KR101263483B1 - A composition for skin hyper-pigmented diseases and skin whitening containing a compound from calcium tree prunus humilis - Google Patents

Abstract

PURPOSE: A composition including a compound from calcium tree is provided to enhance skin lightening and preventing melanogenesis. CONSTITUTION: A pharmaceutical composition for treatment or prevention of hyperpigmentation disorders caused by melanin oversecretion includes hexacosanedioic acid of chemical formula 1 or a pharmaceutically acceptable salt thereof. A health functional food composition for improvement or prevention of hyperpigmentation disorders caused by melanin oversecretion includes hexacosanedioic acid of chemical formula 1. A cosmetic composition for skin lightening includes hexacosanedioic acid of chemical formula 1. A calcium fraction H has an enzyme activity inhibition effect.

Description

A composition for skin hyper-pigmented diseases and skin whitening containing a compound from Calcium tree Prunus humilis}

The present invention relates to a composition for the treatment of skin hyperpigmentation diseases and skin whitening containing a calcium wood-derived compound, and more specifically to the prevention of skin hyperpigmentation disease due to melanin excess formation containing hexacosanedioic acid, a compound derived from calcium wood, or The present invention relates to a therapeutic pharmaceutical composition and health functional food composition and a composition for skin whitening.

Melanin, which determines human skin color, is made from skin cells called melanocytes and migrates to epidermal cells called keratinocytes. Here, melanin forms a hat-like structure around the nucleus to protect genes from ultraviolet rays and remove free radicals to play an important role in protecting intracellular proteins.

Melanogenesis refers to the biosynthesis of melanocytes in melanocytes, which are regulated by intracellular and extracellular environments. This melanin production is triggered by tyrosinase, which oxidizes tyrosine to dopaquinone, which evkznlshs subsequently converts to a pale yellow / red compound, pheomelanin and light brown to black section eumelanin. do. In the skin, melanin affects pigmentation and protects the skin from the causes of ultraviolet rays and other skin diseases, while abnormal accumulation of melanin can cause spots and freckles.

Skin hyperpigmentation diseases caused by melanin hyperplasia include genetic skin disorders such as alum, Badenburgh syndrome, acquired skin diseases such as infectious white nasal mucus, unexplained droplet melaninosis, blemishes and skin diseases resulting from drug treatment such as minocycline , Bleomycin, busulfan, zidovudine, and skin diseases metastasized through infection such as scrubs and the like.

There is no enzyme in the body that breaks down melanin, but when keratinocytes break off the epidermis, they are removed by falling off the skin with melanin. However, if more melanin is produced than necessary, it causes hyperpigmentation, such as blemishes, freckles, and spots, and it is a cosmetically bad result.

On the other hand, as the leisure population increases, the number of people who enjoy working outside increases the demand to prevent melanin pigmentation caused by ultraviolet rays. Therefore, it was necessary to develop a whitening agent that prevents excessive melanin production and many efforts have been made.

Until now, the development of whitening agents has been mainly focused on finding a substance that reduces melanin by inhibiting the activity of tyrosinase, a basic and most important enzyme in melanin biosynthesis. Recent studies on synthetic whitening agents have explored the use of alkoxy benzoates and alkoxy cinnamate derivatives, and hydroxamic acid. Other studies have shown derivatives of known tyrosinase inhibitors, such as 5-[(3-aninopropyl) phosphinooxy] -2- (hydroxymethyl) -4H-pyran-4-one ((5- Focus on the synthesis of ((3-aminopropyl) phophinooxy) -2- (hydroxymethyl) -4H-pyran-4-one)) and N-hydroxycinnamoylphenalkyl-amide derivatives there was.

The whitening agents developed in this way include kojic acid, arbutin, glutathione, vitamin A, and vitamin C, but they do not have a satisfactory whitening effect. Therefore, it is an urgent time to develop a whitening agent that is more effective and safer than ever.

Tyrosinase, an enzyme that synthesizes melanin, is a glycoprotein and is produced through glycosylation, a glycoprotein synthesis in vivo. If there is a problem in the glycosylation process and the sugar part of the tyrosinase is abnormal, the tyrosinase cannot move to the melanosome, which is an intracellular melanin biosynthesis site, and the melanin production cannot be achieved.

Many enzymes are involved in the glycosylation process, an important enzyme of which is glucosidase. If the enzyme can inhibit the enzymatic activity, it can inhibit the melanin biosynthesis and thus bring about a whitening effect.

