Cosmetic composition and food composition having an Anti-glycation effect comprising morus alba leaf extract and cornus officinalis fruit extract and manufacturing therefor

Abstract

The present invention relates to a composition having an anti-glycation effect, and more specifically, to a composition having an anti-glycation effect that includes Cornus fruit extract and mulberry leaf extract as active ingredients to inhibit the glycation of proteins, thereby reducing the glycation reaction and / or the formation of glycation end products. The composition may be provided as a cosmetic composition and a food composition, and a method for manufacturing the same is also provided.

Description

Technology Field

[0001] The present invention relates to a composition having an anti-glycation effect, and more specifically, to a composition having an anti-glycation effect that includes Cornus fruit extract and mulberry leaf extract as active ingredients to inhibit the glycation of proteins, thereby reducing the glycation reaction and / or the formation of glycation end products. The composition may be provided as a cosmetic composition and a food composition, and a method for manufacturing the same is also provided. Background Technology

[0002] In general, human skin ages due to various factors such as ultraviolet rays, free radicals, environmental pollutants, and lifestyle habits, and structural denaturation of dermal proteins, such as collagen and elastin, plays a significant role in this skin aging. In particular, glycation, which occurs when reducing sugars such as glucose non-enzymatically bind to the amino groups of proteins, and the resulting Advanced Glycation End Products (AGEs) are known to accelerate signs of aging—such as wrinkles, reduced elasticity, and dull skin tone—by causing collagen cross-linking, protein stiffness, yellowing, and decreased elasticity. Therefore, anti-glycation technology, which inhibits the glycation reaction of skin proteins or reduces the accumulation of already formed AGEs, is attracting attention as an effective approach to prevent and mitigate skin aging.

[0003] Furthermore, as glycation reactions and the generation and accumulation of AGEs are reported to be associated with changes in biological proteins, including skin, the potential for anti-glycation approaches is being discussed not only for topical compositions but also for ingestible compositions.

[0004] In the past, compositions using low-molecular-weight compounds such as aminoguanidine and carnosine, or specific peptides, as anti-glycation agents have been proposed to control glycation and aging phenomena of the skin. For example, Chinese published patent CN10396127A discloses a technology for providing an anti-glycation cosmetic composition for the skin by combining an anti-glycation agent and an antioxidant. The composition is intended to alleviate signs of skin aging, such as wrinkles and reduced elasticity, by inhibiting the progression of glycation reactions and removing free radicals.

[0005] Meanwhile, while there may be attempts to provide active ingredients with anti-glycation properties for ingestion, there is a need to develop food compositions based on applicable natural materials, considering the safety of raw materials, consumer acceptability, and the stability of the composition.

[0006] However, synthetic compound-based anti-glycation ingredients may have limitations in terms of safety, skin irritation, and stability within formulations during long-term use, and have the limitation of not fully reflecting consumers' natural preferences.

[0007] In addition, even when provided as a food composition, there may be limitations on the application of synthetic compound-based ingredients in terms of the naturalness of the raw materials, water solubility for long-term consumption, and manufacturing and storage stability.

[0008] To address these issues, cosmetic compositions using naturally derived extracts with excellent anti-glycation and anti-aging activities have been proposed. For example, Korean Registered Patent No. 10-1339915 discloses a technology in which a biotransformed carob extract is prepared by enzymatically treating a carob (Ceratonia siliqua) extract, and this is used as an active ingredient in an anti-glycation and anti-aging cosmetic composition.

[0009] The aforementioned registered Korean patent states that enzyme-treated carob extract exhibits not only anti-glycation activity but also antioxidant, collagenase expression inhibition, and anti-inflammatory activities, making it useful as an anti-aging cosmetic composition.

[0010] However, the technology described in the aforementioned Korean Registered Patent No. 10-1339915 focuses technically on carob beans and their enzyme-treated extracts, and is based on a raw material composition heavily concentrated on a specific plant (carob beans). Furthermore, while it is useful in that it provides a combination of anti-glycation activity, antioxidant, collagenase expression inhibition, and anti-inflammatory activities, it does not present a composition that combines medicinal plants known for their metabolism-related efficacy, such as *Morus alba* leaf, or *Cornus officinalis*, which has been traditionally widely used for health and vitality, with the unique physiological activities of these plants focused on inhibiting the glycation of skin proteins.

[0011] Furthermore, technical configurations for designing and formulating compositions containing mulberry leaves and Cornus fruit as active ingredients are not sufficiently presented to expand their applicability beyond cosmetic compositions for topical application to include food compositions for consumption.

[0012] Furthermore, there is currently insufficient disclosure of technology regarding natural cosmetic compositions designed to include mulberry leaves and Cornus fruit as active ingredients, and to provide a balanced combination of antioxidant, anti-inflammatory, and collagenase inhibitory effects, with a focus on anti-glycation effects.

[0013] Furthermore, there is a current situation where technology regarding food compositions capable of providing anti-glycation effects based on the same or similar combination of active ingredients is not sufficiently disclosed.

[0014] Therefore, there is a need for a new composition that complements the safety and consumer acceptance limitations of synthetic anti-glycation agents, effectively inhibits the glycation reaction of proteins based on naturally derived materials, and further provides complex effects such as antioxidant, anti-inflammatory, and inhibition of collagenase activity.

[0015] In particular, it is necessary to develop a composition configured to effectively utilize the biological activities of mulberry leaf extract and Cornus fruit extract, respectively, from the perspective of anti-glycation, and the composition needs to be designed to be provided as a cosmetic composition for skin application and a food composition for consumption. Prior art literature

[0016] Chinese Published Patent Application CN103961276A (Published Aug. 6, 2014) "Anti-glycation cosmetic composition and anti-glycation cosmetic containing the same" Korean Registered Patent Publication No. 10-1339915 (Registered Dec. 4, 2013) "Method for preparing carob bean extract having excellent anti-glycation and anti-aging activity and anti-aging cosmetic composition containing the extract as an active ingredient" Korean Registered Patent Publication No. 10-0940133 (Registered Jan. 26, 2010) "Composition for treating diabetes containing *Citrus aurantium* extract and health supplement having the same as an active ingredient" The problem to be solved

[0017] In accordance with the above requirements, the present invention aims to provide an anti-glycation composition that effectively inhibits protein glycation by utilizing Cornus officinalis extract and mulberry leaf extract while avoiding safety and irritation issues of synthetic anti-glycation agents, and thereby improves signs of aging related to glycation.

[0018] More specifically, the present invention aims to provide a composition and a method for manufacturing the same, which can improve signs of skin aging caused by glycation, such as skin wrinkles and loss of elasticity, by providing the anti-glycation composition as a cosmetic composition for topical application, and at the same time, can also provide the anti-glycation composition as a food composition for consumption. means of solving the problem

[0019] To achieve the above objectives, the present invention provides a composition having an anti-glycation effect comprising a Cornus fruit extract and a mulberry leaf extract, wherein the Cornus fruit extract is prepared by crushing the Cornus fruit, roasting it at 70 to 90°C for 1 to 3 hours, and the mulberry leaf extract is prepared by crushing the mulberry leaf, roasting it at 80 to 100°C for 10 to 30 minutes, and then extracting it.

[0020] In addition, the Cornus fruit extract is prepared by ripening the Cornus fruit at 40 to 50°C for 3 to 4 days, then crushing and roasting it, and then extracting it, and the mulberry leaf extract is prepared by ripening the mulberry leaf at 40 to 50°C for 1 to 2 days, then crushing and roasting it, and then extracting it.

[0021] In addition, the extraction of the Cornus fruit extract is a process of extracting the Cornus fruit, which has been ripened, crushed, and roasted, in purified water or an aqueous ethanol solution for 2 to 3 hours under ultrasonic conditions of 350 to 450 watts, 35 to 45 kHz, and a temperature of 35 to 45°C, and the extraction of the mulberry leaf extract is a process of extracting the mulberry leaf, which has been ripened, crushed, and roasted, in purified water or an aqueous ethanol solution for 1 to 2 hours under ultrasonic conditions of 350 to 450 watts, 35 to 45 kHz, and a temperature of 35 to 45°C.

[0022] In addition, the Cornus fruit extract is prepared by concentrating the extract to 55 to 65 weight% relative to the weight of the Cornus fruit raw material after extraction to produce a Cornus fruit extract concentrate, and then freeze-drying the Cornus fruit extract concentrate to 35 to 45 weight% relative to the weight of the Cornus fruit raw material, and the mulberry leaf extract is prepared by concentrating the extract to 18 to 25 weight% relative to the weight of the mulberry leaf raw material after extraction to produce a mulberry leaf extract concentrate, and then freeze-drying the mulberry leaf extract concentrate to 13 to 17 weight% relative to the weight of the mulberry leaf raw material.

[0023] In addition, the composition of the present invention is characterized in that it can be provided as a cosmetic composition and / or a food composition.

[0024] At this time, the composition is characterized by providing an anti-glycation effect, and when the composition is a cosmetic composition, providing an antioxidant effect, an anti-inflammatory effect and an effect that inhibits collagenase activity, and when the composition is a food composition, providing an α-amylase and α-glucosidase inhibitory effect.

[0025] Furthermore, a method for manufacturing a composition according to the present invention is provided. Effects of the invention

[0026] According to the present invention, by inhibiting the glycation reaction of proteins through a composition containing Cornus fruit extract and mulberry leaf extract as active ingredients, it is possible to contribute to reducing changes associated with glycation and the generation and accumulation of Advanced Glycation End Products (AGEs).

