Composition for improving, preventing or treating vaginitis or vulvitis comprising lactobacillus salivarius HHuMin-U

Abstract

The present invention relates to a composition for ameliorating, preventing or treating vaginitis or vulvitis, comprising Lactobacillus salivarius HHuMin-U, the composition of the present invention exhibiting an excellent antibacterial power against pathogenic bacteria inducing vaginitis and vulvitis, and being very effective in ameliorating and treating vaginitis and vulvitis.

Description

Technical Field

[0001] This invention relates to compositions containing Lactobacillus salivarius HHuMin-U for improving, preventing or treating vaginitis or vulvitis. Background Technology

[0002] Inflammation of the vagina and vulva is the most common cause of women seeking gynecological treatment. The most representative and common types of vaginal and vulva inflammation include bacterial vaginosis, trichomonal vaginitis, and candidal vaginitis. Other causes of vulvar ulcers include herpes virus infection and genital warts.

[0003] Vaginitis is an inflammation caused by bacterial infection of the vaginal mucosa. Pathogens can include gonococci, streptococci, staphylococci, Escherichia coli, Candida, and Trichomonas vaginalis, and it can also be a symptom of infectious diseases such as tuberculosis, syphilis, diphtheria, typhoid fever, and measles. Common symptoms include increased vaginal discharge, which is white or yellowish-white and cloudy, sometimes purulent or bloody, and often has a foul odor. The vulva often feels moist, itchy, and burning. When erosion occurs due to the discharge, urination may be painful. Although some cases can be cured by inserting vaginal suppositories containing antibiotics, complete cure in the early stages depends on proper diagnosis and treatment by a doctor. The most important preventative measure is maintaining local hygiene.

[0004] Vulvitis is an inflammation of the vulva, the external genitalia of the female reproductive system. The normally separated portion is called the vulvar cleft (vaginal fissure). It includes the labia majora, labia minora, clitoris, vaginal opening, and urethral opening. The main causes of vulvitis can be divided into infectious and non-infectious. Infectious causes are most commonly bacterial infections, largely due to an imbalance in the normal vaginal flora. Bacterial vaginosis is frequently associated with this condition. Fungal infections are also common, often caused by yeasts called Candida albicans. Symptoms include itching, redness, swelling, and abnormal discharge. Herpes simplex virus (HSV) or human papillomavirus (HPV) infections can also cause vulvitis. Known sexually transmitted pathogens such as Chlamydia, gonorrhea, and Trichomonas vaginalis can also cause vulvar inflammation. Non-infectious vulvitis can be caused by allergic reactions; inflammation can be triggered by chemicals found in washes, perfumes, sanitary napkins, and condoms that irritate the vulva. Furthermore, hormonal changes are also a significant factor, especially during menopause, when decreased estrogen levels cause the skin of the vulva to become thinner and drier, making it more sensitive to irritation. In addition, physical stimulation also plays a role, as the vulva can be irritated by tight clothing or friction.

[0005] The most common symptoms include itching, burning sensation, and vulvar pain, accompanied by redness and swelling of the vulva, as well as abnormal vaginal discharge.

[0006] Under normal conditions, the vagina of women has a slightly acidic environment due to the many lactobacilli inside the vagina, which helps to inhibit the growth of pathogenic microorganisms or fungi.

[0007] The slightly acidic environment of the vagina can be disrupted by various external factors such as excessive vaginal cleaning, long-term use of antibiotics or contraceptives, pregnancy, childbirth, and menopause, which can alter the balance of hormones and cause changes in the amount of lactobacilli, which are normally present in the vagina, thus changing the slightly acidic environment.

[0008] Treatment for vaginitis often involves sulfonamides, antibiotics, and chemically synthesized disinfectants. However, these medications are not only harmful to bacteria but also to beneficial lactobacilli. In particular, antibiotic use can induce antibiotic resistance. Furthermore, the use of chemically synthesized disinfectants (such as povidone-iodine) carries the risk of allergic reactions to iodine or damage to the mucous membranes, leading to itching, burning sensations, dermatitis, and potentially triggering hypothyroidism due to elevated iodine levels.

[0009] Therefore, it is necessary to find a method that can effectively improve and treat vaginitis and vulvitis without affecting the intestinal flora, and to prevent secondary vaginitis and vulvitis caused by imbalance of intestinal flora.

[0010] Existing technical documents

[0011] Patent documents

[0012] Patent Document 1: Korean Patent No. 10-1860513 (May 16, 2018) discloses a composition for the prevention or treatment of candidal vaginitis containing Lactobacillus salivarius MG242 isolated from the vagina.

[0013] Patent Document 2: Korean Patent No. 10-2337950 (December 7, 2021) discloses a strain of Lactobacillus salivarius HHuMin-U for improving, preventing or treating urinary tract infections and a composition containing the strain. Summary of the Invention

[0014] Technical issues

[0015] The object of the present invention is to provide a composition comprising Lactobacillus salivarius HHuMin-U (KCCM13001P) for use in improving, preventing or treating vaginitis or vulvitis.

[0016] Technical solution

[0017] The present invention provides a pharmaceutical composition for the prevention or treatment of vaginitis or vulvitis, comprising Lactobacillus salivarius HHuMin-U (KCCM13001P) as an active ingredient.

[0018] In this case, for example, the strain may be one or more of the following: live bacteria, fragments thereof, culture medium thereof, culture thereof, extract thereof, and dead cells of Lactobacillus salivarius HHuMin-U (KCCM13001P).

[0019] In this case, for example, the vaginitis or vulvitis may be selected from bacterial vaginosis, trichomonal vaginitis, candidal vaginitis, atrophic vaginitis, herpes virus infection, and genital warts.

[0020] This invention provides a health food containing Lactobacillus salivarius HHuMin-U (KCCM13001P) for improving vaginitis or vulvitis.

[0021] This invention provides a topical skin agent containing Lactobacillus salivarius HHuMin-U (KCCM13001P) for improving vaginitis or vulvitis.

[0022] The present invention provides a wash composition containing Lactobacillus salivarius HHuMin-U (KCCM13001P) for improving vaginitis or vulvitis.

