JP7445910B2 - Oral compositions, oral care products, and foods containing oral resident bacteria regulators - Google Patents
Oral compositions, oral care products, and foods containing oral resident bacteria regulators Download PDFInfo
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- JP7445910B2 JP7445910B2 JP2017128123A JP2017128123A JP7445910B2 JP 7445910 B2 JP7445910 B2 JP 7445910B2 JP 2017128123 A JP2017128123 A JP 2017128123A JP 2017128123 A JP2017128123 A JP 2017128123A JP 7445910 B2 JP7445910 B2 JP 7445910B2
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- bacteria
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- Coloring Foods And Improving Nutritive Qualities (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Medicinal Preparation (AREA)
- Cosmetics (AREA)
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Description
本発明は、口腔常在菌調整剤を含む口腔用組成物に関し、詳しくは、口腔内における有用常在菌を残存させるとともに、選択的に口腔内の病原性細菌に対して抗菌効果を奏するようにして口腔内の常在菌のバランスを良好に保つように調整可能な、優れた選択的抗菌効果を奏する、口腔常在菌調整剤を含む口腔用組成物、口腔ケア製品及び食品に関する。 The present invention relates to an oral cavity composition containing an oral resident bacteria regulating agent, and more specifically, to a composition that maintains useful resident bacteria in the oral cavity and selectively exerts an antibacterial effect against pathogenic bacteria in the oral cavity. The present invention relates to an oral composition, an oral care product, and a food product containing an oral resident bacteria regulating agent that exhibits an excellent selective antibacterial effect and can be adjusted to maintain a good balance of resident bacteria in the oral cavity.
ヒトの口腔内には様々な細菌が共生しており、常在細菌叢と呼ばれる細菌集団を形成している。常在細菌叢には病原菌や日和見感染菌が存在しているが、細菌叢のバランスが保たれている状態では、これらの細菌を原因とする疾患は抑制されており、いわゆる健康な状態にある。しかし、食生活の変化や抗生剤の摂取、ストレス、加齢などを原因として細菌叢のバランスが崩れることにより、病原性細菌や日和見感染菌が増加し疾患を発症する。そのため、疾患の抑制には病原性細菌を選択的に抑制し、非病原性の常在菌(特に有用常在菌)を優勢にすることで常在細菌叢のバランスを保つことが重要である。 Various bacteria coexist in the human oral cavity, forming a bacterial population called the resident bacterial flora. Pathogenic bacteria and opportunistic bacteria exist in the resident flora, but when the flora is in balance, diseases caused by these bacteria are suppressed, and the body is in a so-called healthy state. . However, when the balance of the bacterial flora is disrupted due to changes in diet, intake of antibiotics, stress, aging, etc., pathogenic bacteria and opportunistic bacteria increase, leading to the development of diseases. Therefore, in order to control the disease, it is important to maintain the balance of the resident flora by selectively suppressing pathogenic bacteria and making non-pathogenic resident bacteria (especially beneficial resident bacteria) predominant. .
ヒトの口腔内では700種を超える極めて多種類の口腔内細菌が見出されている。ヒトの口腔内に存在する細菌の中で、病原性を示す細菌は、歯周病原性細菌であるポルフィロモナス ジンジバリス(Porphyromonas gingivalis)、う蝕の原因菌であるストレプトコッカス ミュータンス(Streptococcus mutans)を代表とする数種類の細菌に過ぎない。口腔内細菌の残りのほとんどは、通常、ヒトへの病原性を示さないストレプトコッカス ミティス(Streptococcus mitis、以下、S.mitisとも呼ぶ。)、ストレプトコッカス オラリス(Streptococcus oralis、以下、S.oralisとも呼ぶ。)を代表とするミティス(Mitis)グループなどのストレプトコッカス(連鎖球菌)属細菌などの通常、口腔内においては非病原性の常在菌である。従って、口腔疾患の予防には、口腔内において、非病原性の口腔内常在菌の生育に影響を与えることなく選択的に口腔内病原性細菌の生育を抑制することが重要となる。 More than 700 types of oral bacteria have been found in the human oral cavity. Among the bacteria that exist in the human oral cavity, pathogenic bacteria include Porphyromonas gingivalis, a periodontal pathogen, and Streptococcus mutans, a caries-causing bacterium. These are just a few representative types of bacteria. Most of the remaining oral bacteria are Streptococcus mitis (hereinafter also referred to as S. mitis) and Streptococcus oralis (hereinafter also referred to as S. oralis), which are not normally pathogenic to humans. Bacteria belonging to the genus Streptococcus, such as the Mitis group typified by Mitis, are normally non-pathogenic and resident bacteria in the oral cavity. Therefore, in order to prevent oral diseases, it is important to selectively suppress the growth of oral pathogenic bacteria in the oral cavity without affecting the growth of non-pathogenic oral bacteria.
特に、上述したストレプトコッカス ミティスやストレプトコッカス オラリスなどの非病原性常在菌のミティスグループの連鎖球菌は、上記したう蝕や歯周病の原因菌の歯面付着に関与したり、抗菌作用を有することが知られており(非特許文献1参照)、口腔疾患抑制に作用する有用な常在菌である。 In particular, streptococci belonging to the non-pathogenic resident bacteria group Streptococcus mitis, such as Streptococcus mitis and Streptococcus oralis mentioned above, are involved in the attachment of the above-mentioned caries and periodontal disease-causing bacteria to tooth surfaces, and have antibacterial effects. (Refer to Non-Patent Document 1), and is a useful resident bacteria that acts to suppress oral diseases.
続いて、上述したストレプトコッカス ミティス及びストレプトコッカス オラリスについて具体的に説明する。非特許文献1に記載されているように、バイオフィルムは以下のように形成される。歯の表面を構成するエナメル質の表面には、まずペリクル(歯の表面上の唾液タンパク質の層)が形成される。ペリクルのもとになるのは唾液中のカルシウム結合タンパク質(calcium-binding proteins;CBP)であり、この物質はエナメル質のカルシウムに親和性をもっている。これが歯のエナメル質上に並ぶと、それに親和性をもつ細菌、すなわちストレプトコッカス ミティス(Streptococcus mitis)、ストレプトコッカス オラリス(Streptococcus oralis)などの菌が選択的に付着してくる。これらの菌は、early colonizer(早期定着菌群)と呼ばれる。これが常在菌、すなわちわれわれが守り育てなければならない細菌群で、例えば大腸におけるビフィズス菌に相当する。とくにS.mitisは、口腔内細菌を培養した場合に、口腔内細菌全量の9割を占める。 Next, the above-mentioned Streptococcus mitis and Streptococcus oralis will be specifically explained. As described in Non-Patent Document 1, biofilms are formed as follows. A pellicle (a layer of salivary protein on the tooth surface) is first formed on the surface of the enamel that makes up the tooth surface. The source of the pellicle is calcium-binding proteins (CBP) in saliva, and this substance has an affinity for calcium in enamel. When these are lined up on tooth enamel, bacteria that have an affinity for them, such as Streptococcus mitis and Streptococcus oralis, selectively adhere to them. These bacteria are called early colonizers. These are the resident bacteria, a group of bacteria that we must protect and nurture, and correspond to, for example, the bifidobacteria in the large intestine. Especially S. When oral bacteria are cultured, S. mitis accounts for 90% of the total amount of oral bacteria.
また、非特許文献1では、無菌状態は実は非常に危険な状態であるとの理解が必要であり、あくまで常在菌が口腔内に存在することが大切であり、常在菌叢の維持という観点からは、強力な抗菌剤を口腔内全体に使うのはよくないことであると述べられている。 In addition, Non-Patent Document 1 states that it is necessary to understand that a sterile state is actually a very dangerous state, and that it is important that resident bacteria exist in the oral cavity, and that it is important to maintain the resident bacterial flora. From this point of view, it is stated that it is not a good idea to use strong antibacterial agents throughout the oral cavity.
歯周病等の予防用途として例えば、一般的な呼称としてマウスウォッシュと呼ばれる抗菌作用を有する洗口液が市販されているが、このような洗口液のなかには、種々の抗菌剤が含まれている。洗口液に含まれる幾つかの抗菌剤においては口腔内の細菌全般に対して広く作用し、有用常在菌をも殺菌してしまい、口腔内における常在菌のバランスが崩れてしまうものもある。そのため、口腔内における有用常在菌を死滅させること無く残存させるとともに、選択的に口腔内の病原性細菌に対して抗菌効果を奏するようにして口腔内の常在菌のバランスを良好に保つように調整可能な、優れた選択的抗菌効果を奏する口腔用組成物、口腔ケア製品及び食品が求められている。 For example, mouthwashes with antibacterial effects, commonly called mouthwashes, are commercially available for use in preventing periodontal disease, etc., but these mouthwashes contain various antibacterial agents. There is. Some antibacterial agents contained in mouthwashes have a wide range of effects against all bacteria in the oral cavity, sterilizing even beneficial resident bacteria, and some may disrupt the balance of resident bacteria in the oral cavity. be. Therefore, we maintain a good balance of the indigenous bacteria in the oral cavity by allowing the beneficial bacteria in the oral cavity to remain without being killed, and by selectively exerting an antibacterial effect against pathogenic bacteria in the oral cavity. There is a need for oral compositions, oral care products, and foods that exhibit superior selective antibacterial effects that can be tailored to specific needs.
