WO2023038418A1 - Souche de bifidobacterium bifidum eps da-laim pour le maintien de la santé intestinale, possédant l'effet de favoriser la croissance des lactobacilles, et polysaccharides qui en sont issus - Google Patents

Souche de bifidobacterium bifidum eps da-laim pour le maintien de la santé intestinale, possédant l'effet de favoriser la croissance des lactobacilles, et polysaccharides qui en sont issus Download PDF

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WO2023038418A1
WO2023038418A1 PCT/KR2022/013421 KR2022013421W WO2023038418A1 WO 2023038418 A1 WO2023038418 A1 WO 2023038418A1 KR 2022013421 W KR2022013421 W KR 2022013421W WO 2023038418 A1 WO2023038418 A1 WO 2023038418A1
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eps
bifidobacterium bifidum
laim
kctc
exopolysaccharide
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PCT/KR2022/013421
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English (en)
Korean (ko)
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한상덕
한영선
박민주
조현일
이은석
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동아제약 주식회사
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    • 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
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/66—Microorganisms or materials therefrom
    • A61K35/74—Bacteria
    • A61K35/741—Probiotics
    • A61K35/744—Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
    • A61K35/745—Bifidobacteria
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
    • A61K8/99—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution from microorganisms other than algae or fungi, e.g. protozoa or bacteria
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P39/00—General protective or antinoxious agents
    • A61P39/06—Free radical scavengers or antioxidants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • 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
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P19/00—Preparation of compounds containing saccharide radicals
    • C12P19/04—Polysaccharides, i.e. compounds containing more than five saccharide radicals attached to each other by glycosidic bonds
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00—Microorganisms ; Processes using microorganisms
    • C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales

Definitions

  • the present invention relates to an intestinal health strain, Bifidobacterium bifidum EPS DA-LAIM, and a polysaccharide thereof having a Lactobacillus lactic acid bacterium growth promoting effect.
  • Probiotics is a general term for microorganisms that are beneficial to the human body, and typically include lactic acid bacteria and bifidobacteria. After entering the human body, they adhere to intestinal epithelial cells to inhibit the growth of harmful microorganisms, improve diarrhea and constipation, and exhibit intestinal microbial balance, such as controlling intestinal microbial balance, as well as exhibiting physiological effects such as immune activity enhancement and anticancer activity. It maintains the balance of intestinal microorganisms and has a beneficial effect on human health.
  • exopolysaccharide EPS
  • the polysaccharide produced by Bifidobacterium has various and unique physical properties, so it can be used as a material for physical and functional functions, and it has excellent physiological functions such as anticancer effect, cholesterol suppression, immune regulation, and constipation improvement. this is gradually increasing.
  • EPS is divided into a capsular form attached to the cell wall and a ropy form secreted extracellularly according to the structure and degree of polymerization.
  • Ropy-EPS is characterized by the formation of a viscous material in the form of solid lines in colonies, and provides relatively high viscosity, water retention and soft density.
  • literature has been reported that Ropy-EPS significantly increases the survival and attachment of lactic acid bacteria compared to Non-ropy-EPS in previous studies, lowers serum cholesterol, and suppresses hyperimmune reactions.
  • An object of the present invention is to discover and provide bifidus bacteria that produce Ropy-EPS.
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP) is provided.
  • the strain is preferably characterized by producing exopolysaccharide (EPS) as a postbiotics, and the EPS (exopolysaccharide) is preferably EPS in the form of ropy secreted extracellularly. It is characterized in that it contains (exopolysaccharide).
  • EPS exopolysaccharide
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP), a pharmaceutical composition for preventing or treating diseases caused by excessive accumulation of active oxygen containing any one selected from dead cells and cultures thereof is provided.
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP), dead cells thereof, and a food composition for improving diseases caused by excessive accumulation of active oxygen containing any one selected from its culture are provided.
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP), a health functional food for improving diseases caused by excessive accumulation of active oxygen containing any one selected from dead cells and culture thereof is provided.
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP), provides a cosmetic composition for improving skin aging caused by excessive accumulation of active oxygen containing any one selected from dead cells and cultures thereof.
