KR102020817B1 - Composition for skin cell regeneration, anti-wrinkle, antioxidant and anti-imflamation - Google Patents

Abstract

The present invention relates to a composition for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory. Camperol-7-O-beta-D-glucopyranoside according to the present invention promotes collagen synthesis of fibroblasts of the skin, thereby inhibiting skin regeneration, wrinkle improvement, and free radical production, thereby inhibiting antioxidant and NO production. By exhibiting an anti-inflammatory effect, it can be used as a pharmaceutical composition, external skin preparation, cosmetic composition or health food.

Description

Composition for skin cell regeneration, anti-wrinkle, antioxidant and anti-imflamation

The present invention relates to a composition for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory.

Collagen is a major substrate protein produced by skin fibroblasts and is present in extracellular epilepsy.Its important functions include mechanical firmness of skin, resistance of connective tissue and tissue binding, support of cell adhesion, cell division and differentiation. Induction of growth or wound healing) is known. This collagen is reduced by age and light aging by ultraviolet irradiation, which is known to be closely related to the wrinkle formation of the skin. In addition, in recent years, extensive research on skin aging has revealed an important function of collagen in skin.

Active ingredients are known that promote collagen synthesis and exhibit wrinkle improvement. For example, retinoic acid, transforming growth factor (TGF) [non-patent document 1], protein derived from animal placenta [patent document 1], betulinic acid (betulinic acid) [patent document 2], chlorella extract [ Patent Documents 3 and 4 are known as collagen synthesis promoting substances. However, the active ingredients are limited in the amount of use due to safety problems such as irritation and redness when applying the skin, or there is a problem in that the effect of improving the skin function is not expected by substantially promoting collagen synthesis of the skin.

On the other hand, free radicals introduced from outside the living body or generated in the living body cause many problems such as promoting aging of the living body or generating cancer. Therefore, many researches and developments on antioxidants that inhibit oxidation by active oxygen have been made. Antioxidants are widely distributed in copper and plant systems, and many phenolic compounds, flavonoids, tocopherols, vitamin C, and selenium are known in fruits and vegetables. However, the antioxidants present in nature are not expected to have a substantial enough effect on skin application. Therefore, many synthetic antioxidants having excellent antioxidant power and low cost are used, but their use is limited due to safety concerns such as human side effects.

In addition, inflammation is an immune response of the human body in response to a wound or disease, and oxidative stress such as ultraviolet rays, free radicals, and free radicals activates inflammatory factors to cause various diseases and aging of the skin. Vascular activating polypeptides such as kinin, plasmin, and complement act on vasodilation, contraction and chemotaxis, and other lymphokines such as interleukin-6 (IL-6). Phosphorus and arachidonic acid are responsible for the inflammatory response. Arachidonic acid is metabolized to prostaglandin and leukotriene, which are inflammatory mediators, through two pathways, cyclooxygenase or lipooxygenase, to mediate various inflammatory reactions.

On the other hand, the anti-inflammatory agent that acts to remove the inflammatory source, reduce the biological response and symptoms to eliminate inflammation. Materials used for anti-inflammatory purposes so far are nonsteroidal flufenamic acid, ibuprofen, benzydamine, indomethacin, and indonethacin, and prednisolone as a steroid system. , Dexamethasone, and the like. In addition, allantoin, azene, hydrocortisone, etc. are known to be effective in anti-inflammatory, but these substances have a limited amount of use as a matter of safety to the skin, or a problem that can not be expected to substantially reduce the effect of the inflammation. There is this.

Therefore, the development of ingredients that are safe for the living body, the active ingredient is stable, and above all, the skin regeneration, anti-wrinkle, antioxidant and anti-inflammatory activity is superior to the existing skin regeneration, anti-wrinkle, antioxidant and anti-inflammatory substances. This is desperately desired.

1. Japanese Patent Application Laid-Open No. 8-231370 2. Japanese Patent Application Laid-Open No. 8-208424 3. JP 9-40523 A 4. Japanese Patent Application Laid-Open No. 10-36283

Cardinale G. et al, Adv. Enzymol., 41, p. 425, 1974

Accordingly, the present inventors have found that kaempferol-7-O-beta-D-glucoside promotes collagen synthesis and inhibits collagenase activity of skin fibroblasts. By promoting the regeneration of the skin and improve wrinkles, by scavenging free radicals to exhibit an antioxidant effect, inhibiting the production of nitric oxide (NO) to confirm the anti-inflammatory effect has been completed the present invention.

Accordingly, an object of the present invention to provide a composition for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory comprising a compound represented by the following formula (1) as an active ingredient.

[Formula 1]

As a means for solving the above problems, the present invention provides a pharmaceutical composition for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory comprising a compound represented by the following formula (1) as an active ingredient.

 [Formula 1]

As another means for solving the above problems, the present invention provides a skin external preparation for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory comprising a compound represented by the following formula (1) as an active ingredient.

[Formula 1]

As another means for solving the above problems, the present invention provides a cosmetic composition for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory comprising a compound represented by the following formula (1) as an active ingredient.