そこで、本発明は、上記現状の課題に鑑みてなされたものであり、口腔内における有用常在菌を残存させるとともに、選択的に口腔内の病原性細菌に対して抗菌効果を奏するようにして口腔内の常在菌のバランスを良好に保つように調整可能な、優れた選択的抗菌効果を奏する口腔常在菌調整剤を含む口腔用組成物、口腔ケア製品及び食品を提供することを目的とする。 Therefore, the present invention has been made in view of the above-mentioned current problems, and is designed to maintain useful indigenous bacteria in the oral cavity and selectively exert an antibacterial effect against pathogenic bacteria in the oral cavity. The purpose of the present invention is to provide oral compositions, oral care products, and foods that contain an oral resident bacteria regulating agent that exhibits excellent selective antibacterial effects that can be adjusted to maintain a good balance of resident bacteria in the oral cavity. shall be.
本発明の解決しようとする課題は以上の如くであり、次にこの課題を解決するための手段を説明する。 The problem to be solved by the present invention is as described above, and next, means for solving this problem will be explained.
本発明者らは、前記課題を解決するため種々の検討を重ねた結果、ヒトの唾液中に存在する特定の乳酸菌株について、所定の処方において口腔内における有用常在菌を残存させるとともに、選択的に口腔内の病原性細菌に対して抗菌効果を奏するものがあることを見出し、本発明を完成するに至った。 As a result of various studies in order to solve the above problems, the present inventors have determined that a specific lactic acid bacteria strain present in human saliva can be used in a predetermined formulation to retain useful indigenous bacteria in the oral cavity and to select The present inventors have discovered that there are some substances that have an antibacterial effect against pathogenic bacteria in the oral cavity, and have completed the present invention.
即ち、本発明の口腔常在菌調整剤を含む口腔用組成物においては、
ラクトバチルス・ラムノーサスKO3株(NITEBP-771)の菌体、菌体培養物、又はこれらの抽出物を口腔常在菌調整剤として含有し、
前記ラクトバチルス・ラムノーサスKO3株の組成物全体に対する配合率が0.6~2.5重量%であり、
口腔内の非病原性細菌の生育を抑制することなく病原性細菌の生育を抑制するように選択的に作用する上記口腔常在菌調整剤を含む口腔用組成物であって、
前記病原性細菌が、歯周病原性細菌であるポルフィロモナス ジンジバリス及び/又はプレボテラ インターメディアであり、
前記非病原性細菌が、ストレプトコッカス ミティス及びストレプトコッカス オラリスから選ばれるミティスグループの連鎖球菌であり、
前記歯周病原性細菌と共に前記非病原性細菌を殺菌する非選択的な殺菌作用を奏する非選択的殺菌剤を含まない、口腔内の非病原性細菌の生育を抑制することなく病原性細菌の生育を抑制するように選択的に作用するものである。
That is, in the oral composition containing the oral resident bacteria regulator of the present invention,
Contains Lactobacillus rhamnosus KO3 strain (NITEBP-771) bacterial cells, bacterial cell culture, or extracts thereof as an oral resident bacteria regulator,
The blending ratio of the Lactobacillus rhamnosus KO3 strain to the entire composition is 0.6 to 2.5% by weight,
An oral composition comprising the oral cavity resident bacteria regulator that selectively acts to suppress the growth of pathogenic bacteria without suppressing the growth of non-pathogenic bacteria in the oral cavity,
The pathogenic bacteria are periodontal pathogenic bacteria Porphyromonas gingivalis and/or Prevotella intermedia,
the non-pathogenic bacterium is a streptococcus of the mitis group selected from Streptococcus mitis and Streptococcus oralis;
It does not contain a non-selective bactericidal agent that has a non-selective bactericidal effect that sterilizes the non-pathogenic bacteria together with the periodontal pathogenic bacteria. It acts selectively to suppress growth .
本発明の口腔常在菌調整剤を含む口腔用組成物においては、
前記非選択的殺菌剤は、塩化セチルピリジニウム、塩化ベンゼトニウム、塩化ベンザルコニウム、又はイソプロピルメチルフェノールから選択されるものである。
In the oral composition containing the oral resident bacteria regulator of the present invention,
The non-selective disinfectant is selected from cetylpyridinium chloride, benzethonium chloride, benzalkonium chloride, or isopropylmethylphenol .
本発明の口腔内の非病原性細菌の生育を抑制することなく病原性細菌の生育を抑制するように選択的に作用する口腔ケア製品においては、上記口腔用組成物を含むものである。 The oral care product of the present invention that selectively acts to suppress the growth of pathogenic bacteria without suppressing the growth of non-pathogenic bacteria in the oral cavity contains the above-mentioned oral composition.
本発明の口腔内の非病原性細菌の生育を抑制することなく病原性細菌の生育を抑制するように選択的に作用する食品においては、上記口腔用組成物を含むものである。 The food of the present invention that selectively acts to inhibit the growth of pathogenic bacteria without inhibiting the growth of non-pathogenic bacteria in the oral cavity contains the oral composition described above.
本発明によれば、口腔内における非病原性細菌を残存させるとともに、選択的に口腔内の病原性細菌に対して抗菌効果を奏するようにして口腔内の常在菌のバランスを良好に保つように調整可能な、優れた選択的抗菌効果を奏する口腔常在菌調整剤を含む口腔用組成物を提供することができる。 According to the present invention, non-pathogenic bacteria in the oral cavity remain, and an antibacterial effect is selectively exerted on pathogenic bacteria in the oral cavity to maintain a good balance of resident bacteria in the oral cavity. It is possible to provide an oral cavity composition containing an oral resident bacteria regulating agent that exhibits excellent selective antibacterial effects that can be adjusted to
次に、発明の実施の形態を説明する。 Next, embodiments of the invention will be described.
本発明の口腔用組成物は、口腔常在菌調整作用を奏する口腔常在菌調整剤の一例であるラクトバチルス・ラムノーサスに属する乳酸菌の菌株を含むものである。詳細は後述するが、本発明の口腔用組成物は、口腔内における非病原性細菌である有用常在菌を残存させるとともに、選択的に口腔内の病原性細菌に対して抗菌効果を奏するようにして口腔内の常在菌のバランスを良好に保つように調整するように作用する。すなわち、本発明の口腔用組成物は、選択的抗菌作用を有しており、口腔内の非病原性細菌の生育に影響を与えることなく選択的に歯周病原性細菌の生育を抑制する優れた抗菌効果を奏する。 The oral composition of the present invention contains a strain of lactic acid bacteria belonging to Lactobacillus rhamnosus, which is an example of an oral resident bacteria regulating agent that exhibits an oral resident bacteria regulating effect. Although the details will be described later, the oral composition of the present invention not only retains useful non-pathogenic bacteria in the oral cavity, but also selectively exerts an antibacterial effect against pathogenic bacteria in the oral cavity. It acts to maintain a good balance of resident bacteria in the oral cavity. That is, the oral composition of the present invention has a selective antibacterial effect, and has the advantage of selectively suppressing the growth of periodontal pathogenic bacteria without affecting the growth of non-pathogenic bacteria in the oral cavity. It also has an antibacterial effect.
ここで、「口腔常在菌調整」とは、上述のように口腔内における有用常在菌を残存させるとともに、選択的に口腔内の病原性細菌に対して抗菌効果を奏するようにして口腔内の常在菌のバランスを良好に保つように調整することであり、具体的には、病原性細菌に対して選択的抗菌作用を奏することで非病原性常在菌の生育に影響が及ばないことによって、口腔内の常在菌叢の菌叢バランスを病原性の低い菌叢に調整することである。
また、「選択的抗菌」とは、口腔内の病原性細菌に対して抗菌活性を有し、その一方で非病原性細菌に対する抗菌活性は有さないことによって、非病原性細菌の生育を抑制することなく病原性細菌の生育を選択的に抑制する抗菌作用である。
Here, "regulating oral resident bacteria" refers to maintaining useful resident bacteria in the oral cavity as described above, as well as selectively exerting an antibacterial effect against pathogenic bacteria in the oral cavity. The aim is to maintain a good balance of resident bacteria in the bacteria, and specifically, by exerting a selective antibacterial effect against pathogenic bacteria, the growth of non-pathogenic resident bacteria is not affected. By doing so, the balance of the normal bacterial flora in the oral cavity is adjusted to a bacterial flora with low pathogenicity.
In addition, "selective antibacterial" means that it has antibacterial activity against pathogenic bacteria in the oral cavity, but does not have antibacterial activity against non-pathogenic bacteria, thereby suppressing the growth of non-pathogenic bacteria. It has an antibacterial effect that selectively suppresses the growth of pathogenic bacteria without causing any damage.
本発明における口腔常在菌調整剤は、抗菌作用を有するため口腔用組成物の抗菌成分として使用可能である。
なお、本実施形態において、「抗菌」とは、細菌及び真菌類の殺菌又除菌、もしくはこれらの発生・生育・増殖を抑制することを含めて、最も広義に解釈されるべきであり、如何なる意味においても限定されない。
The oral cavity resident bacteria regulating agent of the present invention has an antibacterial effect and can therefore be used as an antibacterial component of an oral composition.
In addition, in this embodiment, "antibacterial" should be interpreted in the broadest sense, including the sterilization or eradication of bacteria and fungi, or the suppression of their occurrence, growth, and proliferation. It is not limited in meaning either.
