Novel lactobacillus acidophilus strain and use thereof

Abstract

The present invention relates to a novel Lactobacillus acidophilus strain. Also, the present invention relates to uses of the Lactobacillus acidophilus strain for antibacterial activity, antioxidation, the suppression of periodontal tissue damage or degradation, and anti-inflammation. Also, the present invention relates to a composition comprising the Lactobacillus acidophilus strain. The Lactobacillus acidophilus strain of the present invention can exhibit antibacterial activity against oral harmful bacteria that cause oral diseases, suppress the production of reactive oxygen species (ROS), inhibit the expression or activity of matrix metalloproteinase-3 (MMP-3) that causes periodontal tissue damage or degradation, and inhibit the expression or activity of inflammatory cytokines IL-6 and IL-8. Therefore, the strain and the composition comprising same, of the present invention, can be effectively used for antibacterial activity, antioxidation, the suppression of periodontal tissue damage or degradation, and anti-inflammation, and can also be used for treating, ameliorating or preventing oral diseases.

Description

Novel Lactobacillus acidophilus strain and uses thereof

[0001] This application claims the benefit of priority based on Korean Patent Application No. 10-2024-0125903 filed on September 13, 2024, and all contents disclosed in the document of said Korean patent application are incorporated herein as part of this specification.

[0002] The present invention relates to a novel strain of Lactobacillus acidophilus. Furthermore, the present invention relates to the antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory uses of the Lactobacillus acidophilus strain. Additionally, the present invention relates to a composition comprising the said Lactobacillus acidophilus strain.

[0003] Oral diseases, including dental caries, halitosis, stomatitis, gingivitis, and periodontitis, are common across all age groups. Gingivitis and periodontal disease affect approximately 30% of adults in South Korea and 10–15% of adults worldwide. According to the Health Insurance Review and Assessment Service, these conditions ranked first among frequently treated outpatient diseases in 2019–2020, with the number of patients confirmed at 18.09 million in 2022. Dental caries, another oral disease, occurs frequently in those under the age of 19 and was found to account for approximately 11% of dental outpatients in 2022. Furthermore, the symptom experience rate for stomatitis, an oral disease involving inflammation of the oral mucosa, increased from 47.9% in 2019 to 63.1% in 2023, showing an average annual growth rate of 7.1%.

[0004] The primary causes of such oral diseases are plaque and tartar, which provide an environment for the proliferation of harmful oral bacteria. These proliferated bacteria induce inflammatory responses and oxidative stress in oral tissues, and by causing plaque and tartar to regenerate within the destroyed tissues, they repeatedly generate and exacerbate oral diseases, ultimately leading to tooth loss and alveolar bone destruction. Therefore, to treat, improve, or prevent oral diseases, it is necessary to prevent the proliferation of harmful oral bacteria, alleviate induced inflammation or oxidative stress, inhibit the damage or degradation of periodontal tissues, and establish a healthy ecological environment within the oral cavity. Accordingly, research on various types of oral lactic acid bacteria with antibacterial, antioxidant, inhibitory, and anti-inflammatory effects is currently underway; however, the need to utilize the microbiome for the treatment, prevention, or improvement of oral diseases remains significant within the technical field to which this invention belongs.

[0005] Accordingly, the inventors have completed the present invention by experimentally demonstrating, through various studies, that a specific strain of Lactobacillus acidophilus exhibits antibacterial activity against oral harmful bacteria that cause oral diseases, inhibits the production of reactive oxygen species (ROS) induced by LPS (Lipopolysaccharide) of oral harmful bacteria, and inhibits the expression or activity of MMP-3 (Matrix metalloproteinase-3), a proteolytic enzyme and marker of damage or degradation of periodontal tissue, and IL-6 (Interleukin-6) and IL-8 (Interleukin-8), inflammatory cytokines, thereby exhibiting antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory effects.

[0006] Each description and embodiment disclosed in the present invention may be applied to each other description and embodiment. That is, all combinations of the various elements disclosed in the present invention fall within the scope of the present invention. Furthermore, the scope of the present invention should not be considered limited by the specific descriptions provided below.

[0007] Furthermore, terms not specifically defined in this specification should be understood to have the meanings commonly used in the technical field to which the present invention pertains. Additionally, unless specifically defined in the context, the singular includes the plural, and the plural includes the singular.

[0008]

[0009] The present invention provides a novel Lactobacillus acidophilus strain. The novel Lactobacillus acidophilus strain according to the present invention was deposited by the applicant with the Korea Research Institute of Biotechnology and Bioengineering on April 11, 2023, under accession number KCTC 15402BP. The Korea Research Institute of Biotechnology and Bioengineering is a patent strain deposit institution located at 181, Ipsin-gil, Jeongeup-si, Jeollabuk-do, Republic of Korea.

[0010] The above novel strain is a rod-shaped bacterium belonging to the species Lactobacillus acidophilus, and has a 16S rRNA gene containing the nucleotide sequence represented by SEQ ID NO. 1.

[0011]

[0012] In this specification, "Lactobacillus acidophilus strain" includes live cells, dead cells and cultures thereof, fermented products thereof, lysed products thereof, extracts thereof, and cytoplasmic fractions obtained by lysing the same, and also includes post-processed or post-treated products such as filtration, concentration, drying, extraction, and freezing of said cultures, fermented products, lysed products, extracts, cytoplasmic fractions, etc.

[0013] The term "culture" as used in this specification refers to the entire medium containing the Lactobacillus acidophilus strain of the present invention obtained by culturing the strain for a certain period in a nutrient-supplied medium, metabolites of said strain, or excess nutrients remaining after culturing the strain, and includes the culture medium or culture supernatant (supernatant) from which the strain has been removed after culturing said strain. Additionally, it includes a concentrate of said entire medium or culture medium, or a dried concentrate. Specifically, the culture of the present invention may use a medium easily selected by a person skilled in the art according to the purpose among media used for microbial culture, for example, a medium used for culturing Lactobacillus acidophilus, such as MRS (De Man Rogosa, Sharpe agar) medium or blood agar medium, but is not limited thereto as long as said strain can be cultured.

[0014] The Lactobacillus acidophilus strain of the present invention exhibits antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory effects.

[0015] As used in this specification, the term "antimicrobial" means inhibiting the proliferation of harmful bacteria or inhibiting the activity of harmful bacteria.

[0016] As used herein, the term "antioxidant" means inhibiting the oxidation of cells by free radicals or reactive oxygen species (ROS), and includes the reduction of cell damage caused by the removal of free radicals or reactive oxygen species.

[0017] As used herein, the term "periodontal tissue" refers to connective tissue that surrounds and supports the teeth, such as the gingiva (gums), periodontal ligament, or alveolar bone tissue.

[0018] As used in this specification, the term "anti-inflammatory" means inhibiting the occurrence of inflammation or eliminating or reducing inflammation.

[0019] The Lactobacillus acidophilus strain of the present invention may have one or more of the following characteristics:

[0020] (a) Inhibition of the production of reactive oxygen species (ROS),

[0021] (b) inhibition of expression or activity of MMP-3 (Matrix metalloproteinase-3), and

[0022] (c) Inhibition of expression or activity of IL-6 (Interleukin-6) or IL-8 (Interleukin-8).

[0023] IL-6 and IL-8 are inflammatory cytokines whose expression or activity is induced by external stimuli that cause inflammation, such as microbial infection or tissue damage. These inflammatory cytokines generate nitric oxide and reactive oxygen species, thereby inducing oxidative stress, and it is known that the generated nitric oxide and reactive oxygen species promote cell damage and inflammation (Roman Fischer et al. (2015), Interrelation Oxidative Stress and Inflammation in Neurodegenerative Disease: Role of TNF, Oxidative Medicine and Cellular Longevity, 2015:610813). In addition, MMPs are involved in the degradation of the extracellular matrix within periodontal tissues and are the most important pathway in tissue destruction associated with periodontal disease; it is known that harmful oral bacteria in plaque stimulate host cells to increase MMP release (Checchi et al. (2020), The Role of Matrix Metalloproteinases in Periodontal Disease, Int J Environ Res Public Health, 17(14):4923).

[0024] Accordingly, since the inhibition of the generation of the above-mentioned reactive oxygen species, the inhibition of the expression or activity of MMP-3, and the inhibition of the expression or activity of IL-6 or IL-8 exhibit antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory effects, the Lactobacillus acidophilus strain of the present invention can be usefully used as an active ingredient in a composition for the treatment, improvement, or prevention of oral diseases.

[0025] In addition, the Lactobacillus acidophilus strain of the present invention may have antibacterial activity against harmful oral bacteria. Accordingly, the Lactobacillus acidophilus strain can be usefully used as an active ingredient in an antibacterial composition.

[0026] The aforementioned harmful oral bacteria include Streptococcus mutans, Streptococcus sobrinus, Porphyromonas gingivalis, Porphyromonas catoniae, Fusobacterium nucleatum, Actinomyces viscosus, Eikenella corrodens, Streptococcus sanguis, Streptococcus mitis, Streptococcus ratti, Streptococcus criceti, Streptococcus gordonii, and Streptococcus It may be, but is not limited to, Streptococcus anginosus, Streptococcus oralis, Treponema denticola, Prevotella intermedia, Bacteroides forsythus, Tannerella forsythus, Aggregatibacter actinomycetemcomitans, or Candida albicans, and may include any bacteria that contribute to the development of oral disease.

