KR102763933B1 - Composition for improving skin condition comprising extract of Pisum sativum var. arvense (L.) Poir. - Google Patents

Abstract

The present invention relates to a composition for improving skin condition. The present invention relates to a composition for skin regeneration, skin elasticity improvement, prevention or improvement of skin wrinkles, prevention or improvement of skin aging, improvement of skin inflammation, skin moisturizing and/or skin whitening. The Tutankhamun pea extract according to the present invention exhibits a skin regeneration effect, a wrinkle improvement effect, a skin moisturizing effect by promoting collagen synthesis of skin fibroblasts, an antioxidant effect by inhibiting free radical production, an anti-inflammatory effect by inhibiting NO production, and a whitening effect by inhibiting melanin synthesis, thereby being usable as a composition for improving skin condition. In addition, a composition containing the Tutankhamun pea extract of the present invention or a fraction thereof has an excellent effect in preventing or treating skin diseases.

Description

Composition for improving skin condition comprising extract of Pisum sativum var. arvense (L.) Poir.

The present invention relates to a composition for improving skin condition.

Collagen, a major component of the extracellular matrix, is the main matrix protein produced by fibroblasts in the skin. It is also an important protein that accounts for about 30% of the total weight of biological proteins and has a rigid triple helix structure. Collagen forms most of the organic material of skin, tendons, bones, and teeth, and its content is particularly high in bone and skin (dermis). It exists as fibrous inclusions in most other body structures.

Collagen is a relatively weak immunogen, partly due to the shielding of potential antigenic determinants by the helical structure of collagen, which also makes collagen resistant to proteolysis. The main functions of collagen are known to be mechanical strength of the skin, resistance of connective tissue, cohesion of tissues, support of cell adhesion, and induction of cell division and differentiation (when growing an organism or healing wounds) (Non-patent Document 1).

This collagen decreases with age and photoaging caused by ultraviolet irradiation, and it is known that this is closely related to the formation of wrinkles on the skin (Non-patent literature 2). In addition, collagen plays an important role in wound healing, and can quickly and scarlessly heal wounds by promoting collagen synthesis in damaged epithelium.

In addition, collagen is a matrix protein that plays an important role in retaining moisture within the dermis layer. It is known that these matrix proteins absorb moisture and increase the moisture retention capacity within the structure they form, allowing the skin to maintain an appropriate moisture content (i.e., skin hydration), and through this, it is involved in maintaining skin elasticity.

Previously, products that incorporated collagen into external skin preparations such as cosmetics or ointments were released to utilize the skin moisturizing and wound healing effects of collagen. However, since these products applied collagen itself to the skin surface, it was difficult to expect moisturizing or wound healing effects due to the percutaneous absorption of collagen, which is a high molecular weight substance, and therefore it could not be said to exhibit the essential functions of improving skin function and healing wounds.

To solve these problems, interest in collagen synthesis accelerators has increased, and existing collagen synthesis accelerators include vitamin C, retinoic acid, carcinogenic growth factors (non-patent document 3), proteins derived from animal placenta (patent document 1), betulinic acid (patent document 2), and chlorella extracts (patent documents 3 and 4).

However, these substances have safety issues such as irritation and redness when applied to the skin, so the amount of use is limited, or the effect is minimal, so it is not possible to expect the effect of improving skin function or wound healing. Therefore, there is an urgent need to develop a new skin external composition that is safer for the body and more effective than existing skin external compositions.

Melanin, which determines human skin color, is produced in melanocytes. Specifically, melanin is a dark brown pigment formed by polymerization and oxidation reaction of tyrosine, an amino acid present in the body, by enzymes such as tyrosinase present in melanocytes. Melanin formed in this way moves to epidermal cells called keratinocytes through the dendritic projections of melanocytes. Melanin that moves to keratinocytes can be removed by falling off from the skin when keratinocytes fall off from the epidermis. However, since there is no enzyme that breaks down melanin in the body, melanin once formed is not broken down in the body. Therefore, suppressing the production of melanin is the key to brightening the skin.

The need for research on whitening agents as cosmetics has been increasing recently as the living standards of East Asian countries, which emotionally prefer white skin, have improved and skin darkening has become recognized as skin aging caused by ultraviolet rays. Accordingly, substances that exhibit tyrosinase inhibitory activity, such as ascorbic acid, hydroquinone, glutathione, and arbutin, have been mixed and used in cosmetics and medicines (Patent Documents 5 and 6), but most of them have insufficient effects or are unstable in terms of formulation, which limits their utility. In particular, compounds such as hydroquinone exhibit strong depigmentation effects and have skin sensitization properties themselves, which can cause skin allergies, and cause side effects such as altering the function of normal skin and causing vitiligo, so their use is limited in terms of safety for the skin.

In addition, even if it is a substance that inhibits the activity of tyrosinase as mentioned above, if such a substance acts on keratinocytes, which are keratinocytes, and increases the production of factors that promote the biosynthesis of melanin, the whitening effect may be weakened or even have a negative effect. Therefore, a simple tyrosinase activity inhibitor cannot be considered a good whitening agent. Accordingly, although many tyrosinase inhibitors are being developed, it is known that almost all of them do not exhibit an excellent whitening effect.

When the skin is exposed to excessive ultraviolet rays or pollutants, or when free radicals in the body are activated due to stress, disease, etc., it promotes damage to skin cells and tissues, accelerating skin aging symptoms such as an increase in skin wrinkles. Specifically, when the main components of the skin such as lipids, proteins, polysaccharides, and nucleic acids are oxidized by free radicals, skin cells and tissues are destroyed, and skin aging occurs as a result. In particular, when proteins are oxidized, the connective tissues of the skin such as collagen, hyaluronic acid, elastin, proteoglycan, and fibronectin are cut, causing excessive inflammation and damaging skin elasticity. In addition, skin cells with damaged proteins cannot perform their functions smoothly, so the metabolic activity of the cells decreases, and this functional and structural damage causes wrinkles, which further accelerates skin aging. Therefore, by eliminating free radicals generated during the body's metabolic process, free radicals mediated by ultraviolet irradiation, and inflammatory reactions, cell membranes are protected, while cells that have already been damaged are regenerated through active metabolism to proliferate cells, allowing the skin to recover quickly and maintain healthy skin.