本発明に関するラクトバチルス・ラムノーサスに属する乳酸菌の菌株は乳酸菌の一例である。この菌株は、独立行政法人製品評価技術基盤機構 特許微生物寄託センターに寄託されている。
具体的には、ラクトバチルス・ラムノーサスの一例として、当該特許微生物寄託センターにNITE BP-771として寄託されたラクトバチルス・ラムノーサス(Lactobacillus rhamnosus)KO3株が挙げられる(以下、単にKO3株ともいう)。
The strain of lactic acid bacteria belonging to Lactobacillus rhamnosus related to the present invention is an example of lactic acid bacteria. This strain has been deposited at the Patent Microorganism Depositary of the National Institute of Technology and Evaluation.
Specifically, an example of Lactobacillus rhamnosus is the Lactobacillus rhamnosus KO3 strain deposited at the Patent Microorganism Depositary as NITE BP-771 (hereinafter also simply referred to as KO3 strain).
KO3株は、ヒト唾液中から分離されたものであり、ヒト由来の乳酸菌である。KO3株は、16S rRNAの塩基配列がラクトバチルス・ラムノーサス(Lactobacillus rhamnosus)strain IDCC3201の塩基配列と1485/1485の間で100%の相同性を示し、グラム染色後の顕鏡下においてグラム陽性桿菌の様相を呈することから、ラクトバチルス・ラムノーサス(Lactobacillus rhamnosus)と同定された。KO3株の主な菌学的性質を以下に示す。
1)グラム陽性乳酸桿菌、2)ホモ型乳酸発酵、3)カタラーゼ陰性、4)芽胞形成能無し、5)好気条件下でも培養可、6)菌体外多糖類を産生。
The KO3 strain was isolated from human saliva and is a human-derived lactic acid bacterium. The KO3 strain shows 100% homology between the base sequence of Lactobacillus rhamnosus (Lactobacillus rhamnosus) strain IDCC3201 and 1485/1485, and the KO3 strain shows 100% homology between 1485/1485 and the base sequence of Lactobacillus rhamnosus strain IDCC3201. Based on its appearance, it was identified as Lactobacillus rhamnosus. The main mycological properties of the KO3 strain are shown below.
1) Gram-positive lactobacillus, 2) Homotype lactic acid fermentation, 3) Catalase negative, 4) No spore-forming ability, 5) Can be cultured under aerobic conditions, 6) Produces extracellular polysaccharides.
本発明においては、上記乳酸菌の菌体を、乳酸菌培養の常法に従って培養し、得られた培養物から遠心分離等の集菌手段によって分離されたものをそのまま用いることのみならず、当該培養・発酵液(培養上清)、その濃縮液や、菌体を酵素や物理的手段を用いて処理した細胞質や細胞壁画分も用いることができる。また、生菌体のみならず死菌体であってもよい。 In the present invention, the cells of the lactic acid bacteria described above are cultured according to a conventional method for culturing lactic acid bacteria, and the resulting culture is separated by a bacterial collection means such as centrifugation, and not only can it be used as it is, but also the cultured and Fermentation broth (culture supernatant), its concentrate, and cytoplasm and cell wall fraction obtained by treating bacterial cells with enzymes or physical means can also be used. Furthermore, not only viable cells but also dead cells may be used.
本発明のKO3株を培養する培地には、果汁培地、野菜汁培地、牛乳培地、脱脂粉乳培地又は乳成分を含む培地、これを含まない半合成培地等種々の培地を用いることができる。このような培地としては、脱脂乳を還元して加熱殺菌した還元脱脂乳培地、酵母エキスを添加した脱脂粉乳培地、MRS培地、GAM培地等を例示することができる。
培養方法は、静置培養又はpHを一定にした中和培養や、回分培養及び連続培養等、菌体が良好に生育する条件であれば、特に制限はない。
As a medium for culturing the KO3 strain of the present invention, various media can be used, such as a fruit juice medium, a vegetable juice medium, a milk medium, a skim milk powder medium, a medium containing a milk component, and a semi-synthetic medium not containing this. Examples of such a medium include a reduced skim milk medium obtained by reducing and heat sterilizing skim milk, a skim milk powder medium supplemented with yeast extract, an MRS medium, a GAM medium, and the like.
The culture method is not particularly limited as long as the conditions allow for good growth of the bacterial cells, such as static culture, neutralized culture with a constant pH, batch culture, and continuous culture.
本発明のKO3株の菌体又は菌体培養物の抽出物とは、菌体又は菌体培養物を、溶媒抽出することにより得られる各種溶剤抽出液、その希釈液、その濃縮液又はその乾燥末を意味するものである。 The extract of bacterial cells or bacterial cell cultures of the KO3 strain of the present invention refers to various solvent extracts obtained by solvent extraction of bacterial cells or bacterial cell cultures, diluted solutions thereof, concentrated solutions thereof, or dried extracts thereof. It means the end.
本発明の抽出物を得るために用いられる抽出溶剤としては、極性溶剤、非極性溶剤のいずれをも使用することができ、これらを混合して用いることもできる。例えば、水;メタノール、エタノール、プロパノール、ブタノール等のアルコール類;プロピレングリコール、ブチレングリコール等の多価アルコール類;アセトン、メチルエチルケトン等のケトン類;酢酸メチル、酢酸エチル等のエステル類;テトラヒドロフラン、ジエチルエーテル等の鎖状及び環状エーテル類;ポリエチレングリコール等のポリエーテル類;ヘキサン、シクロヘキサン、石油エーテル等の炭化水素類;ベンゼン、トルエン等の芳香族炭化水素類;ピリジン類等が挙げられ、このうち、酢酸エチル等のエステル類、エタノール等のアルコール類が好ましい。 As the extraction solvent used to obtain the extract of the present invention, either a polar solvent or a non-polar solvent can be used, and a mixture of these solvents can also be used. For example, water; alcohols such as methanol, ethanol, propanol, and butanol; polyhydric alcohols such as propylene glycol and butylene glycol; ketones such as acetone and methyl ethyl ketone; esters such as methyl acetate and ethyl acetate; tetrahydrofuran and diethyl ether chain and cyclic ethers such as; polyethers such as polyethylene glycol; hydrocarbons such as hexane, cyclohexane, petroleum ether; aromatic hydrocarbons such as benzene and toluene; Esters such as ethyl acetate and alcohols such as ethanol are preferred.
抽出条件は、使用する溶剤によっても異なるが、例えば、培養液1質量部に対して1~10質量部の溶剤を用い、0~50℃、好ましくは25~37℃の温度で、0.5時間~3時間抽出するのが好ましい。 Extraction conditions vary depending on the solvent used, but for example, using 1 to 10 parts by mass of the solvent per 1 part by mass of the culture solution, at a temperature of 0 to 50 °C, preferably 25 to 37 °C, and 0.5 Preferably, the extraction time is between 3 hours and 3 hours.
上記の抽出物は、そのまま用いることもできるが、当該抽出物を希釈、濃縮若しくは凍結乾燥した後、必要に応じて粉末又はペースト状に調製して用いることもできる。また、液々分配等の技術により、適宜精製して用いることもできる。 The above-mentioned extract can be used as it is, but after diluting, concentrating or lyophilizing the extract, it can also be prepared into a powder or paste form and used, if necessary. Furthermore, it can be used after being appropriately purified by techniques such as liquid-liquid distribution.
本発明における、「病原性細菌」としては、う蝕菌、歯周病菌及びカンジダ菌等が挙げられる。う蝕菌、歯周病菌及びカンジダ菌は、例えば、う蝕症;歯肉炎、歯周炎等の歯周病、舌炎、鵞口瘡、口角炎等の口腔カンジダ症等の口腔内疾患を引き起こす原因となる原因菌である。また、本発明における、「非病原性細菌」としては、上述したように、ストレプトコッカス ミティス(Streptococcus mitis)、ストレプトコッカス オラリス(Streptococcus oralis)を代表とするミティス(Mitis)グループなどのストレプトコッカス(連鎖球菌)属細菌等が挙げられる。 In the present invention, "pathogenic bacteria" include cariogenic bacteria, periodontal disease bacteria, Candida bacteria, and the like. Cariogenic bacteria, periodontal bacteria, and Candida bacteria are causes of oral diseases such as caries; periodontal diseases such as gingivitis and periodontitis; and oral candidiasis such as glossitis, thrush, and angular cheilitis. This is the causative bacteria. In addition, as described above, the "non-pathogenic bacteria" in the present invention include Streptococcus genus, such as the Mitis group represented by Streptococcus mitis and Streptococcus oralis. Examples include bacteria.
また、う蝕菌としては、ストレプトコッカス・ミュータンス(Streptococcus mutans)、ストレプトコッカス・ソブリナス(Streptococcus sobrinus)、歯周病菌としてはポルフィロモナス・ジンジバリス(Porphyromonas gingivalis)、プレボテラ・インターメディア(Prevotella intermedia)、トレポネーマ・デンティコーラ(Treponema denticola)、タンネレラ・フォーサイセンシス(Tannerella forsythensis)、アクチノバチルス・アクチノミセテムコミタンス(Actinobacillus actinomycetemcomitans)、フソバクテリウム・ヌクレアタム(Fusobacterium nucleatum)等が挙げられ、カンジダ菌としては、カンジダ・アルビカンス(Candida albicans)、カンジダ・グラブラータ(Candida glabrata)、カンジダ・トロピカリス(Candida tropicalis)等が挙げられる。 In addition, cariogenic bacteria include Streptococcus mutans and Streptococcus sobrinus, and periodontal disease bacteria include Porphyromonas gingivalis and Prevotella.・Intermedia (Prevotella intermedia), Treponema・Treponema denticola, Tannerella forsythensis, Actinobacillus actinomycetemcomitans, Fusobacterium Examples of Candida include Fusobacterium nucleatum, and Candida albicans. (Candida albicans), Candida glabrata (Candida glabrata), Candida tropicalis (Candida tropicalis), and the like.
