KR102159607B1 - Composition for skin whitening or anti-inflammatory comprising extract of fermented Angelica gigas with improved antioxidant activity as effective component - Google Patents

Abstract

The present invention relates to a skin whitening or anti-inflammatory composition containing a fermented Angelicae extract with enhanced antioxidant activity as an active ingredient, and the fermented Angelicae extract of the present invention has excellent antioxidant, whitening and anti-inflammatory effects, for skin whitening It can be safely and usefully used in health functional foods and pharmaceuticals for the prevention, improvement or treatment of cosmetics, melanin hyperpigmentation diseases or inflammatory diseases.

Description

Composition for skin whitening or anti-inflammatory comprising extract of fermented Angelica gigas with improved antioxidant activity as effective component}

The present invention relates to a skin whitening or anti-inflammatory composition containing a fermented Angelicae extract with enhanced antioxidant activity as an active ingredient.

Recently, it is necessary to develop high value-added beauty products that can meet new expectations with the increase in knowledge and awareness of cosmetics by domestic and foreign consumers. In particular, herbal and fermented cosmetics, which have already become global trends, aim for well-being and naturalism, satisfy consumers' desires for preferring natural ingredients, and at the same time apply an oriental sentiment to the extent that most cosmetic companies have one or more brands per distribution. It is building its own domain. Until only a few years ago, synthetic compounds were the main ingredients for cosmetics manufacturing, but the domestic cosmetic ingredient market is also growing as the use of high value-added ingredients increases due to the expansion of the natural and functional cosmetics market.

The domestic cosmetics industry is showing a trade surplus thanks to overseas expansion, mainly in Asia, and the development of functional cosmetics and natural vegetable cosmetics. However, since 78% of the raw materials used in manufacturing depend on imported raw materials, it is time to lower the degree of dependence on imports. Consumption of products with enhanced functionality is increasing, and research institutes and companies, including consumers, are increasingly interested in developing functional materials with natural physiological activity.

In all living organisms, reactive oxygen species are produced by biochemical reactions and environmental factors in the living body, and the increased production of reactive oxygen species in the living body causes oxidative stress, which may functionally damage intracellular proteins and lipid components. It is also the cause of various diseases such as aging, heart disease, and diabetic arthritis. Natural antioxidants that delay or inhibit oxidative damage to cellular components caused by oxidative stress and functional materials using fermentation have been actively developed.

Angelica gigas is a perennial herb belonging to the Umbelliferae family. It is mainly distributed in Korea, China, and Japan, and is cultivated in each region and used as a herbal material. Angelica grown in our country has been separated from the Chinese angelica (Angelica acutiloba Kitagaw), Japanese angelica (Angelica sinensis Diels). Angelica is a representative herbal medicine in Korea that has been used for disease treatment and health promotion as a herbal medicine with no side effects due to its warm medicinal properties, sweet taste, and non-toxicity. The medicinal ingredients of Angelica Angelica grown in Korea include decursin, decursinol, angelate, nodakenetin, nodakenin, umbelliferone, It contains β-sitosterol, α-pinene, and limonene. In particular, decursin and decursinol angelate are the most abundant constituents among the coumarine derivatives, which are the main components of angelica, and are known for their pharmacological effects such as antibacterial, anti-angiogenic, and anti-cancer.

Plant extracts such as angelica and mulberry leaves, which are used in the development of functional foods or functional cosmetics, have excellent physiological activity, but they often exhibit toxic effects. Recently, fermentation with microorganisms or mushroom mycelium reduces the toxicity of the extract, and at the same time, it is confirmed that a new substance is generated by fermentation or an existing physiologically active substance is converted into another substance, thereby having a different effect than the first. Research related to improvement is actively underway.

On the other hand, Korean Patent No. 1404168 discloses'a pharmaceutical composition for preventing or treating thrombosis containing an extract of Cham Dang-gu as an active ingredient', and Korean Patent No. 1870143 discloses'a pharmaceutical composition containing jujube seed extract as an active ingredient. Skin whitening or anti-inflammatory composition' is disclosed, but there is no description of a skin whitening or anti-inflammatory composition containing the fermented Angelicae extract with enhanced antioxidant activity of the present invention as an active ingredient.

The present invention was derived from the above requirements, and the present inventors prepared a fermented Angelica gigas extract by inoculating and fermenting Angelica gigas powder with Saccharomyces cerevisiae and then extracting with hot water, The present invention was completed by confirming that the prepared fermented Angelicae extract had superior DPPH radical scavenging activity, tyrosinase inhibitory activity, and lipoxygenase inhibitory activity, which is an inflammation-inducing factor, compared to the non-fermented Angelicae extract.

In order to solve the above problems, the present invention provides a cosmetic composition for skin whitening comprising a fermented Angelicae extract with enhanced antioxidant activity as an active ingredient.

