WO2017090704A1 - Novel lactobacillales - Google Patents

Novel lactobacillales Download PDF

Info

Publication number
WO2017090704A1
WO2017090704A1 PCT/JP2016/084894 JP2016084894W WO2017090704A1 WO 2017090704 A1 WO2017090704 A1 WO 2017090704A1 JP 2016084894 W JP2016084894 W JP 2016084894W WO 2017090704 A1 WO2017090704 A1 WO 2017090704A1
Authority
WO
WIPO (PCT)
Prior art keywords
cells
lactic acid
strain
present
lactococcus lactis
Prior art date
Application number
PCT/JP2016/084894
Other languages
French (fr)
Japanese (ja)
Inventor
篤史 大木
洋介 砂田
和也 上原
早苗 岡田
田中　尚人
Original Assignee
日清食品ホールディングス株式会社
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by 日清食品ホールディングス株式会社 filed Critical 日清食品ホールディングス株式会社
Publication of WO2017090704A1 publication Critical patent/WO2017090704A1/en

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C9/00—Milk preparations; Milk powder or milk powder preparations
    • A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L31/00—Edible extracts or preparations of fungi; Preparation or treatment thereof
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/135—Bacteria or derivatives thereof, e.g. probiotics
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor

Definitions

  • the present invention relates to a strain of lactic acid bacteria derived from cranberries having a high immunomodulatory action, and beverages and foods containing the cells.
  • it relates to Lactococcus lactis.
  • Lactic acid bacteria have been used in various processed foods and drinks such as fermented milk products such as yogurt and various pickles for various purposes such as imparting taste and flavor, enhancing nutrition, and improving food storage stability.
  • fermented milk products such as yogurt and various pickles for various purposes such as imparting taste and flavor, enhancing nutrition, and improving food storage stability.
  • attention has been focused on effective physiological activities such as allergy suppression of lactic acid bacteria, and research is being conducted energetically.
  • Th cells that play an important role in the immune system are classified into Th1 cells and Th2 cells according to the cytokines produced.
  • Th1 cells produce cytokines mainly related to cellular immunity such as interferon ⁇ (hereinafter referred to as “IFN- ⁇ ”), and Th2 cells mainly produce cytokines related to humoral immunity such as interleukin 4 (hereinafter referred to as “IL-4”).
  • IFN- ⁇ interferon ⁇
  • IL-4 interleukin 4
  • Th1 cells and Th2 cells antagonize each other and maintain balance to maintain homeostasis of the immune system, but when Th1 cells predominate, autoimmune disease, and when Th2 cells predominate, IgE production increases Allergy develops (Non-patent Document 1).
  • An object of the present invention is to provide a cranberry-derived lactic acid bacterium having a high immunomodulatory action and a food or drink containing the lactic acid bacterium.
  • the first invention of the present application is a Lactococcus genus having an immunomodulatory action derived from cranberries, and more specifically, Lactococcus lactis T21 strain (NITE BP-02164).
  • the applicant of the present application also intends foods and drinks containing the lactic acid bacteria. That is, the second invention of the present application is a food or drink containing Lactococcus lactis T21 strain (NITE BP-02164).
  • the lactic acid bacterium of the present invention has a high immunoregulatory effect. When ingested, allergy can be suppressed by activating immune function.
  • FIG. 1 is a diagram comparing IL-12 production (pg / ml) between the strain of the present invention and a comparative strain.
  • FIG. 2 is a diagram comparing the skim milk medium growth of the strain of the present invention and the comparative strain.
  • Lactococcus lactis T21 strain (NITE BP-02164)
  • the lactic acid bacterium of the present invention is lactococcus lactis.
  • Lactococcus lactis strain T21 Among the lactic acid bacteria belonging to Lactococcus lactis, Lactococcus lactis strain T21.
  • the symbol T21 referred to in the present invention is a number uniquely assigned to the strain by Nissin Foods Holdings. This Lactococcus lactis strain T21 was first isolated from cranberries by one of the inventors.
  • the Lactococcus lactis T21 strain of the present invention is deposited under the following conditions. (1) Depositary Institution: National Institute of Technology and Technology Patent Microorganisms Deposit Center (2) Contact: 2-9-8 Kazusa Kamashi, Kisarazu City, Chiba Prefecture 292-0818 Japan (3) Accession Number: NITE BP -02164 (4) Display for identification: T21 (5) Date of original deposit: November 20, 2015 (6) Date of transfer to deposit under the Budapest Treaty: October 12, 2016
  • the bacteriological properties of the Lactococcus lactis strain T21 of the present invention are as follows: As shown in Tables 1 and 2. This bacteriological property is based on the method described in Bergey's manual of systematic bacteriology Vol. 2 (1986).
