KR20220109664A - Bacillus licheniformis having the effect of preventing or improving for skin inflammation and uses thereof - Google Patents

Abstract

The present invention relates to: a Bacillus licheniformis strain having a skin inflammation inhibition or improvement effect; a microbial preparation for skin inflammation inhibition, improvement or treatment containing the strain or the culture medium thereof as an active ingredient; a probiotic bioactive composition containing the strain or the culture medium thereof as an active ingredient; a pharmaceutical composition containing the strain or the culture medium thereof as an active ingredient; a composition for food or healthy functional food containing the strain or the culture medium thereof as an active ingredient; a quasi-drug composition containing the strain or the culture medium thereof as an active ingredient; a drinking water additive containing the strain or the culture medium thereof as an active ingredient; and a feed additive and a cosmetic composition containing the strain or the culture medium thereof as an active ingredient.

Description

Bacillus licheniformis having the effect of preventing or improving skin inflammation and uses thereof

The present invention relates to Bacillus licheniformis having an anti-inflammatory effect on skin cells, and more particularly, a Bacillus licheniformis strain having an anti-inflammatory effect on skin cells induced by oxidative stress, the strain or A microbial agent having an anti-inflammatory effect on skin cells comprising the culture solution thereof as an active ingredient, a method for producing a microbial agent having an anti-inflammatory effect on skin cells comprising the step of culturing the strain, the strain or its culture medium is effective It relates to a probiotic probiotic active composition, pharmaceutical composition, quasi-drug composition, food (health functional food) composition, drinking water additive, feed additive, and cosmetic composition comprising as a component.

Excessive oxidative stress in the living body is known to cause various diseases such as decreased immune function, inflammation, cancer, arteriosclerosis, and cardiovascular disease, and accelerate aging of all organs. All living things, including humans, need oxygen to produce energy necessary to sustain life, and _the mitochondrial oxidoreductase system or immune cells exposed to foreign antigens ^, It generates various reactive oxygen species (ROS) such as hydrogen peroxide (H ₂ O ₂ ), singlet oxygen (·O ₂ ), and hydroxyl radical (·OH) (Kang KA, Chae SC, Kang DG, Kim). JS, Jin WH. 2005. Screening of antioxidative effect of herbal extracts on oxidative stress. Kor. J. Pharmacogn. 36: 159-163). The living organism maintains homeostasis through the generation and removal of these oxygen species to function as a cell. However, when this harmony is imbalanced, cells and the like are adversely affected by abnormally increased ROS, which is called oxidative stress.

Oxidative stress causes various diseases, and it is known that cancer, arteriosclerosis, or cardiovascular disease damage cells or tissues, participate in aging, and induce progressive memory loss and cognitive impairment (Kamat PK, Kalani A, Rai S, Swarnkar S, Tota S, Nath C, Tyagi N. 2016. Mechanism of Oxidative Stress and Synapse Dysfunction in the Pathogenesis of Alzheimer's Disease: Understanding the Therapeutics Strategies. Mol Neurobiol. 53: 648-661). That is, oxidative stress occurs due to the generation of excessive reactive oxygen species, and various diseases appear through oxidative damage to biological molecules such as proteins, lipids, and DNA. To counter this, there is a mechanism to prevent the accumulation of reactive oxygen species, which can be broadly divided into two categories. Antioxidant enzymes such as superoxide dismutase (SOD), catalse (CAT) and glutathione (GSH) and non-enzymatic free radical scavengers such as vitamin C, vitamic E, and flavonoids prevent the accumulation of reactive oxygen species and consequently (Rhee, S.-J. and Choi, J.-H. 2001. Effects of green tea catechin on the superoxide dismutase, glutathione peroxidase and xanthine oxidase activities of kidney in diabetic rats. Kor (Nutr. Soc. 34, 734-740).

In order to alleviate oxidative stress, research is being conducted on substances that inhibit active oxygen and lipid peroxidation reactions and substances that improve inflammatory responses. such as tannins and flavonoids. In addition, studies on antioxidant effects through natural substances and food materials such as red ginseng and green tea are being actively conducted in Korea.

Inflammation caused by oxidative stress is the expression of a normal and protective in vivo defense mechanism that appears locally against tissue damage caused by harmful chemicals or irritants in in vivo metabolites. Such inflammation is triggered by various chemical mediators produced from damaged tissues and migrating cells, and these chemical mediators are known to vary depending on the type of inflammatory process. In a normal case, the living body neutralizes or removes the onset factor through an inflammatory response and regenerates damaged tissue to restore normal structure and function, but otherwise, it may progress to a disease state such as chronic inflammation.

The most common agents for preventing or treating inflammatory diseases for treating these inflammatory diseases are largely divided into steroidal and non-steroidal inflammatory diseases preventing or treating agents, and most of these synthetic inflammatory diseases preventing or treating agents are various in addition to the main action. There are many disadvantages that are often accompanied by side effects. That is, in order to treat inflammatory diseases, an appropriate non-steroidal inflammatory disease prevention or treatment drug (NSAIDs) is used to relieve inflammation. I will do surgery. In this case, the NSAIDs used mainly inhibit cyclooxygenase (COX) to inhibit the production of prostaglandins involved in the inflammatory response, thereby preventing or treating inflammatory diseases, but gastrointestinal disorders and liver dysfunction. , there is a problem that it is difficult to use for a long period of time by causing side effects such as renal dysfunction.

Atopic dermatitis is a representative disease of skin inflammation, which is a skin inflammatory disease and is a chronic recurrent dermatitis that occurs mainly in infants and children, but recently it is known as a disease that occurs in adolescence or adults (Lehtonen, E.P., Holmberg-Marttkla, D. Kaila, M. 2003. Cumulative prevalence of atopic dermatitis and related skin symptoms in a well-babyclinic: aretrospective cohortstudy. Pediatr Allergy Immunol. 14: 405-408). Atopic dermatitis is chronic, dry, itchy, and recurs repeatedly, and symptoms such as erythema, edema, scab, and keratinization accompanied by exudation appear. It affects daily life such as disability, cough, and shortness of breath. The exact cause and pathogenesis of atopic dermatitis have not been fully elucidated, and it is known that factors such as genetic factors, chemical stimulation due to environmental pollution, and immunological factors are involved (David Boothe W, Tarbox JA). , Tarbox MB. 2017. Atopic dermatitis: Pathophysiology. Adv Exp Med Biol. 1027: 21-37).

