Composition including complex mineral fermented extract as active ingredient for treating, preventing or alleviating inflammatory diseases and blood circulation disorders

Abstract

The present invention relates to a composition including a complex mineral fermented extract of biotite and bentonite as an active ingredient for anti-inflammation and blood circulation improvement, and a use thereof. The mineral complex fermented extract of biotite and bentonite in the present invention exhibits no cytotoxicity, effectively suppresses nitric oxide (NO) production and the expression of iNOS and COX-2 genes involved in inflammation, enhances vascular endothelial cell motility to promote angiogenesis, and inhibits platelet aggregation in a dose-dependent manner by suppressing the production of thromboxane B2, and thus can be advantageously used for the purposes of medicines, quasi-drugs, health functional foods, cosmetics, and the like for alleviating inflammation, improving blood circulation, and inhibiting thrombus production.

Description

Composition for the treatment, prevention or improvement of inflammatory diseases and blood circulation disorders containing mineral complex fermented extract as an active ingredient

[0001] The present invention relates to a composition containing a mineral complex fermentation extract as an active ingredient and a use thereof, and more specifically, to a composition for treating, preventing or improving inflammatory diseases and blood circulation disorders, containing a mineral complex fermentation extract of biotite and bentonite as an active ingredient.

[0002] The inflammatory response is a complex series of physiological reactions, including enzyme activation, secretion of inflammatory mediators, fluid infiltration, cell migration, and tissue destruction, as well as external symptoms such as erythema, edema, fever, and pain, when cells or tissues are damaged or infected with external infectious agents such as bacteria, fungi, viruses, and various types of allergens. Normally, the inflammatory response removes external infectious agents and regenerates damaged tissues to restore organism functions. However, if the antigen is not removed or the inflammatory response is excessive or persistent due to internal substances, it can rather promote mucosal damage, which in some cases can lead to diseases such as cancer.

[0003] It is known that various biochemical phenomena are involved in the cause of inflammation, and in particular, nitric oxide synthase (NOS), an enzyme that generates nitric oxide (NO), and enzymes related to the biosynthesis of prostaglandins are known to play the most important role in mediating the inflammatory response. Therefore, the goal of developing anti-inflammatory treatments is to block inflammation by inhibiting the activity of nitric oxide synthase (NOS), an enzyme that generates nitric oxide from L-arginine, or cyclooxygenase (COX), an enzyme related to the synthesis of prostaglandins from arachidonic acid.

[0004] Meanwhile, platelets exist in an inactive state under normal conditions, but when exposed to collagen in the blood vessel wall in wounds or diseased conditions, they are activated to secrete ADP or thromboxane A2 (TxA2). These secreted substances in turn activate platelets, ultimately inducing thrombosis. These activated platelets are the main cause of heart or cerebral ischemia and play an important role in the development of arteriosclerosis. During the normal wound healing process, thrombosis occurs when platelets adhere to damaged blood vessels, forming a monolayer, undergoing platelet aggregation, stabilization, and fixation. If platelet activation is excessive in the final stage, platelet aggregation continues, blocking the blood vessel.

[0005] Platelet-activating factors include thrombin, thromboxane A2, platelet-activating factor (PAF), and collagen fibers, and these substances are produced not only by activated platelets but also by inflammation.

[0006] Platelet activation causes not only arterial thrombosis but also peripheral vascular disorders, so peripheral vascular circulatory disorders can be overcome by regulating platelet activation. In fact, ginkgo biloba extract is used as a blood circulation improvement agent as a PAF antagonist. Meanwhile, atherothrombotic vascular disease accounts for approximately 40% of deaths in the Western world, and since platelets play a key role in atherothrombosis, research into the development of antiplatelet agents has been focused. Currently, aspirin, a COX-1 and TxA2 inhibitor, and clopidogrel, a P2Y12 receptor blocker, an ADP receptor, have been developed and used. In addition, the above medicine is used to prevent thrombosis during percutaneous transluminal coronary angioplasty (PTCA) and stent insertion, and as a preventive agent for restenosis after stent installation by grafting the medicine having the above platelet aggregation inhibitory effect by applying or embedding it in the stent itself.

[0007] However, despite the availability of various medications, conventionally used drugs like aspirin still have side effects, such as gastrointestinal bleeding. Therefore, the development of novel anti-inflammatory agents or platelet aggregation inhibitors to replace them is urgent. To date, research on the synergistic effects of combinations of extracts composed of natural ingredients on anti-inflammation and blood circulation improvement has been insufficient. Consequently, research on anti-inflammatory or blood circulation-enhancing agents utilizing complex extracts remains limited.

[0008] Against this backdrop, the inventors of the present invention have made great efforts to find a combination of natural extracts that simultaneously have anti-inflammatory activity and blood circulation disorder improvement activity, and as a result, they have found that a mineral complex fermented extract, in which two mineral distillation extracts composed of biotite and bentonite are fermented with a complex strain of Bacillus toyonensis and Bacillus simplex, exhibits a synergistic effect in improving inflammation and blood circulation disorder, and can be usefully utilized as an anti-inflammatory agent and blood circulation improvement agent by replacing conventional medical drugs, thereby completing the present invention.

[0009] The main purpose of the present invention is to provide a composite fermented extract of biotite and bentonite that simultaneously exhibits anti-inflammatory and blood circulation disorder improving effects.

[0010] Another object of the present invention is to provide a method for preventing, treating and / or improving inflammation and blood circulation disorders by administering the above complex fermented extract to a subject.

[0011] Other objects and advantages of the present invention will become more apparent from the detailed description, claims and drawings below.

[0012] According to one aspect of the present invention, the present invention provides a pharmaceutical composition for treating or preventing inflammatory diseases and blood circulation disorders, which contains a mineral complex fermentation extract as an active ingredient.

[0013] In the present invention, the mineral complex fermentation extract is a mineral complex fermentation extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling the fermented extract.

[0014] The present inventors have explored the effects of various natural substances in order to find a substance that simultaneously exhibits excellent effects in improving inflammation and blood circulation disorders, and have discovered that when two types of components, biotite and bentonite, are combined and fermented with a mixed strain of Bacillus toyonensis CNL GAMMA_27 and Bacillus simplex CNL GAMMA_52 to produce a complex fermented extract, the synergistic effect in improving inflammation and blood circulation disorders increases dramatically, and it has also been confirmed that there are no safety issues when applied to the human body.

[0015] The "biotite" of the present invention refers to a mica mineral belonging to the monoclinic system with a crystal structure belonging to the octahedral subfamily, and its color may be black, brownish black, or greenish black. In addition, "bentonite" refers to a light-colored porous rock composed of a clay mineral group of the montmorillonite group and colloidal silica. Bentonite is a colloidal hydrous aluminum silicate that occurs naturally.