[0027] Specifically, when the above composition is provided as a cosmetic composition, it provides the effect of inhibiting the glycation reaction of skin proteins and improving or alleviating glycation-related skin conditions such as wrinkles, loss of elasticity, and deterioration of skin tone associated therewith.

[0028] In addition, even when the above composition is provided as a food composition, it can provide a function from an anti-glycation perspective through the combination of Cornus fruit extract and mulberry leaf extract. Brief explanation of the drawing

[0029] Figure 1 is a graph showing the results of a cytotoxicity confirmation experiment. Figure 2 is a graph showing the results of the antioxidant activity confirmation experiment. Figure 3 is a graph showing the results of an experiment confirming the anti-inflammatory effect. Figure 4 is a graph showing the experimental results of the elastase inhibition effect. Figure 5 is a graph showing the experimental results of the collagenase inhibition effect. Figure 6 is a graph showing the results of the NBT assay experiment to confirm antiglycation activity. Figure 7 is a graph showing the experimental results of AGEs formation to confirm antiglycation activity. Figure 8 is a graph showing the experimental results of the α-amylase inhibitory effect. Figure 9 is a graph showing the experimental results of the α-glucosidase inhibitory effect. Specific details for implementing the invention

[0030] The following detailed descriptions relating to the present invention refer to the accompanying drawings, which are embodiments in which the present invention may be practiced and are illustrated as examples of such embodiments. These embodiments are described in detail to sufficiently enable those skilled in the art to practice the present invention. It should be understood that various embodiments of the present invention are different but need not be mutually exclusive. For example, specific shapes, structures, and characteristics described herein may be implemented in other embodiments without departing from the spirit and scope of the present invention in relation to one embodiment. Furthermore, it should be understood that the location or arrangement of individual components within each described embodiment may be changed without departing from the spirit and scope of the present invention.

[0031] Accordingly, the following detailed description is not intended to be taken in a limiting sense, and the scope of the invention is limited only by the appended claims, including all equivalents to those claimed therein, provided they are properly described. Similar reference numerals in the drawings refer to the same or similar functions across various aspects.

[0032] The terms used in this invention have been selected based on currently widely used general terms, taking into account their functions within the invention; however, these terms may vary depending on the intent of those skilled in the art, case law, the emergence of new technologies, etc. Additionally, in specific cases, terms have been arbitrarily selected by the applicant, and in such cases, their meanings will be described in detail in the relevant description of the invention. Therefore, the terms used in this invention must be defined not merely by their names, but based on their meanings and the overall content of the invention.

[0033] In the present invention, when a part is described as "comprising" a certain component, this means that, unless specifically stated otherwise, it does not exclude other components but may include additional components.

[0035] A composition having an anti-glycation effect comprising Cornus fruit extract and mulberry leaf extract according to the present invention comprises Cornus fruit extract and mulberry leaf extract. Such a composition of the present invention may be provided as a cosmetic composition and / or a food composition.

[0036] That is, the composition of the present invention may be formulated as a cosmetic composition applied to the skin, depending on the purpose of application and form of use, or formulated into an ingestible form and provided as a food composition.

[0037] The above Cornus officinalis is the mature fruit of the Cornus officinalis, which belongs to the Cornaceae family, and contains iridoid glycosides such as morroniside, loganin, and cornidoid glycosides, as well as gallotannins, organic acids, etc.

[0038] These components of Cornus fruit can contribute to alleviating structural denaturation of biological proteins caused by protein glycation and the accompanying deterioration of their condition by regulating glycation and inflammatory responses through antioxidant and anti-inflammatory activities, as well as α-glucosidase inhibition and inhibition of AGEs formation. In particular, when the composition of the present invention is provided as a cosmetic composition, it can contribute to alleviating collagen structural damage caused by glycation and the resulting deterioration of skin conditions, such as wrinkles and reduced elasticity.

[0039] The above-mentioned mulberry leaves are leaves of the mulberry tree (Morus alba) and contain polyphenols such as quercetin, kaempferol, and chlorogenic acid, as well as imino sugar-based components such as 1-deoxynojirimycin (DNJ).

[0040] These components of mulberry leaves can contribute to the expression of the anti-glycation effect of the composition by inhibiting the generation of reactive oxygen species and mitigating protein glycation reactions by regulating pathways related to sugar metabolism and AGEs generation. In particular, when the composition of the present invention is provided as a cosmetic composition, it can contribute to improving skin conditions such as reduced skin elasticity and deterioration of skin tone caused by glycation. Furthermore, even when the composition of the present invention is provided as a food composition, the combination of Cornus fruit extract and mulberry leaf extract can contribute to inhibiting protein glycation reactions.

[0042] The above Cornus fruit extract can be prepared by crushing the Cornus fruit, roasting it at 70 to 90°C for 1 to 3 hours, and then extracting it.

[0043] Specifically, the Cornus fruit extract is obtained by grinding Cornus fruit to 5 to 10 mesh and then roasting it at 70 to 90°C for 1 to 3 hours, and preferably roasting it for 1 hour is appropriate in terms of the balance between the activation of the included components and the prevention of destruction.

[0044] In addition, the extraction may be performed by mixing the Cornus fruit with a solvent in a weight ratio of 1:15 to 25 using purified water or an aqueous ethanol solution as a solvent, and then extracting for 2 to 3 hours at a temperature of 35 to 45°C, and it is preferable to mix the Cornus fruit with a solvent in a weight ratio of 1:20 and then extract at 40°C for 3 hours in order to simultaneously ensure the elution efficiency and stability of the active ingredient.

[0045] At this time, the Cornus fruit extract may be prepared by ripening the Cornus fruit at 40 to 50°C for 3 to 4 days, then grinding and roasting it, and then extracting it.

[0046] Through this, compared to the case where post-ripening is not performed, the distribution of moisture and low molecular weight components within the Cornus fruit is homogenized and the content of specific iridoid and polyphenol components is stabilized, thereby enabling the effect of enhancing the anti-glycation and antioxidant activities of the final Cornus fruit extract.

[0047] Furthermore, the extraction of the Cornus fruit extract may be a process of extracting the Cornus fruit, which has been ripened, ground, and roasted, in purified water or an aqueous ethanol solution for 2 to 3 hours under ultrasonic conditions of 350 to 450 watts, 35 to 45 kHz, and a temperature of 35 to 45°C.

[0048] Specifically, ultrasonic conditions of 350 to 450 W and 35 to 45 kHz are energy conditions within a range that mechanically disturbs the cell walls of Cornus officinalis to shorten the diffusion pathway of active ingredients, without causing degradation or oxidation of the ingredients due to excessive energy. In addition, temperature conditions of 35 to 45°C are a temperature range that increases the extraction rate while minimizing the denaturation of heat-sensitive active ingredients to maintain the structure and activity of anti-glycation related active ingredients.

[0049] Meanwhile, for the extraction of Cornus officinalis extract, it is preferable to extract the Cornus officinalis, which has been ripened, crushed, and roasted, in purified water or an aqueous solution of ethanol for 3 hours under ultrasonic conditions of 400 watts and 40 kHz and a temperature of 40°C.

[0050] At this time, the Cornus fruit extract may be prepared by concentrating the extract to 55 to 65 weight% relative to the weight of the Cornus fruit raw material after the extraction, thereby producing a Cornus fruit extract concentrate, and then freeze-drying the Cornus fruit extract concentrate to 35 to 45 weight% relative to the weight of the Cornus fruit raw material.

[0051] In addition, the weight of the raw Cornus fruit refers to the weight after crushing and roasting.

[0052] Here, the Cornus officinalis extract concentrate is concentrated to 55 to 65 weight percent relative to the weight of the raw Cornus officinalis, because if the concentration is less than 55 weight percent, the content of the active ingredient is relatively low, so sufficient anti-glycation activity is not secured at the same amount and the amount added to the formulation tends to increase excessively; on the other hand, if the concentration exceeds 65 weight percent, the viscosity and solid content ratio increase excessively, which reduces the fluidity of the concentrate and may cause problems such as poor redispersibility of the powdered Cornus officinalis extract due to uneven drying during the subsequent freeze-drying process. Therefore, the range of 55 to 65 weight percent is a critical concentration condition that can balance the concentration of the anti-glycation active ingredient with the process handling and powder redispersibility of the concentrate.

[0053] In addition, freeze-drying the Cornus officinalis extract concentrate to a weight of 35 to 45% relative to the weight of the Cornus officinalis raw material is because if the freeze-drying yield is less than 35% by weight, the loss of active ingredients increases, which reduces the contribution to anti-glycation activity at the same input amount and lowers economic efficiency in terms of process yield; and if the yield exceeds 45% by weight, the proportion of residual moisture and low molecular weight components increases, making the powder prone to aggregation and caking, which makes uniform dispersion difficult during formulation and tends to reduce stability during long-term storage. Therefore, the range of 35 to 45% by weight is a critical freeze-drying yield range to simultaneously satisfy the yield of anti-glycation active ingredients, the physical stability of the powder, and suitability for formulation.

[0054] For example, when producing a Cornus fruit extract from 1 kg of Cornus fruit, the Cornus fruit extract concentrate may be 550 to 650 g at which 60 brix is ​​achieved, and the Cornus fruit extract completed by freeze-drying may be 350 to 450 g.

[0055] In other words, the Cornus fruit extract of the present invention is preferably prepared by ripening the Cornus fruit at 40 to 50°C for 3 to 4 days, then grinding it to 5 to 10 mesh, roasting it at 70 to 90°C for 1 hour, mixing it with purified water or an aqueous ethanol solution in a weight ratio of 1:20, extracting it under ultrasonic conditions of 400 watts and 40 kHz at 40°C for 3 hours, concentrating it to 55 to 65 weight% relative to the weight of the raw Cornus fruit to produce a Cornus fruit extract concentrate, and freeze-drying the Cornus fruit extract concentrate to 35 to 45 weight% relative to the weight of the raw Cornus fruit.

