KR102146879B1 - Composition for skin cell regeneration, anti-wrinkle, anti-imflamation, and skin whitening - Google Patents

Abstract

The present invention relates to a composition for skin regeneration, wrinkle improvement, anti-inflammatory or skin whitening.
Pachymic acid according to the present invention increases the total amount of collagen in the fibroblasts of the skin, thereby reducing the skin regeneration effect, wrinkle improvement effect, and nitric oxide (NO) production to reduce the total amount of melanin, thereby reducing the whitening effect. By indicating, it can be used as a pharmaceutical composition, an external preparation for skin, a cosmetic composition, or a health food.

Description

Composition for skin cell regeneration, anti-wrinkle, anti-imflamation, and skin whitening}

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

Collagen is a major matrix protein produced by fibroblasts of the skin and is present in the extracellular interstitial. Its important functions include skin mechanical firmness, connective tissue resistance and tissue bonding, support for cell adhesion, cell division and differentiation (organic Growth or wound healing) is known. Such collagen is reduced by age and photoaging due to ultraviolet irradiation, which is known to be closely related to the formation of wrinkles in the skin. In addition, in recent years, with the development of extensive research on skin aging, an important function of collagen in the skin has been revealed.

Active ingredients that promote collagen synthesis to improve wrinkles are known. For example, retinoic acid, transforming growth factor (TGF) [Non-Patent Literature 1], protein derived from animal placenta [Patent Literature 1], betulinic acid [Patent Literature 2] and chlorella extract [ Patent Documents 3 and 4] are known as collagen synthesis promoting substances. However, there is a problem in that the amount of the active ingredients is limited due to safety problems such as irritation and redness when applied to the skin, or the effect is insignificant, so that the effect of substantially promoting collagen synthesis in the skin and improving skin function cannot be expected.

In addition, inflammation is an immune response of the human body in response to wounds or diseases, and oxidative stress such as ultraviolet rays, free radicals, etc. activates inflammatory factors, causing various diseases and aging of the skin. Vasoactive polypeptides such as kinin, plasmin, or complement act as 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 or lukotriene, an inflammatory mediator, through two pathways of cyclooxygenase or lipooxygenase, mediating various inflammatory reactions.

On the other hand, in order to eliminate inflammation, it is called an anti-inflammatory agent that acts to remove inflammatory sources, reduce biological reactions, and symptoms. Substances used for anti-inflammatory purposes to date include flufenamic acid, ibuprofen, benzydamine, or indomethacin as non-steroids, and prednisolone as a steroid system. Or dexamethasone. In addition, allantoin, azene, hydrocortisone, etc. are known to be effective in anti-inflammatory, but these substances are limited in use due to safety issues on the skin, or the effect is insignificant, so there is a problem that it is not possible to substantially alleviate inflammation. have.

Also, it is a common desire to have white and fair skin. The color or brightness of the skin is genetically determined according to the concentration and distribution of melanin in a person's skin, but is also affected by environmental or physiological conditions such as solar ultraviolet rays, fatigue or stress. Melanin is produced through a non-enzymatic oxidation reaction after being sequentially converted into dopa and dopaquinone by an enzyme called tyrosinase acting as a catalyst on tyrosine, a kind of amino acid. As such, the pathway by which melanin is produced is known, but the cause of the triggering of tyrosinase in the mechanism that induces melanin synthesis is still not elucidated in detail.

Meanwhile, as a commonly known whitening component, substances that inhibit tyrosinase enzyme activity such as Kojic acid or arbutin, hydroquinone, vitamin C (L-Ascorbic acid), or derivatives thereof, and various There are plant extracts. By inhibiting the synthesis of melanin pigments, they can not only realize skin whitening by brightening the skin tone, but also improve skin hyperpigmentation such as spots and freckles caused by ultraviolet rays, hormones or genetics. However, when applied to the skin, there is a problem in that the amount of use is limited due to safety issues such as irritation and redness, or the effect is insignificant and thus no practical effect can be expected.

Therefore, the development of ingredients that are safe for the living body, have stable active ingredients, and above all have superior skin regeneration, wrinkle improvement, anti-inflammatory or whitening activity than substances that have an existing skin regeneration, wrinkle improvement, anti-inflammatory or whitening effect. This is desperately desired.

