WO2024025173A1 - Cosmetic composition comprising staphylococcus sp. strain, streptococcus sp. strain, cutibacterium sp. strain, enhydrobacter sp. strain, and dna fragment mixture - Google Patents

Abstract

The present invention relates to a cosmetic composition comprising a Staphylococcus sp. strain, a Streptococcus sp. strain, a Cutibacterium sp. strain, a culture broth of an Enhydrobacter sp. strain, and a DNA fragment mixture. The composition can be effectively used in the prevention, amelioration or treatment of skin-related conditions.

Description

A cosmetic composition comprising a mixture of Staphylococcus genus, Streptococcus genus, Cutibacterium genus, and Inhydrobacter genus strains and DNA fragments

It relates to cosmetic compositions.

Microorganisms living on human skin are known to maintain a symbiotic relationship with their human hosts and have various effects on skin aging by interacting with humans. These microorganisms and various information, including their genes, are called skin microbiome. With the recent development of next generation sequencing (NGS), methods for analyzing the microbiome have become easily available, and analysis of the overall distribution of strains existing in the environment has become possible.

Human skin, which is the habitat of these microorganisms, comes in contact with the external environment and serves as a collection site for various microorganisms (e.g., fungi, bacteria, viruses, and small larvae). In addition, the microorganisms prepare a habitat by adapting according to the selection of physical and chemical functions.

Meanwhile, the surface of the skin is cold, shows slightly acidic properties, and remains dry. Structurally, the epidermis forms the skin barrier and plays an important role in blocking the penetration of microorganisms and toxins and maintaining moisture. The uppermost layer of the epidermis is composed of the stratum corneum. The epidermis has a form called a ‘brick and mortar structure’. Skin tissue goes through a continuous self-recovery process, and scales that have completed the differentiation process constantly repeat the process of being shed from the skin tissue. It has been revealed that microorganisms influence these changes in the skin.

In general, in aging skin, wrinkles, decreased skin elasticity, sagging skin, and dryness appear due to changes in matrix proteins present in the dermis. Because the dermis plays a role in supporting the strength and shape of the skin, when morphological changes occur in this area as aging progresses, it plays a critical role in wrinkle formation and skin sagging. Matrix metalloproteinases (MMPs) are representative enzymes that decompose these matrix proteins. Even a single UV irradiation increases the activity of MMPs in the skin, and acts as a major factor in promoting skin aging by decomposing collagen and other matrix proteins in the skin. .

Among skin aging, photoaging is considered the biggest aging factor of the skin, and accordingly, many products to prevent photoaging are being released. However, photoaging prevention technology using natural materials is limited in use and various problems arise when verifying efficacy. There are limitations in many cases.

Therefore, in order to improve the above problems, there is a need to develop functional cosmetics that can exert a synergistic effect in improving skin condition by including a microbial fermentation mixture and functional substances.

One aspect is a Staphylococcus sp. strain, its lysate, culture, extract of the culture, or a mixture thereof; Streptococcus sp. strain, its lysate, culture, extract of the culture, or mixtures thereof; Cutibacterium sp. strain, its lysate, culture, extract of the culture, or mixtures thereof; Enhydrobacter sp. strain, its lysate, culture medium, extract of the culture medium, or mixtures thereof; and a composition for improving skin condition comprising a DNA fragment mixture as an active ingredient.

Another aspect provides a method of preventing, ameliorating, or treating a condition in a subject comprising treating or administering an effective amount of the composition described above to the subject in need thereof.

Another aspect includes the above-mentioned four strains, their lysate, culture medium, extract of the culture medium, or mixtures thereof for use in preparing a composition for improving skin condition; and the use of DNA fragment mixtures.

One aspect is a Staphylococcus sp. strain, its lysate, culture, extract of the culture, or a mixture thereof; Streptococcus sp. strain, its lysate, culture, extract of the culture, or mixtures thereof; Cutibacterium sp. strain, its lysate, culture, extract of the culture, or mixtures thereof; Enhydrobacter sp. strain, its lysate, culture medium, extract of the culture medium, or mixtures thereof; and a DNA fragment mixture as an active ingredient.

As used herein, the term “lysate” may be used interchangeably with “lysate” and may refer to a product obtained by breaking the cell wall of a strain by chemical or physical force. The lysate may include the lysate itself, its concentrate, or its freeze-dried product.

