KR101558184B1 - Anti-aging composition containing red-grape-leaf extract berrylike mixtures and selenium - Google Patents

Abstract

The present invention relates to a composition for antioxidation for improving skin, and more particularly, it relates to a composition for preventing and treating skin, and more particularly, to a composition containing red grape leaf extract, berry mixture and selenium, And is capable of improving photo-aging caused by UV.

Red grape leaf extract, berry mixture, selenium, photo-aging, antioxidant

Description

The present invention relates to an antioxidant composition containing red-grape leaf extract, berri mixture, and selenium.

The present invention relates to a composition for antioxidation for improving skin, and more particularly, it relates to a composition for preventing and treating skin, and more particularly, to a composition containing red grape leaf extract, berry mixture and selenium, And is capable of improving photo-aging caused by UV.

There are many reasons for promoting aging, but Reactive Oxygen species (ROS) is considered to be one of the major causes. This active oxygen is indispensable in energy metabolism, immune response, etc., and it is inevitable irritation caused by external harmful environment. Active oxygen is highly reactive and causes DNA degeneration, excessive signal transduction and protein denaturation in the body, resulting in a series of reactions accumulating adverse health effects. These harmful reactions are supposed to preserve their homeostasis by antioxidants (uric acid, vit.C, vit.E etc.) or antioxidant enzymes (Glutathione peroxidase, superoxide dismutase, catalase, etc.) in vivo. However, the aging of the antioxidant system due to the endogenous aging and the accumulation of active oxygen by the continuous harmful stimuli breaks this balance, harming health, promoting aging, causing various diseases such as skin diseases, skin cancer, arteriosclerosis and thrombosis ( Laure Rittie et al., Aging Research Reviews, 1, 705-720, 2002; Cutler RG, Annals of the New York Academy of Sciences, 1055, 93-135, 2005).

Accordingly, there is an increasing interest in antioxidants that inhibit the formation of active oxygen systems or remove the formed active oxygen system. Antioxidant substances can be divided into two groups: naturally occurring in the body and externally administered. Antioxidants naturally present in the body include superoxide dismutase (SOD), glutathione, peroxidase and Catalase and the like, and phytochemicals such as kaempferol, catechin and genistein are externally administered. Vitamin E, vitamin C and beta carotene; And minerals such as selenium.

However, if the amount of the antioxidant used is too small, a sufficient antioxidative effect can not be expected. If the amount of the antioxidant is excessively increased, the active oxygen produced in the body can not be properly removed due to side effects such as inhibition of activity of some enzymes. Which may cause damage to the tissue. Therefore, there is a growing interest in compositions that exhibit synergy by using antioxidants in combination.

Korean Patent No. 0835866 discloses an antioxidant composition capable of evaluating the reactivity of an antioxidant in the body over time and increasing the duration of its efficacy in the "antioxidative composition preventing side effects as an oxidation promoter of antioxidants" . As for the red grape leaf extract, US Patent Application Publication No. 2005/048008 discloses an anti-aging cosmetic composition containing an extract of Vitis Vinifera as an anti-inflammatory composition as an anti-inflammatory composition, Patent Publication No. 2001-081008 discloses a composition for preventing rough skin, a composition for improving skin or an anti-aging composition having an anti-aging effect, which comprises grape leaf extract and the like and sugar alcohol having 4 to 6 carbon atoms, The synergistic effects of other antioxidant ingredients are in short use.

Accordingly, the inventors of the present invention evaluated various synergistic effects on body reactivity and antioxidant efficacy by adding various antioxidants to the grape leaf extract, and as a result, they have developed a composition for antioxidation which is ultimately effective for improving photoaging, .

Accordingly, an object of the present invention is to provide a composition for antioxidation having an excellent effect for improving photoaging.

In order to achieve the above object, the present invention provides an antioxidative composition comprising a red grape leaf extract, a berry mixture, and selenium as an active ingredient.