Japanese Patent Application No. Hei 8-231370 Japanese Patent Application No. Hei 8-208424 Japanese Patent Application No. Hei 9-40523 Japanese Patent Application Laid-Open No. Hei 10-36283

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

Thus, the inventors of the present invention believe that Pachymic acid increases the total amount of collagen in fibroblasts in the skin, promotes skin regeneration, improves wrinkles, inhibits nitric oxide (NO) production, and exhibits anti-inflammatory effects, and the total amount of melanin It was confirmed that the whitening effect was significantly reduced, and the present invention was completed.

Accordingly, an object of the present invention is to provide a composition for skin regeneration, wrinkle improvement, anti-inflammatory or skin whitening 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

It provides a pharmaceutical composition for skin regeneration, wrinkle improvement, anti-inflammatory, or skin whitening comprising a compound represented by the following Formula 1 as an active ingredient.

[Formula 1]

As another means for solving the above problem, the present invention

It provides a cosmetic composition for skin regeneration, wrinkle improvement, anti-inflammatory or skin whitening comprising a compound represented by the following Formula 1 as an active ingredient.

[Formula 1]

Pachymic acid according to the present invention promotes skin regeneration by increasing the total amount of collagen in fibroblasts in the skin, suppresses the production of nitric oxide (NO) involved in the inflammatory reaction, has excellent anti-inflammatory effects, and has excellent melanin. It can be used in medicines or cosmetics by showing a whitening effect that significantly reduces

Hereinafter, the configuration of the present invention will be described in detail.

In order for skin regeneration, wrinkle improvement, anti-inflammatory or whitening ingredients to exhibit excellent effects when applied to actual skin, they exhibit highly active skin regeneration, wrinkle improvement, anti-inflammatory or whitening activity at low concentration, and have the ability to penetrate and absorb the skin. It is desirable to be excellent. In addition, it has low volatility so that it can stay for a sufficient time to exhibit skin regeneration, wrinkle improvement, anti-inflammatory or whitening effect, and the active ingredient remains stable on the composition or skin, and it is easy to formulate into medicine or cosmetics. It is desirable to be safe. However, among the known components, it is not common to find components that satisfy all of the above properties. For example, some skin regeneration, anti-wrinkle, anti-inflammatory or whitening ingredients are excellent in skin regeneration, wrinkle improvement, anti-inflammatory or whitening activity even at low concentrations when tested in vitro, but their ability to penetrate through the skin and be absorbed is low. It is difficult to apply. Other active ingredients have low hydrophilic properties, making it difficult to formulate into pharmaceuticals or cosmetics. In addition, some skin rejuvenation, anti-wrinkle, anti-inflammatory or whitening ingredients may be decomposed or transformed into other compounds when exposed to heat, light, or oxygen, and their effects disappear before application to the skin.

As can be seen in the following examples, Pachymic acid has the effect of significantly increasing the total amount of collagen at low concentration, the effect of remarkably reducing the total amount of melanin, and the effect of inhibiting nitric oxide production. It can be used as an active ingredient in medicines, cosmetics or health foods for inflammation or skin whitening.

Accordingly, the present invention provides a pharmaceutical composition for skin regeneration, wrinkle improvement, anti-inflammatory or skin whitening comprising a compound represented by the following Formula 1 as an active ingredient.

[Formula 1]

The compound of Chemical Formula 1 is referred to as pachymic acid, and may include all possible isomers.

The compound of Formula 1 may be used separately or synthesized from a plant extract, or a commercially available compound may be used.

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

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, 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, Benzenesulfonic acid, p-toluenesulfonic acid, or methanesulfonic acid-based salts. Inorganic acids include, for example, hydrochloric acid, bromic acid, sulfuric acid, phosphoric acid, nitric acid, carbonic acid or boric acid-based salts. Preferably, it may be in the form of hydrochloride or acetate, and more preferably in the form of hydrochloride.

The above-mentioned acid addition salt is a) directly mixing the compound of Formula 1 and an acid, b) dissolving and mixing one of them in a solvent or aqueous solvent, or c) adding the compound of Formula 1 to a solvent or dropping It is prepared by a general salt preparation method by placing them in an acid in a solvent and mixing them.

In addition to the above, additional salts are possible: gaba salt, gabapentin salt, pregabalin salt, nicotinate, adipate salt, hemimalonate, cysteine salt, acetylcysteine salt, methionine salt, arginine salt, lysine salt, ornithine salt or And aspartate.