As used herein, the term “culture” may be used interchangeably with “culture supernatant,” “conditioned culture,” or “conditioned medium,” and may be used interchangeably with the genus Staphylococcus, Streptococcus, Cutibacterium, and/or Inhydro. It may refer to the entire medium containing the strain, its metabolites, extra nutrients, etc. obtained by culturing the strain for a certain period of time in a medium that can supply nutrients so that the Bacter strain can grow and survive in vitro. Additionally, the culture medium may refer to a culture medium obtained by removing the bacterial cells from the bacterial culture medium obtained by culturing the strain. On the other hand, the liquid from which the bacteria have been removed from the culture medium is also called "supernatant", and the supernatant is obtained by leaving the culture solution still for a certain period of time and taking only the upper liquid excluding the part that has settled in the lower layer, or by removing the bacteria through filtration, or by removing the culture liquid. It can be obtained by centrifuging to remove the sediment at the bottom and taking only the liquid at the top. The "bacteria" refers to the strain itself of the present invention, and includes the strain itself separated and selected from a sample, etc., or a strain isolated from the culture medium by culturing the strain. The bacterial cells can be obtained by centrifuging the culture medium and taking the part that has sunk to the lower layer. Alternatively, since they sink to the lower layer of the culture medium by gravity, they can be obtained by leaving them still for a certain period of time and then removing the upper liquid.

Culture media and culture conditions for cultivating the above strain can be appropriately selected or modified by those skilled in the art. For example, the culture medium includes strains of the genus Staphylococcus, Streptococcus, Cutibacterium, and/or Inhydrobacter in a medium (e.g., R2A, TSB, TSA, BHI or RCM medium) at an appropriate temperature (e.g. : It may be obtained by culturing for a certain period of time (e.g., 4 to 120 hours) at any temperature above 10°C or below 40°C.

The culture medium may include the culture medium itself, its concentrate, or freeze-dried product obtained by cultivating the strain, or the culture supernatant obtained by removing the strain from the culture medium, its concentrate, or freeze-dried product.

In one embodiment, the culture supernatant of the strain may be obtained by removing the strain by centrifuging or filtering the strain culture fluid.

In another embodiment, the concentrate may be obtained by concentrating the strain culture itself or the supernatant obtained after centrifuging or filtering the culture.

As used herein, the term "culture extract" refers to an extract from the culture medium or its concentrate, and may include extracts, diluted or concentrated extracts, dried products obtained by drying the extracts, crude or purified products thereof, and fractions thereof. You can.

The strain, its lysate, culture medium, or extract of the culture medium may be biologically pure. The strain may be obtained by pure culture.

In one embodiment, a mixture of strains and DNA fragments of the genus Staphylococcus, Streptococcus, Cutibacterium, and Inhydrobacter increase the expression of genetic markers COL1A1 and elastin (ELN), which are reduced by ultraviolet rays, and increase the expression of genes related to skin moisturization. It was confirmed that the expression of factors (eg, HAS3) and skin barrier-related factors (eg, ABCA12) was increased. Therefore, the composition can be used to improve skin condition, improve skin beauty, prevent, improve or treat skin diseases, etc.

The improvement of skin condition or skin beauty may be preventing skin aging (anti-aging), improving skin wrinkles (anti-wrinkle), improving skin elasticity, moisturizing skin, or strengthening the skin barrier.

As used herein, the term “skin aging” refers to both tangible and intangible changes that occur in the skin as one ages. Skin aging includes, for example, a thinning of the epidermis, a decrease in the number of cells or blood vessels in the dermis, a decrease in the ability to repair DNA damage, an increase in the cell turnover cycle, a decrease in the ability to heal wounds, a weakening of the skin barrier function, a decrease in the moisture retention ability of the epidermis, These include reduced sweat secretion, reduced sebum secretion, reduced vitamin D production, reduced ability to defend against physical damage, reduced ability to eliminate chemicals, reduced immune response, weakened sensory function, and reduced body temperature regulation.

The prevention of skin aging may mean improving skin aging caused by extrinsic or endogenous factors. The exogenous factors refer to various external factors, such as ultraviolet rays (light). The endogenous factors are also referred to as chronological factors and mainly refer to factors that occur due to the passage of time. In other words, the skin aging is specifically not only a symptom of premature aging induced by external stimuli such as ultraviolet rays, pollution, cigarette smoke, chemicals, etc., but also a natural aging phenomenon that occurs as the proliferation of skin cells decreases with age. It is a concept that includes wrinkles, loss of elasticity, skin sagging and dryness, etc. In addition, wrinkles include stimulation caused by changes in internal and external factors that change the components that make up skin tissue, causing wrinkles.

The aging may be photoaging. The term “photoaging” is a phenomenon caused by external environmental factors, the most representative factor being ultraviolet rays. Ultraviolet rays cause damage to biological components such as activation of proteolytic enzymes, chain cutting of matrix proteins, and abnormal cross-linking, and repetition of this mechanism causes skin aging that is evident in appearance.

