KR102034371B1 - Novel Strain of Lactobacillus brevis BNT 11 Having Antimicrobial Effect Against Pathogenic Microorganisms, and Uses thereof - Google Patents
Novel Strain of Lactobacillus brevis BNT 11 Having Antimicrobial Effect Against Pathogenic Microorganisms, and Uses thereof Download PDFInfo
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- KR102034371B1 KR102034371B1 KR1020180016806A KR20180016806A KR102034371B1 KR 102034371 B1 KR102034371 B1 KR 102034371B1 KR 1020180016806 A KR1020180016806 A KR 1020180016806A KR 20180016806 A KR20180016806 A KR 20180016806A KR 102034371 B1 KR102034371 B1 KR 102034371B1
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- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
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- A—HUMAN NECESSITIES
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- A—HUMAN NECESSITIES
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Abstract
본 발명은 김치에서 분리한 신균주 락토바실러스 브레비스 BNT 11과, 상기 균주 또는 그 배양액을 유효성분으로 포함하는 질염 치료 또는 예방용 조성물에 관한 것이다.
본 발명의 락토바실러스 브레비스 BNT 11 균주 또는 그 배양액을 포함하는 조성물은 유해세균에 의해 발생하는 다양한 질내 세균 감염증에 적용이 가능하다. 또한, 본 발명의 조성물은 건강기능식품 조성물, 세정제품 등의 다양한 분야에서 항진균 및 항염증 목적으로 유용하게 이용가능하다.The present invention relates to a composition for treating or preventing vaginitis comprising the new strain Lactobacillus brevis BNT 11 isolated from kimchi and the strain or its culture as an active ingredient.
The composition comprising the Lactobacillus brevis BNT 11 strain or a culture thereof of the present invention can be applied to various vaginal bacterial infections caused by harmful bacteria. In addition, the composition of the present invention can be usefully used for antifungal and anti-inflammatory purposes in various fields, such as health food composition, cleaning products.
Description
본 발명은 항균 활성을 갖는 락토바실러스 브레비스 균주에 관한 것으로, 더욱 구체적으로 질내 병원성 미생물에 대한 증식 억제활성을 갖는 신균주 락토바실러스 브레비스 BNT 11 및 이의 용도에 관한 것이다.The present invention relates to a Lactobacillus brevis strain having an antibacterial activity, and more particularly to a new strain Lactobacillus brevis BNT 11 and its use having a proliferation inhibitory activity against intravaginal pathogenic microorganisms.
생물체의 항상성(homeostasis)을 유지하기 위한 과정에서 중요한 역할을 담당하고 있는 물질들 중 일부가 각종 생물체 유래의 생리 활성 물질이다. 지금까지 수많은 생리 활성 물질에 대해 많은 연구가 진행되고 있으며, 그 중, 미생물, 식물, 또는 동물체 등에서 분리된 항균물질에 대한 연구는 생명과학 및 의학 분야에서 매우 중요한 영역이다.Some of the substances that play an important role in the process of maintaining homeostasis are biologically active substances from various organisms. Many studies have been conducted on a number of physiologically active substances, and among them, research on antimicrobial substances isolated from microorganisms, plants, or animals is a very important area in the life sciences and medicine fields.
모든 생명체는 생존을 위하여 항균 및 항진균 물질을 생산하는 것으로 알려져 있는데, 항균 및 항진균 물질을 생산하는 미생물에 대해서 많은 연구가 이루어지고 있으며, 항균 및 항진균 물질을 생산하는 미생물 또는 이들에게서 분리된 물질을 이용해 항균제 또는 항진균제를 개발하려는 연구들이 오래전부터 시도되고 있다. All living organisms are known to produce antimicrobial and antifungal substances for survival. Much research has been conducted on microorganisms that produce antimicrobial and antifungal substances. Research into the development of antibacterial or antifungal agents has long been attempted.
항균물질은 미생물을 죽이면서 인체 또는 동물에게는 독성이 낮고 체내의 효소 등에 의해 비활성화되지 않는 선택적 독성작용(selective toxicity)을 갖는 물질로서, 이는 주로 DNA의 복제, 유전정보의 전사 및 해독, 전자에너지의 수송, 세포벽의 생합성 등을 저해함으로써 미생물의 증식을 억제하는 기전을 통해 효과를 나타낸다.Antimicrobial substances have a selective toxicity that kills microorganisms and are not toxic to humans or animals and are not inactivated by enzymes in the body. These include mainly DNA replication, transcription and translation of genetic information, and electronic energy. It inhibits transport, biosynthesis of cell walls, and the like, through the mechanism of inhibiting the growth of microorganisms.
현재까지 개발된 주된 항균물질들은 미생물, 식물 등 천연에서 유래한 것과 화학적으로 합성된 것으로 나뉠 수 있는데, 천연 유래 또는 화학적으로 합성된 많은 항균물질이 유해균에 의해 유발된 질병의 치료에 사용되고 있으나, 최근에는 화학적으로 합성된 항균물질에 대한 내성 증가로 인해 천연에서 유래한 항균물질을 이용한 항균제 개발에 관심이 증가하고 있는 추세이고, 최근 삶의 질이 향상됨에 따라, 여성용 질염의 치료 또는 예방에 효과가 있는 천연유래 물질을 탐색하고자 하는 연구가 활발하게 진행되고 있다. The main antimicrobial agents developed to date can be divided into microorganisms, plants, etc., and chemically synthesized. Many antimicrobial substances, both natural and chemically synthesized, are used for the treatment of diseases caused by harmful bacteria. There is a growing interest in the development of antimicrobial agents using natural antimicrobial substances due to increased resistance to chemically synthesized antimicrobial substances. Recently, as the quality of life is improved, it is effective in treating or preventing women's vaginitis. There is an active research to search for natural derivatives.
이와 관련하여 천연 식물추출물을 함유하는 생리대 조성물과 관련된 기술(특허출원 제10-2006-0012029호)과 한방에서 사용되던 식물 추출물을 이용한 생리대 조성물과 관련된 기술(특허출원 제10-2005-0074562호, 특허출원 제10-2010-0066270호)들이 개발된바 있고, 유용 미생물을 이용하여 질염 병원균을 억제하고자 하는 관련 연구(특허공개 제10-2018-0006998호, 특허공개 제10-2017-0109446호, 특허등록 제10-0712286호)들이 진행된 바 있었으나, 이들 선행기술들은 질염의 원인균인 협기성 세균, 호기성 세균 및 진균류에 대한 항균 활성이 낮아, 질 세정제 제품에 산업적으로 활용되기 어려운 단점이 있었다. In this regard, a technique related to a sanitary napkin composition containing a natural plant extract (Patent Application No. 10-2006-0012029) and a technique related to a sanitary napkin composition using a plant extract used in oriental medicine (Patent Application No. 10-2005-0074562, Patent Application No. 10-2010-0066270 has been developed, and related studies that seek to inhibit vaginitis pathogens using useful microorganisms (Patent Publication No. 10-2018-0006998, Patent Publication No. 10-2017-0109446, Patent registration No. 10-0712286), but the prior art has a low antibacterial activity against aerobic bacteria, aerobic bacteria and fungi that are causative agents of vaginitis, there is a disadvantage that it is difficult to be industrially used in vaginal cleaning products.
