Lacticaseibacillus rhamnosus ICL-46 strain derived from oat and its use for Anti-inflammation, Anti-allergy, skin immunity enhancement, atopic dermatitis improvement, skin soothing and skin microorganism regulation

Abstract

To provide a novel Lacticaseibacillus rhamnosus strain.SOLUTION: The present invention relates to a novel Lacticaseibacillus rhamnosus ICL-46 strain and its use for anti-inflammation, anti-allergy, skin soothing, and skin microorganism regulation. According to one embodiment of the strain, since it has effects of anti-inflammation, anti-allergy, skin immunity enhancement, atopic dermatitis improvement, skin soothing, and skin microorganism regulation, it can be variously applied to cosmetics, pharmaceuticals, foods, and the like.SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] The present invention relates to an oat-derived Lacticase I Bacillus rhamnosus ICL-46 strain and its uses for anti-inflammatory, anti-allergic, skin immunity enhancement, atopic dermatitis amelioration, skin soothing and skin microorganism regulation, and more particularly to a novel oat-derived microorganism and its uses for anti-inflammatory, anti-allergic, skin immunity enhancement, atopic dermatitis amelioration, skin soothing and skin microorganism regulation. [Background technology]

[0002] Probiotics refer to live microorganisms that provide health benefits. Representative probiotic strains include Lactobacillus and Bifidobacterium strains. Recently, active research has been conducted into the application of probiotic microorganisms to cosmetics to produce functional cosmetics (see Patent Document 1). Oats (Avena sativa) also contain ingredients that are beneficial to the skin and are widely used in the field of cosmetics in the form of oat extract, oat oil, oat peptides, oat kernel flour, etc. However, there is currently insufficient research into oat-derived microorganisms and their skin beauty effects. [Prior art documents] [Patent documents]

[0003] [Patent Document 1] Special Publication No. 2024-543178 Summary of the Invention [Problem to be solved by the invention]

[0004] An object of the present invention is to provide a novel Lacticaseibacillus rhamnosus strain. Another object of the present invention is to provide a lysate, culture medium, extract of the culture medium, fermentation medium, or any combination thereof of the strain. Another object of the present invention is to provide a method for producing the disrupted product of the strain. Yet another object of the present invention is to provide a composition comprising the Lacticaseibacillus rhamnosus strain, its lysate, culture medium, extract of the culture medium, fermentation medium, or any combination thereof. A further object of the present invention is to provide uses of the composition for anti-inflammatory, anti-allergic, skin immunity promotion, skin soothing and skin microorganism regulation. It is still another object of the present invention to provide a method for preventing, ameliorating, or treating a skin condition in an individual, comprising administering to the individual in need thereof an effective amount of the composition. [Means for solving the problem]

[0005] The present invention is characterized by the Lacticaseibacillus rhamnosus ICL-46 strain deposited under accession number KCTC15702BP.

[0006] The strain may be isolated from oat (Avena sativa). As used herein, the term "isolated" means something that is not naturally occurring, but is artificially isolated and available for use. The strain may be isolated from oat (Oat or Avena sativa). The strain is preferably isolated from oat kernel, oat bran, or a mixture thereof.

[0007] The strain may be isolated by inoculating oats into an MRS medium, culturing the resulting colonies, and then isolating and culturing them. In this specification, "Lacticaseibacillus rhamnosus isolated from oats" can be used interchangeably with "Lacticaseibacillus rhamnosus derived from oats."

[0008] The strain may be a strain belonging to the genus Lacticaseibacillus.

[0009] The strain may contain a 16S rRNA that has about 95% or more, about 96% or more, about 97% or more, about 98% or more, about 99% or more, about 99.5% or more, or about 99.9% or more sequence identity to SEQ ID NO: 1. Preferably, the strain contains the 16S rRNA of SEQ ID NO: 1.

[0010] As used herein, the term "sequence identity" refers to the degree of identity of amino acid residues or bases between sequences after aligning the two sequences for maximum correspondence in a specific comparison region. The percentage of sequence identity can be determined using known sequence comparison programs, such as BLASTN (NCBI), CLC Main Workbench (CLC bio), and MegAlign™ (DNASTAR Inc.).

[0011] The Lacticaseibacillus rhamnosus strain is preferably the strain deposited under accession number KCTC15702BP.

[0012] The strain may have anti-inflammatory, anti-allergic, skin immunity enhancing, skin soothing or skin microbiota regulating effects.