In addition, the present invention provides a health functional food composition for preventing or improving melanin hyperpigmentation disease comprising a fermented angelica extract as an active ingredient.

In addition, the present invention provides a pharmaceutical composition for the prevention or treatment of melanin hyperpigmentation disease comprising a fermented angelica extract as an active ingredient.

In addition, the present invention provides a health functional food composition for preventing or improving inflammatory diseases comprising a fermented angelica extract as an active ingredient.

In addition, the present invention provides a pharmaceutical composition for the prevention or treatment of inflammatory diseases comprising a fermented angelica extract as an active ingredient.

The fermented Angelicae extract of the present invention is a natural substance that is safe for the human body and has no side effects even when used for a long period of time, so functional cosmetics for skin whitening, foods or treatments that can prevent or improve skin pigmentation disorders, and for improving inflammation It can be usefully used as a food and therapeutic agent.

1 is a thin-layer chromatography analyzing the content of monosaccharides and polysaccharides in non-fermented Angelicae extract (NF), Angelicae extract fermented for 0 days (F/0), and Angelicae extract fermented for 3 days (F/3) (Thin- layer chromatography, TLC) results. STD (standard); Standard malto-oligosaccharide (G1-G7), G1; Glucose, G2; Maltose, G3; Maltotriose, G4; Maltotetraose, G5; Maltopentaose, G6; Maltohexaose and G7; Maltoheptaose. NF; Saccharomyces cerevisiae ) extract of Angelicae powder without mixing strain, F/0; Extract immediately after mixing non-fermented Angelicae and Saccharomyces cerevisiae strains.
Figure 2 is a non-fermented Angelicae extract (NF), an extract of Angelicae fermented for 0 days (F/0), an extract of Angelicae fermented for 1-5 days (F/1, F/2, F/3, F/4, This is the result of measuring DPPH radical scavenging activity of F/5). BHT (butylated hydroxytoluene 0.05%); Positive control.
Figure 3 is a measurement of the tyrosinase inhibition activity (tyrosinase inhibition activity) of non-fermented Angelicae extract (NF), Angelicae extract fermented for 0 days (F/0), and Angelicae extract fermented for 3 days (F/3) It is the result. Arbutin (0.05%); Positive control.
Figure 4 is a measurement of the lipoxygenase inhibition activity of non-fermented Angelicae extract (NF), Angelicae extract fermented for 0 days (F/0), and Angelicae extract fermented for 3 days (F/3). It is the result. indomethacin (0.05%); Positive control.

In order to achieve the object of the present invention, the present invention provides a cosmetic composition for skin whitening comprising a fermented Angelicae extract with enhanced antioxidant activity as an active ingredient.

In the cosmetic composition for skin whitening according to an embodiment of the present invention, the fermented Angelica may be fermented by inoculating yeast bacteria in Angelica gigas powder, and preferably Saccharomyces cerevisiae in Angelica gigas powder ( Saccharomyces cerevisiae ) may be inoculated and fermented for 2 to 4 days, preferably 3 days, but is not limited thereto.

The fermented Angelicae extract may be extracted using water, a lower alcohol having 1 to 4 carbon atoms, or a mixture thereof as an extraction solvent, and preferably extracted using water as an extraction solvent, but is not limited thereto.

The fermented Angelicae extract according to the present invention has a higher content of total polyphenols and flavonoids compared to the non-fermented Angelicae extract, and has excellent DPPH radical scavenging activity, as well as reduced tyrosinase production, which is involved in the production of melanin. It is effective in whitening.

The cosmetic composition for skin whitening of the present invention includes skin, skin softener, skin toner, lotion, milk lotion, moisture lotion, nutrition lotion, massage cream, nutrition cream, eye cream, moisture cream, hand cream, essence, nutrition essence, A pack, cleansing foam, cleansing water, cleansing cream, body lotion, body cleanser, soap, and powder may have any one formulation selected from, but is not limited thereto. The cosmetic composition composed of each of these formulations may contain various bases and additives necessary and appropriate for the formulation of the formulation, and the types and amounts of these components can be easily selected by those skilled in the art.

When the formulation of the cosmetic composition of the present invention is a paste, cream or gel, animal fibers, plant fibers, wax, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide as carrier components Etc. may be used.

When the formulation of the cosmetic composition of the present invention is a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component. In particular, in the case of a spray, additionally chlorofluorohydro Propellants such as carbon, propane-butane or dimethyl ether.

When the formulation of the cosmetic composition of the present invention is a solution or emulsion, a solvent, a solvating agent or an emulsifying agent is used as a carrier component, such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene Glycol, 1,3-butylglycol oil, glycerol aliphatic ester, polyethylene glycol or fatty acid ester of sorbitan.

When the formulation of the cosmetic composition of the present invention is a suspension, as a carrier component, a liquid diluent such as water, ethanol or propylene glycol, an ethoxylated isostearyl alcohol, a suspending agent such as polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, Microcrystalline cellulose, aluminum metahydroxide, bentonite, agar or tracant, and the like may be used.