  • the Lactococcus lactis T21 strain of the present invention has a high immunomodulating action, that is, a high production induction ability of IL-12, as shown in the experimental examples described later. Confirmation of IL-12 production-inducing ability was performed by the following test method.
  • ⁇ Preparation of bacterial cells The cells were cultured in MRS medium (Difco Lactobacilli MRS Broth) shown in Table 3 at 30 ° C. for 48 hours. Next, the grown cells were collected by centrifugation, and the separated cells were washed three times with sterilized water and frozen. Then, it lyophilized
  • MRS medium Difco Lactobacilli MRS Broth
  • J774.1 cells a mouse-derived macrophage-like cell line, were used as cells.
  • J774.1 cells were cultured under conditions of 37 ° C. and CO 2 5%.
  • 2 ml each of 2.5 ⁇ 10 5 cells / ml J774.1 cells was added to each well of the 24-well culture plate, and the dried cell powder obtained above was added to the 0.1 mg / ml J774.1 cell culture medium.
  • the suspension was suspended to a concentration of ml, 100 ⁇ l was added, and the cells were cultured at 37 ° C. under 5% CO 2 for 24 hours. After the culture, the medium was collected and centrifuged at 3500 rpm for 2 minutes to obtain a culture supernatant. IL-12 in the culture supernatant was measured using Mouse Total IL-12 ELISA kit (Thermo scientific).
  • the lactic acid bacteria of the present invention can be used by containing them in food / beverage products. Although the lactic acid bacteria of this invention can be used especially suitably for a drink, fermented milk and a lactic acid bacteria drink are considered, for example.
  • the component standard should be fermented milk (non-fat milk solid content of 8.0% or more) or dairy lactic acid bacteria beverage (non-fat milk solid content of 3.0% or more) 1.0 ⁇ 10 7 production induction capacity / ml or more, 1.0 ⁇ 10 6 cfn / ml or more is required for lactic acid bacteria beverages (non-fat milk solid content less than 3.0%), but in fermented liquid such as milk
  • the above-mentioned number of bacteria can be realized by growing or growing in the form of a final product.
  • the food of the present invention may be in the form of a formulation (for example, powder, granule, capsule, tablet, etc.) by adding an appropriate carrier and additives as necessary together with the lactic acid bacteria. .
  • the lactic acid bacteria of the present invention are also useful for inclusion in foods for specified health use, dietary supplements and the like in addition to general beverages and foods.
  • the lactic acid bacteria of the present invention can also be used in cosmetics such as lotion, pharmaceuticals such as intestinal preparations, daily necessities such as toothpaste, silage, animal feed, and animal feed and plant fertilizers such as plant liquid fertilizer. Applicable.
  • the cranberry-derived lactic acid bacteria (Lactococcus lactis T21 strain) of the present invention have a high immunoregulatory effect. When ingested, allergy can be suppressed by activating immune function.
  • IL-12 production induction evaluation was carried out for Lactococcus lactis T21 strain of the present invention and three Lactococcus lactis comparative strains (LL1, LL2, LL3) owned by the company. .
  • IL-12 production induction evaluation was performed according to the following procedure.
  • the cells were cultured in MRS medium (Difco Lactobacilli MRS Broth) shown in Table 3 at 30 ° C for 48 hours, and the proliferated cells were collected by centrifugation and the separated cells were collected. Washed 3 times with sterilized water and frozen. Then, it lyophilized
  • MRS medium Difco Lactobacilli MRS Broth
  • J774.1 cells were cultured under conditions of 37 ° C. and CO 2 5%. Then, 2 ml each of 2.5 ⁇ 10 5 cells / ml J774.1 cells was added to each well of the 24-well culture plate, and the dried cell powder obtained above was added to the 0.1 mg / ml J774.1 cell culture medium. Each suspension was suspended to a concentration of ml, 100 ⁇ l was added, and the cells were cultured at 37 ° C.
  • IL-12 in the culture supernatant was measured using Mouse Total IL-12 ELISA kit (Thermo scientific).
  • the amount of IL-12 produced by the lactic acid bacterium of the present invention is 6056 pg / ml, compared with other in-house owned Lactococcus lactis. It was confirmed to have a high ability to induce IL-12 production.
  • skim Milk Medium Proliferation Test A skim milk medium proliferation test was performed on the Lactococcus lactis T21 strain of the present invention and three Lactococcus lactis comparative strains (LL1, LL2, LL3) owned by the company.
  • SM skimmed milk
  • 10% SM (skimmed milk) medium was inoculated and cultured at 30 ° C. for 48 hours, and the growth on the skimmed milk medium was evaluated by the number of lactic acid bacteria and pH.
  • the number of strains of the strain of the present invention (Lactococcus lactis T21 strain) was 7.6 ⁇ 10 8 cfu / ml, which was a level with no problem in producing fermented milk and lactic acid bacteria beverages.