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS (exopolysaccharide) produced by EPS DA-LAIM (KCTC 14638BP) is provided, and the EPS (exopolysaccharide) preferably includes exopolysaccharide (EPS) in the form of ropy secreted extracellularly do.
  • EPS exopolysaccharide
  • the EPS exopolysaccharide
  • the EPS is preferably a) Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP) Precipitating and removing proteins by adding an acidic material to the culture supernatant; b) extracting the polysaccharide by adding a hydrophilic organic solvent to the protein removal liquid of step a);
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum )
  • EPS exopolysaccharide
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum )
  • EPS exopolysaccharide
  • KCTC 14638BP KCTC 14638BP
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) Provides a health functional food for improving diseases caused by excessive accumulation of active oxygen containing EPS (exopolysaccharide) produced by EPS DA-LAIM (KCTC 14638BP).
  • EPS exopolysaccharide
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum )
  • EPS exopolysaccharide
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP), its dead cell, its culture and Bifidobacterium bifidum ( Bifidobacterium bifidum )
  • EPS exopolysaccharide
  • KCTC 14638BP EPS DA-LAIM
  • the composition for promoting the growth of probiotics preferably Lactobacillus plantarum ( Lactobacillus plantarum ), Lactobacillus fermentum ( Lactobacillus fermentum ), Lactobacillus helveticus ( Lactobacillus helveticus ) Growth of any one or more probiotics selected It is good to promote
  • the strain of the present invention is characterized by producing EPS (exopolysaccharide) as a postbiotics, and has the advantage of providing excellent antioxidant efficacy and prebiotics action ability to promote the growth of the genus Lactobacillus.
  • Figure 1 shows the result of evaluating the antioxidant capacity of the strain of the present invention, scavenging activity (Scavening activity).
  • Figure 2 is a result of evaluating the prebiotic effect of the strain of the present invention, Lactobacillus plantarum ( Lactobacillus plantarum ), Lactobacillus fermentum ( Lactobacillus fermentum ), Lactobacillus helveticus ( Lactobacillus helveticus ) for prebiotics It represents the prebiotic index.
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP) is provided.
  • the strain discovered in the present invention is preferably characterized in that it produces EPS (exopolysaccharide) as a postbiotics, and the EPS (exopolysaccharide) is preferably secreted extracellularly. Characterized in that it contains exopolysaccharide (EPS) in the form of ropy. It has been confirmed that the strain of the present invention has an excellent antioxidant effect and has a prebiotics action ability to promote the growth of the genus Lactobacillus.
  • EPS exopolysaccharide
  • the above strain is 'Bifidobacterium bifidum ( Bifidobacterium bifidum ) It was named 'EPS DA-LAIM', and was deposited with the Korea Research Institute of Bioscience and Biotechnology and assigned the accession number KCTC 14638BP on July 15, 2021.
  • the strain preferably produces EPS (exopolysaccharide). Therefore, the present invention is a bifidobacterium bifidum ( Bifidobacterium bifidum ) Provides EPS (exopolysaccharide) produced by EPS DA-LAIM (KCTC 14638BP).
  • the EPS exopolysaccharide
  • the EPS is preferably a) Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP) Precipitating and removing proteins by adding an acidic material to the culture supernatant; b) extracting the polysaccharide by adding a hydrophilic organic solvent to the protein removal liquid of step a);
  • trichloroacetic acid may be used as an acidic material in step a), and in this case, it is preferable to react at a low temperature for 1 to 6 hours.
  • ethanol may be used as an example of the hydrophilic organic solvent in step b. At this time, ethanol is preferably added slowly in a volume ratio of 1 to 3 times that of the supernatant using cooled 80 to 100% (v/v) ethanol and reacted at low temperature for 12 to 24 hours to precipitate the precipitate.
  • the trichloroacetic acid is added to a final concentration of 4 to 10% (w/v) and reacted at a low temperature of 0 to 6 ° C, and the ethanol is cooled to a temperature of -20 to -15 ° C. It is good to use what has been done.