特許文献1に示すように、KO3株は、う蝕菌であるストレプトコッカス・ミュータンス(Streptococcus mutans)及びストレプトコッカス・ソブリナス(Streptococcus sobrinus)、歯周病菌であるポルフィロモナス・ジンジバリス(Porphyromonas gingivalis)及びカンジダ菌であるカンジダ・アルビカンス(Candida albicans)の何れに対しても増殖抑制効果を有する。また、口腔内の歯肉縁下プラークフォーマーとして知られるFusobacterium. nucleatum等のFusobacterium属細菌の増殖を抑制する作用を有する。
従って、当該KO3株の菌体若しくは菌体培養物又はこれらの抽出物は、口腔内疾患の予防、改善又は治療剤として、或いはう蝕菌、歯周病菌及びカンジダ菌の増殖抑制剤となり得る。斯かる口腔内疾患の予防、改善又は治療剤、蝕菌、歯周病菌及びカンジダ菌の増殖抑制剤は、それ自体で、当該口腔内の病原微生物により引き起こされる、う蝕症、歯周病、口腔カンジダ症等の口腔内疾患の予防、改善又は治療のための、或いはう蝕菌、歯周病菌及びカンジダ菌の増殖抑制のための食品、医薬品、口腔用組成物等として使用でき、或いは食品、医薬品、口腔用組成物に配合するための素材として使用可能である。また、食品は、虫歯、歯周病、その他の口腔内感染症の予防・改善等をコンセプトとし、必要に応じてその旨を表示した健康食品、サプリメント若しくは特定保健用食品等の機能性食品とすることも可能である。
なお、特許文献1には、KO3株による口腔内の非病原性細菌への影響についての記載はない。
As shown in Patent Document 1, the KO3 strain is a cariogenic bacterium, Streptococcus mutans and Streptococcus sobrinus, and a periodontal disease bacterium, Porphyromonas gingivalis. givalis) and Candida It also has a growth-inhibiting effect on the fungus Candida albicans. In addition, Fusobacterium, which is known as a subgingival plaque former in the oral cavity. It has the effect of suppressing the growth of Fusobacterium bacteria such as Nucleatum.
Therefore, the bacterial cells or bacterial cell culture of the KO3 strain, or their extracts can be used as a prophylactic, ameliorating, or therapeutic agent for oral diseases, or as an agent for inhibiting the growth of cariogenic bacteria, periodontal disease bacteria, and Candida bacteria. Such a preventive, ameliorating or therapeutic agent for oral diseases, an agent for inhibiting the growth of caries, periodontal disease bacteria and candida bacteria, can itself be used to prevent caries, periodontal disease, etc. caused by the pathogenic microorganisms in the oral cavity. It can be used as a food, medicine, oral composition, etc. for the prevention, improvement, or treatment of oral diseases such as oral candidiasis, or for inhibiting the growth of cariogenic bacteria, periodontal disease bacteria, and Candida bacteria, or as a food. It can be used as a material for blending into , pharmaceuticals, and oral compositions. In addition, foods are functional foods such as health foods, supplements, and foods for specified health uses that are designed to prevent and improve tooth decay, periodontal disease, and other oral infections, and are labeled accordingly as necessary. It is also possible to do so.
Note that Patent Document 1 does not describe the influence of the KO3 strain on non-pathogenic bacteria in the oral cavity.
KO3株は、口膣内疾患の予防、改善または治療等に優れた効果を発揮する有効成分となる。したがって、口膣内疾患の予防、改善又は治療剤として有用である。本発明にかかわるKO3株は、選択的抗菌作用を有する口腔常在菌調整剤として口腔用組成物に好適に配合することができる。 The KO3 strain is an active ingredient that exhibits excellent effects in preventing, improving, or treating orovaginal diseases. Therefore, it is useful as a preventive, ameliorating or therapeutic agent for orovaginal diseases. The KO3 strain according to the present invention can be suitably incorporated into oral compositions as an oral cavity resident bacteria regulator having selective antibacterial activity.
なお、本発明において、口腔用組成物とは、一般的な歯磨剤、洗口剤、口腔用スプレー、口腔用パスタ、軟膏剤、貼付剤等に加え、口腔内に含んで適用する錠剤、グミ、キャンディー、チューインガム、ゲル状の食品なども含むものである。さらに、これらの口腔用組成物を、ペート状、ゲル状、液状、固体状などの各種形態に適宜調製して用いることができる。 In the present invention, oral compositions include general dentifrices, mouth rinses, oral sprays, oral pasta, ointments, patches, etc., as well as tablets and gummies that are applied in the oral cavity. It also includes candy, chewing gum, gel-like foods, etc. Furthermore, these oral compositions can be suitably prepared and used in various forms such as paste, gel, liquid, and solid.
より具体的には、口腔用組成物として練歯磨、液状歯磨、液体歯磨、潤製歯磨等の歯磨剤、洗口液及びマウスウォッシュ等を含む洗口剤、口腔用スプレーの一例であるマウススプレー、口腔用パスタ、口中清涼剤、うがい薬、デンタルリンス、口腔保湿剤、義歯安定剤、義歯潤滑剤、人口唾液、軟膏剤、貼付剤、歯磨きシート/ナップや、錠菓、トローチ、タブレット等の錠剤、カプセル剤、グミ、キャンディー、顆粒、チューインガム、ゲル状の食品、飲料、保存食等の各種剤型に調製し得る。すなわち、本実施形態における口腔用組成物は、その剤型を口腔ケア製品、食品または食品形態等にすることができる。特に、歯磨剤、洗口剤、錠剤、グミ、キャンディー、ゲル状の食品として、特に歯磨剤、洗口剤として好適に調製される。なお、抗菌効果付与の点で錠剤、グミ、キャンディーなどは、口腔内に留まる時間が比較的長いため、好適に使用することができる。
なお、口腔ケア製品とは、健全な口腔、歯、及び歯肉を促進するか、息を爽やかにするか、又は口腔疾患を予防若しくは治療するために使用される、口腔内をケアする製品をいう。
また、本実施形態において挙げた、口腔用組成物を含むもしくは口腔用組成物からなる口腔ケア製品又は食品を構成することもできる。これにより、本発明の効果を有する口腔ケア製品又は食品を実現することができる。
More specifically, oral compositions include toothpastes, liquid toothpastes, liquid toothpastes, dentifrices such as Junsei toothpastes, mouthwashes including mouthwashes and mouthwashes, and mouthsprays which are examples of oral sprays. , oral pasta, mouth fresheners, gargles, dental rinses, oral moisturizers, denture stabilizers, denture lubricants, artificial saliva, ointments, patches, toothpaste sheets/naps, tablets, pastilles, tablets, etc. It can be prepared into various dosage forms such as tablets, capsules, gummies, candies, granules, chewing gum, gel foods, drinks, and preserved foods. That is, the oral composition according to the present embodiment can be in the form of an oral care product, a food product, a food product, or the like. In particular, it is suitably prepared as a dentifrice, a mouthwash, a tablet, a gummy, a candy, a gel-like food, especially as a dentifrice or a mouthwash. Note that tablets, gummies, candies, and the like can be preferably used because they remain in the oral cavity for a relatively long time in terms of imparting antibacterial effects.
Oral care products refer to oral care products used to promote a healthy mouth, teeth, and gums, to freshen breath, or to prevent or treat oral diseases. .
Moreover, it is also possible to constitute an oral care product or food containing or consisting of the oral composition mentioned in this embodiment. Thereby, an oral care product or food having the effects of the present invention can be realized.
また、このような種々の剤型の製剤を調製するには、本発明の菌体や培養物の作用を妨げない範囲で、他の薬学的に許容される賦形剤、結合剤、増量剤、崩壊剤、界面活性剤、滑沢剤、分散剤、緩衝剤、保存剤、香料、被膜剤、担体、希釈剤等を適宜組み合わせて用いることができる。 In addition, in order to prepare such various dosage forms, other pharmaceutically acceptable excipients, binders, and fillers may be added to the extent that they do not interfere with the action of the bacterial cells or cultures of the present invention. , a disintegrant, a surfactant, a lubricant, a dispersant, a buffer, a preservative, a fragrance, a coating agent, a carrier, a diluent, and the like can be used in appropriate combinations.
上記口腔用組成物において、KO3株の菌体若しくは菌体培養物又はこれらの抽出物の組成物全体に対する配合率としては、病原性細菌を抗菌するために0.001~10%(重量%、以下同様)が好ましく、より好ましくは0.01~5.0%である。
また、上記口腔用組成物の剤型が、例えば液体の場合には、KO3株の濃度を0.001g/ml~0.025g/mlとすることが好ましい。
In the above-mentioned oral composition, the proportion of the KO3 strain bacterial cells or bacterial cell culture or their extracts in the entire composition is 0.001 to 10% (wt%, (same below) is preferred, and more preferably 0.01 to 5.0%.
Furthermore, when the dosage form of the oral composition is, for example, a liquid, the concentration of the KO3 strain is preferably 0.001 g/ml to 0.025 g/ml.