[0016] The present inventors confirmed the synergistic effect of the composite extract according to the combination of biotite and bentonite on improving inflammation and blood circulation disorder, and in the process of conducting in-depth fermentation research to further increase this, they comprehensively investigated the characteristics of strains optimized for fermentation and confirmed that the strains with the most optimized efficiency for fermentation of a mixture of biotite and bentonite are Bacillus toyonensis and Bacillus simplex strains, and named them "Bacillus toyonensis CNL GAMMA_27" strain and "Bacillus simplex CNL GAMMA_52" strain, and deposited them with the Biological Resource Center of the Korea Research Institute of Bioscience and Biotechnology on May 10, 2023, and were assigned the deposit numbers KCTC 15435BP and KCTC 15437BP. In the present invention, 'Bacillus toyonensis CNL GAMMA_27 strain' or 'CNL GAMMA_27 strain' is used with the same meaning as Bacillus toyonensis (Accession No.: KCTC 15435BP) strain. In addition, 'Bacillus simplex CNL GAMMA_52 strain' or 'CNL GAMMA_52 strain' is used with the same meaning as Bacillus simplex (Accession No.: KCTC 15437BP) strain.

[0017] The term 'fermented extract' used in the present invention refers to an extract obtained by inoculating a medium in which Bacillus toyonensis CNL GAMMA_27 and Bacillus simplex CNL GAMMA_52 strains can grow and obtaining a culture result. In the present invention, it refers to a culture supernatant obtained from a culture obtained by inoculating a Bacillus toyonensis strain into a mixture of biotite and bentonite or a distilled extract thereof, a dried product of the culture supernatant, or a distilled extract thereof.

[0018] The above fermentation may be carried out in a solid-state fermentation manner, in which the Bacillus toyonensis CNL GAMMA_27 and Bacillus simplex CNL GAMMA_52 strains are inoculated into a mixture of powdered biotite and bentonite, and then solid-state culture is performed at a temperature of 25 to 55°C, 25 to 50°C, 25 to 48°C, 25 to 45°C, 25 to 44°C, or 25 to 43°C for 12 to 48 hours, 12 to 36 hours, 12 to 24 hours, 18 to 48 hours, 18 to 42 hours, 18 to 36 hours, 18 to 30 hours, or 18 to 24 hours to produce a fermented product. More preferably, the solid-state culture may be performed at a temperature range of 25 to 45°C for 12 to 36 hours. The above solid-phase culturing step may be culturing under anaerobic conditions, preferably under facultative anaerobic conditions.

[0019] In addition, the fermentation may be carried out in a liquid phase fermentation manner, in which the Bacillus toyonensis CNL GAMMA_27 and Bacillus simplex CNL GAMMA_52 strains are inoculated into the distilled extract of biotite and bentonite, and then the fermented product is produced by culturing in a liquid phase at a temperature of 25 to 55°C, 25 to 50°C, 25 to 48°C, 25 to 45°C, 25 to 44°C, or 25 to 43°C for 12 to 48 hours, 12 to 36 hours, 12 to 24 hours, 18 to 48 hours, 18 to 42 hours, 18 to 36 hours, 18 to 30 hours, or 18 to 24 hours. More preferably, the liquid culture may be performed at a temperature range of 25 to 45°C for 12 to 36 hours. The liquid culture step may be performed under anaerobic conditions, preferably under facultative anaerobic conditions.

[0020] The term "extract" used in the present invention includes all formulations that can be formed using the extract, such as the extract obtained by extracting a mineral or minerals, or extracting a mineral or mineral fermentation product, a distillate, dilution, or concentrate of the extract, a dried product obtained by drying the extract, a conditioned or purified product of the extract, or a mixture thereof. The extract of the present invention can preferably be prepared and used in the form of a diluted solution after extraction.

[0021] In the present invention, the extraction method for extracting biotite or bentonite, or a fermented product thereof, is not particularly limited, and extraction can be performed according to a method commonly used in the relevant technical field. Non-limiting examples of the extraction method include distillation extraction, hot water extraction, ultrasonic extraction, filtration, and reflux extraction, and these may be performed alone or in combination of two or more extraction methods.

[0022] In the present invention, the type of extraction solvent used to extract the biotite or bentonite, or the fermented product thereof, is not particularly limited, and any solvent known in the art can be used. Non-limiting examples of the extraction solvent include water; lower alcohols having 1 to 4 carbon atoms, such as methanol, ethanol, propyl alcohol, and butyl alcohol; polyhydric alcohols, such as glycerin, butylene glycol, and propylene glycol; and hydrocarbon solvents, such as methyl acetate, ethyl acetate, acetone, benzene, hexane, diethyl ether, and dichloromethane; or mixtures thereof. Preferably, water, lower alcohols, 1,3-butylene glycol, and ethyl acetate can be used alone or in combination of two or more.

[0023] In the pharmaceutical composition for treating or preventing inflammatory diseases and blood circulation disorders of the present invention, the combination of biotite and bentonite is not limited thereto, but biotite and bentonite may be mixed in a weight ratio of 4:6 to 6:4 based on the raw material components. In a preferred embodiment of the present invention, it has been confirmed that a composite extract of two types of mineral components mixed in the range of the above raw material mixing ratio or a composite fermented extract thereof exhibits excellent anti-inflammatory and blood circulation improving effects.

[0024] As used herein, the term "inflammation" refers to a protective response to a harmful stimulus, encompassing the period from the stimulus' onset to the repair process. Specifically, it encompasses vasodilation and increased vascular permeability, leukocyte migration and adhesion, granulation, and angiogenesis. Local symptoms include redness, swelling, fever, pain, and signs of functional impairment.

[0025] In addition, "inflammatory disease" is a general term for diseases with inflammation as the main lesion, and in the present invention, the inflammatory disease is characterized as an inflammatory disease mediated by increased expression of nitric oxide (NO) or iNOS and COX-2 genes. Preferably, the inflammatory disease is atopic dermatitis, psoriasis, sinusitis, rhinitis, conjunctivitis, asthma, dermatitis, inflammatory collagen vascular disease, glomerulonephritis, encephalitis, inflammatory enteritis, chronic obstructive pulmonary disease, sepsis, septic shock, pulmonary fibrosis, undifferentiated spondyloarthropathy, undifferentiated arthropathy, arthritis, inflammatory osteolysis, chronic inflammatory disease caused by viral or bacterial infection, colitis, ulcerative colitis, inflammatory bowel disease, arthritis, rheumatoid arthritis, reactive arthritis, osteoarthritis, atherosclerosis, myocarditis, endocarditis, pericarditis, cystic fibrosis, Hashimoto's thyroiditis, Graves' disease, lupus, chilblain lupus, tuberculous lupus, lupus nephritis, systemic lupus erythematosus, macular degeneration, uveitis, irritable bowel syndrome, Crohn's disease, Sjogren's syndrome, fibromyalgia, chronic fatigue syndrome, Examples include chronic fatigue immunodeficiency syndrome, myalgic encephalomyelitis, amyotrophic lateral sclerosis, Parkinson's disease, Alzheimer's disease, and multiple sclerosis.

