Cosmetic Composition for Enhanced Antioxidant Stability Containing Natural Extracts and Method for Preparing the Same

Abstract

The present invention relates to a cosmetic composition comprising a sphagnum extract, a fermented liquid derived from a microorganism of the genus *Psychobacter* or an extract thereof, and ellagic acid. The composition effectively inhibits metal ion-mediated oxidation reactions even in humid environments and under conditions where metal ions are present, thereby maintaining the stability of the antioxidant component. Accordingly, the generation of reactive oxygen species is reduced, and formulation stability is improved, providing an excellent antioxidant effect even during long-term storage and use.

Description

Technology Field

[0001] The present invention relates to a cosmetic composition for improving antioxidant stability containing a natural extract and a method for preparing the same.

[0002] The present invention relates to a cosmetic composition for improving antioxidant stability containing a natural extract, characterized by including a sphagnum extract, a Psychrobacter spp. microbial fermentation liquid, and ellagic acid, and a method for preparing the same.

[0003] More specifically, the present invention relates to a cosmetic composition for improving antioxidant stability containing a natural extract in which oxidation reactions mediated by metal ions are suppressed, thereby maintaining antioxidant stability even in a humid environment where moisture is present, and a method for manufacturing the same.

[0004] In particular, the present invention relates to a cosmetic composition for skincare that effectively suppresses oxidation reactions promoted by the presence of metal ions, such as Fe(II) ions or Fe(III) ions, on the skin surface or within a cosmetic formulation, thereby ensuring that the stability of the antioxidant component and the skin protective effect are not reduced even under conditions where the skin is moist, such as immediately after washing the face, when sweat or sebum secretion is high, or when wearing a mask. Background Technology

[0005] Generally, cosmetic compositions for skincare contain various antioxidant ingredients to inhibit skin aging and protect the skin from the external environment. These antioxidant ingredients have the function of alleviating skin damage by eliminating free radicals or inhibiting oxidation reactions.

[0006] However, conventional antioxidant cosmetic compositions have a problem in that the stability of the antioxidant component is reduced in a humid environment where moisture is present. In particular, under conditions such as wet skin immediately after washing, skin conditions with excessive sweat or sebum secretion, or high humidity of the skin surface due to wearing a mask, oxidation reactions can be promoted by metal ions such as Fe(II) ions or Fe(III) ions present in the cosmetic composition or on the skin surface.

[0007] Such metal ions can induce oxidation pathways, such as the Fenton reaction, which generates reactive oxygen species by reacting with peroxides in the presence of moisture. This can lead to the premature oxidation and decomposition of the antioxidant components themselves, or cause oxidative stress on the skin. As a result, the sustainability of the antioxidant effect is reduced, and in the case of sensitive skin, there is a possibility of redness or irritation.

[0008] Conventional technologies have attempted to solve this problem by increasing the content of antioxidant components or by using simple combinations of antioxidants, but there were limitations in fundamentally suppressing oxidation reactions occurring in a humid environment where moisture and metal ions are present simultaneously. In other words, although existing cosmetic compositions possess antioxidant functions, they still contain the problem that antioxidant stability is not sufficiently ensured under humid conditions, which are the actual usage environment. The problem to be solved

[0009] The present invention was devised to solve the problems of the aforementioned prior art, and aims to solve the problem of reduced stability of antioxidant components due to oxidation reactions mediated by metal ions such as Fe(II) ions or Fe(III) ions in a humid environment where moisture is present.

[0010] Specifically, the present invention aims to solve the problem in which the sustainability of the antioxidant effect is reduced and skin irritation or redness may be induced under conditions of high skin surface humidity, such as wet skin immediately after washing, a state of excessive sweat or sebum secretion, or wearing a mask, as the antioxidant component in the cosmetic composition is easily oxidized and decomposed by a metal ion-mediated oxidation reaction.

[0011] Accordingly, the objective of the present invention is to provide a cosmetic composition in which the stability of the antioxidant component and the skin protective effect are stably maintained even in a humid environment by effectively suppressing metal ion-mediated oxidation reactions. means of solving the problem

[0012] To solve the above-mentioned problem, the present invention provides a cosmetic composition comprising a sphagnum extract, a fermented liquid or extract obtained by fermenting microorganisms of the genus Psychrobacter spp., and ellagic acid.

[0013] Specifically, the cosmetic composition according to the present invention forms a stable moisture matrix in a humid environment through a spargnum extract containing a polysaccharide component derived from a porous cell structure, and maintains relatively stable physical properties even in the presence of moisture through a fermented component derived from a microorganism of the genus *Psychobacter* adapted to a low-temperature environment. In addition, by including ellagic acid having a polyphenolic structure, it effectively inhibits metal ion-mediated oxidation reactions through the formation of complex compounds with Fe(II) ions or Fe(III) ions.