Therefore, in the past, there have been many efforts to obtain anti-aging effects on skin by mixing antioxidants such as superoxide dismutase (SOD), vitamin E derivatives, ascorbic acid and its derivatives, and flavonoids into cosmetics. However, problems have arisen in terms of skin safety and stability when mixing cosmetics, and thus their use has been limited.

Inflammation is the body's immune response to injury or disease, and oxidative stress such as ultraviolet rays, active oxygen, and free radicals activate inflammatory factors, causing various diseases and skin aging. Vasoactive polypeptides such as kinin, plasmin, and complement cause vasodilation and contraction and chemotaxis, and lymphokines such as interleukin-6 (IL-6) and arachidonic acid are responsible for the inflammatory response. Arachidonic acid is metabolized into prostaglandins and leukotrienes, which are inflammatory mediators, through two pathways, cyclooxygenase and lipooxygenase, and mediate various inflammatory responses.

On the other hand, an anti-inflammatory agent is an agent that has the effect of removing the source of inflammation, reducing biological responses, and symptoms in order to eliminate inflammation. Currently, substances used for anti-inflammation purposes include non-steroidal agents such as flufenamic acid, ibuprofen, benzydamine, and indomethacin, and steroidal agents such as prednisolone and dexamethasone. In addition, allantoin, azulene, and hydrocortisone are known to be effective in anti-inflammation, but these substances have limitations in the amount of use due to safety issues on the skin, or the effect is minimal, so there is a problem that it is difficult to expect a practical anti-inflammatory effect.

Japanese Patent Publication No. 8-231370 Japanese Patent Publication No. 8-208424 Japanese Patent Publication No. 9-40526 Japanese Patent Publication No. 10-36283 Republic of Korea Patent Publication No. 10-0923141 Republic of Korea Patent Publication No. 10-0921947

Van der Rest et al., Ann NY Acad Sci, 1990 Arthur K. Balin et al., Aging and the skin, 1989 Cardinale G. 35 al, Adv. Enzymol., 41, p.425, 1974

Therefore, the problem to be solved by the present invention is to solve the above problems, and to provide a new active ingredient that has excellent effects of skin regeneration, skin elasticity improvement, wrinkle prevention or improvement, skin aging prevention or improvement, skin inflammation improvement, skin moisturizing or skin whitening while having few side effects and being safe for the human body, i.e., such useful uses of the active ingredient.

In other words, the problem that the present invention seeks to solve is to provide a composition containing the above-mentioned excellently effective active ingredient as an effective ingredient.

In order to solve the above problems, the present invention provides a composition for skin regeneration, skin elasticity improvement, prevention or improvement of skin wrinkles, prevention or improvement of skin aging, improvement of skin inflammation, skin moisturizing and/or skin whitening, preferably a cosmetic composition, a pharmaceutical composition or a health functional food composition, comprising a Tutankhamun pea extract or a fraction thereof as an effective ingredient.

That is, the present invention provides a use of a Tutankhamun pea extract or a fraction thereof as a cosmetic composition, a pharmaceutical composition or a health functional food composition, and new uses of skin regeneration, skin elasticity improvement, skin wrinkle prevention or improvement, skin aging prevention or improvement, skin inflammation improvement, skin moisturizing and/or skin whitening.

The inventors of the present invention conducted research on substances that are non-toxic and have excellent skin condition improvement effects from natural resources existing in nature, and discovered that Tutankhamun pea extract exhibits the effects of promoting collagen synthesis, inhibiting collagenase activity, inhibiting melanin synthesis, scavenging free radicals, promoting cell proliferation, and inhibiting NO production, and completed a composition for improving skin condition utilizing the same.

The Tutankhamun pea used in the present invention is a propagated version of the pea excavated at the time of the discovery of King Tutankhamun's tomb in Egypt in the early 1990s. The common pea currently cultivated has white flowers and green pods, but unlike these, this one has red flowers and purple pods. Taxonomically, the Tutankhamun pea ( Pisum sativum Var. arvense (L.) Poir.) is listed as a different species from the common pea ( Pisum sativum L.), and the first nominator, Linnaeus, named the pea as Pisum sativum L. in 1753, and the red pea as Pisum arvense L. was named. Later, in 1804, Poiret. gave it the current scientific name, Pisum. sativum var. arvense (L.) Poir., but the term variant usually refers to a species with differences that came from geographically similar regions, but in the case of the above two peas, there is no report that they are geographically the same.

The Tutankhamun pea of the present invention is not limited to the type of Tutankhamun pea, and any Tutankhamun pea can be used regardless of its source, such as cultivated ones or commercially purchased ones, and all parts of the Tutankhamun pea, such as leaves, stems, roots, and seeds, can be used.

The term "extract" used in the present invention includes the extract itself and all formulations that can be formed using the extract, such as an extract obtained by the extraction treatment of the Tutankhamun pea, a diluted or concentrated extract, a dried product obtained by drying the extract, a controlled or purified product of the extract, or a mixture thereof.

In the above Tutankhamun pea extract of the present invention, the method of extracting the Tutankhamun pea is not particularly limited, and extraction can be performed according to a method commonly used in the relevant technical field. Non-limiting examples of the extraction method include solvent extraction, hot water extraction, ultrasonic extraction, filtration, and reflux extraction, and these can be performed alone or in combination of two or more methods.