[0026] Additionally, the term "anti-inflammatory" is used to mean suppressing the "inflammatory" response or treating or preventing an "inflammatory disease".

[0027] In addition, the term "blood circulation disorder" in the present invention refers to a disease that occurs when blood does not flow smoothly throughout the body. When blood circulation in the arteries such as the brain, eyes, and peripheral blood vessels is disturbed, symptoms such as tinnitus, dizziness, numbness in the hands and feet, and cold sensation appear, and when blood circulation in the veins is disturbed, symptoms such as leg swelling, varicose veins, and hemorrhoids appear. In addition, "blood circulation agent" or "blood circulation improving agent" refers to a drug that allows blood to flow smoothly through the blood vessels of the body. The "blood circulation disorder disease" of the present invention may be a blood circulation disorder such as decreased alertness, dizziness, tinnitus, headache, visual impairment, memory loss, anxiety, depression, limb dystrophy, edema, and numbness, and further includes thrombosis-related diseases such as angina pectoris, acute myocardial infarction, or stroke caused by the formation of blood clots. Specifically, the blood circulation disorder disease may be one or more selected from the group consisting of tinnitus, edema, limb dystrophy, angina pectoris, cerebral infarction, stroke, hyperlipidemia, hypertension, diabetes, arteriosclerosis, and myocardial infarction, but is not limited thereto.

[0028] In the pharmaceutical composition for treating or preventing inflammatory diseases and blood circulation disorders of the present invention, the mineral complex fermented extract is characterized by exhibiting anti-inflammatory activity by inhibiting the production of nitric oxide and inhibiting the expression of iNOS and COX-2 genes, thereby being effectively used for the prevention, treatment and improvement of inflammatory reactions or inflammatory diseases. In a preferred embodiment of the present invention, the nitric oxide production inhibition activity of a single mineral extract of biotite or bentonite was minimal, but it was confirmed that a synergistic effect in inhibiting nitric oxide production occurred depending on the combination of these two components, and it was confirmed that the nitric oxide production inhibition effect was further improved through fermentation of Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) (see Fig. 2). In addition, it was confirmed that two types of mineral fermented extracts fermented with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) more effectively suppressed the expression of iNOS and COX-2 genes, which are inflammatory factors, compared to two types of mineral extracts of biotite and bentonite (see Figs. 3 and 4).

[0029] In the pharmaceutical composition for treating or preventing inflammatory diseases and blood circulation disorders of the present invention, the mineral complex fermented extract is characterized by promoting angiogenesis and blood circulation by promoting the migration of vascular endothelial cells, and inhibiting platelet aggregation by suppressing the production of thromboxane B2, thereby being effectively used for the prevention, treatment, and improvement of blood circulation disorders or diseases related to thrombosis. In a preferred embodiment of the present invention, it was confirmed that the two-mineral complex distilled extract can further promote the migration of human umbilical vein endothelial cells (HUVEC) compared to a single mineral component extract, and it was confirmed that the mineral complex fermented extract of the present invention has a more excellent cell migration promoting effect than the two-mineral complex distilled extract (see Fig. 5). In addition, the thromboxane B2 production inhibition effect of the two-mineral complex distillation extract did not significantly increase compared to the single mineral distillation extract, but it was confirmed that the thromboxane B2 production inhibition effect of the mineral complex fermentation extract of the present invention manufactured through the fermentation process of the two strains significantly increased (see Fig. 6).

[0030] The pharmaceutical composition of the present invention comprises a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier included in the pharmaceutical composition of the present invention is one commonly used in formulation, and includes, but is not limited to, lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia gum, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. In addition to the above components, the pharmaceutical composition of the present invention may further include a lubricant, a wetting agent, a sweetening agent, a flavoring agent, an emulsifier, a suspending agent, a preservative, etc., which may be appropriately changed and applied as needed.

[0031] The pharmaceutical composition of the present invention can be administered orally or parenterally, and in the case of parenteral administration, it can be administered by intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, transdermal administration, etc.

[0032] The appropriate dosage of the pharmaceutical composition of the present invention may vary depending on factors such as the formulation method, administration method, patient age, weight, sex, pathological condition, food, administration time, administration route, excretion rate, and response sensitivity. The daily dosage of the pharmaceutical composition of the present invention is, for example, 0.001-100 mg / kg.

[0033] The pharmaceutical composition of the present invention can be manufactured in the form of a unit dose by formulating it using a pharmaceutically acceptable carrier and / or excipient according to a method that can be easily performed by a person having ordinary skill in the art to which the present invention pertains, or can be manufactured by placing it in a multi-dose container. In this case, the formulation may be in the form of a solution, suspension, syrup or emulsion in an oil or aqueous medium, or in the form of an extract, powder, granule, tablet or capsule, and may additionally include a dispersing agent or stabilizer.

[0034] According to another aspect of the present invention, the present invention provides a pharmaceutical composition for improving inflammatory diseases and blood circulation disorders, which contains a mineral complex fermented extract as an active ingredient. The mineral complex fermented extract is a mineral complex fermented extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling the fermented extract. The mineral complex fermented extract and its effect of improving inflammatory diseases and blood circulation disorders are as described in the pharmaceutical composition.

[0035] The term "quasi-drug" used in the present invention refers to products that are used for the purpose of diagnosing, treating, improving, alleviating, managing or preventing diseases of humans or animals, and have a milder effect than pharmaceutical products. For example, according to the Pharmaceutical Affairs Act, quasi-drugs are products other than those used for pharmaceutical purposes, and include products used for the treatment or prevention of diseases of humans or animals, products that have a mild effect on the human body or do not directly act on the human body, etc. The quasi-drug composition of the present invention is used for the purpose of improving inflammatory diseases and blood circulation disorder diseases, and there is no particular limitation on its formulation, but it may preferably be a formulation for oral administration, skin administration or transdermal administration.

[0036] Additionally, for each formulation, the quasi-drug composition may optionally select and combine other ingredients according to the formulation or intended use of the other quasi-drug. The amount of active ingredients mixed may be appropriately determined depending on the intended use (inhibition or relief). For example, it may include conventional excipients such as thickeners, stabilizers, solubilizers, vitamins, pigments, and fragrances, and carriers.

[0037] According to another aspect of the present invention, the present invention provides a food composition for improving inflammatory diseases and blood circulation disorders, which contains a mineral complex fermented extract as an active ingredient. The mineral complex fermented extract is a mineral complex fermented extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling the fermented extract. The mineral complex fermented extract and its effect of improving inflammatory diseases and blood circulation disorders are as described in the pharmaceutical composition. The food composition may be used in the form of a health functional food, but is not limited thereto.