The term “wrinkle” refers to a state in which the skin loses its elasticity and becomes loose, for example, the skin may be folded. The term “prevention or improvement of skin wrinkles” may refer to any action that prevents or improves wrinkles by suppressing the expression of factors related to wrinkles, or increases the total amount of collagen.

The term “skin moisturizing” may refer to any action that maintains skin moisture or prevents moisture loss.

The “skin barrier strengthening” may refer to any action that improves the function of the skin barrier, which is located on the outermost layer of the skin and prevents moisture and nutrition loss.

The “skin disease” may be a disease caused by damage to the skin barrier function, skin aging, skin wound, skin scar, or skin inflammation.

The term “prevention” includes suppressing the occurrence of a disease. The term “treatment” includes inhibiting, alleviating, or eliminating the development of a disease. The term “improvement” may refer to any action that results in at least reducing the severity of a parameter associated with the alleviation or treatment of a condition, such as symptoms.

The damage to the skin barrier function may mean any change that appears in the skin due to decreased or damaged skin barrier function. For example, it may include increased skin wrinkles, dryness, dermatitis, atopic dermatitis, allergic dermatitis, acne, etc.

The Staphylococcus sp. strains include, for example, Staphylococcus epidermidis , Staphylococcus hominis , Staphylococcus warneri , Staphylococcus caprae , It may be Staphylococcus capitis , etc.

In one embodiment, the Staphylococcus genus strain may be Staphylococcus epidermidis (Accession Number: KCCM12559P).

In another embodiment, the Staphylococcus genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof may be included in an amount of 5 to 20% by weight based on the total weight of the composition. The Staphylococcus genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof is, for example, 5 to 20% by weight, 5 to 18% by weight, 5 to 16% by weight, 7 to 7% by weight, based on the total weight of the composition. It may be included at 20% by weight, 7 to 17% by weight, 8 to 15% by weight, or 10 to 13% by weight. At this time, if the content of the Staphylococcus genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof is less than or exceeds the above range, there is a problem that the effect of improving skin condition cannot be sufficiently exerted.

The Streptococcus genus ( Streptococcus sp.) strains are, for example, Streptococcus thermophiles, Streptococcus mitis , Streptococcus infantis , It may be Streptococcus pneumoniae , etc.

In one embodiment, the Streptococcus genus strain may be Streptococcus thermophiles (Accession number: KCCM12004P).

In another embodiment, the Streptococcus genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof may be included in an amount of 5 to 30% by weight based on the total weight of the composition. The Streptococcus genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof is contained in an amount of 5 to 30% by weight, 5 to 28% by weight, 5 to 25% by weight, 5 to 20% by weight, 8%, based on the total weight of the composition. It may be included in an amount of from 30% by weight, 8 to 24% by weight, 8 to 18% by weight, 10 to 27% by weight, 10 to 20% by weight, or 10 to 15% by weight. At this time, if the content of the Streptococcus genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof is less than or exceeds the above range, there is a problem that the effect of improving skin condition cannot be sufficiently exerted.

The Cutibacterium sp. strain may be, for example, Cutibacterium acnes , Cutibacterium granulosum , etc.

In one embodiment, the strain of the Cutibacterium genus may be Cutibacterium acnes (Accession Number: KCCM12680P).

In another embodiment, the Cutibacterium genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof may be included in an amount of 50 to 90% by weight based on the total weight of the composition. The Cutibacterium genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof is, for example, 50 to 90% by weight, 50 to 80% by weight, 50 to 70% by weight, 55 to 55% by weight, based on the weight of the species of the composition. It may be included at 90% by weight, 55 to 85% by weight, 55 to 75% by weight, 60 to 90% by weight, 60 to 80% by weight, or 70 to 75% by weight. At this time, if the content of the Cutibacterium genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof is less than or exceeds the above range, there is a problem that the skin condition improvement effect cannot be sufficiently exerted.

The Enhydrobacter sp. strain may be, for example, Enhydrobacter aerosaccus (Accession number: KCCM13073P).

In one embodiment, the Inhydrobacter genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof may be included in an amount of 0.5 to 10% by weight based on the total weight of the composition. The Inhydrobacter genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof is, for example, 0.5 to 10% by weight, 0.5 to 8% by weight, 0.5 to 5% by weight, 1 to 1% by weight, based on the total weight of the composition. It may be included at 7% by weight, 1 to 6% by weight, 2 to 9% by weight, or 2 to 5% by weight. At this time, if the content of the Inhydrobacter genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof is less than or exceeds the above range, there is a problem that the effect of improving skin condition cannot be sufficiently exerted.