When the formulation of the cosmetic composition of the present invention is a surfactant containing cleansing, as carrier components, aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, acetionate, imidazolinium derivative, methyltaurate, sarcosinate , Fatty acid amide ether sulfate, alkylamidobetaine, aliphatic alcohol, fatty acid glyceride, fatty diethanolamide, vegetable oil, linoline derivative or ethoxylated glycerol fatty acid ester, and the like may be used.

The cosmetic composition of the present invention may further contain excipients including fluorescent substances, fungicides, hydrophobic substances, moisturizers, fragrances, fragrance carriers, proteins, solubilizers, sugar derivatives, sunscreen agents, vitamins, plant extracts, etc. .

The present invention also provides a health functional food composition for preventing or improving melanin hyperpigmentation disease comprising a fermented angelica extract as an active ingredient.

The health functional food composition of the present invention has a higher content of total polyphenols and flavonoids than non-fermented Angelicae extract, and increased DPPH radical scavenging activity and inhibitory activity of tyrosinase involved in melanogenesis. As an active ingredient, it is effective against melanin hyperpigmentation diseases.

In the present invention, the term "hyperpigmentation" means that a specific area of the skin or nails or nails becomes blacker or darker than other areas due to an excessive increase in melanin. The melanin hyperpigmentation disorders include freckles, senile spots, liver spots, melasma, brown or dark spots, solar pigment spots, cyanicmelasma, hyperpigmentation after drug use, gravidic chloasma, or scratches and burns. It may be hyperpigmentation after inflammation due to wounds or dermatitis including, but is not limited thereto.

The health functional food composition for preventing or improving the melanin hyperpigmentation disease may be prepared in any one formulation selected from powders, granules, pills, tablets, capsules, candy, syrup, effervescent tablets, and beverages, but is not limited thereto. . The health functional food composition is not particularly limited as long as it can be ingested to prevent or improve melanin hyperpigmentation disease.

When the fermented Angelicae extract of the present invention is used as a food additive, the fermented Angelicae extract 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 active ingredient may be appropriately used depending on the purpose of use (prevention or improvement). In general, in the manufacture of food or beverage, the fermented Angelicae extract 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, in the case of long-term intake for the purpose of health control, it can be used in an amount within a range without any problems in terms of safety.

There is no particular limitation on the type of the food. Examples of foods to which the health functional food composition 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 There are drinks, alcoholic beverages, and vitamin complexes, and all health foods in the usual sense are included. The beverage includes carbonated beverages, functional ionic beverages, juices (eg, apples, pears, grapes, aloe, tangerines, peaches, carrots, tomato juice, etc.), sikhye, and the like.

Functional foods of the present invention include ingredients that are commonly added during food production, and include, for example, proteins, carbohydrates, fats, nutrients, and seasonings. For example, when prepared as a drink, natural carbohydrates or flavoring agents may be included as additional ingredients in addition to the active ingredient. The natural carbohydrates are monosaccharides (eg, glucose, fructose, etc.), disaccharides (eg, maltose, sucrose, etc.), oligosaccharides, polysaccharides (eg, dextrins, cyclodextrins, etc.) or sugar alcohols (eg , Xylitol, sorbitol, erythritol, etc.). The flavoring agent may be a natural flavoring agent (eg, taumatin, stevia extract, etc.) and a synthetic flavoring agent (eg, saccharin, aspartame, etc.). In addition to the health functional food composition, various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonic acid It may further contain a carbonation agent used in beverages.

In addition to the above health functional food composition, various nutrients, vitamins, inhibitors, flavoring agents, coloring agents, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, It may further contain a carbonating agent used in carbonated beverages. Although the ratio of the ingredients to be added is not very important, it is generally selected from 0.01 to 0.1 parts by weight based on 100 parts by weight of the health functional food composition of the present invention.

The present invention also provides a pharmaceutical composition for the prevention or treatment of melanin hyperpigmentation disease comprising a fermented angelica extract as an active ingredient.

The melanin hyperpigmentation disorders include freckles, senile spots, liver spots, melasma, brown or dark spots, solar pigment spots, cyanicmelasma, hyperpigmentation after drug use, gravidic chloasma, or scratches and burns. It may be hyperpigmentation after inflammation due to wounds or dermatitis including, but is not limited thereto.

The pharmaceutical composition of the present invention is effective in a fermented Angelicae extract having a higher content of total polyphenols and flavonoids compared to the non-fermented Angelicae extract and increased DPPH radical scavenging activity and inhibitory activity of tyrosinase involved in melanogenesis. It is included as an ingredient, and is effective against melanin hyperpigmentation diseases.

The pharmaceutical composition comprising the fermented Angelicae extract of the present invention may further include a suitable carrier, excipient, or diluent commonly used in the preparation of a pharmaceutical composition.