이에, 본 발명자들은 다양한 질염의 원인이 되는 유해 세균들과 진균을 효과적으로 억제하는 천연 미생물들을 스크리닝하고 그 효과를 검증하기 위해 예의 연구노력한 결과, 신균주 락토바실러스 브레비스 BNT 11이 질염의 원인균인 세균 및 진균을 효과적으로 억제하여 질염 치료 또는 예방용 조성물이나 질 세정제 제품으로 유용하게 사용할 수 있음을 확인하고, 본 발명을 완성하게 되었다.Accordingly, the present inventors have conducted extensive research to screen the harmful microorganisms causing various vaginitis and natural microorganisms effectively inhibiting fungi and to verify the effects of the bacterium, and the bacterium Lactobacillus brevis BNT 11 is the causative agent of vaginitis. It has been confirmed that the present invention can be effectively used as a composition for vaginitis treatment or prevention or as a vaginal detergent product by effectively inhibiting fungi, thereby completing the present invention.
따라서, 본 발명의 주된 목적은 질염의 원인균인 유해 세균과 진균을 효과적으로 억제할 수 있는 장점을 갖는 질염 치료 또는 예방용 조성물을 제공하는 데 있다.Therefore, the main object of the present invention is to provide a composition for treating or preventing vaginitis, which has an advantage of effectively inhibiting harmful bacteria and fungi which are causative agents of vaginitis.
본 발명의 다른 목적은 상기 질 세정제 조성물을 유효성분으로 함유하여 질염의 치료 및 예방에 효과가 있는 질염 치료 또는 예방용 조성물을 제공하는데 있다.Another object of the present invention is to provide a vaginitis treatment or prevention composition containing the vaginal detergent composition as an active ingredient effective in the treatment and prevention of vaginitis.
본 발명의 한 양태에 따르면, 본 발명은 항균 활성을 갖는 신균주 락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)(수탁번호: KCCM12167P)을 제공한다.According to one aspect of the present invention, the present invention provides Lactobacillus brevis BNT 11 (Accession No .: KCCM12167P) having antibacterial activity.
본 발명의 용어 "락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)"은 본 발명자들에 의해 김치에서 새롭게 분리 동정된 신규한 락토바실러스 브레비스 균주로서, "BNT 11 균주" 또는 "BNT 11"과 혼용하여 사용할 수 있다. The term " Lactobacillus brevis BNT 11" of the present invention is a novel Lactobacillus brevis strain newly identified and identified in kimchi by the present inventors, and used in combination with "BNT 11 strain" or "BNT 11". Can be used.
본 발명자들은 신규 항균제의 개발을 위해 발효식품에서 분리된 유용 미생물의 항균 효과에 대해 수년간 조사 분석하였고, 그 결과 김치에서 유래한 락토바실러스 브레비스 균주가 우수한 항균 효과가 있음을 검증한 바 있고, 질염의 원인균인 세균 및 칸디다균의 생장 억제에 사용한 결과, 그 효과가 획기적임을 발견하였고, 또한 인체에 적용시 안전성에도 문제가 없음을 확인하였다The inventors have investigated and analyzed the antimicrobial effect of useful microorganisms isolated from fermented foods for the development of new antimicrobial agents for many years, and as a result, the Lactobacillus brevis strain derived from kimchi has an excellent antimicrobial effect. As a result of use in inhibiting the growth of the causative bacteria and Candida, it was found that the effect was remarkable, and it was confirmed that there is no problem in safety when applied to the human body.
본 발명에서, 상기 락토바실러스 브레비스 BNT 11 균주는 서열번호 1의 16s rDNA 서열을 갖는 것을 특징으로 한다.In the present invention, the Lactobacillus brevis BNT 11 strain is characterized by having a 16s rDNA sequence of SEQ ID NO: 1.
본 발명자들은 상기 유전자 서열을 종래 공지된 균주들의 유전자 서열들과 비교 분석한 결과, 본 발명의 균주가 락토바실러스 브레비스에 속하는 새로운 균주임을 확인하여 락토바실러스 브레비스 BNT 11로 명명하였고, 한국미생물보존센터에 기탁하고, 수탁번호 KCCM 12167P를 부여받았다.The present inventors compared the gene sequences with those of conventionally known strains, and as a result, the strain of the present invention was identified as a new strain belonging to Lactobacillus brevis, and was named as Lactobacillus brevis BNT 11, and the Korea Microbial Conservation Center It was deposited and was given accession number KCCM 12167P.
본 발명의 상기 락토바실러스 브레비스 BNT 11 균주는 질염을 유발하는 다양한 유해균에 대해 항균 활성이 있다. 본 발명의 상기 락토바실러스 브레비스 BNT 11 균주는 그람 양성균, 그람 음성균 및 항생제 내성 균주에 대하여 항균 활성을 갖고 있는데, 구체적으로 질의 입구에서 염증을 유발하는 호기성 세균인 에스체리치아 콜라이(Escherichia coli)와, 질의 내부에서 염증을 유발하는 혐기성 세균인 슈도모나스 에루기노사(Pseudomonas aeruginosa) 및 스타필로코커스 아우레우스(Staphylococcus aureus) 등의 세균에 대하여 항균 활성을 나타낸다. 이외에도 살모넬라 파라티피(Salmonella paratyphi), 살모넬라 타이피뮤리움(Salmonella typhimurim) 등의 병원성 세균에도 항균 활성을 보인다.The Lactobacillus brevis BNT 11 strain of the present invention has antimicrobial activity against various harmful bacteria causing vaginitis. The Lactobacillus brevis BNT 11 strain of the present invention has antimicrobial activity against Gram-positive bacteria, Gram-negative bacteria and antibiotic-resistant strains, specifically Escherichia coli , which is an aerobic bacterium that causes inflammation at the entrance of the vagina, It shows antibacterial activity against bacteria such as Pseudomonas aeruginosa and Staphylococcus aureus , which are inflammation-causing anaerobes within the vagina. In addition, it shows antimicrobial activity against pathogenic bacteria such as Salmonella paratyphi and Salmonella typhimurim .
또한, 본 발명의 상기 락토바실러스 브레비스 BNT 11 균주는 질내에 서식하는 다양한 진균류에 대해 항진균 활성이 있고, 특히, 질염 유발에 가장 큰 원인으로 지목되고 있는 칸디다 속 진균에 대해 항진균 활성을 나타내며, 보다 구체적으로는 칸디다 알비칸스(Candida albicans)에 대해 항진균 활성을 갖는다. In addition, the Lactobacillus brevis BNT 11 strain of the present invention has antifungal activity against various fungi inhabiting the vagina, and particularly, shows antifungal activity against fungi of the genus Candida, which is considered to be the biggest cause of vaginitis. It has antifungal activity against Candida albicans .
본 발명의 다른 양태에 따르면, 본 발명은 항균 활성을 갖는 신균주 락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)(수탁번호: KCCM12167P), 또는 그 배양액을 유효성분으로 함유하는 질염 치료 또는 예방용 약학적 조성물을 제공한다.According to another aspect of the present invention, the present invention is a new strain Lactobacillus brevis BNT 11 (Accession No .: KCCM12167P) having an antimicrobial activity, or a pharmaceutical preparation for treating or preventing vaginitis containing the culture medium as an active ingredient. To provide a composition.
본 발명의 질염 치료 또는 예방용 약학적 조성물에서, 상기 균주는 서열번호 1의 16s rDNA 서열을 갖는 것을 특징으로 한다.In the pharmaceutical composition for treating or preventing vaginitis of the present invention, the strain is characterized by having a 16s rDNA sequence of SEQ ID NO: 1.