[0038] The food composition of the present invention may be comprised of a fraction or processed product of the mineral complex fermentation extract. Furthermore, the composition may include, in addition to the active ingredient, a food additive acceptable from a scientific perspective.

[0039] In the present invention, "food auxiliary additive" means a component that can be added to food as an auxiliary, and can be appropriately selected and used by those skilled in the art as added in the manufacture of health functional foods of each formulation. Examples of food auxiliary additives include various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavoring agents and natural flavoring agents, coloring agents and fillers, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, etc., but the types of food auxiliary additives of the present invention are not limited by the above examples.

[0040] The food composition of the present invention may include a health functional food. In the present invention, "health functional food" refers to a food manufactured and processed in the form of tablets, capsules, powders, granules, liquids, pills, etc. using raw materials or ingredients having functional properties useful to the human body. Here, "functionality" means obtaining a beneficial effect for health purposes, such as regulating nutrients or physiological effects on the structure and function of the human body, and in the present invention, it means anti-inflammation, increasing angiogenesis, inhibiting thrombosis, and improving blood circulation. The health functional food of the present invention can be manufactured by a method commonly used in the art, and during the manufacturing process, it can be manufactured by adding raw materials and ingredients commonly added in the art.

[0041] In addition, the formulation of the health functional food can be manufactured without limitation as long as it is a formulation recognized as a health functional food. The food composition of the present invention can be manufactured in various forms of formulation, and unlike general drugs, it has the advantage of not having side effects that may occur with long-term use of drugs, etc., as it is made from edible natural extracts as raw materials, and it is highly portable, and can be taken as a supplement to enhance the effect of improving diseases caused by inflammatory diseases, thrombosis, or blood circulation disorders.

[0042] There is no limitation on the form that the health functional food of the present invention can take, and it can include all foods in the conventional sense, and can be used interchangeably with terms known in the art such as functional food. In addition, the health functional food of the present invention can be manufactured by mixing other appropriate auxiliary ingredients that can be included in foods and known additives according to the choice of a person skilled in the art. Examples of foods that can be added include dairy products including meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, ice cream, various soups, beverages, tea, drinks, alcoholic beverages, and vitamin complexes, and it can be manufactured by adding it to juice, tea, jelly, and juice manufactured using the mineral complex fermentation extract of biotite and bentonite according to the present invention as a main ingredient. In addition, foods used as feed for animals are also included.

[0043] According to another aspect of the present invention, the present invention provides an anti-inflammatory cosmetic composition containing a mineral complex fermented extract as an active ingredient. The mineral complex fermented extract is a mineral complex fermented extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling the fermented extract. The mineral complex fermented extract and its anti-inflammatory effect are as described in the pharmaceutical composition.

[0044] The cosmetic composition of the present invention includes, in addition to the mineral complex fermentation extract of biotite and bentonite as an active ingredient, ingredients commonly used in cosmetic compositions, such as conventional auxiliary agents such as antioxidants, stabilizers, solubilizers, vitamins, pigments, and fragrances, and carriers.

[0045] The cosmetic composition of the present invention can be manufactured into any formulation commonly manufactured in the art, and for example, can be formulated into a solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing, oil, powder foundation, emulsion foundation, wax foundation, and spray, but is not limited thereto. More specifically, it can be manufactured into the formulation of a flexible toner (skin), a nourishing toner (milk lotion), a nourishing cream, a massage cream, an essential oil, an eye cream, a cleansing cream, a cleansing foam, a cleansing water, a pack, a spray, or a powder.

[0046] When the formulation of the present invention is a paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide may be used as a carrier component.

[0047] When the formulation of the present invention is a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component, and particularly in the case of a spray, a propellant such as chlorofluorohydrocarbon, propane / butane or dimethyl ether may be additionally included.

[0048] When the formulation of the present invention is a solution or emulsion, a solvent, solubilizer or emulsifier is used as a carrier component, and examples thereof include water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butyl glycol oil, glycerol aliphatic ester, polyethylene glycol or fatty acid ester of sorbitan.

[0049] When the formulation of the present invention is a suspension, a liquid diluent such as water, ethanol or propylene glycol, a suspending agent such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar or tragacanth, etc. can be used as a carrier component.

[0050] When the formulation of the present invention is a surfactant-containing cleansing agent, a fatty alcohol sulfate, a fatty alcohol ether sulfate, a sulfosuccinic acid monoester, an isethionate, an imidazolinium derivative, a methyl taurate, a sarcosinate, a fatty acid amide ether sulfate, an alkyl amidobetaine, a fatty alcohol, a fatty acid glyceride, a fatty acid diethanolamide, a vegetable oil, a lanolin derivative, or an ethoxylated glycerol fatty acid ester may be used as a carrier component.

[0051] Specific examples of the cosmetics of the present invention include hand sanitizers, facial creams, facial foams, cleansing creams, cleansing milk, cleansing lotions, massage creams, cold creams, moisturizing creams, emulsions, toners, packs, aftersaving creams, anti-sunburn creams, sunburn oils, soaps, body shampoos, hair shampoos, hair rinses, hair treatments, hair conditioners, hair growth conditioners, hair creams, hair liquids, set lotions, hair sprays, hair brushes, color rinses, color sprays, permanent wave liquids, press powders, loose powders, eye shadows, hand creams, and lipsticks.

[0052] According to another aspect of the present invention, a method for treating inflammatory diseases and blood circulation disorders is provided, comprising a step of administering to a subject a composition comprising a mineral complex fermentation extract, wherein the mineral complex fermentation extract is a mineral complex fermentation extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling and extracting the same.

[0053] The combination of the above biotite and bentonite is not limited thereto, but biotite and bentonite may be mixed in a weight ratio of 4:6 to 6:4 based on the raw material components.

[0054] In addition, the above-mentioned blood circulation disorder disease may be one or more selected from the group consisting of tinnitus, edema, limb dystrophy, angina pectoris, cerebral infarction, stroke, hyperlipidemia, hypertension, diabetes, arteriosclerosis, and myocardial infarction, but is not limited thereto.

[0055] In the present invention, the "subject" includes an animal or human having an inflammatory disease and / or a blood circulation disorder whose symptoms can be improved by administration of a composition containing a mineral complex fermentation extract according to the present invention.

[0056] The term "administration" in the present invention refers to introducing a given substance into a human or animal by any suitable method. The composition according to the present invention may be administered orally or parenterally via any common route as long as it can reach the target tissue. Furthermore, the composition according to the present invention may be administered by any device capable of transporting the active ingredient to target cells. The preferred dosage of the composition according to the present invention varies depending on the patient's condition and weight, the severity of the disease, the drug form, the route of administration, and the duration of administration, but can be appropriately selected by those skilled in the art.