In this regard, the conventional composition for skin whitening containing the actinomycetes culture extract of Korea Patent Application No. 10-2004-0025053 actinomycetes strains or actinomycetes culture extract tyrosinase is an enzyme involved in melanin biosynthesis Inhibits the enzymatic activity of β-glucosidase, an important enzyme involved in the process of glycosylation, thereby inhibiting glycosylation of tyrosinase and producing melanin. By inhibiting the fact that the whitening effect is shown.

Calcium trees are rosaceae trees native to China's Heilongjiang, Jilin, Liaoning and Inner Mongolia. Cherry fruits contain large amounts of calcium, iron, vitamins, amino acids, and essential minerals. It appears to have been named `` calcium tree '' when brought in. The tree is short and is called 'Chinese dwarf cherry' in the West.

Calcium TreePrunus humilis) Is a bush-like rose fruit, whose fruit is densely fruited from the bottom up like grapes, and its shape is similar to plum. It is the newest variety grown from wild plum trees in Europe. It belongs to deciduous shrubs. It is resistant to barrens and droughts, and can be cultivated nationwide from Jeju to northern Gangwon because of its cold resistance. Calcium is rich in pulp, sweet and sour, with a unique taste and aroma. Calcium trees bloom very beautifully in April-May, and the red-green berries (6-15g), which are slightly larger than the cherry trees ripening in July-September, have a similar taste to plums. Good for People in Inner Mongolia dry their leaves and drink them with tea.

The fruit of the calcium tree is a nutritional report. It contains 60 mg of active calcium, 47 mg of vitamin C, 1.5 mg of iron, and 400 mg of 17 amino acids per 100 g of fruit, and it contains a large amount of various trace elements. Varieties. The fruit contains not only a large amount of various nutrients, but also the absorption rate of calcium and iron in the human body is more than twice as high as milk, making it the ideal fruit for calcium supplementation and blood donation of children, examinees, mothers, and the elderly.

The fruit of the calcium tree not only has excellent taste and flavor, but also has a variety of useful values such as fruit juice, juice, fruit wine, 쨈, dried fruit, extract, and raw materials of medicines using the calcium fruit containing various functional ingredients. We raise high expectation as functional fruit tree.

The stems and leaves of the calcium tree grow well and tenderly, and contain high calcium and various nutrients, so it can be used as a functional grass feed of excellent quality. Yields are good 2 years after planting seedlings, fruit, leaves, stems, as well as beautiful flowers and good color of the fruit, gardening, military planting, eruption cultivation, bonsai material is ideal.

Looking at the effect of the calcium tree according to the recent scientific research results have been disclosed in Chinese Patent Publication No. 1010884072 regarding the hair health improvement effect.

Also, the dried fruit of the calcium tree is called Uriin, which is mainly made of seeds of Prunus humilis, Prunus japonica, Prunus pedunculata, and Prunus ishidoyana. It has been used as a medicine.

On the other hand, the domestic registered patent using Uk-riin, two patents used as a composition for preventing and treating diabetes of CJ CheilJedang Co., Ltd., one patent for promoting hair growth of KT & G, and chlorophyll a of Gwangju Institute of Science and Technology as active ingredients As for a composition for the treatment of atopic diseases, there is no example of verifying the whitening efficacy with only Uri, and there is no patent for this.

As described above, the present inventors searched for whitening efficacy through the inhibition of intracellular melanin production in various plant extracts in order to develop a new whitening material that is more effective and safer than the whitening agent so far, and the calcium tree extract has excellent whitening efficacy, This was found to be due to hexacosine dioxic acid (hexacosanedioic acid) to complete the present invention.

It is an object of the present invention to provide a pharmaceutical composition for preventing or treating skin hyperpigmentation disease caused by melanin excess production using calcium tree-derived hexacosanedioic acid as an active ingredient.

Another object of the present invention is to provide a health functional food composition for improving dermatological pigmentation disease caused by melanin overproduction containing calcium-derived hexacosanedioic acid as an active ingredient.

Still another object of the present invention is to provide a cosmetic composition for skin whitening containing hexacosanedioic acid derived from calcium wood as an active ingredient.

The above object of the present invention was achieved by preparing an extract from the calcium tree and again preparing an extract from the extract, deriving a fraction having excellent melanin production inhibitory effect among these fractions, and then identifying the chemical structure of the fraction.