[0023] The effects of the invention

[0024] The composition of the present invention exhibits excellent antibacterial activity against bacteria that cause vaginitis and vulvitis, and is very effective in improving and treating vaginitis and vulvitis. Attached Figure Description

[0025] Figure 1 To confirm the results of the antibacterial activity of the Lactobacillus salivarius HHuMin-U strain of the present invention against the Candida albicans strain.

[0026] Figure 2 To evaluate the effect of the Lactobacillus salivarius HHuMin-U strain of the present invention on the adhesion of Candida albicans strain to vaginal epithelial cells.

[0027] Figure 3 To evaluate the effect of the Lactobacillus salivarius HHuMin-U strain of the present invention on the ability of Candida albicans strain to inhibit vaginal epithelial cell killing.

[0028] Figure 4 To evaluate the effect of the present invention's *Lactobacillus salivarius* HHuMin-U strain on the growth of *Candida albicans* strains in vaginal epithelial cells.

[0029] Figure 5The results were used to evaluate the effect of the Lactobacillus salivarius HHuMin-U strain of the present invention on the expression of antimicrobial peptides in vaginal epithelial cells (top image: mRNA, bottom image: protein).

[0030] Figure 6 Results for evaluating the effect of the Lactobacillus salivarius HHuMin-U strain of the present invention on the expression of immunologically active substances in vaginal epithelial cells (top image: mRNA, bottom image: protein).

[0031] Figure 7 To evaluate the effects of oral administration of the present invention of Lactobacillus salivarius HHuMin-U strain on the improvement and treatment of vaginitis (Part A: weight change, Part B: CFU measurement of Candida albicans in vaginal wash, Part C: PAS staining of vaginal tissue).

[0032] Figure 8 To evaluate the effects of intravaginal administration of the present invention's *Lactobacillus salivarius* HHuMin-U strain on the improvement and treatment of vaginitis. Detailed Implementation

[0033] In the following description, to avoid confusion, repeated content will be omitted. That is, the content of this invention is not limited to the following description, and the content of this invention should be interpreted in its entirety.

[0034] In one example of the present invention, the strain Lactobacillus salivarius HHuMin-U (KCCM13001P) was confirmed to have excellent antibacterial activity against strains that induce vaginitis and vulvitis, thereby completing the present invention.

[0035] Furthermore, the strains of the present invention inhibit the adhesion of vaginitis and vulvitis-inducing strains in human vaginal epithelial cells, reduce the degree of killing of human vaginal epithelial cells induced by vaginitis and vulvitis-inducing strains, and promote the expression of antimicrobial peptides and immune-active substances in related cells.

[0036] Furthermore, in animal experiments, oral or intravaginal administration of the strains of the present invention effectively killed vaginitis and vulvitis-inducing strains, thereby effectively inhibiting the infection of vaginal tissues by vaginitis and vulvitis-inducing strains.

[0037] Therefore, the present invention provides a pharmaceutical composition for the prevention or treatment of vaginitis or vulvitis containing Lactobacillus salivarius HHuMin-U (KCCM13001P) as an active ingredient, a health food / topical skin agent / lotion composition for improving vaginitis or vulvitis.

[0038] In this case, the strain may be one or more of the following: live bacteria, fragments thereof, culture medium thereof, culture thereof, extract thereof, and dead bacterial cells of Lactobacillus salivarius HHuMin-U (KCCM13001P).

[0039] The term "fractures thereof" as used in this invention refers to the fractures of a strain, specifically the fractures of a strain broken by physical force or chemical treatment.

[0040] The term "culture medium" as used in this invention refers to the culture medium after the bacterial strain has been cultured. Conventional lactic acid bacteria culture media can be used to culture the strain. Furthermore, the term "culture medium" in this invention should be interpreted to include concentrated culture media.

[0041] The term "culture thereof" as used in this invention refers to a powder prepared by drying or evaporating the culture medium of the cultured strain.

[0042] The term "extract thereof" as used in this invention refers to an extract obtained by adding an extraction solvent to the strain.

[0043] The term "extract thereof" as used in this invention refers to a powder prepared by drying or evaporating the extraction solvent from an extract obtained by adding an extraction solvent to the strain.

[0044] The term "dead bacterial cells" used in this invention refers to the killed bacterial strain. The killing of the strain refers to killing by heating, applying pressure, grinding with grinding beads, enzyme or chemical treatment, and should be interpreted to include the concept of so-called "dead bacterial fragments" that are broken down by killing treatment.

[0045] In this case, for example, the vaginitis or vulvitis may be any one of bacterial vaginosis, trichomonal vaginitis, candidal vaginitis, atrophic vaginitis, herpes virus infection, and genital warts.

[0046] The Lactobacillus salivarius HHuMin-U (KCCM13001P) of this invention is a strain disclosed in Korean Patent No. 10-2337950 (December 7, 2021).

[0047] As one embodiment for achieving the above objectives, the present invention provides a pharmaceutical composition for the prevention or treatment of vaginitis or vulvitis, comprising Lactobacillus salivarius HHuMin-U (KCCM13001P) as an active ingredient.

[0048] The term "prevention" as used in this invention refers to all actions that suppress or delay the onset of affected diseases (i.e., vaginitis and vulvitis) by administering the pharmaceutical compositions of this invention.

[0049] Furthermore, the term "treatment" as used in this invention refers to all actions that improve or cure the symptoms of vaginitis and vulvitis by administering the pharmaceutical composition of this invention.

[0050] The strain of the present invention may also contain one or more active ingredients exhibiting the same or similar functions in the composition.

[0051] In addition to the strains of the present invention, the pharmaceutical compositions of the present invention may also contain pharmaceutically acceptable carriers.

[0052] The types of carriers that can be used in this invention are not particularly limited, and carriers commonly used in related technical fields can be used. Non-limiting examples of the carriers include lactose, glucose, sucrose, sorbitol, mannitol, physiological saline, sterile water, Ringer's solution, buffered physiological saline, albumin injection solution, xylitol, erythritol, maltitol, maltodextrin, glycerol, ethanol, etc. Two or more of the aforementioned carriers can be used alone or in combination.