[0013] In other embodiments, the strain may be provided as a lysate, a culture medium, an extract of the culture medium, a fermentation medium, or any combination thereof.

[0014] The specific details of the strain are as described above. As used herein, the term "lysate" refers to a disruption product of useful components, such as intracellular biological components contained in microorganisms, e.g., cell walls or membranes, and a culture medium containing metabolites released extracellularly during microbial cultivation. The lysate can be prepared by disruption using pressure or ultrasound, excluding methods that induce transformation of microbial metabolites, such as irradiation or heat inactivation. In one embodiment, the disrupted strain of the strain can be prepared using a microfluidizer (MF). Because useful components of microorganisms are susceptible to deterioration due to excessive heat and energy, it is recommended to prepare the disrupted product using a microfluidizer, which is the most stable method for obtaining useful components from microorganisms among existing production processes. The disrupted material may include the disrupted material itself, as well as a concentrate or freeze-dried product thereof.

[0015] As used herein, the term "culture medium" can be used interchangeably with "culture supernatant," "conditioned culture medium," or "conditioned medium" and refers to the entire medium containing the Lacticase Bacillus rhamnosus strain, its metabolites, excess nutrients, etc., obtained by culturing the strain for a certain period of time in a medium that provides nutrients so that the strain can grow and survive in a test tube. The term "culture medium" also refers to the culture medium obtained by culturing a strain and removing the bacterial cells from the bacterial cell culture medium. The liquid from which the bacterial cells have been removed is called the "supernatant." The culture medium can be left to stand for a certain period of time and the upper layer can be collected by removing the precipitated portion, filtering the bacterial cells, or centrifuging the culture medium to remove the lower precipitate and obtain the upper liquid. The "bacterial cells" refers to the bacterial strain of the present invention itself, and includes the bacterial strain itself isolated and selected from a sample, etc., or the bacterial strain isolated from the culture medium after culturing the strain. The bacterial cells can be obtained by centrifuging the culture medium and removing the portion that sinks to the bottom, or by leaving the culture medium to stand for a certain period of time and removing the liquid above, since the bacterial cells sink to the bottom due to gravity.

[0016] The culture medium includes the culture medium itself obtained by culturing the strain, a concentrate or freeze-dried product thereof, or a culture supernatant obtained by removing the strain from the culture medium, a concentrate or freeze-dried product thereof. The culture medium of the strain can be separated into cells and a supernatant by centrifugation. As used herein, the term "lysate" can refer to a mixture of the culture medium of the strain and the lysed cells.

[0017] The culture medium and conditions for culturing the Lacticase Bacillus rhamnosus strain can be appropriately selected or modified by those skilled in the art. For example, the medium may be prepared with one or a mixture of glucose, yeast extract, malt extract, peptone, glutamic acid, KHPO, KHPO, NaCl, MnSO, and MgSO. The culture conditions include, but are not limited to, culturing at a temperature of 30 to 40°C for 12 to 72 hours.

[0018] As used herein, the term "culture extract" refers to a substance extracted from the culture medium or a concentrate thereof, and may include an extract, a diluted or concentrated extract, a dried product obtained by drying the extract, or a preparation or purified product thereof, or a fraction obtained by fractionating any of these.

[0019] Another aspect is to provide a method for producing a lysate of said strain. The method includes the steps of inoculating a Lacticase Bacillus rhamnosus strain into a medium, culturing the strain, and obtaining a culture solution of the strain; centrifuging the culture medium to separate the cells and the supernatant; The method may further include a step of mixing the disrupted bacterial cells with the supernatant to obtain a disrupted bacterial cell product.

[0020] The method further includes a step of sterilizing the medium before the step of obtaining the strain culture solution. The sterilization can be carried out by a method commonly used in the art, for example, at 110 to 130°C for 1 minute to 1 hour.

[0021] The medium is not limited to any particular type, as long as it can culture the Lacticase Bacillus rhamnosus strain. The culture medium and conditions for culturing the Lacticase Bacillus rhamnosus strain can be appropriately selected or modified by those skilled in the art. For example, the medium may be prepared with one or a mixture of glucose, yeast extract, malt extract, peptone, glutamic acid, KHPO, KHPO, NaCl, MnSO, and MgSO. The culture conditions may include, but are not limited to, culturing the strain at a temperature of 30 to 40°C for 12 to 72 hours. A strain culture solution can be obtained by the culture.