The pharmaceutical dosage form of the pharmaceutical composition of the present invention may be used alone or in combination with other pharmaceutically active compounds, as well as in an appropriate combination.

The pharmaceutical composition according to the present invention may be formulated and used in the form of oral preparations such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc., external preparations, suppositories, and injections according to a conventional method. . Carriers, excipients and diluents that may be included in the pharmaceutical composition include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose , Methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oils. In the case of formulation, it is prepared using diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrants, and surfactants that are usually used. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and these solid preparations contain at least one excipient, such as starch, calcium carbonate, sucrose or lactose, gelatin, etc. in the fermented product. It is prepared by mixing. 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, syrups, and other excipients, such as water and liquid paraffin, which are commonly used simple diluents, such as wetting agents, sweeteners, fragrances, and preservatives. have. Preparations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized preparations, and suppositories. Propylene glycol, polyethylene glycol, vegetable oils such as olive oil, injectable esters such as ethyl oleate, etc. may be used as the non-aqueous solvent and suspension. As a base for suppositories, Witepsol, Macrogol, Tween 61, cacao butter, laurin paper, glycerogelatin, and the like can be used.

The preferred dosage of the pharmaceutical composition of the present invention varies depending on the condition and weight of the patient, the degree of disease, the form of the drug, the route of administration and the duration, but may be appropriately selected by those skilled in the art. The pharmaceutical composition of the present invention can be administered to mammals such as mice, mice, livestock, and humans by various routes. All modes of administration can be expected and may be administered by, for example, oral, rectal or intravenous, intramuscular, subcutaneous, intrauterine dura mater or intracerebroventricular injection.

The present invention also provides a health functional food composition for preventing or improving inflammatory diseases comprising a fermented Angelicae extract as an active ingredient.

The health functional food composition of the present invention contains, as an active ingredient, fermented Angelicae extract having an increased inhibitory activity of lipoxugenase, which is an inflammation-inducing factor, and is effective against inflammatory diseases. In addition, the fermented Angelicae extract according to the present invention has excellent antioxidant effect.

The health functional food composition for preventing or improving inflammatory diseases may be prepared in any one formulation selected from powders, granules, pills, tablets, capsules, candy, syrup, effervescent tablets, and beverages, but is not limited thereto. The health functional food composition is not particularly limited as long as it can be consumed to increase anti-inflammatory activity.

When the fermented Angelicae extract of the present invention is used as a food additive, the types of food and ingredients commonly added during food production are as described above.

The present invention also provides a pharmaceutical composition for the prevention or treatment of inflammatory diseases comprising a fermented Angelicae extract as an active ingredient.

In the pharmaceutical composition for preventing or treating inflammatory diseases of the present invention, the inflammatory diseases are atopic dermatitis, edema, dermatitis, allergy, asthma, conjunctivitis, periodontitis, rhinitis, otitis media, pharyngitis, tonsillitis, pneumonia, gastric ulcer, gastritis, Crohn's disease, colitis , Hemorrhoids, gout, hepatic spondylitis, rheumatoid fever, lupus, fibromyalgia, psoriatic arthritis, osteoarthritis, rheumatoid arthritis, peri-shoulderitis, tendinitis, tendonitis, myositis, hepatitis, cystitis, nephritis, sjogren's syndrome and multiple It may be selected from the group consisting of sclerosis, but is not limited thereto.

The pharmaceutical composition of the present invention contains, as an active ingredient, fermented Angelicae extract having increased inhibitory activity of lipoxugenase, which is an inflammatory factor, as an active ingredient, and is effective against inflammatory diseases. In addition, the fermented Angelicae extract according to the present invention has excellent antioxidant effect.

The pharmaceutical composition comprising the fermented Angelicae extract of the present invention may further include a suitable carrier, excipient, or diluent commonly used in the preparation of a pharmaceutical composition, and may be included in a pharmaceutical composition comprising an extract, an excipient, and The diluent is as described above.

The pharmaceutical dosage form of the pharmaceutical composition of the present invention may be used alone or in combination with other pharmaceutically active compounds, as well as in an appropriate combination, and the preferred dosage of the pharmaceutical composition of the present invention is as described above.

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.

Materials and methods

1. Experimental materials

Angelica gigas was purchased from the Jecheonhwanggi Union Farming Association (Jecheon-si, Chungcheongbuk-do) and dried the roots of angelica and used as a sample. Fermented strain three Mai as a kind of yeast Saccharomyces process Levy jiae (Saccharomyces cerevisiae ) M1 (KCTC 10121BP) was selected and used, and the strain pre-cultured in Yeast Malt broth (Difco) medium was inoculated at a 1:1 weight ratio to Angelicae powder, and then fermented at 37°C for 48 hours and then at 60°C for 8 hours. During hot air drying.