The composition for antioxidation according to the present invention has a synergistic effect as an antioxidant agent when each component is used in combination with a combination of red grape leaf extract, berry mixture and selenium as an effective ingredient. And the antioxidant activity in the body was increased. In addition, even when the composition of the optimal combination ratio was administered for a long period of time, antioxidant efficacy was further improved in various antioxidant indexes compared to the case of using the same amount alone. Accordingly, the antioxidant composition according to the present invention is expected to have a great effect on health promotion and anti-aging as a free radical scavenger (antioxidant) when applied to ordinary beverages, health supplements or skin aesthetic agents .

The red grape leaf extract used in the present invention is a material obtained by extracting a water-soluble component from a red leaf ( Folia vitis viniferae ) of a grape (Vitis vinifera L.) It contains not less than 10% of total polyphenol as a whole powder and is attracting attention as a functional material. The grape leaf extract used in the present invention can be extracted with water or an organic solvent through a method well known in the art. The organic solvent used in the present invention may be selected from the group consisting of ethanol, methanol, butanol, ether, ethyl acetate, chloroform and a mixed solvent of these organic solvents and water, preferably 80% ethanol. At this time, the extraction temperature is preferably 10 to 80 캜, and it can be extracted for 6 to 24 hours. If the extraction temperature and the extraction time are exceeded, the extraction efficiency may be deteriorated or the component may be changed.

The berry mixture used in the present invention may be a mixture of two or more kinds of fruits selected from the group consisting of pomegranate, raspberry, blackberry, acai, cranberry and blueberry. The powder may be prepared by drying the powdered fruits and then mixing the powders, or by mixing the powdered fruits of each fruit with freeze-dried powder. However, the present invention is not limited thereto.

The selenium used in the present invention is selenium source material extracted from paradise nut, and 0.1% selenium is contained per weight of paradise nut extract powder.

Components such as Quercetin or Flavone, which are representative polyphenols contained in red grape leaf extract, are excellent antioxidant activity. However, antioxidant components are superior to each other in continuous use and superior Effects often occur. Therefore, in order to develop a composition having an antioxidative effect that is more optimal than other antioxidant components, the present invention provides an antioxidant composition comprising an antioxidant mineral, selenium and an anthocyanin-rich berry mixture, .

The present invention provides an antioxidative composition containing red grape leaf extract, berry mixture and selenium simultaneously.

In the antioxidant composition of the present invention, the grape leaf extract, the berry mixture and the selenium are preferably mixed in a weight ratio of 1: 1: 1 to 10: 1: 1. At this time, the content of the mixture of the active ingredients is preferably 0.1 to 80% by weight based on the total weight of the composition. When the content of the mixture is less than 0.1% by weight, it is difficult to expect effective efficacy. When the content is more than 80% by weight, formulation is difficult.

The antioxidant composition according to the present invention can prevent aging by eliminating the active oxygen system in the body and preventing the activity of the antioxidant enzyme from being lowered.

The antioxidant according to the present invention may be contained in a composition stabilized in tablets and capsules by a technique well known in the art depending on the ratio and properties of each substance, But is not limited thereto.

Further, the antioxidant composition according to the present invention has a great effect on health promotion and anti-aging as a free radical scavenging agent (antioxidant) when it is applied to ordinary beverages, health supplements and skin aesthetic agents.

Hereinafter, the present invention will be described more specifically with reference to examples and test examples. It is to be understood that the scope of the present invention is not limited to these embodiments and that variations, substitutions, and insertions conventionally known in the art can be carried out, And this is included in the scope of the present invention.

[Reference Example 1] Preparation of red grape leaf extract

1 kg of red grape leaves were collected and 5 L of 80% ethanol aqueous solution was added, and the mixture was refluxed three times and immersed at 15 ° C for 1 day. Thereafter, the residue and the filtrate were separated by filtration through a filter cloth and centrifugation, and the separated filtrate was concentrated under reduced pressure to obtain 155 g of a grape leaf extract.

[Reference Example 2] Preparation of berry mixture

The same amount of pomegranate, raspberry, blackberry, acai, cranberry and blueberry fruit were dried and powdered and mixed to prepare a berry mixture.