[Formula 1]

As another means for solving the above problems, the present invention provides a health food for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory comprising a compound represented by the following formula (1) as an active ingredient.

[Formula 1]

Camperol-7-O-beta-D-glucopyranoside according to the present invention promotes collagen synthesis of skin fibroblasts and inhibits collagenase activity, thereby promoting skin regeneration, improving wrinkles, and free radicals. It can be used in medicines, cosmetics or health foods by eliminating anti-oxidant effect, suppressing NO production involved in inflammatory reaction, and anti-inflammatory effect.

Hereinafter, the structure of this invention is demonstrated concretely.

Skin regeneration, wrinkle improvement, anti-oxidation and anti-inflammatory ingredients have excellent effect when applied to the actual skin, showing high activity of skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory activity at low concentration, and the ability to penetrate and absorb skin Excellent, low volatility to stay for a sufficient time to show skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory effects, keep the active ingredient stable on the composition or skin, easy to formulate into medicine or cosmetic, It is desirable to be safe for the skin. However, among the known components, components that satisfy all of the above properties are rare. For example, some skin rejuvenation, wrinkle improvement, antioxidant and anti-inflammatory components have excellent skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory activity even at low concentrations in vitro, but are less effective in penetrating and absorbing the skin. Difficult to apply Other active ingredients are less hydrophilic and difficult to formulate in medicine or cosmetics. In addition, some skin regeneration, anti-wrinkle, antioxidant and anti-inflammatory ingredients may decompose or transform into other compounds when exposed to heat, light, or oxygen, and lose their effects before application to the skin.

As can be seen in the following examples, camphorol-7-O-beta-D-glucopyranoside (kaempferol-7-O-beta-D-glucoside) is excellent collagen synthesis promoting effect, antioxidant effect at low concentrations And because it shows an anti-inflammatory effect, it can be used as an active ingredient in medicines, cosmetics, health foods for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory.

Therefore, the present invention therefore provides a pharmaceutical composition for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory comprising the compound represented by the following formula (1) as an active ingredient.

[Formula 1]

The compound name of the compound of Formula 1 is 3,5-Dihydroxy-2- (4-hydroxyphenyl) -7-[(2S, 3R, 4S, 5S, 6R) -3,4,5-trihydroxy-6- (hydroxymethyl) oxan-2-yl] oxychromen-4-one; It is called Kaempferol-7-O-glucoside, also known as camphorol-7-O-beta-D-glucopyranoside (kaempferol-7-O-beta-D-glucoside).

The compound of Formula 1 may be synthesized or used a commercially available compound.

The pharmaceutical composition for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory of the present invention may include a pharmaceutically acceptable salt of the compound of Formula 1.

The pharmaceutically acceptable salt of the compound of Formula 1 may be an acid addition salt formed using an organic acid or an inorganic acid, and the organic acid may be, for example, formic acid, acetic acid, propionic acid, lactic acid, butyric acid, isobutyric acid or trifluoroacetic acid. , Malic acid, maleic acid, malonic acid, fumaric acid, succinic acid, succinic acid monoamide, glutamic acid, tartaric acid, oxalic acid, citric acid, glycolic acid, glucuronic acid, ascorbic acid, benzoic acid, phthalic acid, salicylic acid, anthranilic acid, dichloroacetic acid, aminooxyacetic acid And benzenesulfonic acid, p-toluenesulfonic acid and methanesulfonic acid salts and inorganic acids include, for example, hydrochloric acid, bromic acid, sulfuric acid, phosphoric acid, nitric acid, carbonic acid and boric acid salts. It may preferably be in the form of hydrochloride or acetate, more preferably in the form of hydrochloride.

The above-mentioned acid addition salts can be a) directly mixed with the compound of formula 1 and an acid, b) dissolved and mixed with one of them in a solvent or a hydrous solvent, or c) a solvent with a solvent or an aqueous solvent. It is prepared by the general salt preparation method of placing in an acid in the mixture and mixing them.

In addition to the above, salts that can be additionally salted are Gabar salt, Gabapentin salt, Pregabalin salt, Nicotinate, Adipate salt, Hemimalonate, Cysteine salt, Acetylcysteine salt, Methionine salt, Arginine salt, Lysine salt, Ornithine salt, Aspartate.

In addition, when using the compound of Formula 1 of the present invention as a medicine, it may further contain one or more active ingredients exhibiting the same or similar functions. For example, it may include known skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory components. Inclusion of additional skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory components may further enhance skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory effects of the compositions of the present invention. When the ingredient is added, skin safety, ease of formulation, and stability of the active ingredients may be considered. In one embodiment of the invention, the composition is a skin regeneration or wrinkle improvement component known in the art, Retinoic acid, TGF, protein from animal placenta, betulinic acid and chlorella extract; Antioxidant ingredients known in the art include tocopherol, selenium, vitamin C and phenolic compounds; As anti-inflammatory components known in the art, COX-2 inhibitors, prednisolone and allantoin; And one or two or more components selected from the group consisting of derivatives and various plant extracts thereof may be further included. Additional ingredients may be included from 0.0001% to 10% by weight relative to the total composition weight, and the content range may be adjusted according to requirements such as skin safety, ease of formulating the compound of Formula 1, and the like. .