[0022] The step of centrifuging the culture medium to separate the cells and the supernatant may further include a step of filtering the culture medium after centrifugation. The filtering may be performed according to a conventional method in the art.

[0023] In the step of obtaining the disrupted strain, the bacterial cells can be disrupted using, but not limited to, a microfluidizer. In another aspect, there is provided use of the Lacticase Bacillus rhamnosus strain, its lysate, culture medium, extract of the culture medium, fermentation liquid, or any combination thereof. More specifically, there can be provided a composition comprising the Lacticase Bacillus rhamnosus strain, its lysate, culture medium, extract of the culture medium, fermentation liquid, or any combination thereof.

[0024] In the above-mentioned use or composition, the Lacticase Bacillus rhamnosus strain may be the Lacticase Bacillus rhamnosus ICL-46 strain according to the above-mentioned embodiment. The uses of the strain may include anti-inflammatory, anti-allergic, skin immunity enhancement, skin soothing, skin microbiome regulation, and prevention or treatment of skin diseases.

[0025] The strain can suppress or reduce the expression of genes associated with inflammation (e.g., COX-2, iNOS), suppress or reduce the release of β-hexose aminase, a key factor associated with allergy or immunity, and suppress or reduce harmful skin bacteria (e.g., P. aeruginosa, S. aureus) while leaving beneficial skin bacteria (e.g., S. epidermidis) unaffected.

[0026] The term "anti-inflammatory" can be used interchangeably with "inflammatory improvement" and "inflammatory suppression," and refers to any action that alleviates immune responses and suppresses the production of COX-2 or iNOS. The term "antiallergic" can be used interchangeably with "allergy improvement" and "allergy suppression," and refers to any action that suppresses or improves the body's allergic reaction. The term "skin immunity enhancement" refers to any action that enhances the skin's immunity, such as, but not limited to, suppressing allergy-inducing substances (e.g., β-hexoaminase) or suppressing harmful bacteria in the skin.

[0027] The term "skin disease" may be a skin inflammatory disease, a skin allergic disease, or an immune-mediated skin disease. The term "prevention" includes inhibiting the occurrence of a disease. The term "treatment" includes inhibiting, alleviating, or eliminating the onset of a disease. The skin inflammatory disease may be any one or more selected from the group consisting of skin wounds, dermatitis, atopic dermatitis, pruritus, eczematous skin diseases, dry eczema, erythema, hives, psoriasis, mild rash, and acne. The skin allergic disease may be any one or more selected from the group consisting of atopic dermatitis, urticaria, mild rash, and pruritus.

[0028] The immune-mediated skin disease is a general term for skin diseases related to immunity, and may include, for example, atopic dermatitis, seborrheic dermatitis, psoriasis, and the like. The term "skin soothing" means any action that soothes and stabilizes the heat or pain of irritated skin. The skin soothing includes soothing skin that has become sensitive due to inflammation, allergies, or a weakened immune system. The term "skin microbiota control" refers to increasing beneficial skin bacteria and decreasing harmful skin bacteria, or suppressing or decreasing only harmful skin bacteria without affecting beneficial skin bacteria. Specifically, skin microbiota control means suppressing or decreasing the growth of harmful skin bacteria while not affecting the growth of beneficial skin bacteria. The beneficial and harmful skin bacteria can include any known strains. In one embodiment, the beneficial skin bacteria can be, but are not limited to, StapHylococcus epidermidis. The harmful skin bacteria can be, but are not limited to, Pseudomonas aeruginosa or StapHylococcus aureus.

[0029] A composition according to one embodiment can suppress or reduce the expression of one or more of COX-2 and iNOS by containing the Lacticase I Bacillus rhamnosus strain, its lysate, culture medium, extract from the culture medium, fermentation medium, or any combination thereof, and therefore can exert anti-inflammatory effects and preventive or therapeutic effects on inflammatory skin diseases. A composition according to one embodiment contains the Lacticase I Bacillus rhamnosus strain, its homogenate, culture medium, culture medium extract, fermentation medium, or any combination thereof, and can suppress or reduce β-hexoaminase, a major factor in allergies. Therefore, the composition can exert anti-allergic effects, enhance skin immunity, and prevent or treat allergic skin diseases and immune-mediated skin diseases.