[0053] Furthermore, when necessary, the pharmaceutical compositions of the present invention may be used with the addition of other pharmaceutically acceptable additives such as antioxidants, excipients, diluents, buffers or antibacterial agents, and may also be used with the addition of surfactants, binders, fillers, extenders, wetting agents, disintegrants, dispersants or lubricants.

[0054] In the pharmaceutical composition of the present invention, based on the total weight of the pharmaceutical composition, it may contain 0.00001 to 9.99% by weight of the strain of the present invention, preferably 0.1 to 90% by weight, more preferably 0.1 to 70% by weight, and even more preferably 0.1 to 50% by weight, but is not limited thereto, and can be varied in many ways depending on the condition of the recipient, the type of specific disease, the degree of progression, etc. When necessary, it may also be included as part of the total content of the pharmaceutical composition.

[0055] That is, the pharmaceutically effective amount and effective dosage of the pharmaceutical composition of the present invention can vary depending on the preparation method, administration method, administration time and / or route of administration of the pharmaceutical composition, etc., and can vary depending on various factors including the type and extent of the response to be achieved by administration of the pharmaceutical composition, the type of individual being administered, age, weight, semi-health status, symptoms or severity of disease, sex, diet, metabolism, and components of other pharmaceutical compositions used simultaneously or not by the relevant individuals, as well as other similar factors well known in the pharmaceutical field. Those skilled in the art can easily determine and prescribe an effective dosage based on the desired outcome. For example, the daily dosage of the pharmaceutical composition of the present invention can be from 0.01 mg / kg to 1000 mg / kg, preferably from 0.1 mg / kg to 100 mg / kg, and can be administered once a day or divided into several doses.

[0056] The pharmaceutical composition of the present invention can be administered once daily or divided into several doses. The pharmaceutical composition of the present invention can be administered as a single therapeutic agent or in combination with other therapeutic agents, and can be administered sequentially or simultaneously with existing therapeutic agents. The dosage that achieves maximum effect with minimal side effects can be determined by those skilled in the art after taking the aforementioned factors into account.

[0057] To prevent or treat vaginitis and vulvitis, the pharmaceutical composition of the present invention can also be used in combination with various methods such as hormone therapy and drug therapy.

[0058] The term "administration" as used in this invention refers to the act of introducing the pharmaceutical composition of the present invention into a patient by any suitable method. The route of administration and method of administration of the pharmaceutical composition of the present invention can be independent of each other. Any route of administration and method of administration can be used without restriction as long as the pharmaceutical composition can be introduced into the target site.

[0059] The pharmaceutical composition can be administered orally or parenterally, and can be formulated into a variety of suitable dosage forms for oral or parenteral administration.

[0060] Non-limiting examples of oral administration preparations using the pharmaceutical compositions of the present invention may include oily suspensions, troches, lozenges, tablets, water-soluble suspensions, crude powders, granules, emulsions, hard capsules, soft capsules, syrups, or liquids.

[0061] To formulate the pharmaceutical composition of the present invention for oral administration, binders such as sorbitol, mannitol, starch, amylopectin, cellulose, lactose, sucrose, or gelatin, lubricants such as magnesium stearate, calcium stearate, sodium stearate fumarate, or polyethylene glycol wax, excipients such as calcium hydrogen phosphate, disintegrants such as corn starch or sweet potato starch, and flavoring agents, syrups, and sweeteners may also be used. Furthermore, in the case of capsules, in addition to the substances mentioned above, liquid carriers such as fatty oils may also be used.

[0062] The parenteral administration method of the pharmaceutical composition of the present invention can be intramuscular, transdermal, intravenous, intraperitoneal or subcutaneous, etc., and the composition can be applied to the diseased site, or it can be used by spray, inhalation or other methods, but is not limited thereto.

[0063] Non-limiting examples of parenteral preparations utilizing the pharmaceutical compositions of the present invention may include injections, suppositories, ointments, topical powders, oils, ventilator inhalation powders, aerosol sprays, creams, etc.

[0064] In order to formulate the pharmaceutical composition of the present invention for parenteral administration, sterile aqueous solutions, non-aqueous solvents, suspending agents, oils, freeze-dried preparations, topical agents, etc. can be used. The non-aqueous solvents and suspending agents can be vegetable oils such as olive oil, propylene glycol, polyethylene glycol, ethyl oleate, etc., which are injectable esters.

[0065] When the pharmaceutical composition of the present invention is formulated as an injection, the pharmaceutical composition of the present invention can be mixed with a stabilizer or buffer in water to prepare a solution or suspension and formulated into a unit for administration in ampoules or vials.

[0066] When the pharmaceutical composition of the present invention is formulated as an aerosol, it can be combined with additives such as accelerators to disperse the dispersed concentrate or wet powder.

[0067] As another embodiment for achieving the above objectives, the present invention provides a health food containing Lactobacillus salivarius HHuMin-U (KCCM13001P) for improving vaginitis or vulvitis.

[0068] The term "health functional food" refers to food prepared and processed using raw materials or ingredients that have beneficial functions to the human body, in accordance with Korean Law No. 6727 concerning health functional foods. "Functionality" means that the food is consumed for the purpose of achieving health benefits, such as regulating or physiological effects of nutrients related to human structure and function.

[0069] In the health-functional food of the present invention, the content of the strains of the present invention can be varied in many ways depending on the condition of the recipient, the type of specific disease, the degree of progression, etc. When necessary, it can also be included as part of the total content of the food.

[0070] In the health food product of the present invention, preferably, the product may contain 0.00001 to 50% by weight of the strain of the present invention, relative to the health food product. If the weight percentage is less than 0.00001%, the effect is negligible; if the weight percentage is greater than 50%, the increase in effect relative to the amount used is negligible, and therefore uneconomical.

[0071] The health-functional food of the present invention can be, for example, a combination of probiotics, synbiotics, and postbiotics. Furthermore, it can be prepared in one or more forms selected from noodles, chewing gum, dairy products, ice cream, meat, grains, caffeinated beverages, regular beverages, chocolate, bread, snacks, candy, biscuits, jelly, sauces, alcoholic beverages, wine, vitamin complexes, and other health supplements, but is not limited thereto.