본 발명의 질염 치료 또는 예방용 약학적 조성물에서, 상기 균주는 슈도모나스 에루기노사(Pseudomonas aeruginosa), 에스체리치아 콜라이(Escherichia coli), 살모넬라 파라티피(Salmonella paratyphi), 살모넬라 타이피뮤리움(Salmonella typhimurim), 스타필로코코스 아우레우스(Staphylococcus aureus) 및 칸디다 알비칸스(Candida albicans)에서 선택된 하나 이상의 병원성 균주에 대해 항균 활성을 갖는 것을 특징으로 한다.In the salt treatment or prevention a pharmaceutical composition of the present invention, the strain is rugi labor (Pseudomonas aeruginosa), S. Cherry Escherichia coli (Escherichia coli), Salmonella para typhimurium (Salmonella paratyphi), S. typhimurium (Salmonella typhimurim) Pseudomonas , Staphylococcus aureus ( Staphylococcus aureus ) and Candida albicans ( Candida albicans ) is characterized by having an antimicrobial activity against one or more pathogenic strains.
본 발명에 따른 약학적 조성물은 인간을 포함하는 포유동물에 다양한 경로로 투여될 수 있다. 투여 방식은 통상적으로 사용되는 모든 방식일 수 있으며, 예컨대, 경구, 피부, 정맥, 근육, 피하 등의 경로로 투여될 수 있으며, 바람직하게는 경구로 투여될 수 있다. The pharmaceutical compositions according to the invention can be administered to a mammal, including humans, by various routes. The mode of administration may be any of the methods commonly used, and may be administered by, for example, oral, skin, vein, muscle, subcutaneous, and the like, and preferably, orally.
본 발명의 약학적 조성물은 각각 통상의 방법에 따라 산제, 과립제, 정제, 캡슐제, 연고제, 현탁액, 에멀젼, 시럽, 에어로졸 등의 경구형 제형, 또는 경피제, 좌제 및 멸균 주사용액의 형태의 비경구 제형 등으로 제형화하여 사용될 수 있다. Pharmaceutical compositions of the present invention may be prepared in the form of powders, granules, tablets, capsules, ointments, suspensions, emulsions, syrups, aerosols, or the like, or in the form of transdermal, suppository, and sterile injectable solutions, respectively, according to conventional methods. It may be used in the form of a formulation.
본 발명의 약학적 조성물은 약제학적으로 적합하고 생리학적으로 허용되는 담체, 부형제 및 희석제 등의 보조제를 추가로 함유하는 것일 수 있다. The pharmaceutical composition of the present invention may further contain an adjuvant such as a pharmaceutically suitable and physiologically acceptable carrier, excipient and diluent.
본 발명의 약학적 조성물에 포함될 수 있는 담체, 부형제 및 희석제로는 락토즈, 덱스트로즈, 수크로스, 솔비톨, 만니톨, 자일리톨, 에리스리톨, 말티톨, 전분, 아카시아 고무, 알지네이트, 젤라틴, 칼슘 포스페이트, 칼슘 실리케이트, 셀룰로즈, 메틸 셀룰로즈, 미정질 셀룰로스, 폴리비닐 피롤리돈, 물, 메틸히드록시벤조에이트, 프로필히드록시벤조에이트, 탈크, 마그네슘 스테아레이트 및 광물유를 들 수 있다. 제제화할 경우에는 보통 사용하는 충진제, 증량제, 결합제, 습윤제, 붕해제, 계면활성제 등의 희석제 또는 부형제를 사용할 수 있다Carriers, excipients and diluents which may be included in the pharmaceutical compositions of the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium Silicates, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. When formulated, diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents and surfactants which are commonly used may be used.
본 발명에 따른 질염의 치료 또는 예방용 약학적 조성물을 비경구용으로 제공하는 경우, 일예로, 액제, 겔(gel)제, 세정 조성물, 질 내 삽입용 정제, 좌제 형태, 크림, 연고, 드레싱 용액, 분무제, 기타 도포제등의 국소 투여제, 용액형, 현탁형, 유제형 등의 액상 제형일 수 있으며, 멸균된 수용액, 비수성용제, 현탁제, 유제, 동결건조제제, 좌제, 크림, 연고, 젤리, 거품, 세척제 또는 질 삽입물, 바람직하게는 액제, 겔(gel)제, 세정 조성물, 질 내 삽입용 정제 등의 피부 외용제가 포함될 수 있다. 상기 제형은 일예로 멸균수에 용해보조제, 유화제, pH 조절을 위한 완충제 등을 첨가하여 제조할 수 있다.When providing a pharmaceutical composition for treating or preventing vaginitis according to the present invention for parenteral use, for example, liquids, gels, cleaning compositions, intravaginal tablets, suppository forms, creams, ointments, dressing solutions It may be a liquid dosage form such as topical administration, spray, or other coating agent, solution type, suspension type, emulsion type, etc., sterile aqueous solution, non-aqueous solvent, suspension, emulsion, lyophilized agent, suppository, cream, ointment, jelly External skin preparations, such as foams, detergents or vaginal inserts, preferably liquids, gels, cleaning compositions, intravaginal tablets, and the like. For example, the formulation may be prepared by adding a dissolving aid, an emulsifier, a buffer for pH adjustment, and the like in sterile water.
상기 비수성용제 또는 현탁용제로는 프로필렌글리콜(propylene glycol), 폴리에틸렌글리콜(polyethylene glycol), 올리브 오일과 같은 식물성 기름, 에틸올레이트와 같은 주사 가능한 에스테르 등이 사용될 수 있다.As the non-aqueous solvent or suspending solvent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like may be used.
본 발명의 약학적 조성물을 인간에게 적용하는 구체예에 있어서, 본 발명의 약학적 조성물은 단독으로 투여될 수 있으나, 일반적으로 투여방식과 표준 약제학적 관행(standard pharmaceutical practice)을 고려하여 선택된 약제학적 담체와 혼합되어 투여될 수 있다. In an embodiment in which the pharmaceutical composition of the present invention is applied to a human, the pharmaceutical composition of the present invention may be administered alone, but is generally selected in consideration of the mode of administration and standard pharmaceutical practice. It can be administered in admixture with a carrier.
예를 들면, 본 발명의 락토바실러스 속 균주 함유 조성물은 전분 또는 락토오즈를 함유하는 정제 형태로, 또는 단독 또는 부형제를 함유하는 캡슐 형태로, 또는 맛을 내거나 색을 띄게 하는 화학 약품을 함유하는 엘릭시르 또는 현탁제 형태로 경구, 구강 내 또는 혀 밑 투여될 수 있다. For example, the Lactobacillus sp. Strain-containing compositions of the present invention may be in the form of tablets containing starch or lactose, in the form of capsules alone or in excipients, or elixirs containing chemicals that flavor or color. Or in suspension, orally, orally or sublingually.