In addition, the pharmaceutical composition for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory of the present invention may further comprise a pharmaceutically acceptable carrier.

Pharmaceutically acceptable carriers may contain various components such as buffers, sterile water for injection, general saline or phosphate buffered saline, sucrose, histidine, salts, polysorbates and the like.

The pharmaceutical composition of the present invention may be administered orally or parenterally, and may be administered in the form of a general pharmaceutical preparation, for example, in various dosage forms, orally and parenterally, during clinical administration. , Diluents or excipients such as extenders, binders, wetting agents, disintegrants, surfactants and the like.

Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and such solid preparations include at least one excipient in the pharmaceutical composition of the present invention, for example, starch, calcium carbonate, It may be prepared by mixing sucrose or lactose, gelatin and the like.

In addition to simple excipients, lubricants such as magnesium styrate talc are also used. Oral liquid preparations include suspensions, solvents, emulsions, and syrups, and may include various excipients, such as wetting agents, sweeteners, fragrances, and preservatives, in addition to commonly used simple diluents such as water and liquid paraffin. .

Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, suppositories. As the non-aqueous solvent and the suspension solvent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like can be used. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin and the like can be used.

In the present invention, the "skin regeneration effect" refers to the recovery of skin tissue against damage caused by external and internal causes of the skin. The damage caused by the external causes may include ultraviolet rays, external pollutants, wounds, traumas, and the like, and the damage caused by the internal causes may include stress.

In the present invention, the "wrinkle improvement effect" refers to inhibiting or inhibiting the generation of wrinkles on the skin, or alleviating wrinkles already generated.

In the present invention, the term "antioxidative effect" refers to a cell caused by free radicals or reactive oxygen species (ROS) that are highly reactive according to oxidative stress caused by intracellular metabolism or ultraviolet light. Refers to the inhibition of oxidation, including the removal of free radicals or reactive oxygen species, thereby reducing damage to cells.

In the present invention, the "anti-inflammatory effect" refers to inhibiting inflammation, which is one of the defense responses of biological tissues to any stimulus, and includes three types of tissue degeneration, circulatory disorder and exudation, and tissue proliferation. To say complex lesions. More specifically, inflammation is part of innate immunity and, as in other animals, human innate immunity recognizes patterns of cell surfaces that are specific to pathogens. Phagocytes recognize cells with such surfaces as nonmagnetic devices and attack pathogens. If the pathogen breaks through the body's physical barrier, an inflammatory reaction occurs. Inflammatory reactions are nonspecific defenses that create a hostile environment against invading microbes. In inflammatory reactions, when wounded or an external infectious agent enters the body, white blood cells in the early stages of immune response rush to express cytokines. Therefore, the expression level of cytokines in cells is an indicator of inflammatory response activation. Examples of skin diseases related to inflammation include atopic dermatitis, psoriasis, radiation, chemicals, burns, etc., erythematous disease, acid burns, bullous skin disease, mammary gland-type diseases, allergies, itching, seborrhea, eczema, Vulgaris, polymorphic exudative erythema, nodular erythema, glansitis, vulvitis, inflammatory hair loss such as alopecia areata, cutaneous T-cell lymphoma, and the like.

The pharmaceutical composition of the present invention may provide desirable skin regeneration effect, wrinkle improvement effect, antioxidant effect and anti-inflammatory effect when the effective amount of the compound of Formula 1 is included. In the present invention, the term "effective amount" means an amount of a compound capable of promoting regeneration of damaged skin, improving wrinkles, inhibiting or alleviating oxidation of cells, or inhibiting inflammation. The effective amount of the compound of formula 1 included in the composition of the present invention will vary depending on the form in which the composition is commercialized, how the compound is applied to the skin, the time it stays on the skin, and the like. For example, when the composition is manufactured as a medicine, it may include the compound of Formula 1 in a higher concentration than when manufactured as a cosmetic that is routinely applied to the skin. Therefore, the daily dosage is 0.1 to 100 mg / kg, preferably 30 to 80 mg / kg, more preferably 50 to 60 mg / kg, 1 to 1 day, based on the amount of the compound of Formula 1 It may be administered six times.

The pharmaceutical composition of the present invention may be used alone or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy and biological response modifiers.

The present invention can also be provided in the formulation of the external preparation for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory including the compound of Formula 1 as an active ingredient.

When the compound of Formula 1 is used as an external preparation for skin, it is further added to fatty substances, organic solvents, solubilizers, thickening and gelling agents, emollients, antioxidants, suspending agents, stabilizers, foaming agents, fragrances, surfactants. Common in water, ionic or nonionic emulsifiers, fillers, metal ion sequestrants and chelating agents, preservatives, vitamins, blockers, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic active agents, lipid vesicles or external preparations for the skin It may contain adjuvants commonly used in the field of dermatology, such as any other ingredients used as. The ingredients may also be introduced in amounts generally used in the field of dermatology.