[0057] The above mulberry leaf extract can be prepared by crushing the mulberry leaves, roasting them at 80 to 100°C for 10 to 30 minutes, and then extracting.

[0058] Specifically, the mulberry leaf extract is obtained by grinding the mulberry leaves to a mesh of 5 to 10 and then roasting them at 80 to 100°C for 10 to 30 minutes, and preferably roasting them for 20 minutes rapidly removes moisture from the mulberry leaf tissue while minimizing thermal denaturation of polyphenol and flavonoid components, thereby providing a proper balance in terms of the releaseability and stability of the active ingredients.

[0059] In addition, the extraction of the mulberry leaf extract may be performed by mixing the mulberry leaves and the solvent in a weight ratio of 1:35 to 45 using purified water or an aqueous ethanol solution as a solvent, and then extracting at a temperature of 35 to 45°C for 2 to 3 hours, and it is preferable to mix the mulberry leaves with the solvent in a weight ratio of 1:40 and then extract at 40°C for 3 hours in order to simultaneously secure the extraction efficiency and structural stability of flavonoid and iminosugar series components.

[0060] At this time, the mulberry leaf extract may be prepared by ripening the mulberry leaves at 40 to 50°C for 1 to 2 days, then grinding and roasting them, and then extracting.

[0061] Through this, compared to the case without the ripening process, the distribution of moisture and low-molecular-weight components within the mulberry leaf tissue becomes uniform, and some precursor components are converted into a stable form, thereby increasing and activating the content of active ingredients that contribute to anti-glycation and antioxidant activity, resulting in an enhanced anti-glycation activity of the mulberry leaf extract.

[0062] Furthermore, the extraction of the mulberry leaf extract may be a process of extracting the mulberry leaves, which have been ripened, ground, and roasted, with purified water or an aqueous ethanol solution for 1 to 2 hours under ultrasonic conditions of 350 to 450 Watt, 35 to 45 kHz, and a temperature of 35 to 45°C.

[0063] Specifically, ultrasonic conditions of 350 to 450 W and 35 to 45 kHz effectively destroy the cell walls of the upper leaves to promote the diffusion of the active ingredient, while preventing the active ingredient from being decomposed due to the generation of free radicals or local overheating caused by excessive ultrasonic energy.

[0064] In addition, the temperature condition of 35 to 45°C is a temperature range for securing the extraction rate while maintaining the thermal stability of the active ingredients derived from mulberry leaves, and since it is classified as a low-temperature extraction, the extraction efficiency drops significantly when the temperature is lower than this, and when the temperature is higher, there is a risk that heat-sensitive anti-glycation related ingredients will decompose.

[0065] Meanwhile, considering the balance between the amount of active ingredient released, extraction time, and ingredient stability, it is preferable to extract the mulberry leaf extract by extracting the mulberry leaves, which have been ripened, crushed, and roasted, in purified water or an aqueous solution of ethanol for 2 hours under ultrasonic conditions of 400W and 40kHz and a temperature of 40℃.

[0066] At this time, the mulberry leaf extract may be prepared by concentrating the extract to 18 to 25 weight percent relative to the weight of the raw mulberry leaf after extraction to produce a mulberry leaf extract concentrate, and then freeze-drying the mulberry leaf extract concentrate to 13 to 17 weight percent relative to the weight of the raw mulberry leaf.

[0067] In addition, the weight of the raw mulberry leaves refers to the weight after crushing and roasting.

[0068] Here, when the mulberry leaf extract concentrate is concentrated to less than 18% by weight relative to the weight of the raw mulberry leaf, the content of the active ingredient per unit weight is low and the contribution to anti-glycation activity is insufficient, whereas when it is concentrated to more than 25% by weight, the solid content ratio increases excessively, leading to increased viscosity and a tendency to gel, which can result in reduced handling of the concentrate and uneven moisture movement within the sample during freeze-drying, causing the formation of a powder with uneven particle size and density.

[0069] Therefore, the range of 18 to 25 weight percent is a critical concentration condition to simultaneously secure the concentration of anti-glycation active ingredients derived from mulberry leaves, process handling properties of the concentrate, and uniform drying properties.

[0070] For example, when preparing a mulberry leaf extract from 1 kg of mulberry leaves, the mulberry leaf extract concentrate may be 180 to 250 g at which 60 brix is ​​achieved, and the finished mulberry leaf extract may be 130 to 170 g after freeze-drying.

[0071] In other words, the mulberry leaf extract of the present invention is preferably obtained by ripening mulberry leaves at 40 to 50°C for 1 to 2 days, then grinding them to 5 to 10 mesh, roasting them at 80 to 100°C for about 20 minutes, mixing them with purified water or an aqueous ethanol solution in a weight ratio of 1:40, extracting them under ultrasonic conditions of 400 watts and 40 kHz at 40°C for 2 hours, concentrating the extract to 18 to 25 weight% relative to the weight of the raw mulberry leaves to produce a mulberry leaf extract concentrate, and freeze-drying the mulberry leaf extract concentrate to 13 to 17 weight% relative to the weight of the raw mulberry leaves.

[0073] Meanwhile, the composition of the present invention may be a mixture of the Cornus officinalis extract and the mulberry leaf extract in a weight ratio of 1 to 3:1 to 3, and preferably, the Cornus officinalis extract and the mulberry leaf extract are mixed in equal amounts of 1 to 1, as polyphenols and deoxynojirimycin contained in the mulberry leaf contribute to alleviating sugar metabolism and protein glycation reactions, and iridoid glycosides and gallotannins contained in the Cornus officinalis contribute to alleviating the glycation process and the accompanying oxidation and inflammatory reactions, as well as collagen structural damage, thereby ensuring that the different functional characteristics of the two extracts are reflected together without being biased toward one side, which is appropriate within the above weight ratio range.

[0074] At this time, as described above, the composition of the present invention may be provided as a cosmetic composition and / or a food composition depending on the purpose of application and form of use.

[0075] Specifically, when the composition of the present invention is provided as a cosmetic composition, it may be formulated into known cosmetic formulations such as toner, skin, lotion, cream, foundation, essence, gel, pack, emulsified sunscreen cream, emulsified foundation, emulsified makeup base, oil cake foundation, two-way cake, or powder pact.

[0076] These formulations may include ingredients commonly used in addition to the cosmetic composition of the present invention, and may include, for example, conventional auxiliary agents such as antioxidants, stabilizers, solubilizers, vitamins, pigments, colorants, and fragrances, and carriers.

[0077] For example, in the case of a paste, cream, or gel containing the cosmetic composition of the present invention, one or more of animal oil, vegetable oil, wax, paraffin, starch, tracanth, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc, or zinc oxide may be selected and used as carrier components.

[0078] In addition, when the composition of the present invention is provided as a food composition, it may be provided in an ingestible formulation such as powder, granules, tablets, capsules, pills, beverages, concentrates, syrups, or jellies.

[0079] At this time, the composition of the present invention may additionally include auxiliary components and / or carriers commonly used in the food industry to ensure palatability, dispersibility, fluidity, formulation stability, and storage stability.

[0080] For example, the above auxiliary components and / or carriers may be appropriately selected according to the formulation of the composition and may include excipients (e.g., starch, dextrin, maltodextrin, etc.), fluidizing agents or anti-caking agents in the case of powder or granule formulations, binders, disintegrants and lubricants in the case of tablet or capsule formulations, and solubilizers, emulsifiers, thickeners and stabilizers in the case of beverage or concentrate formulations.

[0081] Additionally, additives for improving palatability and stability, such as sweeteners, flavorings, acidifiers, pH adjusters, antioxidants, and preservatives, may be further included as needed. In this case, the type and content of the auxiliary component and / or carrier may be selected within a range that does not substantially impede the anti-glycation effect of the active ingredient of the present invention.

[0083] Below, we examine a method for preparing a composition having an anti-glycation effect comprising Cornus fruit extract and mulberry leaf extract according to the present invention.

[0084] At this time, the explanation has been omitted or brief to avoid repetitive descriptions, but it is clear that the manufacturing method of the present invention shares the same technical concept and features as the composition of the present invention described above. For example, the setting of preferred numerical values ​​described above can be applied in the same way to the manufacturing method described later, and vice versa.

[0086] The manufacturing method of the present invention includes a Cornus fruit preparation step (S10), a mulberry leaf preparation step (S20), a Cornus fruit extract manufacturing step (S30), a mulberry leaf extract manufacturing step (S40), and a completion step (S50).

[0088] The Cornus fruit preparation step (S10) is a step of preparing a Cornus fruit extract raw material by crushing the Cornus fruit and then roasting it at 70 to 90°C for 1 to 3 hours, and the mulberry leaf preparation step (S20) is a step of preparing a mulberry leaf extract raw material by crushing the mulberry leaf and then roasting it at 80 to 100°C for 10 to 30 minutes.

[0089] At this time, the Cornus fruit preparation step (S10) is a step of preparing a Cornus fruit extract raw material by ripening the Cornus fruit at 40 to 50°C for 3 to 4 days, crushing it, and then roasting it at 70 to 90°C for 1 to 3 hours, and the mulberry leaf preparation step (S20) may be a step of preparing a mulberry leaf extract raw material by ripening the mulberry leaf at 40 to 50°C for 1 to 2 days, crushing it, and then roasting it at 80 to 100°C for 10 to 30 minutes.