본 발명의 신균주 락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)(수탁번호: KCCM12167P), 또는 그 배양액을 함유하는 약학적 조성물의 투여 용량은 환자의 나이, 몸무게, 성별, 투여형태, 건강상태 및 질환 정도에 따라 달라질 수 있으며, 의사 또는 약사의 판단에 따라 일정 시간간격으로 1일 1회 내지 수회로 분할 투여할 수도 있다. 예컨대, 유효성분 함량을 기준으로 1일 투여량이 0.1 내지 500㎎/kg, 바람직하게는 0.5 내지 300㎎/kg일 수 있다. 상기한 투여량은 평균적인 경우를 예시한 것으로서 개인적인 차이에 따라 그 투여량이 높거나 낮을 수 있다. 본 발명의 신균주 락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)(수탁번호: KCCM12167P), 또는 그 배양액을 함유하는 조성물의 1일 투여량이 상기 투여 용량 미만이면 유의성 있는 효과를 얻을 수 없으며, 그 이상을 초과하는 경우 비경제적일 뿐만 아니라 상용량의 범위를 벗어나므로 바람직하지 않은 부작용이 나타날 우려가 발생할 수 있으므로, 상기 범위로 하는 것이 좋다.The dosage of the present invention strain Lactobacillus brevis BNT 11 (Accession No .: KCCM12167P), or a pharmaceutical composition containing the culture medium, is determined according to the age, weight, sex, dosage form, health condition and It may vary depending on the degree of the disease, and may be divided once or several times a day at regular time intervals according to the judgment of a doctor or a pharmacist. For example, the daily dosage may be 0.1 to 500 mg / kg, preferably 0.5 to 300 mg / kg, based on the active ingredient content. The above dosages are illustrative of the average case and may be high or low depending on individual differences. Lactobacillus brevis BNT 11 (Accession No .: KCCM12167P) of the present invention, or if the daily dose of the composition containing the culture medium is less than the dose, a significant effect can not be obtained, or more If it exceeds, not only is uneconomical, it is out of the range of the normal dose, so it may cause undesirable side effects.
본 발명의 다른 양태에 따르면, 본 발명은 신균주 락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)(수탁번호: KCCM12167P), 또는 그 배양액을 유효성분으로 함유하는 질염 치료 또는 예방용 건강기능식품을 제공한다.According to another aspect of the present invention, the present invention provides a new functional strain Lactobacillus brevis BNT 11 (Accession No .: KCCM12167P), or a functional food for treating or preventing vaginitis containing the culture medium as an active ingredient. do.
상기 건강 기능성 식품은 각종 음료, 발효유, 식품 첨가제 등일 수 있다. The health functional food may be various beverages, fermented milk, food additives, and the like.
상기 락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)(수탁번호: KCCM12167P) 균주는 상술한 바와 같다. The Lactobacillus brevis BNT 11 (Accession No .: KCCM12167P) strain is as described above.
상기 건강 기능성 식품에 함유된 유효성분으로서의 락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)(수탁번호: KCCM12167P) 균주의 함량은 식품의 형태, 소망하는 용도 등에 따라 적절하게 특별한 제한이 없으며, 예컨대, 전체 식품 중량의 0.01 내지 15 중량%로 가할 수 있으며, 건강 음료 조성물은 100㎖를 기준으로 0.02 내지 10 g, 바람직하게는 0.3 내지 1g의 비율로 가할 수 있다.The content of the strain Lactobacillus brevis BNT 11 (Accession No .: KCCM12167P) as an active ingredient contained in the health functional food is not particularly limited according to the form of the food, desired use, etc. It can be added at 0.01 to 15% by weight of the food weight, the health beverage composition may be added in a ratio of 0.02 to 10 g, preferably 0.3 to 1g based on 100ml.
본 발명의 건강기능식품 중 음료에는 지시된 비율로 필수 성분으로서 상기 락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)(수탁번호: KCCM12167P) 균주를 함유하는 것 외에 액체성분에는 특별한 제한은 없으며, 통상의 음료와 같이 여러 가지 향미제 또는 천연 탄수화물 등을 추가 성분으로서 함유할 수 있다. In the beverage of the health functional food of the present invention, in addition to containing the strain Lactobacillus brevis BNT 11 (Accession No .: KCCM12167P) as an essential ingredient in the indicated ratio, there is no particular limitation on the liquid component, Various flavors or natural carbohydrates, such as beverages, may be included as additional ingredients.
상술한 천연 탄수화물의 예는 모노사카라이드, 예를 들어, 포도당, 과당 등의 디사카라이드, 예를 들어 말토스, 슈크로스 등의 및 폴리사카라이드, 예를 들어 덱스트린, 시클로덱스트린 등과 같은 통상적인 당 및 자일리톨, 소르비톨, 에리트리톨 등의 당알콜이다. 상술한 것 이외의 향미제로서 천연 향미제(타우마틴, 스테비아 추출물(예를 들어 레바우디오시드 A, 글리시르히진등) 및 합성 향미제(사카린, 아스파르탐 등)를 유리하게 사용할 수 있다. 상기 천연 탄수화물의 비율은 본 발명의 조성물 100 ㎖당 일반적으로 약 1 내지 20g, 바람직하게는 약 5 내지 12g이다. Examples of the above-mentioned natural carbohydrates are conventional monosaccharides such as disaccharides such as glucose and fructose, such as maltose, sucrose and the like, and polysaccharides such as dextrin, cyclodextrin and the like. Sugars and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those mentioned above, natural flavoring agents (tauumatin, stevia extract (for example, rebaudioside A, glycyrrhizin, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used. The proportion of natural carbohydrates is generally about 1-20 g, preferably about 5-12 g per 100 ml of the composition of the present invention.
상기 외에 본 발명의 건강기능식품은 여러 가지 영양제, 비타민, 광물(전해질), 합성 풍미제 및 천연 풍미제 등의 풍미제, 착색제 및 증진제(치즈, 초콜릿 등), 펙트산 및 그의 염, 알긴산 및 그의 염, 유기산, 보호성 콜로이드 증점제, pH 조절제, 안정화제, 방부제, 글리세린, 알콜, 탄산 음료에 사용되는 탄산화제 등을 함유할 수 있다. In addition to the above, the health functional food of the present invention includes various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, colorants and enhancers (such as cheese and chocolate), pectic acid and salts thereof, alginic acid and Salts, organic acids, protective colloid thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonation agents used in carbonated drinks, and the like.
그밖에 본 발명의 건강기능식품은 천연 과일 쥬스 및 과일 쥬스 음료 및 야채 음료의 제조를 위한 과육을 함유할 수 있다. 이러한 성분은 독립적으로 또는 조합하여 사용할 수 있다. 이러한 첨가제의 비율은 그렇게 중요하진 않지만 본 발명의 건강기능식품 100 중량부 당 0.1 내지 약 20 중량부의 범위에서 선택되는 것이 일반적이다.In addition, the health functional food of the present invention may contain natural fruit juice and fruit flesh for producing fruit juice beverages and vegetable beverages. These components can be used independently or in combination. The proportion of such additives is not so critical but is generally selected in the range of 0.1 to about 20 parts by weight per 100 parts by weight of the health functional food of the present invention.
본 발명의 또 다른 양태에 따르면, 본 발명은 신균주 락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)(수탁번호: KCCM12167P), 또는 그 배양액을 유효성분으로 함유하는 질염 치료 또는 예방용 세정제품을 제공한다. 상기 세정제품은 고체화장비누, 질 세정제, 액체샴푸, 액체비누, 바디 세정제, 크림상 비누 등 일 수 있다. 상기 신균주 락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)(수탁번호: KCCM12167P) 균주는 상술한 바와 같다. 상기 세정제품은 이를 제형화하기 위한 담체를 추가로 포함할 수 있다. 상기 담체는 결합제, 활탁제, 현탁용제, 가용화제, 완충제, 보존제, 윤활제, 등장제, 부형제, 안정화제, 분산제, 현탁화제, 색소, 향료 등을 사용할 수 있다.According to another aspect of the present invention, the present invention provides a cleaning product for treating or preventing vaginitis, which contains the new strain Lactobacillus brevis BNT 11 (Accession Number: KCCM12167P), or a culture solution thereof as an active ingredient. do. The cleaning product may be a solidifying equipment soap, vaginal cleaner, liquid shampoo, liquid soap, body cleaner, creamy soap, or the like. The strain Lactobacillus brevis BNT 11 (Lactobacillus brevis BNT 11) (Accession No .: KCCM12167P) strain is as described above. The cleaning product may further comprise a carrier for formulating it. The carrier may be a binder, a suspending agent, a suspending agent, a solubilizer, a buffer, a preservative, a lubricant, an isotonic agent, an excipient, a stabilizer, a dispersant, a suspending agent, a coloring agent, a fragrance, and the like.