[0057] According to another aspect of the present invention, a mineral complex fermented extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling the fermented extract is provided for use in treating, preventing or improving inflammatory diseases and blood circulation disorders.

[0058] The combination of the above biotite and bentonite is not limited thereto, but biotite and bentonite may be mixed in a weight ratio of 4:6 to 6:4 based on the raw material components.

[0059] In addition, the above-mentioned blood circulation disorder disease may be one or more selected from the group consisting of tinnitus, edema, limb dystrophy, angina pectoris, cerebral infarction, stroke, hyperlipidemia, hypertension, diabetes, arteriosclerosis, and myocardial infarction, but is not limited thereto.

[0060] According to another aspect of the present invention, there is provided a use for producing a composition for treating, preventing or improving inflammatory diseases and blood circulation disorders, characterized in that the mineral complex fermented extract comprises a mineral complex fermented extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling the fermented extract.

[0061] The combination of the above biotite and bentonite is not limited thereto, but biotite and bentonite may be mixed in a weight ratio of 4:6 to 6:4 based on the raw material components.

[0062] In addition, the above-mentioned blood circulation disorder disease may be one or more selected from the group consisting of tinnitus, edema, limb dystrophy, angina pectoris, cerebral infarction, stroke, hyperlipidemia, hypertension, diabetes, arteriosclerosis, and myocardial infarction, but is not limited thereto.

[0063] The composition for treating, preventing or improving the above inflammatory disease and blood circulation disorder disease may be a pharmaceutical composition or a food composition, but is not limited thereto.

[0064] The features and advantages of the present invention are summarized as follows:

[0065] (a) The present invention provides a composition for treating, preventing or improving inflammatory diseases and blood circulation disorders, which contains a mineral complex fermentation extract of biotite and bentonite as an active ingredient.

[0066] (b) The mineral complex fermented extract of the present invention is non-cytotoxic, inhibits the production of nitric oxide (NO) involved in the occurrence of inflammation, effectively inhibits the expression of iNOS and COX-2 genes, increases the mobility of vascular endothelial cells to promote angiogenesis, and inhibits platelet aggregation in a concentration-dependent manner by inhibiting thromboxane B2 production, and can be usefully used for the purposes of drugs, quasi-drugs, health functional foods, cosmetics, etc. for the treatment, prevention, or improvement of inflammatory diseases and blood circulation disorders.

[0067] (c) In addition, the mineral complex fermentation extract of the present invention is not only very safe for the human body, but also has excellent stability.

[0068] Figure 1 is a graph showing the change in cell viability measured after treating RAW264.7 cells with biotite extract (Preparation Example 1), bentonite extract (Preparation Example 2), two types of mineral complex extract (Preparation Example 3), and mineral complex fermentation extract (Preparation Example 4) at different concentrations.

[0069] Figure 2 is a graph showing the change in nitric oxide (NO) production after treating RAW 264.7 cells with biotite extract (Preparation Example 1), bentonite extract (Preparation Example 2), two-mineral complex extract (Preparation Example 3), and mineral complex fermentation extract (Preparation Example 4).

[0070] Figure 3 is a graph showing the change in expression of the COX-2 gene after treating RAW 264.7 cells with two types of mineral complex extract (Preparation Example 3) and mineral complex fermentation extract (Preparation Example 4).

[0071] Figure 4 is a graph showing changes in the expression of the iNOS gene after treating RAW 264.7 cells with two types of mineral complex extract (Preparation Example 3) and mineral complex fermentation extract (Preparation Example 4).

[0072] Figure 5 is a graph showing the migration of human umbilical vein endothelial cells (HUVEC) measured according to treatment with biotite extract (Preparation Example 1), bentonite extract (Preparation Example 2), two-mineral complex extract (Preparation Example 3), and mineral complex fermentation extract (Preparation Example 4).

[0073] Figure 6 is a graph showing the amount of thromboxane B2 produced by measuring the amount of biotite extract (Preparation Example 1), bentonite extract (Preparation Example 2), two-mineral complex extract (Preparation Example 3), and mineral complex fermentation extract (Preparation Example 4).

[0074] Hereinafter, the present invention will be described in more detail through examples. These examples are intended merely to illustrate the present invention, and therefore, the scope of the present invention is not to be construed as being limited by these examples.

[0075] Example 1. Preparation of mineral complex fermentation extract

[0076] 1-1. Preparation of biotite distillation extract

[0077] 0.5 kg of biotite was washed with purified water, dried, pulverized, and finely ground to obtain a powder. 10 times the weight of water was added to the powdered biotite, heated to 100°C, and then cooled by collecting water vapor to obtain a distillation extract (Preparation Example 1).

[0078] 1-2. Preparation of bentonite distillation extract

[0079] 0.5 kg of bentonite was washed with purified water, dried, crushed, and finely ground to obtain a powder. 10 times the weight of water was added to the powdered bentonite, heated to 100°C, and then cooled by collecting the water vapor to obtain a distillation extract (Preparation Example 2).

[0080] 1-3. Preparation of mineral complex extract

[0081] 0.25 kg of biotite and 0.25 kg of bentonite were washed with purified water, dried, pulverized, and finely ground to obtain a powder. 10 times the weight of water was added to the powdered biotite and bentonite, heated to 100°C, and then cooled by collecting the water vapor to obtain a mineral complex distillation extract (Preparation Example 3).

[0082] 1-4. Manufacturing of mineral complex fermented extract (Minexin)

[0083] 0.25 kg of biotite and 0.25 kg of bentonite were washed with purified water, dried, pulverized, and finely ground to obtain powder. The powdered raw materials were inoculated with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP), and then cultured on solid under facultative anaerobic conditions.

[0084] The culture was sterilized at 121°C for 15 minutes to inactivate the strain, then 10 times the weight of water was added, heated to 100°C, and then the steam was collected and cooled to produce a mineral complex fermentation extract (Minexin) (Preparation Example 4).

[0085] Example 2. Cell protective activity

[0086] The cytoprotective effect of the mineral complex fermentation extract prepared in Example 1 above was confirmed through the following method through the effect on the growth of RAW 264.7 cells, a macrophage cell line.

[0087] RAW 264.7 cells, a macrophage cell line, were obtained from the Korea Cell Line Bank (KTCC, Seoul, Korea) and used. The cells were seeded in a medium containing 10% FBS (Fetal bovine serum), DMEM (Dulbecco's modified Eagle's medium), 10% FBS, 100 ㎍ / ㎖ penicillin, and 100 ㎍ / ㎖ streptomycin, and cultured at 37°C and 5% CO2.

