KR101201916B1 - Preparing method for fagopyrum tataricym sprout extracts using enzyme reaction and cosmetic composition containing the same - Google Patents

Abstract

The present invention relates to a method for preparing bitter buckwheat sprout extract by enzyme reaction and to a cosmetic composition for whitening and antioxidant containing the extract as an active ingredient. Specifically, a method of converting rutin glycosides to quercetin to treat bitter buckwheat sprout extract with glycolytic enzymes to produce extracts excellent in inhibiting melanogenesis and antioxidant activity, and using them for the preparation of cosmetic compositions for whitening and antioxidants .

Description

PREPARING METHOD FOR FAGOPYRUM TATARICYM SPROUT EXTRACTS USING ENZYME REACTION AND COSMETIC COMPOSITION CONTAINING THE SAME}

The present invention relates to a method for producing bitter buckwheat sprout extract by enzyme reaction and to a cosmetic composition for whitening and antioxidant containing the extract as an active ingredient, specifically, a glycoside degrading enzyme by treating glycoside degrading enzyme in bitter buckwheat sprout extract To convert to quercetin to produce an extract excellent in inhibiting melanin production and antioxidant activity, and relates to a technique for use in the preparation of cosmetic compositions for whitening and antioxidant.

Since modern women prefer white and clean skin, there is a growing interest in the treatment of abnormal skin pigmentation and whitening to satisfy beauty needs. Skin is a very important tissue that protects the human body while being in direct contact with the external environment and has biochemical and physical functions. Melanin is a pigment in the epidermal layer of the skin, made from melanocytes in the basal layer of the epidermis and migrates to epidermal cells called keratinocytes. There are no enzymes that break down melanin in vivo, but they are removed by falling off the skin as keratinocytes fall off the epidermis. Tyrosine is converted to dopaquinone via DOPA by tyrosinase enzyme in the endoplasmic reticulum of melanocytes in melanocytes, and through dopachrome through dopachrome by automatic oxidation and enzymatic reaction of dopaquinone. Melanin, a copolymer of, is produced. The melanin thus produced is transferred to keratinocytes through the melanosomes, where it is lost to the surface of the skin along with the keratinization process for about 28 days. Melanin absorbs the light energy of sunlight UV to protect skin organs under the dermis from UV damage, and it protects the skin from external harmful factors such as trapping harmful oxygen and free radicals generated in skin skin. Play a role. In addition, melanin in skin cells determines the color of a person's skin. People with high melanin pigments have brown or black skin, while people with low melanin pigments have white skin.

Currently, as a whitening method for skin beauty, a method of reducing the generated melanin pigment by decolorization and a method of inhibiting the activity of tyrosinase, an enzyme that forms melanin pigment, are known. The whitening agent using tocopherol, vitamins, etc. to reduce the melanin pigment has a slight decolorizing effect of the melanin pigment, and therefore, an inhibitor that inhibits the production of melanin pigment by inhibiting the activity of tyrosinase has attracted attention.

In the field of conventional cosmetics, as a whitening component, for example, substances that inhibit tyrosinase enzyme activity such as kojic acid, arbutin, hydroquinone, vitamin C (L-Ascorbic acid) and these Derivatives of and various plant extracts have been used. Kojic acid inhibits enzymatic activity by adsorbing copper ions present in the active site of tyrosinase, but it is known that there are problems of instability, skin side effects, and liver cancer in the formulation. Vitamin C and its derivatives are difficult to use as raw materials for cosmetics due to their oxidative instability, and hydroquinone has excellent whitening effect on skin but allergic properties, toxicity to melanocytes, and permanent depigmentation to skin. It is highly irritating to, and is recently prescribed as a carcinogenic substance, so its use is banned and only a limited concentration is allowed in each country. In addition, arbutin is a derivative in which hydroquinone is conjugated with glucopyranoside. As a result, hydroquinone is less toxic to human body, and inhibits the synthesis of melanin pigment, resulting in an increase in melanin pigmentation However, it has a disadvantage in that it is partially decomposed by skin enzymes.