[0030] A composition according to one embodiment contains the Lacticase I Bacillus rhamnosus strain, its disruptant, culture medium, extract of the culture medium, fermentation medium, or any combination thereof, and thus does not affect the growth of beneficial skin bacteria (S. epidermidis), but can suppress or reduce the growth of harmful skin bacteria (P. aeruginosa, S. aureus). Thus, the composition can exert a skin microbiome-regulating effect. The composition according to one embodiment can exert a skin soothing effect by any one or more of the anti-inflammatory, anti-allergic, skin immunity enhancing, and skin microbiota regulating effects. In other embodiments, the strain may be used in combination with other strains that have the effect of improving skin conditions to achieve a synergistic effect.

[0031] The composition may contain, relative to the total weight of the composition, 0.001% by weight to 80% by weight, for example, 0.01% by weight to 60% by weight, 0.01% by weight to 40% by weight, 0.01% by weight to 30% by weight, 0.01% by weight to 20% by weight, 0.01% by weight to 10% by weight, 0.01% by weight to 5% by weight, 0.05% by weight to 60% by weight, 0.05% by weight to 40% by weight, 0.05% by weight to 30% by weight, 0. The composition may contain 0.05% to 20% by weight, 0.05% to 10% by weight, 0.05% to 5% by weight, 0.1% to 60% by weight, 0.1% to 40% by weight, 0.1% to 30% by weight, 0.1% to 20% by weight, 0.1% to 10% by weight, or 0.1% to 5% by weight of the strain, its disruptant, culture medium, extract of the culture medium, fermentation medium, or any combination thereof.

[0032] The term "containing as an active ingredient" as used herein means that the strain, its disrupted product, culture medium, extract of the culture medium, fermentation medium, or any combination thereof is added to an extent that the aforementioned effect can be exerted, and is also intended to mean that various components are added as accessory ingredients for drug delivery, stabilization, etc., and that the formulation is made in various forms.

[0033] The composition may be in a liquid or dry state, or in one embodiment, in the form of a dry powder. The drying method for producing the composition in a dry state can be any method commonly used in the art, but is not particularly limited thereto. Non-limiting examples of the drying method include air drying, natural drying, spray drying, freeze drying, etc. These methods can be used alone or at least two methods can be used in combination.

[0034] The composition may include an additive, such as, but not limited to, a binder, in an effective amount sufficient to reduce deterioration of the strain, its disruptant, culture medium, extract of the culture medium, fermentation medium, or any combination thereof. The composition may further comprise a cosmetically, pharmaceutically, or food-grade acceptable carrier. The composition may be formulated together with the carrier and provided as a cosmetic, drug, food additive, or the like.

[0035] The composition may be a cosmetic composition. In addition to the active ingredients disclosed herein, the cosmetic composition may further include ingredients, functional additives, etc. that are commonly used in cosmetic compositions, such as antioxidants, stabilizers, solubilizers, surfactants, dispersants, preservatives, vitamins, pigments, fragrances, and other common adjuvants and carriers. The cosmetic composition is not limited to a specific formulation and can be appropriately selected depending on the purpose. The cosmetic composition can include, for example, a solubilized formulation, an emulsion formulation, or a dispersion formulation. The cosmetic composition can include, for example, a softening lotion, a nourishing lotion, a massage cream, a nourishing cream, an essence, a pack, a gel, an ampoule, or a skin adhesive type cosmetic formulation.

[0036] The composition may be a composition for external use on the skin. In this specification, the topical skin preparation may be a cream, gel, ointment, skin emulsion, skin suspension, transdermal delivery patch, drug-containing bandage, lotion, or a combination thereof. The topical skin preparation may contain ingredients typically used in topical skin preparations such as cosmetics or pharmaceuticals, such as aqueous ingredients, oily ingredients, powder ingredients, alcohols, moisturizers, thickeners, UV absorbers, whitening agents, preservatives, antioxidants, surfactants, fragrances, colorants, various skin nutrients, or combinations thereof, as needed.

[0037] The composition can be a pharmaceutical composition. The pharmaceutical composition may further comprise a pharmaceutically acceptable diluent or carrier. The diluent may be lactose, corn starch, soybean oil, microcrystalline cellulose, mannitol, or a combination thereof. The carrier may be an excipient, disintegrant, binder, lubricant, or a combination thereof. The excipient may be microcrystalline cellulose, lactose, low-substituted hydroxycellulose, or a combination thereof. The disintegrant may be calcium carboxymethylcellulose, sodium starch glycolate, anhydrous calcium phosphate, or a combination thereof. The binder is preferably polyvinylpyrrolidone, low-substituted hydroxypropylcellulose, hydroxypropylcellulose, or a combination thereof. The lubricant may be magnesium stearate, silicon dioxide, talc, or a combination thereof. The pharmaceutical composition can be formulated into oral or parenteral dosage forms. Oral dosage forms may be granules, acid preparations, liquid preparations, tablets, capsules, dry syrups, etc. Parenteral dosage forms may be injections, ointments, etc.