Most effective steroids are used for the treatment of atopic dermatitis, but their use is very limited due to their side effects. . Therefore, various studies are being conducted to develop a therapeutic agent that can effectively and safely treat atopic dermatitis, especially natural substances using plants and food. Representatively, research on phenolic, polyphenol, flavone, flavonoid, and tanin among plant-derived substances has been actively conducted. It has also been reported that these substances relieve skin inflammation due to their antibacterial and anti-inflammatory actions. In addition to natural substances derived from plants, many studies have been conducted for the treatment of atopy using probiotics such as lactic acid bacteria, and clinically effective strains have been reported.

On the other hand, probiotics are, when consumed in an appropriate amount, alleviation of lactose intolerance, immune activity and regulation, alleviation of allergic reactions and inflammation, reduction of blood cholesterol, inhibitory effect on colorectal cancer, atopic dermatitis, Crohn's disease, diarrhea, Candida infection and Microorganisms that confer beneficial effects on the health of the host through reduction of clinical symptoms of urethral infection and competitive inhibition of pathogens (Cho, Jung-Whan, Kim, Dong-Hyeon, Kim, Hyun-Sook, Kim, Hong) -Seok, Hwang, Dae-Geun, Song, Kwang-Young, Yim, Jin-Hyuk, Choi, Dasom, Lim, Jong-Soo and Seo, Kun-Ho Seo. 2014. Effect of Probiotics on Risk Factors for Human Disease: A Review. Korean J. Dairy Sci. Technol. 32: 17-29).

Probiotics are microorganisms that have a useful action in the animal digestive tract, and have a useful action on the host animal, such as improving the intestinal flora. The strains used as probiotics include Lactobacillus, Enterococcus, Bifidobacterium, Bacillus, Clostridium, Saccharomyces, and Aspergillus genera. The mechanism of action of probiotics is normalization of the intestinal flora, production of antibiotics, nutrient competition to prevent the establishment of pathogens, production of organic acids by lowering pH, inhibition of harmful enzymes of pathogens such as β-glucuronidase, azoreductase, and nitroreductase, inhibition of the production of decay products and toxins, Digestive enzyme production, vitamin synthesis, immune stimulation, etc.

Early in Europe and Japan, strains such as L. rhumnosus GG (Valio, Finland) and L. casei Shirota (Yakult, Japan) were isolated and probiotics with excellent functions were developed. Research is actively underway. Similarly in Korea, research to isolate fermented foods such as kimchi and soy sauce, as well as animal and human-derived strains and use them as probiotics, is being actively conducted, but securing the strain is the most important key.

The present inventors completed the present invention by discovering the effect of Bacillus licheniformis inhibiting oxidative stress in vivo and preventing or improving skin inflammation while studying the physiologically active properties of probiotics.

The present invention was derived from the above needs, and the present inventors discovered for the first time that a Bacillus licheniformis strain relieves skin inflammation caused by oxidative stress, and the skin comprising the strain or its culture medium as an active ingredient A microbial agent for preventing or improving inflammation, a method for producing a microbial agent for preventing or improving skin inflammation comprising the step of culturing the strain, a probiotic probiotic active composition comprising the strain or a culture solution thereof as an active ingredient, The present invention was completed by confirming that it can be applied as a pharmaceutical composition, a composition for quasi-drugs, a composition for food or health functional food, an additive for drinking water, a composition for feed, and a cosmetic composition.

In order to solve the above problems, the present invention provides a Bacillus licheniformis strain having an effect of preventing or improving skin inflammation.

The Bacillus licheniformis strain was isolated from conventional soybean paste purchased from a public market in Gunsan-si, Jeollabuk-do. As a result of performing a 16S rDNA gene sequence test for strain identification, it was confirmed as Bacillus licheniformis (KCTC1918).

In order to achieve the object of the present invention, the present invention provides a Bacillus licheniformis strain having an effect of preventing or improving skin inflammation.

As used herein, the term 'oxidative stress' refers to superoxide radicals (·O ₂ ^- ), hydrogen peroxide (H ₂ O ₂ ), singlet oxygen (·O ₂ ), and hydroxyl generated during respiration and metabolism of living things. It means that cells are subjected to harmful effects by various reactive oxygen species (ROS) such as radical (·OH) or substances with high peroxidation power.

As used herein, the term 'inflammation' refers to the expression of a normal and protective in vivo defense mechanism that appears locally against tissue damage caused by harmful chemicals or irritants in in vivo metabolites.

As used herein, the term 'skin inflammation' refers to a phenomenon in which the inflammatory reaction is expressed in the skin, and may refer to atopic dermatitis, contact dermatitis, infectious dermatitis, and the like.

As used herein, the term “prevention” may refer to any action of inhibiting or delaying skin inflammation by administering the composition for preventing or improving skin inflammation according to the present invention to an individual.

As used herein, the term “improvement” may refer to any action that at least reduces a parameter related to the condition being treated, for example, the severity of symptoms.

The Bacillus licheniformis strain showed high antioxidant activity and oxidative stress inhibitory effect (Table 2, Table 3), and confirmed that it had a very strong anti-inflammatory effect in skin cells derived from atopic dermatitis-induced experimental animal models (Table 4, 1, 2) It was confirmed that the excellent bioactive effect, in particular, the possibility of use as a biological material for the prevention or improvement of skin inflammation.

In addition, the present invention provides a microbial preparation for preventing or improving skin inflammation comprising the strain or a culture solution thereof as an active ingredient.

The microbial preparation for preventing or improving skin inflammation of the present invention can be prepared using a Bacillus licheniformis strain as an active ingredient. The microbial agent for preventing or improving skin inflammation according to the present invention may be prepared as a solution, powder, suspension, dispersion, emulsion, oily dispersion, paste, dust, dispersion material or granule, but is not limited thereto.

In addition, the present invention provides a method for preparing a microbial agent for preventing or improving skin inflammation comprising the step of culturing the strain. The method of culturing the strain of the present invention may be cultured according to a method commonly used in the art.

In addition, the present invention provides a probiotic probiotic composition comprising the strain or a culture medium thereof as an active ingredient. The strain of the present invention exhibited a high thermal stability of about 77.1% (Table 5) and a survival rate of 39.1% or more in artificial gastric juice adjusted to pH 2.0 (Table 6) when exposed at 70 ° C for 10 minutes in a general characteristic test (Table 6), In resistance to bile acid, the survival rate was 41.5% or more in a 1.0% bile solution (Table 7), and it was confirmed that it had excellent properties as probiotic live bacteria.

In addition, the present invention provides a pharmaceutical composition for preventing or improving skin inflammation comprising the strain or a culture solution thereof as an active ingredient.