  • EPS exopolysaccharide
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP), its dead cells, its culture, and EPS (exopolysaccharide) containing any one selected from the above-mentioned to provide a pharmaceutical composition for preventing or treating diseases caused by excessive accumulation of active oxygen .
  • Antioxidant function is to suppress or slow down the oxidation reaction that occurs in the body.
  • the oxidation reaction that occurs in the body induces aging and cell mutation to generate cancer cells. Therefore, the public is also interested in antioxidant action as a preventive measure to suppress such negative reactions in the body.
  • diseases caused by excessive accumulation of active oxygen include, for example, cancer, inflammation, anemia, myocardial infarction, cerebral palsy, arteriosclerosis, diabetes, rheumatoid arthritis, Parkinson's disease, and autoimmune diseases It may be any one selected from, but is not limited thereto and may include all diseases accompanied by a reaction in which active oxygen is excessively accumulated without limitation.
  • prevention refers to any action that suppresses or delays the onset of diseases affected by the administration of the pharmaceutical composition according to the present invention, that is, diseases caused by excessive accumulation of active oxygen.
  • treatment refers to all activities that improve or beneficially change symptoms caused by diseases caused by excessive accumulation of active oxygen by the administration of the pharmaceutical composition according to the present invention.
  • the Bifidobacterium bifidum of the present invention ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP), a composition comprising any one selected from dead cells, cultures thereof, and EPS (exopolysaccharide) as an active ingredient, in addition to the above ingredients, an active ingredient exhibiting the same or similar function
  • EPS exopolysaccharide
  • the pharmaceutical composition of the present invention may further include a pharmaceutically acceptable carrier in addition to Bifidobacterium bifidum EPS DA-LAIM.
  • the type of carrier that can be used in the present invention is not particularly limited, and any carrier commonly used in the art may be used.
  • Non-limiting examples of the carrier include lactose, dextrose, sucrose, sorbitol, mannitol, saline, sterile water, Ringer's solution, buffered saline, albumin injection solution, xylitol, erythritol, maltitol, maltodextrin, glycerol, ethanol, and the like. can These may be used alone or in combination of two or more.
  • the pharmaceutical composition of the present invention may be used by adding other pharmaceutically acceptable additives such as antioxidants, excipients, diluents, buffers or bacteriostats, surfactants, binders, fillers, bulking agents, wetting agents, disintegrants , a dispersant or a lubricant may be additionally added and used.
  • other pharmaceutically acceptable additives such as antioxidants, excipients, diluents, buffers or bacteriostats, surfactants, binders, fillers, bulking agents, wetting agents, disintegrants , a dispersant or a lubricant may be additionally added and used.
  • Bifidobacterium bifidum EPS DA-LAIM may be included in an amount of 0.00001% to 99.99% by weight based on the total weight of the pharmaceutical composition, preferably 0.1% by weight. Weight% to 90% by weight, more preferably 0.1% to 70% by weight, more preferably 0.1% to 50% by weight, but not limited thereto, the condition of the subject to be administered, the type of specific disease, and the progress Depending on the degree, etc., it may be variously changed. If necessary, it may be included in the entire content of the pharmaceutical composition.
  • the pharmaceutically effective amount and effective dosage of the pharmaceutical composition of the present invention may vary depending on the formulation method, administration method, administration time and/or route of administration of the pharmaceutical composition, and Type and degree of reaction, type of subject to be administered, age, weight, general health condition, symptom or severity of disease, sex, diet, excretion, drugs used simultaneously or at different times in the subject, or components of other compositions, etc. It can vary according to various factors including, and similar factors well known in the medical field, and those skilled in the art can easily determine and prescribe an effective dosage for the desired treatment.
  • the daily dosage of the pharmaceutical composition of the present invention is about 0.01 to 1,000 mg/kg, preferably 0.1 to 100 mg/kg, and may be administered once or several times a day.
  • Administration of the pharmaceutical composition of the present invention may be administered once a day, or may be divided and administered several times.
  • the pharmaceutical composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with conventional therapeutic agents. Considering all of the above factors, it can be administered in an amount that can obtain the maximum effect with the minimum amount without side effects, which can be easily determined by those skilled in the art.