[0027]

[0028] Another aspect of the present invention provides a composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory purposes, comprising a Lactobacillus acidophilus strain deposited as KCTC 15402BP.

[0029] In the composition according to the present invention, each term has the same meaning as described above unless specifically stated otherwise.

[0030] The Lactobacillus acidophilus strain of the present invention or a composition containing the same can be used for treating, improving, or preventing oral diseases.

[0031] As used herein, the term “oral disease” refers to a disease occurring in the tissues within the oral cavity, such as the lips, tongue, lingual papillae, teeth, gingiva, alveolar bone, inner cheek, hard and soft palate, uvula, lingual tonsils, and palatine tonsils, and may be caused by infection by harmful oral bacteria, inflammation resulting therefrom, or damage or breakdown of periodontal tissues. The oral disease may be dental caries, bad breath, or periodontal disease, but is not limited thereto.

[0032] As used herein, the term "dental caries" refers to a disease in which the hard tissues of a tooth are eroded and lost. Symptoms include the dissolution and destruction of a portion of the tooth's calcified tissue, which may cause pain and tooth loss. Dental caries may be caused by plaque, a bacterial film formed on the tooth surface. In this specification, the term "dental caries" may be used interchangeably with the term "cavity."

[0033] As used in this specification, the term "halitosis" refers to an odor originating from the oral cavity and adjacent organs, which may be caused by substances produced when proteins, food debris, and other substances mixed in saliva are decomposed by microorganisms within the oral cavity. The primary cause of bad breath is volatile sulfur compounds, 90% of which consist of hydrogen sulfide derived from cysteine ​​and methylmercaptan and dimethyl sulfide derived from methionine. These components are primarily produced by protein enzymes secreted by anaerobic bacteria, which mainly inhabit the back of the tongue. The back of the tongue is not effectively cleaned by saliva and contains many small depressions, making it an easy habitat for bacteria and thus allowing bad breath to easily occur. In addition, bad breath can also be caused by other oral diseases such as dental caries, periodontitis, and xerostomia.

[0034] As used in this specification, the term "periodontial disease" refers to any disease of the periodontal tissues surrounding and supporting the teeth, such as the gingiva (gums), periodontal ligament, or alveolar bone tissue. The periodontal disease may be caused by microbial infection, inflammation, or damage or degradation of the periodontal tissues. When plaque and tartar, which is formed when the plaque hardens, accumulate on the teeth, the gingiva and the teeth separate; microbial infection and inflammation occur between them, forming a periodontal pocket. As the depth of the periodontal pocket increases, inflammation spreads to the periodontal ligament, leading to alveolar bone loss and the development of periodontal disease. The above periodontal disease may be oral inflammation such as gingivitis, periodontitis, stomatitis, pulpitis, periapical disease, periodontal abscess, periapical abscess with sinus, pericoronitis, and pericementitis, alveolar bone osteodystrophy, or gingival retraction, but is not limited thereto. In this specification, the term "periodontal disease" may be used interchangeably with the term "gum disease."

[0035] As used herein, the term "gingivitis" refers to inflammation occurring in the soft tissue of the gingiva (gum).

[0036] As used herein, the term "periodontitis" refers to inflammation occurring in the gingiva as well as in the periodontal ligament, alveolar bone, or cementum. Such inflammation damages the gingiva, periodontal ligament, alveolar bone, or cementum and can further lead to tooth loss. An example of such alveolar bone damage is alveolar bone osteodystrophy.

[0037] As used herein, the term "alveolar bone osteodystrophy" refers to a disease resulting from a decrease in alveolar bone mass due to an imbalance between bone formation and bone resorption in the alveolar bone, and includes alveolar bone osteoporosis, alveolar bone osteomalacia, and alveolar bone osteopenia.

[0038] As used in this specification, the term "stomatitis" refers to a disease in which inflammation occurs in the oral mucosa, such as the tongue, gums, lips, or inside of the cheeks, due to an infection by microorganisms in the oral cavity. The stomatitis includes candidal stomatitis (oral candidiasis, thrush) caused by Candida albicans infection, herpetic gingivostomatitis caused by the herpes simplex virus, and aphthous stomatitis caused by a decrease in immunity. In this specification, the term "stomatitis" may be used interchangeably with the term "oral mucosal ulcer."

[0039] As used herein, the term "pulpitis" refers to a disease in which the dental pulp becomes infected by microorganisms or inflamed. The pulp is the innermost part of the tooth, where nerves and blood vessels are located. The most common causes of pulpitis are infection and tooth damage resulting from dental caries, and pulpitis causes severe toothache.

[0040] As used herein, the term "periapical disease" refers to a disease caused by inflammation of the tooth root and surrounding tissues resulting from the progression of pulpitis.

[0041] As used in this specification, the term "periodontial abscess" refers to a purulent inflammation that occurs within the periodontal tissue and may occur due to oral microorganisms, dental trauma, or when the drainage of inflammatory secretions from the periodontal pocket is not smooth.

[0042] As used in this specification, the term "periapical abscess with sinus" refers to inflammation occurring around the root (apex) of a tooth, which can occur when dental caries becomes severe and inflammation develops in the nerve within the tooth. Symptoms of an oral abscess include toothache, tooth sensitivity (sensitive teeth), swelling of the tooth root, or gingival fistula. In this specification, the term "periapical abscess" may be used interchangeably with the term "apical abscess."

[0043] As used herein, the term "pericoronitis" refers to inflammation caused by microbial infection of the gingiva (gums) or surrounding tissues around a wisdom tooth. Symptoms of pericoronitis include bad breath, pain, and congestion or pus in the gingiva or surrounding tissues. In this specification, the term "pericoronitis" may be used interchangeably with the term "pericoronitis."

[0044] As used herein, the term "pericementitis" refers to inflammation of the periodontal ligament, which is a membrane surrounding the tooth roots. Pericementitis damages or destroys the tooth and surrounding tissues, thereby causing tooth loosening.

[0045] The term "gingival retraction" as used in this specification refers to a condition in which the gingival (gum) tissue that anchors the teeth is damaged or broken down, exposing the tooth roots. Such gingival retraction can cause tooth sensitivity (sensitive teeth) or tooth decay.

[0046] As used in this specification, the term "treatment" refers to any act in which symptoms of a disease are improved or completely cured by the administration, ingestion, or application of the composition according to the present invention.

[0047] Furthermore, as used herein, the term "improvement" refers to any act in which the symptoms of a disease are at least reduced or improved or benefited by the administration, ingestion, or application of the composition according to the present invention.

[0048] Furthermore, as used herein, the term "prevention" refers to any act of suppressing or delaying the symptoms of a disease by administering, ingesting, or applying the composition according to the present invention.

[0049] As used herein, the term "administration" means physically introducing the composition to a subject using any of the various methods and delivery systems known to a person skilled in the art. Routes of administration for the composition of the present invention include, but are not limited to, oral administration routes, or oral administration such as sublingual administration, administration through the oral mucosa, oral spray, oral application, or intraoral injection, inhalation, intravenous, intramuscular, subcutaneous, intraperitoneal, spinal, or other parenteral administration routes, such as injection, infusion, nasal spray, or application. The number of administrations for the composition of the present invention may be, for example, one time, multiple times, and / or over one or more extended periods.

[0050] The content of the active ingredient included in the composition according to the present invention can be appropriately determined by considering various factors such as the purpose of use (prevention, improvement, or therapeutic treatment), duration of use, type of formulation, route of administration, condition of the subject, symptoms of the disease, degree of progression of symptoms, age, and gender. For example, the content of the Lactobacillus acidophilus strain of the present invention may be included in an amount of about 0.0001 to 100 weight% based on the total weight of the composition.

[0051] As used herein, the term "subject" includes humans or any non-human animals, and said non-human animals may be vertebrates, such as primates, dogs, cattle, horses, pigs, rodents, such as mice, rats, guinea pigs, etc. In this specification, said "subject" is used interchangeably with "individual" and "patient." There are no restrictions on gender, age, etc. for subjects using the composition according to the present invention. Specifically, it may be used by any subject who wishes to obtain antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory effects.

[0052]

[0053] According to one embodiment of the present invention, a pharmaceutical composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory effects is provided, comprising the Lactobacillus acidophilus strain of the present invention.

[0054] In the pharmaceutical composition according to the present invention, each term has the same meaning as the foregoing unless specifically stated otherwise.

[0055] The above pharmaceutical composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory purposes may be a pharmaceutical composition used for the treatment or prevention of oral diseases.

[0056] The pharmaceutical composition of the present invention may be prepared in a unit dose form or contained in a multi-dose container by formulation using a pharmaceutically acceptable carrier and / or excipient, according to a method that can be easily carried out by a person skilled in the art to which the invention belongs. In this case, the formulation may be in the form of a solution, suspension, syrup, or emulsion in an oil or aqueous medium, or may be in the form of an extract, powder, powder, granule, tablet, or capsule, and may additionally include a dispersant or a stabilizer.