As used herein, the term "DNA fragment mixture" is a form in which DNA corresponding to a biopolymer composed of phosphoric acid, four types of bases, and deoxyribose is extracted and exists in a mixture of DNA fragments having various molecular weights.

In one embodiment, the DNA fragment mixture may be hydrolyzed.

The DNA fragment mixture may be, for example, polydeoxyribonucleotide (PDRN), polynucleotide, etc. The polydeoxyribonucleotide (PDRN) is a mixture of short deoxyribonucleotides, a low molecular weight DNA complex made by fractionating DNA chains to a certain size. It's kind of like that. In addition, the polynucleotide refers to a DNA or RNA strand of a certain length or more, which is a polymer of nucleotides in which nucleotide monomers are connected in a chain shape by covalent bonds. For example, the polynucleotide may have a relatively longer nucleic acid length or a higher molecular weight than a polydeoxyribonucleotide, but is not limited thereto.

In another embodiment, the molecular weight of the DNA fragment mixture may be 50 to 10,000 kDa. The molecular weight of the DNA fragment mixture may be, for example, 50 to 10,000 kDa, 50 to 5,000 kDa, 50 to 3,000 kDa, 50 to 1,500 kDa, 1,000 to 10,000 kDa, 1,000 to 5,000 kDa, or 5,000 to 10,000 kDa.

In another embodiment, the DNA fragment mixture may be a mixture of DNA fragments obtained by extraction from testes or semen of a fish, and the fish may be a salmonid, specifically salmon or trout.

In another embodiment, the DNA fragment mixture may be included in an amount of 0.01 to 1% by weight based on the total weight of the composition. The DNA fragment mixture may be included, for example, at 0.01 to 1% by weight, 0.01 to 0.7% by weight, 0.03 to 0.7% by weight, 0.1 to 0.8% by weight, or 0.3 to 0.6% by weight, based on the total weight of the composition. At this time, if the content of the DNA fragment mixture is less than or exceeds the above range, there is a problem that the skin condition improvement effect cannot be sufficiently exhibited.

The composition may have synergistic activity in improving skin condition by containing strains of the genus Staphylococcus, Streptococcus, Cutibacterium, and Inhydrobacter and a mixture of DNA fragments in a specific weight ratio.

In one embodiment, the Staphylococcus genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof; Streptococcus strains, their lysate, culture medium, extract of the culture medium, or mixtures thereof; Cutibacterium genus strains, their lysate, culture medium, extract of the culture medium, or mixtures thereof; Inhydrobacter genus strains, their lysate, culture medium, extract of the culture medium, or mixtures thereof; And the DNA fragment mixture may be mixed at a weight ratio of 1:1 to 2:5 to 7:0.1 to 0.5:0.01 to 0.06. The mixture of strains and DNA fragments of the Staphylococcus genus, Streptococcus genus, Cutibacterium genus, and Inhydrobacter genus is, for example, 1:1 to 2:5 to 7:0.1 to 0.5:0.01 to 0.06, 1:1 It may be mixed at a weight ratio of 1.5:5 to 6.5:0.1 to 0.3:0.01 to 0.05, or 1:1 to 1.3:5.5 to 7:0.2 to 0.5:0.03 to 0.05. At this time, when the mixing weight ratio of the Staphylococcus genus, Streptococcus genus, Cutibacterium genus, Inhydrobacter genus strain and DNA fragment mixture is less than or exceeds the above range, the skin condition improvement effect can be sufficiently exhibited. There is a problem that there is no.

Accordingly, the composition according to one aspect includes a Staphylococcus sp. strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof; Streptococcus sp. strain, its lysate, culture, extract of the culture, or mixtures thereof; Cutibacterium sp. strain, its lysate, culture, extract of the culture, or mixtures thereof; Enhydrobacter sp. strain, its lysate, culture medium, extract of the culture medium, or mixtures thereof; and a DNA fragment mixture in a specific content and weight ratio can exert a synergistic effect in improving skin condition.

As used herein, the term "included as an active ingredient" refers to the strain of the present specification, its lysate, culture medium, or extract of the culture medium to the extent that it can exhibit the above-mentioned effects; and/or DNA fragment mixture is added, and includes formulation in various forms by adding various ingredients as sub-ingredients for drug delivery and stabilization.

In one embodiment, the composition may be a cosmetic composition.

In addition to the active ingredients disclosed in this specification, the cosmetic composition may further include ingredients commonly used in cosmetic compositions, functional additives, etc., such as antioxidants, stabilizers, solubilizers, surfactants, dispersants, emulsifiers, and preservatives. , conventional auxiliaries such as vitamins, pigments, fragrances, etc., and carriers.