[0038] The composition may be a food composition. In this case, it may be formulated using a conventional health functional food formulation known in the art. Therefore, the food composition may be a health functional food composition.

[0039] The food composition may be prepared by using the strain, its disrupted product, culture medium, culture medium extract, fermentation liquid, or any combination thereof, alone or in combination with other foods or food ingredients, and may be prepared according to conventional methods. The amount of active ingredient to be added may be determined appropriately depending on the intended use (prevention, health, or therapeutic treatment). The type of functional health food is not particularly limited. Among the types of functional health foods, beverage compositions may contain various flavorings or natural carbohydrates as additional ingredients, similar to ordinary beverages. Natural carbohydrates include monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, polysaccharides such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. Sweeteners may include natural sweeteners such as thaumatin and stevia extract, or synthetic sweeteners such as saccharin and aspartame. The food composition may also include nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonation agents used in carbonated beverages, or combinations thereof. The food composition may also include natural fruit juice, fruit juice drinks, fruit pulp for producing vegetable drinks, or combinations thereof.

[0040] Another aspect provides a method of preventing, ameliorating, or treating a skin condition in an individual comprising administering to the individual in need thereof an effective amount of the composition. The individual's skin condition may be an inflammatory, allergic, or immune-related condition.

[0041] As used herein, the terms "administering," "introducing," and "implanting" are used interchangeably and refer to the placement of a composition according to an embodiment into an individual by a method or route that results in at least partial localization of the composition according to an embodiment to a desired site. Administration can be performed according to methods known to those skilled in the art. Administration can be performed directly to an individual by any means, such as intravenous, intramuscular, oral, transdermal, mucosal, intranasal, intratracheal, or subcutaneous administration. The administration can be systemic or local.

[0042] The individual may be a mammal, such as a human, cow, horse, pig, dog, sheep, goat, or cat, and may be an individual in need of improved skin condition, such as an individual in need of anti-inflammatory, anti-allergic, skin immune-boosting, skin soothing, or skin microbiome-regulating effects.

[0043] The administration may be 0.1 mg to 1,000 mg of the composition according to one embodiment administered per individual per day, for example, 0.1 mg to 500 mg, 0.1 mg to 100 mg, 0.1 mg to 50 mg, 0.1 mg to 25 mg, 1 mg to 1,000 mg, 1 mg to 500 mg, 1 mg to 100 mg, 1 mg to 50 mg, 1 mg to 25 mg, 5 mg to 1,000 mg, 5 mg to 500 mg, 5 mg to 100 mg, 5 mg to 50 mg, 5 mg to 25 mg, 10 mg to 1,000 mg, 10 mg to 500 mg, 10 mg to 100 mg, 10 mg to 50 mg, or 10 mg to 25 mg. However, dosages can vary depending on factors such as formulation method, administration method, patient age, weight, sex, condition, diet, administration time, administration route, excretion rate, and reaction sensitivity. Those skilled in the art can appropriately adjust dosages taking these factors into account. Administration can be once daily or two or more times per day within the range of clinically acceptable side effects. Administration can be at one or more sites, daily or every 2 to 5 days, with a total administration period of one to 30 days. If necessary, the same treatment can be repeated at an appropriate time. For non-human animals, the dosage can be the same per kg as for humans, or can be calculated based on, for example, the volume ratio (e.g., average) of organs (e.g., heart) between the target animal and humans. [Effects of the Invention]

[0044] According to one embodiment, the novel Lacticasei Bacillus rhamnosus strain has anti-inflammatory, anti-allergic, skin immunity enhancing, atopic dermatitis improving, skin soothing, or skin microbiota regulating effects, and can therefore be widely used in cosmetics, pharmaceuticals, foods, etc. [Brief explanation of the drawings]