When the compound of Formula 1 is provided as an external preparation for skin, it is not limited thereto, and may have a formulation such as an ointment, a patch, a gel, a cream, or a spray.

The present invention can also be provided in the formulation of a cosmetic for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory comprising the compound of Formula 1 as an active ingredient.

When the compound of Formula 1 is used as a cosmetic, the cosmetic prepared by containing the compound of Formula 1 as an active ingredient may be prepared in the form of a general emulsion formulation and solubilized formulation. For example, lotions such as flexible lotions or nourishing lotions, emulsions such as facial lotions, body lotions, creams such as nourishing creams, moisture creams, eye creams, essences, cosmetic ointments, sprays, gels, packs, sunscreens, makeup bases, liquids Formulations such as foundations, powders, cleansing creams, cleansing lotions, makeup removers such as cleansing oils, cleansing foams, soaps, body washes and the like.

In addition, the cosmetics are in addition to the compound of Formula 1, fatty substances, organic solvents, solubilizers, thickening and gelling agents, emollients, antioxidants, suspending agents, stabilizers, foaming agents, fragrances, surfactants, water Commonly used in ionic or nonionic emulsifiers, fillers, metal ion sequestrants and chelating agents, preservatives, vitamins, blockers, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic active agents, lipid vesicles or cosmetics It may contain adjuvants conventionally used in the cosmetic field, such as any other ingredient.

The compound of Formula 1 may be a relatively high concentration of the compound of Formula 1 in the case of a wash-off type cosmetic such as a make-up remover, a detergent, etc., in which the active ingredient stays on the skin within a short time when it is commercialized. It may include. On the other hand, in the case of a leave-on type cosmetic such as a lotion, an emulsion, a cream, an essence, etc., in which the active ingredient stays on the skin for a long time, the compound of Chemical Formula 1 has a lower concentration than the wash-off type cosmetic. It may be included. Although not limited thereto, in one embodiment of the present invention, the composition may include the compound of Formula 1 in an amount of 0.0001% to 10% by weight (preferably 0.0001% to 1% by weight) based on the total weight of the composition. Can be. When the composition of the present invention contains less than 0.0001% by weight of the compound of Formula 1, sufficient skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory effects cannot be expected, and when it contains more than 10% by weight, allergy and the like This is to prevent the reaction because it may occur or there may be a problem in skin safety.

The present invention also relates to a health food for skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory comprising the compound of Formula 1.

In the present specification, "health food" is a food prepared by adding the compound of Formula 1 to food materials, such as beverages, teas, spices, gums, confectionery, encapsulated, powdered, suspensions, etc. It means to bring a certain effect, but unlike the general medicine has the advantage that there are no side effects that can occur when taking a long-term use of the drug as a raw material.

Since the health food of the present invention thus obtained can be consumed on a daily basis, high skin regeneration, wrinkle improvement, antioxidant and anti-inflammatory effects can be expected and are very useful.

When the compound of Formula 1 is used as a food additive, the compound of Formula 1 may be added as it is or used with other foods or food ingredients, and may be appropriately used according to a conventional method. The mixed amount of the active ingredient can be suitably determined according to the purpose of use (prevention, health or therapeutic treatment). Generally, in the preparation of food or beverages, the composition of the present invention is added in an amount of up to 15 parts by weight, preferably up to 10 parts by weight based on the raw materials. However, in the case of long-term intake for health and hygiene or health control, the amount may be below the above range, and the active ingredient may be used in an amount above the above range because there is no problem in terms of safety. .

There is no particular limitation on the kind of food. Examples of the food to which the substance may be added include meat, sausages, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gums, dairy products including ice cream, various soups, beverages, teas, drinks, Alcoholic beverages and vitamin complexes, and the like and include all of the health foods in the conventional sense.

The health beverage composition of the present invention may contain various flavors or natural carbohydrates, etc. as additional components, as in the general beverage. The above-mentioned natural carbohydrates are glucose, monosaccharides such as fructose, disaccharides such as maltose and sucrose, and polysaccharides such as dextrin and cyclodextrin, sugar alcohols such as xylitol, sorbitol and erythritol. As the sweetening agent, natural sweetening agents such as tautin and stevia extract, synthetic sweetening agents such as saccharin and aspartame, and the like can be used. The proportion of said natural carbohydrate is generally about 0.01 to 0.04 g, preferably about 0.02 to 0.03 g per 100 mL of the composition of the present invention.

In addition to the above, the health food of the present invention includes various nutrients, vitamins, electrolytes, flavors, coloring agents, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloid thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols. And carbonation agents used in carbonated beverages. In addition, the health food of the present invention may contain a flesh for preparing natural fruit juice, fruit juice drink and vegetable drink. These components can be used independently or in combination. The proportion of such additives is not critical but is generally selected in the range of 0.01 to 0.1 parts by weight per 100 parts by weight of the composition of the present invention.

Hereinafter, the present invention will be described in detail by way of examples. However, the following examples are merely to illustrate the invention, but the content of the present invention is not limited to the following examples.