[0090] In other words, the raw material for Cornus fruit and mulberry leaf extract refers to an intermediate substance for extraction obtained by ripening, crushing, and roasting the raw materials of Cornus fruit and mulberry leaves.

[0092] The step of manufacturing Cornus officinalis extract (S30) is a step of manufacturing Cornus officinalis extract by extracting the above Cornus officinalis extract raw material, and the step of manufacturing mulberry leaf extract (S40) is a step of manufacturing mulberry leaf extract by extracting the above mulberry leaf extract raw material.

[0093] At this time, the step of preparing the Cornus fruit extract (S30) is a step of preparing the Cornus fruit extract by extracting the Cornus fruit extract raw material in purified water or an aqueous ethanol solution for 2 to 3 hours under ultrasonic conditions of 350 to 450 watts, 35 to 45 kHz, and a temperature condition of 35 to 45°C, and the step of preparing the mulberry leaf extract (S40) is a step of preparing the mulberry leaf extract by extracting the mulberry leaf extract raw material in purified water or an aqueous ethanol solution for 1 to 2 hours under ultrasonic conditions of 350 to 450 watts, 35 to 45 kHz, and a temperature condition of 35 to 45°C.

[0094] The subsequent finishing step (S50) is a step of preparing the composition of the present invention by mixing the Cornus fruit extract and the mulberry leaf extract. The composition prepared in the finishing step (S50) may be provided as a cosmetic composition or as a food composition depending on the purpose of application and form of use.

[0096] According to one embodiment of the present invention, the step of preparing Cornus officinalis extract powder (S31) and the step of preparing mulberry leaf extract powder (S41) may be included.

[0097] The step of manufacturing Cornus officinalis extract powder (S31) is to prepare a Cornus officinalis extract concentrate by concentrating the Cornus officinalis extract to a weight of 55 to 65% relative to the weight of the Cornus officinalis extract raw material, and then freeze-drying the Cornus officinalis extract concentrate to a weight of 35 to 45% relative to the weight of the Cornus officinalis extract raw material to produce Cornus officinalis extract powder.

[0098] The step of manufacturing mulberry leaf extract powder (S41) is to concentrate the mulberry leaf extract to 18 to 25 weight% relative to the weight of the mulberry leaf extract raw material to produce a mulberry leaf extract concentrate, and then freeze-dry the mulberry leaf extract concentrate to 13 to 17 weight% relative to the weight of the mulberry leaf extract raw material to produce a mulberry leaf extract powder.

[0099] In one embodiment including the step of preparing Cornus fruit extract powder (S31) and the step of preparing mulberry leaf extract powder (S41), the completion step (S50) may be a step of completing the composition of the present invention by mixing the Cornus fruit extract powder and the mulberry leaf extract powder.

[0101] The effects of the composition according to the present invention will be examined in detail below through examples and experimental examples.

[0103] According to Table 1 below, Cornus fruit extract and mulberry leaf extract were prepared for the preparation of the example.

[0104] At this time, the extraction solvent was purified water, the concentration was performed using the Rotavapor R-300 rotary vacuum concentrator from BUCHI, and the freeze-drying was performed using the FD 8508 laboratory freeze dryer from IlShin BioBase.

[0105] The Cornus fruit extracts in Table 1, 3, and 5 below were prepared by grinding 1 kg of Cornus fruit to 10 mesh, roasting it on a hot plate set to 80°C for 1 hour, mixing it with an extraction solvent in a weight ratio of 1:20, and extracting it at 40°C for 3 hours. In the case of 3 and 5, the extracts were aged for 3 days in a drying room set to 43°C before grinding, and in the case of 5, 400 watt, 40 kHz ultrasound was applied during extraction.

[0106] In addition, for the mulberry leaf extracts of 2, 4, and 6, 1 kg of mulberry leaves were ground to 10 mesh, roasted on a hot plate set to 90°C for 20 minutes, mixed with an extraction solvent in a weight ratio of 1:40, and extracted at 40°C for 2 hours; for 4 and 6, the leaves were ripened for 1 day in a drying room set to 43°C before grinding, and for 6, 400 watt, 40 kHz ultrasound was applied during extraction.

[0107] Raw Material Classification Ultrasonic treatment ripening Concentrate (g) Brix 60 Concentration rate (%) Freeze-dried (g) Drying yield (%) 1 Cornelian cherry - - 627 62.7 435 43.5 2 Sangyeop - - 200 20.0 151 15.1 3 Post-ripening Cornus fruit - o 596 59.6 408 40.8 4 After ripening leaves - o 188 18.8 142 14.2 5 Post-ripening milk (ultrasound) o o 615 61.5 416 41.6 6 Post-ripening leaves (ultrasound) o o 198 19.8 147 14.7

[0108] Here, the concentration yield (%) is calculated by dividing the weight of the concentrate obtained when 1 kg of raw Cornus fruit or mulberry leaves is concentrated to Brix 60 by the weight of the raw material and multiplying the result by 100.

[0109] Next, experimental cosmetic compositions of Examples 1 to 5 were prepared according to Table 2 below.

[0110] Cornelian cherry extract Mulberry leaf extract Mixed weight ratio Example 1 After ripening cherry (3) Sangyeop (2) 1 : 1 Example 2 Cornelian cherry (1) After ripening leaves (4) 1 : 1 Example 3 After ripening cherry (3) After ripening leaves (4) 1 : 1 Example 4 Post-ripening breast milk (ultrasound, 5) Post-ripening leaves (ultrasound, 6) 1 : 1 Example 5 Cornelian cherry (1) Sangyeop (2) 1 : 1

[0111] Examples 1 to 5 in powder form prepared in this way were mixed with purified water at various concentrations or applied as powder in order to be used as samples for the experimental examples to be described later.

[0113] The following experimental examples are intended to evaluate the safety and efficacy of the composition of the present invention and were performed separately according to the purpose of application.

[0114] Specifically, Experimental Example 1 is a common experimental example for confirming cytotoxicity, which is the basic safety of the composition of the present invention.

[0115] Experimental Examples 2 to 7 are experimental examples to confirm effects such as antioxidant, anti-glycation, anti-inflammatory, and improvement of skin-related indicators when the composition of the present invention is provided as a cosmetic composition, and present the results of an efficacy evaluation based on external application to the skin.

[0116] Experimental Examples 8 and 9 are experimental examples to confirm the effects when the composition of the present invention is provided as a food composition, and present the results of an efficacy evaluation based on the premise of ingestion application.

[0118] Experimental Example 1. Confirmation of Cytotoxicity

[0119] 1) Experimental Method

[0120] To confirm the cytotoxicity of the composition containing Cornus fruit extract and mulberry leaf extract according to the present invention, an MTT assay was performed using Raw264.7 cells, a mouse-derived macrophage cell line.

[0121] Raw264.7 cells were cultured and maintained in Dulbecco Modified Eagle Medium (DMEM) containing 10 percent heat-inactivated fetal bovine serum and 1 percent penicillin streptomycin. Cells in culture were harvested and transferred to 24-well plates at a ratio of 1 × 10⁶ per well. 5 After dispensing cells to form a number of wells, they were cultured for 24 hours at 37°C under 5% carbon dioxide conditions. Subsequently, samples according to the embodiments of the present invention were added to each well at different concentrations and cultured again for 24 hours.

[0122] After 24 hours of culture, the cell supernatant was removed, and MTT reagent adjusted to a final concentration of 1 milligram per milliliter was added to each well, followed by an additional 2 hours of incubation in the dark. Subsequently, the MTT reagent was removed, and dimethyl sulfoxide (DMSO) was added to each well to completely dissolve the formed formazan crystals.

[0123] The solution dissolved in formazan was transferred to a 96-well plate at a rate of 100 µl per well, and the cell viability was calculated by measuring the absorbance at a wavelength of 540 nm.

[0124] The above cytotoxicity evaluation was conducted independently three times under the same conditions, and the cell viability for each condition was expressed as the average value of the measurements obtained from three repeated experiments.

[0126] 2) Experimental Results

[0127] The experimental results are shown in Table 3 below, and the unit of cell viability is percentage.

[0128] Concentration (µg / ml) Example 1 Example 2 Example 3 Example 4 Example 5 0 100.0 100.0 100.0 100.0 100.0 10 101.5 101.5 102.6 102.9 102.4 25 102.7 105.2 102.9 103.9 103.8 50 103.7 106.0 104.6 105.1 104.4 100 107.1 108.3 106.6 108.6 109.8 250 110.4 111.7 109.6 111.2 110.5 500 113.8 114.5 112.2 115.1 113.7 1,000 118.1 117.1 115.2 117.6 123.1

[0129] As shown in Table 3, all of Examples 1 to 5 did not show significant cytotoxicity, so it can be seen that they can be applied as external skin agents or ingested as food.

[0131] Experimental Example 2. DPPH assay

[0132] 1) Experimental Method

[0133] DPPH radical scavenging activity analysis was performed to confirm the antioxidant activity of a composition containing Cornus fruit extract and mulberry leaf extract according to the present invention.

[0134] First, the powder sample for each example was dissolved in purified water to prepare a final concentration of 10% (weight / volume) solution, and then filtered using a 0.22 μm syringe filter. Each filtered sample solution was then diluted stepwise with purified water to prepare test samples at different concentrations.

[0135] 100 µl of samples diluted to different concentrations were dispensed into each well of a 96-well plate, and then an equal amount (100 µl) of DPPH solution prepared at a concentration of 0.2 mM was added to each well. Subsequently, the reaction was carried out at room temperature for 30 minutes in a dark environment with light blocked.

[0136] After the reaction was completed, DPPH radical scavenging activity was evaluated by measuring the absorbance at a wavelength of 517 nm for each well. Radical scavenging activity at each concentration was calculated as a % using the rate of decrease in absorbance compared to the control group.