[0088] The above RAW 264.7 cells were 1Х10 6 Cells / ㎖ concentration (in DMEM) was inoculated into a 96-well plate and cultured for 24 hours. Then, samples prepared in Example 1 (preparation example 1), bentonite extract (preparation example 2), two-mineral complex extract (preparation example 3), and mineral complex fermentation extract (preparation example 4) were diluted to concentrations of 1, 5, and 20% (v / v), respectively, and a new medium containing LPS (1 μg / ㎖), known as endotoxin, were simultaneously treated and cultured for 24 hours.

[0089] Afterwards, 1 mg / ㎖ of MTT reagent was treated, and after 2 hours, the formazan produced by MTT treatment in the cells was dissolved in DMSO solution and the absorbance was measured at 570 nm. For comparative experiments, 10% (v / v) dexamethasone (Dex) was treated and used as a positive control.

[0090] As a result of the experiment, it was shown that the mineral complex fermentation extract had a more excellent cell viability increasing effect than the mineral distillation extract. The mineral complex fermentation extract of the present invention (Preparation Example 4), which was fermented by inoculating two strains, showed improved cell viability of 110%, 117%, and 127% (based on the negative control group treated with 1 μg / ㎖ of LPS) at concentrations of 1, 5, and 20% (v / v), respectively, confirming that the cell protection effect on RAW 264.7 cells was more excellent than that of the mineral distillation extract before fermentation (see Fig. 1).

[0091] Example 3. Anti-inflammatory activity

[0092] 3-1. Inhibition of nitric oxide production

[0093] In order to confirm the anti-inflammatory effect of the mineral complex fermentation extract prepared in Example 1, the inhibition rate of nitric oxide (NO), a representative cytotoxic substance involved in inducing inflammation, was measured.

[0094] First, RAW 264.7 cells, a macrophage cell line, were obtained from the Korea Cell Line Bank (KTCC, Seoul, Korea) and used. The cells were seeded in a medium containing 10% FBS (Fetal bovine serum), DMEM (Dulbecco's modified Eagle's medium), 10% FBS, 100 ㎍ / ㎖ penicillin, and 100 ㎍ / ㎖ streptomycin, and cultured at 37°C and 5% CO2.

[0095] Specifically, the RAW 264.7 cells were treated with 1Х10 6 After inoculating a 96-well plate at a concentration of 10 cells / ㎖ (in DMEM) and culturing for 24 hours, the biotite single extract (Preparation Example 1), bentonite single extract (Preparation Example 2), two-mineral complex extract (Preparation Example 3), and mineral complex fermentation extract (Preparation Example 4) prepared in Example 1 were diluted to concentrations of 1, 5, and 20% (v / v), respectively, and a new medium containing LPS (1 μg / ㎖), known as endotoxin, was simultaneously treated, and cultured for 24 hours. Afterwards, 100 ㎕ of cell culture supernatant and 100 ㎕ of Griess reagent [1% (w / v) sulfanilamide, 0.1% (w / v) naphtylethylenediamine in 2.5% (v / v) phosphoric acid] were mixed and reacted in a 96-well plate for 10 minutes, and the amount of nitric oxide produced was measured by measuring the absorbance at 540 nm using an ELISA reader.

[0096] The concentration of nitrite produced was calculated using a standard curve of sodium nitrite dissolved in DMEM medium. The inhibitory activity of each sample against nitric oxide production was determined based on the difference in the amount of nitrite produced between the LPS-treated and LPS-untreated control groups. For comparative experiments, 10% (v / v) dexamethasone (Dex) was used as a positive control.

[0097] Referring to [Figure 2], which summarizes the experimental results, it was confirmed that the mineral complex fermentation extract of the present invention, which is produced by inoculating two strains and fermenting, has a superior effect in inhibiting nitrogen oxide production compared to the single or composite distillation extracts of biotite or bentonite at concentrations of 1, 5, and 20% (v / v).

[0098] Specifically, the positive control group treated with 10% (v / v) dexamethasone (Dex) showed a 43% inhibition of nitric oxide production compared to the negative control group treated with only LPS. In addition, compared to the negative control group treated with only LPS, the biotite single extract treatment group of Manufacturing Example 1 showed nitric oxide production inhibition activities of 11%, 19%, and 26% at concentrations of 1, 5, and 20% (v / v), respectively, the bentonite single extract treatment group of Manufacturing Example 2 showed nitric oxide production inhibition activities of 8%, 16%, and 19% at concentrations of 1, 5, and 20% (v / v), respectively, the biotite and bentonite two-composite extract treatment group of Manufacturing Example 3 showed nitric oxide production inhibition activities of 17%, 28%, and 44% at concentrations of 1, 5, and 20% (v / v), respectively, and the mineral complex fermentation extract treatment group of Manufacturing Example 4 showed nitric oxide production inhibition activities of 24%, 45%, and 24% at concentrations of 1, 5, and 20% (v / v), respectively. A 57% level of inhibition activity of nitric oxide production was observed.

[0099] Through the above results, it was confirmed that although the inhibition activity of single mineral extracts of biotite or bentonite on nitric oxide production was minimal, a synergistic effect on nitric oxide production inhibition occurred to some extent depending on the combination of these two components, and it was confirmed that the inhibition effect on nitric oxide production was further improved through fermentation with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP).

[0100] 3-2. Inhibition of the expression of inflammatory factors

[0101] The effect of mineral complex fermented extract on regulating the expression of iNOS and COX-2 genes, which are inflammatory factors, was verified.

[0102] First, RAW 264.7 cells were seeded in a 24-well cell culture dish at a density of 1X10 in DMEM (Dulbeccos modified Eagles medium), a dedicated medium containing 10% FBS (fetal bovine serum, Cambrex). 5 After inoculating at a cell concentration of 10 cells, the cells were cultured for 24 hours at 37°C in a humidified atmosphere of 5% CO2. Afterwards, the medium was removed and diluted with serum-free DMEM medium, and then samples diluted to 1, 5, and 20% (v / v) concentrations of two types of mineral complex extract (Preparation Example 3) and mineral complex fermentation extract (Preparation Example 4) and new medium containing LPS (1 μg / ml), known as endotoxin, were simultaneously treated and cultured for 24 hours.

[0103] The experiment used RT-PCR to determine the mRNA expression levels of iNOS and COX-2. The primers used are shown in Table 1 below:

[0104] Gene primer sequence (5'-3') Sequence number COX-2 Forward 5'-TGC TGG TGG AAA AAC CTC GT-3'1 Reverse 5'- AAA ACC CAC TTC GCC TCC AA-3'2 iNOS Forward 5'-AAT GGC AAC ATC AGG TCG GCC ATC ACT-3'3 Reverse 5'-GCT GTG TGT CAC AGA AGT CTC GAA CTC-3'4 GAPDH Forward 5'-TGA AGG TCG GTG TGA ACG GAT TTT GGC-3'5 Reverse 5'-TGG TTC ACA CCC ATC ACA AAC ATG G-3'6

[0105] RT-PCR was performed using the primers described above.