본 발명의 특징 및 이점을 요약하면 다음과 같다:The features and advantages of the present invention are summarized as follows:
(ⅰ) 본 발명은 김치에서 분리한 신균주 락토바실러스 브레비스 BNT 11과, 상기 균주 또는 그 배양액을 유효성분으로 포함하는 질염 치료 또는 예방용 조성물을 제공한다.(Iii) The present invention provides a composition for treating or preventing vaginitis comprising the new strain Lactobacillus brevis BNT 11 isolated from kimchi and the strain or its culture as an active ingredient.
(ⅱ) 본 발명의 락토바실러스 브레비스 BNT 11 균주 또는 그 배양액을 포함하는 조성물은 유해세균에 의해 발생하는 다양한 질내 세균 감염증에 적용이 가능하다. (Ii) The composition comprising the Lactobacillus brevis BNT 11 strain of the present invention or a culture thereof can be applied to various vaginal bacterial infections caused by harmful bacteria.
(ⅲ) 또한, 본 발명의 조성물은 건강기능식품 조성물, 세정제품 등의 다양한 분야에서 항진균 및 항염증 목적으로 유용하게 이용가능하다.(Iii) In addition, the composition of the present invention can be usefully used for antifungal and anti-inflammatory purposes in various fields such as health functional food compositions and cleaning products.
도 1은 락토바실러스 브레비스 BNT 11 균주의 동결 건조 분말의 처리가 세포 생존율에 영향을 미치지 않는 것을 보여주는 그래프이다.1 is a graph showing that treatment of lyophilized powder of Lactobacillus brevis BNT 11 strain does not affect cell viability.
이하, 실시예를 통하여 본 발명을 더욱 상세히 설명하기로 한다. 이들 실시예는 단지 본 발명을 예시하기 위한 것이므로, 본 발명의 범위가 이들 실시예에 의해 제한되는 것으로 해석되지는 않는다.Hereinafter, the present invention will be described in more detail with reference to Examples. Since these examples are only for illustrating the present invention, the scope of the present invention is not to be construed as being limited by these examples.
실시예 1. 균주분리 및 항균 활성 테스트Example 1. Strain Separation and Antimicrobial Activity Test
1-1. 균주 분리1-1. Strain isolate
10여일 상온에서 발효 숙성 중인 김치에서 미생물의 분리를 위해 멸균 식염수에 편취한 김치와 국물의 일부를 넣고 호모게나이저로 30초간 작동한 뒤, MRS agar 고체배지(Difco사, USA)에 도말하여 37℃에서 24시간 배양하였다. 노란색 집락을 백금이로 취하여 액상 MRS배지에 배양하기를 10회 반복실험을 거쳐 미생물을 순수 분리 한 후, 이를 20% 글리세린과 함께 -70℃에서 냉동 보관하였다.After fermentation at room temperature for 10 days, put kimchi and broth in sterile saline solution for 30 minutes to separate microorganisms, and operate it with homogenizer for 30 seconds, and then plate on MRS agar solid medium (Difco, USA) 37 Incubated for 24 hours at ℃. Yellow colonies were taken as platinum and cultured in a liquid MRS medium after 10 repeated experiments to separate microorganisms purely, which were then stored frozen at -70 ° C with 20% glycerin.
1-2. 분리된 균주의 항균 및 항진균활성 테스트1-2. Antimicrobial and Antifungal Activity Test of Isolated Strains
분리된 균주들의 배양을 위해 MRS 배지(Difco,USA) 사용하였으며, 분리 균주들의 항균활성을 검증하기 위해, 종래 항균활성이 뛰어난 것으로 검증된 락토바실러스 파라카제이 균주와 류코노스톡 메센토로이데스 균주를 비교 균주로 하여 분리된 미생물들의 항균 활성을 검증하였다. MRS medium (Difco, USA) was used for the cultivation of the isolated strains, and in order to verify the antimicrobial activity of the isolated strains, the Lactobacillus paracasei strains and the L. aureus stock mesentoroides strains, which have been proven to have excellent antimicrobial activity, As a comparative strain, the antimicrobial activity of the isolated microorganisms was verified.
실험은 한천 확산법(Agar disc diffusion assay)을 이용하여 아래와 같이 진행하였다. 먼저, MRS 배지를 멸균하고, 분리된 균주와 비교 균주들을 별도로 37℃에서 18시간 동안 배양하였고, 균주배양액을 원심분리(3,000xg,15분)하여 회수한 상등액을 0.45㎛ 멤브레인 필터로 제균한 배양액을 준비하였다.The experiment was carried out as follows using the Agar disc diffusion assay. First, the MRS medium was sterilized, and the isolated strains and the comparative strains were separately incubated at 37 ° C. for 18 hours, and the supernatant recovered by centrifugation (3,000 × g, 15 minutes) of the strain culture was sterilized with a 0.45 μm membrane filter. Was prepared.
그리고, 각 병원성 공시균주들을 0.5 MacFarland 농도(세균수 약 1×108 CFU/mL)로 배양하고, 이를 2번 정도 희석하여 1×105 CFU/mL 농도로 조정한 후, Nutrient agar(Difco, USA)에 60㎕ 용량으로 도말하여 이를 건조시키고, 멸균된 paper disc(6mm, Whatman, UK)를 올려놓았다.In addition, each pathogenic specimens were incubated at 0.5 MacFarland concentration (about 1 × 10 8 CFU / mL), diluted twice and adjusted to 1 × 10 5 CFU / mL, followed by Nutrient agar (Difco, USA) was plated at 60 μl volume and dried, and a sterile paper disc (6 mm, Whatman, UK) was placed.
상기 준비된 분리 균주들과 비교 균주들의 배양액을 paper disc에 30㎕ 용량으로 점적한 후, 37℃, 5% CO2 조건으로 인큐베이터에서 24시간 배양하여 억제환의 지름을 측정하였다. The culture medium of the prepared isolated strains and the comparative strains was instilled in a 30 μl volume on a paper disc, and then cultured in an incubator at 37 ° C. and 5% CO 2 for 24 hours to determine the diameter of the inhibitory rings.
그 결과, 아래 [표 1]에 나타난 바와 같이, 상기 김치에서 분리되어 BNT 11로 넘버링 된 균주의 배양액을 처리한 실험군은 종래 항균 효과가 있는 것으로 알려져 비교균주로 사용된 락토바실러스 파라카제이 균주와 류코노스톡 메센토로이데스 균주의 배양액과 대비하여 병원성 공시균주 5종 모두에 대하여 우수한 항균 효과가 있음이 관찰되었으며, 이러한 결과를 통해, BNT 11 균주 및 그 배양액이 유해미생물의 성장을 억제하는 효능을 갖는 것을 확인할 수 있었다.As a result, as shown in Table 1 below, the experimental group treated with the culture solution of the strain numbered with BNT 11 separated from the kimchi is known to have a conventional antimicrobial effect and the Lactobacillus paracaze strain used as a comparative strain. It was observed that the antimicrobial effect was excellent against all five pathogenic test strains compared to the culture medium of the Leuconosestock Mesentoroides strain. Through these results, the BNT 11 strain and the culture medium inhibit the growth of harmful microorganisms. It was confirmed to have.