The cosmetic composition is not particularly limited to a specific formulation, and the formulation may be appropriately selected depending on the purpose. The cosmetic composition may have, for example, a solubilized formulation, an emulsified formulation, or a dispersed formulation. The cosmetic composition includes, for example, lotion, cream, essence, cleansing foam, cleansing water, pack, ampoule, body lotion, body oil, body gel, shampoo, rinse, hair conditioner, hair gel, foundation, lipstick, mascara, and makeup. It may have a base or skin-adhesive cosmetic formulation. The skin-adhesive cosmetic formulation may be, for example, a mask pack, but is not limited thereto.

In another embodiment, the composition may be a composition for external application to the skin.

The external skin preparation may be a cream, gel, ointment, skin emulsifier, skin suspension, transdermal delivery patch, drug-containing bandage, lotion, or a combination thereof. The skin external preparations include ingredients commonly used in external skin preparations such as cosmetics and medicines, such as aqueous ingredients, oil-based ingredients, powder ingredients, alcohols, moisturizers, thickeners, ultraviolet absorbers, whitening agents, preservatives, antioxidants, surfactants, and fragrances. , colorants, various skin nutrients, metal ion sequestrants, sugars, or combinations thereof may be appropriately mixed as needed.

In another embodiment, the composition may be a health functional food composition.

The health functional food composition may be used alone or in combination with other foods or food ingredients, and may be used appropriately according to conventional methods. The mixing amount of the active ingredient can be appropriately determined depending on the purpose of use (prevention, health, or therapeutic treatment). In general, when manufacturing food or beverages, the composition of the present specification may be added in an amount of 15 parts by weight or less based on the raw materials. There are no particular restrictions on the types of health functional foods. Among the types of health functional foods, beverage compositions may contain various flavoring agents or natural carbohydrates as additional ingredients like ordinary beverages. The natural carbohydrates include monosaccharides such as glucose and fructose; Disaccharides such as maltose and sucrose; and polysaccharides such as dextrins and cyclodextrins; Sugar alcohols such as xylitol, sorbitol, and erythritol. As a sweetener, natural sweeteners such as thaumatin and stevia extract or synthetic sweeteners such as saccharin and aspartame can be used. The health food composition also contains nutrients, vitamins, electrolytes, flavors, colorants, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, and carbonated beverages. It may contain the carbonating agent used, or a combination thereof. The health functional food composition may also contain pulp for the production of natural fruit juice, fruit juice beverage, vegetable beverage, or a combination thereof.

Another aspect provides a method of preventing, ameliorating, or treating a condition in a subject comprising treating or administering an effective amount of the composition described above to the subject in need thereof.

The condition of the subject may be a condition related to skin-related conditions, and specifically, may be a disease caused by damage to the skin barrier function or skin aging.

The term “effective amount” refers to an amount effective enough to produce the above-mentioned effects.

As used herein, the terms “administering,” “introducing,” and “implanting” are used interchangeably and refer to an individual by a method or route that results in at least partial localization of the composition according to one embodiment to the desired site. It may refer to the arrangement of the composition according to one embodiment into the inner space.

Administration may be by methods known in the art. The administration method, such as administration route and administration frequency, can be appropriately selected by a person skilled in the art. Administration may be administered directly to the subject by any means, such as, for example, intravenous, intramuscular, oral, transdermal, mucosal, intranasal, intratracheal or subcutaneous administration. You can. The administration may be administered systemically or locally. The administration may include application to the skin.

The subject may be a mammal, such as a human, cow, horse, pig, dog, sheep, goat, or cat. The subject may be an entity that requires improvement of skin condition, for example, an entity that requires anti-aging skin, improvement of skin wrinkles, enhancement of skin elasticity, skin moisturization, strengthening of skin barrier, and inhibition of skin inflammation.

The administration of the composition according to one embodiment is 0.1 mg to 1,000 mg, for example, 0.1 mg to 500 mg, 0.1 mg to 100 mg, 0.1 mg to 50 mg, 0.1 mg to 25 mg, 1 mg to 1 mg per day. 1,000 mg, 1 mg to 500 mg, 1 mg to 100 mg, 1 mg to 50 mg, 1 mg to 25 mg, 5 mg to 1,000 mg, 5 mg to 500 mg, 5 mg to 100 mg, 5 mg to 50 mg , 5 mg to 25 mg, 10 mg to 1,000 mg, 10 mg to 500 mg, 10 mg to 100 mg, 10 mg to 50 mg, or 10 mg to 25 mg. However, the dosage may be prescribed in various ways depending on factors such as formulation method, administration method, patient's age, weight, gender, pathological condition, food, administration time, administration route, excretion rate, and reaction sensitivity, and those skilled in the art will Taking these factors into consideration, the dosage can be adjusted appropriately. The frequency of administration can be once a day or two or more times within the range of clinically acceptable side effects. Regarding the administration site, it can be administered at one or two or more locations, and the total number of administration days can be from 1 to 30 days for one treatment, either daily or at intervals of 2 to 5 days. If necessary, the same treatment can be repeated after an appropriate period. For animals other than humans, the dosage per kg is the same as for humans, or the above dosage is converted into, for example, the volume ratio (e.g., average value) of the organs (heart, etc.) between the target animal and human. One dose can be administered.