Abstract

[Problem] The purpose of the present invention is to provide cranberry-derived lactobacillales that have high immunomodulatory effects and that can suitably be used in food and drink, and to provide food and drink containing the lactobacillales. [Solution] Lactococcus lactis strain T21 (NITE BP-02164), which contains lactobacillales having high immunomodulatory effects, is provided. In addition, food and drink containing these lactobacillales are provided.

Description

新規乳酸菌New lactic acid bacteria
　本発明は、高い免疫調節作用を有するクランベリー由来の乳酸菌の菌株及びその菌体を含有する飲料、食品に関するものである。特に、ラクトコッカス・ラクティスに関するものである。 The present invention relates to a strain of lactic acid bacteria derived from cranberries having a high immunomodulatory action, and beverages and foods containing the cells. In particular, it relates to Lactococcus lactis.
　乳酸菌は、ヨーグルトなどの発酵乳製品、各種漬物類など、多くの加工飲食品において、味や風味の付与、栄養の強化、食品の保存性改善など様々な目的で用いられてきた。また、乳酸菌のアレルギー抑制といった有効な生理活性について近年注目が集まっており、精力的に研究が進められている。 Lactic acid bacteria have been used in various processed foods and drinks such as fermented milk products such as yogurt and various pickles for various purposes such as imparting taste and flavor, enhancing nutrition, and improving food storage stability. In recent years, attention has been focused on effective physiological activities such as allergy suppression of lactic acid bacteria, and research is being conducted energetically.
　免疫系において重要な役割を担っているTh細胞は、産生するサイトカインによりTh1細胞とTh2細胞に分類される。Th1細胞はインターフェロンγ（以下「IFN-γ」という）など主に細胞性免疫関わるサイトカインを、Th2細胞はインターロイキン4（以下「IL-4」という）など主に液性免疫に関わるサイトカインを産生する。Th2細胞の産生するIL-4は、B細胞からのIgE産生を促進するが、Th1細胞の産生するIFN-γは、IgE産生を抑制する。Th1細胞とTh2細胞はお互いに拮抗し、バランスを保つことにより免疫系の恒常性が維持されているが、Th1細胞が優位に傾くと自己免疫疾患、Th2細胞が優位に傾くとIgE産生が増えてアレルギーが発症される（非特許文献１）。 Th cells that play an important role in the immune system are classified into Th1 cells and Th2 cells according to the cytokines produced. Th1 cells produce cytokines mainly related to cellular immunity such as interferon γ (hereinafter referred to as “IFN-γ”), and Th2 cells mainly produce cytokines related to humoral immunity such as interleukin 4 (hereinafter referred to as “IL-4”). To do. IL-4 produced by Th2 cells promotes IgE production from B cells, whereas IFN-γ produced by Th1 cells suppresses IgE production. Th1 cells and Th2 cells antagonize each other and maintain balance to maintain homeostasis of the immune system, but when Th1 cells predominate, autoimmune disease, and when Th2 cells predominate, IgE production increases Allergy develops (Non-patent Document 1).
　したがって、アレルギーを抑制するためには、Th1細胞の数あるいはTh1細胞由来のサイトカイン産生量を増やすことによりTh1細胞とTh2細胞のバランスを正常に戻す必要があり、Th1細胞とTh2細胞の共通の前駆細胞（Th0細胞）をTh1細胞に分化誘導することによりTh1細胞を優位にする働きのある、抗原提示細胞由来のインターロイキン12（以下「IL-12」という）産生を促進する乳酸菌株が求められている。 Therefore, in order to suppress allergies, it is necessary to restore the balance between Th1 cells and Th2 cells by increasing the number of Th1 cells or the production of cytokines derived from Th1 cells, and a common precursor for Th1 and Th2 cells. There is a need for a lactic acid strain that promotes the production of interleukin 12 (hereinafter referred to as “IL-12”) derived from antigen-presenting cells, which has the function of predominating Th1 cells by inducing differentiation of cells (Th0 cells) into Th1 cells. ing.
　本発明は、高い免疫調節作用を有するクランベリー由来の乳酸菌及び当該乳酸菌を含有する飲食品を提供することを目的とする。 An object of the present invention is to provide a cranberry-derived lactic acid bacterium having a high immunomodulatory action and a food or drink containing the lactic acid bacterium.
　本発明者らは、種々の果実、野菜等を分離源として、乳酸菌について検討を行った結果、クランベリー由来の乳酸菌に高い免疫調節作用を有する株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 studies on lactic acid bacteria using various fruits, vegetables and the like as separation sources, the present inventors have found that cranberry-derived lactic acid bacteria have high immunoregulatory strains and have completed the present invention. It was.
　すなわち、本願第一の発明は、クランベリー由来の免疫調節作用を有するラクトコッカス属菌であり、より詳細には、ラクトコッカス・ラクティスT21株（NITE BP-02164）である。 That is, the first invention of the present application is a Lactococcus genus having an immunomodulatory action derived from cranberries, and more specifically, Lactococcus lactis T21 strain (NITE BP-02164).