[0045] [Figure 1] 1 is a graph showing the expression level of COX-2 mRNA depending on the type of lactic acid bacteria strain derived from oats. [Figure 2] 1 is a graph showing the expression level of iNOS mRNA depending on the type of lactic acid bacteria strain derived from oats. [Figure 3] 1 is a graph showing the β-hexaminase inhibitory effect depending on the type of lactic acid bacteria strain derived from oats. [Figure 4] 1 is a graph comparing the growth of beneficial skin bacteria S. epidermidis and harmful skin bacteria P. aeruginosa when co-cultured and treated with crushed Lacticase Bacillus rhamnosus ICL-46. [Figure 5] 1 is a graph comparing the growth of beneficial skin bacteria S. epidermidis and harmful skin bacteria S. aureus when co-cultured and treated with a homogenate of Lacticase I Bacillus rhamnosus ICL-46. DETAILED DESCRIPTION OF THE INVENTION

[0046] Hereinafter, the present invention will be described in more detail through embodiments thereof. However, the embodiments are for illustrative purposes only and the scope of the present invention is not limited to the embodiments.

[0047] Embodiment 1. Isolation and identification of oat-derived strains Microorganisms were isolated and identified from oats. Specifically, undried oat kernels and oat bran were crushed using a sterilized crusher, and 10 g of the crushed material was added to 90 mL of MRS (de Man, Rogosa, and Sharpe) liquid medium and cultured at 37°C for approximately 48 hours. The cultured liquid medium was diluted ten-fold with saline, smeared on MRS plates, and cultured under the same culture conditions for 48 hours or more. After that, bacterial strains were randomly isolated from the resulting colonies.

[0048] Randomly isolated strains were subcultured three or more times, and pure colonies isolated were dropped onto BCP (Bromocresol Purple) Agar Plates (manufacturer: Eiken Chemical, product name: BCP plate count agar) to confirm acid production. Strains with milky white colonies and yellow surrounding areas were selected. The selected colonies were analyzed for the 16S rRNA gene sequence for identification and classification. A similarity search on NCBI for the analyzed 16S rRNA sequence showed 99.67% homology with Lacticaseibacillus rhamnosus. The novel microbial strain Lacticaseibacillus rhamnosus isolated from the oats (designated "ICL-46") was deposited with the Korea Center for Bioscience and Biotechnology (KCTC) on November 20, 2023, and was assigned the accession number KCTC15702BP. The ICL-46 strain has a 16S rRNA sequence of SEQ ID NO: 1 (complementary DNA).

[0049] Embodiment 2: Production of Lacticase Bacillus rhamnosus ICL-46 Strain Disruption The medium was sterilized for 15 minutes at 121° C. The medium can be appropriately selected depending on the type of microorganism to be inoculated, and for example, a medium prepared with one or a mixture of glucose, yeast extract, malt extract, peptone, glutamic acid, KHPO, KHPO, NaCl, MnSO, and MgSO can be used. The Lacticase Bacillus rhamnosus ICL-46 strain of Example 1 was inoculated into a sterilized medium and then cultured at 37°C for 24 hours. After the cultivation was completed, the cells and the supernatant were separated by centrifugation. The cells were disrupted by passing them twice through a microfluidizer at 800 bar. The disrupted cells and the supernatant were mixed to obtain a disrupted product of Lacticase I Bacillus rhamnosus ICL-46 strain.

[0050] Comparative Examples 1 to 4. Isolation of lactic acid bacteria from oats and production of disrupted products Lactic acid bacteria were isolated from oats using the same method as in Example 1. Analysis of the nucleotide sequence of the 16S rRNA gene confirmed the presence of Lactiplantibacillus plantarum, Lactococcus lactis, Latilactobacillus sakei, and Lacticaseibacillus rhamnosus. The Lacticaseibacillus rhamnosus strains are the same species as the ICL-46 strain of Example 1, but have different 16S rRNA sequences.

[0051] Lysates of the oat-derived lactic acid bacteria were produced in the same manner as in Example 2. The lysate of the Lactoplantobacillus plantarum strain was designated Comparative Example 1, the lysate of the Lactococcus lactis strain was designated Comparative Example 2, the lysate of the Lactyllactobacillus sakei strain was designated Comparative Example 3, and the lysate of the Lactocaseibacillus rhamnosus strain was designated Comparative Example 4.