In the present invention, the type of extraction solvent used to extract the Tutankhamun pea is not particularly limited, and any solvent known in the art can be used. Non-limiting examples of the extraction solvent include water; lower alcohols having C ₁ to C ₄ such as methanol, ethanol, propyl alcohol, butyl alcohol; polyhydric alcohols such as glycerin, butylene glycol, propylene glycol; and hydrocarbon solvents such as methyl acetate, ethyl acetate, acetone, benzene, hexane, diethyl ether, dichloromethane; or mixtures thereof. Preferably, water, lower alcohols, 1,3-butylene glycol, and ethyl acetate can be used alone or in combination of two or more. In the present invention, as a solvent for extracting Tutankhamun pea, water (or distilled water), methanol or a mixed solvent thereof can be more preferably used, and even more preferably, a solvent in which water (or distilled water) and methanol are mixed in a volume ratio of 1:1 to 10, preferably about 3:7, can be used. Tutankhamun pea can be extracted one or more times using the solvent to prepare a solvent extract of Tutankhamun pea, and the solvent extract can be distilled under reduced pressure and then freeze-dried or spray-dried to prepare a dried extract.

In the present invention, the extract obtained by heat extraction or cold extraction can be used as is after filtering to remove floating solid particles, for example, filtering the particles using nylon or the like, or filtering using a cryofiltration method, etc., or after drying using freeze drying, hot air drying, spray drying, etc.

The term "fraction" as used in the present invention means a result obtained by performing fractionation to separate a specific component or a specific group of components from a mixture containing various components.

The fractionation method for obtaining the fraction in the present invention is not particularly limited, and can be performed according to a method commonly used in the relevant technical field. A non-limiting example of the fractionation method is a method of obtaining a fraction from an extract obtained by extracting Tutankhamun pea by treating the extract with a predetermined solvent.

In the present invention, the type of fractionation solvent used to obtain the fraction is not particularly limited, and any solvent known in the art can be used. Non-limiting examples of the fractionation solvent include polar solvents such as water, distilled water, and alcohol; non-polar solvents such as hexane, ethyl acetate, chloroform, and dichloromethane. These may be used alone or in combination of two or more. When using alcohol among the fractionation solvents, it is preferable to use a C ₁ to C ₄ alcohol.

The composition according to the present invention contains Tutankhamun pea extract as an active ingredient.

In the present invention, the meaning of "included as an effective ingredient" means that an extract is added to an extent that the composition according to the present invention can exhibit a skin condition improvement effect, and includes the meaning that various ingredients can be added as auxiliary ingredients for skin delivery and stabilization, etc., and can be formulated in various forms.

In the present invention, the Tutankhamun pea extract may be included in an amount of 0.0001 to 50 wt% based on the total weight of the composition, and preferably 0.0005 to 10 wt%. Within the above range, there is an advantage of exhibiting excellent skin regeneration, skin elasticity improvement, skin wrinkle prevention or improvement, skin aging prevention or improvement, skin inflammation improvement, skin moisturizing or skin whitening effects, and there is an advantage of stabilizing the formulation of the composition.

The composition according to the present invention has biological activity and exhibits excellent effects in improving skin condition.

More specifically, the composition according to the present invention contains a Tutankhamun pea extract or a fraction thereof as an effective ingredient and has effects of improving skin condition, specifically skin regeneration, improving skin elasticity, preventing or improving skin wrinkles, preventing or improving skin aging, improving skin inflammation, moisturizing the skin, and/or whitening the skin.

The term "improvement of skin condition" as used in the present invention means improvement of skin condition, specifically skin regeneration, improvement of skin elasticity, prevention or improvement of skin wrinkles, prevention or improvement of skin aging, improvement of skin inflammation, and/or skin whitening.

The term "skin regeneration" used in the present invention refers to the recovery of skin tissue from damage caused by external and internal causes of the skin. Damage caused by external causes may include ultraviolet rays, external pollutants, wounds, trauma, etc., and damage caused by internal causes may include stress, etc.

The term "improvement of skin elasticity" used in the present invention means increasing the elasticity of the skin, suppressing or inhibiting the loss of skin elasticity, or alleviating elasticity that has already been reduced.

The term "prevention or improvement of skin wrinkles" used in the present invention refers to suppressing or inhibiting the formation of wrinkles on the skin, or alleviating wrinkles that have already formed.

The above skin regeneration, improvement of skin elasticity, prevention or improvement of skin wrinkles are not limited thereto, but it can be confirmed through the efficacy of the Tutankhamun pea extract or its fractions to effectively increase the total amount of collagen, and specifically, the efficacy of inhibiting collagenase activity, and the efficacy of inhibiting the production of MMP-1, which is the collagenase enzyme related to the increase in the total amount of collagen.

According to a specific embodiment of the present invention in relation to the above effects, when human fibroblasts were treated with Tutankhamun pea extract or fractions thereof at various concentrations, it was confirmed that the total amount of collagen produced in the human fibroblasts increased in a concentration-dependent manner.

In another embodiment, when human fibroblasts were treated with various concentrations of the Tutankhamun pea extract or fractions thereof and then treated with tumor necrosis factor-α (TNF-α), a substance known to produce collagenase MMP-1, it was confirmed that the amount of MMP-1 synthesized from the human fibroblasts was inhibited in a concentration-dependent manner of the Tutankhamun pea extract or fractions thereof.

The term "skin aging" used in the present invention means all diseases caused by an increase in active oxygen, and non-limiting examples of the skin aging include wrinkles, sagging skin, and decreased elasticity.

The term "improvement of skin inflammation" as used in the present invention means any action that alleviates an immune response and suppresses NO production.

The term "skin whitening" used in the present invention means any action that inhibits or prevents the deposition of melanin on the skin by inhibiting the synthesis of melanin.

The term "skin pigmentation" as used in the present invention means any action or condition in which the color of the skin darkens due to increased synthesis of melanin.

The term "skin moisturizing" used in the present invention means all actions that supply moisture to the skin or block the evaporation of moisture to maintain the flexibility of the skin and induce uniform exfoliation of dead skin cells to maintain a smooth surface. The term "skin dryness" used in the present invention is a term that comprehensively means a state in which the skin lacks moisture.

As used herein, the term "improvement" means any action that at least reduces a parameter related to alleviation or treatment of a condition, for example, the degree of a symptom.