本発明では、KO3株の菌体若しくは菌体培養物又はこれらの抽出物を配合する場合、その配合率が上記範囲内であると、より優れた選択的抗菌効果を付与することができる。なお、前記KO3株の菌体若しくは菌体培養物又はこれらの抽出物の配合率が高いほど、抗菌効果が向上するが、配合率が高すぎると有用な常在菌を減少させることになる。 In the present invention, when the bacterial cells of the KO3 strain, the bacterial cell culture, or their extracts are blended, a superior selective antibacterial effect can be imparted if the blending ratio is within the above range. Incidentally, the higher the blending ratio of the bacterial cells of the KO3 strain, the bacterial cell culture, or their extracts, the better the antibacterial effect will be, but if the blending ratio is too high, useful resident bacteria will be reduced.
また、口腔用組成物には、上記成分に加えて、本発明の効果を妨げない範囲において、剤型に応じた適宜なその他の公知成分を必要に応じて配合し、通常の方法で調製できる。 In addition to the above-mentioned components, the oral composition can be prepared by a conventional method by adding other known components suitable for the dosage form as necessary, within a range that does not impede the effects of the present invention. .
歯磨剤には、例えば、研磨剤、粘稠剤、粘結剤、界面活性剤、更に必要によって甘味剤、着色剤、防腐剤、保湿剤、香料、有効成分等を配合できる。洗口剤には、粘稠剤、粘結剤、界面活性剤、更に必要によって甘味剤、着色剤、防腐剤、香料、有効成分等を配合できる。錠剤には、ソルビトール、マルチトール等の糖アルコール、セルロース、乳糖、デキストリン等の賦形剤、微粒二酸化ケイ素、酸味料、甘味剤、着色剤、乳化剤、増粘剤、ゲル化剤、果汁、香辛料、有効成分等を配合できる。グミには、グリセリン、ゼラチン等の増粘剤やゲル化剤、糖類、酸味料、甘味剤、着色剤、乳化剤、果汁、有効成分等を配合できる。チューインガムには、ガムベース、食用ガム質等の結合剤、甘味剤、着色剤、酸味料、保存料、光沢剤、香料、有効成分等を配合でき、必要に応じて糖衣で被覆してもよい。 The dentifrice may contain, for example, an abrasive, a thickener, a binder, a surfactant, and if necessary, a sweetener, a coloring agent, a preservative, a humectant, a fragrance, an active ingredient, and the like. The mouthwash may contain a thickening agent, a binder, a surfactant, and if necessary, a sweetener, a coloring agent, a preservative, a fragrance, an active ingredient, and the like. Tablets contain sugar alcohols such as sorbitol and maltitol, excipients such as cellulose, lactose, and dextrin, fine silicon dioxide, acidulants, sweeteners, colorants, emulsifiers, thickeners, gelling agents, fruit juice, and spices. , active ingredients, etc. can be blended. Gummies can contain thickeners and gelling agents such as glycerin and gelatin, sugars, acidulants, sweeteners, colorants, emulsifiers, fruit juice, active ingredients, and the like. The chewing gum may contain a gum base, a binder such as an edible gum, a sweetening agent, a coloring agent, an acidulant, a preservative, a brightening agent, a flavoring agent, an active ingredient, etc., and may be coated with a sugar coating if necessary.
歯磨剤に用いられる研磨剤としては、例えば、シリカ系研磨剤、リン酸カルシウム系研磨剤、炭酸カルシウム系研磨剤などが挙げられ、その配合量は、通常、練歯磨では2~50%、液状歯磨では0~30%である。 Examples of abrasives used in toothpastes include silica-based abrasives, calcium phosphate-based abrasives, and calcium carbonate-based abrasives, and the amount of these is usually 2 to 50% in toothpastes and 2% to 50% in liquid toothpastes. It is 0-30%.
粘稠剤としては、例えば、ソルビトール、キシリトール等の糖アルコール、グリセリン、プロピレングリコール等の多価アルコールが挙げられる。その配合量は、通常、5~50%である。 Examples of the thickening agent include sugar alcohols such as sorbitol and xylitol, and polyhydric alcohols such as glycerin and propylene glycol. Its blending amount is usually 5 to 50%.
粘結剤としては、例えば、カルボキシメチルセルロースナトリウム等のセルロース誘導体、キサンタンガム等のガム類、ゲル化性シリカ、ゲル化性アルミニウムシリカなどの有機又は無機粘結剤を配合できる。その配合量は、通常、0.5~10%である。 As the binder, for example, cellulose derivatives such as sodium carboxymethylcellulose, gums such as xanthan gum, organic or inorganic binders such as gelling silica, gelling aluminum silica, etc. can be blended. Its blending amount is usually 0.5 to 10%.
また、界面活性剤としては、例えば、口腔用組成物に一般的に用いられるアニオン性界面活性剤、ノニオン性界面活性剤、カチオン性界面活性剤、両性界面活性剤を配合できる。アニオン性界面活性剤としては、ラウリル硫酸ナトリウム等のアルキル硫酸塩、N-アシルサルコシン酸塩などが挙げられる。ノニオン性界面活性剤としては、糖脂肪酸エステル、糖アルコール脂肪酸エステル、ソルビタン脂肪酸エステル、グリセリン脂肪酸エステル、ポリオキシエチレン硬化ヒマシ油等のポリオキシエチレン脂肪酸エステル、ポリオキシエチレン高級アルコールエーテル、脂肪酸アルカノールアミドなどが挙げられる。カチオン性界面活性剤としては、アルキルアンモニウム塩等、両性界面活性剤としては、ベタイン系やイミダゾリン系のものが挙げられる。界面活性剤の配合量は、通常、0~10%、特に0.01~5%である。 Furthermore, as the surfactant, for example, anionic surfactants, nonionic surfactants, cationic surfactants, and amphoteric surfactants commonly used in oral compositions can be blended. Examples of the anionic surfactant include alkyl sulfates such as sodium lauryl sulfate, N-acyl sarcosinates, and the like. Examples of nonionic surfactants include sugar fatty acid esters, sugar alcohol fatty acid esters, sorbitan fatty acid esters, glycerin fatty acid esters, polyoxyethylene fatty acid esters such as polyoxyethylene hydrogenated castor oil, polyoxyethylene higher alcohol ethers, fatty acid alkanolamides, etc. can be mentioned. Examples of cationic surfactants include alkylammonium salts, and examples of amphoteric surfactants include betaine and imidazoline surfactants. The amount of surfactant added is usually 0 to 10%, particularly 0.01 to 5%.
甘味剤としては、例えば、キシリトール、スクラロース、サッカリンナトリウム等が挙げられる。着色剤としては、赤色2号、青色1号、黄色4号等が、防腐剤としては、パラオキシ安息香酸エステル等が挙げられる。 Examples of sweeteners include xylitol, sucralose, and sodium saccharin. Examples of the coloring agent include Red No. 2, Blue No. 1, Yellow No. 4, etc., and examples of the preservative include paraoxybenzoic acid ester.
香料または着香剤としては、例えば、ペパーミント油、スペアミント油、アニス油、ユーカリ油、ケイヒ油、ウイキョウ油、チョウジ油(丁字油)、ウィンターグリーン油、カシア油、クローブ油、タイム油、セージ油、レモン油、オレンジ油、ハッカ油、カルダモン油、コリアンダー油、マンダリン油、ライム油、ラベンダー油、ローズマリー油、ローレル油、カモミル油、キャラウェイ油、マジョラム油、ベイ油、レモングラス油、オリガナム油、パインニードル油、ネロリ油、ローズ油、ジャスミン油、グレープフルーツ油、スウィーティー油、柚油、イリスコンクリート、アブソリュートペパーミント、アブソリュートローズ、オレンジフラワー等の天然香料、及びこれら天然香料の加工処理(前溜部カット、後溜部カット、分留、液液抽出、エッセンス化、粉末香料化等)した香料、及び、メントール、カルボン、アネトール、シネオール、サリチル酸メチル、シンナミックアルデヒド、オイゲノール、3-l-メントキシプロパン-1,2-ジオール、チモール、リナロール、リナリールアセテート、リモネン、メントン、メンチルアセテート、N-置換-パラメンタン-3-カルボキサミド、ピネン、オクチルアルデヒド、シトラール、プレゴン、カルビールアセテート、アニスアルデヒド、エチルアセテート、エチルブチレート、アリルシクロヘキサンプロピオネート、メチルアンスラニレート、エチルメチルフェニルグリシデート、バニリン、ウンデカラクトン、ヘキサナール、ブタノール、イソアミルアルコール、ヘキセノール、ジメチルサルファイド、シクロテン、フルフラール、トリメチルピラジン、エチルラクテート、エチルチオアセテート等の単品香料、更に、ストロベリーフレーバー、アップルフレーバー、アップルミントフレーバー、バナナフレーバー、パイナップルフレーバー、グレープフレーバー、マンゴーフレーバー、バターフレーバー、ミルクフレーバー、フルーツミックスフレーバー、トロピカルフルーツフレーバー等の調合香料等が挙げられ、口腔用組成物に用いられる公知の香料素材を使用できる。
これら香料の配合量は、通常、歯磨剤、洗口剤では0.0001~1%であり、また、錠剤、グミ、チューインガムでは0.001~50%であり、また、上記香料素材を使用した賦香用香料は、組成中に0.1~10%使用するのが好ましい。
Flavoring or flavoring agents include, for example, peppermint oil, spearmint oil, anise oil, eucalyptus oil, cinnamon oil, fennel oil, clove oil, wintergreen oil, cassia oil, clove oil, thyme oil, sage oil. , lemon oil, orange oil, peppermint oil, cardamom oil, coriander oil, mandarin oil, lime oil, lavender oil, rosemary oil, laurel oil, chamomile oil, caraway oil, marjoram oil, bay oil, lemongrass oil, origanum. natural fragrances such as oil, pine needle oil, neroli oil, rose oil, jasmine oil, grapefruit oil, sweetie oil, yuzu oil, iris concrete, absolute peppermint, absolute rose, orange flower, and processing of these natural fragrances (pre-distilled). Fragrances such as partial cut, rear distillation cut, fractional distillation, liquid-liquid extraction, essence, powder fragrance, etc.), and menthol, carvone, anethole, cineole, methyl salicylate, cinnamic aldehyde, eugenol, 3-l-menth. Xypropane-1,2-diol, thymol, linalool, linaryl acetate, limonene, menthone, menthyl acetate, N-substituted-paramenthane-3-carboxamide, pinene, octylaldehyde, citral, pulegone, carbyl acetate, anisaldehyde, Ethyl acetate, ethyl butyrate, allyl cyclohexane propionate, methyl anthranilate, ethyl methyl phenyl glycidate, vanillin, undecalactone, hexanal, butanol, isoamyl alcohol, hexenol, dimethyl sulfide, cyclotene, furfural, trimethyl pyrazine, ethyl Single flavorings such as lactate and ethylthioacetate, as well as combinations of strawberry flavor, apple flavor, apple mint flavor, banana flavor, pineapple flavor, grape flavor, mango flavor, butter flavor, milk flavor, mixed fruit flavor, tropical fruit flavor, etc. Examples include perfumes, and known perfume materials used in oral compositions can be used.