[0072] When the health-functional food of the present invention is used as a food additive, it can be added directly or used together with other foods or food ingredients, and can be used appropriately according to conventional methods.

[0073] The health-functional food of the present invention may also contain additional ingredients that can enhance odor, taste, and appearance. For example, it may contain biotin, folic acid, pantothenic acid, vitamin A, vitamin C, vitamin D, vitamin E, vitamin B1, vitamin B2, vitamin B6, vitamin B12, niacin, etc. Furthermore, it may contain minerals such as chromium (Cr), magnesium (Mg), manganese (Mn), copper (Cu), zinc (Zn), iron (Fe), and calcium (Ca). It may also contain amino acids such as cysteine, valine, lysine, and tryptophan. Furthermore, preservatives (potassium sorbate, sodium benzoate, salicylic acid, sodium dehydroacetate, etc.), colorants (tar pigments), color developers (sodium nitrite, sodium nitrite, etc.), bleaching agents (sodium sulfite), bactericides (bleaching powder and high-concentration bleaching powder, sodium hypochlorite, etc.), leavening agents (alum, potassium hydrogen tartrate, etc.), fortifying agents, emulsifiers, thickeners (pastes), coating agents, antioxidants (butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), etc.), flavorings (monosodium glutamate, etc.), sweeteners (glycine, cyclamate, saccharin, sodium, etc.), fragrances (vanillin, lactones, etc.), chewing gum base, antifoaming agents, solvents, improvers, and other food additives can be added. These additives can be selected according to the type of food and used in appropriate amounts.

[0074] As another embodiment for achieving the above objectives, the present invention provides a topical skin agent containing Lactobacillus salivarius HHuMin-U (KCCM13001P) for improving vaginitis or vulvitis.

[0075] The term "improvement" in this invention refers to all actions that improve or cure vaginitis and vulvitis by administering the compositions of this invention.

[0076] Examples of the topical skin preparations of the present invention include creams, gels, masks, sprays, ointments, plasters, emulsions, liniments, pastes, poultices, etc., but are not limited thereto.

[0077] In addition, other additives may include oils, moisturizers, emollients, surfactants, organic and inorganic pigments, organic powders, UV absorbers, preservatives, bactericides, antioxidants, plant extracts, pH adjusters, alcohol, colorants, fragrances, circulation enhancers, cooling agents, deodorants, and purified water. Organic ingredients may include ester oils, hydrocarbon oils, silicone-based oils, fluorinated oils, animal fats, and plant oils.

[0078] Ester oils can include glyceryl tri-2-ethylhexanoate, hexadecyl 2-ethylhexanoate, isopropyl myristate, butyl myristate, isopropyl palmitate, ethyl stearate, octyl palmitate, isochetol isostearate, butyl stearate, ethyl linoleate, isopropyl linoleate, ethyl oleate, isochetol myristate, isostearyl palmitate, octyl dodecyl myristate, isochetol isostearate, diethyl sebacate, diisopropyl adipate, isoalkyl neopentanoate, tri(caprylic, caprylic)glyceryl, trimethylolpropane tri-2-ethylhexanoate, trimethylolpropane triisostearate, and pentylene glycol tetra-2-ethylhexanoate. - Ethylhexanoate, hexadecyl caprylate, decyl laurate, hexyl laurate, decyl myristate, myristyl myristate, hexadecyl myristate, stearyl stearate, decyl oleate, hexadecyl castor oil, isostearyl laurate, isotrimethylene myristate, isochetyl palmitate, isostearyl laurate, isotrimethylene myristate, isochetyl palmitate, octyl stearate, isochetyl stearate, isodecanyl oleate, octyl dodecyl oleate, octyl dodecyl linoleate, isopropyl isostearate, cetearyl 2-ethylhexanoate, stearyl 2-ethylhexanoate, hexyl isostearate, ethylene glycol dioctyl ester ... Dioleate, Propylene glycol didecanoate, Propylene glycol di(caprylic / capric) ester, Propylene glycol dicaprylate, Neopentyl glycol didecanoate, Neopentyl glycol dicaprylate, Tricaprylic acid glyceride, Tricaprylic acid glyceride, Triisopalmitoyl glyceride, Triisostearate glyceride, Octyldodecyl neopentanoate, Octyl isostearate, Octyl isostearate, Isostearate isostearate, Octyl decyl isostearate, Polyglyceryl oleate, Polyglyceryl isostearate, Triisohexyl citrate, Triisoalkyl citrate, Triisooctyl citrate, Laureth lactate, Myristyl lactate, Lactic acid Esters such as cetyl ester, octyl decyl lactate, triethyl citrate, acetyl triethyl citrate, acetyl tributyl citrate, trioctyl citrate, diisostearyl maleate, 2-ethylhexyl hydroxystearate, di-2-ethylhexyl succinate, diisobutyl adipate, diisopropyl sebacate, dioctyl sebacate, cholesterol stearate, cholesterol isostearate, cholesterol hydroxystearate, cholesterol oleate, dihydrocholesterol oleate, phytosterol isostearate, phytosterol oleate, isochetol 12-stearoylhydroxystearate, stearyl 12-stearoylhydroxystearate, isostearyl 12-stearoylhydroxystearate, etc.

[0079] Hydrocarbon oils can be squalene, liquid paraffin, α-olefin oligomers, isoalkanes, ceresin, paraffin, liquid isoparaffin, polybutene, microcrystalline wax, petrolatum, etc.

[0080] Silicone-based greases can be polymethylsiloxane, methylphenylsiloxane, methylcyclopolysiloxane, octamethylpolysiloxane, decamethylpolysiloxane, dodecylcyclosiloxane, dimethylsiloxane / methylcetoxysiloxane copolymer, dimethylsiloxane / methylstearoxysiloxane copolymer, alkyl-modified silicone oil, amino-modified silicone oil, etc.

[0081] Fluorinated greases can be perfluoropolyethers, etc.