[표 1]TABLE 1
실시예 2. BNT 11 균주의 생리학적 특성 분석 및 동정Example 2. Physiological Characterization and Identification of BNT 11 Strains
2-1. API 키트 분석2-1. API Kit Analysis
상기 실시예 1에서 분리된 BNT 11 균주의 당 이용 특성을 분석하기 위하여 API 50 CHL 키트(Biomerieux, France)를 이용하였다. API 50 CHL kit (Biomerieux, France) was used to analyze the sugar utilization characteristics of the BNT 11 strain isolated in Example 1.
상기 BNT 11 균주를 MRS agar plate에 도말하여 37℃에서 24시간 배양한 후, 매우 높은 탁도를 유지하기 위해 백금이로 콜로니를 수득하여 2ml 멸균 생리식염수에 고농도로 현탁하였고, 이를 5ml 멸균 생리식염수에 n방울 떨어뜨려 Mcfarland 2로 조정하였다. 다시 n방울을 API 50 CHL 배지에 부유시켜 스트립 큐브에 분주하고 미네랄오일을 첨가하여 혐기적 상태로 35℃에서 24시간과 48시간 동안 양성과 음성반응을 검사하였고, 그 결과를 하기 [표 2]에 정리하여 나타내었다. The BNT 11 strain was plated on an MRS agar plate and incubated at 37 ° C. for 24 hours, and colonies were obtained with platinum to maintain a very high turbidity. Drop n drops to adjust to Mcfarland 2. Again n drops were suspended in API 50 CHL medium and dispensed into strip cubes, and mineral oil was added to test for positive and negative reactions at 35 ° C. for 24 and 48 hours in anaerobic conditions. In summary, it is shown.
[표 2]TABLE 2
2-2. 염기서열 분석을 통한 균주 동정2-2. Identification of strains through sequencing
상기 실시예 1에서 분리된 BNT 11 균주의 동정을 위하여 유전자 염기서열 분석을 수행하였다. BNT 11 균주의 16s rRNA에 대한 Sequencing primer로서 785F 5'(CGA TTA GAT ACC CTG GTA) 및 907R5'(CCT TCA ATT CMT TTR AGT TT)를 사용하였다. 분석을 위한 데이터베이스는 NCBI-DB와 SILVA-DB를 사용하였으며, 계통발생학적으로 동정하였다.Gene sequencing was performed to identify the BNT 11 strain isolated in Example 1. As a sequencing primer for 16s rRNA of BNT 11 strain, 785F 5 '(CGA TTA GAT ACC CTG GTA) and 907R5' (CCT TCA ATT CMT TTR AGT TT) were used. For analysis, NCBI-DB and SILVA-DB were used and phylogenetically identified.
그 결과, BNT 11 균주는 공시균주 Lactobacillus brevis(gi:636560178)와 98% 수준의 염기서열 상동성이 있는 것으로 나타났으며, 락토바실러스 브레비스 균주와 가장 높은 상동성을 보였으며, 이들과 단일계통을 이루는 것으로 나타났다As a result, the BNT 11 strain showed 98% sequence homology with the test strain Lactobacillus brevis (gi: 636560178), and showed the highest homology with the Lactobacillus brevis strain. Appeared to achieve
이러한 결과를 바탕으로 BNT 11 균주는 락토바실러스 브레비스(Lactobacillus brevis)에 속하는 균주임을 확인할 수 있었다. 본 발명자들은 우수한 항균 활성을 갖는 본 발명의 균주를 락토바실러스 브레비스 BNT11 (Lactobacillus brevis BNT11)로 명명하고, 한국미생물보존센터에 기탁번호(KCCM 12167P)로 기탁하였다. Based on these results, it was confirmed that the BNT 11 strain belongs to Lactobacillus brevis. The present inventors named the strain of the present invention having excellent antimicrobial activity as Lactobacillus brevis BNT11, and deposited it with the accession number (KCCM 12167P) to the Korea Microorganism Conservation Center.
실시예 4. 섬유아세포의 MTT assay 실험을 통한 안전성 평가 Example 4. Safety evaluation through MTT assay experiment of fibroblasts
실험에 사용한 mouse embryo 유래 3T3-L1세포는 ATCC(American Type Culture Collection, Manassas, VA, USA)에서 분양 받아 사용하였다. 3T3-L1세포는 10% FBS와 100units/ml의 penicillin-streptomycin이 함유된 Dulbeco’s modified Eagle’s medium(DMEM) 배지를 넣고 37℃, 5% CO2 incubator에서 배양하였다. 배양중인 세포를 PBS(phosphate buffered saline) 버퍼로 세척한 뒤 0.05% trypsin-0.02% EDTA(GIBCO, Buffalo, NY, USA)로 부착된 세포를 분리하여 원심분리 한 후, 집적된 세포에 배지를 첨가하여 피펫으로 세포가 골고루 분산되도록 잘 혼합하여 100mm3 세포배양 접시에 10 ml씩 일정한 수로 분할하여 주입하고 일주일에 2회 계대배양 하여 실험에 사용하였다. The mouse embryo-derived 3T3-L1 cells used in the experiments were used by receiving from ATCC (American Type Culture Collection, Manassas, VA, USA). 3T3-L1 cells were incubated in Dulbeco's modified Eagle's medium (DMEM) medium containing 10% FBS and 100 units / ml penicillin-streptomycin in 37 ° C and 5% CO 2 incubator. After culturing the cells in culture with PBS (phosphate buffered saline) buffer, the cells attached with 0.05% trypsin-0.02% EDTA (GIBCO, Buffalo, NY, USA) were separated and centrifuged. The cells were mixed well by pipetting to evenly disperse the cells, and then divided into 100 ml 3 cell culture dishes in a predetermined number of 10 ml and passaged twice a week to be used for the experiment.
이렇게 배양된 세포를 96 well 세포 배양 플레이트에 5×104(cells/ml) 농도가 되도록 100μl씩 분주하여, 37℃, 5% CO2 incubator에서 24시간 배양한 후, 배지는 제거하고, 락토바실러스 브레비스 BNT 11 균주의 동결건조 분말 시료를 각 농도별로 희석하여 well당 100μl씩 첨가하였다. 대조군에는 시료 대신 배지를 100μl씩 첨가하였다. 이 plate를 다시 37℃, 5% CO2 incubator 에서 24시간 배양하였다. 배양 후 MTT assay를 위하여 3-(4,5-dimethylthiazole)-2,5-diphenyltetrazolium bromide(MTT) 시약 4mg을 PBS로 녹인 후, 배지양의 1/10인 10 μl씩 첨가하고 3 내지 4시간 동안 추가적으로 배양하여 MTT가 환원되도록 하였다. 배양 종료 후 생성된 formazan 결정을 가라앉힌 후, 각 well에 형성된 결정이 흐트러지지 않도록 주의하면서 반응 후 남은 MTT가 처리된 배지를 제거하였다. 배지가 제거된 각 well에 formazan 결정을 용해시키기 위하여 DMSO를 100μl씩 분주하여 5 내지 10분간 반응시켜 microplate reader(EL808, BIO-TEK instrument Inc, Winooski VT, USA)로 540nm에서 흡광도를 측정하였다. 이 흡광도는 MTT가 세포에 의해서 환원된 양을 나타내며, 따라서 각 well에 존재하는 세포의 생존 수와 비례한다. 세포생존율은 다음의 식에 따라 계산하였고, 그 결과를 도 1에 나타내었다. The cultured cells were dispensed in 100 μl aliquots to 5 × 10 4 (cells / ml) in 96 well cell culture plates, incubated in 37 ° C., 5% CO 2 incubator for 24 hours, and then the medium was removed. Lyophilized powder samples of Brevis BNT 11 strain were diluted at each concentration and added to 100 μl per well. 100 μl of medium was added to the control instead of the sample. The plate was incubated again at 37 ° C. in a 5% CO 2 incubator for 24 hours. After incubation, 4 mg of 3- (4,5-dimethylthiazole) -2,5-diphenyltetrazolium bromide (MTT) reagent was dissolved in PBS for MTT assay, and then 10 μl of 1/10 of the medium was added thereto for 3 to 4 hours. Further incubation allowed MTT to be reduced. After the formazan crystals were immersed after the incubation, the MTT-treated media remaining after the reaction was removed while being careful not to disturb the crystals formed in each well. In order to dissolve formazan crystals in each well from which the medium was removed, 100 μl of DMSO was dispensed and reacted for 5 to 10 minutes, and the absorbance was measured at 540 nm with a microplate reader (EL808, BIO-TEK instrument Inc, Winooski VT, USA). This absorbance represents the amount of MTT reduced by the cells and is therefore proportional to the number of cells present in each well. Cell viability was calculated according to the following equation, the results are shown in FIG.