  • the pharmaceutical composition of the present invention may be additionally used in combination with various methods such as hormone therapy and drug therapy to prevent or treat diseases caused by excessive accumulation of active oxygen.
  • the term "administration” means introducing the pharmaceutical composition of the present invention to a patient by any suitable method, and the route and method of administration of the pharmaceutical composition of the present invention may be independent, respectively, and the purpose Any administration route and administration method may be followed without particular limitation, as long as the pharmaceutical composition can reach the corresponding site.
  • the pharmaceutical composition may be administered by oral administration or parenteral administration, and may be formulated into various dosage forms suitable for oral administration or parenteral administration.
  • Non-limiting examples of preparations for oral administration using the pharmaceutical composition of the present invention include oily suspensions, troches, lozenges, tablets, aqueous suspensions, prepared powders, granules, emulsions, hard capsules, and soft capsules, syrups or elixirs; and the like.
  • a binder such as sorbitol, mannitol, starch, amylopectin, cellulose lactose, saccharose or gelatin; lubricating oils such as magnesium stearate, calcium stearate, sodium stearyl fumarate or polyethylene glycol wax; excipients such as dicalcium phosphate and the like; A disintegrant such as corn starch or sweet potato starch may be used, and aromatics, syrups, sweeteners, and the like may also be used.
  • a liquid carrier such as fatty oil may be additionally used in addition to the above-mentioned materials.
  • intramuscular administration As a method for parenteral administration of the pharmaceutical composition of the present invention, intramuscular administration, transdermal administration, intravenous administration, intraperitoneal administration, or subcutaneous administration may be used, and a method of applying, spraying, or inhaling the composition to a diseased area It can also be used, but is not limited thereto.
  • Non-limiting examples of parenteral preparations using the pharmaceutical composition of the present invention include injection solutions, suppositories, ointments, powders for application, oils, powders for respiratory inhalation, aerosols for sprays, creams, and the like.
  • aqueous solutions In order to formulate the pharmaceutical composition of the present invention for parenteral administration, sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, freeze-dried preparations, external preparations, etc. may be used. Vegetable oils, propylene glycol, polyethylene glycol, injectable esters such as ethyl oleate, and the like can be used.
  • the pharmaceutical composition of the present invention is formulated as an injection solution
  • the pharmaceutical composition of the present invention is mixed in water together with a stabilizer or buffer to prepare a solution or suspension, which is used for unit administration in an ampoule or vial. can be formulated.
  • a propellant or the like may be mixed with additives so that the water-dispersed concentrate or wet powder is dispersed.
  • composition of the present invention When the pharmaceutical composition of the present invention is formulated into ointments, oils, creams, powders for application, skin external preparations, etc., animal oils, vegetable oils, waxes, paraffins, polyethylene glycols, silicones, bentonites, silicas, talc, starch, tras It can be formulated using Kant, cellulose derivatives, zinc oxide and the like as carriers.
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP), its dead cells, its culture, and EPS (exopolysaccharide) containing any one selected from the above provides a food composition for improving diseases caused by excessive accumulation of active oxygen.
  • the term “improvement” refers to all activities that improve or beneficially change diseases caused by excessive accumulation of active oxygen by administration of the composition of the present invention.
  • the Bifidobacterium bifidum EPS DA-LAIM is preferably included in an amount of 0.00001 to 50% by weight relative to the food composition . If it is less than 0.00001% by weight, the effect is insufficient, and if it exceeds 50% by weight, the increase in effect compared to the amount used is insignificant, which is uneconomical.
  • the food composition of the present invention is, for example, noodles, gums, dairy products, ice creams, meats, grains, caffeinated beverages, general beverages, chocolates, breads, snacks, confectionery products, candies, pizzas, jellies, alcoholic beverages, alcohol, and vitamin complexes. And it may be any one selected from other health supplements, but is not necessarily limited thereto.
  • the food composition of the present invention when used as a food additive, it may be added as it is or used together with other foods or food ingredients, and may be appropriately used according to conventional methods.