[0137] The above DPPH radical scavenging activity evaluation was performed independently three times under the same conditions, and the measurement value for each condition was expressed as the average value of the values ​​obtained from the three repeated experiments, and 100 μg / mL ascorbic acid was used as the control group.

[0139] 2) Experimental Results

[0140] The experimental results are shown in Table 4 below, and the units are percentages.

[0141] Concentration (µg / ml) control group Example 1 Example 2 Example 3 Example 4 Example 5 control group 88.9 10 16.1 17.2 19.8 21.5 15.5 50 35.1 39.4 40.7 42.7 32.3 100 60.3 63.6 65.5 67.6 56.1 250 70.5 78.8 79.0 80.0 69.9 500 76.5 81.7 83.3 84.6 73.1

[0142] As shown in Table 4, in the evaluation of DPPH radical scavenging activity with ascorbic acid as the control, Examples 1 to 5 of the present invention showed a tendency for DPPH radical scavenging activity to increase in a concentration-dependent manner across all concentration ranges.

[0143] When looking at the differences between the examples, Example 4, a combination in which both ripening and ultrasonic treatment were applied, showed the highest DPPH radical scavenging activity across the entire concentration range, followed by Example 3, in which only ripening was applied, and Examples 1 and 2, in which only one of Cornus fruit or mulberry leaf was ripened, showing the next best activity, while Example 5, in which neither ripening nor ultrasonic treatment was applied, showed a relatively low value.

[0144] In addition, at the same concentration, radical scavenging activity was greater in the combination where both raw materials were ripened as in Example 3 compared to the combination where only Cornus fruit or mulberry leaf was ripened as in Examples 1 and 2. Furthermore, the antioxidant activity was best when ultrasonic extraction was applied after ripening as in Example 4. This result supports the fact that the process of combining ripening and ultrasonic treatment on the Cornus fruit extract and mulberry leaf extract of the present invention is effective in efficiently extracting antioxidant-related active ingredients such as polyphenols, iridoid glycosides, and imino sugar series components, thereby enhancing free radical scavenging ability.

[0145] Therefore, from the results of Experimental Example 2, it can be confirmed that the cosmetic composition containing Cornus fruit extract and mulberry leaf extract according to the present invention exhibits concentration-dependent and excellent DPPH radical scavenging activity, and in particular, the combination of ripening and ultrasonic treatment of the raw materials is the most advantageous in terms of antioxidant activity.

[0147] Experimental Example 3. NO assay

[0148] 1) Experimental Method

[0149] To confirm the anti-inflammatory effect of the composition containing Cornus fruit extract and mulberry leaf extract according to the present invention, an evaluation of NO production inhibition was performed using Raw264.7 cells, a mouse-derived macrophage cell line.

[0150] Raw264.7 cells were cultured and maintained in DMEM medium containing 10% heat-inactivated fetal bovine serum (HI-FBS) and 1% penicillin-streptomycin (P / S). Cells in culture were harvested and transferred to a 24-well plate at a rate of 1 × 10⁶ per well. 5 After dispensing as many cells as possible, they were cultured for 24 hours at 37℃ and 5% CO2 conditions.

[0151] Subsequently, samples according to the embodiments of the present invention were treated at different concentrations in each well, and all treatment groups except the untreated group were treated with lipopolysaccharide (LPS) at a concentration of 1 μg / ml simultaneously with the sample treatment and cultured for an additional 24 hours.

[0152] After incubation, 100 µl of the cell supernatant from each well was taken and dispensed into a 96-well plate, and an equal amount of Griess reagent was added to each well. After reacting at room temperature for 30 minutes, the absorbance was measured at a wavelength of 540 nm. The measured absorbance values ​​were substituted into a pre-prepared NaNO2 standard curve to convert them into the amount of NO produced (nitrate concentration) for each sample treatment group.

[0153] The above evaluation of NO production was conducted independently three times under the same conditions, and the NO production amount for each condition was expressed as the average value of the measurements obtained from the three repeated experiments. The control groups below are the sample-untreated groups, consisting of the LPS-untreated group and the LPS-only treated group.

[0155] 2) Experimental Results

[0156] The experimental results are shown in Table 5 below, and the units are percentages.

[0157] Concentration (µg / ml) LPS control group Example 1 Example 2 Example 3 Example 4 Example 5 - - 3.0 - - - - - - o 50.5 - - - - - 10 o 49.7 50.1 49.3 46.5 49.9 50 o 48.3 48.0 46.7 43.5 49.2 100 o 47.1 46.6 44.7 38.2 47.7 250 o 43.7 42.4 40.4 34.9 44.4 500 o 39.2 38.8 35.2 31.4 41.3

[0158] As can be seen from Table 5, the amount of NO produced in the control group that was not treated with LPS was about 3.0%, whereas the amount of NO produced in the control group treated only with LPS increased significantly to about 50.5%, confirming that NO, an inflammatory mediator, is significantly induced by LPS stimulation.

[0159] In contrast, Examples 1 to 5 showed a tendency for NO production to decrease compared to the LPS-alone treatment group in all concentration ranges when treated with LPS, and exhibited a concentration-dependent inhibition pattern in which NO production gradually decreased as the sample concentration increased.

[0160] In addition, as a result of comparing the examples, Example 4, which applied both ripening and ultrasonic treatment, showed the lowest NO production amount across the entire concentration range, indicating the best NO production inhibition effect; Example 3, which applied only ripening to both raw materials, showed the next strongest inhibition effect; and Example 5, a combination without ripening or ultrasonic treatment, showed a relatively weak inhibition effect. This suggests that applying ripening and ultrasonic extraction processes to Cornus fruit and mulberry leaf raw materials increases the release of active ingredients and the expression of activity that can more effectively inhibit inflammatory responses related to NO production.

[0161] Accordingly, through Experimental Example 3, it can be confirmed that the cosmetic composition containing Cornus fruit extract and mulberry leaf extract according to the present invention exhibits anti-inflammatory activity that reduces NO production induced by LPS in a concentration-dependent manner, and in particular, the extraction process combined with ripening and ultrasonic treatment is advantageous for maximizing the anti-inflammatory effect.

[0163] Experimental Example 4. Elastase inhibition assay

[0164] 1) Experimental Method

[0165] To confirm whether the composition containing Cornus fruit extract and mulberry leaf extract according to the present invention has an elastase inhibitory effect, elastase inhibitory activity was measured using the Neutrophil Elastase Inhibitor Screening Kit (Fluorometric) (BioVision, Catalog No. K782-100).

[0166] First, the powder sample for each example was dissolved in purified water to prepare a final concentration of 10% solution, then filtered through a 0.22 μm syringe filter, and then appropriately diluted with purified water to match the usable concentration range of the kit and used as an experimental sample.

[0167] The evaluation of elastase inhibitory activity was performed according to the method described in the instructions for use of the kit. Specifically, the neutrophil-derived elastase enzyme, fluorescent substrate, buffer solution, and samples of each concentration provided in the kit were dispensed into a reaction plate according to the kit instructions to form a reaction system. Then, as soon as the final dispensing was completed, the primary fluorescence intensity was measured using a fluorescent plate reader (Varioskan™ ALF Multimode Microplate Reader, ThermoFisher, USA) under conditions of an excitation wavelength of 345 nm and a fluorescence wavelength of 450 nm.

[0168] Subsequently, the reaction plate was reacted at 37°C for 30 minutes in a light-blocked state, and after the reaction was completed, the secondary fluorescence intensity was measured under the same measurement conditions (Ex 345 nm, Em 450 nm). For each sample, the value obtained by subtracting the primary fluorescence intensity R1 from the secondary fluorescence intensity R2 was defined as ΔR, and the elastase inhibitory activity was calculated according to Equation 1 below using the ΔR values ​​of the sample treatment group and the blank group.

[0169]

[0170] Here, the blank group is a reaction system containing only the elastase enzyme, fluorescent substrate, and reaction buffer solution, without adding the sample of the present invention or other inhibitors, and was used as a standard for indicating the maximum activity of the elastase.

[0171] The above evaluation of elastase inhibitory activity was performed independently three times under the same conditions, and the elastase inhibitory activity value for each condition was expressed as the average value of the measurements obtained from the three repeated experiments.

[0173] 2) Experimental Results

[0174] The experimental results are shown in Table 6 below, and the units are percentages.

[0175] Concentration (µg / ml) Example 1 Example 2 Example 3 Example 4 Example 5 10 7.9 7.8 12.3 13.4 1.3 50 16.1 17.2 21.5 26.9 5.5 100 20.6 21.9 33.9 37.8 12.2 250 26.0 29.0 45.6 50.2 16.3 500 36.3 38.4 55.2 60.7 20.5

[0176] As can be seen from Table 6, Examples 1 to 5 of the present invention exhibit a concentration-dependent trend in which the elastase inhibitory activity (%) gradually increases with increasing concentration across all concentration ranges from 10 to 500 μg / ml. That is, Example 1 starts at 7.9% at 10 μg / ml and increases to 36.3% at 500 μg / ml, and Example 2 also increases from 7.8% at 10 μg / ml to 38.4% at 500 μg / ml, confirming a trend in which the inhibitory activity increases stepwise with increasing concentration.

[0177] In particular, as a result of comparison between examples, Example 4, which applied both ripening and ultrasonic treatment, showed the highest elastase inhibitory activity across the entire concentration range.