In addition, when the compound of Formula 1 of the present invention is used as a pharmaceutical product, it may further contain one or more active ingredients exhibiting the same or similar function. For example, it may include known skin regeneration, wrinkle improvement, anti-inflammatory or skin whitening ingredients. If additional skin regeneration, wrinkle improvement, anti-inflammatory or skin whitening ingredients are included, the skin regeneration, wrinkle improvement, anti-inflammatory or skin whitening effect of the composition of the present invention may be further enhanced. When the above ingredients are added, skin safety, ease of formulation, and stability of active ingredients can be considered in combination with use. In one embodiment of the present invention, the composition is a skin regeneration component known in the art, including retinoic acid, TGF, protein derived from animal placenta, betulinic acid or chlorella extract; As a whitening component known in the art, substances that inhibit tyrosinase enzyme activity such as kojic acid or arbutin, hydroquinone, vitamin-C (L-Ascorbic acid); Derivatives thereof; And it may further include one or two or more components selected from the group consisting of various plant extracts. Additional components may be included in an amount of 0.0001% by weight or more and 10% by weight or less based on the total weight of the composition. For example, 0.0001% by weight or more and 1% by weight or more, 0.0001% by weight or more and 0.1% by weight or less, 0.0001% by weight or more and 0.001% by weight or less, 0.001% by weight or more and 10% by weight or less, 0.001% by weight or more and 1 It may be less than or equal to 0.001% by weight and less than or equal to 0.1% by weight, less than or equal to 0.01% and less than or equal to 10%, and less than or equal to 0.01% and less than 1% by weight. The content range may be adjusted according to requirements such as skin safety and ease of formulation of the compound of Formula 1.

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

The pharmaceutically acceptable carrier may contain various components such as a buffer solution, sterile water for injection, general saline or phosphate buffered saline, sucrose, histidine, or polysorbate.

The pharmaceutical composition of the present invention may be administered orally or parenterally, and may be administered in the form of a general pharmaceutical formulation, for example, in various oral or parenteral formulations during clinical administration. In the case of formulation, a filler commonly used , Extenders, binders, wetting agents, disintegrants or surfactants, such as diluents or excipients.

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

In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid preparations for oral use include suspensions, liquid solutions, emulsions or syrups, and various excipients such as wetting agents, sweetening agents, fragrances or preservatives may be included in addition to water or liquid paraffin, which are simple diluents commonly used.

Preparations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized preparations, or suppositories. As the non-aqueous solvent or suspension solvent, vegetable oils such as propylene glycol, polyethylene glycol, or olive oil; Injectable esters such as ethyl oleate may be used. As a base for suppositories, witepsol, macrogol, tween 61, cacao butter, laurin paper or glycerogelatin may be used.

In the present invention, the term'skin regeneration effect' refers to recovery of skin tissue against damage caused by external and internal causes of the skin. The damage caused by the external cause may include ultraviolet rays, external pollutants, wounds or trauma, and the damage caused by the internal cause may include stress.

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

In the present invention, "anti-inflammatory effect" refers to suppressing inflammation. The inflammation is one of the defense responses of living tissues against certain stimuli, and refers to a complex lesion that co-occurs three types of tissue deterioration, circulatory disorders, effusion, and tissue proliferation. More specifically, inflammation is part of innate immunity, and as in other animals, innate immunity in humans recognizes patterns on the cell surface that are specific to pathogens. Phagocytes recognize cells with such surfaces as non-magnetic and attack pathogens. If pathogens break through the body's physical barriers, an inflammatory reaction occurs. The inflammatory reaction is a nonspecific defense action that creates a hostile environment for microorganisms invading the wound. In the inflammatory reaction, when a wound or an external infectious agent enters the body, white blood cells responsible for the initial stage immune response gather and express cytokines. Therefore, the amount of expression of cytokines in cells becomes an indicator of activation of the inflammatory response. Examples of skin diseases related to inflammation include atopic dermatitis, psoriasis, erythematous disease caused by radiation, chemical substances, or burns; Itching due to acid burns, bullous skin disease, lichen planus disease or allergies; Inflammatory hair loss such as seborrheic eczema, rose acne, pemphigus vulgaris, polymorphic exudative erythema, nodular erythema, balanitis, vulvitis or alopecia areata; Cutaneous T-cell lymphoma and the like, but are not limited thereto.

In the present invention, the term'whitening effect' refers to not only brightening skin tone by inhibiting the synthesis of melanin pigment, but also improving skin hyperpigmentation such as spots and freckles caused by ultraviolet rays, hormones or genetics.