　さらに、本願出願人は、当該乳酸菌を含む飲食品も意図している。すなわち、本願第二の発明は、ラクトコッカス・ラクティスT21株（NITE BP-02164）を含有する飲食品である。 Furthermore, the applicant of the present application also intends foods and drinks containing the lactic acid bacteria. That is, the second invention of the present application is a food or drink containing Lactococcus lactis T21 strain (NITE BP-02164).
　本発明の乳酸菌は、高い免疫調節作用を有する。摂取した場合に免疫機能を賦活化することによりアレルギーを抑制することが可能となる。 The lactic acid bacterium of the present invention has a high immunoregulatory effect. When ingested, allergy can be suppressed by activating immune function.
図１は、本発明の菌株と比較菌株のIL-12産生量（pg/ml）を比較した図である。FIG. 1 is a diagram comparing IL-12 production (pg / ml) between the strain of the present invention and a comparative strain. 図２は、本発明の菌株と比較菌株のスキムミルク培地増殖性を比較した図である。FIG. 2 is a diagram comparing the skim milk medium growth of the strain of the present invention and the comparative strain.
　以下、本発明を詳細に説明する。
１．ラクトコッカス・ラクティスT21株（NITE BP-02164）
　本発明の乳酸菌は、ラクトコッカス・ラクティス（lactococcus lactis）である。特にラクトコッカス・ラクティスに属する乳酸菌のうち、ラクトコッカス・ラクティスT21株である。本発明にいうT21の記号は日清食品ホールディングス株式会社で独自に菌株に付与した番号である。本ラクトコッカス・ラクティスT21株はクランベリーより本発明者の一人によって初めて分離されたものである。 Hereinafter, the present invention will be described in detail.
1. Lactococcus lactis T21 strain (NITE BP-02164)
The lactic acid bacterium of the present invention is lactococcus lactis. Among the lactic acid bacteria belonging to Lactococcus lactis, Lactococcus lactis strain T21. The symbol T21 referred to in the present invention is a number uniquely assigned to the strain by Nissin Foods Holdings. This Lactococcus lactis strain T21 was first isolated from cranberries by one of the inventors.
　本発明のラクトコッカス・ラクティスT21株は、下記の条件で寄託されている。
（１）寄託機関名：独立行政法人製品技術基盤機構　特許微生物寄託センター
（２）連絡先：〒２９２－０８１８　千葉県木更津市かずさ鎌足２－５－８　１２２号室
（３）受託番号：ＮＩＴＥ　ＢＰ－０２１６４
（４）識別のための表示：Ｔ２１
（５）原寄託日：２０１５年１１月２０日
（６）ブダペスト条約に基づく寄託への移管日：２０１６年１０月１２日
　本発明のラクトコッカス・ラクティスT21株の菌学的性質は、以下の表１及び２に示す通りである。本菌学的性質は、Bergey’s manual of systematic bacteriology Vol.2(1986)に記載の方法による。 The Lactococcus lactis T21 strain of the present invention is deposited under the following conditions.
(1) Depositary Institution: National Institute of Technology and Technology Patent Microorganisms Deposit Center (2) Contact: 2-9-8 Kazusa Kamashi, Kisarazu City, Chiba Prefecture 292-0818 Japan (3) Accession Number: NITE BP -02164
(4) Display for identification: T21
(5) Date of original deposit: November 20, 2015 (6) Date of transfer to deposit under the Budapest Treaty: October 12, 2016 The bacteriological properties of the Lactococcus lactis strain T21 of the present invention are as follows: As shown in Tables 1 and 2. This bacteriological property is based on the method described in Bergey's manual of systematic bacteriology Vol. 2 (1986).
Figure JPOXMLDOC01-appb-T000001
Figure JPOXMLDOC01-appb-T000001
Figure JPOXMLDOC01-appb-T000002
Figure JPOXMLDOC01-appb-T000002
２．IL-12産生能試験
　本発明のラクトコッカス・ラクティスT21株は、後述する実験例に示すように、高い免疫調節作用、すなわちIL-12の高い産生誘導能力を有する。IL-12の産生誘導能力の確認については以下の試験方法によって行った。 2. IL-12 Production Ability Test The Lactococcus lactis T21 strain of the present invention has a high immunomodulating action, that is, a high production induction ability of IL-12, as shown in the experimental examples described later. Confirmation of IL-12 production-inducing ability was performed by the following test method.
＜菌体の調製＞
　菌体を表３に示すMRS培地（Difco Lactobacilli MRS Broth）で30℃ ・48時間培養した。次いで、増殖した菌体を遠心分離して集菌し、分離した菌体を滅菌水にて3回洗浄し、凍結した。その後、凍結乾燥機を用いて凍結乾燥し、乾燥菌体粉末を得た。 <Preparation of bacterial cells>
The cells were cultured in MRS medium (Difco Lactobacilli MRS Broth) shown in Table 3 at 30 ° C. for 48 hours. Next, the grown cells were collected by centrifugation, and the separated cells were washed three times with sterilized water and frozen. Then, it lyophilized | freeze-dried using the freeze dryer and the dry microbial cell powder was obtained.
Figure JPOXMLDOC01-appb-T000003
Figure JPOXMLDOC01-appb-T000003
＜IL-12産生誘導評価＞
　IL-12産生誘導評価では、細胞としてマウス由来マクロファージ様細胞株であるJ774.1細胞を使用した。はじめに、10%の非動化FBS（Invitrogen）とPenicillin-Streptomycin（Invitrogen）、NaHCO3（Sigma）、L- glutamine（Invitrogen）を加えたRPMI1640培地（日水製薬）を用いて、J774.1細胞を37℃ CO2 5%の条件下で培養した。そして、24穴の培養プレートの1穴ごとに2.5×105 cells/mlのJ774.1細胞を2mlずつ加え、そこに上記で得た乾燥菌体粉末をJ774.1細胞培養培地で0.1 mg/mlの濃度になるよう懸濁して100μl添加し37℃ 5%CO2条件下で24時間培養した。培養後培地を回収し3500 rpmで2分間遠心分離を行い、培養上清を得た。Mouse Total IL-12 ELISA kit（Thermo scientific）を用いて培養上清中のIL-12を測定した。 <IL-12 production induction evaluation>