[0014] Accordingly, the cosmetic composition of the present invention inhibits the decomposition of antioxidant components and reduces the generation of reactive oxygen species even in a humid environment where moisture is present and under conditions where metal ions are present, thereby ensuring both antioxidant stability and formulation stability simultaneously. Effects of the invention

[0015] The cosmetic composition according to the present invention comprises a sphagnum extract, a fermented liquid derived from microorganisms of the genus Psychrobacter spp. or the extract thereof, and ellagic acid, thereby providing the effect of effectively inhibiting metal ion-mediated oxidation reactions even in a humid environment where moisture is present and under conditions where metal ions are present.

[0016] Specifically, the cosmetic composition according to the present invention has a significantly improved retention rate of antioxidant components under moist conditions, so that antioxidant stability is maintained even in long-term storage or usage environments. In addition, the generation of reactive oxygen species is significantly reduced in the presence of metal ions, so that oxidative stress induced by metal ions can be effectively relieved.

[0017] In addition, the cosmetic composition according to the present invention has the effect of maintaining excellent formulation stability by suppressing phase separation, precipitation, or microstructural collapse even under conditions of humidity and the presence of metal ions. Accordingly, the composition can maintain a stable appearance and physical properties in actual usage environments.

[0018] Accordingly, the cosmetic composition according to the present invention can simultaneously solve the problems of the prior art, such as reduced stability of antioxidant components in a humid environment and metal ion-mediated oxidation reactions, and can be usefully applied to skincare products that require both antioxidant stability and formulation stability. Brief explanation of the drawing

[0019] FIG. 1 is a graph showing the comparison of the ellagic acid retention rates of the test group and the comparison group under wet conditions according to Test Example 1 of the present invention. FIG. 2 is a graph showing the amount of active oxygen (ROS) produced in the test group and the control group in the presence of metal ions under wet conditions according to Test Example 2 of the present invention. FIG. 3 is a drawing showing photographs comparing the appearance of cosmetic compositions before storage in the test group and the comparison group according to Test Example 3 of the present invention. FIG. 4 is a diagram showing photographs comparing the appearance of the test group and the control group after storage under conditions of moisture and metal ion presence (moisture 20 wt%, Fe(III) 100 μM, 40℃, relative humidity 90%, 7 days) according to Test Example 3 of the present invention. FIG. 5 is a photograph showing the results of optical microscope observation of cosmetic compositions of the test group and the comparison group after storage under conditions of moisture and the presence of metal ions according to Test Example 3 of the present invention, and illustrates an exemplary sample in which a change in microstructure was representatively observed among the test group and the comparison example. Specific details for implementing the invention

[0020] Prior to the description of the present invention, the following specific structural or functional descriptions are provided merely for the purpose of illustrating embodiments according to the concept of the present invention. Embodiments according to the concept of the present invention may be implemented in various forms and should not be interpreted as being limited to the embodiments described herein.

[0021] In addition, since embodiments according to the concept of the present invention may be subject to various modifications and may take various forms, specific embodiments are to be described in detail in this specification. However, this is not intended to limit the embodiments according to the concept of the present invention to specific disclosed forms, and it should be understood that they include all modifications, equivalents, and substitutions that fall within the spirit and scope of the present invention.

[0023] The present invention relates to a cosmetic composition for improving antioxidant stability containing a natural extract.

[0024] The cosmetic composition for improving antioxidant stability containing the above natural extract comprises Sphagnum extract, a fermented liquid or extract obtained by fermenting microorganisms of the genus Psychrobacter spp., and ellagic acid as essential components.

[0025] Although each of the above components can perform a certain function on its own, the present invention combines them to suppress metal ion-mediated oxidation reactions even in a humid environment where moisture is present, thereby exhibiting an effect of improved antioxidant stability.

[0027] The spagnum extract (10) of the present invention will be described.

[0028] The above spagnum extract (10) may be an extract obtained using the whole spagnum plant as a raw material, and preferably contains a polysaccharide component derived from a porous cell structure.

[0030] As used in this specification, Sphagnum refers to a plant of the genus Sphagnum belonging to the phylum Bryophyta, which is a moss that grows in wetlands, peatlands, or high-humidity environments.

[0031] The above-mentioned spargnum is characterized by having a porous cell structure, possessing the ability to store a large amount of water within the cell, and due to this structure, maintains relatively stable physical properties even in humid environments. Additionally, spargnum is known to have a cell wall structure containing polysaccharide components. In the present invention, spargnum is used as a concept encompassing the entire plant body (thallus, stem, and mixtures thereof) and can be used in any one of the forms of fresh, dried, or ground. Furthermore, spargnum is not limited to a single species but may include various species belonging to the genus Spagnamus.

[0032] The above-mentioned Spagnum may be *Sphagnum squarrosum Crome*. This *Sphagnum squarrosum* is characterized by having a scale-like arrangement structure in which the thallus spreads outward. Combined with a porous cell structure, it effectively stores moisture while maintaining relatively excellent structural stability in humid environments. Furthermore, compared to other plants of the genus *Sphagnum*, *Sphagnum squarrosum* exhibits a better balance between moisture storage and diffusion, resulting in relatively less change in viscosity or physical property instability of the cosmetic composition when the extract is applied. Accordingly, it is more suitable for the purpose of the present invention, which requires the maintenance of antioxidant stability even in humid environments. Moreover, as a plant adapted to high humidity and relatively low temperatures, *Sphagnum squarrosum* has high environmental compatibility with the low-temperature fermented microorganism-derived components included in the present invention, which is advantageous for ensuring the overall stability and consistent quality of the composition. Therefore, among plants of the genus *Sphagnum*, it is particularly preferable to use *Sphagnum squarrosum* in the present invention.