[0106] Referring to [Figure 3], which summarizes the results of COX-2 gene expression, compared to the negative control group treated with only LPS, the group treated with the two types of biotite and bentonite complex extracts of Manufacturing Example 3 showed 9% and 25% inhibition of COX-2 gene expression at 5% and 20% (v / v) concentrations, respectively, and the group treated with the mineral complex fermentation extract of Manufacturing Example 4 showed 35% and 60% inhibition of COX-2 gene expression at 5% and 20% (v / v) concentrations, respectively.

[0107] In addition, referring to [Figure 4] which summarizes the iNOS gene expression results, the group treated with the two types of biotite and bentonite complex extracts of Manufacturing Example 3 showed iNOS gene expression levels that were suppressed by 13% and 39% at concentrations of 5% and 20% (v / v), respectively, compared to the negative control group treated with only LPS, and the group treated with the mineral complex fermentation extract of Manufacturing Example 4 showed iNOS gene expression levels that were suppressed by 36% and 72% at concentrations of 5% and 20% (v / v), respectively.

[0108] Through the above results, it was confirmed that two types of mineral fermented extracts fermented with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) more effectively suppressed the expression of iNOS and COX-2 genes, which are inflammatory factors, compared to two types of mineral extracts of biotite or bentonite, and thus could be effectively used for the prevention, treatment, and improvement of inflammatory responses.

[0109] Example 4. Effect of promoting vascular endothelial cell migration (HUVECs migration)

[0110] The mobility of vascular endothelial cells, which is used as a general method for measuring angiogenesis, was measured and analyzed. The cell mobility analysis was performed using Boyden's chamber (Transwell). 600 ㎕ of serum-free M199 culture medium was added to each 24-well plate, and the biotite distillation extract (Preparation Example 1), bentonite distillation extract (Preparation Example 2), two-mineral complex extract (Preparation Example 3), and mineral complex fermentation extract (Preparation Example 4) prepared in Example 1 were diluted to concentrations of 1, 5, and 20% (v / v), respectively, and then gelatin (1 mg / ml) was spread on the lower side of the Transwell, and 2 x 10 4 Human umbilical vein endothelial cells (HUVEC) were attached at a cell concentration of 100 cells. For comparative experiments, 1 ng / ml of VEGF (vascular endothelial growth factor) was treated and used as a positive control.

[0111] After about 4 hours, the cells on the upper surface were removed using a cotton swab, and the cells that had moved to the lower surface were stained with hematoxylin and eosin, and the number of stained cells was counted and measured.

[0112] Referring to [Figure 5], which summarizes the experimental results, it was confirmed that the two-type complex mineral extract can promote the migration of human umbilical vein endothelial cells (HUVEC) more than the single mineral distillation extract, and the mineral complex fermentation extract was confirmed to have a superior cell migration promotion effect compared to the two-type mineral complex extract.

[0113] Specifically, in the biotite distillate extract (Preparation Example 1) treatment group, the migration rates of human umbilical vein endothelial cells were observed to increase by 110%, 125%, and 137%, respectively, compared to the untreated group at concentrations of 1, 5, and 20% (v / v), and in the bentonite distillate extract (Preparation Example 2) treatment group, the migration rates of human umbilical vein endothelial cells were observed to increase by 101%, 103%, and 108%, respectively, compared to the untreated group at concentrations of 1, 5, and 20% (v / v). In contrast, the group treated with the two-mineral complex extract (Preparation Example 3) showed a migration rate of human umbilical vein endothelial cells that was increased by 119%, 135%, and 161%, respectively, compared to the untreated group at concentrations of 1, 5, and 20% (v / v), and the group treated with the mineral complex fermented extract (Preparation Example 4) showed a migration rate of human umbilical vein endothelial cells that was increased by 137%, 180%, and 221%, respectively, compared to the untreated group at concentrations of 1, 5, and 20% (v / v).

[0114] Through the above results, it was confirmed that the angiogenesis-promoting activity of single mineral distillation extracts of biotite or bentonite was minimal, but that a certain synergistic effect occurred in promoting angiogenesis depending on the combination of these two mineral components, and that the angiogenesis-promoting effect was further improved through fermentation of Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP).

[0115] Example 5. Inhibitory activity against thromboxane B2 production

[0116] Platelets of experimental rats (4 Х 10 8 Platelet / mL) was diluted to concentrations of 1, 5, and 20% (v / v) each of the biotite distillation extract (Preparation Example 1), bentonite distillation extract (Preparation Example 2), two types of mineral complex extract (Preparation Example 3), and mineral complex fermentation extract (Preparation Example 4) prepared in Example 1, and then treated. After pre-adapting at 37°C for 2 minutes, platelet aggregation was induced by stimulation with collagen (10 μg / mL).

[0117] After the reaction, the reaction was stopped by adding a thromboxane A2 (TxA2) production inhibitor (5 mM EDTA, 0.2 mM indomethacin). The amount of thromboxane A2 produced was determined by measuring the amount of thromboxane B2 produced, and the amount of thromboxane B2 produced was measured using a thromboxane B2 EIA kit.

[0118] Referring to [Figure 6], which summarizes the experimental results, the inhibitory effect of the two-type complex mineral distillation extract on thromboxane B2 production was not significantly increased compared to the single mineral distillation extract, but the inhibitory effect of the mineral complex fermentation extract produced through the fermentation process of the two strains was confirmed to have significantly increased.

[0119] Specifically, the biotite distillate extract (Preparation Example 1) treatment group was observed to have thromboxane B2 production suppressed by 12%, 16%, and 24% compared to the collagen treatment group at concentrations of 1, 5, and 20% (v / v), respectively, the bentonite distillate extract (Preparation Example 2) treatment group was observed to have thromboxane B2 production suppressed by 2%, 9%, and 19% compared to the collagen treatment group at concentrations of 1, 5, and 20% (v / v), respectively, and the two-mineral complex extract (Preparation Example 3) treatment group was observed to have thromboxane B2 production suppressed by 12%, 26%, and 40% compared to the collagen treatment group at concentrations of 1, 5, and 20% (v / v), respectively. In contrast, in the mineral complex fermented extract (preparation example 4) treatment group, the production of thromboxane B2 was observed to be suppressed by 20%, 33%, and 50%, respectively, compared to the collagen treatment group at concentrations of 1, 5, and 20% (v / v).

[0120] Through the above results, it was confirmed that the platelet aggregation inhibitory activity of single or complex mineral extracts of biotite or bentonite was minimal, but the platelet aggregation inhibitory effect of the mineral complex extract was further improved through fermentation of Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP), and it was confirmed that it can be effectively used for the purpose of improving blood circulation by suppressing platelet aggregation in a concentration-dependent manner through inhibition of thromboxane B2 production.