In the evaluation of IL-12 production induction, J774.1 cells, a mouse-derived macrophage-like cell line, were used as cells. First, using 77% RPMI1640 medium (Nissui Pharmaceutical) with 10% immobilized FBS (Invitrogen), Penicillin-Streptomycin (Invitrogen), NaHCO 3 (Sigma), and L-glutamine (Invitrogen), J774.1 cells Was cultured under conditions of 37 ° C. and CO 2 5%. Then, 2 ml each of 2.5 × 10 5 cells / ml J774.1 cells was added to each well of the 24-well culture plate, and the dried cell powder obtained above was added to the 0.1 mg / ml J774.1 cell culture medium. The suspension was suspended to a concentration of ml, 100 μl was added, and the cells were cultured at 37 ° C. under 5% CO 2 for 24 hours. After the culture, the medium was collected and centrifuged at 3500 rpm for 2 minutes to obtain a culture supernatant. IL-12 in the culture supernatant was measured using Mouse Total IL-12 ELISA kit (Thermo scientific).
３．スキムミルク培地での増殖性試験
＜スキムミルク培地増殖性試験＞
　前培養液を10％SM（スキムミルク）培地に植菌し、これを30℃ ・48時間培養し、乳酸菌数、pHによってミルク培地での増殖性を評価した。 3. Proliferation test on skim milk medium <Skim milk medium growth test>
The preculture was inoculated into 10% SM (skimmed milk) medium, which was cultured at 30 ° C. for 48 hours, and the growth on the milk medium was evaluated by the number of lactic acid bacteria and pH.
４．飲食品
　本発明の乳酸菌は飲食品に含有せしめて使用することができる。本発明の乳酸菌は特に飲料に好適に用いることができるが、例えば、発酵乳及び乳酸菌飲料が考えられる。現行の乳及び乳製品の成分規格等に関する省令では、成分規格として発酵乳（無脂乳固形分8.0％以上のもの）や乳製品乳酸菌飲料（無脂乳固形分3.0％以上のもの）であれば1.0×107産生誘導能力/ml以上、乳酸菌飲料（無脂乳固形分3.0％未満のもの）であれば1.0×106cfn/ml以上必要とされるが、乳などのはっ酵液中で増殖させたり、最終製品の形態で増殖させたりすることによって上記の菌数を実現することができる。また、発酵乳及び乳酸菌飲料以外にも、バター等の乳製品、マヨネーズ等の卵加工品、バターケーキ等の菓子パン類等にも利用することができる。また、即席麺やクッキー等の加工食品にも好適に利用することができる。上記の他、本発明の食品は、前記乳酸菌と共に、必要に応じて適当な担体及び添加剤を添加して製剤化された形態（例えば、粉末、顆粒、カプセル、錠剤等）であってもよい。 4). Food / Beverage Products The lactic acid bacteria of the present invention can be used by containing them in food / beverage products. Although the lactic acid bacteria of this invention can be used especially suitably for a drink, fermented milk and a lactic acid bacteria drink are considered, for example. According to the current ministerial ordinance regarding milk and dairy product component specifications, the component standard should be fermented milk (non-fat milk solid content of 8.0% or more) or dairy lactic acid bacteria beverage (non-fat milk solid content of 3.0% or more) 1.0 × 10 7 production induction capacity / ml or more, 1.0 × 10 6 cfn / ml or more is required for lactic acid bacteria beverages (non-fat milk solid content less than 3.0%), but in fermented liquid such as milk The above-mentioned number of bacteria can be realized by growing or growing in the form of a final product. In addition to fermented milk and lactic acid bacteria beverages, it can also be used for dairy products such as butter, processed egg products such as mayonnaise, and confectionery breads such as butter cake. Moreover, it can utilize suitably also for processed foods, such as instant noodles and a cookie. In addition to the above, the food of the present invention may be in the form of a formulation (for example, powder, granule, capsule, tablet, etc.) by adding an appropriate carrier and additives as necessary together with the lactic acid bacteria. .
　本発明の乳酸菌は、一般の飲料や食品以外にも特定保健用食品、栄養補助食品等に含有させることも有用である。 The lactic acid bacteria of the present invention are also useful for inclusion in foods for specified health use, dietary supplements and the like in addition to general beverages and foods.
　また、本発明の乳酸菌は、食品以外にも化粧水等の化粧品分野、整腸剤等の医薬品分野、歯磨き粉等の日用品分野、サイレージ、動物用餌、植物液体肥料等の動物飼料・植物肥料分野においても応用可能である。 In addition to food, the lactic acid bacteria of the present invention can also be used in cosmetics such as lotion, pharmaceuticals such as intestinal preparations, daily necessities such as toothpaste, silage, animal feed, and animal feed and plant fertilizers such as plant liquid fertilizer. Applicable.