The pharmaceutical composition for preventing, improving or treating skin inflammation according to the present invention may further include a pharmaceutically acceptable carrier, and is formulated with the carrier to provide pharmaceuticals, food or health functional foods, feed additives, drinking water additives, and It may be provided as a cosmetic composition or the like.

As used herein, the term 'pharmaceutically acceptable carrier' may refer to a carrier or diluent that does not inhibit the biological activity and properties of the administered compound without irritating the organism.

The type of carrier usable in the present invention is not particularly limited, and any carrier commonly used in the art and pharmaceutically acceptable may be used. Non-limiting examples of the carrier include saline, sterile water, Ringer's solution, buffered saline, albumin injection solution, dextrose solution, maltodextrin solution, glycerol, ethanol, and the like. These may be used alone or in combination of two or more.

In addition, if necessary, other conventional additives such as antioxidants, buffers and/or bacteriostats may be added and used, and diluents, dispersants, surfactants, binders, lubricants, etc. It can be used by formulating it into a dosage form, pill, capsule, granule, or tablet.

The method of administering the pharmaceutical composition for preventing or treating skin inflammation according to the present invention is not particularly limited, and may follow a method commonly used in the art. As a non-limiting example of the administration method, the composition may be administered by oral administration or parenteral administration. The pharmaceutical composition for preventing or improving skin inflammation of the present invention may be prepared in various formulations according to the desired administration method.

The pharmaceutical composition of the present invention can be used as a single formulation, and can be prepared and used as a combination formulation by additionally including a drug known to have an approved skin inflammation treatment effect, and can be formulated using a pharmaceutically acceptable carrier or excipient. It may be prepared in unit dose form or by introduction into a multi-dose container.

As another aspect, the present invention provides a method for preventing or improving skin inflammation, comprising administering to an individual a pharmaceutical composition for the prevention or treatment of skin inflammation.

As used herein, the term "individual" may refer to any animal, including humans, that has or is likely to develop skin inflammation.

Specifically, the prevention or treatment method of the present invention may include administering the composition in a pharmaceutically effective amount to an individual who has or is at risk of developing skin inflammation.

As used herein, the term "pharmaceutically effective amount" means an amount sufficient to treat a disease at a reasonable benefit/risk ratio applicable to medical treatment, and generally the number of bacteria of the present invention per 1 g (or mL) When converted to, the amount of 7 log CFU to 10 log CFU, preferably 8 log CFU / g to 9 log CFU / g can be administered once a day divided into several times. However, for the purpose of the present invention, a suitable total daily amount of the composition containing the extract may be determined by the treating physician within the scope of correct medical judgment, and may be administered once or divided into several doses. However, for the purposes of the present invention, a specific therapeutically effective amount for a specific patient depends on the type and extent of the response to be achieved, the specific composition, including whether other agents are used, depending on the specific composition, the patient's age, weight, general health, It is preferable to apply differently depending on various factors including sex and diet, administration time, administration route and secretion rate of the composition, treatment period, drugs used together with or concurrently with a specific composition, and similar factors well known in the pharmaceutical field.

The pharmaceutical composition of the present invention may be administered as an individual therapeutic agent or may be administered in combination with other therapeutic agents, and may be administered sequentially or simultaneously with conventional therapeutic agents. and may be administered single or multiple. Taking all of the above factors into consideration, it is important to administer an amount that can obtain the maximum effect with a minimum amount without causing side effects, and can be easily determined by those skilled in the art.

As used herein, the term “administration” means introducing the pharmaceutical composition of the present invention to a patient by any suitable method, and the administration route of the composition of the present invention is oral or parenteral as long as it can reach the target tissue. It can be administered through various routes.

The method of administration of the pharmaceutical composition according to the present invention is not particularly limited, and may follow a method commonly used in the art. As a non-limiting example of the administration method, the composition may be administered by oral administration or parenteral administration. The pharmaceutical composition according to the present invention may be prepared in various dosage forms depending on the desired administration method.

The frequency of administration of the composition of the present invention is not particularly limited thereto, but may be administered once a day or administered several times in divided doses.

In addition, the present invention provides a composition for food or health functional food that can prevent or improve skin inflammation comprising the strain or a culture solution thereof as an active ingredient. Since ingestion of the strain of the present invention exhibits an anti-inflammatory effect in skin cells, it can be provided as a food for preventing or improving skin inflammation using this as an active ingredient. The health functional food for preventing or improving skin inflammation may be ingested for the prevention and treatment of dermatitis diseases including atopic dermatitis, allergic dermatitis, contact dermatitis, and the like.

When the strain or its culture solution obtained in the step of culturing the strain of the present invention is used as a food additive, the strain or its culture solution can be added as it is or used with other foods or food ingredients, and can be used appropriately according to a conventional method. can The mixing amount of the active ingredient may be appropriately determined depending on the purpose of its use (prophylactic, health or therapeutic treatment).

There is no particular limitation on the type of the food. Examples of foods to which the above substances can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gums, dairy products including ice cream, various soups, beverages, tea, drinks, Alcoholic beverages and vitamin complexes are available.

When the composition is used as a food additive, the composition may be added as it is or used together with other foods or food ingredients, and may be appropriately used according to a conventional method.

The food composition may include a food pharmaceutically acceptable carrier.

The food composition may be a health functional food. Functional food is the same term as food for special health use, FoSHU. , The food may be prepared in various forms such as tablets, capsules, powders, granules, liquids, pills, etc. in order to obtain a useful effect in the prevention or improvement of skin inflammation.

At this time, the content of the strain or its culture medium contained in the food is not particularly limited thereto, but may be included in an amount of 0.01 to 100% by weight, more preferably 1 to 80% by weight, based on the total weight of the food composition. When the food is a beverage, it may be included in a ratio of 1 to 30 g, preferably 3 to 20 g, based on 100 mL.

The health functional food can be prepared by a method commonly used in the art, and during the manufacture, it can be prepared by adding raw materials and components commonly added in the art. In addition, unlike general drugs, there is an advantage in that there are no side effects that may occur when taking the drug for a long time by using food as a raw material, and it can be excellent in portability.

According to another embodiment of the present invention, the present invention provides a quasi-drug composition for preventing or improving skin inflammation comprising the strain or a culture solution thereof. The composition of the present invention may be added to the quasi-drug composition for the purpose of preventing or treating skin inflammation.