2. Extraction Conditions and Yield of Fermented Angelicae

The water-soluble extract was refluxed three times for 3 hours at room temperature after adding 20 times of distilled water to 500 g of unfermented angelica and dried angelica powder (Toyo 2A; Toyo Roshi). The extract was collected and extracted with Whatman No. , Japan) and concentrated at 40°C with a rotary vacuum concentrator (Buchi Rotavapor R-215, Flawil, Switzerland) to obtain a crude extract from which the solvent is completely removed, and then freeze-drier (Eyela FUD-2100, Tokyo Rikakikai Co., Japan). ) To obtain the extraction yield (%) by lyophilization. The experimental group was performed by setting the extract of Angelicae (NF) that was not fermented, the extract of Angelicae on Day 0 of fermentation (F/0), and the Extract of Angelicae on Day 3 (F/3) of fermentation. The extract of the fermented Angelicae on Day 0 is an extract extracted immediately after mixing non-fermented Angelicae and Saccharomyces cerevisiae strains.

3. Soluble solids (° Brix ) And pH

As for the soluble solid, 1.5 ml of each extract sample was taken, centrifuged at 2,000 x g for 10 minutes, and the supernatant was measured using a sugar content meter (ATAGO, PR-10, Japan), and the pH was measured with a pH meter (Satrious, PB-101, Germany) was measured three times.

4. Thin layer chromatography (Thin-layer chromatography, TLC)

Thin-layer chromatography was performed to confirm the polysaccharide in the non-fermented Angelicae extract and the fermented Angelicae extract. Thin layer chromatography was performed by dropping the extracts on a silica gel plate (25 DC-Alufolien Kieselgel 60, Merck Co., Ltd.), developing with 1-butanol/ethanol/water (5:5:3) as a developing solvent, and then silica gel. The plate was completely dried, and a color developing solution prepared by adding 3% sulfuric acid to methanol was sprayed, and then placed on a hot plate of 100°C or higher and heated for 10 minutes. The standard substance used at this time is standard malto-oligosaccharide (G1-G7), and G1, G2, G3, G4, G5, G6 and G7 are glucose, maltose, and maltotriose, respectively. ), maltotetraose, maltopentaose, maltohexaose and maltoheptaose.

5. Measurement of phenolic compound content

The content of phenolic compounds was measured by the Folin-Denis method using the principle that a phenolic substance reacts with phosphormolybdic acid to give a blue color. To 0.5 ml of a 0.1% (w/v) sample solution, 2.5 ml of a Folin-ciocalteu phenol solution was added, mixed, and allowed to stand at room temperature for 5 minutes. After reacting for exactly 5 minutes, 2 ml of 7.5% sodium carbonate (Na ₂ CO ₃ ) was added and mixed, followed by color development at 50°C for 5 minutes, and then 760 nm using a spectrophotometer (Hitachi U-2900, Hitachi High-Technologies Co., Japan). Absorbance was measured at. At this time, the content of phenolic compounds was calculated from a standard curve prepared by measuring tannic acid at a concentration of 0 to 500 ㎍/㎖ in the same manner as in the sample.

6. Flavonoid content measurement

To measure the flavonoid content, 1.25 ml of purified water and 5 ml of 5% sodium nitrite (NaNO ₂ ) solution were added to 0.25 ml of 0.1% (w/v) sample solution. After 5 minutes, 10% aluminum chloride hexahydrate ( 0.15 ml of AlCl ₃ ·6H ₂ O) was mixed, and the absorbance of the mixture was measured at 510 nm using a spectrophotometer. At this time, the flavonoid content was calculated from the standard curve prepared by measuring (+)-catechin hydrate as a standard substance at a concentration of 20 to 200 µg/ml in the same manner as the sample.

7. Measurement of mineral content

The mineral content of the leaves and stem mixture of Angelicae and the dried root powder were measured according to the AOAC (Association of Official Analytical Chemists) analysis method. Accurately take 1 g of the dry powder and incinerate each in an incinerator at 550° C. for 3 hours, dissolve in 6N hydrochloric acid (HCl) to completely decompose acid, completely remove the acid in a water bath, add 3N hydrochloric acid to the dried material, and filter it. Accordingly, each was diluted at a certain ratio and measured using an atomic absorption spectrophotometer (Analyst 300, Perkin Elmer, Norwalk CT, USA).

8. Deckersin ( decursin ) And Deckercinol Angelate ( decursinol angelate ) Content measurement

After reflux extraction three times for 3 hours with 95% ethanol, which is 20 times the weight of the dried Angelicae or the dried powder of fermented Angelicae, filtered through a Sartorius Minisart 0.45 μm filter (Sartorius Stedim, Germany) and filtered with a vacuum concentrator (Buchi Rotavapor). R-215, Flawil, Switzerland). The concentrated sample was dissolved in methanol, filtered through a Sartorius Minisart 0.2 μm filter, and used as an analysis sample. The sample dissolved in methanol was HPLC (Agilent 1200, Agilent Technologies, USA), the column was a Waters symmetry C18 column (4.5 × 250 mm, 5 μm), a temperature of 25° C., a mobile phase rate of 1 ml/min, and a UV wavelength of 340 nm, The analysis solvents were solvent A (purified water) and solvent B (acetonitrile), and the analysis conditions were gradient analysis with 0 to 100% of solvent B until 0 to 20 minutes (Table 1).