[0121] In summary of the above experimental results, the mineral complex fermented extract of the present invention, which is obtained by fermenting a mixture of biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and then distilling and extracting it, i) is non-cytotoxic and promotes cell growth, ii) effectively inhibits the production of nitric oxide, an inflammatory mediator, and the expression of iNOS and COX-2 genes, iii) promotes angiogenesis by increasing the mobility of vascular endothelial cells, and iv) suppresses platelet aggregation in a concentration-dependent manner by suppressing thromboxane B2 production, so that it can be effectively used for the purpose of improving blood circulation. Therefore, it was confirmed that it can be usefully utilized in foods, pharmaceuticals, and quasi-drugs for simultaneously improving inflammation and blood circulation disorders.

[0122] While specific aspects of the present invention have been described in detail above, it should be apparent to those skilled in the art that these specific descriptions are merely preferred embodiments and do not limit the scope of the present invention. Therefore, the substantial scope of the present invention is defined by the appended claims and their equivalents.

[0123] [Accession number]

[0124] Name of depositor: Korea Research Institute of Bioscience and Biotechnology, Biological Resource Center (KCTC)

[0125] Accession number: KCTC15435BP

[0126] Date of acceptance: 20230510

[0127] Name of depositor: Korea Research Institute of Bioscience and Biotechnology, Biological Resource Center (KCTC)

[0128] Accession number: KCTC15437BP

[0129] Date of acceptance: 20230510

Claims

1. Contains mineral complex fermented extract as an effective ingredient, The above mineral complex fermentation extract is characterized by being a mineral complex fermentation extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling the fermented biotite and bentonite. A pharmaceutical composition for the treatment or prevention of inflammatory diseases and blood circulation disorders.

2. In paragraph 1, The above biotite and bentonite are characterized in that they are mixed in a weight ratio of 4:6 to 6:4 based on the raw material components. A pharmaceutical composition for the treatment or prevention of inflammatory diseases and blood circulation disorders.

3. In paragraph 1, A pharmaceutical composition for treating or preventing inflammatory diseases and blood circulation disorders, characterized in that the above mineral complex fermented extract exhibits anti-inflammatory activity by inhibiting the production of nitric oxide and inhibiting the expression of iNOS and COX-2 genes.

4. In paragraph 1, The above mineral complex fermented extract is characterized by promoting angiogenesis by promoting the movement of vascular endothelial cells and improving blood circulation by inhibiting platelet aggregation through inhibition of thromboxane B2 production. A pharmaceutical composition for the treatment or prevention of inflammatory diseases and blood circulation disorders.

5. A pharmaceutical composition for the treatment or prevention of inflammatory diseases and blood circulation disorders, wherein the blood circulation disorder disease in paragraph 1 is at least one disease selected from the group consisting of tinnitus, edema, limb dystrophy, angina pectoris, cerebral infarction, stroke, hyperlipidemia, hypertension, diabetes, arteriosclerosis, and myocardial infarction.

6. Contains mineral complex fermented extract as an effective ingredient, The above mineral complex fermentation extract is characterized by being a mineral complex fermentation extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling the fermented biotite and bentonite. A pharmaceutical composition for inhibiting thrombosis.

7. Contains mineral complex fermented extract as an effective ingredient, The above mineral complex fermentation extract is characterized by being a mineral complex fermentation extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling the fermented biotite and bentonite. A pharmaceutical composition for improving inflammatory diseases and blood circulation disorders.

8. Contains mineral complex fermented extract as an effective ingredient, The above mineral complex fermentation extract is characterized by being a mineral complex fermentation extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling the fermented biotite and bentonite. Food composition for improving inflammatory diseases and blood circulation disorders.

9. Contains mineral complex fermented extract as an effective ingredient, The above mineral complex fermentation extract is characterized by being a mineral complex fermentation extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling the fermented biotite and bentonite. Anti-inflammatory cosmetic composition.

10. A step of administering to a subject a composition containing a mineral complex fermentation extract, A method for treating inflammatory diseases and blood circulation disorders, characterized in that the above mineral complex fermented extract is a mineral complex fermented extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and then distilling the fermented extract.

11. In paragraph 10, A method for treating inflammatory diseases and blood circulation disorders, characterized in that the biotite and bentonite are mixed in a weight ratio of 4:6 to 6:4 based on the raw material components.

12. In clause 10, the blood circulation disorder disease is at least one disease selected from the group consisting of tinnitus, edema, limb dystrophy, angina pectoris, cerebral infarction, stroke, hyperlipidemia, hypertension, diabetes, arteriosclerosis, and myocardial infarction, a method for treating inflammatory diseases and blood circulation disorder diseases.

13. Use of a mineral complex fermented extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling the extract for the treatment, prevention or improvement of inflammatory diseases and blood circulation disorders.

14. In paragraph 13, The above biotite and bentonite are characterized by being mixed in a weight ratio of 4:6 to 6:4 based on the raw material components, and are used for the treatment, prevention or improvement of inflammatory diseases and blood circulation disorders.

15. In the 13th paragraph, the blood circulation disorder disease is at least one disease selected from the group consisting of tinnitus, edema, limb dystrophy, angina pectoris, cerebral infarction, stroke, hyperlipidemia, hypertension, diabetes, arteriosclerosis, and myocardial infarction, and is for use in the treatment, prevention, or improvement of inflammatory diseases and blood circulation disorder diseases.

16. For the purpose of manufacturing a composition for treating, preventing or improving inflammatory diseases and blood circulation disorders using a mineral complex fermentation extract, The above mineral complex fermented extract comprises a mineral complex fermented extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and Bacillus simplex CNL GAMMA_52 strain (Accession No. KCTC 15437BP) and distilling the extracted extract. The use of the composition for the treatment, prevention or improvement of inflammatory diseases and blood circulation disorders is characterized in that the above mineral complex fermented extract comprises a mineral complex fermented extract obtained by fermenting biotite and bentonite with Bacillus toyonensis CNL GAMMA_27 strain (Accession No. KCTC 15435BP) and distilling the extracted extract.

17. In paragraph 16, A composition for the manufacture of a composition for the treatment, prevention or improvement of inflammatory diseases and blood circulation disorders, characterized in that the biotite and bentonite are mixed in a weight ratio of 4:6 to 6:4 based on the raw material components.

18. In the 16th paragraph, the blood circulation disorder disease is at least one disease selected from the group consisting of tinnitus, edema, limb dystrophy, angina pectoris, cerebral infarction, stroke, hyperlipidemia, hypertension, diabetes, arteriosclerosis, and myocardial infarction, a composition for the manufacture of a treatment, prevention, or improvement of inflammatory diseases and blood circulation disorder diseases.

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