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP), its dead cells, its culture, and EPS (exopolysaccharide) containing any one selected from the above provides a health functional food for improving diseases caused by excessive accumulation of active oxygen.
  • Bifidobacterium bifidum Bifidobacterium bifidum
  • EPS DA-LAIM KCTC 14638BP
  • EPS exopolysaccharide
  • health functional food refers to food manufactured and processed using raw materials or ingredients having functional properties useful for the human body in accordance with the Health Functional Food Act No. 6727, and “functional” refers to the structure of the human body. And it refers to intake for the purpose of obtaining useful effects for health purposes such as regulating nutrients for functions or physiological functions.
  • the food composition and health functional food of the present invention may include additional ingredients that are commonly used and can improve odor, taste, and visual properties.
  • it may include biotin, folate, panthotenic acid, vitamins A, C, D, E, B1, B2, B6, B12, niacin, and the like.
  • minerals such as chromium (Cr), magnesium (Mg), manganese (Mn), copper (Cu), zinc (Zn), iron (Fe), and calcium (Ca) may be included.
  • amino acids such as cysteine, valine, lysine, and tryptophan may be included.
  • preservatives potassium sorbate, sodium benzoate, salicylic acid, sodium dihydroacetate, etc.
  • coloring agents tar colorant, etc.
  • coloring agents sodium nitrite, sodium nitrite, etc.
  • bleaching agents sodium sulfite
  • disinfectants bleaching powder and high bleaching powder, sodium hypochlorite, etc.
  • swelling agent alum, D-potassium hydrogen tartrate, etc.
  • strengthening agent emulsifier, thickener (thickener), coating agent, antioxidant (butylhydroxyanisole (BHA), butylhydroxytoluene (BHT), etc.
  • Food additives such as seasonings (MSG monosodium glutamate, etc.), sweeteners (dulcin, cyclemate, saccharin, sodium, etc.), flavorings (vanillin, lactones, etc.), gum base, antifoaming agent, solvent, improver, etc. can be added.
  • the additive may be selected according to the type of food and
  • the content of Bifidobacterium bifidum EPS DA-LAIM is not particularly limited, and can be varied depending on the condition of the subject to be administered, the type of specific disease, the degree of progression, etc. there is. If necessary, it may also be included in the total content of food.
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP), its dead cell, its culture and Bifidobacterium bifidum ( Bifidobacterium bifidum )
  • EPS exopolysaccharide
  • the composition for promoting the growth of probiotics preferably Lactobacillus plantarum ( Lactobacillus plantarum ), Lactobacillus fermentum ( Lactobacillus fermentum ), Lactobacillus helveticus ( Lactobacillus helveticus ) Growth of any one or more probiotics selected It is good to promote
  • Bifidobacterium bifidum Bifidobacterium bifidum
  • EPS exopolysaccharide
  • KCTC 14638BP EPS DA-LAIM
  • KCTC 14638BP EPS DA-LAIM
  • the composition for promoting the growth of probiotics claimed in the present invention is also called prebiotics in the art.
  • the present invention is Bifidobacterium bifidum ( Bifidobacterium bifidum ) EPS DA-LAIM (KCTC 14638BP), its dead cells, its culture, and EPS (exopolysaccharide) containing any one selected from the above provides a cosmetic composition for improving skin aging due to excessive accumulation of active oxygen.
  • Bifidobacterium bifidum Bifidobacterium bifidum
  • EPS DA-LAIM KCTC 14638BP
  • EPS exopolysaccharide
  • the antioxidant effect of cosmetics means the ability to improve or prevent skin functions damaged by oxidation, through which skin aging can be prevented or improved.
  • the Bifidobacterium bifidum EPS DA-LAIM is preferably contained in an amount of 0.0001 to 30.0% by weight based on the total weight of the cosmetic composition. More preferably, it is good to contain 0.01 to 10% by weight based on the total weight of the cosmetic composition.
  • the content of Bifidobacterium bifidum EPS DA-LAIM is less than 0.0001% by weight, the antioxidant effect is insignificant, and when it exceeds 30.0% by weight, the obvious effect does not increase with the increase in the content.