The skin is always in contact with oxygen and is exposed to ultraviolet radiation, which continues to undergo photochemical damage induced by reactive oxygen species (ROS). Free radicals are highly reactive ¹ O ₂ And OH, including O ^{2 −} , H ₂ O ₂ , ROO ·, RO ·, ROOH, HOCl and the like. They can be produced in cells and tissues through various processes including high energy radiation, photosensitization and some enzymatic reactions. Among these reactive oxygen species, ¹ O ₂ and OH are known to play an important role in skin photodamage. They accelerate skin aging, including skin antioxidant breakdown, initiation of lipid peroxidation, protein oxidation, DNA oxidation, chain breakdown of connective tissue collagen and hyaluronic acid, wrinkle formation by abnormal crosslinking, and melanogenesis process. Let's do it.

In human skin cells, reactive oxygen species have been described to include UVA (320-380 nm) dependent apoptosis and gene activation, as well as oxidative damage of biological components such as lipids, proteins and DNA. Since collagen is the most abundant component of the matrix of the dermal layer of the skin, the control of biosynthesis and degradation of collagen is the key during skin aging process. This suggests an important role of ROS including ¹ O ₂ in human skin fibroblasts. ^Since ROS, including ¹ O ₂ , are associated with photoaging, the reduction of ROS after exposure to ultraviolet light by antioxidants is an effective way to prevent and minimize photoaging.

Buckwheat (Fagopyrum esculentum) is a yearly herb from the grass family, and is widely distributed in Europe, America, South Africa, and Australia. It is cultivated all over the country in Korea, and it is used a lot as a raw material for food. One reason why buckwheat is a new functional health food is growing in demand because it contains high amounts of bioactive substances rutin, quercetin, quercetrin and myricetin. . Rutin, a functional ingredient of buckwheat, is a flavonoid, a yellow or pale yellow polyphenolic compound, which combines quercetin with rutinose, controls lipid metabolism, and has antihypertensive effects. A variety of functions have been reported, including reports of their effectiveness in the prevention and treatment of cancer. In addition, the use of buckwheat has been diversified as the function of various phenolic substances including quercetin as a natural antioxidant is revealed. In particular, various studies have been attempted as the routine content of bitter buckwheat is found to be higher than that of sweet buckwheat, but systematic research is insufficient.

Korean Unexamined Patent Publication No. 2003-0068969 discloses a composition for alleviating and preventing skin allergy containing buckwheat flour extract or buckwheat seed extract, and Korean Patent Publication No. 2007-0014412 discloses a buckwheat extract obtained from a buckwheat or stem of buckwheat. A functional cosmetic composition containing buckwheat extract, which is included as an active ingredient and has an antioxidant activity, a toxic effect of heavy metals, and an antiwrinkle effect, is disclosed. However, the buckwheat extract had a problem that the effect is lowered due to problems such as not being effectively absorbed by the skin when applied to the skin.

Accordingly, the present inventors have completed the present invention by confirming that when the sprout extract of the bitter buckwheat extract is prepared by the glycolysis degrading enzyme reaction, the specific active ingredient is strengthened and the skin absorption is promoted so that the whitening and antioxidant effects are excellent.

It is an object of the present invention to provide a method for producing a bitter buckwheat sprout extract excellent in whitening and antioxidant activity by strengthening a specific active ingredient by an enzymatic reaction.

In another aspect, the present invention is to provide a bitter buckwheat sprout extract produced by the enzyme reaction having excellent melanin production inhibitory and antioxidant activity.

In another aspect, the present invention is to provide a cosmetic composition for whitening and antioxidant containing bitter buckwheat sprout extract prepared by the above method as an active ingredient.

In order to achieve the above object, according to the present invention,

Bitter buckwheat sprout extract manufacturing step of extracting the dried bitter buckwheat sprout with at least one solvent selected from the group consisting of water, ethanol, methanol, butanol, ether, ethyl acetate and chloroform;

An enzyme addition step of dissolving the bitter buckwheat sprout extract in a solvent and adding a sugar-binding degrading enzyme; And

There is provided a method for preparing a bitter buckwheat sprout extract by an enzyme reaction comprising an enzyme reaction step of proceeding with an enzyme reaction while stirring the extract to which the enzyme is added.