[0057] The pharmaceutical composition according to the present invention can be formulated into various oral, intraoral, or parenteral administration forms.

[0058] Formulations for oral administration or administration within the mouth may include tablets, pills, hard / soft capsules, liquids, suspensions, emulsifiers, syrups, granules, elixirs, etc., and these formulations may use one or more diluents or excipients such as fillers, extenders, humectants, disintegrants, lubricants, binders, and surfactants that are commonly used in addition to the strain according to the present invention. Agar, starch, alginic acid or its sodium salt, calcium monohydrogen phosphate, etc. may be used as disintegrants; silica, talc, stearic acid or its magnesium or calcium salt, polyethylene glycol, etc. may be used as lubricants; and magnesium aluminum silicate, starch paste, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose, polyvinylpyrrolidine, low-substituted hydroxypropylcellulose, etc. may be used as binders. In addition, lactose, dextrose, sucrose, mannitol, sorbitol, cellulose, glycine, etc. may be used as diluents, and in some cases, generally known absorbents, coloring agents, flavoring agents, sweeteners, etc. may be used together.

[0059] In addition, parenteral formulations may include injections, creams, lotions, topical ointments, oils, moisturizers, gels, sprays, topicals, aerosols, oral patches, dressing solutions, and nasal inhalers, and these formulations may use one or more commonly used carriers, stabilizers, or preservatives in addition to the active ingredient. Carriers may include water, suitable oils, saline solution, aqueous glucose, or glycol; stabilizers may include antioxidants such as sodium bisulfite, sodium sulfite, or ascorbic acid; and preservatives may include benzalkonium chloride, methyl-paraben, propyl-paraben, or chlorobutanol.

[0060] The above pharmaceutical composition may be sterilized or contain preservatives, stabilizers, hydrating agents or emulsification promoters, salts for osmotic pressure regulation, buffers, and other therapeutically useful substances, and may be formulated according to commonly used mixing, granulation, or coating methods.

[0061] The above-mentioned Lactobacillus acidophilus strain may be included alone, or may further include a suitable carrier, excipient, or diluent commonly used in the preparation of pharmaceutical compositions. Specifically, the carrier, excipient, and diluent that may be included in the above-mentioned pharmaceutical composition may be, for example, lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil, but are not limited thereto. These may be used alone or in a mixture of two or more. In addition, the above pharmaceutical composition may further include other conventional additives such as antioxidants, buffers and / or bacteriostatic agents if necessary, and may additionally include dispersants, surfactants, binders, lubricants, etc. to be formulated into powders, granules, tablets, liquids, capsules, suspensions, emulsions, syrups, creams, lotions, gels, ointments, aerosols, powder sprays, creams, suppositories, pills, etc.

[0062]

[0063] According to another embodiment of the present invention, a quasi-drug composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory purposes is provided, comprising the Lactobacillus acidophilus strain of the present invention.

[0064] In the composition of a quasi-drug according to the present invention, each term has the same meaning as described above unless specifically stated otherwise.

[0065] The above-mentioned antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory quasi-drug composition may be a quasi-drug composition used for the improvement or prevention of oral diseases.

[0066] The quasi-drug composition according to the present invention can be manufactured in the form of general emulsion formulations and solubilized formulations, etc., using commonly known manufacturing methods. At this time, the quasi-drug composition of the present invention can be manufactured in various formulations such as patches, ointments, gels, creams, sprays, detergents, disinfectants, sanitary pads, wet wipes, masks, gauze, absorbent cotton, adhesive bandages, bandages, solid forms, liquid forms, etc., and commonly used manufacturing methods for quasi-drugs may be applied. Furthermore, the types of the above quasi-drugs are not particularly limited. Examples include toothpaste, oral spray, mouthwash, mouthwash, oral rinse, oral ointment, mouthwash, gum massage cream, cleanser, wet wipe, shower foam, soap, mask, gauze, absorbent cotton, adhesive bandage, eye mask, bandage, repellent, disinfectant, detergent, etc., and include all quasi-drugs in the conventional sense.

[0067] The quasi-drug composition according to the present invention may further include additional components in addition to the strain to enhance efficacy. For example, there are no limitations on the additional components provided that they do not offset or reduce the efficacy of the strain according to the present invention. Optionally, carriers, excipients, diluents, etc., commonly used in the field of quasi-drugs may be further included. For example, the carrier, excipient, or diluent may be a filler, extender, binder, wetting agent, disintegrant, surfactant, lubricant, sweetener, fragrance, preservative, etc.

[0068] Optionally, ingredients typically used to add or enhance the function of the quasi-drug may also be added. For example, the quasi-drug composition may additionally include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, maltitol, starch, gelatin, glycerin, acacia gum, alginate, calcium phosphate, calcium carbonate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, mineral oil, propylene glycol, polyethylene glycol, vegetable oil, injectable ester, Witepsol, macrogol, Tween 61, cocoa dough, laurize, etc.

[0069]

[0070] According to another embodiment of the present invention, a food composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory properties is provided, comprising the Lactobacillus acidophilus strain of the present invention.

[0071] In the food composition according to the present invention, each term has the same meaning as the foregoing unless specifically stated otherwise.

[0072] The above foods include processed forms of all natural materials, such as nutraceutical foods, medical foods, health functional foods, nutritional supplements, and food additives.

[0073] The above-mentioned food composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory purposes may be a food composition used for the improvement or prevention of oral diseases.

[0074] As used in this specification, the term "Nutraceutical food" refers to a food prepared from raw materials or ingredients that perform a beneficial function for a subject, and which improves or maintains the subject's health by maintaining normal function or activating physiological function.

[0075] As used in this specification, the term "medical food" refers to a food for special medical purposes, specifically a food in a special formulation for the specific dietary management of a subject suffering from a disease, and includes foods for special medical purposes prescribed by the Ministry of Food and Drug Safety of the Republic of Korea. The medical food alleviates the subject's disease or symptoms by satisfying specific nutritional requirements that cannot be achieved by a regular diet alone, and may be consumed as part of a meal or as a meal replacement. The special formulation is a formulation that can be taken orally or via enteral nutrition, and enteral nutrition refers to supplying nutrition by inserting a nasogastric tube into the gastrointestinal tract through the nose. In this specification, the term "medical food" may be used interchangeably with the term "medical food."

[0076] As used in this specification, the term "health functional food" refers to a food manufactured and processed in the form of tablets, capsules, powders, granules, liquids, pills, etc., using raw materials or ingredients that have functional properties useful to the human body. Herein, "functionality" means obtaining effects useful for health purposes, such as regulating nutrients or physiological actions on the structure and function of the human body.

[0077] The types of the above foods are not specifically limited. Specific examples include yogurt, dairy products, meat, sausage, bread, chocolate, candies, snacks, confectionery, pizza, ramen, other noodles, chewing gum, dairy products including ice cream, various soups, beverages, tea, drinks, alcoholic beverages, vitamin complexes, etc., and include all foods in the conventional sense.

[0078] The food composition according to the present invention can be manufactured by methods commonly used in the art, and during such manufacturing, raw materials and ingredients commonly added in the art may be added. The added ingredients include, for example, proteins, carbohydrates, fats, nutrients, seasonings, and flavoring agents. The carbohydrates include all conventional sugars such as glucose, fructose, maltose, sucrose, oligosaccharides, dextrin, cyclodextrin, etc., or sugar alcohols such as xylitol, sorbitol, erythritol, etc. As flavoring agents, natural flavoring agents such as taumatin and stevia extract, or synthetic flavoring agents such as saccharin and aspartame may be used. For example, when the food composition is manufactured as a drink, in addition to the strain according to the present invention, citric acid, liquid fructose, sugar, glucose, acetic acid, malic acid, fruit juice, jujube extract, or licorice extract may be additionally included, and various natural carbohydrates, etc., may be included as additional ingredients, as in conventional beverages.

[0079] In addition, unlike general pharmaceuticals, when food is used as a raw material, there is an advantage of not having side effects that may occur during long-term use of the drug, and since it is highly portable, the food composition of the present invention can be used as an adjuvant to enhance or improve the preventive or therapeutic effects of oral diseases, and it is also possible to use it simultaneously or sequentially with the pharmaceutical composition, quasi-drug composition, and / or other composition or other therapy according to the present invention for the purpose of maximizing the above effects.

[0080] The content of the active ingredient included in the food composition of the present invention may be appropriately determined according to the purpose of use (prevention, improvement, or therapeutic treatment), the duration of use, the condition of the subject, etc. For example, when manufacturing food, the content of the strain according to the present invention may be included in the food composition in an amount of 0.001 to 20 weight%, 0.001 to 15 weight%, or 0.001 to 10 weight%. In the case of health drinks, it may be included in an amount of 0.01 to 2 g, specifically 0.02 to 2 g, and more specifically 0.3 to 1 g based on 100 ml. However, in the case of long-term consumption for the purpose of health and hygiene or health control, it may be used in an amount less than the above range. During the process of manufacturing the food composition of the present invention, the content of the active ingredient added to the food composition may be appropriately increased or decreased as needed.