[0034] The above spargnum extract (10) can be used after removing foreign substances from the spargnum plant body after collection, and after undergoing washing and drying processes. Subsequently, extraction is performed under conditions of 20°C to 60°C using purified water, ethanol, or a mixed solvent thereof. The extraction method may be one or more of immersion extraction, stirring extraction, or reflux extraction.

[0035] After extraction, insoluble solids can be removed through filtration or centrifugation, and the spargnum extract can be obtained by concentrating the solution by removing the solvent under reduced pressure, or by performing freeze-drying, spray-drying, or reduced-pressure drying as necessary. The spargnum extract can be applied to a cosmetic composition in the form of a liquid, concentrate, or powder.

[0037] Example 1: Spagnum extract (10)

[0038] In this embodiment, the spagnum extract (10) was prepared in the following manner.

[0039] First, the above-mentioned spagnum extract was obtained using *Gaksimulflaekki*.

[0040] As a raw material preparation step (A1), the entire sphagnum plant body was collected, external foreign substances were removed, and it was naturally dried. The moisture content of the dried sphagnum plant body was about 8.5% by weight, and it was crushed to produce a crushed material with an average particle size of 2 to 5 mm.

[0041] In the extraction step (A2), 500 g of the ground material was used as the extraction raw material. 5,000 mL of extraction solvent was prepared by mixing purified water and ethanol in a volume ratio of 70:30, and then added to the ground sphagnum plant material. At this time, the ratio of solvent to raw material was 1:10 (w / v). The mixture was stirred and extracted for 6 hours at a stirring speed of 200 rpm under conditions maintained at 50°C. After the extraction was completed, the extract was naturally cooled to room temperature.

[0042] In the solid removal step (A3), the cooled extract was first filtered using a 100 μm mesh filter, and then centrifuged at 6,000 rpm, 20 minutes, and 10°C to remove insoluble solids. A clear yellowish-brown supernatant was recovered from this.

[0043] In the vacuum concentration step (A4), the recovered supernatant was vacuum concentrated using a rotary evaporator. The vacuum conditions were set to 150 mbar and the water bath temperature to 50°C, and the volume of the extract after concentration was adjusted to approximately 1,000 mL. As a result, a viscous concentrated extract was obtained.

[0044] In the drying step (A5), the concentrated extract was placed in a freeze dryer and dried. The freezing conditions were set to -45℃, the drying pressure to less than 0.05 mbar, and the drying time to 48 hours. As a result, a light brown powder form of sphagnum extract (10) was obtained.

[0045] In the result of the acquisition and physical property analysis step (A6), the amount of the finally obtained sphagnum extract (10) was 78.5 g, which corresponded to a yield of 15.7 wt% based on the dried sphagnum plant. As a result of analyzing the basic physical properties of the obtained sphagnum extract (10), the moisture content was 3.2 wt% and the pH was 5.6 based on a 1 wt% aqueous solution. In addition, as a result of analysis using the phenol-sulfuric acid method, the polysaccharide content was confirmed to be 41.3 wt%.

[0047] The present invention describes a fermented liquid or extract (20) obtained by fermenting microorganisms of the genus Psychrobacter spp.

[0048] The fermented liquid (20a) obtained by fermenting the above-mentioned Psychrobacter genus microorganisms is a fermented culture liquid obtained by inoculating Psychrobacter genus microorganisms into a liquid medium containing a carbon source and a nitrogen source, and then culturing and fermenting them under low-temperature conditions. Psychrobacter genus microorganisms are microorganisms adapted to low-temperature environments and have the characteristic of producing relatively stable metabolic products even under conditions where moisture is present.

[0049] Specifically, the liquid medium may contain one or more of glucose, fructose, glycerol, or maltodextrin as a carbon source, and may contain yeast extract or peptone as a nitrogen source. After the microorganisms of the genus *Phycorobacter* are inoculated into this liquid medium, they are cultured and fermented for a certain period of time in a temperature range of 4°C to 20°C. At this time, fermentation may be carried out under aerobic or microaerotic conditions.

[0050] After the fermentation is finished, the culture medium can be recovered as is to obtain a fermented liquid (20a) of the genus *Psychobacter*, and the fermented liquid contains water-soluble metabolites generated during the fermentation process. These metabolites are components derived from microorganisms adapted to low temperature or oxidative stress environments, and have the characteristic of maintaining relatively stable physical properties even in humid environments.

[0051] Meanwhile, a fermentation culture extract (20b) derived from microorganisms of the genus *Psychobacter* may be obtained by recovering the supernatant from which the cells have been removed by filtration or centrifugation, and then extracting it using purified water, ethanol, or a mixture of these solvents. The fermentation culture extract obtained in this way may contain effective metabolites generated during the fermentation process in a concentrated form.