The enzyme reaction is preferably carried out under the conditions of reaction pH 3.5 ~ 5.0, reaction temperature 25 ~ 50 ℃ and reaction time 12 ~ 60 hours.

Glucosidase, glucosidase, xylosidase, xylanase, xylanase, cellulose, galactosidase, pectinase, pectinase, At least one selected from the group consisting of naringinase may be used, and more preferably, pectinase may be used. The bitter buckwheat sprout extract with enhanced quercetin component is prepared by selectively removing the sugars of the routine glycosides contained in the bitter buckwheat sprout extract through the enzyme reaction.

In order to achieve the other object of the present invention, according to the present invention is provided with a cosmetic composition containing 0.005-50% by weight, preferably 0.01-20% by weight of the bitter buckwheat sprout extract prepared by the enzymatic reaction do. Since the cosmetic composition exhibits an excellent tyrosinase inhibitory effect and antioxidant effect, it can be used as a composition for whitening and antioxidant.

The cosmetic composition containing bitter buckwheat sprout extract by an enzyme reaction as a skin lotion, skin softener, skin toner, astringent, lotion, milk lotion, moisturizing lotion, nutrition lotion, massage cream, nutrition cream, moisture cream, hand It may be applied in any one formulation selected from the group consisting of creams, essences, packs, soaps, shampoos, cleansing foams, cleansing lotions, cleansing creams, body lotions, body cleansers, emulsions, press powders, loose powders and eye shadows.

Bitter buckwheat sprout extract by the enzyme reaction prepared by the production method according to the present invention is enhanced in the quercetin component, and excellent in melanogenesis suppression and antioxidant activity, showing a good whitening and antioxidant effect even at a small amount is useful as a cosmetic composition Do.

The present invention, in the production of bitter buckwheat sprout extract, enzymes, such as glucosidase, xylosidase, xylanase, xylanase, cellulose By using at least one enzyme selected from the group consisting of galactosidase (galactosidase), pectinase and naringinase (naringinase) to prepare an extract having a specific active ingredient quercetin strengthening effect by the enzymatic reaction And it relates to a technique for applying it to cosmetics.

Buckwheat is generally classified into Fagopyrum escylentum and Bagopyrum tataricym. Buckwheat contains a large amount of bioactive substances, rutin, quercetin, quercetrin, and myricetin, and bitter buckwheat contains more bioactive substances, including rutin. have. The routine represented by the following formula (2-phenyl-3,5,7,3 ', 4'-pentahydroxy benzopyrone) is a kind of flavonoid which is a yellow or pale yellow polyphenol compound and quercetin (5,7,3', 4). '-tetrahydroxy flavone) combines rutinoside.

Quercetin represented by the following chemical formula (Quercetin) is a kind of flavonoids, one of the highest physiological activity among the flavonoids.

Quercetin is a powerful antioxidant that prevents oxidative damage of cells and rancidity of fats, and has been recognized as a preventive effect of hypertension. Quercetin has this effect because it is highly reactive in vivo, eliminating free radicals that damage many components of the cell, reducing low-density lipoprotein oxidation, the cause of atherosclerosis and heart disease, and antioxidants in the body. It is known to play a role in protecting and regenerating zein vitamin E and inactivating the harmful action of metal ions.

The types and amounts of these buckwheat bioactive substances differ depending on the buckwheat species and the extraction sites. In particular, there is a difference between the bioactive material of buckwheat seeds and germinated buckwheat, that is, the bioactive material of the sprout extract of buckwheat. Plant seeds increase physiological activity and change in composition as germination progresses. In other words, the plant stores energy in the seed to produce the next life, so that when it germinates to produce new shoots, it is equipped with various nutrients. Eventually, the sprout stage will contain much more vitamins, minerals and amino acids than seeds and mature vegetables. As the seed of the plant grows, the physiological activity increases as the germination progresses and the change of the components occurs, so that the high physiological activity starts to appear by germination in terms of physiological activity.

The present invention in the preparation of bitter buckwheat sprout extract to extract quercetin, a specific bioactive substance contained in buckwheat with high efficiency, by enzymatic reaction using a specific enzyme to enhance the specific biologically active substance, promote skin absorption It is characterized by preparing an extract and applying it to the cosmetic composition for whitening and antioxidant.