[0081]

[0082] According to another embodiment of the present invention, a cosmetic composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory properties is provided, comprising the Lactobacillus acidophilus strain of the present invention.

[0083] In the cosmetic composition according to the present invention, each term has the same meaning as described above unless specifically stated otherwise.

[0084] The above-mentioned antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory cosmetic composition may be a cosmetic composition used for the improvement or prevention of oral diseases.

[0085] The cosmetic composition according to the present invention can be prepared in the form of a general emulsion formulation and a solubilized formulation, etc., using a commonly known manufacturing method. Specifically, it can be prepared in various formulations such as patches, ointments, skin adhesive gels, creams, packs, lotions, essences, sprays, masks, foundations, makeup bases, cleansers, water (W) type, oil (O) type, silicone (S) type, oil-in-water (O / W) type, water-in-oil (W / O) type, silicone-in-water (W / S) type, silicone-in-water (S / W) type, solid phase, liquid phase, etc. In addition, it can be prepared in the form of a foam or in the form of an aerosol containing a compressed propellant.

[0086] The cosmetic composition according to the present invention may further include additional ingredients in addition to the active ingredients. There are no limitations on the additional ingredients provided that they do not offset or reduce the antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory effects. Optionally, adjuvants, carriers, etc., commonly used in the field of cosmetics may be further included. For example, it may be one or more selected from fatty substances, organic solvents, solvents, thickeners, gelling agents, softeners, antioxidants, suspending agents, stabilizers, foaming agents, fragrances, surfactants, water, ionic or nonionic emulsifiers, thickeners, humectants, liquid crystal film strengtheners, pH adjusters, antimicrobial agents, water-soluble polymers, film-forming agents, amino acids, organic amines, polymer emulsions, pH adjusters, skin nutrients, antioxidants, antioxidant aids, preservatives, fragrances, oils and fats, waxes, hydrocarbon oils, higher fatty acid oils, higher alcohols, synthetic ester oils, silicone oils, fillers, metal ion chelating agents, opacifiers, anti-foaming agents, bulking agents, skin softeners, skin conditioning agents, chelating agents, preservatives, vitamins, blockers, humectants, essential oils, dyes, pigments, hydrophilic or lipophilic active agents, and lipid vesicles. The above-mentioned adjuvants and carriers are introduced in amounts commonly used in the fields of cosmetics or dermatology.

[0087]

[0088] According to another embodiment of the present invention, a feed composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory purposes is provided, comprising the Lactobacillus acidophilus strain of the present invention.

[0089] In the feed composition according to the present invention, each term has the same meaning as the foregoing unless specifically stated otherwise.

[0090] The above-mentioned feed composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory purposes may be a feed composition used for the improvement or prevention of oral diseases.

[0091] The above feed is a substance that supplies organic or inorganic nutrients necessary for the maintenance of life and rearing of subjects other than humans, and includes both feed and feed additives.

[0092] The types of the above feed are not particularly limited. Specific examples include grains, plant-based protein feeds, animal-based protein feeds, dairy products, nutritional supplements, digestion and absorption enhancers, growth promoters, etc., and include all feed or feed additives in the conventional sense. The above grains include, for example, ground or crushed rice, wheat, oats, barley, or corn. The above plant-based protein feed includes, for example, feeds with rapeseed (rapeseed), sunflower, or soybeans as the main components. The above animal-based protein feed includes, for example, feeds with blood meal, bone meal, meat meal, or fish meal as the main components. The above dairy products include, for example, feeds with powdered milk or whey powder as the main components.

[0093] The feed composition according to the present invention can be manufactured by a method commonly used in the art, and in addition to the active ingredient, the composition may further include a suitable carrier, preservative, stabilizer, wetting agent, emulsifier, solution accelerator, excipient, organic acid, phosphate, antioxidant, etc. commonly used in the art. Specifically, the carrier, preservative, stabilizer, wetting agent, emulsifier, solution accelerator, excipient, organic acid, phosphate, antioxidant that may be included in the feed composition may be one or more selected from, for example, corn starch, lactose, sucrose, soybean flakes, olive oil, sesame oil, peanut oil, propylene glycol, lactic acid, malic acid, citric acid, fumaric acid, adipic acid, sodium phosphate, potassium phosphate, polyphenol, vitamin C, green tea extract, licorice extract, tocopherol, tannic acid, and chitosan. The above carrier, preservative, stabilizer, wetting agent, emulsifier, solution accelerator, excipient, organic acid, phosphate, antioxidant, etc. are introduced in amounts commonly used in the industry.

[0094] The feed composition according to the present invention may be formulated using commonly known manufacturing methods. For example, it may be in the form of a powder, liquid, or granular formulation. The content of the active ingredient included in the feed composition of the present invention may be appropriately determined according to the purpose of use (prevention, improvement, or therapeutic treatment), duration of use, condition of the subject, etc. For example, when manufacturing the feed, the content of the strain according to the present invention may be 20 to 90 weight percent of the feed composition. The feed composition may be used by immersion, spraying, or mixing with other feed compositions.

[0095]

[0096] Another aspect of the present invention provides a method for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory, comprising the step of administering to a subject a composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory, the composition comprising a Lactobacillus acidophilus strain.

[0097] In the antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory methods according to the present invention, each term has the same meaning as the foregoing unless specifically stated otherwise.

[0098] According to one embodiment of the present invention, a method for treating, improving, or preventing an oral disease of a subject is provided, comprising the step of administering the composition to the subject.

[0099] The composition according to the present invention contains an effective amount of the Lactobacillus acidophilus strain of the present invention and can be administered to a subject who requires treatment or prevention of oral diseases.

[0100] The above effective amount may be a "therapeutic effective amount" or a "preventive effective amount."

[0101] As used herein, the term “therapeutic effective dose” means any amount in which, when a drug or therapeutic agent is used alone or in combination with other therapeutic agents, it may result in a reduction in the severity of disease symptoms, an increase in the frequency and duration of symptom-free periods, or the prevention of damage or disability caused by disease suffering.

[0102] As used herein, the term "preventive effective dose" refers to any amount that suppresses the occurrence or recurrence of disease in a subject. The level of said effective dose may be determined based on factors including the subject's severity, age, gender, drug activity, sensitivity to the drug, time of administration, route of administration and elimination rate, duration of treatment, concurrently used drugs, and other factors well known in the medical field.

[0103] The dosage of the above composition may vary depending on the subject's age, gender, body weight, formulation method, administration route, or severity of the disease, excretion rate, administration time, pathological condition, and response sensitivity, for example, depending on the subject's symptoms, the composition of the present invention may be administered at a dose of 0.1 to 100 mg / kg once or several times a day, or at intervals of several days to several months.

[0104] In the composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory effects according to the present invention, the composition may be administered to a subject simultaneously, sequentially, or individually with other components exhibiting antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory effects. The "simultaneous" administration means administering the composition and other components at once as a single preparation, or administering the composition and other components at once as separate preparations; in this case, the administration route of the composition and the administration route of the other components may differ from each other. The "sequential" administration means administering the composition and other components relatively continuously, allowing for the minimum possible time consumed in the administration interval. The "individual" administration means administering the composition and other components at regular time intervals. The method of administration of the composition and other components may be appropriately selected by an expert in the art, taking into consideration the efficacy and side effects on the subject.

[0105] The composition of the present invention may be administered in combination with other therapeutic agents. In this case, the composition of the present invention and the other therapeutic agents may be administered simultaneously, sequentially, or individually. The other therapeutic agents may be drugs, such as compounds or proteins, having effects of preventing, treating, and / or improving oral diseases, but are not limited thereto.

[0106] In addition, the above composition may be formulated to be administered simultaneously, sequentially, or individually with other therapeutic agents. For example, the strain and other therapeutic agents may be administered simultaneously as a single formulation, or they may be administered simultaneously, sequentially, or individually as separate formulations. To administer simultaneously, sequentially, or individually, the strain and other therapeutic agents included in the composition of the present invention may be formulated separately in individual containers, or formulated together in the same container. Furthermore, the pharmaceutically effective amount, administration time, administration interval, administration route, treatment period, etc., of the strain and other therapeutic agents included in the composition of the present invention may be the same or different from each other.

[0107]

[0108] Another aspect of the present invention provides the use of a Lactobacillus acidophilus strain deposited at KCTC 15402BP, or a composition comprising said strain, for treating, improving, or preventing oral diseases. In the use of the Lactobacillus acidophilus strain or a composition comprising said strain according to the present invention, each term has the same meaning as above unless specifically stated otherwise.

[0109] According to one embodiment of the present invention, the strain or composition may be used in pharmaceuticals, quasi-pharmaceuticals, cosmetics, or food for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, or anti-inflammatory purposes.