[0052] The fermented liquid derived from a genus of microorganisms or the extract (20) according to the present invention maintains a relatively stable state even in an oxidation reaction environment induced by metal ions under conditions where moisture is present, and when combined with spagnum extract and ellagic acid, it performs the function of further improving the antioxidant stability of the cosmetic composition.

[0054] Example 2: Fermented culture extract derived from microorganisms of the genus *Psychobacter* (20b)

[0055] In this embodiment, a fermented culture extract (20b) derived from Psychrobacter spp. microorganisms used in the cosmetic composition according to the present invention was prepared in the following way.

[0056] First, a liquid medium was prepared for culturing microorganisms of the genus Psychrobacter spp. The liquid medium was prepared by dissolving 10.0 g of glucose, 5.0 g of yeast extract, 5.0 g of peptone, and 5.0 g of sodium chloride in 1,000 mL of purified water, and the prepared medium was sterilized by autoclaving at 121°C for 15 minutes.

[0057] After inoculating 10 mL of a genus of Psychorobacter microorganism starter into 1,000 mL of sterile liquid medium, culture and fermentation were carried out for 72 hours under aerobic conditions at a fermentation temperature of 10°C and a stirring speed of 150 rpm. After the fermentation was completed, 1,000 mL of fermented culture medium containing cells and metabolites was obtained.

[0058] The above fermentation culture solution was centrifuged at 8,000 rpm and 10°C for 20 minutes to remove the cells, and 950 mL of the supernatant was recovered after centrifugation.

[0059] Ethanol was added to 950 mL of the recovered supernatant to adjust the final ethanol concentration to 50 wt%, and then stirring extraction was performed for 4 hours at a stirring speed of 300 rpm under conditions of 25℃.

[0060] The extracted mixture was filtered using a 0.45 μm membrane filter, and the filtrate was concentrated under reduced pressure using a rotary evaporator under conditions of a pressure of 120 mbar and a water bath temperature of 45℃. The volume of the extract after reduced pressure concentration was 200 mL.

[0061] 200 mL of the above concentrate was placed in a freeze-dryer and dried for 48 hours under freezing temperature -50°C and drying pressure 0.03 mbar. As a result, a fermented culture extract (20b) derived from a microorganism of the genus *Psychorobacter* in the form of a pale yellow powder was obtained.

[0062] The amount of the final fermented culture extract (20b) obtained was 12.4 g, which corresponds to a yield of 1.24 wt% based on 1,000 mL of fermented culture. As a result of analyzing the physical properties of the obtained fermented culture extract (20b), the moisture content was 4.1 wt%, and the pH was measured to be 6.2 based on a 1 wt% aqueous solution. In addition, the total solid content was confirmed to be 95.9 wt%.

[0064] The ellagic acid of the present invention will be described.

[0065] The above ellagic acid (30) is a polyphenol compound having a polyvalent phenol structure and performs the function of inhibiting metal ion-mediated oxidation reactions by forming a complex with metal ions in the cosmetic composition. Since ellagic acid contains a number of phenolic hydroxyl groups in its molecular structure, it performs the function of reducing the activity of free metal ions that promote oxidation reactions by binding with Fe(II) ions or Fe(III) ions.

[0066] The ellagic acid (30) in the present invention may be included in a dissolved or finely dispersed state in the aqueous component of the cosmetic composition, and a uniform dispersion state may be maintained by using a dissolving aid or a dispersing agent together as needed. By this form of inclusion, the ellagic acid acts stably even under conditions where moisture is present, thereby inhibiting the initiation of a metal ion-mediated oxidation reaction.

[0067] In particular, when ellagic acid (30) is combined with a sphagnum extract (10) and a fermented liquid or extract thereof derived from a sphagnum genus microorganism (20, 20b), it contributes to effectively maintaining antioxidant stability even under oxidative stress conditions that may occur in a humid environment. That is, by interacting with the humid matrix environment formed by the sphagnum extract and the stabilizing effect of the fermented components, the metal ion chelation function of ellagic acid can be expressed more efficiently.

[0068] Accordingly, the ellagic acid (30) included in the cosmetic composition according to the present invention functions as a key component that structurally blocks metal ion-mediated oxidation reactions, going beyond the role of a simple antioxidant component, and plays an important role in achieving the technical objective of the present invention, which ensures antioxidant stability even in a humid environment.

[0070] Example 3: Ellagic acid (30)

[0071] In this embodiment, ellagic acid (30) used in the cosmetic composition according to the present invention was prepared in the following way.

[0072] First, ellagic acid powder with a purity of 98.0 wt% was prepared as a raw material for ellagic acid. 10.0 g of the ellagic acid powder was weighed using a precision balance.

[0073] Next, a solvent composition for dissolution and dispersion was prepared. The solvent composition was prepared by mixing 800 mL of purified water, 200 mL of ethanol, and 50 mL of propylene glycol to a total volume of 1,050 mL.

[0074] The above-described solvent composition was introduced into a stirrer under conditions maintained at 40°C, and 10.0 g of metered ellagic acid powder was slowly added while the stirring speed was set to 400 rpm. After the addition of the ellagic acid powder was completed, the mixture was continuously stirred for 60 minutes under the same conditions to uniformly disperse the ellagic acid.