When the pharmaceutical composition of the present invention contains an effective amount of the compound of Formula 1, it can provide desirable effects such as skin regeneration, wrinkle improvement, anti-inflammatory or whitening. In the present invention, "effective amount" refers to the amount of a compound capable of promoting regeneration of damaged skin, improving wrinkles, suppressing inflammation, or exhibiting a whitening effect. 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, the method in which the compound is applied to the skin, or the time it stays on the skin. For example, when the composition is commercialized as a pharmaceutical, it may contain the compound of Formula 1 at a higher concentration than when it is commercialized 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, and 1 to 100 mg/kg per day based on the amount of the compound of Formula 1 above. It can be administered 6 times.

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

The present invention may also be provided in a formulation of a skin external preparation for skin regeneration, wrinkle improvement, anti-inflammatory or whitening, comprising the compound of Formula 1 as an active ingredient.

When the compound of Formula 1 is used for external skin, additionally fatty substances, organic solvents, solubilizers, thickening and gelling agents, emollients, antioxidants, suspending agents, stabilizers, foaming agents, fragrances, surfactants , Water, ionic emulsifier, nonionic emulsifier, filler, sequestering agent, chelating agent, preservative, vitamin, blocker, wetting agent, essential oil, dye, pigment, hydrophilic activator, lipophilic activator or lipid vesicle, etc. May contain adjuvants commonly used in the field of dermatology such as any other ingredients commonly used in In addition, the above ingredients may 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 may be a formulation such as an ointment, patch, gel, cream, or spray, but is not limited thereto.

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

When the compound of Formula 1 is used as a cosmetic, a cosmetic prepared by containing the compound of Formula 1 as an active ingredient may be prepared in the form of a general emulsified formulation or a solubilized formulation. For example, a lotion such as a flexible lotion or a nutritional lotion; Emulsions such as facial lotion or body lotion; Creams such as nourishing cream, moisture cream, or eye cream; essence; Cosmetic ointment; spray; Gel; pack; Sunscreen; Makeup base; Foundations such as liquid type, solid type or spray type; powder; Makeup removers such as cleansing cream, cleansing lotion, and cleansing oil; It may have a formulation such as a detergent such as a cleansing foam, soap, and body wash.

In addition, in addition to the compound of Formula 1, the cosmetics include fatty substances, organic solvents, solubilizers, thickeners, gelling agents, emollients, antioxidants, suspending agents, stabilizers, foaming agents, fragrances, surfactants, water, Ionic emulsifiers, nonionic emulsifiers, fillers, sequestering agents, chelating agents, preservatives, vitamins, blockers, wetting agents, essential oils, dyes, pigments, hydrophilic activators, lipophilic activators or lipid vesicles, etc. It may contain auxiliaries used.

The compound of Formula 1 is a relatively high concentration of the compound of Formula 1 in the case of a wash-off type cosmetic such as a makeup remover or detergent in which the active ingredient stays on the skin within a short period of time when it is commercialized as a cosmetic. Could be included. On the other hand, in the case of leave-on type cosmetics such as lotions, emulsions, creams, or essences in which the active ingredient stays on the skin for a long period of time, the compound of Formula 1 is contained in a lower concentration than the wash-off type cosmetics. It would be fine. Although not limited thereto, in one embodiment of the present invention, the composition comprises 0.0001% to 10% by weight (preferably 0.0001% to 1% by weight) of the compound of Formula 1 based on the total weight of the composition. I can. 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, anti-inflammatory or whitening effects cannot be expected, and when it contains more than 10% by weight, allergy, etc. This is to prevent a reaction that does not occur or there may be a problem with skin safety.

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

In the present specification, the term'health food' refers to food prepared by adding the compound of Formula 1 to food materials such as beverages, teas, spices, chewing gum, confectionery, or encapsulating, powdering, and suspension, etc. It means to bring about a specific effect, but unlike general drugs, it has the advantage of not having side effects that may occur when long-term use of drugs is made using food as a raw material.

Since the health food of the present invention obtained in this way can be consumed on a daily basis, high skin regeneration, wrinkle improvement, anti-inflammatory or whitening effects can be expected, and thus it is 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 may be used together with other foods or food ingredients, and may be appropriately used according to a conventional method. The mixed amount of the active ingredient may be appropriately determined according to the purpose of use (prevention, health or therapeutic treatment). In general, in the manufacture of food or beverage, the composition of the present invention is added in an amount of 15 parts by weight or less, preferably 10 parts by weight or less based on the raw material. However, when ingested for a long time for health and hygiene purposes, the content below the above range may be included, and since there is no problem in terms of safety, the active ingredient may be used in an amount above the above range.

There is no particular limitation on the type of food. 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, There are alcoholic beverages or vitamin complexes, and all health foods in the usual sense are included.