[0110] The Lactobacillus acidophilus strain of the present invention exhibits antibacterial activity against oral harmful bacteria that cause oral diseases, inhibits the generation of reactive oxygen species (ROS), inhibits the expression or activity of MMP-3 (Matrix metalloproteinase-3) which causes damage or degradation of periodontal tissue, and inhibits the expression or activity of inflammatory cytokines IL-6 and IL-8. Accordingly, the strain of the present invention and the composition containing it can be usefully utilized for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory purposes, and can also be utilized for the treatment, improvement, or prevention of oral diseases.

[0111] Figure 1 is a photograph showing the hemolytic result of the Lactobacillus acidophilus strain of the present invention, taken after spreading the strain on a blood agar medium (Sheep blood agar) and culturing it.

[0112] Figure 2 shows the cytotoxicity results of the Lactobacillus acidophilus strain of the present invention. NONE represents negative control group 1, which was not treated with LPS (Lipopolysaccharide) derived from Porphyromonas gingivalis, dexamethasone, and the sample; CON represents negative control group 2, which was treated with LPS; DEXA represents positive control group, which was treated with LPS and dexamethasone; 10 MOI represents experimental group 1, which was treated with LPS and 10 MOI of Lactobacillus acidophilus strain powder as the sample; 50 MOI represents experimental group 2, which was treated with LPS and 50 MOI of Lactobacillus acidophilus strain powder as the sample; and 100 MOI represents experimental group 3, which was treated with LPS and 100 MOI of Lactobacillus acidophilus strain powder as the sample.

[0113] Figure 3 shows the results of inhibiting the production of reactive oxygen species (ROS) by the Lactobacillus acidophilus strain of the present invention. Each term has the same meaning as defined in Figure 2.

[0114] Figure 4 shows the results of inhibiting MMP-3 (Matrix metalloproteinase-3) expression of the Lactobacillus acidophilus strain of the present invention. Each term has the same meaning as defined in Figure 2.

[0115] Figure 5 shows the results of IL-6 (Interleukin-6) expression inhibition by the Lactobacillus acidophilus strain of the present invention. Each term has the same meaning as defined in Figure 2.

[0116] Figure 6 shows the results of IL-8 (Interleukin-8) expression inhibition by the Lactobacillus acidophilus strain of the present invention. Each term has the same meaning as defined in Figure 2.

[0117] Figure 7 shows the antimicrobial activity results of the Lactobacillus acidophilus strain of the present invention, the Lactobacillus acidophilus ATCC 4356 strain, and the LA5 strain. KCTC 15402BP is used as a sample containing 5 x 10 of the original powder of the Lactobacillus acidophilus strain of the present invention. 8 This represents Experimental Group 1 treated to a CFU / well, and ATCC 4356 is the sample, consisting of 5x10 original powder of the Lactobacillus acidophilus ATCC 4356 strain. 8 This represents Experimental Group 2 treated to a CFU / well, where LA5 is the original powder of Lactobacillus acidophilus LA5 strain as the sample at a rate of 5 x 10 8 This represents experimental group 3 treated to a CFU / well, and P represents the significance (P value).

[0118] Figure 8 shows the results of ROS production inhibition of the Lactobacillus acidophilus strain of the present invention, the Lactobacillus acidophilus JCM1021 strain, the ATCC 4356 strain, and the LA5 strain. NONE represents Negative Control Group 1, which was not treated with LPS, dexamethasone, or the sample; CON represents Negative Control Group 2, which was treated with LPS; DEXA represents Positive Control Group, which was treated with LPS and dexamethasone; KCTC 15402BP represents Experimental Group 1, which was treated with LPS and a suspension of the Lactobacillus acidophilus strain of the present invention as the sample at a concentration of 100 MOI per cell; JCM1021 represents Experimental Group 2, which was treated with LPS and a suspension of the Lactobacillus acidophilus JCM1021 strain as the sample at a concentration of 100 MOI per cell; ATCC 4356 represents Experimental Group 3, which was treated with LPS and a suspension of the Lactobacillus acidophilus ATCC 4356 strain as the sample at a concentration of 100 MOI per cell; and LA5 represents LPS and as the sample This represents experimental group 4, treated with a suspension of Lactobacillus acidophilus LA5 strain at 100 MOI per cell number, and P represents the significance (P value).

[0119] Figure 9 shows the results of MMP-3 expression inhibition by the Lactobacillus acidophilus strain of the present invention, the Lactobacillus acidophilus JCM1021 strain, the ATCC 4356 strain, and the LA5 strain. Each term has the same meaning as defined in Figure 8.

[0120] The present invention will be explained in more detail below through examples. These examples are intended to explain the invention more specifically, and the scope of the invention is not limited by these examples.

[0121]

[0122] Example 1. Identification of Lactobacillus acidophilus strains

[0123] After collecting 1 mL of the pure culture of the isolated microorganism, the 16S rRNA gene sequence was determined and is shown in Table 1 below, and the primer sequences used are shown in Table 2 below.

[0124] Type 16S rRNA gene SEQ ID NO: Microorganism AGTTCCCTTCGGGGACACTAAGACAGGTGGTGCATGGCTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTGTCATTAGTTGCCAGCATTAAGTTGGGCACTCTAATGAGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAGTCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGACAGTACAA CGAGGAGCAAGCCTGCGAAGGCAAGCGAATCTCTTAAAGCTGTTCTCAGTTCGGACTGCAGTCTGCAACTCGACTGCACGAAGCTGGAATCGCTAGTAATCGCGGATCAGCACGCCGCG GTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCATGGGAGTTCTGCAATGCCCAAAGCCGGTGGCCTAACCTTCGGGAAGGAGCCGTCTAAGGCAGGGCAGATGACTSEQ ID NO. 1

[0125]

[0126] Type Sequence Sequence Number Forward Primer 27F (for PCR) AGAGTTTGATCMTGGCTCAG Sequence Number 2 Reverse Primer 1492R (for PCR) TACGGYTACCTTGTTACGACTT Sequence Number 3 Forward Primer 785F (for Sequencing) GGATTAGATACCCTGGTA Sequence Number 4 Reverse Primer 907R (for Sequencing) CCGTCAATTCMTTTRAGTTT Sequence Number 5

[0127]

[0128] As a result, it was confirmed that the isolated microorganism exhibited 100% homology with Lactobacillus acidophilus. Accordingly, the microorganism was deposited with the Korea Research Institute of Bioscience and Biotechnology (KRIBB), a patent strain depositary, on April 11, 2023, and was assigned accession number KCTC 15402BP. KRIBB is a patent strain depositary located at 181, Ipsin-gil, Jeongeup-si, Jeollabuk-do, South Korea.

[0129]

[0130] Example 2. Preparation of Lactobacillus acidophilus strain powder

[0131] 1% (v / v) of the Lactobacillus acidophilus strain of the present invention was inoculated into a 50mL tube (50mL TPP Tube with 0.2㎛ Vent Cap (EF)) containing 10mL of MRS (De Man Rogosa, Sharpe agar) medium, and then cultured in an incubator at 37℃ and 0rpm for 24 hours. After culture, 1% (v / v) of the Lactobacillus acidophilus strain culture solution was inoculated into a 125mL Erlenmeyer flask (125mL Erlenmeyer Flask with 0.2㎛ Vent Cap (EF)) containing 50mL of MRS medium, and then cultured at 37℃ and 0rpm for 24 hours. Subsequently, the sample was centrifuged, filtered through a 0.22㎛ filter, and the supernatant was removed to prepare a cell precipitate, which was then freeze-dried.

[0132]

[0133] Example 3. Safety of Lactobacillus acidophilus strain

[0134] Example 3-1. Hemolytic activity of Lactobacillus acidophilus strains

[0135] In order to confirm the safety of the Lactobacillus acidophilus strain of the present invention for the human body, hemolytic activity, which is the ability to destroy cells such as red blood cells present in the blood, was confirmed.

[0136] Specifically, a Lactobacillus acidophilus strain pure-cultured in MRS medium was collected using a loop, streaked onto Sheep Blood Agar, and incubated aerobically at 37°C for 48 hours. Subsequently, hemolytic activity was determined by the formation of a clear zone around the cells.

[0137] As a result, as can be seen in Figure 1, it was confirmed that the Lactobacillus acidophilus strain cultured on blood agar medium was non-hemolytic as no clear halo formed around the cell.

[0138] Through the above results, it was confirmed that the Lactobacillus acidophilus strain of the present invention is harmless to the human body and has excellent safety, so it can be usefully utilized in medicines, cosmetics, or food applied to the human body.

[0139]

[0140] Example 3-2. Cytotoxicity of Lactobacillus acidophilus strains

[0141] As confirmed through Example 3-1 above that the Lactobacillus acidophilus strain of the present invention does not exhibit toxicity to the human body, toxicity was checked against HGF-1 cells, which are human gingival fibroblasts, to confirm whether the original Lactobacillus acidophilus strain also does not exhibit toxicity.