　本発明のクランベリー由来の乳酸菌（ラクトコッカス・ラクティスT21株）は、高い免疫調節作用を有する。摂取した場合に免疫機能を賦活化することによりアレルギーを抑制することが可能となる。 The cranberry-derived lactic acid bacteria (Lactococcus lactis T21 strain) of the present invention have a high immunoregulatory effect. When ingested, allergy can be suppressed by activating immune function.
　以下、本発明の実施例を示すが、本発明は以下の実施例に限定されるものではない。
＜試験例１＞IL-12産生誘導評価
　本発明のラクトコッカス・ラクティスT21株と、自社保有の3つのラクトコッカス・ラクティス比較菌株（LL1、LL2、LL3）についてIL-12産生誘導評価を実施した。 Examples of the present invention will be described below, but the present invention is not limited to the following examples.
<Test Example 1> IL-12 production induction evaluation IL-12 production induction evaluation was carried out for Lactococcus lactis T21 strain of the present invention and three Lactococcus lactis comparative strains (LL1, LL2, LL3) owned by the company. .
　IL-12産生誘導評価は次の手順により行った。本発明の菌株と比較菌株のそれぞれについて、表３に示すMRS培地（Difco Lactobacilli MRS Broth）で30℃ ・48時間培養し、増殖した菌体を遠心分離して集菌し、分離した菌体を滅菌水にて3回洗浄し、凍結した。その後、凍結乾燥機を用いて凍結乾燥し、乾燥菌体粉末を得た。 IL-12 production induction evaluation was performed according to the following procedure. For each of the strains of the present invention and the comparative strain, the cells were cultured in MRS medium (Difco Lactobacilli MRS Broth) shown in Table 3 at 30 ° C for 48 hours, and the proliferated cells were collected by centrifugation and the separated cells were collected. Washed 3 times with sterilized water and frozen. Then, it lyophilized | freeze-dried using the freeze dryer and the dry microbial cell powder was obtained.
　別途、10%の非動化FBS（Invitrogen）とPenicillin-Streptomycin（Invitrogen）、NaHCO3（Sigma）、L- glutamine（Invitrogen）を加えたRPMI1640培地（日水製薬）を用いて、J774.1細胞を37℃ CO2 5%の条件下で培養した。そして、24穴の培養プレートの1穴ごとに2.5×105 cells/mlのJ774.1細胞を2mlずつ加え、そこに上記で得た乾燥菌体粉末をJ774.1細胞培養培地で0.1 mg/mlの濃度になるよう懸濁してそれぞれ100 μl添加し、37℃ 5%CO2条件下で24時間培養した。培養後培地を回収し3500 rpmで2分間遠心分離を行い、培養上清を得た。Mouse Total IL-12 ELISA kit（Thermo scientific）を用いて培養上清中のIL-12を測定した。 Separately, using 77% RPMI1640 medium (Nissui Pharmaceutical) supplemented with 10% immobilized FBS (Invitrogen), Penicillin-Streptomycin (Invitrogen), NaHCO 3 (Sigma), and L-glutmine (Invitrogen), J774.1 cells Was cultured under conditions of 37 ° C. and CO 2 5%. Then, 2 ml each of 2.5 × 10 5 cells / ml J774.1 cells was added to each well of the 24-well culture plate, and the dried cell powder obtained above was added to the 0.1 mg / ml J774.1 cell culture medium. Each suspension was suspended to a concentration of ml, 100 μl was added, and the cells were cultured at 37 ° C. under 5% CO 2 for 24 hours. After the culture, the medium was collected and centrifuged at 3500 rpm for 2 minutes to obtain a culture supernatant. IL-12 in the culture supernatant was measured using Mouse Total IL-12 ELISA kit (Thermo scientific).
　その結果を表４及び図１に示す。 The results are shown in Table 4 and FIG.
Figure JPOXMLDOC01-appb-T000004
Figure JPOXMLDOC01-appb-T000004
　表４及び図１からも明らかなように、本発明の乳酸菌（ラクトコッカス・ラクティスT21株）のIL-12産生量は6056 pg/mlであり、他の自社保有のラクトコッカス・ラクティスと比べて高いIL-12産生誘導能力を有していることが確認された。 As is clear from Table 4 and FIG. 1, the amount of IL-12 produced by the lactic acid bacterium of the present invention (Lactococcus lactis T21 strain) is 6056 pg / ml, compared with other in-house owned Lactococcus lactis. It was confirmed to have a high ability to induce IL-12 production.
＜試験例２＞スキムミルク培地増殖性試験
　本発明のラクトコッカス・ラクティスT21株と、自社保有の3つのラクトコッカス・ラクティス比較菌株（LL1、LL2、LL3）について、スキムミルク培地増殖性試験を実施した。 <Test Example 2> Skim Milk Medium Proliferation Test A skim milk medium proliferation test was performed on the Lactococcus lactis T21 strain of the present invention and three Lactococcus lactis comparative strains (LL1, LL2, LL3) owned by the company.
　10％SM（スキムミルク）培地に植菌し、これを30℃ ・48時間培養し、乳酸菌数、pHによってスキムミルク培地での増殖性を評価した。 10% SM (skimmed milk) medium was inoculated and cultured at 30 ° C. for 48 hours, and the growth on the skimmed milk medium was evaluated by the number of lactic acid bacteria and pH.
　結果を表５及び図２に示す。 The results are shown in Table 5 and FIG.
Figure JPOXMLDOC01-appb-T000005
Figure JPOXMLDOC01-appb-T000005
　本発明の菌株（ラクトコッカス・ラクティスT21株）の菌数は7.6×108cfu/mlであり、はっ酵乳、乳酸菌飲料を生産する上で問題のないレベルであった。 The number of strains of the strain of the present invention (Lactococcus lactis T21 strain) was 7.6 × 10 8 cfu / ml, which was a level with no problem in producing fermented milk and lactic acid bacteria beverages.