The Tutankhamun pea extract or fraction thereof of the present invention has the effect of effectively inhibiting the production of melanin, and specifically exhibits the effect of reducing the total amount of melanin synthesized.

Melanin is a dark brown pigment produced by melanocytes and determines human skin color. It can be expected to have the effect of brightening the skin by inhibiting melanin pigmentation.

According to a specific example of the present invention with respect to the above efficacy, when mouse melanoma cells (B-16 mouse melanoma cells) were treated with the Tutankhamun pea extract or fractions thereof at various concentrations, it was confirmed that the melanin content of the melanoma cells decreased in a concentration-dependent manner depending on the treatment concentration of the Tutankhamun pea extract or fractions thereof.

The Tutankhamun pea extract of the present invention or a fraction thereof exhibits excellent skin cell activation efficacy. Specifically, the Tutankhamun pea extract of the present invention or a fraction thereof exhibits efficacy in promoting skin cell proliferation.

According to a specific embodiment of the present invention with respect to the above efficacy, it was confirmed that when human fibroblasts were treated with the Tutankhamun pea extract or a fraction thereof, the effect of promoting the proliferation of human fibroblasts was exhibited.

The Tutankhamun pea extract of the present invention or a fraction thereof exhibits excellent antioxidant efficacy. Specifically, the Tutankhamun pea extract of the present invention or a fraction thereof exhibits an efficacy in scavenging free radicals, and is particularly excellent in the efficacy in scavenging DPPH.

Free radicals are either introduced from outside the body or generated within the body, and they cause many problems, such as accelerating aging or causing cancer.

According to a specific embodiment of the present invention with respect to the above efficacy, it was confirmed that when DPPH was added to the Tutankhamun pea extract and left to stand, it exhibited excellent free radical scavenging ability.

In addition, the Tutankhamun pea extract of the present invention or a fraction thereof exhibits excellent anti-inflammatory efficacy. Specifically, the Tutankhamun pea extract of the present invention or a fraction thereof exhibits efficacy in inhibiting the production of NO, and is particularly excellent in the efficacy in inhibiting NO production, such as L-NMMA, which is known as an NO production inhibitor.

NO is produced by the enzyme NOS (NO synthase), and this production process can be activated by immune stimuli such as IFNγ, TNFα, and LPS.

According to a specific example of the present invention with respect to the above efficacy, when mouse macrophage RAW264.7 cells were treated with various concentrations of the Tutankhamun pea extract and then treated with LPS (Lipopolysaccharide) as an immune stimulant, it was confirmed that the amount of NO produced decreased in a concentration-dependent manner depending on the treatment concentration of the Tutankhamun pea extract or a fraction thereof.

The present invention provides a cosmetic composition for skin regeneration, skin elasticity improvement, prevention or improvement of skin wrinkles, prevention or improvement of skin aging, improvement of skin inflammation, skin moisturizing and/or skin whitening, comprising a Tutankhamun pea extract or a fraction thereof as an effective ingredient.

The cosmetic composition of the present invention can be prepared in a formulation selected from the group consisting of, but not limited to, a solution, an external ointment, a cream, a foam, a nourishing toner, an emollient toner, a pack, an emollient, an emulsion, a makeup base, an essence, a soap, a liquid cleanser, a bath agent, a sunscreen cream, a sun oil, a suspension, an emulsion, a paste, a gel, a lotion, a powder, a soap, a surfactant-containing cleansing, an oil, a powder foundation, an emulsion foundation, a wax foundation, a patch, and a spray.

The cosmetic composition of the present invention may additionally contain one or more cosmetically acceptable carriers that are blended with general skin cosmetics, and may appropriately blend conventional ingredients such as oil, water, surfactants, moisturizers, lower alcohols, thickeners, chelating agents, pigments, preservatives, fragrances, etc., but is not limited thereto.

The cosmetically acceptable carrier included in the cosmetic composition of the present invention varies depending on the formulation of the cosmetic composition.

When the formulation of the present invention is an ointment, paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide, etc. can be used as a carrier component, but are not limited thereto. These can be used alone or in a mixture of two or more.

When the formulation of the present invention is a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate, polyamide powder, etc. can be used as carrier components, and especially in the case of a spray, a propellant such as chlorofluorocarbons, propane/butane, or dimethyl ether can be additionally included, but is not limited thereto. These can be used alone or in a mixture of two or more.

When the formulation of the present invention is a solution or an emulsion, a solvent, a solubilizer or an emulsifier can be used as a carrier component, and for example, water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butyl glycol oil, and the like can be used, and in particular, cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil and sesame oil, glycerol aliphatic ester, polyethylene glycol or fatty acid ester of sorbitan can be used, but is not limited thereto. These can be used alone or in a mixture of two or more.

When the formulation of the present invention is a suspension, liquid diluents such as water, ethanol or propylene glycol, suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar or tragacanth may be used as carrier components, but are not limited thereto. These may be used alone or in combination of two or more.

When the formulation of the present invention is soap, alkali metal salts of fatty acids, fatty acid hemiester salts, fatty acid protein hydrolysates, isethionates, lanolin derivatives, fatty alcohols, vegetable oils, glycerol, sugars, etc. can be used as carrier components, but are not limited thereto. These can be used alone or in a mixture of two or more.

In addition, the present invention provides a pharmaceutical composition for skin regeneration, skin elasticity improvement, prevention or improvement of skin wrinkles, prevention or improvement of skin aging, improvement of skin inflammation, skin moisturizing and/or skin whitening, comprising a Tutankhamun pea extract or a fraction thereof as an effective ingredient.

The term "quasi-drug" used in the present invention refers to products that are used for the purpose of diagnosing, treating, improving, alleviating, managing or preventing diseases of humans or animals, and have a milder effect than a pharmaceutical product. For example, according to the Pharmaceutical Affairs Act, quasi-drugs are products other than those used for the purpose of pharmaceutical products, and include products used for the treatment or prevention of diseases of humans or animals, products that have a mild effect on the human body or do not act directly, etc.