The amount of these fragrances is usually 0.0001 to 1% in toothpastes and mouthwashes, and 0.001 to 50% in tablets, gummies, and chewing gum. It is preferable to use 0.1 to 10% of the fragrance in the composition.
有効成分としては、口腔用組成物に通常配合される公知のものを本発明の効果を妨げない範囲で配合できる。有効成分としては、イソプロピルメチルフェノール等の非イオン性殺菌剤、塩化セチルピリジニウム等のカチオン性殺菌剤、トラネキサム酸、イプシロンアミノカプロン酸等の抗炎症剤、デキストラナーゼ等の酵素、フッ化ナトリウム、モノフルオロリン酸ナトリウム等のフッ化物、水溶性リン酸化合物、銅化合物、硝酸カリウム、乳酸アルミニウム、各種ビタミン類、植物抽出物などが挙げられる。なお、上記有効成分の配合量は、本発明の選択的抗菌効果を妨げない範囲で設定される。 As the active ingredient, any known active ingredient that is commonly blended into oral compositions can be blended within a range that does not impede the effects of the present invention. Active ingredients include non-ionic disinfectants such as isopropyl methylphenol, cationic disinfectants such as cetylpyridinium chloride, anti-inflammatory agents such as tranexamic acid and epsilon aminocaproic acid, enzymes such as dextranase, sodium fluoride, monomers, etc. Examples include fluorides such as sodium fluorophosphate, water-soluble phosphoric acid compounds, copper compounds, potassium nitrate, aluminum lactate, various vitamins, and plant extracts. The amount of the above-mentioned active ingredient is set within a range that does not interfere with the selective antibacterial effect of the present invention.
なお、本発明の口腔用組成物においては、歯周病原性細菌と共に非病原性細菌をも殺菌する非選択的殺菌剤の配合は制限することが好ましい。前記非選択的殺菌剤は、従来から広く口腔用製剤に使用されている殺菌剤に多く見られ、例えば塩化セチルピリジニウム、塩化ベンゼトニウム、塩化ベンザルコニウム等のカチオン性殺菌剤が挙げられる。また、イソプロピルメチルフェノール等の非イオン性殺菌剤なども挙げられる。 In addition, in the oral composition of the present invention, it is preferable to limit the amount of non-selective sterilizers that sterilize non-pathogenic bacteria as well as periodontal pathogenic bacteria. The non-selective disinfectants are commonly found in disinfectants that have been widely used in oral preparations, and examples include cationic disinfectants such as cetylpyridinium chloride, benzethonium chloride, and benzalkonium chloride. Also included are nonionic disinfectants such as isopropylmethylphenol.
これら非選択的殺菌剤は、本発明の口腔用組成物が選択的抗菌効果を有する範囲で配合することができるが、配合する場合は組成物全体の0.1%以下、特に0.05%以下が好ましく、配合しないことが最も好ましい。これらの非選択的殺菌剤は、病原性の歯周病原性細菌と共に非病原性の口腔内常在菌をも殺菌してしまう、非選択的な殺菌作用を奏するため、これら殺菌剤が配合されていると、口腔内の菌バランスが崩れて口腔内常在菌叢が制御できなくなり、菌交代症の可能性が生じてくる場合がある。従って、本発明においては、上記非選択的殺菌剤が配合されていないほうが、選択的抗菌効果による口腔内常在菌叢のバランス制御にはより好ましい。
以下に、実施例及び抗菌力の試験結果を示し、本発明についてより詳細に説明する。
These non-selective bactericidal agents can be blended within the range in which the oral composition of the present invention has a selective antibacterial effect, but when blended, they account for 0.1% or less, particularly 0.05% of the total composition. The following are preferred, and it is most preferred not to include it. These non-selective bactericidal agents have a non-selective bactericidal effect that kills non-pathogenic oral bacteria as well as pathogenic periodontal pathogenic bacteria, so these bactericidal agents are combined. If this occurs, the bacterial balance in the oral cavity may be disrupted, and the normal oral flora may become uncontrollable, leading to the possibility of bacterial replacement. Therefore, in the present invention, it is more preferable that the above-mentioned non-selective bactericidal agent is not blended in order to control the balance of the oral flora due to the selective antibacterial effect.
Examples and antibacterial activity test results are shown below, and the present invention will be explained in more detail.
以下、試験例などにより本発明をさらに詳しく説明するが、本発明はこれらによりなんら限定されるものではない。また、特に断らない限り「%」は重量%を意味する。 Hereinafter, the present invention will be explained in more detail using test examples and the like, but the present invention is not limited to these in any way. Further, unless otherwise specified, "%" means % by weight.
(菌体の調製)
MRS培地(Difco社)を121℃で20分間滅菌した後、ラクトバチルス・ラムノーサス(Lactobacillus rhamnosus)KO3株(NITE BP-771)を接種し、37℃で、48時間大気下で培養した後、蒸留水・超純水・緩衝液などで洗浄後、菌体を調製した。
(Preparation of bacterial cells)
After sterilizing MRS medium (Difco) at 121°C for 20 minutes, Lactobacillus rhamnosus KO3 strain (NITE BP-771) was inoculated, cultured at 37°C for 48 hours in the atmosphere, and then distilled. After washing with water, ultrapure water, buffer, etc., bacterial cells were prepared.
以上に示されるKO3株は、公知の手法によって調整することができ、その方法を限定するものではない。 The KO3 strain shown above can be prepared by a known method, and the method is not limited.
<口腔内の非病原性細菌に対するKO3株の抗菌力試験>
口腔内の非病原性細菌であるストレプトコッカス ミティス及びストレプトコッカス オラリスの2種の各菌に対するKO3株の抗菌力を試験する。
<Antibacterial activity test of KO3 strain against non-pathogenic bacteria in the oral cavity>
The antibacterial activity of strain KO3 against two non-pathogenic bacteria in the oral cavity, Streptococcus mitis and Streptococcus oralis, will be tested.
[試験(1)]
KO3株の抗菌力を評価した。なお、対照としてKO3株を配合しない精製水を用いた。
[Test (1)]
The antibacterial activity of the KO3 strain was evaluated. Note that purified water without the KO3 strain was used as a control.
1.検体
本発明に係る口腔用組成物の一例としてKO3株を2.5%の配合率で配合した評価用のKO3株含有洗口液(基材:精製水)を調整して、市販されている洗口液である市販品A、Bと合わせて抗菌力試験を行った。
検体1)2.5%KO3株含有株洗口液(ノンアルコール系、基材:水、殺菌剤無し)
検体2)市販品A(ノンアルコール系、薬用成分として塩化セチルピリジニウム(殺菌剤CPC)配合)
検体3)市販品B(ノンアルコール系、薬用成分として1,8‐シネオール、塩化亜鉛等配合)
2.試験概要
検体に試験菌液を接種後(以下、「試験液」という。)、所定時間後に試験液中の生菌数を測定した。また、あらかじめ予備試験(中和条件の確認)を行い、検体の影響を受けずに生菌数を測定できる条件を確認した。
3.試験条件
a)試験菌として、以下の非病原性細菌の菌種を用いた。
(1)ストレプトコッカス ミティス(Streptococcus mitis NBRC 106071)
(2)ストレプトコッカス オラリス(Streptococcus oralis JCM 12997)
b)試験菌液:試験菌をTrypticase soy agar(BBL)で37℃±1℃、18~24時間後嫌気培養した後、精製水に浮遊させ、菌数が108~109/mLとなるように調整した。
c)試験液:検体10mLに試験菌液0.1mLを接種
d)保存条件:30秒、3分(室温)
e)対照:精製水
f)中和条件:SCDLP培地(日本製薬株式会社)で10倍希釈(なお、試験菌がストレプトコッカス ミティスの場合の検体2は100倍希釈)
g)生菌数測定:Trypticase soy agar、混釈平板培養法
4.試験結果
試験菌としてストレプトコッカス ミティス及びストレプトコッカス オラリスを用いたそれぞれの試験結果を表1、表2に示す。なお、試験液をSCDLP培地で10倍(試験菌がストレプトコッカス ミティスの場合の検体2は100倍希釈)に希釈することにより、検体の影響を受けずに生菌数の測定ができることを予備試験により確認した。
なお、以下の表1、表2の「対象」における検体1)、検体2)、検体3)とは、それぞれ上述した2.5%KO3株含有株洗口液、市販品A、市販品Bのことである。
1. Specimen As an example of the oral composition according to the present invention, a mouthwash containing the KO3 strain for evaluation (base material: purified water) containing the KO3 strain at a blending rate of 2.5% was prepared and commercially available. An antibacterial activity test was conducted with commercially available mouthwashes A and B.