In the present invention, the term "quasi-drug" refers to fibers, rubber products, or the like, which are used for the purpose of treating, alleviating, treating or preventing diseases of humans or animals, have weak action on the human body, or do not directly act on the human body. Or non-machines and the like, as an article corresponding to one of the preparations used for sterilization, insecticide and similar purposes for the prevention of infection type, for the purpose of diagnosing, treating, alleviating, treating or preventing diseases of humans or animals. It can refer to items other than instruments, machines, or devices among used items, and items used for the purpose of pharmacologically affecting the structure and function of humans or animals, excluding items that are not instruments, machines, or devices.

In addition, the quasi-drugs may include external preparations for skin and personal care products. Preferably, it may be a disinfectant cleaner, shower foam, gargrin, wet tissue, detergent soap, hand wash, or ointment, but is not limited thereto.

When the composition according to the present invention is used as a quasi-drug additive, the composition may be added as it is or used together with other quasi-drugs or quasi-drug ingredients, and may be appropriately used according to a conventional method. The mixing amount of the active ingredient may be suitably determined according to the purpose of use.

According to another embodiment of the present invention, the present invention is the strain or its culture

It provides a drinking water additive for preventing or improving skin inflammation, including. The drinking water additive of the present invention may be used by separately preparing a composition containing the strain or its culture solution in the form of a drinking water additive and mixing it with drinking water, or by directly adding the composition to drinking water. The drinking water additive of the present invention may be in a liquid or dry state, preferably in a dried powder form. A drying method for preparing the drinking water additive of the present invention in a dried powder form is not particularly limited, and a method commonly used in the art may be used. The drinking water additive of the present invention may further include other additives as needed. Non-limiting examples of the additives that can be used include binders, emulsifiers, preservatives, etc. added to prevent deterioration of the quality of drinking water, amino acids, vitamins, enzymes, probiotics, flavoring agents added to increase the utility of feed or drinking water , a non-protein nitrogen compound, a silicate agent, a buffer, a colorant, an extractant, or an oligosaccharide, and may further include a feed mixture, etc. in addition. These may be used alone or two or more may be added together.

Since the Bacillus licheniformis strain of the present invention is very effective in preventing or improving skin inflammation, the Bacillus licheniformis strain and its culture medium are foods or health functional foods requiring the prevention or improvement of skin inflammation or treatment. It can be very usefully used in pharmaceuticals, quasi-drugs, cosmetics, feed additives for improving skin inflammation of livestock or companion animals, or veterinary medicines.

1 is a result confirming the reduction of the inflammatory transcription factor NFAT5 and cytokines by LPS of the Bacillus licheniformis strain.
2 is a result confirming the reduction of skin inflammatory transcription factors and cytokines by TNF-α of the Bacillus licheniformis strain.

Hereinafter, the present invention will be described in detail by way of Examples and Experimental Examples. However, the following Examples and Experimental Examples are merely illustrative of the present invention, and the content of the present invention is not limited to the following Examples and Experimental Examples.

[Example 1]

<Preparation of cells and culture medium of Bacillus licheniformis>

Brain heart infusion (BHI, Difco Laboratories, U.S.A.) medium was inoculated with Bacillus licheniformis, grown by shaking culture at 30 ° C. for 48 hours, and then centrifuged to obtain a strain and a culture supernatant, respectively, and then It was used as a sample.

[Experimental Example 1]

<Test for measuring the anti-inflammatory activity of skin cells of Bacillus licheniformis>

<1-1> Experimental method

① Resistance test to H ₂ O ₂

Bacillus licheniformis was inoculated and cultured in BHI broth medium, centrifuged (7,000 × g , 20 minutes, 4℃), and washed 3 times with PBS solution to adjust the cell number to about 10 ⁸ CFU/mL in the same buffer. suspended. 500 μL of 0.5 mM H ₂ O ₂ solution and the same amount of cell suspension were immediately mixed and reacted for a total of 5 hours at 60-minute intervals, followed by centrifugation again to remove residual H ₂ O ₂ (20,000 × g , 1 min, 4 °C) to recover the pellet. After resuspending the pellets in buffer, diluting them step by step and plated on BHI agar (30°C, 24 hours) to measure the number of colonies produced, the reduction rate for the initial number of bacteria was shown.

② Antioxidant activity test by DPPH method

Antioxidant substances donate electrons or hydrogen to free radicals to form a complex, and 1,1-diphenyl-2-picyryl hydrazyl (DPPH) is an antioxidant. Since it receives electron hydrogen from a sexual substance to form an irreversibly stable molecule, it is possible to measure antioxidant activity from electron donating ability (Seo, Y.-H., Kim, I.-.J., Min). , H.-K. and Park, S.-U. 1999. Fatty acid composition and antioxidative activity in wax corn (Zea mays L.) F1s. Kor. J. Food Sic. Technol. 31, 1415-1420).

The hydrogen donating ability for 1,1-diphenyl-2-picyryl hydrazyl (1,1-diphenyl-2-picyryl hydrazyl; DPPH) was 100 M DPPH solution (6 mg of DPPH was completely dissolved in 100 mL of ethanol) mL of distilled water) Add 200 µL of each of the culture supernatant and sample of Bacillus licheniformis to 1000 µL, shake for 10 seconds, leave for 10 minutes, and then measure the absorbance at 528 nm.

③ NO production inhibition effect test by LPS

Raw 264.7 macrophages were aliquoted in DMEM medium containing 10% FBS in a 96-well plate at 1×10 ⁴ /well and cultured for 24 hours. After 24 hours, the adherent cells were pre-treated with a culture solution of 2.5% and 5% concentration for 1 hour, and then treated with LPS (100 ng/mL) and cultured for 20 hours. After 20 hours, the supernatant was taken and absorbance was measured at 540 nm using Geiess reagent (0.2% N-(1-naphthy) ethylene diamine solution: 0.2% sulfanilamide solution = 1:1). Inhibitory activity was evaluated by comparing with the group treated only with LPS.

Separately, in order to check whether the NO inhibitory activity is due to the cytotoxicity of the culture medium, the supernatant was taken to measure the amount of NO production, and the tetrazolium bromide salt (MTT, Sigma, St.Louis, MO, USA) solution was added to the final concentration of the culture medium. After incubating for about 4 hours, the MTT solution was removed, and 100 μL of DMSO was treated in each well to dissolve all the fomazan generated in the wells, and absorbance was measured at 570 nm. At this time, the ratio of the absorbance average of the culture medium-treated group to the absorbance average of the LPS-only group was calculated, and the survival rate (%) was calculated.

④ Inflammation and oxidative stress model development in skin cells

When HaCat cells, which are human keratinocytes, were aliquoted in DMEM medium containing 10% FBS in a 6-well plate and grown to about 80%, the culture solution of 2.5% and 5% concentration was treated for 1 hour. After 1 hour, 10 ng/mL TNF-α and 500 μM H ₂ O ₂ were treated and cultured for 24 hours.