The health beverage composition of the present invention may contain various flavoring agents or natural carbohydrates as an additional component, like a conventional beverage. The natural carbohydrates described above include monosaccharides such as glucose and fructose; Disaccharides such as maltose and sucrose; Polysaccharides such as dextrin and cyclodextrin; It may be a sugar alcohol such as xylitol, sorbitol, and erythritol. Sweeteners include natural sweeteners such as taumatin and stevia extract; Synthetic sweeteners such as saccharin and aspartame can be used. The proportion of the 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, flavoring agents, colorants, pectic acid, salts of pectic acid, alginic acid, salts of alginic acid, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, It may contain glycerin, alcohol or a carbonating agent. In addition, the health food of the present invention may contain flesh for the manufacture of natural fruit juice, fruit juice beverage, or vegetable beverage. These ingredients may be used independently or in combination. The ratio of these additives is not very important, but it is generally selected from 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 examples. However, the following examples are only illustrative of the present invention, and the contents of the present invention are not limited to the following examples.

Reference example One: Pachymic acid substance Information

Name: Pachymic acid; 3-acetyltumulosic acid ; 2-(3-acetoxy-16-hydroxy-4,4,10,13,14-pentamethyl-2,3,5,6,7,11,12,15,16,17-decahydro-1H-cyclopenta[a ]phenanthren-17-yl)-5-isopropyl-hex-5-enoic acid

CAS No.: 29070-92-6

Where to buy: Tauto Biotech Co., Ltd.

Example 1: Effect of increasing the total amount of collagen

The substances in Table 1 were added to the culture medium of human-derived fibroblasts to measure the total amount of collagen at the cell level. The total amount of collagen was measured using the PICP EIA kit (Procollagen Type I C-Peptide Enzyme ImmunoAssay KIT). Cytotoxicity was evaluated at concentrations of 10 ppm, 1 ppm, 0.1 ppm, 0.01 ppm, and 0.001 ppm for 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], and a concentration without cytotoxicity was selected to measure the total amount of collagen.

In the experiment, the final concentration of the compound of Formula 1 was 1 ppm and 10 ppm, respectively, and each sample was added to the culture medium of human fibroblasts and incubated for 1 day, and then the culture solution was taken and collagen at each concentration using the PICP EIA kit. The total amount was measured at 450 nm using a spectrophotometer. In order to compare the effect, the total amount of collagen was measured in the same manner for the culture medium (control) of fibroblasts to which nothing was added and the sample to which vitamin C was added to a final concentration of 52.8 μg/ml. The total amount of collagen was measured as UV absorbance, and the rate of increase of the total amount of collagen was calculated as the ratio of the total amount of collagen relative to the control, and the results are summarized in Table 1 below.

Effect of increasing the total amount of collagen of the compound of formula 1 sample Absorbance average Collagen increase rate (%) Compound of formula 1
(1 ㎍/ml) 548 123.4 Compound of formula 1
(10 ㎍/ml) 772 173.9 Control
(No additives) 444 - Vitamin C
(52.8 μg/ml) 578 130.2

As can be seen from the results of Table 1, the compound of Formula 1 showed an increase in the total amount of collagen that was better at a lower concentration than when vitamin C, which was known to have collagen synthesis ability, was applied.

Therefore, it was found that the compound of Formula 1 can be used for skin regeneration and wrinkle improvement.

Example 2: Collagen crab Inhibitory effect

For the substances in Table 2 below, the effect of inhibiting collagenase activity was confirmed as follows.

Cytotoxicity was evaluated at concentrations of 10 ppm, 1 ppm, 0.1 ppm, 0.01 ppm, and 0.001 ppm for 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], and a collagenase evaluation method was performed by selecting a concentration without cytotoxicity.

Fibroblasts, which are normal human skin cells, were inoculated into 24-well microplates to be 2.5×10 ⁴ cells per well, cultured for 24 hours in 10% serum DMEM medium and 37°C conditions, and then 10% serum DMEM medium was removed. After washing once with a phosphate buffer solution, it was further cultured for 30 minutes in a serum-free DMEM medium to which the compound of Formula 1 was added and a serum-free DMEM medium that did not contain the compound of Formula 1 as a control.

After 30 minutes of sample treatment, it was cultured for 24 hours after stimulation with 50 ng/ml of TNF-α (tumor necrosis factor-α), a substance known to produce collagenase MMP-1. At this time, the TNF-α untreated group and the treated group were the untreated group not treated with TNF-α and the treated group treated with TNF-α among the control groups that did not contain the compound of Formula 1.

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

The amount of MMP-1 in the experimental group refers to the group to which the substance was added at each concentration.