The present invention provides a pharmaceutical composition for preventing or treating skin hyperpigmentation disease due to melanin overproduction, which is derived from calcium tree-derived hexacosanedioic acid of Formula 1 or a pharmaceutically acceptable salt thereof.

≪ Formula 1 >

The present invention provides a health functional food composition for preventing or improving skin hyperpigmented disease caused by melanin overproduction, which is derived from calcium hexacosanedioic acid of Formula 1 or a pharmaceutically acceptable salt thereof.

In the present invention, the term "active ingredient" includes a substance or a group of substances (a pharmacologically active ingredient or the like, which is expected to express the efficacy or effect of the drug directly or indirectly by intrinsic pharmacological action). ) Means containing the main component.

In the present invention, the health functional food means a food manufactured and processed using raw materials or ingredients having functional properties useful for the human body according to Act No. 6767 of the Health Functional Food Act, and functional means the structure and function of the human body. It means the ingestion for the purpose of obtaining a useful effect for health use such as nutrient control or physiological action.

The compound of formula 1 according to the present invention, by inhibiting melanin biosynthesis in the skin cells can increase the skin whitening activity, and at the same time inhibit the excessive formation of melanin, thereby preventing, improving or treating skin hyperpigmentation disease have.

In the present invention, the skin hyperpigmentation disease caused by the melanin hyperplasia is genetic skin diseases, such as alum, Badenburg syndrome, acquired skin diseases, for example, post-inflammatory white nasal mucus, unexplained water droplet melaninosis, For example, blemishes, skin diseases due to drug treatment, such as minocycline, bleomycin, busulfan, zidovudine, and skin diseases transmitted through infection, such as stumps.

Pharmaceutical compositions comprising a compound of formula 1 of the present invention may further comprise suitable carriers, excipients and diluents commonly used in the manufacture of pharmaceutical compositions.

Pharmaceutical compositions comprising the compound of formula 1 of the present invention, oral formulations, such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, external preparations, suppositories, and sterile injectable solutions, respectively, according to conventional methods Formulated in the form of can be used.

Carriers, excipients and diluents which may be included in pharmaceutical compositions comprising the compound of formula 1 of the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate , Gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. In the case of formulation, a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and such solid preparations include at least one excipient such as starch, calcium carbonate, sucrose in the extract. Or lactose, gelatin, or the like is mixed. In addition to simple excipients, lubricants such as magnesium stearate and 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 sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Examples of the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin and the like can be used.

The preferred dosage of the g compound of formula 1 according to the present invention depends on the condition and weight of the patient, the extent of the disease, the form of the drug, the route of administration and the duration, and may be appropriately selected by those skilled in the art. However, for the desired effect, the compound of formula 1 of the present invention is preferably administered at 0.0001 to 100 /, preferably 0.001 to 100 / day. The administration may be carried out once a day or divided into several times. The dose is not intended to limit the scope of the invention in any way.

The compound of formula 1 according to the present invention can be administered to mammals such as rats, mice, livestock, humans, etc. by various routes. All modes of administration can be expected, for example, oral, rectal or intravenous, intramuscular, subcutaneous, intrauterine dural or intracerebroventricular injections and directly to the skin.

In addition, the compound of formula 1 according to the present invention may be added to food or beverage for the purpose of preventing or improving skin hyperpigmentation disease caused by melanin overproduction. Examples of the food to which the compound of formula 1 may be added include various foods, beverages, gums, teas, vitamin complexes, health functional foods, powdered meals, granules, tablets, capsules or beverages. At this time, the amount of the compound of formula 1 in the food or beverage may be added in 0.01 to 15% by weight of the total food weight, the health beverage composition is added in a ratio of 0.02 to 5g, preferably 0.3 to 1g based on 100 Can be.

The food composition of the present invention is not particularly limited to other ingredients except for containing the compound of formula 1 as an essential ingredient in the indicated ratio, and may contain various flavors or natural carbohydrates as additional ingredients, such as ordinary beverages. Examples of the above-mentioned natural carbohydrates are monosaccharides such as glucose, fructose and the like and disaccharides such as maltose and sucrose and the like, and conventional sugars and xylitol such as polysaccharides such as dextrin and cyclodextrin, Sugar alcohols such as sorbitol and erythritol. As flavoring agents other than those mentioned above, natural flavoring agents, such as taumartin, stevia extracts, for example, rebaudioside A, glycyrrhizin, and the like, synthetic flavoring agents such as saccharin, aspartame and the like can be advantageously used. The ratio of the natural carbohydrate is generally about 1 to 20 g, preferably about 5 to 12 g per 100 mL of the composition of the present invention.