[0082] Animal or vegetable oils can include avocado oil, almond oil, olive oil, sesame oil, rice bran oil, safflower oil, soybean oil, corn oil, rapeseed oil, almond oil, palm kernel oil, palm oil, castor oil, sunflower seed oil, grape seed oil, cottonseed oil, coconut oil, macadamia nut oil, wheat germ oil, rice germ oil, shea butter, evening primrose oil, macadamia nut oil, wheat germ oil, egg yolk oil, beef tallow, horse oil, mink oil, orange peel oil, jojoba oil, candelilla wax, carnauba wax, liquid lanolin, hydrogenated castor oil, and other animal or vegetable oils.

[0083] Moisturizers can be water-soluble low-molecular-weight moisturizers, lipid-soluble molecular moisturizers, water-soluble polymers, lipid-soluble polymers, etc.

[0084] Water-soluble low-molecular-weight moisturizers can include serine, glutamine, sorbitol, mannitol, sodium pyrrolidone carboxylate, glycerin, propylene glycol, 1,3-butanediol, ethylene glycol, polyethylene glycol B (degree of polymerization n is 2 or higher), polypropylene glycol (degree of polymerization n is 2 or higher), polyglycerol B (degree of polymerization n is 2 or higher), lactic acid, lactate, etc.

[0085] Lipid-soluble low-molecular-weight moisturizers can be cholesterol, cholesterol esters, etc.

[0086] Water-soluble polymers can include carboxyvinyl polymers, polyaspartate, tragacanth gum, xanthan gum, methylcellulose, hydroxymethylcellulose, hydroxyethylcellulose, hydroxypropylcellulose, carboxymethylcellulose, water-soluble chitin, chitosan, dextrin, etc.

[0087] Fat-soluble polymers can be polyvinylpyrrolidone / eicosene copolymer, polyvinylpyrrolidone / hexadecene copolymer, nitrocellulose, dextrin fatty acid ester, high molecular weight siloxane, etc.

[0088] Emollients can include long-chain acylglutamate cholesterol ester, hydroxystearic acid cholesterol ester, 12-hydroxystearic acid, stearic acid, rosin acid, lanolin fatty acid cholesterol ester, etc.

[0089] Surfactants can be nonionic surfactants, anionic surfactants, cationic surfactants, amphoteric surfactants, etc.

[0090] Nonionic surfactants can be self-emulsifying glyceryl monostearate, propylene glycol fatty acid esters, glyceryl fatty acid esters, polyglycerol fatty acid esters, sorbitan fatty acid esters, polyoxyethylene (POE) sorbitan fatty acid esters, polyoxyethylene sorbitan fatty acid esters, polyoxyethylene glycerol fatty acid esters, polyoxyethylene alkyl ethers, polyoxyethylene fatty acid esters, polyoxyethylene hydrogenated castor oil, polyoxyethylene castor oil, polyoxyethylene (POE) / polyoxypropylene (POP) copolymers, polyoxyethylene / polyoxypropylene alkyl ethers, polyether-modified silicone oils, lauric acid alkanolamides, alkylamine oxides, hydrogenated soybean lecithin, etc.

[0091] Anionic surfactants can be fatty acid soaps, α-acylsulfonates, alkylsulfonates, alkylallylsulfonates, alkylnaphthalenesulfonates, alkyl sulfates, polyoxyethylene alkyl ether sulfates, alkylamide sulfates, alkyl phosphates, polyoxyethylene alkyl phosphates, alkylamide phosphates, alkylacylalkyl taurates, N-acyl amino acid salts, polyoxyethylene alkyl ether carboxylates, alkyl sulfosuccinates, sodium alkyl sulfoacetate, acylated hydrolyzed collagen peptide salts, perfluoroalkyl phosphates, etc.

[0092] Cationic surfactants can be alkyltrimethylammonium chloride, stearyltrimethylammonium chloride, stearyltrimethylammonium bromide, cetearyltrimethylammonium chloride, distearate dimethylammonium chloride, stearyldimethylbenzylammonium chloride, behenyltrimethylammonium bromide, benzalkonium chloride, diethylaminoethyl amide stearate, dimethylaminopropyl amide stearate, quaternary ammonium salts of lanolin derivatives, etc.

[0093] Amphoteric surfactants can be of various types, including carboxybetaine, amide betaine, sulfobetaine, hydroxysulfobetaine, amide sulfobetaine, phosphate betaine, aminocarboxylate, imidazoline derivative, and amide amine.

[0094] Organic and inorganic pigments can include: silicic acid, anhydrous silicic acid, magnesium silicate, talc, sericite, mica, kaolin, ferric oxide, clay, bentonite, titanium pearlite, bismuth oxychloride, zirconium oxide, magnesium oxide, zinc oxide, titanium dioxide, aluminum oxide, calcium sulfate, barium sulfate, magnesium sulfate, calcium carbonate, magnesium carbonate, iron oxide, ultramarine, chromium oxide, chromium hydroxide, calamine, and their complexes, etc.; organic pigments such as polyamide, polyester, polypropylene, polystyrene, polyurethane, vinyl resin, urea resin, phenolic resin, fluororesin, silicone resin, acrylic resin, melamine resin, epoxy resin, polycarbonate resin, divinylbenzene / styrene copolymer, silk powder, cellulose, CI pigment yellow, CI pigment orange, etc.; and composite pigments of these inorganic and organic pigments, etc.

[0095] Organic powders can be: metal soaps such as calcium stearate; alkyl phosphate metal salts such as sodium cetyl phosphate, zinc lauryl phosphate, and calcium lauryl phosphate; polyvalent metal salts of acyl amino acids such as calcium N-lauroyl-β-alanine, zinc N-lauroyl-β-alanine, and calcium N-lauroylglycinate; polyvalent metal salts of amide sulfonic acids such as calcium N-lauroyl-taurate and calcium N-palmitoyl-taurate; N-acyl basic amino acids such as N-ε-lauroyl-L-lysine, N-ε-palmitoyllysine, N-α-palmitoylornithine, N-α-lauroylarginine, and N-α-hydrogenated tallow fatty acid acylarginine; N-acyl polypeptides such as N-lauroylglycine; α-amino fatty acids such as α-aminooctanoic acid and α-aminolauric acid; polyethylene, polypropylene, nylon, polymethyl methacrylate, polystyrene, divinylbenzene / styrene copolymer, and polytetrafluoroethylene, etc.