[0052] Experimental Example 1. Confirmation of anti-inflammatory effects - Analysis of COX-2 and iNOS expression To confirm the anti-inflammatory efficacy of the lysate of Bacillus rhamnosus ICL-46 strain, the expression of COX-2 (cyclooxygenase-2) and iNOS (nitric oxide synthase) genes was analyzed by real-time PCR. Specifically, mouse macrophages Raw 264.7 were treated with LPS (liposaccharides), and then 0.5% (v / v) of the disrupted strains of Example 2 and Comparative Examples 1 to 4 was added to check the expression levels of COX-2 and iNOS genes. A group that was not treated with any agent (MO: media only) was used as the negative control, and a group that was treated with LPS only was used as the positive control.

[0053] FIG. 1 is a graph showing the expression level of COX-2 mRNA depending on the type of lactic acid bacteria strain derived from oats. FIG. 2 is a graph showing the expression level of iNOS mRNA depending on the type of lactic acid bacteria strain derived from oats. As a result, as shown in Figures 1 and 2, it was confirmed that the lysate of Lacticaceae Bacillus rhamnosus ICL-46 strain had the most effective inhibitory effect on the expression of COX-2 and iNOS genes compared to other lactic acid bacteria strains derived from oats. From the above results, it was confirmed that the homogenate of Lacticase I Bacillus rhamnosus ICL-46 strain exhibits skin soothing effects due to its significantly excellent anti-inflammatory effects.

[0054] Experimental Example 2: Confirmation of anti-allergic effect - Inhibition of β-hexose aminase release To confirm the anti-allergic effect of the homogenate of Bacillus rhamnosus ICL-46 strain, we tested its inhibitory effect on the release of β-hexosaminidase, a key factor associated with allergy or immunity. Inhibition of β-hexosaminidase release means that it prevents the degranulation of immune cells.

[0055] Specifically, RBL-2H3 cells were uniformly seeded into a 24-well plate at 2 x 10 cells / well and stabilized at 37°C and 5% CO2 for 24 hours. After washing the cells with PBS, each well was treated with anti-DNP-IgE (50 ng / mL) and cultured for 24 hours. The anti-DNP-IgE-sensitized cells were washed twice with MEM medium supplemented with 1% FBS, and 160 μL of the medium was added and cultured at 37°C for 20 minutes. Subsequently, 0.5% (v / v) lysates of the bacterial strains of Example 2 and Comparative Examples 1 to 4 were added, and 20 μL of 1 μg / mL DNP-BSA was added. The cells were incubated at 37°C for 1 hour to stimulate granule formation, and the supernatant was centrifuged at 13,000 rpm at 4°C for 10 minutes. 50 μL of 0.1 M citrate buffer (containing 10 mM p-nitrophenyl-N-acetyl-β-D-glucosamide, pH 4.5) was added to 25 μL of the supernatant after centrifugation, and the mixture was incubated for 1 hour at 37° C. The reaction was then stopped by adding 100 μL of stop buffer (0.1 M NaCO / NaHCO, pH 10.0), and the absorbance was measured at 405 nm using a microplate reader.

[0056] FIG. 3 is a graph showing the β-hexose aminase inhibitory effect depending on the type of lactic acid bacteria strain derived from oats. As a result, as shown in Figure 3, it was confirmed that the disrupted product of Lacticase Bacillus rhamnosus ICL-46 strain had the most effective effect in reducing β-hexose aminase activity compared to other lactic acid bacteria strains derived from oats. From the above results, it was confirmed that the disrupted product of Lacticase I Bacillus rhamnosus ICL-46 exhibits significantly excellent anti-allergic or skin immunity enhancing effects.

[0057] Experimental Example 3: Confirmation of the effect of skin microbiota regulation Experiments were conducted to confirm the effect of the homogenate of Lacticase I Bacillus rhamnosus ICL-46 strain, which was confirmed to have the best anti-inflammatory and anti-allergic effects in Experimental Examples 1 and 2, on regulating skin microorganisms. Specifically, TSB (tryptic soy broth, BD Difco, USA) was used as the microbial medium. The bacterial growth was evaluated by inoculating the strain into the appropriate medium at a concentration of 1 x 10 CFU / mL and culturing at 37°C for 24 hours, followed by measuring the absorbance at 600 nm to compare the growth rate, or by measuring the actual number of viable bacteria using a serial dilution method. The growth rate was compared as a percentage based on an experimental group to which the same amount of phosphate-buffered saline as the test sample was added.