According to the present invention, the step of extracting the dried bitter buckwheat sprout with at least one solvent selected from the group consisting of water, ethanol, methanol, butanol, ether, ethyl acetate and chloroform to prepare a bitter buckwheat sprout extract;

An enzyme addition step of dissolving the bitter buckwheat sprout extract in a solvent, preferably water, and adding a sugar-binding enzyme; And

There is provided a method for preparing a bitter buckwheat sprout extract by an enzyme reaction comprising an enzyme reaction step of proceeding with an enzyme reaction while stirring the extract to which the enzyme is added.

The enzyme reaction is preferably carried out under the conditions of reaction pH 3.5 ~ 5.0, reaction temperature 25 ~ 50 ℃ and reaction time 12 ~ 60 hours.

The method is described in detail as follows.

The bitter buckwheat sprout extract provided for the enzymatic reaction comprises dried bitter buckwheat sprout with one or more organic solvents selected from the group consisting of water or organic solvents, preferably water, ethanol, methanol, butanol, ether, ethyl acetate and chloroform or It may be prepared by extraction with a conventional extraction method using a mixture of water and more preferably 70% ethanol.

The bitter buckwheat sprout extract is an enzyme that performs a glycolytic decomposition, preferably glucosidase, xylosidase, xylanase, cellulase, cellulose, galactosidase ( galactosidase), pectinase (pectinase) and naringinase (naringinase) is prepared by proceeding the enzymatic reaction by treatment with one or more enzymes selected from the group, more preferably pectinase (pectinase). The sugars of the routine glycosides contained in the buckwheat sprout extract by the sugar-binding enzymes are selectively degraded to increase the content of quercetin in the extract. The enzymes include the genus Aspergillus, genus Bacillus, genus penicillium, genus rhizopus, genus rhizomucor, genus talomyces, genus Bifi Obtained from microorganisms of the genus bifidobactertium, mortierella.

The bitter buckwheat sprout extract prepared by the enzyme reaction was compared with the extract of buckwheat seeds or simple germinated buckwheat extract, it was confirmed that the quercetin component is enhanced to have a better whitening and antioxidant activity even at a small amount.

As described above, according to the present invention, a buckwheat sprout extract is prepared by the enzymatic reaction and has excellent melanin production inhibitory activity and antioxidant activity. According to the present invention, there is provided a cosmetic composition for whitening and antioxidant, containing 0.005 to 50% by weight, preferably 0.01 to 20% by weight of the bitter buckwheat sprout extract by the enzymatic reaction.

The cosmetic composition containing bitter buckwheat sprout extract by an enzyme reaction as a skin lotion, skin softener, skin toner, astringent, lotion, milk lotion, moisturizing lotion, nutrition lotion, massage cream, nutrition cream, moisture cream, hand It may be applied in any one formulation selected from the group consisting of creams, essences, packs, soaps, shampoos, cleansing foams, cleansing lotions, cleansing creams, body lotions, body cleansers, emulsions, press powders, loose powders and eye shadows.

Hereinafter, the present invention will be described in detail with reference to the following examples and test examples, but the present invention is not limited to these examples.

Preparation Example 1: Preparation of Bitter Buckwheat Sprout Extract

100 g of bitter buckwheat sprout was extracted from 600 g of 70% ethanol at 50 ° C. for 3 hours to obtain a bitter buckwheat sprout extract, and it was concentrated under reduced pressure and freeze-dried to prepare 3.4 g of bitter buckwheat sprout extract.

Examples 1-7: Preparation of Bitter Buckwheat Sprout Extract by Enzyme Reaction

Concentration and bitter buckwheat sprout extract prepared in Preparation Example 1 concentrate (solid content per 100g 1.47g) in 50g of purified water to 400ml and a mixture of 50ml enzyme solution in table 1 enzyme with stirring for 48 hours at pH 4.5, 45 ℃ conditions After the reaction was conducted, the mixture was concentrated under reduced pressure, thereby preparing a bitter buckwheat sprout extract by enzyme reaction.