[0096] UV absorbers can be p-aminobenzoic acid, ethyl p-aminobenzoate, amyl p-aminobenzoate, octyl p-aminobenzoate, ethylene glycol salicylate, phenyl salicylate, octyl salicylate, benzyl salicylate, butyl phenyl salicylate, horamyl salicylate, benzyl cinnamate, 2-ethoxyethyl p-methoxycinnamate, octyl p-methoxycinnamate, mono-2-ethylhexyl glycerol bis-p-methoxycinnamate, isopropyl p-methoxycinnamate, diisopropyl / diisopropyl cinnamate. Mixtures, uric acid, ethyl uric acid, hydroxymethoxybenzophenone, hydroxymethoxybenzophenone sulfonic acid and its salts, dihydroxymethoxybenzophenone, sodium dihydroxymethoxybenzophenone disulfonate, dihydroxybenzophenone, tetrahydroxybenzophenone, 4-tert-butyl-4'-methoxybenzoylmethane, 2,4,6-triphenylamino-p-(carbon-2'-ethylhexyl-1'-oxy)-1,3,5-triazine, 2-(2-hydroxy-5-methylphenyl)benzotriazole, etc.

[0097] Bactericides can include pyriproxyfen, triclosan, trichlorohydroxydiphenyl ether, chlorhexidine gluconate, phenoxyethanol, resorcinol, isopropyl methylphenol, chamomile, salicylic acid, zinc pyrithione, benzalkonium chloride, photosensitizer 301, sodium guaiacol mononitrate, undecanoic acid, etc.

[0098] Antioxidants can include butylated hydroxyanisole, propyl gallate, isoascorbic acid, etc.

[0099] pH adjusters can include citric acid, sodium citrate, malic acid, sodium malate, fumaric acid, sodium fumarate, succinic acid, sodium succinate, sodium hydroxide, sodium dihydrogen phosphate, etc.

[0100] The alcohols can be higher alcohols such as cetyl alcohol.

[0101] Furthermore, the additional ingredients that can be added are not limited to this, and any of the ingredients may be used in combination without impairing the purpose and effect of the present invention.

[0102] The topical skin preparations of the present invention can vary widely depending on various factors, including the activity of the specific active ingredient used, age, weight, general health condition, gender, diet, application time, drug combination, and the severity of the specific disease to be prevented or treated. The applicable dosage of the topical skin preparation can vary depending on the patient's condition, weight, disease severity, drug form, route of application, and duration, but can be appropriately selected by relevant practitioners. Preferably, after considering all the aforementioned factors, the dosage that achieves the maximum effect with the minimum amount without side effects can be applied. More preferably, it can be administered at a dose of 1–10,000 μg / kg / day, and even more preferably, at an effective dose of 10–1,000 mg / kg / day once a day or repeated several times. However, in no way are the dosages described herein limiting the scope of the present invention.

[0103] As another embodiment for achieving the above objectives, the present invention provides a wash composition containing Lactobacillus salivarius HHuMin-U (KCCM13001P) for improving vaginitis or vulvitis.

[0104] In this invention, preferably, the wash composition may contain 0.00001 to 30.0% by weight of the bacterial strain relative to the total weight. More preferably, the wash composition may contain 0.01 to 10% by weight of the bacterial strain relative to the total weight. If the content of the bacterial strain is less than 0.00001% by weight, no effect on improving vaginitis and vulvitis is observed; if it is greater than 30.0% by weight, no significant increase in effect with increasing content is observed.

[0105] On the other hand, the detergent composition of the present invention may contain, in addition to the strain of the present invention as an active ingredient, components commonly used in detergent compositions, such as antioxidants, stabilizers, solubilizers, vitamins, pigments and fragrances, and carriers.

[0106] The cleansing compositions of the present invention can be prepared in any dosage form commonly prepared in the technical field to which the present invention pertains, for example, as solutions, suspensions, emulsions, patches, gels, creams, soaps, surfactant-containing facial cleansers, oils, and sprays, but are not limited thereto.

[0107] When the dosage form of the lotion composition of the present invention is a patch, cream or gel, the carrier component may be animal oil, vegetable oil, wax, paraffin, starch, tragacanth gum, cellulose derivatives, polyethylene glycol, silica gel, bentonite, silica, talc or zinc oxide, etc.

[0108] When the detergent composition of the present invention is in the form of a solution or emulsion, the carrier component may be a solvent, cosolvent, or emulsifier, for example, water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butanediol oil, glycerol aliphatic ester, polyethylene glycol, or fatty acid esters of sorbitol.

[0109] When the detergent composition of the present invention is in the form of a suspension, the carrier component may be a liquid diluent such as water, ethanol or propylene glycol, a suspending agent such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitan ester and polyoxyethylene dehydrated sorbitan ester, microcrystalline cellulose, aluminum aluminate hydroxide, bentonite, agar or tragacanth gum, etc.

[0110] When the detergent composition of the present invention is in the form of a spray, the carrier component may be lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder. In particular, when it is a spray, it may also contain propellants such as chlorofluorocarbons, propane / butane or dimethyl ether.

[0111] When the detergent composition of the present invention is in the form of a surfactant-containing facial cleanser, the carrier component may be fatty alcohol sulfate, fatty alcohol ether sulfate, sulfosuccinate monoester salt, hydroxyethyl sulfonate, imidazoline derivative, methyl taurine, sarcosine salt, fatty acid amide ether sulfate, alkylamide betaine, fatty alcohol, fatty acid glycerides, fatty acid diethanolamide, vegetable oil, lanolin derivative, or ethoxylated glycerol fatty acid ester.

[0112] When the cleansing composition of the present invention is a soap, a surfactant-containing facial cleanser, or a surfactant-free facial cleanser, it can be wiped off, peeled off, or rinsed off with water after application to the skin. Specifically, the soap can be liquid soap, powdered soap, solid soap, or oil-based soap, and the surfactant-containing / surfactant-free facial cleanser can be a cleansing cream, makeup remover, or cleansing gel, but is not limited thereto.