[Test Example 1] In vitro free radical scavenging activity

Experiments were conducted to evaluate changes in free radical scavenging ability of the single substance or complex substance of the control group, Comparative Examples 1 to 3 and Example 1 shown in the following Table 1.

division Contents Control group DMSO without sample was prepared. Comparative Example 1 The red grape leaf extract of Reference Example 1 was dissolved in solvent DMSO to a concentration of 100,000 ppm (10% w / v). Comparative Example 2 The berry mixture of Reference Example 2 was dissolved in solvent DMSO to 100,000 ppm (10% w / v). Comparative Example 3 Selenium was dissolved in solvent DMSO to 100,000 ppm (10% w / v). Example 1 The grape leaf extract of Reference Example 1, the berry mixture of Reference Example 2 and selenium were mixed in a ratio of 6: 1: 1 and dissolved in solvent DMSO to 100,000 ppm.

The measurement method was 1,1-diphenyl-2-picryl-hydrazyl (DPPH) method. The reagent was purchased from Sigma, USA. Since this reagent exists as a relatively stable free radical, it is used primarily as an in vitro method in confirming the free radical scavenging action.

The concentration of the components used for measuring DPPH free radical scavenging activity was 4 ppm, 20 ppm and 100 ppm, respectively. The components of the above concentrations were placed in a 96-well plate, and DPPH prepared from 100 μM ethanol solution was added thereto to make the total volume of the solution 200 ml. After incubation at 37 ° C for 30 minutes, absorbance was measured with a 520 nm ELISA reader. The free radical scavenging activity was calculated by the following equation (1).

Free radical scavenging activity (%) = 100 - B / A × 100

(Where A is the absorbance of the control well not treated with the components and B is the absorbance of the wells of Comparative Examples 1 to 3 and Example 1 in which the respective components were treated)

The experiment was repeated four times, and the results are shown in Table 2 below.

DPPH cancellation capability measurement Test substance Free radical scavenging activity 4 ppm 20 ppm 100ppm Comparative Example 1 18.2% 22.1% 57.3% Comparative Example 2 6.8% 12.1% 24.6% Comparative Example 3 10.0% 16.3% 28.8% Example 1 15.1% 25.4% 55.7%

The results of Table 2 above indicate that DPPH elimination ability of Comparative Example 1 containing red grape leaf extract was excellent and it was thought to affect many physiological actions related to the oxidation-reduction reaction in the body. In addition, the antioxidant activity of Example 1 in which the berry mixture and selenium were mixed with the grape leaf extract was also significantly increased.

[Test Example 2] Effect of active oxygen scavenging in vitro

In order to confirm the effect of antioxidants on the action of intracellular antioxidants, experiments were carried out to evaluate the scavenging ability of active oxygen generated by UV in vitro.

Cell culture and material treatment are as follows. The Hacat cell line (human keratinocyte cell line) (Dulbecco's Modification of Eagles Medium, Sigma) containing 10% FBS, 100 IU / ml penicillin and 100 μg / ml streptomycin was used as a culture medium and incubated at 37 ° C In the incubator, a mixed gas of air (95%) and CO ₂ (5%) was continuously supplied and cultured. Hacat cells were transplanted into a culture vessel, and after 24 hours, the culture medium was replaced with non-FBS-treated DMEM, and a sample to be assayed was treated with cultured cells. After incubation for 24 hours, the cells were washed with HEPES-buffered control salt solution to remove the remaining medium. DCFH-DA (2 ', 7'-dichlorodihydro-fluorescein diacetate, Molecular Probes, Inc) was added to each well and incubated at 37 ° C and 5% CO ₂ for 20 minutes and washed with HCSS. The fluorescence intensity of DCF (dichlorofluorescein) initially oxidized to ROS after adding 100 ml of HCSS was measured with a fluorescent plate reader (Ex = 485 nm, Em = 530 nm). The fluorescence intensity was measured with a fluorescent plate reader (Ex = 485 nm, Em = 530 nm) immediately after the treatment and after 3 hours of treatment with UVB 30 mJ / Each experimental group used 1, 10 and 100 ppm of Comparative Examples 1 to 3 and Example 1 in Table 1, and the results are shown in Table 3 below.

The composition of the HCSS (HEPES-buffered control salt solution) used in the above procedure was as follows: NaCl 120 mM (7.0128 g / L), KCl 5 mM (0.37275 g / L), MgCl ₂ 1.6 mM (327 ul / , CaCl ₂ 2.3 mM (0.2553 g / L), glucose 15 mM (2.703 g / L), HEPES 20 mM (4.766 g / L) and NaOH 10 mM (0.4 g / L).