Enzyme Origin Example 1 Amylase Aspergillus oryzae Example 2 Beta-glucosidase Almond Example 3 Cellulase-A Aspergillus niger Example 4 Beta-galactosidase Aspergillus oryzae Example 5 Beta-xylosidase Aspergillus niger Example 6 Pectinase Rhizopus sp Example 7 Naringinase Penicillium decumbens

Test Example 1 Measurement of Quercetin Formation Rate

Quercetin production rate was measured after 48 hours of the enzyme reaction extract prepared in Examples 1 to 7 using HPLC. The production rate of quercetin indicates the increase rate of quercetin content when the extract of Preparation Example 1 was used as a control, and was calculated as in the following formula.

Production rate (%) = Quercetin Standard sample amount (mg) x Example peak area value ÷ Quercetin Standard peak area value ÷ Example sample amount (mg)

The results are shown in Table 2 below.

% Creation Example 1 32 Example 2 45 Example 3 59 Example 4 56 Example 5 28 Example 6 79 Example 7 12

Examples 8-13: Preparation of Bitter Buckwheat Sprout Extract by Enzyme Reaction

Bitter buckwheat sprout extract was prepared by enzyme reaction using the pectinase of Example 6, which had the best quercetin production rate. 50 g of the concentrated solution of the bitter buckwheat sprout extract prepared in Preparation Example 1 (solid content of 1.47 g per 100 g) was mixed with 400 ml of purified water and 50 ml of Pectinase (AFP-L4, Novozyme) enzyme solution, and the conditions as shown in Table 3 below. After proceeding the enzyme reaction in, concentrated under reduced pressure to prepare a bitter buckwheat sprout extract by the enzyme reaction.

Reaction temperature (℃) Reaction time (hr) Reaction pH Example 8 25 48 4.5 Example 9 30 48 4.5 Example 10 35 48 4.5 Example 11 40 48 4.5 Example 12 45 48 4.5 Example 13 50 48 4.5

Examples 14-19: Preparation of bitter buckwheat sprout extract by enzyme reaction

Bitter buckwheat sprout extract was prepared by the enzyme reaction using the pectinase of Example 6, which had the best quercetin production rate.

50 g of the concentrated solution of the bitter buckwheat sprout extract prepared in Preparation Example 1 (solid content of 1.47 g per 100 g) was mixed with 400 ml of purified water and 50 ml of Pectinase (AFP-L4, Novozyme) enzyme solution, as shown in Table 4 below. After proceeding the enzyme reaction in, concentrated under reduced pressure to prepare a bitter buckwheat sprout extract by the enzyme reaction.

Reaction temperature (℃) Reaction time (hr) Reaction pH Example 14 45 12 4.5 Example 15 45 18 4.5 Example 16 45 24 4.5 Example 17 45 36 4.5 Example 18 45 48 4.5 Example 19 45 60 4.5

Examples 20-23: Preparation of bitter buckwheat sprout extract by enzyme reaction

Bitter buckwheat sprout extract was prepared by the enzyme reaction using the pectinase of Example 6, which had the best quercetin production rate, by varying the reaction pH.

50 g of the concentrated solution of the bitter buckwheat sprout extract prepared in Preparation Example 1 (solid content of 1.47 g per 100 g) was mixed with 400 ml of purified water and 50 ml of pectinase (AFP-L4, Novozyme) enzyme solution, as shown in Table 5 below. After proceeding the enzyme reaction in, concentrated under reduced pressure to prepare a bitter buckwheat sprout extract by the enzyme reaction.

Reaction temperature (℃) Reaction time (hr) Reaction pH Example 20 45 48 3.5 Example 21 45 48 4.0 Example 22 45 48 4.5 Example 23 45 48 5.0

Test Example 2: Measurement of Quercetin Production Rate

The production rate of bitter buckwheat sprout extract by the enzyme reaction prepared in Examples 8 to 23 was measured and the results are shown in Table 6 below.