[0142] Specifically, 2 x 10 HGF-1 cells in a 96-well cell culture plate 4Cells were attached to the cells / well for 24 hours. Subsequently, LPS (Lipopolysaccharide) derived from Porphyromonas gingivalis, dexamethasone, and the original strain powder of Example 2 were treated according to the control and experimental group conditions below and cultured for 24 hours at 37°C and 5% CO2. LPS is a substance that forms the cell membrane of Gram-negative bacteria and is known to act as an endotoxin in cells to induce inflammation. Dexamethasone is a synthetic adrenocortical hormone and a type of corticosteroid drug known to have an anti-inflammatory effect.

[0143] (1) Negative Control 1 (NONE): LPS, dexamethasone, and sample not treated

[0144] (2) Negative control group 2 (CON): Treated with 1 μg / mL of LPS

[0145] (3) Positive control (DEXA): Treated with LPS 1 μg / mL and dexamethasone 1 μM

[0146] (4) Experimental group 1 (10 MOI): LPS 1 μg / mL and Lactobacillus acidophilus strain powder as a sample were treated at 10 MOI (Multiplicity of infection).

[0147] (5) Experimental group 2 (50 MOI): LPS 1 µg / mL and Lactobacillus acidophilus strain powder as a sample were treated at 50 MOI.

[0148] (6) Experimental group 3 (100 MOI): LPS 1 μg / mL and Lactobacillus acidophilus strain powder as a sample were treated at 100 MOI.

[0149] After the culture was completed, the cultured cell medium was removed and washed with PBS. Then, 100 µl of FBS-free DMEM medium was added, followed by the addition of 10 µl of EZ-Cytox reagent, and the mixture was reacted for 1 hour. Optical density (OD) was measured at 450 nm using a microplate reader. Subsequently, the cell viability was calculated based on the measured absorbance using the following Equation 1 and presented as a graph in Figure 2.

[0150] [Mathematical Formula 1]

[0151] Cell viability = (Absorbance of control or experimental group / Absorbance of negative control 1 (NONE)) X 100

[0152] As a result, as can be seen in Figure 2, the original Lactobacillus acidophilus strain sample did not affect the viability of HGF-1 cells even at a high throughput of 100 MOI.

[0153] Through the above results, it was confirmed that the Lactobacillus acidophilus strain and the original powder thereof of the present invention have no cytotoxicity, are harmless to the human body, and have excellent safety, so they can be usefully utilized in cosmetics, medicines, or food applied to the human body.

[0154]

[0155] Example 4. Antimicrobial activity of Lactobacillus acidophilus strain

[0156] To confirm the antibacterial activity of the Lactobacillus acidophilus strain of the present invention, Porphyromonas gingivalis and Fusobacterium nucleatum, which are representative bacteria that cause periodontitis or gingivitis, were used.

[0157] Specifically, the Porphyromonas gingivalis strain (KCTC 5352) was pure-cultured in TSB (Tryptic soy broth with 5% sheep blood defibrinated, 2% hemin-vitamin K1 and 0.05% L-cysteine ​​hydrochloride) medium, and then evenly plated over the entire TSA (Tryptic soy agar with 5% sheep blood defibrinated, 2% hemin-vitamin K1 and 0.05% L-cysteine ​​hydrochloride) medium. Fusobacterium nucleatum subsp. The nucleatum (Fusobacterium nucleatum subsp. nucleatum) strain (KGMB 05490) was pure-cultured in RCM broth (Reinforced Clostridial Medium broth with 3.98 μM resazurin sodium salt) and then evenly spread over the entire surface of RCM agar (Reinforced Clostridial Medium agar with 3.98 μM resazurin sodium salt). After creating holes in the solid medium inoculated with the periodontitis or gingivitis-causing bacteria using the agar well diffusion method with a flame-sterilized Pasteur pipette, 1X PBS, a cryopreservative, and the strain powder of Example 2 as a sample were dispensed onto the solid medium according to the control and experimental group conditions below. At this time, the cryopreservative and sample were added to and treated with 1X PBS. The cryopreservative was the same one used when preparing the strain powder of Example 2.

[0158] (1) Negative control 1: 1X PBS, cryopreservative, and untreated sample

[0159] (2) Negative control group 2: Treated with 80 µl of 1X PBS

[0160] (3) Negative control group 3: Treated with 80 µl of cryopreservative

[0161] (4) Experimental group: 1 x 10⁶ Lactobacillus acidophilus strain powder as a sample 9 Treats CFU / 80 µl

[0162] After dispensing, the samples were cultured anaerobically for 48 hours at 37°C, 0 rpm, and in a light-blocked environment. After culture, the clear zone formed around the holes of the recovered solid medium was examined, and the shortest diameter (mm) was measured and shown in Table 3.

[0163] Porphyromonas gingivalis strain (KCTC 5352) Fusobacterium nucleatum strain (KGMB 05490) Negative control group 10 mm 0 mm Negative control group 20 mm 0 mm Negative control group 30 mm 0 mm Experimental group 14 mm 25 mm

[0164] As a result, as can be seen in Table 3, the sample Lactobacillus acidophilus strain exhibited antibacterial activity against both of the aforementioned periodontitis or gingivitis-causing bacteria.

[0165] The above results suggest that the Lactobacillus acidophilus strain of the present invention inhibits the growth or proliferation of bacteria causing periodontitis or gingivitis, thereby exhibiting an effect of treating, improving, or preventing oral diseases.

[0166]

[0167]

[0168] Example 5. Inhibitory effect of Lactobacillus acidophilus strain on the production of reactive oxygen species (ROS).

[0169] In order to determine whether the Lactobacillus acidophilus strain of the present invention can treat, improve, or prevent oral diseases, particularly inflammatory diseases such as periodontitis, gingivitis, or oral inflammation, the inhibitory effect of said strain on ROS production was confirmed. It is known that high ROS concentrations cause apoptosis and induce chronic inflammation through the secretion of inflammatory substances such as cytokines (Roman Fischer et al. (2015), Interrelation Oxidative Stress and Inflammation in Neurodegenerative Disease: Role of TNF, Oxidative Medicine and Cellular Longevity, 2015:610813).

[0170] Specifically, 2 x 10 HGF-1 cells in a 96-well cell culture plate 4 The cells were dispensed at a concentration of cells / well and attached for 24 hours. Afterward, the cell medium was removed and washed with PBS. Then, LPS derived from Porphyromonas gingivalis, dexamethasone, and the original strain powder of Example 2 were treated according to the control and experimental group conditions below, and cultured for 24 hours at 37°C and 5% CO2. At this time, the LPS, dexamethasone, and the sample were added to DMEM medium that did not contain FBS.

[0171] (1) Negative Control 1 (NONE): LPS, dexamethasone, and sample not treated

[0172] (2) Negative control group 2 (CON): Treated with 1 μg / mL of LPS

[0173] (3) Positive control (DEXA): Treated with LPS 1 μg / mL and dexamethasone 1 μM

[0174] (4) Experimental group 1 (10 MOI): LPS 1 μg / mL and Lactobacillus acidophilus strain powder as a sample were treated at 10 MOI.

[0175] (5) Experimental group 2 (50 MOI): LPS 1 µg / mL and Lactobacillus acidophilus strain powder as a sample were treated at 50 MOI.

[0176] (6) Experimental group 3 (100 MOI): LPS 1 μg / mL and Lactobacillus acidophilus strain powder as a sample were treated at 100 MOI.

[0177] After the culture was completed, the amount of ROS produced in each well was measured and shown as a graph in Figure 3.

[0178] As a result, as can be seen in Figure 3, the amount of ROS generated in experimental groups 1 (10 MOI) to 3 (100 MOI) decreased in a concentration-dependent manner compared to negative control group 2 (CON). In particular, experimental group 3 (100 MOI) showed a higher ROS generation inhibitory effect than the positive control group (DEXA).

[0179] The above results suggest that the Lactobacillus acidophilus strain of the present invention exhibits antioxidant activity by inhibiting ROS production, and that this effect has the effect of treating, improving, or preventing oral diseases such as periodontitis, gingivitis, or oral inflammation.

[0180]

[0181] Example 6. Inhibitory effect of MMP-3 (Matrix metalloproteinase-3) expression in Lactobacillus acidophilus strains

[0182] To determine whether the Lactobacillus acidophilus strain of the present invention can treat, improve, or prevent oral diseases, particularly inflammatory diseases such as periodontitis, gingivitis, or oral inflammation, the inhibitory effect of said strain on the expression or activity of MMP-3 was examined. Since MMPs are involved in the degradation of the extracellular matrix within periodontal tissues, they are considered the most important pathway in tissue destruction associated with periodontal disease. MMPs and TIMPs (tissue inhibitors) that inhibit them play a key role in the physiological tissue remodeling of periodontal tissues, and it has been shown that periodontal disease and tissue destruction occur when their balance is disrupted. Harmful oral bacteria in plaque stimulate host cells to increase MMP release, which is one of the mechanisms of tissue destruction in periodontitis (Checchi et al. (2020), The Role of Matrix Metalloproteinases in Periodontal Disease, Int J Environ Res Public Health, 17(14):4923).