Claims (4)

  1. 　クランベリー由来の免疫調節作用を有するラクトコッカス属菌。 Lactococcus spp. That have an immunomodulatory effect derived from cranberries.
  2. 　ラクトコッカス属菌がラクトコッカス・ラクティスである、請求項１に記載の菌。 The bacterium according to claim 1, wherein the genus Lactococcus is Lactococcus lactis.
  3. 　前記ラクトコッカス・ラクティスがラクトコッカス・ラクティスT21株（NITE BP-02164）である、請求項２に記載の菌。 The bacterium according to claim 2, wherein the Lactococcus lactis is Lactococcus lactis T21 strain (NITE BP-02164).
  4. 　請求項３に記載のラクトコッカス・ラクティスT21株（NITE BP-02164）を含有する飲食品。 A food or drink containing the Lactococcus lactis T21 strain (NITE BP-02164) according to claim 3.
PCT/JP2016/084894 2015-11-26 2016-11-25 Novel lactobacillales WO2017090704A1 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
JP2015230538A JP6345642B2 (en) 2015-11-26 2015-11-26 New lactic acid bacteria
JP2015-230538 2015-11-26

Publications (1)

Publication Number Publication Date
WO2017090704A1 true WO2017090704A1 (en) 2017-06-01

Family

ID=58763519

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/JP2016/084894 WO2017090704A1 (en) 2015-11-26 2016-11-25 Novel lactobacillales

Country Status (2)

Country Link
JP (1) JP6345642B2 (en)
WO (1) WO2017090704A1 (en)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20220411836A1 (en) * 2021-06-22 2022-12-29 SAIB & CO Inc. Methods for extracting anthocyanins to improve urinary health by using cranberries and plant-based lactobacillus which enhance female reproductive health and extracts of thereby

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH10139674A (en) * 1996-11-11 1998-05-26 Yakult Honsha Co Ltd Production promoter of interleukin 12
JP2009256312A (en) * 2008-03-26 2009-11-05 Fujicco Co Ltd Immunomodulating composition and food and drink or material for food and drink using the composition