The inhibitory effect of the compound of formula 1 on collagenase activity (repeat number = 4) sample Amount of MMP-1 (ng/ml) Inhibition rate (%) 10 ppm of compound of formula 1 6.46 94.71% 1 ppm of compound of formula 1 16.28 20.48% Negative control (TNF-α no treatment) 5.76 - Positive control (TNF-α treatment) 18.99 -

As can be seen from the results in Table 2, it was found that when the compound of Formula 1 and TNF-α were treated, the amount of MMP-1 produced was decreased compared to the positive control group treated with TNF-α without the compound of Formula 1 I can. That is, the compound of Formula 1 exhibited an excellent inhibitory effect on collagenase activity.

Therefore, it was found that the compound of Formula 1 can be used for skin regeneration and wrinkle improvement.

Example 3: whitening effect-reducing the total amount of melanin

The whitening effect was confirmed by measuring the total amount of melanin at the cell level by adding the compound of Formula 1 to the culture medium of mouse melanoma cells (B-16 mouse melanoma cells) (Lotan R., Lotan D. Cancer) Res . 40:3345-3350, 1980). Before the experiment, toxicity was evaluated for the melanoma cells of mice, and a non-toxic concentration was selected to perform whitening evaluation.

The final concentration of the compound of Formula 1 was 5ppm, 10ppm, and 20ppm in the culture medium, and arbutin, a control, was added to the medium so that 200ppm was treated to B-16 melanoma cells and cultured for 3 days.

Thereafter, the cells were treated with trypsin, removed from the culture vessel, centrifuged, and melanin was extracted. The detached cells were boiled for 10 minutes by adding 1 mL of sodium hydroxide solution (1N concentration) to dissolve melanin, and the absorbance was measured at 400 nm using a spectrophotometer to measure the amount of melanin produced.

The amount of melanin was measured by a method expressed in terms of absorbance per unit cell count (1×10 ⁶ cells), and the total amount of melanin relative to the control was calculated as an inhibition rate (%), and the results are summarized in Table 3. The experiment was repeated three times.

The effect of reducing the total amount of melanin at the cellular level according to the concentration sample Total melanin (Abs.) Inhibition rate (%) Control (no addition) 0.212 - Control 1: Arbutin (200ppm) 0.140 33.96 Compound of Formula 1 (20ppm) 0.099 53.3 Compound of Formula 1 (10ppm) 0.125 41.0 Compound of Formula 1 (5ppm) 0.159 25.0

As can be seen from the results of Table 3, when compared to arbutin, which is known as a whitening material, the compound of Formula 1 exhibits a similar total amount of melanin to the melanoma cells of mice cultured at a small concentration, indicating that it can be used for whitening purposes. Could know.

Example 4: Anti-inflammatory effect- NO Production inhibitory effect

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

Specifically, RAW264.7 cells, which are mouse macrophages, were subcultured several times, placed in a 24-well plate so as to enter 3×10 ⁵ cells in each well, and cultured for 24 hours. Subsequently, the compound of Formula 1 was diluted to the concentration shown in Table 4 and replaced with a cell medium containing. At this time, the NO-production inhibitor L-NMMA (L-NG-Monomethylarginine) was treated together as a positive control and incubated for 30 minutes, and 1 μg of LPS (Lipopolysaccharide) was treated as an stimulating source and incubated for 24 hours. . Take 100 μl of the supernatant and transfer it to a 96-well plate, add 100 μl of the GRIESS solution to react at room temperature for 10 minutes, and measure the absorbance at 540 nm to determine the NO inhibitory effect of the compound of Formula 1, and the result It is shown in Table 4 below.

NO production inhibitory effect sample NO production inhibition rate (%) L-NMMA (positive control, 0.001%) 41.38 Compound of Formula 1 (0.001 %) 23.42 Compound of Formula 1 (0.01 %) 29.10 Compound of formula 1 (0.1%) 33.23 Compound of Formula 1 (1%) 40.09

As can be seen from Table 4, the compound of Formula 1 has low activity compared to L-NMMA, which is a representative anti-inflammatory drug, but exhibits excellent activity as a natural substance, so that the anti-inflammatory effect plays a secondary role in whitening or improving wrinkles. It can be expected to act as a synergistic effect.

Formulation example 1: Preparation of pharmaceutical formulation

1. Preparation of powder

0.001 g of compound of formula 1

1g lactose

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

2. Preparation of tablets

0.2 mg of compound of formula 1

100mg corn starch

100mg lactose

Stearic acid 2 mg of magnesium

After mixing the above ingredients, tablets were prepared by tableting according to a conventional tablet preparation method.