Sample 1) Strain mouthwash containing 2.5% KO3 strain (non-alcoholic, base material: water, no disinfectant)
Sample 2) Commercial product A (non-alcoholic, containing cetylpyridinium chloride (bactericide CPC) as a medicinal ingredient)
Sample 3) Commercial product B (non-alcoholic, contains medicinal ingredients such as 1,8-cineole and zinc chloride)
2. Test Overview After inoculating the specimen with the test bacterial solution (hereinafter referred to as the "test solution"), the number of viable bacteria in the test solution was measured after a predetermined period of time. In addition, a preliminary test (confirmation of neutralization conditions) was conducted in advance to confirm the conditions under which the number of viable bacteria could be measured without being affected by the sample.
3. Test conditions a) The following non-pathogenic bacterial species were used as test bacteria.
(1) Streptococcus mitis (Streptococcus mitis NBRC 106071)
(2) Streptococcus oralis (Streptococcus oralis JCM 12997)
b) Test bacteria solution: After culturing the test bacteria anaerobically in trypticase soy agar (BBL) at 37°C ± 1°C for 18 to 24 hours, suspend it in purified water until the number of bacteria reaches 10 8 to 10 9 /mL. I adjusted it as follows.
c) Test solution: Inoculate 10 mL of sample with 0.1 mL of test bacterial solution d) Storage conditions: 30 seconds, 3 minutes (room temperature)
e) Control: Purified water f) Neutralization conditions: 10-fold dilution with SCDLP medium (Nippon Pharmaceutical Co., Ltd.) (In addition, when the test bacteria is Streptococcus mitis, sample 2 is diluted 100-fold)
g) Measurement of viable bacteria count: Trypticase soy agar, pour plate culture method 4. Test Results Tables 1 and 2 show the test results using Streptococcus mitis and Streptococcus oralis as test bacteria. Preliminary tests have shown that by diluting the test solution 10 times with SCDLP medium (100 times dilution for sample 2 when the test bacterium is Streptococcus mitis), the number of viable bacteria can be measured without being affected by the sample. confirmed.
In addition, sample 1), sample 2), and sample 3) in "Target" in Tables 1 and 2 below refer to the above-mentioned strain mouthwash containing 2.5% KO3 strain, commercial product A, and commercial product B, respectively. It is about.
表1の結果に基づく棒グラフを図1に示す。図1(a)では、各検体における保存時間(30秒、3分)毎の生菌数(対数表示)を比較して表示した。図1(b)では、試験開始時を100%とし、各検体における保存時間毎の生菌数の変化率を比較して表示した。ここで、図1においては、開始時において、4本の棒グラフを示しているが、左から右に順に、2.5%KO3株含有洗口液、市販品A、市販品B、対照に関する試験結果である。 A bar graph based on the results in Table 1 is shown in FIG. In FIG. 1(a), the number of viable bacteria (logarithm display) for each specimen at each storage time (30 seconds, 3 minutes) is compared and displayed. In FIG. 1(b), the time at the start of the test is set as 100%, and the rate of change in the number of viable bacteria for each specimen at each storage time is compared and displayed. Here, in FIG. 1, four bar graphs are shown at the beginning, and in order from left to right, tests regarding 2.5% KO3 strain-containing mouthwash, commercial product A, commercial product B, and control. This is the result.
表2の結果に基づく棒グラフを図2に示す。図2(a)では、各検体における保存時間(30秒、3分)毎の生菌数(対数表示)を比較して表示した。図2(b)では、試験開始時を100%とし、各検体における保存時間毎の生菌数の変化率を比較して表示した。ここで、図2においては、開始時において、4本の棒グラフを示しているが、左から右に順に、2.5%KO3株含有洗口液、市販品A、市販品B、対照に関する試験結果である。 A bar graph based on the results in Table 2 is shown in FIG. In FIG. 2(a), the number of viable bacteria (logarithm display) for each specimen at each storage time (30 seconds, 3 minutes) is compared and displayed. In FIG. 2(b), the time at the start of the test is set as 100%, and the rate of change in the number of viable bacteria for each specimen at each storage time is compared and displayed. Here, in FIG. 2, four bar graphs are shown at the beginning, and in order from left to right, the test for mouthwash containing 2.5% KO3 strain, commercial product A, commercial product B, and control. This is the result.
表1(図1)、表2(図2)においては、保存時間毎において生菌数が多いほど有用な常在菌であるストレプトコッカス ミティスやストレプトコッカス オラリスが残存していることを表しており、好ましい結果となる。また、本実施例では口腔用組成物の一例として洗口液を試験したものであるが、洗口液は、一般的に短時間(例えば、30秒程度)で口腔内全体に行き渡らしてから口腔外に吐き出すようにして使用される。そのため、例えば、開始後30秒後に着目すると、表1及び表2では市販品A、市販品Bはすでにストレプトコッカス ミティス及びストレプトコッカス オラリスが検出できないレベルにまで減少しており、有用な常在菌がほぼ残存していないが、本発明に係る2.5%KO3株含有株洗口液では、まだ多くの有用な常在菌が残存しているが確認できた(30秒後の生菌率において、ストレプトコッカス ミティスが83%、ストレプトコッカス オラリスが75%)。
このことから、本発明における選択的抗菌効果は、本試験方法において、評価に用いた非病原性細菌に対して、30秒時に生菌率が0.7以上となる効果である。
In Table 1 (Figure 1) and Table 2 (Figure 2), the higher the number of viable bacteria at each storage time, the more useful resident bacteria Streptococcus mitis and Streptococcus oralis remain, which is preferable. result. In addition, in this example, a mouthwash was tested as an example of an oral composition, but mouthwashes are generally spread throughout the oral cavity in a short period of time (for example, about 30 seconds) and then It is used by spitting it out of the mouth. Therefore, for example, if we focus on 30 seconds after the start, in Tables 1 and 2, Streptococcus mitis and Streptococcus oralis have already decreased to an undetectable level in Commercial Product A and Commercial Product B, and almost all useful resident bacteria are present. However, in the strain mouthwash containing 2.5% KO3 strain according to the present invention, it was confirmed that many useful indigenous bacteria still remained (in the viable bacteria rate after 30 seconds, 83% for Streptococcus mitis and 75% for Streptococcus oralis).
From this, the selective antibacterial effect in the present invention is an effect in which the viable bacteria rate is 0.7 or more at 30 seconds with respect to the non-pathogenic bacteria used for evaluation in this test method.
[試験(2)]
上述した抗菌力試験方法に基づいて、検体として2.5%KO3株含有株洗口液と、市販品Aを用いて、かつ保存条件を1分、5分、10分(室温)として抗菌力試験を行った結果を図3に示す。図3のグラフより明らかなように、本発明に係る2.5%KO3株含有株洗口液の場合は、10分経過後においても有用な常在菌であるストレプトコッカス ミティスやストレプトコッカス オラリスが残存している。一方、市販品Aにおいては1分後においてすでに残存していない。すなわち、2.5%KO3株含有株洗口液においては有用常在菌に対して抗菌作用が緩和されていることが確認された。それに対して、市販品Aにおいては殺菌剤である塩化セチルピリジニウムの影響により歯周病等の原因菌だけでなく有用常在菌までも殺菌してしまったと推察される。
[Test (2)]
Based on the above-mentioned antibacterial activity test method, antibacterial activity was determined using a strain mouthwash containing 2.5% KO3 strain and commercially available product A as specimens, and under storage conditions of 1 minute, 5 minutes, and 10 minutes (room temperature). The results of the test are shown in Figure 3. As is clear from the graph in Figure 3, in the case of the strain mouthwash containing 2.5% KO3 strain according to the present invention, useful indigenous bacteria such as Streptococcus mitis and Streptococcus oralis remain even after 10 minutes. ing. On the other hand, commercially available product A does not remain after 1 minute. That is, it was confirmed that the antibacterial effect against useful indigenous bacteria was alleviated in the strain mouthwash containing 2.5% KO3 strain. On the other hand, in commercially available product A, it is presumed that not only the causative bacteria of periodontal disease, etc. but also useful resident bacteria were sterilized due to the effect of cetylpyridinium chloride, which is a bactericide.