[0183] Specifically, 5 x 10 HGF-1 cells in a 24-well cell culture plate 4 The cells were dispensed at a concentration of cells / well and attached for 24 hours. Afterward, the cell medium was removed and washed with PBS. Then, LPS derived from Porphyromonas gingivalis, dexamethasone, and the original powder of the Lactobacillus acidophilus strain of Example 2 were treated according to the control and experimental group conditions below and cultured at 37°C and 5% CO2 for 24 hours. At this time, the LPS, dexamethasone, and the sample were added to DMEM medium that did not contain FBS.

[0184] (1) Negative Control 1 (NONE): LPS, dexamethasone, and sample not treated

[0185] (2) Negative control 2 (CON): Treated with 1 μg / mL of LPS

[0186] (3) Positive control (DEXA): Treated with LPS 1 μg / mL and dexamethasone 1 μM

[0187] (4) Experimental group 1 (10 MOI): LPS 1 μg / mL and Lactobacillus acidophilus strain powder as a sample were treated at 10 MOI.

[0188] (5) Experimental group 2 (50 MOI): LPS 1 μg / mL and Lactobacillus acidophilus strain powder as a sample were treated at 50 MOI.

[0189] (6) Experimental group 3 (100 MOI): LPS 1 μg / mL and Lactobacillus acidophilus strain powder as a sample were treated at 100 MOI.

[0190] After the culture was completed, the amount of MMP-3 in each well was measured and shown as a graph in Figure 4.

[0191] As a result, as can be seen in Figure 4, the MMP-3 expression levels of experimental groups 1 (10 MOI) to 3 (100 MOI) treated with the strain of the present invention decreased in a concentration-dependent manner. Compared to negative control group 2 (CON), experimental group 1 (10 MOI) decreased by approximately 40%, experimental group 2 (50 MOI) by approximately 47%, and experimental group 3 (100 MOI) by approximately 65%. In particular, experimental group 3 (100 MOI) was approximately 36% lower than the positive control (DEXA).

[0192] The above results suggest that the Lactobacillus acidophilus strain of the present invention reduces MMP-3 and exhibits activity that inhibits periodontal tissue damage or degradation, and that this effect has the effect of treating, improving, or preventing oral diseases such as periodontitis, gingivitis, or oral inflammation.

[0193]

[0194] Example 7. Inhibitory effect of Lactobacillus acidophilus strain on inflammatory cytokine expression

[0195] To determine whether the Lactobacillus acidophilus strain of the present invention can treat, improve, or prevent oral diseases, particularly inflammatory diseases such as periodontitis, gingivitis, or oral inflammation, the inhibitory effect of said strain on the expression of inflammatory cytokines IL-6 (Interleukin-6) and IL-8 (Interleukin-8) was confirmed. IL-6 and IL-8 are inflammatory cytokines whose expression or activity is induced by external stimuli that cause inflammation, such as microbial infection or tissue damage. These inflammatory cytokines generate nitric oxide and reactive oxygen species, thereby inducing oxidative stress, and it is known that the generated nitric oxide and reactive oxygen species promote cell damage and inflammation (Roman Fischer et al. (2015), Interrelation Oxidative Stress and Inflammation in Neurodegenerative Disease: Role of TNF, Oxidative Medicine and Cellular Longevity, 2015:610813).

[0196] Specifically, 5 x 10 HGF-1 cells in a 24-well cell culture plate 4 The cells were dispensed at a concentration of cells / well and attached for 24 hours. Afterward, the cell medium was removed and washed with PBS. Then, LPS derived from Porphyromonas gingivalis, dexamethasone, and the original strain powder of Example 2 were treated according to the control and experimental group conditions below and cultured at 37°C and 5% CO2 for 24 hours. At this time, the LPS, dexamethasone, and the sample were added to DMEM medium that did not contain FBS.

[0197] (1) Negative Control 1 (NONE): LPS, dexamethasone, and sample not treated

[0198] (2) Negative control group 2 (CON): Treated with 1 μg / mL of LPS

[0199] (3) Positive control (DEXA): Treated with LPS 1 μg / mL and dexamethasone 1 μM

[0200] (4) Experimental group 1 (10 MOI): LPS 1 μg / mL and Lactobacillus acidophilus strain powder as a sample were treated at 10 MOI.

[0201] (5) Experimental group 2 (50 MOI): LPS 1 µg / mL and Lactobacillus acidophilus strain powder as a sample were treated at 50 MOI.

[0202] (6) Experimental group 3 (100 MOI): LPS 1 μg / mL and Lactobacillus acidophilus strain powder as a sample were treated at 100 MOI.

[0203] After the culture was completed, the amounts of inflammatory cytokines IL-6 and IL-8 were measured and shown in graphs in Figures 5 and 6.

[0204] As a result, as can be seen in FIGS. 5 and 6, the expression levels of IL-6 and IL-8 in experimental groups 1 (10 MOI) to 3 (100 MOI) treated with the strain of the present invention were significantly reduced by more than 40%. The expression level of IL-6 decreased by approximately 42% in experimental group 1 (10 MOI), approximately 43% in experimental group 2 (50 MOI), and approximately 46% in experimental group 3 (100 MOI), while the expression level of IL-8 decreased by approximately 49% in experimental group 1 (10 MOI), approximately 52% in experimental group 2 (50 MOI), and approximately 53% in experimental group 3 (100 MOI). In particular, the expression level of IL-8 in experimental groups 1 (10 MOI) to 3 (100 MOI) decreased to a level similar to that of the positive control (DEXA).

[0205] The above results suggest that the Lactobacillus acidophilus strain of the present invention exhibits anti-inflammatory activity by reducing inflammatory cytokines, and that this effect is effective in treating, improving, or preventing oral diseases such as periodontitis, gingivitis, or oral inflammation.

[0206]

[0207] Example 8. Confirmation of the superiority of the strain according to the present invention against oral diseases or inflammatory diseases

[0208] In order to determine whether the Lactobacillus acidophilus strain of the present invention can exhibit superior effects in treating, improving, or preventing oral diseases, particularly inflammatory diseases such as periodontitis, gingivitis, or oral inflammation, the Lactobacillus acidophilus strain of the present invention was compared with Lactobacillus acidophilus JCM1021 (ATCC 4355), ATCC 4356, or LA5 strains, which are of the same species but different strains, regarding antibacterial activity, ROS production inhibition effect, and MMP-3 expression inhibition effect.

[0209] Lactobacillus acidophilus ATCC 4356 and JCM1021 strains were obtained from KCTC (Korea Biotechnology Research Institute Biological Resource Center) on January 14, 2025, and Lactobacillus acidophilus LA5 strain was obtained from Bigsol Co., Ltd., a food ingredient supplier, on March 14, 2025, after obtaining a distribution license through the headquarters of Chr. Hansen A / S in Denmark.

[0210]

[0211] Example 8-1. Antimicrobial activity

[0212] To compare the antibacterial activity of the strain of the present invention, the ATCC 4356 strain, and the LA5 strain, a Porphyromonas gingivalis strain (KCTC 5352), a representative bacterium that causes periodontitis or gingivitis, was used.

[0213] Specifically, a culture of the Porphyromonas gingivalis strain was evenly spread over the entire TSA medium. After creating holes in the solid medium inoculated with the periodontitis or gingivitis-causing bacteria using the agar well diffusion method with a flame-sterilized Pasteur pipette, the strain of the present invention, the ATCC 4356 strain, and the LA5 strain were dispensed into the solid medium as samples according to the control and experimental group conditions below. At this time, the strain according to the present invention, the ATCC 4356 strain, and the LA5 strain were treated according to Example 2.

[0214] (1) Experimental group 1 (KCTC 15402BP): 5x10 original strain powder of the present invention 8 Busy to achieve CFU / well

[0215] (2) Experimental group 2 (ATCC 4356): ATCC 4356 strain powder 5x10 8 Busy to achieve CFU / well

[0216] (3) Experimental group 3 (LA5): LA5 strain powder 5x10 8 Busy to achieve CFU / well

[0217]

[0218] After dispensing, the samples were cultured anaerobically for 72 hours at 37°C under light-blocking conditions. After culture, the clear zone formed around the holes of the recovered solid medium was examined, the shortest diameter (mm) was measured, and the average was calculated and shown in Table 4 and Figure 7.

[0219] Experimental group average clear zone diameter: 119.86 mm, 217.00 mm, 316.43 mm

[0220]

[0221] As a result, as can be seen in Table 4 and Figure 7, the antibacterial activity of experimental group 1 (KCTC 15402BP) against bacteria causing periodontitis or gingivitis increased compared to experimental group 2 (ATCC 4356) and experimental group 3 (LA5), and the difference was statistically significant at the P < 0.0001 level.

[0222] The above results suggest that compared to other Lactobacillus acidophilus strains of the same species but different strains, the Lactobacillus acidophilus strain of the present invention (KCTC 15402BP) has an excellent effect in inhibiting the growth or proliferation of bacteria that cause periodontitis or gingivitis, and that this effect is excellent for treating, improving, or preventing oral diseases such as periodontitis, gingivitis, or oral inflammation.

[0223]

[0224] Example 8-2. Effect of inhibiting ROS generation

[0225] In order to compare the antioxidant activity of the strain of the present invention, the ATCC 4356 strain, and the LA5 strain, the amount of ROS produced by each strain was determined.