[0075] After stirring was finished, insoluble particles were removed by filtering the mixture using a 0.45 μm membrane filter. The total volume of the ellagic acid preparation obtained after filtration was 1,000 mL.

[0076] The ellagic acid preparation prepared as described above was in the form of a uniform dispersion with an ellagic acid content of 1.0 weight%, and was used as ellagic acid (30) in the preparation of the cosmetic composition according to the present invention and in subsequent test examples.

[0077] The results of measuring the basic properties of the above ellagic acid (30) are as follows.

[0078] - Ellagic acid content: 1.0 wt%

[0079] - pH: 5.8

[0080] - Appearance: Pale yellow transparent dispersion

[0081] - Precipitation (25℃, left for 24 hours): No precipitation.

[0083] A method for preparing a cosmetic composition for improving antioxidant stability containing a natural extract of the present invention will be described.

[0084] The above manufacturing method includes a step of preparing a spagnum extract (M1), a step of preparing a fermented liquid or extract derived from a genus of microorganisms of the genus Cyclobacter (M2), a step of preparing ellagic acid (M3), and a mixing step (M4).

[0085] First, the step of preparing the sphagnum extract (M1) is a step of preparing the sphagnum extract (10) using the entire sphagnum plant body as a raw material. The sphagnum extract can be obtained by collecting the sphagnum plant body, removing foreign substances, extracting it at a temperature of 20°C to 60°C using purified water, ethanol, or a mixed solvent thereof, removing solids from the extract by filtration or centrifugation, and then concentrating it by removing the solvent under reduced pressure, or by performing freeze-drying, spray-drying, or reduced-pressure drying as necessary.

[0086] Next, the step of preparing a fermented liquid derived from a Psychrobacter genus microorganism or its extract (M2) is a step of preparing a fermented liquid derived from a Psychrobacter genus microorganism or its extract (20, 20b) using a Psychrobacter genus microorganism (Psychrobacter spp.) as a raw material. The fermented liquid derived from a Psychrobacter genus microorganism can be obtained by inoculating a Psychrobacter genus microorganism into a liquid medium containing a carbon source and a nitrogen source, and then culturing and fermenting it under temperature conditions of 4°C to 20°C, and recovering the fermented culture liquid obtained as is. Alternatively, the supernatant from which the cells have been removed from the fermented culture liquid through filtration or centrifugation can be recovered, and then extracted using purified water, ethanol, or a mixed solvent thereof, and, if necessary, subjected to reduced pressure concentration and freeze-drying, spray-drying, or reduced pressure drying to obtain a fermented culture liquid extract (20b) derived from a Psychrobacter genus microorganism.

[0087] Next, the ellagic acid preparation step (M3) is a step of preparing ellagic acid (30) in a form applicable to a cosmetic composition. The ellagic acid is a polyphenol compound having a polyvalent phenol structure, which is a component capable of inhibiting metal ion-mediated oxidation reactions by forming a complex with metal ions, and can be prepared in a state dissolved in or finely dispersed in the aqueous component of the cosmetic composition. To this end, the ellagic acid is prepared in the form of an ellagic acid preparation solution by dissolving or dispersing it in a solvent composition containing purified water, ethanol, and a polyvalent alcohol, and can be adjusted to a uniform state by including stirring, heating, and filtration processes as needed.

[0088] Finally, the mixing step (M4) is a step of preparing a cosmetic composition by mixing the spargnum extract (10), a fermented liquid derived from a genus of microorganisms or its extract (20, 20b), and ellagic acid (30). In the mixing step, the spargnum extract (10) is first added to the aqueous component and uniformly dispersed, and then the fermented liquid derived from a genus of microorganisms or its extract (20, 20b) is sequentially added and stirred to form a mixture. Subsequently, ellagic acid (30) is added in a dissolved or dispersed state so that the entire composition is uniformly mixed. The mixing can be performed under stirring conditions according to a conventional cosmetic manufacturing process, and the stirring speed and stirring time can be adjusted so that each component is stably dispersed without phase separation or precipitation. If necessary, a heating or cooling process may be included during the mixing process. The cosmetic composition obtained through the mixing step (M4) in this manner has the characteristic of ensuring antioxidant stability by suppressing oxidation reactions mediated by metal ions even in a humid environment where moisture is present, and can be applied to various skincare formulations such as skin, toner, lotion, essence, serum, cream, or mask pack.

[0090] Example 4: Method for preparing a cosmetic composition

[0092] In this embodiment, a cosmetic composition for improving antioxidant stability containing a natural extract according to the present invention was prepared using the spargnum extract (10) prepared in Example 1, the fermented culture extract (20b) derived from a genus of microorganisms prepared in Example 2, and the ellagic acid (30) prepared in Example 3.

[0093] The above manufacturing method consists of a basic formulation preparation step of the composition (S1), a step of adding spagnum extract (S2), a step of adding fermented culture extract (S3), a step of adding ellagic acid preparation solution (S4), a final composition adjustment step (S5), and a step of checking the state of the final composition (S6).