⑤ Protein expression through Western blot analysis

Protein concentration in the cell lysate was measured using a BCA protein assay kit (Thermo scientific, Waltham, USA) by extracting intracellular proteins with a passive lysis buffer. After separating 40 μg of protein through 10% SDS-PAGE, blotting the separated protein on a nitrocellulose membrane, the membrane was reacted with primary antibody at 4°C for 18 hours, washed 3 times for 10 minutes, and then HRP was conjugated The secondary antibody was reacted at room temperature for 1 hour. At this time, protein expression was analyzed using the ECL detection system (AI680, GE healthcare, Uppsala, Sweden).

⑥ RNA isolation and gene analysis using qRT-PCR

Cells were cultured in a 6 well plate, the culture medium was pretreated for 1 hour, and then treated with TNF-α. After 24 hours, the cells were washed twice with PBS, and total RNA was isolated from the cells using TRIsol reagent. cDNA was synthesized using PCR thermal cycler dice (Takara Bio Inc., Japan), and quantitative real-time PCR was performed using SYBR Green mastermix in CFX connect Real-time PCR detection system (Bio-rad, USA). Gene expression was corrected for cyclophilin A using the threshold change method. Table 1 shows the specific primer sequences selected for the real-time PCR reaction.

Specific primer sequences for real-time PCR reactions Primer sequence TNF-α Forward 5’-CAGAGGGCCTGTACCTCATC-3’ Reverse 5’-GGAAGACCCCTCCCAGATAG-3’ IL-6 Forward 5’-CCACTCACCTCTTCAGAACG-3’ Reverse 5’-CATCTTTGGAAGGTTCAGGTT-3’ IL-1β Forward 5’-GCTACGAATCTCCGACCACCA-3’ Reverse 5’-AACCAGCATCTTCCTCAGCTTG-3’

<1-2> Experiment result

① Resistance test to H ₂ O ₂

Results showing the resistance to oxidative stress induced by H ₂ O ₂ of the Bacillus licheniformis culture are shown in Table 2.

Results of resistance test to oxidative stress induced by H ₂ O ₂ processing time 0 1 hours 3 hours 5 hours Inhibition rate (%) 0 8.4±1.2 ^a 35.6±2.1 ^b 78.2±2.1 ^c

Different English shoulder letters ( ^ac ) of resistance values to oxidative stress indicate statistical significance (p<0.05).

As a result of oxidative stress resistance evaluation, the inhibition rate was about 8.4% after 1 hour of reaction, 35.6% after 3 hours, and 78.2% after 5 hours. As a result of measuring the resistance of lactic acid bacteria to oxidative stress by cell viability according to the reaction time, it showed about 85.6 % cell viability after 5 hours of reaction. It was confirmed, and this is considered to increase resistance by detoxifying reactive oxygen species in the culture medium or by secreting antioxidant enzymes such as SOD and Catalase.

② Antioxidant activity test by DPPH method

Table 3 shows the results of measuring the electron donating ability by DPPH as the antioxidant power of the Bacillus licheniformis culture medium.

Antioxidant activity (DPPH radical scavenging ability) evaluation result culture medium live bacteria dead cells vitamin C DPPH radical scavenging ability (%) 54.9±3.2 ^a 64.2±4.1 ^ab 65.7±3.9 ^b 100 ^c

Different English shoulder letters ( ^ac ) of DPPH radical scavenging activity values indicate statistical significance (p<0.05).

The DPPH radical scavenging ability of the Bacillus licheniformis culture medium and live and dead cells was 54.9, 64.2, and 65.7%, respectively, as a result of measuring the BHI broth as a control. Various ROS and toxic products, including free radicals generated during oxidation, damage cells and molecules exhibiting biological activity in vivo. When the DPPH radical scavenging ability is high, the ability to reduce or offset free radicals is high, which means that it has antioxidant activity. Therefore, it can be said that the Bacillus licheniformis culture medium, live cells, and dead cells have antioxidant power, and based on this, antioxidant functional products can be developed.

③ NO production inhibitory effect by LPS

Raw 264.7 macrophages were treated with LPS to evaluate the inhibitory efficacy of the Bacillus licheniformis culture on NO that is generated inducing iNOS. Table 4 shows the NO production rate together with the cell viability indicative of cytotoxicity.

Inhibition of NO production and cell viability by LPS NO production rate Cell viability (%) control 0 100 ^d LPS treatment group 27.6±2.4 ^c 72.8±1.3 ^a LPS + culture medium 2.5% 12.6±2.1 ^b 75.2±1.9 ^ab LPS + 5% culture medium 10.1±2.5 ^ab 78.1±0.8 ^b LPS + culture medium 10% 8.7±1.1 ^a 82.9±1.4 ^c

Different English shoulder letters ( ^ad ) of the NO production inhibition value and the cell viability value indicate that there is a statistical significance (p<0.05).

NO production increased by LPS was decreased in a concentration-dependent manner in the culture medium of Bacillus licheniformis, and as the cell viability was also increased, it was shown that the inflammatory response by LPS was improved without cytotoxicity. In other words, LPS increases the production of NO by iNOS from L-arginine in immune cells, and the excessively generated NO intensifies the inflammatory response and is involved in the mechanisms of chronic inflammatory disease, autoimmune disease, cancer, and other diseases. it is known On the other hand, RAW264.7 macrophages are sensitive to LPS stimulation and produce NO well, so they are used to evaluate the anti-inflammatory effect. As a result of this experiment, it was confirmed that the Bacillus licheniformis culture medium effectively reduces NO excessively generated by LPS, similar to that extracts fermented with Lactobacillus inhibit NO production by regulating the iNOS gene. became

④ Reduction of inflammatory transcription factor NFAT5 and cytokines by LPS

Nuclear factor of activated T-cells 5 (NFAT5) has been found to be an important regulator of NFκB, and as it is activated in inflammatory diseases such as rheumatoid arthritis, severe sclerosis, and diabetic nephropathy, it is known as a major target of anti-inflammatory drug research. As the culture medium of Bacillus licheniformis inhibited NO production by LPS, the expression of NFAT5 and cytokine, an inflammatory transcription factor, was examined to elucidate the mechanism.