  • the ingredients included in the cosmetic composition of the present invention may include ingredients commonly used in cosmetic compositions other than the Bifidobacterium bifidum EPS DA-LAIM of the present invention as active ingredients, such as antioxidants , conventional adjuvants such as stabilizers, solubilizers, vitamins, pigments and flavors, and carriers.
  • the cosmetic composition of the present invention can be prepared in any formulation conventionally prepared in the art, for example, a solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing , Oil, pack, massage cream and spray, etc., but may be formulated, but is not limited thereto. More specifically, it may be prepared in the form of softening lotion, nutrient lotion, nutrient cream, massage cream, essence, cleansing cream, cleansing foam, cleansing water, pack, spray or powder.
  • the formulation of the cosmetic composition of the present invention is a paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tracanth, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide as a carrier component this can be used
  • a solvent, solubilizing agent or emulsifying agent is used as a carrier component, such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene fatty acid esters of glycol, 1,3-butyl glycol oil, glycerol aliphatic esters, polyethylene glycol or sorbitan.
  • the formulation of the cosmetic composition of the present invention is a suspension
  • a liquid diluent such as water, ethanol or propylene glycol, an ethoxylated isostearyl alcohol, a suspending agent such as polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester , microcrystalline cellulose, aluminum metahydroxide, bentonite, agar or tracanth, and the like
  • a suspending agent such as polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester
  • microcrystalline cellulose aluminum metahydroxide
  • bentonite agar or tracanth, and the like
  • lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component, and in particular, in the case of a spray, additional chlorofluorohydro propellants such as carbon, propane/butane or dimethyl ether.
  • the formulation of the cosmetic composition of the present invention is surfactant-containing cleansing
  • carrier components aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyl taurate, sarcosinate, Fatty acid amide ether sulfates, alkylamidobetaines, fatty alcohols, fatty acid glycerides, fatty acid diethanolamides, vegetable oils, lanolin derivatives, or ethoxylated glycerol fatty acid esters may be used.
  • the cosmetic composition of the present invention is a soap, a surfactant-containing cleansing formulation, or a surfactant-free cleansing formulation
  • the soap is liquid soap, powder soap, solid soap, and oil soap
  • the surfactant-containing cleansing formulation is a cleansing foam, cleansing water, cleansing towel, and a cleansing pack
  • the surfactant-free cleansing formulation is a cleansing cream. , cleansing lotion, cleansing water and cleansing gel, but are not limited thereto.
  • the feces were diluted step by step using sterile physiological saline by a decimal dilution method, and then 100 ⁇ l were plated on TOS-MUP selective media ( Bifidobacterium selective media). After anaerobic culture of the plated plate at 37° C. for 48 hours, a single colony was first selected from among the cultured strains, and then the colony was picked and spotted once more on a TOS-MUP selective medium. The plate was incubated anaerobically at 37° C. for 48 hours, and then it was confirmed once again that the colony grew well, and finally 129 bifidus candidate strains were selected.
  • TOS-MUP selective media Bifidobacterium selective media
  • the EPS production ability of 129 bifidobacteria isolated from feces of healthy Korean volunteers was quantitatively evaluated. Inoculate 1% of each strain in MRS broth (De Man, Rogosa and Sharpe, 0.05% L-cysteine) containing 10% sucrose as a sugar source, incubate anaerobically at 37 ° C for 48 hours, and centrifuge the culture medium at 4 ° C ( 10,000 ⁇ g, 25 min) to remove the cells, and to the supernatant, twice the volume of cooled 95% ethanol was slowly added to precipitate the EPS at 4 ° C for 15 to 24 hours. Thereafter, the precipitate was collected by centrifugation (10,000 ⁇ g, 25 min) at 4 ° C., and the remaining ethanol was dried and lyophilized to measure the weight of the solid content.
  • MRS broth De Man, Rogosa and Sharpe, 0.05% L-cysteine
  • Ropy-EPS Ropy-Exopolysaccharide
  • EPS DA-LAIM As a result of measuring the EPS production ability of 129 bifidobacteria isolated from feces of healthy Korean volunteers and the ropy characteristics of the EPS, Bifidobacterium bifidum, one of the best EPS production species, Bifidobacterium bifidum , as shown in Table 1 EPS DA-LAIM was selected.