The above pharmaceutical composition of the present invention can be manufactured as a product selected from the group consisting of body cleanser, foam, soap, mask, ointment, cream, lotion, essence, and spray, but is not limited thereto.

In addition, the present invention provides a health functional food composition for skin regeneration, skin elasticity improvement, prevention or improvement of skin wrinkles, prevention or improvement of skin aging, improvement of skin inflammation, skin moisturizing and/or skin whitening, containing a Tutankhamun pea extract or a fraction thereof as an effective ingredient.

When the health functional food composition of the present invention is used as a food additive, the composition may be added as is or used together with other foods or food ingredients, and may be used appropriately according to a conventional method.

The types of the above foods are not particularly limited and include all foods in the conventional sense. Non-limiting examples of foods to which the above substances can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea, drinks, alcoholic beverages, and vitamin complexes.

If the health functional food composition of the present invention is a beverage composition, it may contain various flavoring agents or natural carbohydrates as additional ingredients, like a typical beverage. Non-limiting examples of the natural carbohydrates include monosaccharides such as glucose and fructose; disaccharides such as maltose and sucrose; natural sweeteners such as dextrin and cyclodextrin; and synthetic sweeteners such as saccharin and aspartame. The ratio of the additional ingredients to be added may be appropriately determined by a person skilled in the art.

In addition to the above, the health functional food composition of the present invention may contain various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH regulators, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, etc. In addition, the health functional food composition of the present invention may contain fruit pulp for producing natural fruit juice, fruit drinks, or vegetable drinks. These ingredients may be used independently or in combination of two or more. The ratio of these additives may also be appropriately selected by those skilled in the art.

In addition, the present invention provides a pharmaceutical composition for skin regeneration, skin elasticity improvement, prevention or improvement of skin wrinkles, prevention or improvement of skin aging, improvement of skin inflammation, skin moisturizing and/or skin whitening, comprising a Tutankhamun pea extract or a fraction thereof as an effective ingredient.

In addition, the present invention provides a pharmaceutical composition for preventing or treating skin diseases, comprising a Tutankhamun pea extract or a fraction thereof as an effective ingredient.

The above skin disease includes at least one selected from the group consisting of skin wounds, skin scars, skin aging, skin inflammation and skin pigmentation.

In the pharmaceutical composition of the present invention, the description of the Tutankhamun pea extract or its fraction, and the description of skin aging, skin inflammation and skin pigmentation are the same as described above.

In the present invention, the skin wound and skin scar refer to all diseases that can be treated by increasing the total amount of collagen in skin tissue. Non-limiting examples of the skin wound and skin scar include abrasions and scars caused by abrasions.

In the present invention, the skin inflammatory disease refers to any disease caused by immune stimulation. Non-limiting examples of the skin inflammatory disease include atopic dermatitis, eczema, seborrheic dermatitis, psoriasis, acne, etc.

In the present invention, the skin pigmentation disease refers to all diseases caused by skin pigmentation due to increased production of melanin. Non-limiting examples of the skin pigmentation disease include blemishes, freckles, spots, solar pigmentation, pigmentation after drug use, pigmentation after inflammation, and pregnancy pigmentation.

The effective content of the Tutankhamun pea extract or fraction thereof in the pharmaceutical composition of the present invention is not particularly limited.

The pharmaceutical composition of the present invention, in addition to containing the Tutankhamun pea extract or a fraction thereof as an active ingredient, may additionally contain a pharmaceutically acceptable carrier.

In the present invention, the term “pharmaceutically acceptable” means something that can be commonly used in the pharmaceutical field, without stimulating a living organism when administered, and without inhibiting the biological activity and properties of the administered compound.

The pharmaceutical composition of the present invention can be formulated together with the carrier and utilized as a food, pharmaceutical, feed additive, drinking water additive, etc.

In the present invention, the type of the carrier is not particularly limited, and any carrier commonly used in the relevant technical field can be used. Non-limiting examples of the carrier include saline solution, sterile water, Ringer's solution, buffered saline, albumin injection solution, lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, maltodextrin, glycerol, ethanol, etc. These may be used alone or in combination of two or more.

In addition, the pharmaceutical composition of the present invention may be used by adding other pharmaceutically acceptable additives such as excipients, diluents, antioxidants, buffers or bacteriostatic agents, if necessary, and may be used by additionally adding fillers, bulking agents, wetting agents, disintegrating agents, dispersing agents, surfactants, binders or lubricants.

The pharmaceutical composition of the present invention can be formulated and used in various dosage forms suitable for oral administration or parenteral administration.

Non-limiting examples of the oral administration preparations include troches, lozenges, tablets, aqueous suspensions, oily suspensions, prepared powders, granules, emulsions, hard capsules, soft capsules, syrups or elixirs.

In order to formulate the pharmaceutical composition of the present invention for oral administration, a binder such as lactose, saccharose, sorbitol, mannitol, starch, amylopectin, cellulose or gelatin; an excipient such as dicalcium phosphate; a disintegrating agent such as corn starch or sweet potato starch; a lubricant such as magnesium stearate, calcium stearate, sodium stearyl fumarate or polyethylene glycol wax; and a sweetener, an air freshener, a syrup, etc. can also be used.

In addition, in the case of capsules, in addition to the substances mentioned above, a liquid carrier such as fatty oil can be additionally used.

Non-limiting examples of the above parenteral preparations include injections, suppositories, powders for respiratory inhalation, aerosols for sprays, oral sprays, mouthwashes, toothpastes, ointments, powders for application, oils, creams, and the like.

In order to formulate the pharmaceutical composition of the present invention for parenteral administration, a sterile aqueous solution, a non-aqueous solvent, a suspension, an emulsion, a freeze-dried preparation, an external preparation, etc. can be used. As the non-aqueous solvent and suspension, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate can be used.