Another aspect includes the above-mentioned four strains, their lysate, culture medium, extract of the culture medium, or mixtures thereof for use in preparing a composition for improving skin condition; and uses of DNA fragment mixtures.

Redundant content is omitted in consideration of the complexity of the present specification, and terms not otherwise defined in the present specification have meanings commonly used in the technical field to which the present invention pertains.

The composition according to one aspect increases the expression of collagen, elastin, HAS3, and ABCA12, and can be usefully used to improve skin conditions such as skin aging, skin moisturizing, skin barrier strengthening, skin wrinkles, or skin elasticity.

Figure 1 is a result showing the effect of a composition according to one aspect on COL1A1 expression.

Figure 2 is a result showing the effect of a composition according to one aspect on ELN expression.

Figure 3 is a result showing the effect of a composition according to one aspect on HAS3 expression.

Figure 4 is a result showing the effect of a composition according to one aspect on ABCA12 expression.

Below, preferred embodiments are presented to aid understanding of the present invention. However, the following examples are provided only to make the present invention easier to understand, and the content of the present invention is not limited by the following examples.

[Manufacturing example]

Preparation Example 1. Preparation of Staphylococcus epidermidis strain culture medium

Staphylococcus epidermidis strain culture medium was prepared. Specifically, Staphylococcus epidermidis strain (Accession number: KCCM12559P) was received from the Korean Culture Center of Microorganisms (KCCM) and grown at 150 rpm/min in R2A (Reasoner's 2A) medium. It was suspended and cultured for enrichment. The temperature was maintained at about 30°C during the culturing period, and after culturing for 72 hours, the culture medium was filtered using a 0.45 ㎛ filter.

Preparation Example 2. Preparation of Streptococcus thermophilus strain culture medium

Streptococcus thermophiles strain (Accession number: KCCM12004P) and TSB (Tryptic Soy Broth) medium were used, except that Streptococcus thermophiles strain culture medium was prepared in the same manner as in Preparation Example 1 . did.

Preparation Example 3. Preparation of Coutibacterium acnes strain culture medium

Cutibacterium acnes strain (Accession number: KCCM12680P) was prepared in the same manner as in Preparation Example 1 except that it was stationary cultured using RCM (Reinforced Clostridial Medium) medium. Culture medium was prepared.

Preparation Example 4. Preparation of Inhydrobacter Aerosarcus strain culture medium

An Enhydrobacter aerosaccus strain culture medium was prepared in the same manner as in Preparation Example 3, except that the Enhydrobacter aerosaccus strain (accession number: KCCM13073P) was used.

[Example]

Example 1. Preparation of a composition comprising strain culture medium and DNA fragment mixture

A composition containing the ingredients and contents shown in Table 1 below was prepared. Specifically, a composition was prepared by mixing the culture media of Preparation Examples 1 to 4 with hydrolyzed DNA (manufactured by Pharma Research Co., Ltd.) with a molecular weight of 50 to 1,500 kDa. The hydrolyzed DNA is a mixture of DNA fragments in a white or off-white powder. Unless otherwise stated herein, content is in weight percent.

ingredient	content
Manufacturing Example 1	11
Production example 2	14
Production example 3	71.5
Production example 4	3
hydrolyzed DNA	0.5

[Comparative example]

Comparative Example 1. Preparation of a composition containing strain culture medium

The composition was prepared in the same manner as Example 1, except that it included the ingredients and contents shown in Table 2 below.

Comparative Example 1 has the same mixing ratio of the strain culture medium as Example 1, but differs in that it contains sterilized distilled water instead of hydrolyzed DNA.

ingredient	Comparative Example 1
Manufacturing Example 1	11
Production example 2	14
Production example 3	71.5
Production example 4	3
sterile distilled water	0.5

Comparative Example 2. Preparation of a composition containing strain culture medium and DNA fragment mixture

The composition was prepared in the same manner as Example 1, except that it included the ingredients and contents shown in Table 3 below.