[試験(3)]
上述した抗菌力試験方法に基づいて、本発明に係る口腔用組成物の一例としてKO3株を0.6%、1.25%、2.5%の各配合率で配合した評価用のKO3株含有洗口液(主成分:精製水)を調製して、かつ保存条件を1分、5分、10分(室温)として抗菌力の濃度依存性試験を行った。試験結果を図4に示す。ここで、図4においては、各時間において、4本の棒グラフを示しているが、左から右に順に、2.5%KO3株含有洗口液、1.25%KO3株含有洗口液、0.6%KO3株含有洗口液、対照に関する試験結果である。図4に示す試験結果により、ストレプトコッカス ミティスに対してはKO3株の濃度依存性があり、KO3株の配合濃度が高いほど、生菌数が減少していることが確認できた。一方、ストレプトコッカス オラリスに対してはKO3株の濃度依存性が確認できなかった。このように、KO3株においては、上述したようにう蝕菌や歯周病菌等の病原性細菌に対して優れた増殖抑制効果を有するが、非病原性細菌で有用な常在菌であるストレプトコッカス ミティスに対しては生菌数を減少させてしまうことが確認できた。このことを考慮すると、KO3株の口腔用組成物全体に対する配合率は0.01~5.0重量%が好ましく、より好ましくは0.1~4.0重量%、更に好ましくは0.5~3.0重量%である。こうして、KO3株は所定の処方において口腔用組成物に含有させることで、選択的抗菌剤として作用し、口腔常在菌調整作用を有することが確認できた。
[Test (3)]
Based on the above-mentioned antibacterial activity test method, the KO3 strain for evaluation was blended with each blending ratio of 0.6%, 1.25%, and 2.5% as an example of the oral composition according to the present invention. A concentration-dependent test of antibacterial activity was conducted by preparing a mouthwash containing the following ingredients (main component: purified water) and storing them for 1 minute, 5 minutes, and 10 minutes (room temperature). The test results are shown in Figure 4. Here, in FIG. 4, four bar graphs are shown at each time, and from left to right, mouthwash containing 2.5% KO3 strain, mouthwash containing 1.25% KO3 strain, These are test results regarding a control mouthwash containing 0.6% KO3 strain. From the test results shown in FIG. 4, it was confirmed that Streptococcus mitis was dependent on the concentration of the KO3 strain, and the higher the concentration of the KO3 strain, the lower the number of viable bacteria. On the other hand, no concentration dependence of the KO3 strain on Streptococcus oralis could be confirmed. In this way, the KO3 strain has an excellent growth-inhibiting effect against pathogenic bacteria such as cariogenic bacteria and periodontal disease bacteria as described above, but it has an excellent growth-inhibiting effect against pathogenic bacteria such as cariogenic bacteria and periodontal disease bacteria, but it also inhibits the growth of Streptococcus, which is a useful non-pathogenic bacteria. It was confirmed that the number of viable bacteria against Mitis was reduced. Considering this, the blending ratio of the KO3 strain to the entire oral composition is preferably 0.01 to 5.0% by weight, more preferably 0.1 to 4.0% by weight, even more preferably 0.5 to 5.0% by weight. It is 3.0% by weight. In this way, it was confirmed that the KO3 strain acts as a selective antibacterial agent and has an effect of regulating oral bacteria when included in an oral composition in a predetermined formulation.
試験(1)、(2)、(3)の結果について考察すると、本発明に係るKO3株含有洗口液と市販品A、Bと比較すると、例えば、図1に示すように、2.5%KO3株水溶液では有用常在菌の生菌数が維持されており、当該有用常在菌に悪影響を及ぼさないことが確認された。これにより、口膣内での使用を想定した口腔用組成物としてのKO3株の優位性が確認された。すなわち、KO3株は有用常在菌の生育阻害が緩和されているのに対して、市販品A、Bは配合されている殺菌剤の影響によって顕著に有用常在菌の生育を阻害していると考えられる。
以上により、KO3株含有洗口液は、市販品A、Bと比較して、有用常在菌であるストレプトコッカス ミティスやストレプトコッカス オラリスを残存させることができることが確認できた。
Considering the results of tests (1), (2), and (3), when comparing the mouthwash containing the KO3 strain according to the present invention with commercially available products A and B, for example, as shown in FIG. It was confirmed that the viable count of useful indigenous bacteria was maintained in the aqueous solution of %KO3 strain, and that it did not have an adverse effect on the useful indigenous bacteria. This confirmed the superiority of the KO3 strain as an oral composition intended for oral or vaginal use. In other words, while the KO3 strain has less inhibition of the growth of useful indigenous bacteria, commercial products A and B significantly inhibit the growth of useful indigenous bacteria due to the effects of the fungicides contained in them. it is conceivable that.
As a result of the above, it was confirmed that the mouthwash containing the KO3 strain was able to retain useful indigenous bacteria such as Streptococcus mitis and Streptococcus oralis compared to commercially available products A and B.
また、本発明に係るKO3株含有洗口液は、上述したように選択的抗菌作用を有し、例えば、口膣内において病原性細菌等の原因菌に対しては抗菌作用が効果的に働くとともに、口腔内常在菌(本実施形態では、ストレプトコッカス ミティス、ストレプトコッカス オラリス)に対しては抗菌作用が緩和される。このように、本発明に係るKO3株含有洗口液は口膣内の有用な常在菌を死滅させることなく安定して存在させて、常在菌の病原性細菌に対する耐性を維持することができる。 In addition, the mouthwash containing the KO3 strain according to the present invention has a selective antibacterial effect as described above, and for example, the antibacterial effect effectively acts against causative bacteria such as pathogenic bacteria in the oral cavity. At the same time, the antibacterial action against bacteria resident in the oral cavity (in this embodiment, Streptococcus mitis and Streptococcus oralis) is alleviated. As described above, the mouthwash containing the KO3 strain according to the present invention can stably exist the useful indigenous bacteria in the oral cavity without killing them, and can maintain the resistance of the indigenous bacteria to pathogenic bacteria. can.
また、本発明に係るKO3株は、口膣内常在菌に影響を与えず、常在菌により病原性微生物の侵入や増殖を抑制させるプロバイオティクスの概念を達成する素材として提供することができる。 In addition, the KO3 strain according to the present invention can be provided as a material that achieves the concept of probiotics that does not affect the resident bacteria in the oral cavity and suppresses the invasion and proliferation of pathogenic microorganisms by the resident bacteria. can.
本発明の口腔用組成物には、添加剤としては、本発明の効果を阻害しない範囲でその形態に応じた、基剤(水、アルコール等)、潤滑材、清掃剤(デキストリン等)、pH調整剤(クエン酸Na等)、清掃剤、香味剤、香料、甘味料(キシリトール、スクラロース等)、可溶可剤、保存剤(メチルパラベン、エチルパラベン、プロピルパラベン等)、保湿剤、乳化剤、分散剤、増粘剤、酸化防止剤、紫外線吸収剤等からなる群から選ばれる1種又は2種以上のものを加えることができる。 The oral composition of the present invention may include a base (water, alcohol, etc.), lubricant, cleaning agent (dextrin, etc.), pH Conditioners (Na citrate, etc.), cleaning agents, flavoring agents, fragrances, sweeteners (xylitol, sucralose, etc.), solubilizers, preservatives (methylparaben, ethylparaben, propylparaben, etc.), humectants, emulsifiers, dispersants One or more selected from the group consisting of additives, thickeners, antioxidants, ultraviolet absorbers, etc. can be added.
以上の如く、本発明によれば、口腔内における非病原性細菌を残存させるとともに、選択的に口腔内の病原性細菌に対して抗菌効果を奏するようにして口腔内の常在菌のバランスを良好に保つように調整可能な、優れた選択的抗菌効果を奏する。 As described above, according to the present invention, non-pathogenic bacteria in the oral cavity remain and the balance of resident bacteria in the oral cavity is maintained by selectively exerting an antibacterial effect on pathogenic bacteria in the oral cavity. Excellent selective antibacterial effect that can be adjusted to maintain good performance.
Claims (4)
前記ラクトバチルス・ラムノーサスKO3株の組成物全体に対する配合率が0.6~2.5重量%であり、
口腔内の非病原性細菌の生育を抑制することなく病原性細菌の生育を抑制するように選択的に作用する上記口腔常在菌調整剤を含む口腔用組成物であって、
前記病原性細菌が、歯周病原性細菌であるポルフィロモナス ジンジバリス及び/又はプレボテラ インターメディアであり、
前記非病原性細菌が、ストレプトコッカス ミティス及びストレプトコッカス オラリスから選ばれるミティスグループの連鎖球菌であり、
前記歯周病原性細菌と共に前記非病原性細菌を殺菌する非選択的な殺菌作用を奏する非選択的殺菌剤を含まない、口腔内の非病原性細菌の生育を抑制することなく病原性細菌の生育を抑制するように選択的に作用する口腔用組成物。 Contains Lactobacillus rhamnosus KO3 strain (NITEBP-771) bacterial cells, bacterial cell culture, or extracts thereof as an oral resident bacteria regulator,
The blending ratio of the Lactobacillus rhamnosus KO3 strain to the entire composition is 0.6 to 2.5% by weight,
An oral composition comprising the oral cavity resident bacteria regulator that selectively acts to suppress the growth of pathogenic bacteria without suppressing the growth of non-pathogenic bacteria in the oral cavity,
The pathogenic bacteria are periodontal pathogenic bacteria Porphyromonas gingivalis and/or Prevotella intermedia,
the non-pathogenic bacterium is a streptococcus of the mitis group selected from Streptococcus mitis and Streptococcus oralis;
It does not contain a non-selective bactericidal agent that has a non-selective bactericidal effect that sterilizes the non-pathogenic bacteria together with the periodontal pathogenic bacteria. An oral composition that selectively acts to inhibit growth .
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