3. Preparation of capsules

0.2 mg of compound of formula 1

100mg corn starch

100mg lactose

Stearic acid 2 mg of magnesium

After mixing the above ingredients, the gelatin capsules were filled according to a conventional capsule preparation method to prepare a capsule formulation.

4. Preparation of pills

0.003 g of compound of formula 1

1.5 g lactose

1 g glycerin

Xylitol 0.5 g

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

5. Preparation of granules

Compound of Formula 1 2 mg

Soybean extract 50 mg

200 mg of glucose

Starch 600 mg

After mixing the above ingredients, 100 mg of 30% ethanol was added, dried at 60°C to form granules, and then filled into a cloth.

Formulation example 2: manufacture of cosmetics

1. Manufacture of flexible lotion (skin lotion)

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

0.1% by weight of the compound of formula 1

1.0% by weight of beta-1,3-glucan

2.0% by weight of butylene glycol

2.0% by weight of propylene glycol

0.1% by weight of carboxyvinyl polymer

PG-12 nonylphenyl ether 0.2% by weight

Polysorbate 80 0.4% by weight

Ethanol 10.0% by weight

0.1% by weight of triethanolamine

0.05% by weight of preservative

0.05% by weight of pigment

Perfume 0.05% by weight

100% by weight of purified water

2. Manufacture of nutrient cosmetic water (milk lotion)

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

0.1% by weight of the compound of formula 1

1.0% by weight of beta-1,3-glucan

Beeswax 4.0% by weight

1.5% by weight of polysorbate 60

1.5% by weight of sorbitansquioleate

0.5% by weight of liquid paraffin

Caprylic/Capric Triglyceride 5.0% by weight

3.0% by weight of glycerin

Butylene glycol 3.0% by weight

Propylene glycol 3.0% by weight

0.1% by weight of carboxyvinyl polymer

0.2% by weight of triethanolamine

0.05% by weight of preservative

0.05% by weight of pigment

Perfume 0.05% by weight

100% by weight of purified water

3. Manufacture of nutrition cream

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

0.2% by weight of the compound of formula 1

5.0% by weight of beta-1,3-glucan

10.0 wt% beeswax

1.5% by weight of polysorbate 60

Sebum 60 hydrogenated castor oil 2.0% by weight

0.5% by weight of sorbitansquioleate

Liquid paraffin 10.0% by weight

5.0% by weight of squalane

Caprylic/Capric Triglyceride 5.0% by weight

5.0% by weight of glycerin

Butylene glycol 3.0% by weight

Propylene glycol 3.0% by weight

0.2% by weight of triethanolamine

0.05% by weight of preservative

0.05% by weight of pigment

Perfume 0.05% by weight

100% by weight of purified water

4. Manufacturing of massage cream

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

0.1% by weight of the compound of formula 1

Beta-1,3-glucan 3.0% by weight

10.0 wt% beeswax

1.5% by weight of polysorbate 60

Sebum 60 hydrogenated castor oil 2.0% by weight

Sorbitansquioleate 0.8% by weight

Liquid paraffin 40.0 wt%

5.0% by weight of squalane

Caprylic/Capric Triglyceride 4.0% by weight

5.0% by weight of glycerin

Butylene glycol 3.0% by weight

Propylene glycol 3.0% by weight

0.2% by weight of triethanolamine

0.05% by weight of preservative

0.05% by weight of pigment

Perfume 0.05% by weight

100% by weight of purified water

5. Manufacturing of the pack

As shown 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% by weight of the compound of formula 1

1.0% by weight of beta-1,3-glucan

13.0% by weight of polyvinyl alcohol

Sodium carboxymethylcellulose 0.2% by weight

5.0% by weight of glycerin

0.1% by weight of allantoin

Ethanol 6.0% by weight

PG-12 nonylphenyl ether 0.3% by weight

Polysorbate 60 0.3% by weight

0.05% by weight of preservative

0.05% by weight of pigment

Perfume 0.05% by weight

100% by weight of purified water

Formulation example 3: External skin Produce

1. Preparation of gel

As shown 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% by weight of the compound of formula 1