Example
8 Example 9 Example 10 Example 11 Example 12 Example 13 Example 14 Example 15 Example 16 Example 17 Example 18 Example 19 Example 20 Example 21 Example 22 Example 23 % Creation 58 61 63 66 72 65 41 46 52 66 67 73 69 71 80 73

As shown in Table 6, when using pectinase as the glycoside degrading enzyme, the reaction rate was 45 ° C., the reaction time was 48 hours, and the reaction pH showed the best quercetin production rate (%).

Test Example 3: Melanin production inhibitory effect

6 ml plate was inoculated with B16F10 cells at a concentration of 2 × 10 ⁵ in 2 ml of DMEM medium added 10% FBS per well to 37 ° C., 5% CO _2. The incubator was incubated for 24 hours. After incubation for 24 hours, the wells were exchanged with fresh medium, and the diluted solution prepared by diluting the bitter buckwheat sprout extract by the enzyme reaction prepared in Example 22 to 62.5, 125, and 250 µg / ml, respectively, was prepared. Incubated at the same conditions for 18 hours by addition. The cells were then washed with PBS, treated with 300 μl of 0.25% trypsin-EDTA, collected and transferred to a microcentrifuge tube, and then centrifuged at 1,000 rpm for 4 minutes. The precipitated cells were washed twice with PBS, and 500 µl of 1N NaOH was added to dissolve melanin. To help dissolve melanin, the reaction was carried out at 100 ° C. for 10 minutes. The amount of melanin in each microcentrifuge tube was determined spectrophotometrically by measuring absorbance at 400 nm.

The test results were obtained as shown in Table 7 below.

Sample concentration (㎍ / ㎖) Melanin synthesis inhibition (%) 0 0.00 62.5 0.00 125 2.30 250 44.24

The higher the melanin synthesis inhibition (%), the more melanin production was inhibited, as shown in Table 7 when the treatment of the extract of bitter buckwheat sprouts by the glycolytic enzyme reaction according to the present invention melanin synthesis Inhibition (%) was found to show an excellent inhibitory effect of melanin production, it was found that there is a synergistic effect as the concentration increases.

Test Example 4: Effect of inhibiting TRP-1, TRP-2 and Mu-tyr expression

B16F10 cells were inoculated in DMEM medium with 10% FBS at a concentration of 5 × 10 ⁵ and incubated in 37 ° C., 5% CO ₂ incubator for 24 hours. Incubated for 24 hours, the dilution solution prepared by diluting the bitter buckwheat sprout extract by the glycolytic enzyme reaction prepared in Example 22 to 50 and 100 ㎍ / ml, respectively, was added to the same conditions for 18 hours. Incubated at. Thereafter, B16F10 cells were recovered using Trizol reagent (invitrogen, USA), mRNA was extracted, and cDNA was synthesized through a series of processes. From the synthesized cDNA, TRP-1 (Tyrosinase related protein-1), TRP-2 (Tyrosinase related protein-2) and Mu-tyrosinase gene regions were amplified to confirm the expression level of the gene by electrophoresis. (GenePro, Hangzhou bioer tech., CHINA) using 10x taq polymerase buffer, 10 mM dNTP, 10 pmol primer (TRP-1 F: AGG AAT CTG GCT TGG GAT TT, TRP-1 R: ATG AGC CAC AAG GGT CAG TC, TRP-2 F: AGC AGA CGG AAC ACT GGA CT, TRP-2 R: CAG GTA GGA GCA TGC TAG GC, Mu-tyr F: GGG CCC AAA TTG TAC AGA GA, Mu-tyr R: GGC AAA TCC TTC CAG TGT GT), taq polymerase was mixed and distilled water was adjusted to 50 uL, then 94 ° 5 min 1 cycle / 94 ° 30 sec / 52 ° 30 sec (TRP-1), 53 ° 30 sec (TRP-2, Mu- tyr) / 72 degrees 30 seconds 35 cycles amplification, was carried out under the conditions for 5 minutes reaction at 72 degrees. The expression inhibition rate of the generated TRP-1, TRP-2 and Mu-tyr was calculated according to the following equation, and the control group used normal B16F10 cells without treatment.