[0178] For example, Example 4 exhibited the highest elastase inhibitory activity across the entire concentration range. For instance, Example 4 showed inhibitory activities of 37.8% at 100 µg / ml, 50.2% at 250 µg / ml, and 60.7% at 500 µg / ml, demonstrating superior values ​​compared to other examples at the same concentration. Additionally, Example 3 showed the next highest inhibitory activity, exhibiting an increasing trend of 33.9% at 100 µg / ml, 45.6% at 250 µg / ml, and 55.2% at 500 µg / ml.

[0179] On the other hand, Example 5 showed 1.3% at 10 μg / ml and 20.5% at 500 μg / ml, indicating overall lower inhibitory activity compared to other examples, so it is determined that differences in raw material pretreatment and extraction processes affect elastase inhibition efficiency.

[0180] Therefore, from the results of Experimental Example 4, it can be confirmed that the composition of the present invention has the effect of inhibiting elastase activity in a concentration-dependent manner, and in particular, the elastase inhibitory activity is most excellent in the combination of raw material processing and extraction conditions applied as in Example 4. Since this elastase inhibitory activity can contribute to alleviating the deterioration of skin conditions, such as reduced skin elasticity and wrinkle formation, by inhibiting the action of elastase involved in the degradation of elastic fibers, this supports the fact that the composition of the present invention can be usefully applied to maintaining skin elasticity and alleviating damage.

[0182] Experimental Example 5. Collagenase inhibition assay

[0183] 1) Experimental Method

[0184] In order to confirm whether a composition comprising Cornus fruit extract and mulberry leaf extract according to the present invention has a collagenase inhibitory effect, EnzChe TMCollagenase inhibitory activity was measured using the Gelatinase / Collagenase Assay Kit (Invitrogen, USA, Catalog No. E12055).

[0185] First, the powder sample for each example was dissolved in purified water to prepare a final concentration of 10% solution, then filtered through a 0.22 μm syringe filter, and then diluted with purified water to match the usable concentration range of the kit to be used as an experimental sample.

[0186] The evaluation of collagenase inhibitory activity was performed according to the method described in the instruction manual of the above kit. Specifically, the reaction system was constructed according to the instructions using the collagenase enzyme, fluorescent substrate, and reaction buffer solution provided in the kit, along with samples of each concentration; the reaction was carried out for 30 minutes starting from the time the last dispensing was completed; and after the reaction was finished, the fluorescent plate reader (Varioskan TM Fluorescence intensity was measured using an ALF Multimode Microplate Reader (ThermoFisher, USA) under conditions of an excitation wavelength of 485 nm and a fluorescence wavelength of 525 nm.

[0187] Collagenase inhibitory activity for each sample was calculated according to the following Equation 2 using the fluorescence values ​​of the blank group and the sample-treated group.

[0188]

[0189] Here, the blank group is a reaction system containing only collagenase enzyme, fluorescent substrate, and reaction buffer solution, without adding the sample of the present invention or other inhibitors, and was used as a standard for showing the maximum activity of collagenase.

[0190] The above evaluation of collagenase inhibitory activity was performed independently three times under the same conditions, and the inhibitory activity value for each condition was expressed as the average value of the measurements obtained from the three repeated experiments.

[0192] 2) Experimental Results

[0193] The experimental results are shown in Table 7 below, and the units are percentages.

[0194] Concentration (µg / ml) Example 1 Example 2 Example 3 Example 4 Example 5 10 4.3 5.5 10.1 13.0 3.3 50 10.1 12.4 17.7 19.3 10.3 100 21.6 23.3 30.7 32.5 18.2 250 35.4 40.4 50.5 57.2 32.3 500 49.5 53.8 63.8 73.9 48.9

[0195] As can be seen from Table 7, in all of Examples 1 to 5, the collagenase inhibitory activity exhibits a concentration-dependent pattern of consistently increasing as the sample concentration increases.

[0196] As a result of the comparison between the examples, Example 4, a combination in which both ripening and ultrasonic treatment were applied, showed the highest collagenase inhibitory activity across the entire concentration range. For example, at 250 μg / ml, Example 4 showed an inhibitory activity of approximately 57.2%, whereas Example 5, which did not apply ripening or ultrasonic treatment, showed only about 32.3%, and Example 3, in which only ripening was applied to both raw materials, showed a level of about 50.5%. At 500 μg / ml, Example 4 showed the highest inhibition rate at 73.9%, followed by Example 3 at 63.8%, Examples 1 and 2 at 49.5% and 53.8%, respectively, and Example 5 at 48.9%, showing relatively low inhibitory activity.

[0197] From these results, it is clearly confirmed that the combination of Cornus fruit and mulberry leaf extracts that undergoes a ripening process and is further extracted using ultrasound tends to inhibit the activity of collagenase, a collagen-degrading enzyme, more strongly.

[0198] In other words, the raw material pretreatment (cooking) and ultrasonic extraction processes can be interpreted as increasing the release and activity expression of active ingredients related to collagen protection, such as polyphenols, iridoid glycosides, and imino sugar series components, thereby enhancing the effect of inhibiting collagen degradation.

[0199] Therefore, the experimental results of Experimental Example 5 support the fact that the cosmetic composition containing Cornus fruit extract and mulberry leaf extract according to the present invention has the effect of reducing collagenase activity in a concentration-dependent manner, and in particular, the combination of Example 4, in which Cornus fruit and mulberry leaf were ripened and then ultrasonically extracted, exhibits the best performance in terms of inhibiting collagen degradation.

[0201] Experimental Example 6. NBT assay

[0202] 1) Experimental Method

[0203] An NBT assay was performed to confirm the antiglycation activity of a composition containing Cornus fruit extract and mulberry leaf extract according to the present invention.

[0204] First, a reaction buffer solution was prepared by adding 5% BSA and 0.8 M glucose to 1.5 M PPB. Samples for each example were diluted to different concentrations and added to the buffer solution, and then reacted in a 35°C incubator for 7 days to carry out a non-enzymatic saccharification reaction between proteins and sugars.

[0205] After the saccharification reaction was completed, the reaction solution was transferred to a 1.5 mL centrifuge tube (E-tube) and mixed with NBT solution at a ratio of 1:100. The mixed tube was reacted in a 37°C incubator for 10 minutes under light blockage. After the reaction was finished, the absorbance was measured at a wavelength of 530 nm to obtain the absorbance values ​​for each reaction system (sample Abs.).

[0206] At this time, in order to correct for background absorbance caused by the color or turbidity of the sample itself, a reaction system containing only the sample and buffer solution was prepared separately for each sample concentration without adding BSA and glucose, and the absorbance was measured at 530 nm in the same way, and this was used as the background absorbance (sample background Abs.) of the corresponding sample.

[0207] In addition, a reaction system containing BSA and glucose but without the addition of the sample was set as the blank group, and the absorbance value of this reaction system at 530 nm was used as blank Abs. To compare antiglycation activity, aminoguanidine was treated to a final concentration of 500 μg / mL as a control, and the glycation reaction and NBT reaction were performed under the same conditions.

[0208] The anti-glycation activity for each sample was calculated using the values ​​obtained by correcting the sample background absorbance from the absorbance values ​​of the blank group and the sample treatment group, as shown in Equation 3 below.

[0209]

[0210] The evaluation of antiglycation activity according to the above NBT assay was performed independently three times for the blank group, aminoguanidine control group, and sample group for each example under the same conditions, and the antiglycation activity value for each condition was expressed as the average value of the measurements obtained from the three repeated experiments.

[0212] 2) Experimental Results

[0213] The experimental results are shown in Table 8 below, and the unit is percentage.

[0214] Concentration (µg / ml) control group Example 1 Example 2 Example 3 Example 4 Example 5 control group 30.0 10 13.2 14.1 17.0 19.4 11.9 50 20.4 22.8 28.1 32.7 19.2 100 26.3 26.9 31.8 38.0 23.2 250 31.5 33.7 39.1 44.5 28.1 500 37.0 38.7 44.0 49.5 31.0

[0215] As can be seen from Table 8, the antiglycation activity of aminoguanidine 500 μg / ml used as a control was about 30.0%, whereas Examples 1 to 5 of the present invention showed antiglycation activity equivalent to or higher than that of aminoguanidine in all concentration ranges, and showed a concentration-dependent trend in which antiglycation activity consistently increased as the sample concentration increased.

[0216] Specifically, at 10 μg / ml, Examples 1 to 5 exhibited antiglycation activity of approximately 11.9% to 19.4%, increased to approximately 23.2% to 38.0% at 100 μg / ml, approximately 28.1% to 44.5% at 250 μg / ml, and approximately 31.0% to 49.5% at 500 μg / ml, so that in the high concentration range, most examples exhibited higher antiglycation activity than aminoguanidine at 500 μg / ml (30.0%). This suggests that the combination of Cornus officinalis extract and mulberry leaf extract according to the present invention can have an equivalent or greater level of glycation inhibition ability compared to aminoguanidine, which is a widely used antiglycation agent.

[0217] In the comparison among the examples, Example 4, which applied both ripening and ultrasonic treatment, exhibited the highest anti-glycation activity across the entire concentration range, while Example 3, in which both raw materials were ripened, showed the next best activity. On the other hand, Example 5, which did not apply ripening or ultrasonic treatment, tended to show the lowest values ​​at the same concentration, indicating that the ripening and ultrasonic extraction processes for Cornus fruit and mulberry leaf raw materials contribute to the improvement of anti-glycation activity. For example, at 500 μg / ml, Example 4 showed the highest anti-glycation activity at 49.5%, whereas Example 5 showed a relatively low value of 31.0%.