HPLC analysis conditions Time (minutes) Flow rate (ml/min) Solvent A (%) Solvent B (%) 0 1.0 50 50 10 1.0 50 50 11 1.0 70 30 31 1.0 70 30 32 1.0 100 0 42 1.0 100 0 43 1.0 0 100 53 1.0 0 100 54 1.0 50 50 70 1.0 50 50

9. DPPH Measurement of antioxidant activity by

To measure the antioxidant activity, 16 mg of DPPH (α,α'-diphenyl-β-picrylhydrazyl) was dissolved in 100 ml of ethanol, and 100 ml of distilled water was mixed. Whatman filter paper NO. Filtered through 2 to prepare a DPPH reaction solution. After making 1 ml of non-fermented Angelicae extract or fermented Angelicae extract to a concentration of 1%, 5 mL of DPPH reaction solution was added and mixed, reacted in the dark for 30 minutes, and absorbance was measured at 528 nm using a spectrophotometer. At this time, the absorbance was measured in the same manner as the sample by adding BHT, a synthetic antioxidant, which has been widely used as an antioxidant, at a concentration of 0.05%. Antioxidant activity using DPPH was expressed as a percentage (%) of the difference in absorbance between the sample added and the non-added group. The equation for obtaining the DPPH radical scavenging activity is as follows; DPPH radical scavenging activity (%)=[1-(sample absorbance _{528 nm} )/control absorbance _{528 nm} ]×100.

10. Tyrosinase ( tyrosinase ) Activity measurement

To measure the in vitro mushroom tyrosinase activity inhibitory activity, 2.5mM L-DOPA (3,4-dihydroxyphenylalanine) 0.3ml, 1% concentration of non-fermented Angelicae extract or 0.05 ml of fermented angelica extract and 0.1M phosphate buffer solution (pH 6.8) were mixed and reacted at 25°C, and then 1,380 units/ml mushroom tyrosinase (2,500 units, Sigma Chemical Co., St. USA) 0.05 ml was added. The absorbance was measured and calculated at 470 nm using a spectrophotometer while reacting at °C for 2 minutes. As a control, a sample dissolution solution containing no sample was used, and as a positive control, 0.6mM arbutin, known as a tyrosinase inhibitor, was used. The formula for obtaining the tyrosinase inhibitory activity is as follows.; Tyrosinase activity inhibition rate (%)=100-[(AB)/A] X 100(A; difference in absorbance between 0.5 and 1 min of the reaction solution without sample, B; 0.5 to 1 of the reaction solution containing sample Absorbance difference between minutes).

11. Lipoxygenase ( lipoxygenase ) Inhibitory activity

For measurement of lipoxygenase activity, 10 µl of non-fermented Angelicae extract or fermented Angelicae extract at a concentration of 1% was added to a 96-well corning plate, and 20 µl of borate buffer was added, followed by linoleic acid. ) Was added and the absorbance value was measured before the lipoxygenase was added, and then 10 μl of lipoxygenase was added and reacted at room temperature for 10 minutes, and the absorbance was measured. The absorbance was measured at 234 nm using a spectrophotometer, and the positive control was used by dissolving indomethacin (Sigma) in ethanol at 0.05% (w/v).

12. Statistical processing

The results obtained from the above experiment were expressed as the mean value and standard deviation (mean±SE) by one-way ANOVA test.

Example 1. Yield, Soluble Solids, and pH of Fermented Angelicae Extract

Extract of non-fermented Angelicae (hereinafter referred to as NF), extract immediately extracted after mixing strains of non-fermented Angelicae and Saccharomyces cerevisiae (hereinafter, F/0), extract of Angelicae fermented for 3 days ( Hereinafter, the yield, soluble solids and pH of F/3) were analyzed.

As a result, the yield was 87.62% in the NF group, 53% in the F/0 group, and 54% in the F/3 group. Although the yield of the NF group was higher than that of the fermented Angelicae extract, it was confirmed that the yield increased as the fermentation period increased (Table 1), which is the fermentation strain, Saccharomyces cerevisiae , which is a nutrient of Angelicae. It was considered to be a part of environmental change that occurs through proliferation based on.

In addition, soluble solids showed a slight increase in sugar content due to fermentation treatment in the NF group to 1°Brix and in the F/0 group to 1.5°Brix, but as the fermentation proceeded to 0.9°Brix in the F/3 group, It was confirmed that it decreased. It was found that the pH increased as the number of fermentation days increased (Table 2).