[0113] The present invention will now be described in more detail through the following embodiments or experimental examples. However, the scope of protection of the present invention is not limited to the following embodiments or experimental examples, but should also include modifications of equivalent technical concepts.

[0114] Example 1: Evaluation of the antibacterial activity of Lactobacillus salivarius strain HHuMin-U against Candida albicans

[0115] Candida albicans strain (KCTC2720), a representative inducing strain for vaginitis and vulvitis, was cultured in YM medium under anaerobic conditions (85% N2, 10% H2 and 5% CO2).

[0116] Candidate strains containing *Lactobacillus salivarius* HHuMin-U (hereinafter referred to as "HMU") were cultured in MRS modified medium for 15 hours under both anaerobic conditions (85% N2, 10% H2, and 5% CO2) at 37°C, pH 5.5, and 150 rpm. The supernatant was then separated by centrifugation at 7000 rpm for 15 minutes. The separated supernatant was filtered through a 0.20 μm filter.

[0117] The degree of inhibition was confirmed by adding supernatant to the Candida albicans culture medium at a concentration of 3%–5% (v / v) and then using a growth curve. The control group used culture medium uninoculated with Lactobacillus salivarius HHuMin-U strain.

[0118] The experimental results showed that, in the antibacterial activity experiments using multiple strains of Candida albicans, the Lactobacillus salivarius strain HHuMin-U (HMU) exhibited the best antibacterial activity. Figure 1 ). Figure 1 To confirm the results of the antibacterial activity of the Lactobacillus salivarius HHuMin-U strain of the present invention against the Candida albicans strain. Figure 1 In the text, "CH88" refers to Lactobacillus paracasei strain CH88, and "number" indicates unidentified candidate strains.

[0119] Example 2: Evaluation of the therapeutic efficacy of vaginitis and vulvitis in human vaginal epithelial cell lines

[0120] In all the experiments conducted below, *Lactobacillus salivarius* strain HHuMin-U was cultured for 15 hours in a jar fermenter (Fermentec, South Korea) using MRS modified medium at 37°C, pH 5.5, 150 rpm, and anaerobic conditions (85% N2, 10% H2, and 5% CO2). After centrifugation at 7000 rpm for 15 minutes to remove the supernatant, the pellets were freeze-dried with cryoprotectant, and the pulverized powder was used as the sample.

[0121] 1. Assess the ability of Candida albicans strains to inhibit adhesion to human vaginal epithelial cells (adhesion assay)

[0122] Human vaginal epithelial cell lines were aliquoted into 6-well cell culture plates and cultured until a monolayer was formed. The cells were then treated with 5 × 10⁶ cells per well. 7 CFU, 1×10 8Four hours after inoculating CFU with HMU or phosphate-buffered saline (PBS), inoculate with 4 × 10⁻⁶ CFU. 4 CFU (Candida albicans) strains.

[0123] Then, after culturing for 3 hours, the culture medium was removed, and unattached Candida albicans strains were removed by washing with PBS. After washing, 1 ml of distilled water was added to the wells after washing and reacted for 15 minutes to remove vaginal epithelial cells. All solutions were recovered and serially diluted before being spread onto Sabouraud dextrose agar plates to measure the CFU of Candida albicans strains.

[0124] Experimental results showed that treatment with Lactobacillus salivarius HHuMin-U strain reduced the adhesion of Candida albicans strains to vaginal epithelial cells. Figure 2 ). Figure 2 To evaluate the effect of the Lactobacillus salivarius HHuMin-U strain of the present invention on the adhesion of Candida albicans strain to vaginal epithelial cells.

[0125] 2. Assess the killing ability of Candida albicans strains against human vaginal epithelial cells (lactate dehydrogenase assay (LDHassay)).

[0126] After aliquoting human vaginal epithelial cell lines into 6-well cell culture plates and culturing until a monolayer is formed, 5 × 10⁻⁶ cells were used. 7 CFU, 1×10 8 Four hours after CFU treatment with HMU or PBS, 2 × 10⁶ cells were inoculated. 6 CFU of Candida albicans strains was cultured.

[0127] The culture medium was then recovered at 48 and 72 hours, and the lactate dehydrogenase (LDH) levels in the culture medium were measured using the Cyquant LDH Cytotoxicity Assay Kit (Thermo Fisher Scientific) to confirm the relative degree of vaginal epithelial cell killing.

[0128] Experimental results showed that treatment with Lactobacillus salivarius HHuMin-U strain reduced the killing effect of Candida albicans strain on vaginal epithelial cells. Figure 3 ). Figure 3 To evaluate the effect of the Lactobacillus salivarius HHuMin-U strain of the present invention on the ability of Candida albicans strain to inhibit vaginal epithelial cell killing.

[0129] 3. Assess antimicrobial activity against Candida albicans strains in human vaginal epithelial cells (growth inhibition assay)

[0130] Cultured until a monolayer is formed, using 1×10 8 After treating CFU with HMU or PBS for 6 hours, the culture medium is recovered.

[0131] Then, after centrifuging the culture medium at 3000 rpm, the supernatant was collected for experimental use. 150 μl of the relevant supernatant was added to each well of a 96-well plate, and 4 × 10⁶ cells were inoculated. 4 CFU of Candida albicans strains were cultured at 37°C, and the OD values ​​were measured at 0, 12, and 24 hours to determine the growth extent of the Candida albicans strains.

[0132] Experimental results confirmed that treatment with *Lactobacillus salivarius* strain HHuMin-U inhibited the growth of *Candida albicans* strains in human vaginal epithelial cells. Figure 4 ). Figure 4 To evaluate the effect of the present invention's *Lactobacillus salivarius* HHuMin-U strain on the growth of *Candida albicans* strains in vaginal epithelial cells.

[0133] 4. Analyze the expression of antimicrobial peptides and immunologically active substances.

[0134] A. Analyze the expression of antimicrobial peptides (real-time quantitative polymerase chain reaction (RT-qPCR)).