Reference Example  One: Camphorol -7-O-beta-D-glucopyranoside ( kaempferol -7-O- beta -D-glucoside) Substance Information

Name: 3,5-Dihydroxy-2- (4-hydroxyphenyl) -7-[(2S, 3R, 4S, 5S, 6R) -3,4,5-trihydroxy-6- (hydroxymethyl) oxan-2-yl] oxychromen-4-one; Kaempferol-7-O-glucoside

CAS No .: 16290-07-6

Where to buy: Tauto Biotech Co., Ltd.

Plant of Origin: Hosta plantaginea ( Lam .) Aschers.

Example  1: collagen synthesis effect

The substances shown in Table 1 below were added to the culture solution of human-derived fibroblasts to test collagen synthesis promoting effects at the cellular level.

The synthesis of collagen was quantified using a PICP EIA kit (Procollagen Type I C-Peptide Enzyme ImmunoAssay KIT). Evaluation of cytotoxicity at concentrations of 10 ppm, 1 ppm, 0.1 ppm, 0.01 ppm, and 0.001 ppm in human-derived fibroblasts before the experiment (method of culturing fibroblasts for MTT test [Reference: Mossman T. (1983) Rapid Colorimetric Assay for Cellular Growth & Survival: application to proliferation & cytotoxicity assays.Journal of Immunological Methods 65, 55-63]).

In the experiment, the final concentrations of camphorol-7-O-beta-D-glucopyranoside were 1 ppm and 10 ppm, and each sample was added to the culture medium of human fibroblasts and cultured for 1 day. The degree of collagen synthesis at each concentration was measured at 450 nm by using a spectrophotometer. For comparison of the effects, the degree of collagen synthesis was measured by the same method for the culture medium (control) of the fibroblasts to which nothing was added and the sample to which the vitamin C was added at a final concentration of 52.8 μg / ml. Collagen production was measured as UV absorbance, the collagen production increase rate was calculated as the ratio of collagen production relative to the control, the results are summarized in Table 1 below.

Collagen Biosynthesis Effect of Camperol-7-O-beta-D-Glucopyranoside (Repeat Number = 3) sample Absorbance average Collagen Growth Rate (%) Control group (no addition) 845 - Vitamin C (52.8 μg / ml) 1056 125 10 ppm of a compound of formula 1 1175 139 1 ppm of a compound of Formula 1 1039 123

As can be seen from the results in Table 1, camphorol-7-O-beta-D-glucopyranoside generally exhibits better collagen synthesis effect at a lower concentration than when vitamin C is known to be capable of collagen synthesis. It was.

Example  2: Collagenase  Active inhibitory effect

It was confirmed as follows the collagenase activity inhibitory effect on the material of Table 2.

Evaluation of cytotoxicity at concentrations of 10 ppm, 1 ppm, 0.1 ppm, 0.01 ppm, and 0.001 ppm in human-derived fibroblasts before the experiment (method of culturing fibroblasts for MTT test [Reference: Mossman T. (1983) Rapid Colorimetric Assay for Cellular Growth & Survival: application to proliferation & cytotoxicity assays.Journal of Immunological Methods 65, 55-63]).

Fibroblasts, which are human normal skin cells, were seeded in a 24-well microplate at 2.5 × 10 ⁴ cells per well, incubated for 24 hours at 10% serum DMEM medium and 37 ° C., and then 10% serum DMEM medium was removed. Washed once with phosphate buffer solution, and then camphorol-7-O-beta-D-glucopyranoside in the serum-free DMEM medium and camphor with the addition of camphorol-7-O-beta-D-glucopyranoside. It was further incubated for 30 minutes in serum-free DMEM medium containing no.

Thirty minutes after the sample was incubated with 50ng / ml of TNF-α (tumor necrosis factor-alpha), a substance known to produce MMP-1 (Matrix metalloproteinase-1). At this time, the TNF-α untreated group and the treated group were treated with TNF-α untreated group and TNF-α not treated with camphorol-7-O-beta-D-glucopyranoside. It was set as a group.

The supernatant of each well was collected to measure the amount (ng / ml) of newly synthesized MMP-1 using an MMP-1 assay kit (Amersham, USA), and the collagenase activity inhibition rate was determined according to Equation 1 below. -1 generation inhibition rate (%) was calculated, the results are shown in Table 2 below.

The amount of MMP-1 in the control group means the amount of MMP-1 in the TNF treatment group, and the amount of MMP-1 in the experimental group refers to the group to which the substance is added at each concentration.

Inhibitory Effect of Camperol-7-O-beta-D-Glucopyranoside on Collagenase Activity (Repeat Number = 4) sample Amount of MMP-1 (ng / mL) % Inhibition Camperol-7-O-beta-D-glucopyranoside 10 ppm 6.1883 66.5545 Camperol-7-O-beta-D-glucopyranoside 1 ppm 17.7778 6.0143 Negative Control (TNF No Processing) 6.3831 - Positive control group (TNF treatment) 18.5452 -

As can be seen from the results in Table 2, when treated with camphorol-7-O-beta-D-glucopyranoside and TNF, camphorol-7-O-beta-D-glucopyranoside was not treated. It can be seen that the amount of MMP-1 production decreased compared to the TNF-treated positive control. That is, camphorol-7-O-beta-D-glucopyranoside exhibited excellent collagenase activity inhibitory effect.