Yield, soluble solids and pH yield(%) Soluble Solids (°Brix) pH NF 87.62±0.12 ^a 1.00±0.00 ^a 5.18±0.00 ^a F/0 54.00±0.40 ^b 1.50±0.02 ^b 4.90±0.02 ^b F/3 66.67±0.37 ^c 0.90±0.01 ^a 6.22±0.01 ^c

Example 2. Analysis of Monosaccharide and Polysaccharide Content of Fermented Angelicae Extract

Malto-oligosaccharides Standard G1-G7 (G1; glucose, G2; maltose, G3; maltotriose, G4; maltotetraose, G5; maltopentase, G6; maltohexase, G7; maltoheptaose) ), a 10% aqueous solution of non-fermented Angelicae powder and fermented Angelicae powder was developed on a TLC plate, and the contents of monosaccharides and polysaccharides in the non-fermented Angelicae extract and the fermented Angelicae extract were analyzed.

As a result, it was confirmed that G2, which showed a dark spot in the NF group, was mostly decomposed into the low-molecular sugar form of G1 in the F/0 group, but G3 was generated in the F/3 group (FIG. 1). Through this, it was found that the content of oligosaccharides can be increased by fermenting Angelicae.

Example 3. Analysis of Total Polyphenol and Flavonoid Contents of Fermented Angelicae Extract

Polyphenol compounds are secondary metabolites that are widely distributed in the plant world and are largely classified into flavonoids, catechins, and tannins. In particular, phenolic compounds are known to exhibit high antioxidant activity due to their electron donating ability.

As a result of measuring the total polyphenol and flavonoid contents of the non-fermented Angelicae extract and the fermented Angelicae extract, the total polyphenol content was 67.66mg/100g in the NF group, 60.71mg/100g in the F/0 group, and 93.61mg in the F/3 group. /100g, and the flavonoid content was 8.43mg/100g in the NF group, 6.53mg/100g in the F/0 group, and 9.46mg/100g in the F/3 group (Table 3). That is, the total polyphenol content and flavonoid content were increased in the fermented Angelicae extract compared to the non-fermented Angelicae extract.

Total polyphenol and flavonoid content Total polyphenol content (mg/100g) Flavonoid content (mg/100g) NF 67.66±3.61 ^a 8.43±0.13 ^a F/0 60.71±1.64 ^b 6.53±0.88 ^b F/3 93.61±4.86 ^c 9.46±0.35 ^c

Example 4. Mineral Content Analysis of Fermented Angelicae Extract

As a result of measuring the mineral content in the non-fermented Angelicae extract and the fermented Angelicae extract, as a result, potassium (K) was the most contained in the NF, F/0 and F/3 groups at 2.67ppm, 2.61ppm and 3.51ppm, respectively, and the following Calcium (Ca) is 0.77ppm, 0.76ppm, and 0.89ppm, respectively, iron (Fe) is 0.43ppm, 0.59ppm and 0.56ppm, respectively, zinc (Zn) is 0.40ppm, 0.32ppm and 0.35ppm, respectively, manganese (Mn) was 0.39 ppm, 0.38 ppm and 0.49 ppm, respectively, magnesium (Mg) was 0.29 ppm, 0.29 ppm and 0.37 ppm, respectively, and sodium (Na) was contained at 0.16 ppm, 0.22 ppm and 0.29 ppm, respectively. Confirmed (Table 4). That is, the content of various minerals in the fermented Angelicae extract increased compared to the non-fermented Angelicae extract.

K, Ca, Fe, Zn, etc. contained in fermented Angelicae extract are essential minerals important to the human body, are very important in nutritional aspect, and can act as an important factor for maintaining the balance of the skin and having antioxidant activity, thus creating value as a functional material. It was thought to be even higher.

Mineral content NF (ppm) F/0 (ppm) F/3 (ppm) K 2.67±0.08 ^a 2.61±0.09 ^b 3.51±0.09 ^c Ca 0.77±0.05 ^a 0.76±0.02 ^a 0.89±0.07 ^b Fe 0.43±0.01 ^a 0.59±0.01 ^b 0.56±0.04 ^c Zn 0.40±0.02 ^a 0.32±0.04 ^b 0.35±0.04 ^c Mn 0.39±0.01 ^a 0.38±0.03 ^a 0.49±0.03 ^b Mg 0.29±0.03 ^a 0.29±0.03 ^a 0.37±0.04 ^b Na 0.16±0.04 ^a 0.22±0.03 ^b 0.27±0.05 ^c

Example 5. Fermented Angelicae Extract Deckersin And Deckercinol Angelate Content analysis

The content of decurcine and decursinol angelate, which are the main effective substances of Angelicae, in the non-fermented Angelicae extract and the fermented Angelicae extract were analyzed using HPLC.