[0218] Therefore, from the results of Experimental Example 6, it can be confirmed that the cosmetic composition containing Cornus fruit extract and mulberry leaf extract according to the present invention has anti-glycation activity that inhibits the non-enzymatic glycation reaction between proteins and sugars in a concentration-dependent manner, and in particular, Example 4, which is extracted using ultrasound after ripening Cornus fruit and mulberry leaves, is the best combination in terms of anti-glycation effect.

[0219] This serves as the basis for the composition of the present invention to work advantageously in reducing the glycation of skin proteins and the resulting structural damage.

[0221] Experimental Example 7. AGEs formation

[0222] 1) Experimental Method

[0223] To further confirm the anti-glycation activity of the composition containing Cornus officinalis extract and mulberry leaf extract according to the present invention, an AGEs formation assay using BSA and glucose was performed.

[0224] First, a buffer solution for the saccharification reaction was prepared by adding 5% BSA and 0.8 M glucose to 1.5 M phosphate buffer (PPB). Samples for each example were diluted to different concentrations and added to the buffer solution, and then reacted in a 35°C incubator for 7 days, during which time a non-enzymatic saccharification reaction between protein (BSA) and glucose was carried out.

[0225] After the reaction, 100 μL of each reaction solution was dispensed into each well of a 96-well black plate, and the fluorescence intensity was measured using a fluorescence plate reader (Varioskan™ ALF Multimode Microplate Reader, ThermoFisher, USA) under conditions of an excitation wavelength (Ex) of 340 nm and a fluorescence wavelength (Em) of 440 nm. The fluorescence intensity value of each reaction system measured at this time was defined as a sample.

[0226] To correct for background signals caused by the fluorescence or turbidity of the sample itself, a reaction system containing only the sample and PPB was separately prepared for each sample concentration without the addition of BSA or glucose, and the fluorescence intensity was measured under the same conditions and used as the sample background. Additionally, a reaction system containing BSA and glucose but without the sample was set as a blank, and the fluorescence intensity value of this reaction system was used as the blank.

[0227] As a control group, a group treated with 500 µg / mL aminoguanidine was provided.

[0228] The anti-glycation activity for each sample was calculated according to the following Equation 4 using the fluorescence values ​​of the blank and sample treatment groups, and the sample background fluorescence.

[0229]

[0230] The evaluation of anti-glycation activity according to the above AGEs formation assay was performed independently three times under the same conditions for the blank group, the aminoguanidine control group, and the sample group for each example, and the anti-glycation activity value for each condition was expressed as the average value of the measurements obtained from the three repeated experiments.

[0232] 2) Experimental Results

[0233] The experimental results are shown in Table 9 below, and the unit is percentage.

[0234] Concentration (µg / ml) control group Example 1 Example 2 Example 3 Example 4 Example 5 control group 40.2 10 22.2 24.3 27.7 35.3 18.3 50 26.6 29.5 34.5 40.6 22.0 100 33.2 35.7 40.8 45.5 28.0 250 37.8 41.2 44.3 49.8 35.3 500 43.9 46.5 52.5 56.8 40.6

[0235] As can be seen from Table 9, the antiglycation activity of aminoguanidine 500 μg / ml used as a control was approximately 40.2%, whereas Examples 1 to 5 of the present invention exhibit a concentration-dependent pattern in which antiglycation activity consistently increases with increasing concentration across all concentration ranges.

[0236] Specifically, at 10 μg / ml, Examples 1 to 5 show antiglycation activity of about 18.3% to 35.3%, which increases to about 22.0% to 40.6% at 50 μg / ml and about 28.0% to 45.5% at 100 μg / ml. At higher concentrations of 250 and 500 μg / ml, they show antiglycation activity of about 35.3% to 49.8% and 40.6% to 56.8%, respectively, confirming that in the high concentration range, most examples reach a level equivalent to or exceeding the antiglycation activity of aminoguanidine 500 μg / ml (40.2%).

[0237] As a result of comparing the examples, Example 4, which applied both ripening and ultrasonic treatment, showed the best anti-glycation activity across the entire concentration range. For example, at 500 μg / ml, Example 4 showed the highest anti-glycation activity at 56.8%, and Example 3, which applied only ripening to both raw materials, showed the next highest value at 52.5%. In contrast, Example 5, which did not apply ripening or ultrasonic treatment, showed a relatively low value of 40.6% at the same concentration, indicating that the ripening and ultrasonic extraction processes applied to the raw materials contribute to the improvement of anti-glycation activity that inhibits AGEs formation.

[0238] Therefore, from the results of Experimental Example 7, it can be confirmed that the cosmetic composition containing Cornus fruit extract and mulberry leaf extract according to the present invention has anti-glycation activity that significantly reduces the formation of AGEs in the BSA-glucose system, and in particular, Example 4, which is a combination of Cornus fruit and mulberry leaf extracted using ultrasound after ripening, is the best combination in terms of anti-glycation effect.

[0239] This suggests that the composition of the present invention can reduce the accumulation of advanced glycation end products of proteins, thereby acting favorably in alleviating skin structural damage and functional decline caused by glycation.

[0241] Experimental Example 8. α-amylase inhibition assay

[0242] 1) Experimental Method

[0243] In order to confirm the function of a composition containing Cornus fruit extract and mulberry leaf extract according to the present invention when provided as a food composition, α-amylase inhibitory activity was evaluated.

[0244] The sample and positive control (100 μg / mL Acarbose) were dissolved in DMSO to prepare a stock, filtered through a 0.22 μm syringe filter, and then diluted with 0.05 M PPB (pH 6.9) for use in the experiment.

[0245] 50 µl of the prepared sample, 100 µl of 5 U / ml α-amylase, 100 µl of 0.05 M PPB (pH 6.9), and 100 µl of 1% starch were added to an E-tube, mixed, and reacted at room temperature for 15 minutes.

[0246] After adding 100 µl of DNS solution, the reaction was stopped and color developed in a 90°C heat-block for 10 minutes, then removed and cooled.

[0247] 900 µl of purified water was added to the reacted E-tube and mixed, then 100 µl was dispensed into a 96-well plate in triplicate, and the absorbance was measured at a wavelength of 540 nm using a plate reader (SpectraMax ABS Plu, Molecular Devices, USA).

[0248] α-amylase inhibitory activity was calculated according to the following mathematical formula 5.

[0249]

[0251] 2) Experimental Results

[0252] The experimental results are shown in Table 10 below, and the unit is percentage.

[0253] Concentration (µg / ml) control group Example 1 Example 2 Example 3 Example 4 Example 5 control group 61.0 10 5.6 4.8 5.9 6.5 4.4 50 19.6 18.6 19.6 20.2 17.2 100 28.0 26.2 29.8 30.6 23.4 250 33.6 32.2 35.3 40.5 30.3 500 37.5 36.3 39.9 43.4 32.2

[0254] As shown in Table 10, Examples 1 to 5 showed a tendency for α-amylase inhibitory activity to increase in a concentration-dependent manner across the entire concentration range.

[0255] This suggests that the combination of Cornus fruit extract and mulberry leaf extract of the present invention influences starch-degrading enzyme activity, thereby providing a function that mitigates the rate of sugar production during the carbohydrate digestion process.

[0256] In the comparison between the examples, Example 4, a combination that applied both ripening and ultrasonic extraction, showed the highest inhibitory activity across the entire concentration range, followed by Example 3, a combination that treated both raw materials with ripening, which showed a high tendency.

[0257] On the other hand, the comparative example without ripening and ultrasonic treatment showed relatively lower inhibitory activity values ​​at the same concentration.

[0258] Through these results, it can be seen that the composition containing Cornus fruit extract and mulberry leaf extract of the present invention exhibits α-amylase inhibitory activity, and in particular, that a combination of ripening of the raw materials and ultrasonic treatment is effective.

[0259] At this time, since the α-amylase inhibitory activity can act to alleviate the burden of sugar production and absorption resulting from starch degradation after a meal, it suggests that when the composition of the present invention is provided as a food composition, it can contribute to alleviating the environment in which the saccharification reaction proceeds.

[0261] Experimental Example 9. α-glucosidase inhibition assay

[0262] 1) Experimental Method

[0263] The α-glucosidase inhibitory activity of a composition containing Cornus fruit extract and mulberry leaf extract according to the present invention was confirmed.

[0264] The sample and positive control (100 μg / mL Acarbose) were dissolved in DMSO and filtered through a 0.22 μm syringe filter to prepare a stock, which was then diluted with 0.2 M PPB (pH 6.8) and used in the experiment.

[0265] 50 µl of the prepared sample or positive control, 50 µl of 0.1 U / ml α-glucosidase, and 50 µl of 0.2 M PPB (pH 6.8) were dispensed into a 96-well plate and reacted at 37 °C for 15 minutes. After the reaction, 100 µl of 2.5 mM pNPG was added and reacted at 37 °C for 15 minutes, after which 50 µl of 0.1 M NaOH was added to stop the reaction.

[0266] Next, absorbance was measured at a wavelength of 405 nm using a plate reader (SpectraMax ABS Plus, Molecular Devices, USA).

[0267] α-glucosidase inhibitory activity was calculated using the same method as Equation 5 above.

[0269] 2) Experimental Results

[0270] The experimental results are shown in Table 11 below.

[0271] Concentration (µg / ml) control group Example 1 Example 2 Example 3 Example 4 Example 5 control group 45.5 10 5.8 5.3 7.5 10.6 3.4 50 22.7 21.5 26.9 39.9 19.7 100 59.1 57.2 66.5 71.3 30.0 250 65.3 63.7 71.8 88.2 39.7 500 72.8 69.7 81.5 92.6 49.1

[0272] As can be seen from Table 11, all of Examples 1 to 5 showed a tendency for α-glucosidase inhibitory activity to increase in a concentration-dependent manner across all concentration ranges.