In addition, the compound of the formula (1) of the present invention is a variety of nutrients, vitamins, minerals (electrolytes), flavors such as synthetic and natural flavors, coloring and neutralizing agents (such as cheese, chocolate), pectic acid and salts thereof, alginic acid And salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerol, alcohols, carbonation agents used in carbonated beverages, and the like. The compound of formula 1 of the present invention may also contain pulp for the production of natural fruit juices, fruit juice drinks and vegetable drinks. These components can be used independently or in combination. The proportion of the additive is not so important, but the extract of the present invention is preferably selected in the range of 0 to about 20 parts by weight per 100 parts by weight.

In the cosmetic composition of the present invention, the composition is characterized in that the formulation of the lotion (skin lotion), lotion, cream, massage cream, essence, pack. The formulations may be prepared using methods known in the art. In addition, the content of the compound of formula 1 in the formulation may be included in 1-10% by weight within the content range known in the art.

Components included in the cosmetic composition of the present invention may include components commonly used in cosmetic compositions in addition to the compound of Formula 1 as an active ingredient, for example, such as stabilizers, solubilizers, vitamins, pigments and flavorings Phosphorus aids and carriers.

In the present invention, 100% ethanol extract of calcium tree (leaf, stem, fruit) raw material and calcium tree (leaf, stem, fruit) are lyophilized, and then 70% ethanol extract of lyophilized calcium tree is prepared, respectively, the two extracts. The skin whitening activity of was evaluated by the following method.

Experimental Example 1 Whitening Activity of Calcium Tree Extracts

Melan-a cells were seeded at 210 ⁵ cells / well in 12 well plates and incubated in cell culture conditions until the cells were about 80% adhered to the bottom of the well. Thereafter, the medium was removed, and the test substance was diluted in RPMI serum free media, treated with cells, and cultured for 72 hours. After removing the medium of the cultured cells and washed with PBS, 200L of 1M NaOH containing 10% DMSO was added to incubate for 1 hour in a 60 ℃ water bath to obtain intracellular melanin. This solution was measured by absorbance at 490nm with a microplate reader to determine the amount of melanin. This solution was again subjected to a bradford assay to determine the total protein amount and to determine the amount of melanin per constant protein. The group treated with the culture medium containing no sample was set as a control group, and the degree of inhibition of melanin production was measured as a percentage. Positive control used 100 uM N-Phenylthiourea (Fig. 1).

As a result of the experiment, 70% ethanol extract of calcium tree lyophilized extract was excellent in inhibiting melanin biosynthesis, and each extract sample was analyzed for effective ingredients by comparing the components through LC-MS analysis (Tables 1 to 5 and FIGS. 5).

[Table 1] LC-MS Analysis Conditions

[Table 2] Sample analysis DBase search result table in positive mode of raw calcium wood sample

[Table 3] Sample analysis DBase search result table in negative mode of raw calcium wood sample

TABLE 4 Mass formula DB analysis results in positive mode of calcium wood lyophilized samples.

[Table 5] Mass formula DB analysis results in negative mode of calcium wood lyophilized samples.

LC-MS results of 100% ethanol extracts of raw calcium and 70% ethanol extracts of calcium lyophilized samples were investigated by dBase of the Kyung Hee University Skin Biotechnology Center. Hexacosanedioic acid in ESI negative mode (TR = 28.937min mass = 426.3709, matching score = 94.79%) and GPEtn (18: 0/18: 3) (Phosphatidylethanolamine, TR = 12.83min, mass = 741.5305, matching score = 99.59%).

Fractions of hexacosanedioic acid from lyophilized samples of calcium wood were collected by HPLC under the same conditions as LC-MS and designated as H_fraction. The fractions were subjected to melanin biosynthesis inhibition test and NMR. It showed the same whitening activity as 70% ethanol extract of dried product, and its structure was identified to be hexacosanedioic acid.

As described above, hexacosidioic acid, a calcium-derived compound according to the present invention, has an excellent melanin inhibitory effect, and thus prevents, treats, or improves skin hyperpigmentation diseases caused by melanin overproduction. Since it is an excellent effect, it is a very useful invention for the pharmaceutical industry and the cosmetic industry.

