴儿双歧杆菌M-63的增殖倾向。另外,在2’-岩藻糖基乳糖和乳铁蛋白共存下培养的情况下,相对于添加2’-岩藻糖基乳糖且未添加乳铁蛋白组,确认到婴儿双歧杆菌M-63的显著的增殖。
[表3]
<实施例3>人乳寡糖和乳铁蛋白对长双歧杆菌的增殖活性的影响的研究
将双歧杆菌替换为长双歧杆菌BB536,除此之外,进行与实施例1同样的操作,进行增殖的评价。
将结果示于表4。在2’-岩藻糖基乳糖和乳铁蛋白共存下培养的情况下,与对照(未添加2’-岩藻糖基乳糖和乳铁蛋白)相比,确认到长双歧杆菌BB536的增殖倾向。
[表4]
<实施例4>人乳寡糖和乳铁蛋白对短双岐杆菌的增殖活性的影响的研究
将双歧杆菌替换为短双岐杆菌MCC1274,除此之外,进行与实施例1同样的操作,进行增殖的评价。
将结果示于表5。在2’-岩藻糖基乳糖和乳铁蛋白共存下培养的情况下,与对照(未添加2’-岩藻糖基乳糖和乳铁蛋白)相比,确认到短双岐杆菌MCC1274的增殖倾向。
[表5]
PCT/RO/134表
Claims (12)
1.一种组合物,其含有乳铁蛋白和/或乳铁蛋白水解物、且含有人乳寡糖。
2.根据权利要求1所述的组合物,其中,人乳寡糖相对于乳铁蛋白和/或乳铁蛋白水解物的质量比为1/100000以上且1/10以下。
3.根据权利要求1或2所述的组合物,其中,乳铁蛋白和/或乳铁蛋白水解物相对于组合物整体的含量为0.001质量%以上且低于100质量%。
4.根据权利要求1~3中任一项所述的组合物,其中,人乳寡糖相对于组合物整体的含量为0.001质量%以上且低于100质量%。
5.根据权利要求1~4中任一项所述的组合物,其中,人乳寡糖包含选自由2’-岩藻糖基乳糖、3-岩藻糖基乳糖和乳糖二岩藻四糖组成的组中的一种或二种以上。
6.根据权利要求1~5中任一项所述的组合物,其用于双歧杆菌属细菌增殖促进。
7.根据权利要求6所述的组合物,其中,所述双歧杆菌属细菌为选自由短双岐杆菌(Bifidobacterium breve)、婴儿双歧杆菌(Bifidobacteriuminfantis)和长双歧杆菌(Bifidobacterium longum)组成的组中的一种或二种以上。
8.一种经口组合物,其含有:乳铁蛋白和/或乳铁蛋白水解物、人乳寡糖、和双歧杆菌属细菌。
9.根据权利要求8所述的经口组合物,其中,所述双歧杆菌属细菌为选自由短双岐杆菌(Bifidobacterium breve)、婴儿双歧杆菌(Bifidobacterium infantis)和长双歧杆菌(Bifidobacterium longum)组成的组中的一种或二种以上。
10.根据权利要求8或9所述的经口组合物,其中,人乳寡糖相对于乳铁蛋白和/或乳铁蛋白水解物的质量比为1/100000以上且1/10以下。
11.根据权利要求8~10中任一项所述的经口组合物,其中,人乳寡糖包含选自由2’-岩藻糖基乳糖、3-岩藻糖基乳糖和乳糖二岩藻四糖组成的组中的一种或二种以上。
12.根据权利要求8~11中任一项所述的经口组合物,其为婴幼儿用配方乳。
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