Expression inhibition rate (%) = {1- (B / A)} × 100 (%)

A: the amount of TRP-1, TRP-2 and Mu-tyr expression in the control group

B: Sample Processing and TRP-1, TRP-2 and Mu-tyr Expression in B16F10 Cells

The test results were obtained as shown in Table 8 below.

Sample concentration
(Μg / ml) Expression inhibition rate (%) TRP-1 TRP-2 Mu-tyr 0 0.00 0.00 0.00 50 9.53 1.28 3.07 100 20.44 20.99 30.44

As confirmed in the results of Table 8, the application of bitter buckwheat sprout extract by the glycolytic enzyme reaction effectively inhibited the expression of TRP-1, TRP-2 and Mu-tyr genes involved in melanin synthesis, As the concentration increased, the effect was better.

Test Example 5: Free radical scavenging activity

The antioxidant capacity of the extract according to the present invention was measured by electron donating ability against DPPH. DPPH shows strong absorption band at 517nm because of its odd number of electrons, but when it reacts with electron donor, it receives electrons or hydrogen radicals and becomes a stable molecule. In addition, the donated electrons are irreversibly bound, and the proportion of the progressive DPPH becomes lighter and the absorbance decreases in proportion to the number.

The extract prepared in Example 22 was used as a sample, and DPPH was purchased from Sigma Co., Ltd., USA. First, a standard DPPH solution of 400 μm concentration was made. And the sample of Example 22 was prepared by dissolving in DMSO so that the concentration of 10, 50, 100, 250, 500ppm, respectively. Ascorbic acid (500 ppm) and non-enzymatic bitter buckwheat sprout extract (500 ppm, Preparation Example 1) were prepared as a control. Add 100 μl of 0.4 mM DPPH to the ascorbic acid and 100 μl of the sample in a 96 well plate and mix. Ascorbic acid and the sample blank were mixed with 100 μl of EtOH instead of 0.4 mM DPPH. After color development at 37 ° C. for 30 minutes, zero calibration was carried out with blanks at 517 nm and the absorbance was measured.

DPPH radical scavenging ability (%) = (absorbance of 1-test group / absorbance of control group) × 100

The results are shown in Table 9 below.

Sample concentration Free radical scavenging effect (%) 500 ppm of Ascorbic acid 95.8 Example 22 10 ppm 18.1 Example 22 50 ppm 29.4 Example 22 100 ppm 54.8 Example 22 250 ppm 72.8 Example 22 500 ppm 93.2 Preparation Example 1 500 ppm 45.3

As confirmed in Table 9, the bitter buckwheat sprout extract by the enzymatic reaction according to the present invention was similar to the existing antioxidant ascorbic acid, or showed an excellent antioxidant capacity compared to the bitter buckwheat sprout extract not treated with enzyme.

Test Example 6: Xanthine oxidase inhibitory activity

Xanthine oxidase inhibitory activity was measured according to SOD-like activity measurement / NBT (Nitroblue tetrazolium) method. 10 μl of the sample extract of Example 22 dissolved at a concentration of 10 to 500 ppm was mixed with 140 μl of 50 mM sodium phosphate buffer (PH 7.5), 40 μl of xanthine (3 mM), and 10 μl of 1.5 mM NBT, followed by xanthine oxidase ( 0.13 U / mL) 10 μl was finally added, mixed, and reacted at 25 ° C. for 40 minutes, and then absorbance was measured at 560 nm.

Xanthine oxidase inhibitory activity of the extract of the present invention was expressed in% the absorbance reduction rate of the addition and no addition of the sample as shown in the following equation. Ascorbic acid (500ppm) and non-enzymatic bitter buckwheat sprout extract (500ppm, Preparation Example 1) was measured as a control.

Xanthine oxidase inhibitory activity (%) = (absorbance of 1-sample added / absorbed sample-free) x 100

The results are shown in Table 10 below.

Sample concentration Xanthine oxidase Inhibitory Activity (%) 500 ppm of Ascorbic acid 88.0 Example 22 10 ppm 16.8 Example 22 50 ppm 22.7 Example 22 100 ppm 39.4 Example 22 250 ppm 61.3 Example 22 500 ppm 89.5 Preparation Example 1 500 ppm 56.4

As confirmed in Table 10, the bitter buckwheat sprout extract by the enzyme reaction according to the present invention was similar to the existing antioxidant ascorbic acid, or showed an excellent antioxidant capacity compared to the bitter buckwheat sprout extract not treated with enzyme.