[0273] When looking at the differences between the examples, Example 4, a combination in which both ripening and ultrasonic treatment were applied, showed the highest inhibitory activity across the entire concentration range, followed by Example 3, a combination in which both raw materials were ripened, and Examples 1 and 2, in which only one side was ripened.

[0274] On the other hand, Example 5, which did not apply ripening and ultrasonic treatment, showed a relatively low inhibitory activity value at the same concentration.

[0275] Therefore, from the results of Experimental Example 9, it can be confirmed that the composition containing Cornus fruit extract and mulberry leaf extract of the present invention exhibits α-glucosidase inhibitory activity, and in particular, the combination of ripening of the raw material and ultrasonic treatment is superior in terms of α-glucosidase inhibitory activity.

[0276] At this time, since the α-glucosidase inhibitory activity can act to alleviate the burden of sugar production and absorption during the carbohydrate breakdown process, it suggests that when the composition of the present invention is provided as a food composition, it can provide an anti-glycation function in terms of alleviating the environment in which the glycation reaction proceeds.

[0278] In summary, the composition comprising Cornus fruit extract and mulberry leaf extract according to the present invention can provide anti-glycation activity that significantly reduces non-enzymatic glycation reactions between proteins and sugars and the formation of Advanced Glycation End Products (AGEs), and together with this, complex physiological activities such as antioxidant activity, anti-inflammatory activity, and inhibition of elasticity-related enzymes can be expressed.

[0279] Accordingly, the composition of the present invention can contribute to mitigating structural denaturation of biological proteins and the accompanying deterioration of their condition that may be induced or exacerbated by glycosylation and oxidative stress.

[0280] In particular, when the composition of the present invention is provided as a cosmetic composition, the antioxidant activity due to free radical scavenging, the anti-inflammatory activity due to inhibition of NO production, and the protective effect on elastic fibers due to inhibition of elastase and collagenase are expressed together with the anti-glycation activity, thereby alleviating the deterioration of skin conditions such as collagen structure damage caused by glycation and the accompanying decrease in skin elasticity, wrinkle formation, roughness of skin texture, and deterioration of skin tone, and contributing to maintaining a smoother and more elastic skin condition.

[0281] In addition, when the composition of the present invention is provided as a food composition, it can act to alleviate the burden of sugar production and absorption after a meal through the expression of function in indicators related to carbohydrate-degrading enzyme activity, and this can provide an anti-glycation effect by alleviating the environment that increases exposure to reducing sugars in vivo, thereby reducing the conditions that promote protein glycation reactions and AGEs formation.

[0282] Furthermore, since the composition of the present invention can be provided as a high-performance active ingredient in powder form through a concentration and freeze-drying process, it is easy to apply it to various cosmetic formulations such as toners, lotions, creams, essences, gels, packs, and foundations, and it is also possible to commercialize it into various food formulations such as powders, granules, tablets, capsules, beverages, and health functional food formulations.

[0283] Therefore, the present invention can be utilized as a substitute or supplementary material for synthetic anti-glycation agents and has useful industrial applicability applicable to cosmetic products for improving skin condition and food products having anti-glycation functions.

[0285] Although the present invention has been described together with the accompanying drawings, this is merely one example among various embodiments containing the gist of the present invention, and is intended to enable those skilled in the art to easily implement it. It is clear that the present invention is not limited to the embodiments described above. Accordingly, the scope of protection of the present invention should be interpreted by the following claims, and all technical ideas within an equivalent scope by modification, substitution, replacement, etc., within the scope that does not deviate from the gist of the present invention shall be included in the rights of the present invention. Furthermore, it is clarified that some components in the drawings are provided in an exaggerated or reduced form compared to the actual form to more clearly explain the configuration. Explanation of the symbols

[0286] (S10): Cornus officinalis preparation stage (S20): Mulberry leaf preparation stage (S30): Step for preparing Cornus fruit extract (S40): Step for preparing mulberry leaf extract (S31): Step for manufacturing Cornus fruit extract powder (S41): Step for manufacturing mulberry leaf extract powder (S50): Completion stage

Claims

Claim 1 A composition having an anti-glycation effect comprising Cornus officinalis extract and mulberry leaf extract, wherein the Cornus officinalis extract and mulberry leaf extract are included in a weight ratio of 1 to 3:1 to 3 (Cornelian cherry extract: mulberry leaf extract), the Cornus officinalis extract is prepared by ripening Cornus officinalis in a drying chamber at 40 to 50°C for 3 to 4 days, grinding it, roasting it at 70 to 90°C for 1 to 3 hours, and then extracting it, and the mulberry leaf extract is prepared by ripening mulberry leaves in a drying chamber at 40 to 50°C for 1 to 2 days, grinding them, roasting them at 80 to 100°C for 10 to 30 minutes, and then extracting them. Claim 2 A composition according to claim 1, characterized in that the composition is a cosmetic composition. Claim 3 A composition according to claim 1, characterized in that the composition is a food composition. Claim 4 delete Claim 5 A composition according to claim 1, wherein the extraction of the Cornus fruit extract is a process of extracting the Cornus fruit, which has been ripened, crushed, and roasted, in purified water or an aqueous ethanol solution for 2 to 3 hours under ultrasonic conditions of 350 to 450 watts, 35 to 45 kHz, and a temperature of 35 to 45°C, and the extraction of the mulberry leaf extract is a process of extracting the mulberry leaf, which has been ripened, crushed, and roasted, in purified water or an aqueous ethanol solution for 1 to 2 hours under ultrasonic conditions of 350 to 450 watts, 35 to 45 kHz, and a temperature of 35 to 45°C. Claim 6 A composition according to claim 1, wherein the Cornus officinalis extract is prepared by concentrating the extract to 55 to 65 weight% relative to the weight of the Cornus officinalis raw material after extraction to produce a Cornus officinalis extract concentrate, and then freeze-drying the Cornus officinalis extract concentrate to 35 to 45 weight% relative to the weight of the Cornus officinalis raw material, and the Mulberry leaf extract is prepared by concentrating the extract to 18 to 25 weight% relative to the weight of the Mulberry leaf raw material after extraction to produce a Mulberry leaf extract concentrate, and then freeze-drying the Mulberry leaf extract concentrate to 13 to 17 weight% relative to the weight of the Mulberry leaf raw material. Claim 7 A composition according to claim 1, characterized in that the composition provides an anti-glycation effect, and when the composition is a cosmetic composition, provides an antioxidant effect, an anti-inflammatory effect and an effect that inhibits collagenase activity, and when the composition is a food composition, provides an α-amylase and α-glucosidase inhibitory effect. Claim 8 A Cornus fruit preparation step (S10) for preparing a Cornus fruit extract raw material by ripening Cornus fruit in a drying chamber at 40 to 50°C for 3 to 4 days, grinding it, and then roasting it at 70 to 90°C for 1 to 3 hours; a mulberry leaf preparation step (S20) for preparing a mulberry leaf extract raw material by ripening mulberry leaves in a drying chamber at 40 to 50°C for 1 to 2 days, grinding them, and then roasting them at 80 to 100°C for 10 to 30 minutes; a Cornus fruit extract preparation step (S30) for preparing a Cornus fruit extract by extracting the Cornus fruit extract raw material; a mulberry leaf extract preparation step (S40) for preparing a mulberry leaf extract by extracting the mulberry leaf extract raw material; and a completion step (S50) for preparing a composition by mixing the Cornus fruit extract and the mulberry leaf extract in a weight ratio of 1 to 3 : 1 to 3 (Cornelian fruit extract : mulberry leaf extract). A method for preparing a composition having an anti-glycation effect, comprising Cornus fruit extract and mulberry leaf extract. Claim 9 A manufacturing method according to claim 8, characterized in that the composition manufactured in the completion step (S50) is a cosmetic composition. Claim 10 A method of manufacturing according to claim 8, characterized in that the composition produced in the completion step (S50) is a food composition. Claim 11 delete Claim 12 A method for manufacturing according to claim 8, wherein the step of preparing the Cornus fruit extract (S30) is a step of preparing the Cornus fruit extract by extracting the Cornus fruit extract raw material in purified water or an aqueous ethanol solution for 2 to 3 hours under ultrasonic conditions of 350 to 450 watts, 35 to 45 kHz, and a temperature condition of 35 to 45°C, and the step of preparing the mulberry leaf extract (S40) is a step of preparing the mulberry leaf extract by extracting the mulberry leaf extract raw material in purified water or an aqueous ethanol solution for 1 to 2 hours under ultrasonic conditions of 350 to 450 watts, 35 to 45 kHz, and a temperature condition of 35 to 45°C. Claim 13 A method for manufacturing according to claim 8 or 12, comprising a step (S31) of manufacturing a Cornus fruit extract powder by concentrating the Cornus fruit extract to 55 to 65 weight% relative to the weight of the Cornus fruit extract source material to produce a Cornus fruit extract concentrate, and then freeze-drying the Cornus fruit extract concentrate to 35 to 45 weight% relative to the weight of the Cornus fruit extract source material to produce a Cornus fruit extract powder; a step (S41) of manufacturing a mulberry leaf extract powder by concentrating the mulberry leaf extract to 18 to 25 weight% relative to the weight of the mulberry leaf extract source material to produce a mulberry leaf extract concentrate, and then freeze-drying the mulberry leaf extract concentrate to 13 to 17 weight% relative to the weight of the mulberry leaf extract source material to produce a mulberry leaf extract powder; and a completion step (S50) characterized by being a step of completing a composition by mixing the Cornus fruit extract powder and the mulberry leaf extract powder.

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