[Cell viability] (%) = (ABS sample / ABS control)×100[Cell viability] (%) = (ABS sample / ABS control) × 100
ABS sample : Absorbance of the experimental sampleABS sample: Absorbance of the experimental sample
ABS control : Absorbance of the controlABS control: Absorbance of the control
그 결과, 락토바실러스 브레비스 BNT 11 균주의 동결건조 분말을 시료로 처리하였을 때, 0 내지 500㎍/㎖ 농도에서 3T3-L1 세포의 생존율에 변화가 없는 것으로 관찰되었다. 이러한 결과를 통해, 본 발명에 신균주 락토바실러스 브레비스 BNT 11 균주는 세포독성을 일으키지 않으며 안전한 것을 확인할 수 있었다.As a result, when lyophilized powder of Lactobacillus brevis BNT 11 strain was treated with a sample, it was observed that there was no change in the survival rate of 3T3-L1 cells at a concentration of 0 to 500 µg / ml. Through these results, the strain Lactobacillus brevis BNT 11 strain in the present invention was confirmed that it does not cause cytotoxic and safe.
실시예 5. 질내 유해 세균에 대한 항균 활성 테스트Example 5 Antimicrobial Activity Test against Harmful Bacteria in the Vagina
BNT 11 균주의 질내 유해 세균에 대한 항균 활성을 아래와 같은 방법으로 검증하였다. The antibacterial activity against harmful bacteria in the vagina of BNT 11 strain was verified by the following method.
실험은 MRS 배지에 37℃에서 90rpm으로 17시간 동안 혐기성 조건으로 배양한 락토바실러스 브레비스 BNT 11 균주의 배양여액과, 상기 락토바실러스 브레비스 BNT 11 균주의 배양여액을 농축기에서 20 Brix 조건으로 농축한 농축액을 이용하여, 질염에 직접적으로 작용하는 대표적인 병원성 세균인 칸디다 알비칸스(Candida albicans ATCC 22972)에 대한 항균활성을 검증하였다.Experiment was carried out in the culture filtrate of Lactobacillus brevis BNT 11 strain cultured in anaerobic conditions for 17 hours at 90 rpm at 37 ℃ in MRS medium, and the concentrate of the culture filtrate of the Lactobacillus brevis BNT 11 strain in a
칸디다 알비칸스(C. albicans)의 배양은 YM Broth(Difco)를 사용하여 0.5 MacFarland 농도(세균수 약 1×108 CFU/mL)로 배양하고, 이를 2번 정도 희석하여 1×105 CFU/mL 농도로 조정한 후, Nutrient agar(Difco, USA)에 60㎕ 용량으로 도말한 후, 이를 건조시키고, 멸균된 paper disc(6mm, Whatman, UK)를 올려놓았다.Culture of Candida albicans ( C. albicans ) was incubated at 0.5 MacFarland concentration (about 1 × 10 8 CFU / mL) using YM Broth (Difco), and diluted twice to 1 × 10 5 CFU / After adjusting to the mL concentration, 60 μl of Nutrient agar (Difco, USA) was plated, dried, and placed on a sterile paper disc (6 mm, Whatman, UK).
상기 준비된 락토바실러스 브레비스 BNT 11 균주의 배양여액과 이를 20 Brix 조건으로 농축한 농축액을 paper disc에 30㎕ 용량으로 점적한 후, 37℃, 5% CO2 조건으로 인큐베이터에서 24시간 배양하여 억제환의 지름을 측정하였다(표 3). The culture filtrate of the prepared Lactobacillus brevis BNT 11 strain and the concentrate concentrated at 20 Brix conditions were instilled in a 30 μl volume on a paper disc, followed by culturing in an incubator at 37 ° C. and 5% CO 2 for 24 hours to determine the diameter of the inhibitory ring. Was measured (Table 3).
[표 3]TABLE 3
그 결과, 본 발명의 락토바실러스 브레비스 BNT 11 균주의 배양여액은 질염에 직접적으로 작용하는 병원성 세균인 칸디다 알비칸스의 생장을 직접적으로 억제하는 효과가 있음을 확인할 수 있었다. 또한, 이러한 BNT 11 균주의 배양여액의 칸디다 알비칸스에 대한 생장억제능은 농축액에서 더욱 증가하는 것으로 나타나, 농도 의존적임을 확인할 수 있었다. As a result, it was confirmed that the culture filtrate of the Lactobacillus brevis BNT 11 strain of the present invention has an effect of directly inhibiting the growth of Candida albicans, a pathogenic bacterium that directly acts on vaginitis. In addition, growth inhibition ability against Candida albicans of the culture filtrate of the BNT 11 strain was shown to increase further in the concentrate, it was confirmed that the concentration dependent.
이상으로 본 발명의 특정한 부분을 상세히 기술하였는 바, 당업계의 통상의 지식을 가진 자에게 있어서 이러한 구체적인 기술은 단지 바람직한 구현예일 뿐이며, 이에 본 발명의 범위가 제한되는 것이 아닌 점은 명백하다. 따라서, 본 발명의 실질적인 범위는 첨부된 청구항과 그의 등가물에 의하여 정의된다고 할 것이다.Having described the specific part of the present invention in detail, it is apparent to those skilled in the art that the specific technology is merely a preferred embodiment, and the scope of the present invention is not limited thereto. Thus, the substantial scope of the present invention will be defined by the appended claims and equivalents thereof.