[0094] First, in the basic formulation preparation step (S1) of the composition, 880.0 g of purified water, 50.0 g of glycerin, and 30.0 g of butylene glycol were mixed as aqueous components of the cosmetic composition, and then a homogeneous aqueous mixture was prepared by stirring at 300 rpm for 10 minutes at 25°C.

[0095] In the step of adding the spagnum extract (S2), 20.0 g of the spagnum extract (10) prepared in Example 1 was added to the aqueous mixture. After addition, the mixture was stirred for 20 minutes at 25°C and 300 rpm to ensure that the spagnum extract was uniformly dispersed.

[0096] Next, in the step of adding the fermented culture extract (S3), 10.0 g of the fermented culture extract (20b) derived from the genus *Psychobacter* microorganism prepared in Example 2 was added to the mixture. After addition, the mixture was stirred for 15 minutes at 25°C and 300 rpm to ensure that the fermented culture extract was uniformly mixed.

[0097] Next, in the step of adding the ellagic acid solution (S4), 10.0 g of the ellagic acid solution (30) prepared in Example 3 (ellagic acid content 1.0 wt%) was added to the mixture. This was set so that the ellagic acid content based on the final composition would be 0.10 wt%. After addition, the mixture was stirred for 20 minutes under the same conditions to ensure that the ellagic acid was uniformly dispersed.

[0098] In the final composition adjustment step (S5), after all ingredients were mixed, the total weight of the final composition was adjusted to 1,000 g. The pH of the composition was measured and found to be 5.7, and no separate pH adjustment process was performed.

[0099] Looking at the state verification step (S6) of the final composition, the appearance of the final composition was a pale yellow translucent liquid, and when left at 25°C for 24 hours, there was no precipitation and no phase separation was observed.

[0100] The cosmetic composition prepared in this example was used as a test group composition in subsequent tests, including an antioxidant stability evaluation test under wet conditions, an inhibition test of active oxygen generation in the presence of Fe(II) ions or Fe(III) ions, and a stability comparison test within the composition.

[0102] Test Example 1: Evaluation of Antioxidant Stability under Wet Conditions

[0103] In this test example, in order to evaluate the antioxidant stability of the cosmetic composition according to the present invention under wet conditions, the cosmetic composition prepared in Example 4 was used as a test group to measure the residual rate of the antioxidant component.

[0104] The cosmetic composition prepared in Example 4 was used as a test group, and the following comparative group was prepared for comparison. Comparative Example 1 is a cosmetic composition having the same composition as Example 4 but not containing ellagic acid (30), Comparative Example 2 is a cosmetic composition having the same composition as Example 4 but not containing a fermented culture extract (20b) derived from a microorganism of the genus *Psychobacter*, and Comparative Example 3 is a cosmetic composition having the same composition as Example 4 but not containing sphagnum extract (10).

[0106] 20.0 g of purified water was added to each sample and mixed uniformly to adjust the moisture content of the sample to 20% by weight. The wet-treated samples were placed in a sealed container and stored for 48 hours in a constant temperature and humidity chamber under conditions of 37°C and 90% relative humidity.

[0107] 1.0 g was taken from each sample before and after treatment under wet conditions and used for analysis. 10.0 mL of ethanol was added to each sample, and stirring extraction was performed for 30 minutes at a stirring speed of 300 rpm. After stirring extraction was completed, insoluble components were removed by filtering the extract using a 0.45 μm membrane filter.

[0108] The residual amount of antioxidant components in the filtered extract was analyzed using high-performance liquid chromatography (HPLC). Ellagic acid was used as the analytical indicator for antioxidant components. HPLC analysis was performed using equipment equipped with a UV detector, and the detection wavelength was set to 254 nm. Separation was carried out using a C18 column (4.6 × 250 mm), and a solvent mixture of water and methanol in a volume ratio of 40:60 was used as the mobile phase. The flow rate of the mobile phase was set to 1.0 mL / min, and the sample injection volume was 20 μL.

[0109] The ellagic acid content prior to treatment under wet conditions was set as the reference value and set to 100%, and the residual antioxidant component of each sample was calculated by converting the ellagic acid content measured after treatment under wet conditions into a percentage relative to the reference value.

[0110] The test results are as follows.

[0111] division Ellagic acid retention rate (%) Test group (Example 4) 89.6 Comparative Example 1 (excluding ellagic acid) Unmeasurable Comparative Example 2 (excluding fermented culture extract) 61.2 Comparative Example 3 (excluding spagnum extract) 58.7

[0112] As can be seen from the results above, the cosmetic composition according to Example 4 maintained an ellagic acid retention rate of 89.6% even under wet conditions, exhibiting significantly superior antioxidant stability compared to the comparative examples. In particular, in Comparative Examples 2 and 3, which lacked either the fermented culture extract derived from the genus *Psychobacter* or the *Spargnum* extract, the ellagic acid retention rates decreased to 61.2% and 58.7%, respectively, confirming that the stability of the antioxidant component was significantly reduced under wet conditions. Accordingly, it was confirmed that the cosmetic composition according to the present invention effectively ensures antioxidant stability even in a wet environment through the combination of *Spargnum* extract, the fermented culture extract derived from the genus *Psychobacter*, and ellagic acid.