In addition, more specifically, when the pharmaceutical composition of the present invention is formulated as an injection, the composition of the present invention may be mixed with a stabilizer or a buffer in water to prepare a solution or suspension, which may be formulated for unit administration in an ampoule or vial. In addition, when the pharmaceutical composition of the present invention is formulated as an aerosol, a propellant or the like may be mixed with additives so that the water-dispersed concentrate or wet powder can be dispersed.

In addition, when the pharmaceutical composition of the present invention is formulated into an ointment, cream, etc., it can be formulated using animal oil, vegetable oil, wax, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide, etc. as a carrier.

The pharmaceutical composition of the present invention may preferably be formulated as a dosage form for parenteral administration, and more preferably may have a dosage form selected from the group consisting of gels, patches, sprays, ointments, pastes, lotions, liniments, pastes, and cataplasmas.

The pharmaceutically effective amount, effective dosage of the pharmaceutical composition of the present invention may vary depending on the formulation method, administration method, administration time, and/or administration route of the pharmaceutical composition, and may vary depending on various factors including the type and degree of the response to be achieved by administration of the pharmaceutical composition, the type, age, weight, general health condition, symptoms or degree of the disease, sex, diet, excretion, drugs used simultaneously or simultaneously in the subject, other components of the composition, and similar factors well known in the medical field, and a person having ordinary knowledge in the relevant technical field can easily determine and prescribe an effective dosage for the intended treatment. The pharmaceutical composition of the present invention may be administered once a day or divided into several times. Therefore, the dosage does not limit the scope of the present invention in any way.

A preferred dosage of the pharmaceutical composition of the present invention may be 1 mg/kg to 1,000 mg/kg per day.

The route and method of administration of the pharmaceutical composition of the present invention may be independent, and are not particularly limited in their methods, and any route and method of administration may be followed as long as the pharmaceutical composition can reach the target site. The pharmaceutical composition may be administered orally or parenterally.

Examples of the parenteral administration method include intravenous administration, intraperitoneal administration, intramuscular administration, transdermal administration, or subcutaneous administration, and methods of applying, spraying, or inhaling the composition to the diseased area may also be used, but are not limited thereto.

The pharmaceutical composition of the present invention can preferably be administered parenterally, and more preferably can be administered by topically applying it to an area where improvement in skin condition or prevention, improvement or treatment of a skin disease is required.

The term "prevention" used in the present invention means any act of inhibiting or delaying the onset of skin diseases including skin wounds, skin scars, skin aging, skin inflammatory diseases, skin pigmentation diseases, and skin dryness by administering the pharmaceutical composition of the present invention to a subject.

The term "treatment" as used in the present invention means all acts of improving or benefiting symptoms of skin wounds, skin scars, skin aging, skin inflammatory diseases, skin pigmentation diseases, and skin dryness by administering the pharmaceutical composition of the present invention to a subject.

In addition, the present invention relates to the Tutankhamun pea ( Pisum sativum Var. arvense (L.) Poir.) extract or a fraction thereof as an effective ingredient, a cosmetic composition, a pharmaceutical composition or a food composition is provided.

In the present invention, the term "subject" means all animals, including mammals such as rats, livestock, and humans.

In the present invention, the cosmetic composition, quasi-drug composition or food composition containing the Tutankhamun pea extract or a fraction thereof as an effective ingredient is the same as described above, and the meaning of improving skin condition is also the same as described above.

In the present invention, the description regarding the dosage, administration route, administration method, etc. of the composition is replaced with the description regarding the pharmaceutical composition of the present invention.

According to another embodiment of the present invention, Tutankhamun pea ( Pisum sativum The present invention provides a method for preventing or treating a skin disease, comprising administering to a subject a pharmaceutical composition containing an extract of var. arvense (L.) Poir.) or a fraction thereof as an active ingredient.

In the present invention, the pharmaceutical composition containing the Tutankhamun pea extract or a fraction thereof as an active ingredient is the same as described above, and the meaning of the object, skin disease, prevention, and treatment is also the same as described above.

In the present invention, the description regarding the dosage, administration route, administration method, etc. of the pharmaceutical composition is the same as that described with respect to the pharmaceutical composition of the present invention.

The composition containing the Tutankhamun pea extract or a fraction thereof of the present invention is excellent in skin regeneration, skin elasticity improvement, skin wrinkle prevention or improvement, skin aging prevention or improvement, skin inflammation improvement, skin moisturizing or skin whitening effects. Specifically, the composition of the present invention is excellent in the effects of increasing the total amount of collagen, decreasing collagenase activity, decreasing the total amount of melanin, promoting cell proliferation, scavenging free radicals, and inhibiting inflammatory reactions.

In addition, a composition containing the Tutankhamun pea extract or a fraction thereof of the present invention has an excellent effect in preventing or treating skin diseases.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, the examples according to the present invention may be modified in various different forms, and the scope of the present invention should not be construed as being limited to the examples described below. The examples of the present invention are provided as examples to help in a specific understanding of the present invention.

Preparation of Tutankhamun Pea Extract

The inventors of the present invention received Tutankhamun peas from the National Arboretum’s Usefulness Propagation Center and, to prepare a Tutankhamun pea extract, crushed Tutankhamun pea seeds, placed 10 g in a flask, and extracted by steeping for 3 days with 100 g of an extraction solvent (distilled water: methanol = 3:7). The steeped extract was filtered through a filter with a pore size of 0.2 μm to prepare a Tutankhamun pea extract.

Experimental example 1: Effect of increasing total collagen amount

The present inventors added Tutankhamun pea extract to a culture medium of human-derived fibroblasts and tested the effect of increasing the total amount of collagen at the cellular level.

Specifically, the measurement of the increased total collagen amount was quantified using a PICP EIA kit (Procollagen Type I C-Peptide Enzyme ImmunoAssay KIT). Before the experiment, the cytotoxicity was evaluated by varying the concentration (㎍/ml) of the Tutankhamun pea extract prepared as described above on human-derived fibroblasts (a method of performing the MTT test by culturing fibroblasts [Reference: Mossman T. (1983). Rapid Colorimetric Assay for Cellular Growth & Survival: application to proliferation & cytotoxicity assays. Journal of Immunological Methods 65, 55-63]), and a concentration without cytotoxicity (10 ㎍/ml or less) was selected to evaluate the degree of increase in the total collagen amount.