1 is a graph showing the results of evaluation of melanin biosynthesis of 100% ethanol extract of calcium wood raw material and 70% ethanol extract of calcium wood lyophilized sample and H fraction of HPLC.
Figure 2 is a chromatogram of the ESI positive result of LC-MS of 100% ethanol extract of the raw calcium wood sample.
Figure 3 is a chromatogram of the ESI Negative result of LC-MS of 100% ethynol extract of raw calcium wood samples.
Figure 4 is a chromatogram of ESI positive results of LC-MS of 70% ethynol extract of calcium wood lyophilized samples.
5 is an ESI Negative chromatogram of LC-MS of 70% ethynol extract of calcium wood lyophilized sample.

Hereinafter, preferred embodiments of the present invention will be described in more detail with reference to Examples. However, the scope of the present invention is not limited to these examples.

Example 1 Calcium Tree Extract Preparation and Separation of Active Substance

5 kg of calcium powder (Prunus humilis) dry powder was extracted twice with 70% aqueous ethanol (18 l x 2) and then filtered through a filter paper. The filtrate was concentrated under reduced pressure at 45 to obtain an EtOH extract. The obtained EtOH extract was separated by HPLC on the same column as LC-MS, and the time was determined so that a mass value of 426 of TR = 28.937min was collected. It was confirmed by LC-MS whether each fraction was fractionated with the same components, and when the concentration of the calcium tree extract showed the whitening efficacy, the whitening efficacy test was performed by the following method.

<Experimental Example 2> Gamelan -a (melan -a) melanogenesis inhibitory effect using a cell

Melan-a cells were seeded at 210 ⁵ cells / well in 12 well plates and incubated in cell culture conditions until the cells were about 80% adhered to the bottom of the well. Thereafter, the medium was removed, and the test substance was diluted in RPMI serum free media, treated with cells, and cultured for 72 hours. After removing the medium of the cultured cells and washed with PBS, 200L of 1M NaOH containing 10% DMSO was added to incubate for 1 hour in a 60 ℃ water bath to obtain intracellular melanin. This solution was measured by absorbance at 490nm with a microplate reader to determine the amount of melanin. This solution was again subjected to a bradford assay to determine the total protein amount and to determine the amount of melanin per constant protein. The group treated with the culture medium containing no sample was set as a control group, and the degree of inhibition of melanin production was measured as a percentage. Positive control was 100 uM N-Phenylthiourea.

As a result, as shown in Figure 1, the calcium fraction _H fraction showed an enzyme activity inhibitory effect.

Experimental Example 3 LC - MS of Extracts Analysis and Dbase search

As shown in FIG. 4 and FIG. 5, each extract was analyzed under the conditions shown in Table 1, and a fraction of 28.937 for the retention time of mass 426 was searched and its structure was confirmed as follows.

The structure of Fraction_H was the same as Compound 1 below. Compound of the Invention 1 [Hexacosanedioic acid]: colorless oil, EI / MS m / z : 426 [M]; 1 H-NMR (400 MHz, CDCL ₃ , _H ) :, 11.20 (2H, carboxlic acidx2), 2.30 (4H, -C (= O) -CH2, carboxlic acid adjacent methylene x2) 1.52-1.26 (42H, CH ₂ 21 ),; ¹³ C-NMR (100 MHz, CDCl ₃ , c): 178.4 (2C, carboxlic acid-C), 34.0, (2C), 29.6 (16C), 29.3 (2C), 29.0 (2C), 24.7 (2C),

&Lt; Formula 1 >

The present invention has the effect of obtaining hexacosine diioic acid extracted and purified from calcium wood, and has the effect of providing a pharmaceutical composition for preventing and treating skin hyperpigmented diseases comprising the compound and providing a functional food for preventing and improving the disease. As well as excellent effect of providing a cosmetic composition for skin whitening is a useful invention in the biological industry.

Claims (3)

A pharmaceutical composition for preventing or treating skin hyperpigmentation disease caused by melanin excess production containing hexacosine diioic acid (Hexacosanedioic acid) of Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.
&Lt; Formula 1 >

Health functional food composition for preventing or improving skin hyperpigmentation disease caused by melanin excess production containing hexacosane diioic acid (Hexacosanedioic acid) as an active ingredient of claim 1 A cosmetic composition for skin whitening containing hexacosine diioic acid (Hexacosanedioic acid) as an active ingredient according to claim 1 as an active ingredient.

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