[0114] Test Example 2: Test of Reactive Oxygen Species (ROS) Generation Inhibition Under Conditions of Metal Ions

[0115] In this test example, to evaluate how effectively the cosmetic composition according to the present invention inhibits the generation of reactive oxygen species (ROS) under conditions in the presence of metal ions, the amount of reactive oxygen species generated was measured under oxidation-inducing conditions containing Fe(II) ions and Fe(III) ions.

[0116] The cosmetic composition prepared in Example 4 was used as a test group, and the following comparative group was prepared for comparison. Comparative Example 1 is a cosmetic composition having the same composition as Example 4 but not containing ellagic acid (30), Comparative Example 2 is a cosmetic composition having the same composition as Example 4 but not containing a fermented culture extract (20b) derived from a microorganism of the genus *Psychobacter*, and Comparative Example 3 is a cosmetic composition having the same composition as Example 4 but not containing sphagnum extract (10).

[0118] To induce a metal ion-mediated oxidation reaction for each sample, conditions containing Fe(II) ions and Fe(III) ions were established, respectively. Iron(II) sulfate (FeSO₄·7H₂O) was used as the source for Fe(II) ions, and iron(III) chloride (FeCl₃) was used as the source for Fe(III) ions. After adjusting the final concentration of Fe(II) ions or Fe(III) ions for each sample to 100 μM, the samples with added metal ions were left at 25°C for 30 minutes to induce an oxidation reaction by the metal ions.

[0119] The amount of reactive oxygen species generated in each sample after metal ion treatment was measured using the DCFH-DA (2′,7′-Dichlorodihydrofluorescein diacetate) fluorescence assay. Specifically, 0.5 g of each sample was taken and diluted in 4.5 mL of purified water, after which DCFH-DA solution was added to adjust the final DCFH-DA concentration to 10 μM. The mixture was reacted at 37°C for 30 minutes, after which the fluorescence intensity was measured using a fluorescence microplate reader. Fluorescence measurements were performed under conditions of an excitation wavelength of 485 nm and an emission wavelength of 535 nm, and the measured fluorescence intensity values ​​were used as an indicator proportional to the amount of reactive oxygen species generated. For data processing, a condition in which metal ions were added but the test sample was not included was designated as the control group, and the fluorescence intensity measured under this condition was defined as 100% of the reactive oxygen species generation. The fluorescence intensity measured in each test group and comparison group was converted into a percentage relative to the control group to calculate the reactive oxygen species generation inhibition rate.

[0120] division ROS generation amount (%) control group 100.0 Test group (Example 4) 38.5 Comparative Example 1 (excluding ellagic acid) 85.1 Comparative Example 2 (excluding fermented culture extract) 70.2 Comparative Example 3 (excluding spagnum extract) 74.6

[0121] As can be seen from the above results, the cosmetic composition of Example 4 according to the present invention showed active oxygen generation amounts of 34.2% and 38.5%, respectively, under conditions where Fe(II) ions and Fe(III) ions were present, confirming that the metal ion-mediated oxidation reaction is effectively suppressed.

[0122] In contrast, in the comparative example that did not contain any of ellagic acid, fermented culture extract derived from the genus *Psychobacter*, or *Spargnum* extract, the inhibitory effect on the generation of reactive oxygen species was significantly reduced. This implies that the cosmetic composition of the present invention structurally blocks metal ion-mediated oxidation reactions through the combination of the three components.

[0124] Test Example 3: Evaluation of Formulation Stability under Wetting and Metal Ion Presence Conditions

[0125] In this experimental example, to evaluate whether the cosmetic composition according to the present invention maintains formulation stability even in a humid environment and under conditions where metal ions are present, phase separation, precipitation, and changes in appearance of the composition were observed under conditions of humid treatment and addition of metal ions.

[0126] The cosmetic composition prepared in Example 4 was used as a test group, and the following comparative group was prepared for comparison. Comparative Example 1 is a cosmetic composition having the same composition as Example 4 but not containing ellagic acid (30), Comparative Example 2 is a cosmetic composition having the same composition as Example 4 but not containing a fermented culture extract (20b) derived from a microorganism of the genus *Psychobacter*, and Comparative Example 3 is a cosmetic composition having the same composition as Example 4 but not containing sphagnum extract (10).

[0127] 50.0 g of each test group and control group sample was taken and placed in a transparent glass container. Then, 10.0 g of purified water was added to each sample to adjust the moisture content in the composition to 20% by weight, and then the mixture was uniformly mixed.

[0128] Next, to introduce conditions for the presence of metal ions, an aqueous solution of iron(III) chloride (FeCl₃) was added to each sample to adjust the final Fe(III) ion concentration to 100 μM. The samples with added metal ions were sealed and stored for 7 days in a constant temperature and humidity chamber at 40°C and 90% relative humidity. After the storage period, the presence of phase separation, precipitation, and changes in appearance were evaluated for each sample through visual inspection and optical microscopy (magnification 200x). In addition, changes in viscosity and color were observed by comparing the samples before and after storage, respectively, while left at 25°C.