Example  3: antioxidant effect Free radical  Erasure rate

Free radical scavenging activity was measured to confirm the antioxidant activity of camphorol-7-O-beta-D-glucopyranoside. Free radical scavenging activity was measured using DPPH (2,2-diphenyl-1-picrylhydrazyl). DPPH was purchased from Sigma Co., Ltd, USA. First, a 1.5 mM (0.06 mg / ml) standard DPPH ethanol solution was made. Then, ethanol was added to the ascorbic acid, an antioxidant, as a compound of Formula 1 and a reference material, respectively, to prepare samples at concentrations of 50 µg / ml, 25 µg / ml, 12.5 µg / ml, 6.25 µg / ml, and 3.125 µg / ml. . Then, the sample and the standard DPPH solution were added in the same ratio, stirred well, and then reacted at 37 ° C. for 30 minutes, and the absorbance was measured at 520 nm. At this time, ethanol was added instead of the sample as a control. The free radical scavenging ability was obtained from IC ₅₀ , which is a half maximal inhibitory concentration, and the results are shown in Table 3 below. IC ₅₀ is a general method of expressing free radical scavenging ability as a concentration of ascorbic acid and a compound of formula (1) required to remove 50% of free radicals from the no addition control.

Free radical scavenging rate (IC ₅₀ ) sample Free radical scavenging rate (ug / ml; ppm) Ascorbic acid 14 Compound of Formula 1 25

As a result of the evaluation, the compound of Formula 1 has lower activity than that of ascorbic acid (Vitamin C), which is a representative strong antioxidant, but it is a natural substance and has excellent stability with excellent level of antioxidant effect. there was.

Example  4: anti-inflammatory effect NO  Generation suppression effect

In order to confirm the anti-inflammatory effect and skin trouble alleviating effect of the compound of Formula 1, nitric oxide (NO) formation inhibitory experiment was performed by GRIESS method using RAW264.7 cell line (ATCC number: CRL-2278).

Specifically, RAW264.7 cells, which are macrophages of mice, were passaged several times, placed in a 24-well plate to enter 3 × 10 ⁵ cells in one well, and then cultured for 24 hours. Subsequently, the compound of formula 1 was diluted and replaced with the cell medium containing the concentration shown in Table 4 below. At this time, L-NMMA (L-NG-Monomethylarginine), a NO-producing inhibitor, was treated with a positive control and incubated for 30 minutes, and 1 μg of LPS (Lipopolysaccharide) was incubated for 24 hours. . NPS treated with 1 μg without any sample was used as a negative control. 100 μl of the supernatant was taken and transferred to a 96-well plate, 100 μl of GRIESS solution was added thereto, reacted at room temperature for 10 minutes, and the absorbance at 540 nm was measured to determine the NO inhibitory effect of Compound 1, and the results are shown in the following table. 4 is shown.

NO production inhibitory effect sample NO production inhibition rate (%) L-NMMA (positive control, 0.001 w / v%) 22.70 Compound of Formula 1 (0.001 w / v%) 9.00 Compound of Formula 1 (0.01 w / v%) 16.73 Compound of formula 1 (0.1 w / v%) 30.76 Compound of Formula 1 (1 w / v%) 39.21

As a result of the evaluation, the compound of Chemical Formula 1 is low in activity compared to NMMA, which is a representative anti-inflammatory substance, but shows excellent activity as a natural substance, and anti-inflammatory effect acts as a secondary role in antioxidation and wrinkle improvement, and thus a synergistic effect can be expected.

Formulation example  1: Preparation of Pharmaceutical Formulations

1. Preparation of powder

0.001 g of compound of Formula 1

1g lactose

The above ingredients were mixed and filled in airtight cloth to prepare a powder.

2. Preparation of Tablets

0.2 mg of compound of Formula 1

Corn Starch 100mg

Lactose 100mg

Stearic acid  Magnesium 2mg

After mixing the above components, tablets were prepared by tableting according to a conventional method for producing tablets.

3. Preparation of Capsule

0.2 mg of compound of Formula 1

Corn Starch 100mg

Lactose 100mg

Stearic acid  Magnesium 2mg

After mixing the above components, the capsule was prepared by filling in gelatin capsules according to the conventional method for producing a capsule.

4. Manufacture of rings

0.003 g of compound of Formula 1

Lactose 1.5 g

1 g of glycerin

Xylitol  0.5 g

After mixing the above components, it was prepared to be 4 g per ring according to a conventional method.

5. Preparation of Granules

2 mg of compound of formula 1

Soybean Extract 50mg

Glucose 200 mg

Starch 600 mg

After mixing the above components, 100 mg of 30% ethanol was added and dried at 60 ° C. to form granules, and then filled in fabric.

Formulation example  2: manufacture of cosmetics

1. Preparation of Soft Cosmetics (Skin Lotion)

Like the following composition, a flexible longevity containing the compound of formula 1 as an active ingredient was prepared according to a conventional method.