Comparative Example 2 differs from Example 1 in the mixing ratio of the strain culture medium.

ingredient	Comparative Example 2
Manufacturing example 1	24.875
Production example 2	24.875
Production example 3	24.875
Production example 4	24.875
hydrolyzed DNA	0.5

[Experimental example]

Experimental Example 1. Confirmation of anti-aging, anti-wrinkle, and skin elasticity promotion activities

The skin aging, wrinkle improvement, and skin elasticity enhancing activities of the composition according to one aspect were confirmed.

1-1. Cell culture and sample processing

Human dermal fibroblast (Hs68) cell line was distributed at 3.5x10 ⁵ in a 6-well plate and cultured in an incubator at 37°C and 5% CO ₂ for 24 hours. Afterwards, the medium was removed, DPBS (Dulbecco's phosphate-buffered saline) was added, and 20 mJ/cm ² UVB was irradiated or not. Immediately after UVB irradiation, DPBS was removed and replaced with medium without FBS fetal bovin serum, and then treated with the composition prepared in the above Examples and Comparative Examples and further cultured for 24 hours. As a negative control group, a group untreated with ultraviolet rays and strain culture was used.

1-2. Gene expression level analysis

Afterwards, RNA was isolated from the cells of each sample using Trizol (RNA iso, DAKARA, Japan), RNA was quantified at 260 nm using a nanodrop, and 2 μg of RNA was used in an amplifier. cDNA was synthesized (C1000 Thermal Cycler, Bio-Rad, USA). Using the synthesized cDNA as a template, Cybergreen (SYBR Green supermix, Applied Biosystems, USA) and primers (primers for COL1A1 or primers for ELN) were added to a real-time PCR machine (Step One Plus, AppliedBiosystems, USA). Real-time polymerase chain reaction was performed. The expression level of the gene was finally analyzed through correction for the β-actin gene.

1-3. result

Figure 1 is a result showing the effect of a composition according to one aspect on COL1A1 expression.

Figure 2 is a result showing the effect of a composition according to one aspect on ELN expression.

As a result, as shown in Figures 1 and 2, the expression of COL1A1 and elastin in the fibroblast cell line was decreased by ultraviolet irradiation, and the expression of COL1A1 and elastin was increased by the treatment of Example 1 and Comparative Examples 1 and 2. I was able to confirm that Specifically, in Example 1, it was confirmed that the expression of COL1A1 and elastin was significantly increased compared to Comparative Examples 1 and 2.

That is, the composition according to one aspect increases the expression of COL1A1 and elastin, which are reduced by ultraviolet rays, by including four types of strain cultures and a mixture of DNA fragments as active ingredients, thereby preventing skin aging (e.g. photoaging) (anti-aging) ), is effective in improving wrinkles (anti-wrinkle) and improving skin elasticity.

Experimental Example 2. Confirmation of skin moisturizing activity

The skin moisturizing activity of the composition according to one aspect was confirmed.

2-1. Cell culture and sample processing

HaCaT cells, a human keratinocyte cell line, were cultured in DMEM medium (Dulbecco's modified Eagle's Medium, Gibco 1210-0038) containing 10% fetal bovine serum, and all cultures were performed at 37°C and 5% CO ₂ It was performed in an incubator. The cultured cell lines were treated with the compositions prepared in the Examples and Comparative Examples and further cultured for 24 hours.

2-2. Gene expression level analysis

The HAS3 gene expression level was analyzed using the same method as Experimental Example 1-2, except that primers for HAS3 (hyaluronic acid synthase) were used.

2-3. result

Figure 3 is a result showing the effect of a composition according to one aspect on HAS3 expression.

As a result, as shown in Figure 3, it was confirmed that HAS3 expression in Example 1 was significantly increased compared to Comparative Examples 1 and 2.

That is, the composition according to one aspect significantly increases the expression of HAS3 by including four types of strain cultures and a mixture of DNA fragments as active ingredients, and can be usefully used to improve skin moisturization.

Experimental Example 3. Confirmation of skin barrier function improvement activity

The skin barrier function improvement activity of the composition according to one aspect was confirmed.

3-1. Cell culture and sample processing

HaCaT cells, a human keratinocyte cell line, were treated with polyIC (polyinosinic:polycytidylic acid; Poly I:C), a synthetic ribonucleic acid that promotes interferon production, and then cultured for 24 hours to stimulate the HaCaT cell line. Thereafter, cells aged by stimulation were treated with 1% (w/w) of the composition prepared in the above Examples and Comparative Examples and further cultured for 24 hours.