[0129] As a result, no phase separation or precipitation was observed in the cosmetic composition of Example 4 during the storage period, and it maintained a uniform appearance. In addition, no change in viscosity or color was observed.

[0130] On the other hand, in the case of Comparative Example 1, fine sediment was observed at the bottom of the container after 7 days of storage, and in the case of Comparative Examples 2 and 3, non-uniformity of color concentration was observed along with partial phase separation.

[0131] Accordingly, it was confirmed that the cosmetic composition according to the present invention maintains excellent formulation stability even in a humid environment and under conditions where metal ions are present, which means that formulation stability is structurally secured by the combination of sphagnum extract, fermented culture extract derived from microorganisms of the genus *Psychobacter*, and ellagic acid. Explanation of the symbols

[0134] Spagnum extract (10), Fermented liquid or extract (20) obtained by fermenting microorganisms of the genus Psycholobacter, Ellagic acid (30).

Claims

Claim 1 Step (S1) of preparing an aqueous mixture by mixing 880.0 g of purified water, 50.0 g of glycerin, and 30.0 g of butylene glycol, and stirring at 300 rpm for 10 minutes at 25°C; collecting the entire sphagnum plant body, removing external foreign substances, drying until the moisture content reaches 8.5 wt%, grinding it to an average particle size of 2 to 5 mm, adding the ground sphagnum plant body to 5,000 mL of an extraction solvent mixed with purified water and ethanol in a volume ratio of 70:30 to prepare a mixture with a solvent-to-raw material ratio of 1:10 (w / v), stirring the mixture for 6 hours at a stirring speed of 200 rpm under conditions maintained at 50°C, cooling the extract, primary filtration of the cooled extract using a 100 μm mesh filter, and centrifugation at 6,000 rpm for 20 minutes at 10°C. Step (S2): removing insoluble solids to recover the supernatant, concentrating the supernatant under reduced pressure conditions of 150 mbar and a water bath temperature of 50℃ using a rotary evaporator so that the volume of the extract after concentration becomes 1,000 mL to obtain a concentrated extract, and drying the concentrated extract by setting the temperature to -45℃, the drying pressure to less than 0.05 mbar, and the drying time to 48 hours to obtain 20.0 g of a Spagnum extract, which is added to the aqueous mixture, and stirring for 20 minutes under conditions of 25℃ and 300 rpm; 10.0 g of glucose, 5.0 g of yeast extract, 5.0 g of peptone, and 5.0 g of sodium chloride.A liquid medium was prepared by dissolving 0 g in 1,000 mL of purified water, and the prepared liquid medium was autoclaved at 121°C for 15 minutes. Then, 10 mL of a microbial inoculum of the genus *Psychobacter* was inoculated into 1,000 mL of the sterilized liquid medium, and culture and fermentation were performed for 72 hours under aerobic conditions at a fermentation temperature of 10°C and a stirring speed of 150 rpm. After the fermentation was completed, 1,000 mL of a fermented culture medium containing cells and metabolites was obtained. The fermented culture medium was centrifuged at 8,000 rpm and 10°C for 20 minutes to remove the cells, and 950 mL of the supernatant was recovered. Ethanol was added to the recovered 950 mL of supernatant to adjust the final ethanol concentration to 50 wt%, and then stirring extraction was performed for 4 hours at 25°C and a stirring speed of 300 rpm. A step (S3) in which 10.0 g of a powdered extract of a fermented culture derived from the genus *Psychobacter*, obtained by filtering the mixture using a 0.45 μm membrane filter to obtain a filtrate by filtering the mixture using a rotary evaporator under reduced pressure conditions of 120 mbar and a water bath temperature of 45℃ to obtain a concentrate with a volume of 200 mL after reduced pressure concentration, and introducing 200 mL of the concentrate into a freeze-dryer and drying for 48 hours under freezing temperature conditions of -50℃ and drying pressure of 0.03 mbar, is added to the mixture obtained through step (S2), and the mixture is stirred for 15 minutes under conditions of 25℃ and 300 rpm; a step (S3) in which a solvent composition prepared by mixing 800 mL of purified water, 200 mL of ethanol, and 50 mL of propylene glycol to obtain a total volume of 1,050 mL is introduced into a stirrer under conditions maintained at 40℃ and the stirring speed is set to 400 rpm Ellagic acid dispersion 10, from which insoluble particles have been removed by adding 10.0 g of ellagic acid powder with a purity of 98.0 wt% and continuously stirring the mixture under the same conditions for 60 minutes, and filtering the mixture using a 0.45 μm membrane filter.A method for preparing a cosmetic composition for improving antioxidant stability containing a natural extract, comprising the step of adding 0 g to the mixture obtained through the above step (S3) and stirring for 20 minutes. Claim 2 A cosmetic composition for improving antioxidant stability comprising a natural extract, manufactured by the method of claim 1, characterized in that metal ion-mediated oxidative decomposition of ellagic acid is inhibited under conditions of a humid environment and the presence of metal ions. Claim 3 delete Claim 4 delete Claim 5 delete Claim 6 delete Claim 7 delete

Images