0.1 wt% of a compound of Formula 1

Beta-1,3-Glucan 1.0 wt%

Butylene Glycol 2.0 wt%

Propylene Glycol 2.0 wt%

Carboxy vinyl polymer 0.1 wt%

Fiji-12 nonylphenyl ether 0.2% by weight

Polysorbate 80 0.4 wt%

Ethanol 10.0 wt%

0.1% by weight of triethanolamine

0.05 wt% preservative

0.05% by weight of pigment

0.05% by weight fragrance

100% by weight of purified water

2. Preparation of Nutrients (Milk Lotion)

As shown in the following composition, nutrient longevity containing the compound of Formula 1 as an active ingredient was prepared according to a conventional method.

0.1 wt% of a compound of Formula 1

Beta-1,3-Glucan 1.0 wt%

Beeswax 4.0 wt%

Polysorbate60 1.5 wt%

Sorbanthesquioleate 1.5 wt%

0.5% by weight of liquid paraffin

Caprylic / Capric Triglycerides 5.0 wt%

Glycerin 3.0 wt%

Butylene Glycol 3.0 wt%

Propylene Glycol 3.0 wt%

Carboxy vinyl polymer 0.1 wt%

0.2% by weight of triethanolamine

0.05 wt% preservative

0.05% by weight of pigment

0.05% by weight fragrance

100% by weight of purified water

3. Preparation of nutrition cream

As in the following composition, a nourishing cream containing the compound of formula 1 as an active ingredient was prepared according to a conventional method.

0.2 wt% of a compound of Formula 1

Beta-1,3-Glucan 5.0 wt%

Beeswax 10.0 wt%

Polysorbate60 1.5 wt%

Sebum 60 Cured Castor Oil 2.0 wt%

0.5 wt% sorbitan sesquioleate

10.0% by weight of liquid paraffin

Squalane 5.0 wt%

Caprylic / Capric Triglycerides 5.0 wt%

Glycerin 5.0 wt%

Butylene Glycol 3.0 wt%

Propylene Glycol 3.0 wt%

0.2% by weight of triethanolamine

0.05 wt% preservative

0.05% by weight of pigment

0.05% by weight fragrance

100% by weight of purified water

4. Preparation of Massage Cream

As in the following composition, a massage cream containing a compound of Formula 1 as an active ingredient was prepared according to a conventional method.

0.1 wt% of a compound of Formula 1

Beta-1,3-Glucan 3.0 wt%

Beeswax 10.0 wt%

Polysorbate60 1.5 wt%

Sebum 60 Cured Castor Oil 2.0 wt%

Sorbanthesquioleate 0.8 wt%

40.0% by weight of liquid paraffin

Squalane 5.0 wt%

Caprylic / Capric Triglyceride 4.0 wt%

Glycerin 5.0 wt%

Butylene Glycol 3.0 wt%

Propylene Glycol 3.0 wt%

0.2% by weight of triethanolamine

0.05 wt% preservative

0.05% by weight of pigment

0.05% by weight fragrance

100% by weight of purified water

5. Manufacture of pack

As in the following composition, a pack containing the compound of formula 1 as an active ingredient was prepared according to a conventional method.

0.2 wt% of a compound of Formula 1

Beta-1,3-Glucan 1.0 wt%

Polyvinyl Alcohol 13.0 wt%

Sodium Carboxymethyl Cellulose 0.2% by weight

Glycerin 5.0 wt%

Allantoin 0.1 wt%

Ethanol 6.0 wt%

0.3% by weight of sebum-12 nonylphenyl ether

Polysorbate 60 0.3 wt%

0.05 wt% preservative

0.05% by weight of pigment

0.05% by weight fragrance

100% by weight of purified water

Formulation example  3: External skin preparations  Produce

1. Preparation of Gel

As in the following composition, a gel containing the compound of formula 1 as an active ingredient was prepared according to a conventional method.

0.1 wt% of a compound of Formula 1

Beta-1,3-Glucan 0.1 wt%

0.05% by weight of ethylenediamine sodium acetate

Glycerin 5.0 wt%

Carboxy vinyl polymer 0.3 wt%

Ethanol 5.0 wt%

Fiji-60 Cured Castor Oil 0.5 wt%

0.3% by weight of triethanolamine

0.05 wt% preservative

0.05% by weight of pigment

0.05% by weight fragrance

100% by weight of purified water

2. Manufacture of ointments

As shown in the following composition, an ointment containing the compound of formula 1 as an active ingredient was prepared according to a conventional method.

0.5% by weight of compound of formula 1

Beta-1,3-Glucan 10.0 wt%

Beeswax 10.0 wt%

Polysorbate 60 5.0 wt%

Sebum 60 Cured Castor Oil 2.0 wt%

Sorbanthesquioleate 0.5 wt%

Vaseline 5.0 wt%

10.0% by weight of liquid paraffin

Squalane 5.0 wt%

Shea Butter 3.0 wt%

Caprylic / Capric Triglycerides 5.0 wt%

Glycerin 10.0 wt%

Propylene Glycol 10.2 wt%

0.2% by weight of triethanolamine

0.05 wt% preservative

0.05% by weight of pigment

0.05% by weight fragrance

100% by weight of purified water

3. Preparation of Topical Drug (Gel Ointment)

As the following composition, a gel ointment comprising the compound of formula 1 as an active ingredient was prepared according to a conventional method.