[0135] After aliquoting human vaginal epithelial cell lines into 6-well cell culture plates and culturing until a monolayer is formed, 5 × 10⁻⁶ cells were used. 7 CFU, 1×10 8 CFU was treated with HMU or PBS for 6 hours. Then, the culture medium was removed and the cells were washed twice with cool PBS to isolate the mRNA. The isolated mRNA was then grouped into cDNA and used for RT-qPCR.

[0136] Experimental results confirmed that treatment with Lactobacillus salivarius strain HHuMin-U increased the expression of antimicrobial peptides in human vaginal epithelial cells. Figure 5 ). Figure 5 The results were used to evaluate the effect of the Lactobacillus salivarius HHuMin-U strain of the present invention on the expression of antimicrobial peptides in vaginal epithelial cells (top image: mRNA, bottom image: protein).

[0137] B. Analyze the expression of immunologically active substances (enzyme-linked immunosorbent assay (ELISA))

[0138] After aliquoting human vaginal epithelial cell lines into 6-well cell culture plates and culturing until a monolayer is formed, 5 × 10⁻⁶ cells were used. 7 CFU, 1×10 8 CFU was treated with HMU or PBS for 6 hours. Then, after recovering the culture medium, the supernatant was centrifuged at 13,000 rpm and used in ELISA analysis.

[0139] Experimental results confirmed that treatment with Lactobacillus salivarius strain HHuMin-U increased the expression of immunologically active substances in human vaginal epithelial cells. Figure 6 ). Figure 6 Results for evaluating the effect of the Lactobacillus salivarius HHuMin-U strain of the present invention on the expression of immunologically active substances in vaginal epithelial cells (top image: mRNA, bottom image: protein).

[0140] Example 3: Evaluation of therapeutic efficacy for vaginitis and vulvitis in animal models

[0141] In all the experiments conducted below, *Lactobacillus salivarius* strain HHuMin-U (hereinafter referred to as "HUM") was cultured for 15 hours in a tank fermenter (Fermentec Corporation, South Korea) using MRS modified medium at 37°C, pH 5.5, 150 rpm, and anaerobic conditions (85% N2, 10% H2, and 5% CO2). After centrifugation at 7000 rpm for 15 minutes to remove the supernatant, the pellets were freeze-dried with cryoprotectant, and the pulverized powder was used as the sample.

[0142] 1. Oral administration

[0143] Six-week-old female C57BL / 6 mice were acclimatized for one week before the experiment. Each group was orally administered 100 μl of PBS or 1×10⁻⁶ PBS. 7 CFU, 1×10 8 The HMU (Hepatobiliary Mutation) of CFU lasted for 13 days. On day 8 of oral administration, 0.5 mg / 100 μl of β-estradiol dissolved in sesame oil was administered subcutaneously. On day 11, 5 × 10⁵ mg / 10⁶ μl of β-estradiol was administered vaginally. 5 Infection was performed using Candida albicans strains in 20 μl PBS.

[0144] Three days after infection, the experiment was completed on day 14, and the vagina was washed with 100 μl of PBS. After serial dilution of the washing solution, the samples were spread onto Sabouraud dextrose agar plates to measure the CFU of Candida albicans strains. Vaginal tissue was collected and analyzed. The vaginal tissue was analyzed as follows: vaginal tissue was fixed with 10% paraformaldehyde, prepared into paraffin blocks, and then sectioned and stained with iodate-Schiff stain. The presence of Candida albicans strains infiltrating the tissue was confirmed by this staining.

[0145] The experimental results confirmed a significant reduction in CFU of Candida albicans strains in vaginal tissue, and infection suppression was also confirmed in tissue iodosylation of Schiff. Figure 7 ). Figure 7 To evaluate the effects of oral administration of the present invention's Lactobacillus salivarius HHuMin-U strain on the improvement and treatment of vaginitis (A: weight change, B: CFU measurement of Candida albicans in vaginal wash, C: Schiff staining of vaginal tissue with iodotic acid).

[0146] 2. Intravaginal administration

[0147] Six-week-old female C57BL / 6 mice were acclimatized for one week before the experiment. Each group was administered PBS or 1×10⁻⁶ PBS intravaginally. 6 The HMU (Hepatocellular Mucosal Administration) of CFU lasted for 5 days. On day 3 of vaginal administration, 0.5 mg / 100 μl of β-estradiol dissolved in sesame oil was administered subcutaneously. On day 6, 5 × 10⁵ CFU was administered vaginally. 5 Infection was performed using Candida albicans strains in 20 μl PBS.

[0148] The experiment ended on day 3 of infection, and the vagina was washed with 100 μl of PBS. After continuous dilution of the relevant washing solution, it was smeared onto Sabouraud dextrose agar plates to measure the CFU of Candida albicans strains.

[0149] Experimental results confirmed a significant reduction in CFU of Candida albicans strains in the guiding washing solution. Figure 8 ). Figure 8 To evaluate the effects of intravaginal administration of the present invention's *Lactobacillus salivarius* HHuMin-U strain on the improvement and treatment of vaginitis.

Claims

1. A pharmaceutical composition for the prevention or treatment of vaginitis or vulvitis, characterized in that, It contains Lactobacillus salivarius HHuMin-U with accession number KCCM13001P as the active ingredient.

2. The pharmaceutical composition for the prevention or treatment of vaginitis or vulvitis according to claim 1, characterized in that, The strain is one or more of the following: live Lactobacillus salivarius HHuMin-U strain with accession number KCCM13001P, its fragments, its culture medium, its culture, its extract, and its dead cells.

3. The pharmaceutical composition for the prevention or treatment of vaginitis or vulvitis according to claim 1, characterized in that, The vaginitis or vulvitis mentioned is selected from any one of bacterial vaginosis, trichomonal vaginitis, candidal vaginitis, atrophic vaginitis, herpes virus infection, and genital warts.

4. A health food product for improving vaginitis or vulvitis, characterized in that, It includes Lactobacillus salivarius HHuMin-U with accession number KCCM13001P.

5. A topical skin agent for improving vaginitis or vulvitis, characterized in that, It includes Lactobacillus salivarius HHuMin-U with accession number KCCM13001P.

6. A wash composition for improving vaginitis or vulvitis, characterized in that, It includes Lactobacillus salivarius HHuMin-U with accession number KCCM13001P.

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