<110> LEE, Tae sun <120> Novel Strain of Lactobacillus brevis BNT 11 Having Antimicrobial Effect Against Pathogenic Microorganisms, and Uses thereof <130> PN180013AN <160> 1 <170> KoPatentIn 3.0 <210> 1 <211> 1511 <212> DNA <213> Lactobacillus brevis <400> 1 ctcaggacga acgctggcgg catgcctaat acatgcaagt cgaacgagct tccgttgaat 60 gacgtgcttg cactgatttc aacaatgaag cgagtggcga actggtgagt aacacgtggg 120 gaatctgccc agaagcaggg gataacactt ggaaacaggt gctaataccg tataacaaca 180 aaatccgcat ggattttgtt tgaaaggtgg cttcggctat cacttctgga tgatcccgcg 240 gcgtattagt tagttggtga ggtaaaggcc caccaagacg atgatacgta gccgacctga 300 gagggtaatc ggccacattg ggactgagac acggcccaaa ctcctacggg aggcagcagt 360 agggaatctt ccacaatgga cgaaagtctg atggagcaat gccgcgtgag tgaagaaggg 420 tttcggctcg taaaactctg ttgttaaaga agaacacctt tgagagtaac tgttcaaggg 480 ttgacggtat ttaaccagaa agccacggct aactacgtgc cagcagccgc ggtaatacgt 540 aggtggcaag cgttgtccgg atttattggg cgtaaagcga gcgcaggcgg ttttttaagt 600 ctgatgtgaa agccttcggc ttaaccggag aagtgcatcg gaaactggga gacttgagtg 660 cagaagagga cagtggaact ccatgtgtag cggtggaatg cgtagatata tggaagaaca 720 ccagtggcga aggcggctgt ctagtctgta actgacgctg aggctcgaaa gcatgggtag 780 cgaacaggat tagataccct ggtagtccat gccgtaaacg atgagtgcta agtgttggag 840 ggtttccgcc cttcagtgct gcagctaacg cattaagcac tccgcctggg gagtacgacc 900 gcaaggttga aactcaaagg aattgacggg ggcccgcaca agcggtggag catgtggttt 960 aattcgaagc tacgcgaaga accttaccag gtcttgacat cttctgccaa tcttagagat 1020 aagacgttcc cttcggggac agaatgacag gtggtgcatg gttgtcgtca gctcgtgtcg 1080 tgagatgttg ggttaagtcc cgcaacgagc gcaaccctta ttatcagttg ccagcattca 1140 gttgggcact ctggtgagac tgccggtgac aaaccggagg aaggtgggga tgacgtcaaa 1200 tcatcatgcc ccttatgacc tgggctacac acgtgctaca atggacggta caacgagtcg 1260 cgaagtcgtg aggctaagct aatctcttaa agccgttctc agttcggatt gtaggctgca 1320 actcgcctac atgaagttgg aatcgctagt aatcgcggat cagcatgccg cggtgaatac 1380 gttcccgggc cttgtacaca ccgcccgtca caccatgaga gtttgtaaca cccaaagccg 1440 gtgagataac cttcgggagt cagccgtcta aggtgggaca gatgattagg gtgaagtcgt 1500 acaaagggta a 1511 <110> LEE, Tae sun <120> Novel Strain of Lactobacillus brevis BNT 11 Having Antimicrobial Effect Against Pathogenic Microorganisms, and Uses <130> PN180013AN <160> 1 <170> KoPatentIn 3.0 <210> 1 <211> 1511 <212> DNA <213> Lactobacillus brevis <400> 1 ctcaggacga acgctggcgg catgcctaat acatgcaagt cgaacgagct tccgttgaat 60 gacgtgcttg cactgatttc aacaatgaag cgagtggcga actggtgagt aacacgtggg 120 gaatctgccc agaagcaggg gataacactt ggaaacaggt gctaataccg tataacaaca 180 aaatccgcat ggattttgtt tgaaaggtgg cttcggctat cacttctgga tgatcccgcg 240 gcgtattagt tagttggtga ggtaaaggcc caccaagacg atgatacgta gccgacctga 300 gagggtaatc ggccacattg ggactgagac acggcccaaa ctcctacggg aggcagcagt 360 agggaatctt ccacaatgga cgaaagtctg atggagcaat gccgcgtgag tgaagaaggg 420 tttcggctcg taaaactctg ttgttaaaga agaacacctt tgagagtaac tgttcaaggg 480 ttgacggtat ttaaccagaa agccacggct aactacgtgc cagcagccgc ggtaatacgt 540 aggtggcaag cgttgtccgg atttattggg cgtaaagcga gcgcaggcgg ttttttaagt 600 ctgatgtgaa agccttcggc ttaaccggag aagtgcatcg gaaactggga gacttgagtg 660 cagaagagga cagtggaact ccatgtgtag cggtggaatg cgtagatata tggaagaaca 720 ccagtggcga aggcggctgt ctagtctgta actgacgctg aggctcgaaa gcatgggtag 780 cgaacaggat tagataccct ggtagtccat gccgtaaacg atgagtgcta agtgttggag 840 ggtttccgcc cttcagtgct gcagctaacg cattaagcac tccgcctggg gagtacgacc 900 gcaaggttga aactcaaagg aattgacggg ggcccgcaca agcggtggag catgtggttt 960 aattcgaagc tacgcgaaga accttaccag gtcttgacat cttctgccaa tcttagagat 1020 aagacgttcc cttcggggac agaatgacag gtggtgcatg gttgtcgtca gctcgtgtcg 1080 tgagatgttg ggttaagtcc cgcaacgagc gcaaccctta ttatcagttg ccagcattca 1140 gttgggcact ctggtgagac tgccggtgac aaaccggagg aaggtgggga tgacgtcaaa 1200 tcatcatgcc ccttatgacc tgggctacac acgtgctaca atggacggta caacgagtcg 1260 cgaagtcgtg aggctaagct aatctcttaa agccgttctc agttcggatt gtaggctgca 1320 actcgcctac atgaagttgg aatcgctagt aatcgcggat cagcatgccg cggtgaatac 1380 gttcccgggc cttgtacaca ccgcccgtca caccatgaga gtttgtaaca cccaaagccg 1440 gtgagataac cttcgggagt cagccgtcta aggtgggaca gatgattagg gtgaagtcgt 1500 acaaagggta a 1511
Claims (7)
서열번호 1의 16s rDNA 서열을 포함하는 신균주 락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)(수탁번호: KCCM12167P). Have antibacterial activity against Salmonella paratyphi and Salmonella typhimurim strains,
Lactobacillus brevis BNT 11 (Accession: KCCM12167P) comprising the 16s rDNA sequence of SEQ ID NO: 1.
서열번호 1의 16s rDNA 서열을 포함하는 신균주 락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)(수탁번호: KCCM12167P), 또는 그 배양액을 유효성분으로 함유하는,
살모넬라 파라티피(Salmonella paratyphi) 및 살모넬라 타이피뮤리움(Salmonella typhimurim) 균주에 의한 감염증의 치료 또는 예방용 약학적 조성물. Has antibacterial activity against Salmonella paratyphi and Salmonella typhimurim strains,
Lactobacillus brevis BNT 11 (Accession No .: KCCM12167P) containing the 16s rDNA sequence of SEQ ID NO: 1, or containing the culture solution as an active ingredient,
Pharmaceutical composition for the treatment or prevention of infection by Salmonella paratyphi ( Salmonella paratyphi ) and Salmonella typhimurim strains.
서열번호 1의 16s rDNA 서열을 포함하는 신균주 락토바실러스 브레비스 BNT 11(Lactobacillus brevis BNT 11)(수탁번호: KCCM12167P), 또는 그 배양액을 유효성분으로 포함하는,
살모넬라 파라티피(Salmonella paratyphi) 및 살모넬라 타이피뮤리움(Salmonella typhimurim) 균주에 의한 감염증의 예방용 건강기능식품. Has antibacterial activity against Salmonella paratyphi and Salmonella typhimurim strains,
Lactobacillus brevis BNT 11 (Accession Number: KCCM12167P) comprising the 16s rDNA sequence of SEQ ID NO: 1 (Accession Number: KCCM12167P), or a culture solution thereof, as an active ingredient,
Health functional food for the prevention of infection by Salmonella paratyphi and Salmonella typhimurim strains.
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