Each sample was added to the culture medium of human fibroblasts and cultured for 1 day. The culture medium was taken and the degree of increase in the total amount of collagen at each concentration was measured using a spectrophotometer at 450 nm using a PICP EIA kit. In order to compare the effect, the degree of increase in the total amount of collagen was measured in the same way for the culture medium of fibroblasts to which nothing was added (control group) and the sample to which vitamin C was added to a final concentration of 52.8 ㎍/ml. The total amount of collagen was measured as UV absorbance, and the increase rate of the total amount of collagen was calculated as the ratio of the total amount of collagen relative to the control group, and the results are shown in Table 1 below.

Sample Absorbance average Collagen Increase Rate (%) Tutankhamun Pea Extract (10 μg/ml) 635 139 Tutankhamun Pea Extract (5 μg/ml) 583 128 Control group (no additives) 456 - Vitamin C (52.8 μg/ml) 552 121

* Effect of Tutankhamun Pea Extract on increasing total collagen content (repetition = 3)

As can be seen from the results in Table 1 above, the Tutankhamun pea extract has the effect of promoting collagen synthesis and increasing the total amount of collagen, and in particular, it was confirmed to increase the total amount of collagen to a degree superior to that of vitamin C, which is known to have the ability to increase the total amount of collagen.

Experimental example 2: Collagenase Active inhibitory effect

The present inventors confirmed the collagenase activity inhibitory effect of Tutankhamun pea extract as follows.

Human normal skin cells, fibroblasts, were seeded at 2.5 × 10 ⁴ cells per well in a 24-well microplate and cultured for 24 hours under conditions of 10% serum DMEM medium and 37°C. The 10% serum DMEM medium was removed, washed once with phosphate buffered solution, and then cultured for an additional 30 minutes in serum-free DMEM medium supplemented with Tutankhamun pea extract and in serum-free DMEM medium without Tutankhamun pea extract as a control.

After 30 minutes of sample treatment, the cells were stimulated with 50 ng/ml of tumor necrosis factor-α (TNF-α), a substance known to produce collagenase MMP-1, and then cultured for 24 hours. At this time, the TNF-α untreated group and the treated group were divided into the untreated group and the treated group among the control group that did not contain the Tutankhamun pea extract and the TNF-α treated group.

The supernatant from each well was collected, and the amount of newly synthesized MMP-1 (ng/㎖) was measured using an MMP-1 assay kit (Amersham, USA), and the collagenase activity inhibition rate was calculated as the MMP-1 production inhibition rate (%) according to the following mathematical formula 1, and the results are as shown in Table 2 below.

The amount of MMP-1 in the negative control group refers to the amount of MMP-1 in the TNF-free group, the amount of MMP-1 in the positive control group refers to the amount of MMP-1 in the TNF-treated group, and the amount of MMP-1 in the experimental group refers to the groups to which the Tutankhamun pea extract was added at each concentration.

[Mathematical formula 1]

MMP-1 production inhibition rate (%) =

(X = amount of MMP-1 in the experimental group, Y = amount of MMP-1 in the positive control group, Z = amount of MMP-1 in the negative control group)

Sample Amount of MMP-1 (ng/㎖) Inhibition rate (%) Tutankhamun Pea Extract (10 μg/ml) 11.38 54.27 Tutankhamun Pea Extract (5 μg/ml) 14.13 28.76 Negative control (no TNF treatment) 6.45 - Positive control (TNF treatment) 17.23 -

* Collagenase activity inhibition effect of Tutankhamun pea extract (repetition number = 3)

As can be seen from the results in Table 2 above, it can be seen that the Tutankhamun pea extract has excellent inhibitory ability on collagenase, MMP-1.

Experimental example 3: Whitening effect - Confirmed effect of reducing total melanin amount

The present inventors added Tutankhamun pea extract to a culture medium of mouse melanoma cells (B-16 mouse melanoma cells) and tested the whitening effect at the cellular level.

Tutankhamun pea extract was added to the culture medium at final concentrations of 5 ㎍/ml and 10 ㎍/ml, and arbutin, as a control, was added to the medium at 100 ㎍/ml, and treated to B-16 melanoma cells, respectively, and cultured for 3 days.

Afterwards, the cells were treated with trypsin to detach them from the culture vessel, centrifuged, and melanin was extracted. The detached cells were boiled in 1 ml of sodium hydroxide solution (1N concentration) for 10 minutes to dissolve the melanin, and the amount of melanin produced was measured by measuring the absorbance at 400 nanometers (nm) using a spectrophotometer.

The above melanin amount was measured by a method expressing the absorbance per unit cell number (10 ⁶ cell number), and the inhibition rate (%) relative to the total melanin amount in the control group was calculated, and the results are shown in Table 3.

Sample Total Melanin Amount (Abs) Inhibition rate (%) Tutankhamun Pea Extract (10 μg/ml) 0.062 24.39 Tutankhamun Pea Extract (5 μg/ml) 0.075 8.54 Control group (no additives) 0.082 - Arbutin (100 μg/ml) 0.042 48.78

* Effect of Tutankhamun Pea Extract on Reduction of Total Melanin Amount (Repetition = 3)

As can be seen from the results in Table 3 above, the Tutankhamun pea extract showed the effect of inhibiting melanin synthesis even at low concentrations, thereby reducing the total amount of melanin.

Experimental example 4: CCK -8 Confirmation of skin cell proliferation promotion effect through evaluation method

The present inventors added Tutankhamun pea extract to a culture medium of human-derived fibroblasts and tested the cell proliferation promoting effect.