3-2. Gene expression level analysis

Afterwards, RNA was extracted and quantified using nanodrop. 16 ㎕ of RNA was quantified and treated at 70°C for 5 minutes, then added to PCR premix and gently vortexed. Afterwards, cDNA was synthesized by treating at 42°C for 55 minutes, at 70°C for 15 minutes, and storing at 4°C. The synthesized cDNA was amplified using PCR. After mixing 2 ㎕ of cDNA, 1 ㎕ of forward primer targeting the ABCA12 (ATP-binding cassette sub-family A member 12) gene, 1 ㎕ of reverse primer, 6 ㎕ of distilled water, and 10 ㎕ of premix sample, the mixture was stored at 95°C. Amplification was performed through 40 cycles of 3 minutes, {95°C 30 seconds, 49 to 53°C 30 seconds, 72°C 1 minute}.

3-3. result

Figure 4 is a result showing the effect of a composition according to one aspect on ABCA12 expression.

As a result, as shown in Figure 4, it was confirmed that ABCA12 expression in Example 1 was significantly increased compared to Comparative Examples 1 and 2.

That is, the composition according to one aspect significantly increases the expression of ABCA12 by including four types of strain cultures and a mixture of DNA fragments as active ingredients, and can be usefully used to improve the skin barrier.

In summary, the composition according to one aspect includes a mixture of four types of strain cultures and DNA fragments, thereby (i) increasing the expression of COL1A1 and ELN reduced by ultraviolet rays, (ii) increasing the expression of HAS3 effect, (iii) has the effect of increasing the expression of ABCA12. In addition, it was confirmed that the effects of (i) to (iii) were significantly better than those of Comparative Example 1, which did not contain a mixture of DNA fragments, or Comparative Example 2, which had a different mixing ratio of the strain culture medium.

The description of the present invention described above is for illustrative purposes, and those skilled in the art will understand that the present invention can be easily modified into other specific forms without changing the technical idea or essential features of the present invention. will be. Therefore, the embodiments described above should be understood in all respects as illustrative and not restrictive.

Claims (11)

1. Staphylococcus sp. strain, its lysate, culture, extract of the culture, or mixtures thereof;

   Streptococcus sp. strain, its lysate, culture, extract of the culture, or mixtures thereof;

   Cutibacterium sp. strain, its lysate, culture, extract of the culture, or mixtures thereof;

   Enhydrobacter sp. strain, its lysate, culture medium, extract of the culture medium, or mixtures thereof; and

   A cosmetic composition for improving skin condition comprising a mixture of DNA fragments as an active ingredient.
2. The cosmetic composition according to claim 1, wherein the improving skin condition includes preventing skin aging, improving skin wrinkles, improving skin elasticity, moisturizing skin, or strengthening the skin barrier.
3. The cosmetic composition according to claim 1, wherein the Staphylococcus genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof is contained in an amount of 5 to 25% by weight based on the total weight of the composition.
4. The cosmetic composition according to claim 1, wherein the Streptococcus genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof is contained in an amount of 5 to 30% by weight based on the total weight of the composition.
5. The cosmetic composition according to claim 1, wherein the Cutibacterium genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof is contained in an amount of 50 to 90% by weight based on the total weight of the composition.
6. The cosmetic composition according to claim 1, wherein the Inhydrobacter genus strain, its lysate, culture medium, extract of the culture medium, or a mixture thereof is contained in an amount of 0.5 to 10% by weight based on the total weight of the composition.
7. The cosmetic composition according to claim 1, wherein the DNA fragment mixture is contained in an amount of 0.01 to 1% by weight based on the total weight of the composition.
8. The cosmetic composition according to claim 1, wherein the DNA fragment mixture is polydeoxyribonucleotide, polynucleotide, or a mixture thereof.
9. The cosmetic composition according to claim 1, wherein the molecular weight of the DNA fragment mixture is 50 to 10,000 kDa.
10. In claim 1,

    The Staphylococcus genus strain, its lysate, culture medium, extract of the culture medium, or mixtures thereof;

    Streptococcus strains, their lysate, culture medium, extract of the culture medium, or mixtures thereof;

    Cutibacterium genus strains, their lysate, culture medium, extract of the culture medium, or mixtures thereof;

    Inhydrobacter genus strains, their lysate, culture medium, extract of the culture medium, or mixtures thereof; and

    A cosmetic composition in which the DNA fragment mixture is mixed at a weight ratio of 1:1 to 2:5 to 7:0.1 to 0.5:0.01 to 0.06.
11. Staphylococcus sp. strain, its lysate, culture, extract of the culture, or mixtures thereof;

    Streptococcus sp. strain, its lysate, culture, extract of the culture, or mixtures thereof;

    Cutibacterium sp. strain, its lysate, culture, extract of the culture, or mixtures thereof;

    Enhydrobacter sp. strain, its lysate, culture medium, extract of the culture medium, or mixtures thereof; and

    An external skin composition for improving skin condition comprising a DNA fragment mixture as an active ingredient.

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