[0226] Specifically, 2 x 10 HGF-1 cells in a 96-well cell culture plate 4 The cells were dispensed at a concentration of cells / well. The next day, after confirming by microscope that the cell confluency was 80% and an even layer had formed, the medium was washed with PBS and replaced with 0.2 mL of serum-free media (without penicillin and streptomycin). Subsequently, LPS, dexamethasone, and as samples, suspensions of the strain according to the present invention, suspensions of the JCM1021 strain, suspensions of the ATCC 4356 strain, and suspensions of the LA5 strain were dispensed into the solid medium according to the following control and experimental group conditions.

[0227] (1) Negative Control 1 (NONE): LPS, dexamethasone, and sample not treated

[0228] (2) Negative control group 2 (CON): Treated with 1 μg / mL of LPS

[0229] (3) Positive control (DEXA): Treated with LPS 1 μg / mL and dexamethasone 1 μM

[0230] (4) Experimental group 1 (KCTC 15402BP): LPS 1 μg / mL and a suspension of the strain according to the present invention as a sample were treated at a concentration of 100 MOI per cell number.

[0231] (5) Experimental group 2 (JCM1021): LPS 1 µg / mL and a suspension of the JCM1021 strain as a sample were treated at 100 MOI per cell number.

[0232] (6) Experimental group 3 (ATCC 4356): LPS 1 µg / mL and a suspension of ATCC 4356 strain as a sample were treated at 100 MOI per cell number.

[0233] (7) Experimental group 4 (LA5): LPS 1 µg / mL and a suspension of LA5 strain as a sample were treated at 100 MOI per cell number.

[0234]

[0235] After a 24-hour reaction, the cell supernatant was removed and the cells were stained with DCFDA (2',7'-dichlorofluorescin diacetate). The absorbance was measured using a microplate reader under the condition of fluorescence Ex / Em = 485 / 535 nm to determine the amount of ROS produced, and the results are shown as a graph in Figure 8.

[0236] As a result, as can be seen in Figure 8, the amount of ROS generated in experimental group 1 (KCTC 15402BP) was reduced compared to experimental group 2 (JCM1021), experimental group 3 (ATCC 4356), and experimental group 4 (LA5), and the difference was statistically significant at the P < 0.05 or P < 0.001 level.

[0237] In particular, experimental group 1 (KCTC 15402BP) showed a higher inhibitory effect on ROS production than the positive control group (DEXA).

[0238] The above results suggest that compared to other Lactobacillus acidophilus strains of the same species but different strains, the Lactobacillus acidophilus strain of the present invention (KCTC 15402BP) exhibits excellent antioxidant activity by inhibiting ROS production, and this effect is excellent for treating, improving, or preventing oral diseases such as periodontitis, gingivitis, or oral inflammation.

[0239]

[0240] Example 8-3. Inhibitory effect on MMP-3 expression

[0241] In order to compare the therapeutic, improvement, or preventive effects of the strain of the present invention, ATCC 4356 strain, and LA5 strain on inflammatory diseases, the inhibitory effect on the expression or activity of MMP-3 according to each strain was confirmed.

[0242] Specifically, 2 x 10 HGF-1 cells in a 96-well cell culture plate 4 The cells were dispensed at a concentration of cells / well. The next day, after confirming by microscope that the cell confluence was 80% and an even layer had formed, the medium was washed with PBS and then replaced with 1 mL of serum-free medium (without penicillin and streptomycin). Subsequently, LPS, dexamethasone, and as samples, suspensions of the strain according to the present invention, suspension of the JCM1021 strain, suspension of the ATCC 4356 strain, and suspension of the LA5 strain were dispensed into the solid medium according to the following control and experimental group conditions.

[0243] (1) Negative Control 1 (NONE): LPS, dexamethasone, and sample not treated

[0244] (2) Negative control group 2 (CON): Treated with 1 μg / mL of LPS

[0245] (3) Positive control (DEXA): Treated with LPS 1 μg / mL and dexamethasone 1 μM

[0246] (4) Experimental group 1 (KCTC 15402BP): LPS 1 μg / mL and a suspension of the strain according to the present invention as a sample were treated at a concentration of 100 MOI per cell number.

[0247] (5) Experimental group 2 (JCM1021): LPS 1 µg / mL and a suspension of the JCM1021 strain as a sample were treated at 100 MOI per cell number.

[0248] (6) Experimental group 3 (ATCC 4356): LPS 1 µg / mL and a suspension of ATCC 4356 strain as a sample were treated at 100 MOI per cell number.

[0249] (7) Experimental group 4 (LA5): LPS 1 µg / mL and a suspension of LA5 strain as a sample were treated at 100 MOI per cell number.

[0250]

[0251] After 24 hours of reaction, the cell supernatant was obtained, and the MMP-3 expression level was measured using a microplate reader and shown as a graph in Figure 9.

[0252] As a result, as can be seen in Figure 9, the MMP-3 expression level of experimental group 1 (KCTC 15402BP) was reduced compared to experimental group 2 (JCM1021), experimental group 3 (ATCC 4356), and experimental group 4 (LA5). The difference was statistically significant at the P < 0.01 level compared to experimental group 2, the P < 0.001 level compared to experimental group 3, and the P < 0.05 level compared to experimental group 4.

[0253] In particular, the MMP-3 expression inhibitory effect of experimental group 1 (KCTC 15402BP) was found to be at a level similar to that of the positive control (DEXA).

[0254] The above results suggest that compared to other Lactobacillus acidophilus strains of the same species but different strains, the Lactobacillus acidophilus strain of the present invention (KCTC 15402BP) exhibits excellent activity in inhibiting periodontal tissue damage or degradation by reducing MMP-3, and that this effect is excellent for treating, improving, or preventing oral diseases such as periodontitis, gingivitis, or oral inflammation.

[0255] [Consignment Number]

[0256]

Claims

1. Lactobacillus acidophilus strain deposited under accession number KCTC 15402BP.

2. In Paragraph 1, A strain in which the 16S rRNA gene of the above strain comprises the nucleotide sequence represented by SEQ ID NO.

1.

3. In Paragraph 1, The strain is one or more selected from the group consisting of live cells, dead cells and cultures thereof, fermented products thereof, lysed products thereof, extracts thereof and cytoplasmic fractions obtained by lysed products.

4. In Paragraph 1, The strain above is a strain having one or more of the following characteristics: (a) Inhibition of the production of reactive oxygen species (ROS), (b) inhibition of expression or activity of MMP-3 (Matrix metalloproteinase-3), and (c) Inhibition of expression or activity of IL-6 (Interleukin-6) or IL-8 (Interleukin-8).

5. In Paragraph 1, The strain above is a strain having antibacterial activity against oral harmful bacteria.

6. In Paragraph 5, The above-mentioned harmful oral bacteria are Streptococcus mutans, Streptococcus sobrinus, Porphyromonas gingivalis, Porphyromonas catoniae, Fusobacterium nucleatum, Actinomyces viscosus, Eikenella corrodens, Streptococcus sanguis, Streptococcus mitis, Streptococcus ratti, Streptococcus criceti, Streptococcus gordonii, and Streptococcus A strain that is one or more selected from the group consisting of Streptococcus anginosus, Streptococcus oralis, Treponema denticola, Prevotella intermedia, Bacteroides forsythus, Tannerella forsythus, Aggregatibacter actinomycetemcomitans, and Candida albicans.

7. A composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory purposes, comprising a strain of Lactobacillus acidophilus deposited under accession number KCTC 15402BP.

8. In Paragraph 7, A composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory, wherein the strain is one or more selected from the group consisting of live cells, dead cells and cultures thereof, fermented products thereof, lysed products thereof, extracts thereof, and cytoplasmic fractions obtained by lysing thereof.

9. In Paragraph 7, The above composition is a composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory purposes, which is used as a pharmaceutical composition, a quasi-drug composition, a food composition, a cosmetic composition, or a feed composition.

10. In Paragraph 9, The above pharmaceutical composition is an antibacterial, antioxidant, inhibitory of damage or degradation of periodontal tissue, and anti-inflammatory composition used for the treatment or prevention of dental caries, bad breath, or periodontal disease.

11. In Paragraph 10, A composition for antibacterial, antioxidant, inhibition of damage or degradation of periodontal tissue, and anti-inflammatory, wherein the above periodontal disease is selected from the group consisting of gingivitis, periodontitis, oral inflammation, periodontal abscess, periapical abscess, pericoronitis, periodontitis, and alveolar bone formation disorder.

12. In Paragraph 9, The above-mentioned quasi-drug composition, food composition, cosmetic composition, or feed composition is an antibacterial, antioxidant, inhibitory of damage or degradation of periodontal tissue, and anti-inflammatory composition used for the improvement or prevention of dental caries, bad breath, or periodontal disease.

13. A method for treating or preventing dental caries, bad breath, or periodontal disease in a subject, comprising the step of administering a composition according to claim 7 to a subject.

14. Use of the Lactobacillus acidophilus strain deposited as KCTC 15402BP for treating, improving, or preventing dental caries, halitosis, or periodontal disease in subjects.

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