As shown in FIG. 1 , the expression of cytokines such as NFAT5 and TNF-α, IL-6, and IL-1β increased by LPS was decreased in a concentration-dependent manner in the Bacillus licheniformis culture medium. - need to be considered

⑤ Reduction of skin inflammatory transcription factors and cytokines by TNF-α

By identifying the anti-inflammatory activity of the Bacillus licheniformis strain in Raw 264.7 macrophages, and establishing a skin inflammatory model by TNF-α in skin cells similar to atopic dermatitis, anti-inflammatory according to the culture medium and live cell treatment of Bacillus licheniformis The inflammatory effect was investigated. As shown in FIG. 2 , the protein expression of NFAT5 and p65 increased by TNF-α was decreased, which resulted in a decrease in inflammatory cytokines such as IL-6 and IL-1β. That is, just as the Bacillus licheniformis strain showed an anti-inflammatory effect that inhibits NO production in Raw 264.7 macrophages, it also reduces the protein expression of NFAT5 and p65, which are inflammation regulators in skin cells, so that the gene expression of inflammatory cytokines is also was found to decrease.

[Experimental Example 2]

<Characteristic evaluation test of Bacillus licheniformis as a live cell>

<2-1> Experimental method

① Test strain

In order to know the basic properties of Bacillus licheniformis as a probiotic, acid resistance, bile resistance and heat resistance were measured. At this time, as a comparative strain, Bacillus subtilis (ATCC23857) was purchased from the American Type Culture Collection (ATCC, USA) and used.

The antibacterial activity of Bacillus licheniformis was determined by four bacterial species ( Salmonella enterica serovar Typhi (ATCC19430), Escherichia coli O157:H7 (ATCC 43895), Staphylococcus aureus (ATCC25923) , Listeria monocytogenes (ATCC 19113)), and one yeast species ( Candida albicans ). ATCC 24433) and one kind of mold ( Aspergilus fumigatus ATCC 96918) were investigated by the paper disk method. The bacteria and fungi were purchased from ATCC and used.

② Resistance to artificial gastric acid (acid resistance) test

Artificial gastric acid was inoculated with 1% of a strain suspension of at least 7.0 log CFU/mL in PPS (0.2% Photassium phosphate solution) adjusted to pH 2, 4, and 6, respectively, and cultured for 2 hours. The number of viable cells was counted by culturing.

③ Resistance to artificial bile fluid (biliary tolerance) test

For artificial bile, each artificial bile was prepared by adding Oxgall to BHI Broth at concentrations of 0.3, 0.5, and 1.0%, respectively. After inoculating 1% of the strain suspension of 8.0 log CFU/mL in the prepared artificial bile, cultured for 2 hours, plated on a plate medium, and then cultured at 37° C. for 24 hours to count the number of viable cells.

④ Heat resistance test

The comparative strain and the Bacillus licheniformis strain were cultured at 36.5°C for 20 hours (using BHI broth) and cultured for at least 7 log CFU/mL, then treated at 50, 60, 70°C temperature conditions (using Waterbath) for 10 minutes and plate After plating on the medium, the number of viable cells was counted by culturing at 37° C. for 24 hours.

⑤ Antibacterial test

After inoculating the Bacillus licheniformis strain in BHI broth, it was cultured at 36.5° C. for about 20 hours, cultured at least 7 log CFU/ml or more, and then the suspension was centrifuged and the supernatant was filtered and prepared as a test sample solution stock solution. After the test sample solution was concentrated 20 times using a reduced pressure concentrator, a 10-fold and 5-fold concentrate were prepared using BHI, respectively, and then used for the following antibacterial activity measurement.

4 types of test bacteria 4 types of test bacteria ( Sal. enterica serovar Typhi, E. coli O157:H7, Sta. aureus, L. monocytogenes ) were plated on BHI (Difco Laboratories) plate medium at least 7 log CFU/mL and prepared above. After immersing the paper disk sufficiently in the Bacillus licheniformis strain culture test sample solution prepared at 4 concentrations (undiluted solution, 5 times, 10 times, and 20 times concentrated solution), attach it to the center, incubate at 37°C for 20 hours, and then center the paper disk By measuring the diameter of the generated clear zone, the presence and degree of antibacterial activity were measured. The test yeast Can. albicans on sabouraud dextrose (Difco Laboratories) plate medium, test fungus Asp. fumigatus is smeared on PDA plate medium at 1×10 ⁶ CFU/mL, respectively, and after immersing the paper disk sufficiently in the test sample solution, attaching it to the center, incubating at 25°C for 72 hours, By measuring the diameter, the presence and degree of antibacterial activity were measured.

<2-2> Experiment result

① Resistance to artificial gastric acid (acid resistance) test result

The results of testing the viability of the Bacillus licheniformis strain in artificial gastric acid conditions are shown in Table 5 below.

Comparison of gastric acid resistance of Bacillus licheniformis strain name Number of viable bacteria (Log CFU/mL) pH 6.0 pH 4.0 pH 2.0 Bacillus lichenifomis 7.92 ± 0.05 ^a 4.13 ± 0.31 ^a,Y 2.74 ± 0.24 ^a,Y Bacillus subtilis 7.94 ± 0.03 ^a 3.67 ± 0.25 ^a,X 1.46 ± 0.15 ^a,X

Among the resistance values to artificial gastric acid, different English shoulder letters ( ^ac ) of the results for each strain indicate that there is a statistical significance (p<0.05).

Among the resistance values to artificial gastric acid, different English shoulder letters ( ^XY ) of the results between strains indicate that there is a statistical significance (p<0.05).

As a result of the test, the test strain, Bacillus licheniformis strain, showed an increase in growth to 7.92 log CFU/mL in the artificial gastric acid condition of pH 6, and 4.1 log CFU/mL in the artificial gastric acid condition of pH 4, showing a survival rate of about 59.0%. , showed a survival rate of about 39.1% at a level of 2.7 log CFU/mL under the condition of pH 2. On the other hand, the comparative strain, Bacillus subtilis, showed a survival rate of about 20.8% at a level of 1.46 log CFU/mL in artificial gastric acid culture conditions of pH 2, and the test strain Bacillus licheniformis showed higher artificial gastric acid resistance.

② Resistance to artificial bile fluid (biliary resistance) test

When Bacillus licheniformis strains were prepared and treated by concentrations of artificial bile solution (Oxgall), the results of testing the survival rate in artificial bile solution conditions are shown in Table 6 below.

Bacillus licheniformis strain resistance to artificial bile (biliary tolerance) test results strain name Number of viable bacteria (Log CFU/mL) oxgall 0.3% oxgall 0.5% oxgall 1.0% Bacillus lichenifomis 7.41 ± 0.03 ^c 4.43 ± 0.14 ^b 3.32 ± 0.25 ^a Bacillus subtilis 7.32 ± 0.04 ^c 4.38 ± 0.08 ^b 3.41 ± 0.08 ^a

Among the resistance values to artificial bile, different English shoulder letters ( ^ac ) of the results for each strain indicate that there is a statistical significance (p<0.05).