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH10139674A (en) * 1996-11-11 1998-05-26 Yakult Honsha Co Ltd Production promoter of interleukin 12
JP2009256312A (en) * 2008-03-26 2009-11-05 Fujicco Co Ltd Immunomodulating composition and food and drink or material for food and drink using the composition

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
KIMOTO, H. ET AL.: "New Lactococcus Strain with Immunomodulatory Activity: Enhancement of Th1- Type Immune Response", MICROBIOL. IMMUNOL., vol. 48, no. 2, 2004, pages 75 - 82, XP002555796, ISSN: 1348-0421 *
YOSUKE SUNADA ET AL.: "Lactococcus lactis T21 Kabu no Kafunsho ni Taisuru Koka", PROCEEDINGS OF THE 2016 ANNUAL MEETING OF JAPAN SOCIETY FOR BIOSCIENCE, BIOTECHNOLOGY, AND AGROCHEMISTRY, 5 March 2016 (2016-03-05), ISSN: 2186-7976 *
YOSUKE SUNADA ET AL.: "Men'eki Chosetsu Sayo o Yusuru Nyusankin no Screening", ASSOCIATE JOURNAL OF JAPANESE SOCIETY FOR MEDICAL USE OF FUNCTIONAL FOODS, vol. 9, no. 4, 1 December 2015 (2015-12-01), pages 262, ISSN: 1348-2564 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20220411836A1 (en) * 2021-06-22 2022-12-29 SAIB & CO Inc. Methods for extracting anthocyanins to improve urinary health by using cranberries and plant-based lactobacillus which enhance female reproductive health and extracts of thereby

Also Published As

Publication number Publication date
JP6345642B2 (en) 2018-06-20
JP2017093380A (en) 2017-06-01

Similar Documents

Publication Publication Date Title
CA2793729C (en) Novel lactobacillus classified as lactobacillus plantarum, and use thereof
CA2721704C (en) Novel strain of lactobacillus paracasei subspecies paracasei having antimicrobial and immunomodulatory properties
US20220064589A1 (en) Novel lactobacillus plantarum strain atg-k2, atg-k6 or atg-k8, and composition for preventing or treating vaginitis comprising same
KR102028744B1 (en) Lactobacillus plantarum HY7717 strain having immune-enhancing activity, antioxidative activity and digestive fluid resistance and use thereof
KR101239806B1 (en) The new Lactobacillus plantarum HY7712 stimulate immunity
KR101511976B1 (en) The new Lactobacillus fermentum HY7301 producing poly saccharides having immune stimulating activity and products containing thereof as effective component
JP4891255B2 (en) Lactic acid bacteria with immunomodulatory action derived from wine fermentation moromi
KR101274467B1 (en) Lactobacillus plantarum JBMI F5 strain having antibacterial activity against harmful bacteria and uses thereof
WO2018012164A1 (en) Lactic acid bacteria having capacity to promote hyaluronic acid production
JP5425337B1 (en) New lactic acid bacteria
JP6345642B2 (en) New lactic acid bacteria
Ricci et al. Cocoa powder as delivery medium for probiotic Lactobacillus strains
KR101969906B1 (en) Composition for maintaing immunological homeostasis by settling immunological imbalance due to fine dust containing Lactobacillus casei HY2782 as effective component
KR20200145941A (en) A food composition for prevention or improvement of diseases related to tight junction protein comprising Lactobacillus casei HY2782 as an effective compotent
JP2020022393A (en) Medium for lactic acid bacteria
WO2021024915A1 (en) Bifidobacterium having low activity of inducing generation of inflammatory cytokines and high activity of inducing generation of anti-inflammatory cytokines
JP2021164435A (en) Bifidobacteria proliferation promotor
JP2018113892A (en) Bifidobacteria with ability to induce production of anti-inflammatory cytokines
JP2020061978A (en) Novel lactic acid bacteria belonging to enterococcus faecium and use thereof
JP5980785B2 (en) New bifidobacteria and their use
KR102587892B1 (en) Lactobacillus Paracasei HY7017 With Enhanced Functional Characteristics and Enhanced Immunity Function by Using Red Ginseng as a Nutrient Source, and Use Thereof
JP6836298B1 (en) Composition containing lactic acid bacteria and natto bacteria
JP2009254236A (en) New lactic acid bacterium and food, drink and immunostimulator using new lactic acid bacterium
JP2022117889A (en) Lactic acid bacteria having oxidative stress resistance imparting ability (il-8 inhibiting ability) and small intestinal stem cell gene regulating function
Walhe Rajan et al. Vitamin B12 production and cholesterol reduction activity of Pediococcus pentosaceus isolates from fermented Indian foods

Legal Events

Date Code Title Description
DPE2 Request for preliminary examination filed before expiration of 19th month from priority date (pct application filed from 20040101)
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 16868647

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 16868647

Country of ref document: EP

Kind code of ref document: A1