Preparation Example 1 Emulsion Base Preparation

An emulsion base was prepared according to the composition of Table 11 as a cosmetic for whitening and antioxidant containing bitter buckwheat sprout extract by the enzyme reaction of the present invention.

ingredient (weight%) glycerin 3.00 Disodium iodide 0.02 Polyglyceryl-3-methylglucose distearate 1.50 Cetearyl Alcohol 0.50 Caprylic / capric triglyceride 7.00 Polyacrylamide & C13-14 Isoparaffin & Laures-7 0.60 Bitter buckwheat sprout extract by enzyme reaction (Example 22) 1.00 Incense, preservative a very small amount Purified water Balance system 100

Preparation Example 2 Flexible Lotion

A flexible lotion containing bitter buckwheat sprout extract by the enzyme reaction according to the present invention with the composition shown in Table 12 was prepared by a conventional method.

ingredient (weight%) Bitter buckwheat sprout extract by enzyme reaction (Example 22) 0.5 1,3-butylene glycol 5.2 Oleyl alcohol 1.5 ethanol 3.2 Polysorbate 20 3.2 Benzophenone-9 2.0 Carboxyl Vinyl Polymer 1.0 glycerin 3.5 incense a very small amount antiseptic a very small amount Purified water Balance system 100

Preparation Example 3: Nutritional Lotion

A nutritious lotion containing bitter buckwheat sprout extract by the enzyme reaction according to the present invention with the composition shown in Table 13 was prepared by a conventional method.

ingredient (weight%) Bitter buckwheat sprout extract by enzyme reaction (Example 22) 2.0 Glyceryl Stearate SE 1.5 Stearyl alcohol 1.5 lanolin 1.5 Polysorbate 1.3 Sorbitan stearate 0.5 Hardened vegetable oil 1.0 Mineral oil 5.0 Squalane 3.0 Trioctanoine 2.0 Dimethicone 0.8 Tocopherol Acetate 0.5 Carboxyvinyl polymer 0.12 glycerin 5.0 1,3-butylene glycol 3.0 Sodium hyaluronate 5.0 Triethanolamine 0.12 Preservative, fragrance, pigment a very small amount Distilled water Balance Sum 100

Preparation Example 4 Nutrition Cream

A nutritious cream containing bitter buckwheat sprout extract by the enzyme reaction according to the present invention with the composition shown in Table 14 was prepared by a conventional method.

ingredient (weight%) Bitter buckwheat sprout extract by enzyme reaction (Example 22) 2.0 Lipophilic monostearate 2.0 Stearic acid 1.6 Wax 1.0 Polysorbate 1.6 Sorbitan stearate 0.6 Hardened vegetable oil 1.0 Squalane 3.0 Mineral oil 5.0 Trioctanoine 5.0 Dimethicone 1.0 glycerin 5.0 Triethanolamine 1.0 Sodium hyaluronate 4.0 Preservative, fragrance, pigment a very small amount Distilled water Balance Sum 100

Claims (7)

A step of preparing bitter buckwheat sprout extract by extracting the dried bitter buckwheat sprout with at least one solvent selected from the group consisting of water, ethanol, methanol, butanol, ether, ethyl acetate and chloroform;
An enzyme addition step of dissolving the bitter buckwheat sprout extract in a solvent and adding pectinase as a sugar-binding enzyme; And
A method of preparing bitter buckwheat sprout extract by an enzyme reaction comprising an enzyme reaction step of performing an enzyme reaction while stirring the extract to which the enzyme is added under conditions of a reaction pH 4.5, a reaction temperature of 45 ° C. and a reaction time of 48 hours. delete delete delete delete It is prepared according to claim 1, bitter buckwheat sprout extract by the enzyme reaction with enhanced quercetin component. A whitening and antioxidant cosmetic composition containing 0.005-50% by weight of the bitter buckwheat sprout extract by the enzyme reaction prepared according to claim 1, based on the total weight of the composition.