As a result of the test, the Bacillus licheniformis strain was about 92.6% at 7.41 log CFU/mL in the condition of oxgall 0.3% artificial bile, and 4.43 log CFU/mL in the condition of 0.5% artificial bile solution, about 55.3%, 1.0%. showed a survival rate of about 41.5% at a level of 3.32 log CFU/mL. The comparative strain, Bacillus subtilis, exhibited a survival rate of about 42.6% at a level of 3.41 log CFU/mL in the condition of 1.0% artificial bile acid, and the test strain, Bacillus licheniformis, is artificial bile acid resistance similar to that of Bacillus subtilis, a representative Bacillus sp. appeared to exist.

③ Heat resistance test

The heat resistance test results of the Bacillus licheniformis strain are shown in Table 7 below.

Heat resistance test result of Bacillus licheniformis strain strain name Number of viable bacteria (Log CFU/mL) 50℃ 60℃ 70℃ Bacillus lichenifomis 8.72 ± 0.06 ^c,Y 7.61 ± 0.03 ^b,Y 5.39 ± 0.03 ^a,Y Bacillus subtilis 7.91 ± 0.03 ^c,X 6.63 ± 0.07 ^b,X 4.78 ± 0.04 ^a,X

Among the result values for the heat resistance test, different English shoulder letters ( ^ac ) of the results for each strain indicate that there is a statistical significance (p<0.05).

Among the result values for the heat resistance test, different English shoulder letters ( ^XY ) between the strains indicate that there is a statistical significance (p<0.05).

As a result of the test, both the test strain, Bacillus licheniformis and the comparative strain, Bacillus subtilis, increased growth at 50°C, and when heat-treated at 60°C for 10 minutes, the test strain was 7.61 log CFU/mL, and the comparison strain was 6.63 log. CFU/mL is shown. When the test strain was exposed to 70 ° C. for 10 minutes, it exhibited a survival rate of about 77% at 5.39 log CFU / mL, and the comparative strain showed a survival rate of about 68.2% at a level of 4.78 log CFU / mL at 70 ° C., the test strain Bacillus licheniformis appeared to have a rather high heat resistance, and it was confirmed that it had excellent properties as a probiotic live cell.

④ Antibacterial test

Antimicrobial test results for 6 pathogens of Bacillus licheniformis strains are shown in Table 8 below.

Antimicrobial test results test strain antibacterial activity undiluted culture solution 5x concentrate 10-fold concentrate 20 times concentrate Salmonella enterica serovar Typhi (ATCC 19430) - - + ++ Escherichia coli O157:H7
ATCC 43895 - - - + Staphylococcus aureus
(ATCC 25923) - - - - Listeria monocytogenes
(ATCC 19113) - - - - Candida albicans
(ATCC 24433) - - - - Aspergilus fumigatus
(ATCC 96918) - - - -

'-' or '+' in Table 8 is determined based on the diameter size of the live cell inhibition ring, '-' means no antibacterial or antifungal activity, and '+' indicates that the diameter size of the inhibition ring is less than 14.0 mm Meaning, '++' means that the diameter of the ring of inhibition is in the range of 14.0 - 17.0 mm, and '+++' means that the diameter of the ring of inhibition exceeds 17.0 mm.

When the culture supernatant of the Bacillus licheniformis strain was concentrated 5 times, 10 times, and 20 times, the culture stock solution did not show antibacterial activity in all 6 types of the evaluated strains. In 20-fold concentrate, Sal. Enterica serover Typhi and E. coli O157:H7 showed only growth inhibitory activity. Bacillus licheniformis was judged not to have high antibacterial activity.

[Statistical Analysis]

Statistical analysis of the experimental results was analyzed using the 'Independent-Sample-T-Test method', a SPSS 13.0 statistical software, and expressed as the mean value ± standard difference. In addition, multiple comparisons were indicated by the LSD method, P>0.05: no significant difference, P<0.05: significant difference.

<Preparation Example 1> Preparation of microbial preparations

A preparation for preventing or improving skin inflammation containing Bacillus licheniformis is prepared by drying Bacillus licheniformis by a conventional freeze-drying method, and then mixing it with an excipient or encapsulating it.

<Production Example 2> Preparation of fermented milk

Fermented milk is prepared using the Bacillus licheniformis of the present invention according to a method commonly performed in the art. Crude milk (74.41%) and powdered skim milk (6.45%), etc. are mixed, and cultured seeds and Bacillus licheniformis are added and cultured to prepare a culture solution (1X10 ⁹ CFU/mL).

<Preparation Example 3> Preparation of food and beverage (health functional food) composition

<3-1> Preparation of beverage

The beverage is prepared by mixing the ingredients of the formulations set forth below according to methods commonly practiced in the art. Honey 522 mg, thioctoacid amide 5 mg, nicotinic acid amide 10 mg, riboflavin hydrochloride 3 mg, pyridoxine hydrochloride 2 mg, inositol 30 mg, ortic acid 50 mg, culture supernatant of Bacillus licheniformis, 200 mL of water.

<3-2> Preparation of powder

Bacillus licheniformis freeze-dried powder (1×10 ⁹ CFU/g) 20 mg, lactose 100 mg, and talc 10 mg are mixed and filled in an airtight bag to prepare a powder.

<3-3> Preparation of tablets

After mixing 10 mg of Bacillus licheniformis freeze-dried powder (1×10 ⁹ CFU/g), 100 mg of corn starch, 100 mg of lactose, and 2 mg of magnesium stearate, tablets are prepared by tableting according to a conventional tablet manufacturing method. .

<3-4> Preparation of capsules

Bacillus licheniformis freeze-dried powder (1×10 ⁹ CFU/g) 10 mg, crystalline cellulose 3 mg, lactose 14.8 mg, and magnesium stearate 0.2 mg were mixed according to a conventional capsule preparation method and filled in a gelatin capsule. Prepare capsules.