0.5% by weight of compound of formula 1

Beta-1,3-Glucan 10.0 wt%

1.5% by weight of polyacrylic acid (Carbopol 940)

Isopropanol 5.0 wt%

Hexylene glycol 25.0 wt%

Triethanolamine 1.7 wt%

100% by weight of deionized water

4. Preparation of medicament (patch) for topical administration

As in the following composition, a patch containing the compound of Formula 1 as an active ingredient was prepared according to a conventional method.

0.5% by weight of compound of formula 1

Beta-1,3-Glucan 3.0 wt%

Hexylene glycol 20.0 wt%

Diethylamine 0.7 wt%

1.0% by weight of polyacrylic acid (Carbopol 934P)

Sodium sulfite 0.1 wt%

1.0 weight% of polyoxyethylene lauryl ether (E.O = 9)

1.0% by weight of polyhydroxyethylene cetyl stearyl ether (Cetomacrogol 1000)

Viscous paraffin oil 2.5% by weight

Caprylic acid ester / capric acid ester (Cetiol LC) 2.5% by weight

Polyethylene glycol 400 3.0 wt%

100% by weight of deionized water

Formulation example  4: manufacture of food

Food containing the compound of formula 1 of the present invention was prepared as follows.

1. Preparation of flour food

0.05 to 1.0 parts by weight of the compound of Formula 1 was added to the flour, and using the mixture to prepare bread, cake, cookies, crackers and noodles to prepare a health food.

2. Manufacturing of Dairy Products

0.2 parts by weight of the compound of Formula 1 was added to milk, and various dairy products such as butter and ice cream were prepared using the milk.

3. Manufacture of wire

Brown rice, barley, glutinous rice, and yulmu were alphad by a known method, and then dried and roasted to prepare a powder having a particle size of 60 mesh. Black beans, black sesame seeds, and perilla were also steamed and dried by a known method, and then ground to a powder having a particle size of 60 mesh. The compound of Chemical Formula 1 was concentrated under reduced pressure in a vacuum concentrator, and the dried product obtained by drying with a spray and a hot air dryer was pulverized with a particle size of 60 mesh to obtain a dry powder.

The dry powder of the grains, seeds and the compound of Formula 1 prepared in the above was prepared by blending at 100 parts by weight of the mixed powder in the following ratio.

Cereals (30 parts by weight brown rice, 15 parts by weight brittle, 20 parts by weight of barley),

Seeds (7 parts by weight perilla, 8 parts by weight black beans, 7 parts by weight black sesame seeds),

Compound of formula 1 (0.1 parts by weight),

Ganoderma lucidum (0.5 parts by weight),

Foxglove (0.5 Parts by weight )

Formulation example  5: manufacture of beverages

1. Preparation of health drinks

0.1 mg of compound of Formula 1

Citric acid 1000 mg

100 g oligosaccharides

Plum concentrate 2 g

1 g of taurine

Purified water  By adding a full 900 mL

After mixing the above components according to the conventional healthy beverage production method, and stirred and heated at 85 ℃ for about 1 hour, the resulting solution is filtered and obtained by sterilization in a sterilized 2L-container, sealed sterilization and then stored in the refrigerator It was used to prepare a healthy beverage composition of the invention.

Although the composition ratio is a composition suitable for a preferred beverage in a preferred embodiment, the composition ratio may be arbitrarily modified according to regional and ethnic preferences such as demand hierarchy, demand country, and intended use.

2. Preparation of Vegetable Juice

1 g of the compound of Chemical Formula 1 of the present invention was added to 1,000 mL of tomato or carrot juice to prepare vegetable juice for health promotion.

3. Preparation of Fruit Juice

1 g of the compound of Formula 1 was added to 1,000 mL of apple or grape juice to prepare fruit juice for health promotion.

Claims (18)

A pharmaceutical composition for improving skin renewal and wrinkles comprising camphorol-7-O-beta-D-glucopyranoside represented by the following Chemical Formula 1 as an active ingredient:
[Formula 1]

Skin external preparation for skin regeneration and wrinkle improvement comprising camphorol-7-O-beta-D-glucopyranoside represented by Formula 1 as an active ingredient:
[Formula 1]

The method of claim 2,
Skin external preparations in the form of ointments, patches, gels, creams or sprays.
A cosmetic composition for skin regeneration and wrinkle improvement comprising camphorol-7-O-beta-D-glucopyranoside represented by Formula 1 as an active ingredient:
[Formula 1]

The method of claim 4, wherein
Cosmetic composition in the form of a lotion, essence, lotion, cream, pack, gel, powder, foundation or detergent.
Health food for skin regeneration and wrinkle improvement comprising camphorol-7-O-beta-D-glucopyranoside represented by the following formula 1 as an active ingredient:
[Formula 1]

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