Human fibroblasts were seeded in 96-well microplates at a density of 1 × 10 ⁴ cells per well and cultured for 24 hours under conditions of 10% serum DMEM medium at 37℃. The 10% serum DMEM medium was removed, and the cells were washed once with phosphate buffered solution. After culturing for 2 days in serum-free DMEM medium supplemented with Tutankhamun pea extract, the cultured cells were evaluated using CCK-8 (Cell counting kit-8). The CCK-8 evaluation is an analysis method that indirectly indicates the density of living cells by measuring the absorbance of formazan produced by dehydrogenase in the intracellular electron transport chain decomposing tetrazolium salt. In order to compare the effect, the culture medium of fibroblasts without any addition (control group) and the sample with the addition of 5% serum culture medium (positive control) were evaluated using the same method.

After treating cells with CCK-8 solution (1/10 of the medium volume treated) at 37℃ for 1 hour and 30 minutes, the absorbance at a wavelength of 450 nm was measured spectrophotometrically. The skin cell proliferation promoting effect of Tutankhamun pea extract was calculated as the proliferation rate (%) according to the following mathematical equation 2 from the measured absorbance value compared to the control group, and the value is shown in Table 4 below.

[Mathematical formula 2]

Cell count (%) = (absorbance of sample treatment group/absorbance of control group) × 100

Sample density Cell proliferation rate (%) compared to control group Tutankhamun Pea Extract 1 ㎍/ml 123 Tutankhamun Pea Extract 0.1 μg/ml 119 Control group (serum-free medium) - 100 Positive control group (5% serum medium) - 150

* Cell proliferation promotion effect (repetition number = 3)

As can be seen from the results in Table 4 above, the Tutankhamun pea extract was confirmed to promote the proliferation of skin cells compared to the untreated control group, and although it is a simple extract of the plant, it showed excellent cell proliferation promotion efficacy even at low concentrations.

Experimental example 5: Antioxidant effect - Free radical scavenging rate

The inventors of the present invention measured free radical scavenging activity to confirm the antioxidant activity of the Tutankhamun pea extract. Free radical scavenging activity was measured using DPPH. DPPH is a relatively stable free radical, and when it exists in a radical state, it shows maximum absorbance at 517 nm, and when the radical is scavenged, it loses absorbance. DPPH from Sigma (Sigma Co., Ltd, USA) was used, and was dissolved in methyl alcohol at a concentration of 0.15 mM.

First, 100 ㎕ of the above Tutankhamun pea extract and ascorbic acid were added to each well of a 96-well plate. 100 ㎕ of DPPH solution was added here, and the mixture was left at room temperature for 30 minutes. The absorbance at 517 nm was measured using a Micro plate reader (BioTek EL-340). At this time, methyl alcohol was added instead of the above sample as a control group. The free radical scavenging activity was calculated as the half maximal inhibitory concentration (inhibition median value), IC ₅₀ , and the results are shown in Table 5 below. IC ₅₀ is the concentration of ascorbic acid and Tutankhamun pea extract required to remove 50% of the free radicals of the non-added control group, and is a general method of expressing free radical scavenging activity.

division Tutankhamun Pea Extract Ascorbic acid IC ₅₀ (%) 0.0169 0.0007

As shown in Table 5 above, it was found that the Tutankhamun pea extract of the present invention has excellent free radical scavenging ability.

Experimental example 6: Anti-inflammatory effect - NO production inhibition effect

In order to confirm the anti-inflammatory effect and skin trouble alleviation effect of Tutankhamun pea extract, the present inventors conducted a nitric oxide (NO) formation inhibition experiment using the GRIESS method using RAW264.7 cell line (ATCC number: CRL-2278).

Specifically, mouse macrophage RAW264.7 cells were passaged several times, placed in a 24-well plate at a density of 3 × 10 ⁵ cells per well, and cultured for 24 hours.

Next, the cell medium was replaced with the diluted Tutankhamun pea extract at the concentration shown in Table 7 below. At this time, L-NMMA (L-NG-Monomethylarginine), an NO production inhibitor, was treated together as a positive control and cultured for 30 minutes, and 10 ㎍ of LPS (Lipopolysaccharide) was treated as a stimulus and cultured for 24 hours. 100 ㎕ of the supernatant was transferred to a 96-well plate, 100 ㎕ of GRIESS solution was added, and the reaction was performed at room temperature for 10 minutes. The absorbance at 540 nm was measured to determine the NO inhibitory effect of the Tutankhamun pea extract, and the results are shown in Table 6 below.

Sample NO production inhibition rate (%) Tutankhamun Pea Extract (50 μg/ml) 41.89 Tutankhamun Pea Extract (5 μg/ml) 22.33 L-NMMA (10 ug/ml) 39.54

As shown in Table 6 above, it was confirmed that the Tutankhamun pea extract used in the cosmetic composition of the present invention exhibited excellent NO production inhibition efficacy even at low concentrations, although it was a simple extract.

Claims (7)

A cosmetic composition for skin regeneration, improvement of skin inflammation or skin whitening, comprising a Tutankhamun pea extract or a fraction thereof as an active ingredient. A pharmaceutical composition for skin regeneration, improvement of skin inflammation or skin whitening, comprising Tutankhamun pea extract or a fraction thereof as an active ingredient. A health functional food composition for skin regeneration, improvement of skin inflammation or skin whitening, comprising Tutankhamun pea extract or a fraction thereof as an effective ingredient. Contains Tutankhamun pea extract or a fraction thereof as an active ingredient,
A pharmaceutical composition for preventing or treating any one or more skin diseases selected from the group consisting of skin wounds, skin scars, skin inflammation and skin pigmentation. delete In any one of claims 1 to 4,
A composition characterized in that the above Tutankhamun pea extract is extracted with at least one solvent selected from the group consisting of water, C ₁ to C ₄ alcohol, 1,3-butylene glycol, and ethyl acetate. In any one of claims 1 to 4,
A composition characterized in that the Tutankhamun pea extract or a fraction thereof is contained in an amount of 0.0001 to 50 wt% based on the total weight of the composition.