  • the EPS DA-LAIM strain showed an excellent EPS production ability that was 3.6 times higher than that of other Bifidobacterium sp. A strand of 50 mm was formed, and it was confirmed that the EPS had ropy characteristics. Its 16S rRNA nucleotide sequence is shown in SEQ ID NO: 1.
  • the DPPH radical scavenging activity was confirmed by colorimetric method. Specifically, the Bifidobacterium stock was inoculated to a concentration of 1% (v/v) in MRS broth (0.05% L-cysteine), and incubated at 32-37 ° C for 24-168 hours under anaerobic conditions. Cells were recovered by centrifuging 1 ml of the cultured bacterial solution at 13,000 rpm and 4° C. for 3 minutes, and at this time, the number of bacteria was approximately 5.0 ⁇ 10 8 CFU/ml. The cells were washed once with sterilized PBS buffer (pH 7.4) and dissolved in 1 ml of the same new buffer.
  • PBS buffer pH 7.4
  • DPPH scavenging activity (%) (1-(sample OD 517 -blank OD 517 )/control OD 517 )*100
  • the DPPH scavenging activity of the EPS DA-LAIM strain was 59.6%, which was 1.9 times higher than that of vitamin C (0.1mg/ml), a representative antioxidant, and 3.2 times higher than that of the same strain of a famous domestic dry food product. (Fig. 1). 1 is a result of evaluating the antioxidant ability of the strain of the present invention.
  • the cell culture medium was centrifuged (10,000 ⁇ g, 25 min) at 4 ° C to remove the cells, and trichloroacetic acid was added to the recovered supernatant to a final concentration of 4 to 4 It was added to 10% and reacted at 4°C for 2 hours.
  • the reaction solution was centrifuged at 4 ° C (10,000 ⁇ g, 25 min) to remove precipitated proteins, and then 2 times of cold 95% ethanol was slowly added to the supernatant to precipitate EPS at 4 ° C for 15 to 24 hours.
  • the precipitate was collected by centrifugation (10,000 ⁇ g, 25 min) at 4 ° C, dissolved in tertiary distilled water, and dialysis-sack (M.W. cut off 10,000, Spectra / Por 6 membrane, Pre-wetted RC Tubing, Spectrum Laboratories, USA), dialyzed at 4° C. for 48 hours, and then lyophilized to prepare purified EPS.
  • MRS broth or TSB broth containing 2% EPS as a sugar source 100 ⁇ l of MRS broth or TSB broth containing 2% EPS as a sugar source was dispensed into a 96-well plate, and then the MRS broth contained intestinal beneficial bacteria activated the previous day ( lactic acid bacteria), the TSB broth was inoculated with 1% of the culture medium of intestinal harmful bacteria ( Escherichia coli ), respectively, and the OD 600 value after 24 hours was observed.
  • the same experiment was conducted on MRS broth or TSB broth with 2% inulin, a representative prebiotic, as a glycogen, and MRS broth or TSB broth to which no glycogen was added.
  • a blank group and a control group in which the same experiment was performed for MRS broth or TSB broth containing 2% glucose as a sugar source were also tested.
  • Prebiotic index Optical density of the growth of probiotic culture / Optical density of the growth of E. coli

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Abstract

La présente invention concerne une souche de bifidobactérium bifidum EPS DA-LAIM (KCTC 14638BP) pour la santé des intestins, ayant l'effet de promouvoir la croissance de lactobacillus, polysaccharides produit de ce dernier, et une composition comportant ces derniers, pour l'atténuation des maladies provoquées à l'accumulation excessive d'oxygène active.
PCT/KR2022/013421 2021-09-10 2022-09-07 Souche de bifidobacterium bifidum eps da-laim pour le maintien de la santé intestinale, possédant l'effet de favoriser la croissance des lactobacilles, et polysaccharides qui en sont issus WO2023038418A1 (fr)

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