0.1% by weight of beta-1,3-glucan

Sodium ethylenediamine acetate 0.05% by weight

5.0% by weight of glycerin

0.3% by weight of carboxyvinyl polymer

Ethanol 5.0% by weight

PEG-60 hydrogenated castor oil 0.5% by weight

0.3% by weight of triethanolamine

0.05% by weight of preservative

0.05% by weight of pigment

Perfume 0.05% by weight

100% by weight of purified water

2. Preparation of ointment

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 the compound of formula 1

Beta-1,3-glucan 10.0% by weight

10.0 wt% beeswax

5.0% by weight of polysorbate 60

PEG 60 hydrogenated castor oil 2.0% by weight

0.5% by weight of sorbitansquioleate

Vaseline 5.0% by weight

Liquid paraffin 10.0% by weight

5.0% by weight of squalane

Shea butter 3.0% by weight

Caprylic/Capric Triglyceride 5.0% by weight

Glycerin 10.0% by weight

10.2% by weight of propylene glycol

0.2% by weight of triethanolamine

0.05% by weight of preservative

0.05% by weight of pigment

Perfume 0.05% by weight

100% by weight of purified water

3. Preparation of drugs for topical administration (gel ointment)

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

0.5% by weight of the compound of formula 1

Beta-1,3-glucan 10.0% by weight

1.5% by weight of polyacrylic acid (Carbopol 940)

5.0% by weight of isopropanol

25.0% by weight of hexylene glycol

1.7 wt% triethanolamine

100% by weight of deionized water

4. Preparation of drugs for topical administration (patches)

As shown 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 the compound of formula 1

3.0% by weight of beta-1,3-glucan

Hexylene glycol 20.0% by weight

0.7% by weight of diethylamine

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

Sodium sulfite 0.1% by weight

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

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

2.5% by weight viscous paraffin oil

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

Polyethylene glycol 400 3.0% by weight

100% by weight of deionized water

Formulation example 4: manufacture of food

Foods containing the compound of Formula 1 of the present invention were prepared as follows.

1. Manufacture of flour food

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

2. Manufacture 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. Manufacturing of Seonsik

Brown rice, barley, glutinous rice, and adlay were gelatinized by a known method, dried, and then roasted, and then prepared into a powder having a particle size of 60 mesh with a grinder. Black soybeans, black sesame seeds, and perilla seeds were also steamed and dried by a known method, and then roasted, and then prepared into a powder having a particle size of 60 mesh with a grinder. The dried product obtained by concentrating the compound of Formula 1 under reduced pressure in a vacuum concentrator, spraying, and drying with a hot air dryer was pulverized to a particle size of 60 mesh with a grinder to obtain a dry powder.

The grains, seeds, and dry powder of the compound of Formula 1 were mixed in the following ratio with respect to 100 parts by weight of the mixed powder.

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

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

A compound of formula 1 (0.1 parts by weight),

Ganoderma lucidum (0.5 parts by weight),

Rehmannia (0.5 Parts by weight )

Formulation example 5: preparation of beverage

1. Manufacturing of health drinks

0.1 mg of the compound of formula 1

Citric acid 1000 mg

100 g oligosaccharides

2 g of plum concentrate

1 g taurine

Purified water By total 900 mL

After mixing the above ingredients according to the general health drink manufacturing method, stirring and heating at 85°C for about 1 hour, the resulting solution is filtered and obtained in a sterilized 2L-container, sealed and sterilized, and stored in a refrigerator. It was used to prepare the health beverage composition of the invention.

The composition ratio is composed of ingredients suitable for a relatively preferred beverage in a preferred embodiment, but the blending ratio may be arbitrarily modified according to regional and ethnic preferences such as the demand class, the country of demand, and the intended use.

2. Manufacture of vegetable juice

Vegetable juice for health promotion was prepared by adding 1 g of the compound of Formula 1 of the present invention to 1,000 mL of tomato or carrot juice.

3. Manufacture of fruit juice

Fruit juice for health promotion was prepared by adding 1 g of the compound of Formula 1 to 1,000 mL of apple or grape juice.

Claims (6)

A pharmaceutical composition for skin regeneration or wrinkle improvement comprising a compound represented by the following Formula 1 as an active ingredient:
[Formula 1]

A cosmetic composition for skin regeneration or wrinkle improvement comprising a compound represented by the following Formula 1 as an active ingredient:
[Formula 1]

Anti-inflammatory pharmaceutical composition comprising a compound represented by the following formula (1) as an active ingredient:
[Formula 1]

Anti-inflammatory cosmetic composition comprising a compound represented by the following formula 1 as an active ingredient:
[Formula 1]

A pharmaceutical composition for skin whitening comprising a compound represented by the following Formula 1 as an active ingredient:
[Formula 1]

A cosmetic composition for skin whitening comprising a compound represented by the following Formula 1 as an active ingredient:
[Formula 1]