[0058] In a 12-well plate for co-culture of beneficial and harmful skin bacteria, 1 mL of 10% TSB was placed and inoculated with harmful bacteria (Pseudomonas aeruginosa, StapHylococcus aureus) at a concentration of 1 x 10 CFU / mL. A 0.4 μm cell culture insert was placed on top, and beneficial bacteria (StapHylococcus epidermidis) was inoculated into 1 mL of 10% TSB at a concentration of 1 x 10 CFU / mL. The 10% TSB was treated with 0.5% (v / v) Lacticase Bacillus rhamnosus ICL-46 homogenate from Example 2, followed by the experiment. A control group was treated with the same amount of phosphate-buffered saline. After 12 hours of incubation at 32°C, the number of bacteria in each group was counted and the growth rate was compared.

[0059] FIG. 4 is a graph comparing the growth of beneficial skin bacteria S. epidermidis and harmful skin bacteria P. aeruginosa when co-cultured and treated with homogenized Lacticase Bacillus rhamnosus ICL-46. FIG. 5 is a graph comparing the growth of beneficial skin bacteria S. epidermidis and harmful skin bacteria S. aureus when co-cultured and treated with disrupted Lacticase Bacillus rhamnosus ICL-46. As a result, as shown in Figures 4 and 5, it was confirmed that treatment with Lacticase Bacillus rhamnosus ICL-46 homogenate did not affect the growth of beneficial bacteria (S. epidermidis), but inhibited or reduced the growth of harmful bacteria (P. aeruginosa, S. aureus). From the above results, it was confirmed that the homogenate of Lacticase I Bacillus rhamnosus ICL-46 exhibited a significantly excellent effect of regulating skin microorganisms.

[0060] In summary, the oat-derived Lacticase Bacillus rhamnosus ICL-46 strain inhibits or reduces the expression of COX-2 and iNOS, and inhibits or reduces the release of β-hexaminase, a key factor associated with allergies and immunity. It has been confirmed that it has no effect on the growth of beneficial skin bacteria (S. epidermidis) but reduces the growth of harmful skin bacteria (P. aeruginosa, S. aureus). Therefore, the oat-derived Lacticase Bacillus rhamnosus ICL-46 strain has the effects of improving skin inflammation (anti-inflammatory), improving skin allergies (anti-allergic), enhancing skin immunity, soothing skin, and regulating skin microbiota.

[0061] The above description is merely an illustrative example of the technical concept of the present invention, and a person having ordinary knowledge in the technical field to which the present invention pertains can make various modifications or variations without departing from the essential characteristics of the present invention. [Accession number]

[0062] KCTC 15702BP TIFF2025114485000002.tif240170

Claims

1. Lacticaseibacillus rhamnosus ICL-46 strain, which has been deposited under accession number KCTC15702BP.

2. 2. The strain of claim 1, which is isolated from oats (Avena sativa).

3. The strain of claim 1, characterized in that it contains the 16S rRNA of SEQ ID NO:

1.

4. a step of inoculating the strain according to claim 1 into a medium, and culturing the medium to obtain a culture solution of the strain; centrifuging the culture medium to separate the cells and the supernatant; 2. The method for producing a disrupted bacterial strain product according to claim 1, further comprising the step of: mixing the disrupted bacterial cells with the supernatant to obtain a disrupted bacterial cell product.

5. A cosmetic composition for improving skin inflammation, alleviating skin allergies, enhancing skin immunity, or soothing skin, comprising one or more of the strain described in claim 1, its disrupted product, culture medium, extract of the culture medium, and fermentation medium.

6. A cosmetic composition for regulating skin microorganisms, comprising at least one of the strain of claim 1, its disrupted product, culture medium, extract of the culture medium, and fermentation medium.

7. 7. The cosmetic composition for regulating skin microorganisms according to claim 6, wherein the skin microorganism regulation is achieved by inhibiting the growth of harmful bacteria on the skin.

8. 8. The cosmetic composition according to claim 7, wherein the harmful bacteria on the skin are Pseudomonas aeruginosa or Staphylococcus aureus.

9. A pharmaceutical composition for preventing or treating skin inflammatory and allergic skin diseases or immune-mediated skin diseases, comprising one or more of the strain described in claim 1, its disrupted material, culture medium, extract of the culture medium, and fermentation medium.

10. The pharmaceutical composition according to claim 9, which inhibits or reduces the expression of one or more of COX-2 (cyclooxygenase-2) and iNOS (nitricoxide synthases), or inhibits or reduces the release of β-hexosaminidase.

11. A functional health food composition comprising at least one of the strain of claim 1, its disrupted product, culture medium, extract of the culture medium, and fermentation medium.

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