As a result, peaks corresponding to decursin and decursinol angelate appeared at the retention times of 17.18 and 17.51 minutes, respectively, and their contents were 270.8 ppm and 276.1 ppm respectively in the NF group, and 288.3 ppm each in the F/0 group. And 302 ppm and F/3 groups were 369.5 ppm and 398.0 ppm, respectively, and in particular, it was confirmed that decursine and decursinol angelate were most frequently contained in the F/3 group (Table 5).

Deckercin and Decercinol Angelate Content Retention time (RT) Area value (Area) Height Content (ppm) NF Deckersin
Decercinol Angelate 17.180
17.513 8386522
8720421 649528
515035 270.856
276.117 F/0 Deckersin
Decercinol Angelate 17.169
17.500 9108256
9552622 712636
563264 288.306
302.624 F/3 Deckersin
Decercinol Angelate 17.185
17.517 11593135
12459208 899058
709015 369.531
398.032

Example 6. Fermented Angelicae Extract DPPH Radical Scavenging activity assay

DPPH radical scavenging activity of the non-fermented Angelicae extract and the fermented Angelicae extract was measured, and the positive control was treated with 0.05% of BHT, a synthetic antioxidant available on the market.

As a result, DPPH radical scavenging activity was 41.89% in the NF group and 39.51% in the F/0 group, showing no significant difference, but increased to 60.26% in the F/3 group. It was confirmed that the DPPH radical scavenging activity was increased in the extracts of Angelicae fermented for 4 to 5 days compared to the extracts of Angelicae fermented for 3 days, but was slightly reduced compared to the extract of Angelicae fermented for 3 days. In the positive control group, the DPPH radical scavenging activity was 90.26% (Fig. 2).

Example 7. Analysis of Tyrosinase Inhibitory Activity of Fermented Angelica Angelica Extract

Melanin is a pigment component that is widely distributed in living organisms including humans such as skin, hair, and eyes. In the human body, it is synthesized from melanosomes in melanocytes in the epidermis, and is a kind of amino acid, tyrosine. As a starting material, it is oxidized to DOPA (3,4-dihydroxy-phenylalanine) or DOPA quinone by tyrosinase enzyme, followed by a non-enzymatic reaction and an automatic oxidation process. Melanin is synthesized through polymerization. Since the melanin produced at this time darkens the color of the skin, an experiment that effectively inhibits the activity of tyrosinase is evaluated as a very useful research method to confirm the whitening effect. The positive control group was treated with 500 μM of commercially available arbutin.

As a result of measuring the tyrosinase inhibitory activity of the non-fermented Angelicae extract and the fermented Angelicae extract, the tyrosinase inhibitory activity was 9.13% in the NF group, 13.46% in the F/0 group, and 25.48% in the F/3 group (Fig. 3).

The tyrosinase inhibitory activity was relatively small in the non-fermented Angelicae extract, but increased by about 2.5 times in the extract of Angelicae fermented for 3 days, which was Saccharomyces cerevisiae ( Saccharomyces) cerevisiae ) synergistically, it was thought that the active ingredient of whitening activity was released.

Example 8. Fermented Angelicae Extract Lipoxygenase Inhibitory activity assay

Lipoxygenase is insufficiently expressed in normal cells, but when inflammation is induced, it exhibits activity due to an increase in leukotrienes substances. The positive control group was treated with 0.05% of commercially available indomethacin.

As a result of measuring the lipoxygenase inhibitory activity of the non-fermented Angelicae extract and the fermented Angelicae extract, the lipoxygenase inhibitory activity was 17.62% in the NF group and 18.83% in the F/0 group. At 37.35%, it was about twice as high. In the positive control group, the lipoxygenase inhibitory activity was 89.58% (Fig. 4).

Through this, for the development of anti-inflammatory functional cosmetics derived from herbal herbal medicines, rather than simply using extracts, fermenting them with microorganisms or mycelium is expected to reduce cytotoxicity, inhibit lipoxygenase, and inhibit melanin biosynthesis. It was thought to be possible.

Claims (8)

A cosmetic composition for skin whitening comprising a fermented Angelicae extract with enhanced antioxidant activity as an active ingredient,
The fermented Angelica gigas extract is a cosmetic composition for skin whitening, which is extracted with water after fermentation by inoculating Saccharomyces cerevisiae in Angelica gigas powder. delete As a health functional food composition for preventing or improving melanin hyperpigmentation disease comprising a fermented angelica extract as an active ingredient,
The fermented Angelica gigas extract is a health functional food composition for preventing or improving melanin hyperpigmentation disease, characterized in that the extract is extracted with water after fermentation by inoculating Saccharomyces cerevisiae in Angelica gigas powder. delete delete As a health functional food composition for preventing or improving inflammatory diseases comprising fermented angelica extract as an active ingredient,
The fermented Angelica gigas extract is a health functional food composition for preventing or improving inflammatory diseases, characterized in that the extract is extracted with water after fermentation by inoculating Saccharomyces cerevisiae in Angelica gigas powder. delete delete

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