<3-5> Preparation of mixed powder

Bacillus licheniformis freeze-dried powder (1×10 ⁹ CFU/g) 1 g, vitamin mixture appropriate amount (vitamin A acetate 70 μg, vitamin E 1.0 mg, vitamin B1 0.13 mg, vitamin B2 0.15 mg, vitamin B6 0.5 mg, vitamin B12 0.2 μg, vitamin C 10 mg, biotin 10 μg), nicotinic acid amide 1.7 mg, folic acid 50 μg, calcium pantothenate 0.5 mg, mineral mixture (ferrous sulfate 1.75 mg, zinc oxide 0.82 mg, magnesium carbonate 25.3 mg, 15 mg of potassium monophosphate, 55 mg of dibasic calcium phosphate, 90 mg of potassium citrate, 100 mg of calcium carbonate, and 24.8 mg of magnesium chloride) were mixed according to a conventional health functional food manufacturing method, and then granules were prepared, and the conventional method It can be used to prepare a health functional food composition according to the present invention. The composition ratio of the above vitamin and mineral mixture is relatively suitable for health functional food, but the mixing composition is a preferred example, but the mixing ratio may be arbitrarily modified.

<Preparation Example 4> Preparation of external preparation for skin (ointment)

The ointment contains 5.00% by weight of Bacillus licheniformis cells or culture medium powder, 10.00% by weight of caprin/caprylic triglyceride, 10.00% by weight of liquid paraffin, 6.00% by weight of sorbitan sesquinoate, It is prepared by mixing 9.00% by weight of octyldodeces-25, 10.00% by weight of cetylethylhexanoate, 1.00% by weight of squalane, 1.00% by weight of salicylic acid, 15.00% by weight of glycerin, 10.00% by weight of sorbitol, and the remainder by weight of purified water.

<Preparation Example 5> Preparation of feed composition

According to a method commonly performed in the art, the composition for feed is corn 45 g, soybean meal 30 g, wheat 15 g, tallow 4 g, molasses 3 g, calcium phosphate 2 g, limestone 0.4 g, salt 0.2 g, freeze-dried Bacillus lychee It is prepared by mixing 5 g of niformis cell powder.

<Production Example 6> Preparation of cosmetics

<6-1> Preparation of lotion

According to a method commonly performed in the art, 0.05 g of polypyrrolidone, 0.1 g of polyol alcohol, 0.2 g of polyoxyethylene monooleate, 0.2 g of fragrance, 0.1 g of methyl paraoxybenzoate, 0.1 g of methyl paraoxybenzoate in 8 g of 95% ethanol It is prepared by mixing and dissolving a small amount of an antioxidant of Bacillus licheniformis culture powder, 5 g of glycerin was dissolved in 85.33 g of purified water, and the mixture was added and stirred to obtain a lotion having an effect of inhibiting and improving skin inflammation.

<6-2> Preparation of lotion

The lotion contains 0.10 wt% of a Bacillus licheniformis culture solution, 3.00 wt% of glycerin, 0.10 wt% of carbomer, 0.05 wt% of xanthan gum, 3.00 wt% of 1,3-butylene glycol, poly Glyceryl-3 methylglucose distearate 1.50% by weight, glyceryl stearate 0.50% by weight, cetylaryl alcohol 0.30% by weight, jojoba oil 3.00% by weight, liquid paraffin 2.00% by weight, squalane 3.00% by weight, dimethicone 0.50% by weight , tocopheryl acetate 0.20% by weight, triethanolamine 0.10% by weight, preservatives, fragrance, trace amounts of color, and purified water residual weight% by mixing.

<6-3> Preparation of emulsion

The emulsion is prepared according to a method conventionally performed in the art: 1.2 g of cetyl alcohol, 10 g of squalane, 2 g of vaseline, 0.2 g of ethyl paraoxybenzoate, 1 g of glycerin monoester, 1 g of polyoxyethylene (addition of 20 moles) 1 g of monooleate and 0.1 g of fragrance were mixed and dissolved at 70 ° C., and 0.05 g of Bacillus licheniformis culture solution powder, 5 g of dipropylene glycol, 2 g of polyethylene glycol 1500, 0.2 g of triethanolamine, and 76.2 g of purified water were heated to 75 ° C. heat to dissolve Both are mixed and emulsified, and then cooled to obtain a water/oil type emulsion that has the effect of improving skin inflammation.

<6-4> Preparation of cosmetic liquid

According to a method conventionally performed in the art, 1.2 g of polyoxyethylene sorbitan monooleate, 0.3 g of kitulose, 0.2 g of sodium hyaluronate, 0.2 g of vitamin E-acetate, licoric acid in 5 g of 95% ethyl alcohol. 0.2 g of sodium, 0.1 g of paraoxybenzoic acid ester ester, 1 g of Bacillus licheniformis culture powder, and an appropriate amount of pigment are mixed to obtain a cosmetic solution with an effect of improving skin inflammation.

From the above description, those skilled in the art to which the present invention pertains will understand that the present invention may be embodied in other specific forms without changing the technical spirit or essential characteristics thereof. In this regard, it should be understood that the manufacturing examples described above are illustrative in all respects and not restrictive. The scope of the present invention should be construed as being included in the scope of the present invention, rather than the above detailed description, all changes or modifications derived from the meaning and scope of the claims described below and their equivalents.

The present invention is a Bacillus licheniformis strain having a skin inflammation inhibition, improvement or therapeutic effect, the strain or its culture medium has high antioxidant and oxidative stress resistance and blocks or expresses factors that cause inflammation in skin cells By providing a microbial agent that can be used for the prevention, improvement or treatment of diseases related to skin inflammation by inhibiting it, it can be widely used in related industries such as pharmaceuticals, food and feed, and cosmetics industries.

Claims (10)

Bacillus licheniformis strain having a preventive or ameliorating effect of skin inflammation ( Bacillus licheniformis ). A microbial preparation for preventing or improving skin inflammation comprising the strain of claim 1 or a culture solution thereof as an active ingredient. A method for producing a microbial agent for preventing or improving skin inflammation comprising the step of culturing the strain of claim 1. A probiotic bioactive composition comprising the strain of claim 1 or a culture solution thereof as an active ingredient. A pharmaceutical composition for preventing, improving or treating skin inflammation comprising the strain of claim 1 or a culture solution thereof as an active ingredient. A food or health functional food composition for preventing or improving skin inflammation comprising the strain of claim 1 or a culture solution thereof as an active ingredient. A quasi-drug composition for preventing or improving skin inflammation comprising the strain of claim 1 or a culture solution thereof as an active ingredient. A drinking water additive for preventing or improving skin inflammation comprising the strain of claim 1 or a culture solution thereof as an active ingredient. A feed additive for preventing or improving skin inflammation comprising the strain of claim 1 or a culture solution thereof as an active ingredient. A cosmetic composition for preventing or improving skin inflammation comprising the strain of claim 1 or a culture solution thereof as an active ingredient.

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