Active oxygen scavenging activity in Hacat cell line Test substance Active oxygen (ROS) scavenging activity 1 ppm 10 ppm 100 ppm Comparative Example 1 8.2% 12.1% 23.2% Comparative Example 2 4.8% 11.1% 12.6% Comparative Example 3 8.0% 9.3% 10.8% Example 1 16.1% 27.4% 48.3%

The results of the above Table 3 show that the active oxygen scavenging activity was excellent in Example 1, and Comparative Examples 1 to 3 were relatively inferior in activity. This means that the action of the active oxygen in the cell is less effective for elimination of the active oxygen, which means that the intracellular substance absorption or signal transduction capacity required for the active oxygen scavenging activity is relatively low. On the other hand, in Example 1, the active oxygen scavenging ability was much better than the actual content of red grape leaf extract. This means that synergy can be obtained by further mixing the berry mixed extract and selenium, Such as intracellular uptake, signaling or role relaxation and stabilization of the oxidizing promoter.

[Test Example 3] Free radical scavenging effect in vitro

In order to examine the effect of the composition of the present invention on the free radical scavenging activity in vivo, hairless mice were selected as animal models. 6-7 week old female hairless mice (SKH, HR-1) were divided into a control group (normal group), a UV control group, and a control group (Comparative Example 4) They were raised during the experimental period.

division Contents of administration Control group As a normal group, physiological saline was administered. UV control group As a UV control group, UV irradiation and physiological saline were administered. Comparative Example 4 UV was applied and the grape leaf extract of Reference Example 1 was mixed with physiological saline. Example 2 UV was applied and mixed with the grape leaf extract of Reference Example 1, the berry mixture of Reference Example 2 and selenium in a ratio of 6: 1: 1 in physiological saline.

In the control and UV control groups, 0.5 ml of physiological saline was orally administered. In Comparative Example 4 and Example 2, 167 mg of powder per kg of body weight was mixed with 0.5 ml of saline per kg of body weight. The administration period was 5 weeks in total and 5 days in a week.

For the free radical scavenging activity, 1 ml of blood samples were taken from the eyeballs of the experimental animals in each experimental group after the test period, and they were separated into blood cells and plasma using a centrifuge (3000 rpm, 10 minutes) The supernatant was taken and stored in 1.7 ml Micro-E tubes, frozen until just before analysis.

The evaluation method is as follows. ABTS (2,2'-Azino-bis (3-ethylbenzthiazoline 6-sulfonic acid)) was added to 0.05 M phosphate-citrate buffer at pH 5.0 to a final concentration of 150 M Respectively. Hydrogen peroxide (H ₂ O ₂ ) and peroxidase were added to the ABTS solution at 75 M and 2.5 M, respectively, and reacted to produce the ABTS radical (ABTS˙ ⁺ ). 20 L of plasma samples were added to 180 L of ABTS radical solution, mixed thoroughly, shaded at room temperature, reacted for 60 min, and absorbance of the sample was measured at 600 nm after completion of the reaction. At the same time, Trolox was added to 0.05 M phosphate-citric acid buffer, pH 5.0, to a final concentration of 10 mM, 2 mM, 1 mM, 100 M and 10 M to prepare a Trolox sample. And the absorbance was measured to prepare a standard curve. The results are shown in FIG. The free radical scavenging ability of the plasma sample was represented by the Trolox Equivalence Antioxidant Activity (TEAC) value using the following equation 2 derived from the prepared standard curve, and the results are shown in Table 5. Here, TEAC is a unit that allows relative comparisons of antioxidant activity, by indicating the free eliminability of the evaluation sample in terms of the equivalent activity of trolox (Miller NJ et al., Clinical Science, 84, 407-412, 1993).

Test substance TEAC value Control group 0.0 ± 12.5 UV control group -34.8 ± 8.6 Comparative Example 4 13.3 ± 9.3 Example 2 22.2 ± 12.1

From the results shown in Table 5, the free radical scavenging ability of Example 2 according to the present invention was much superior to that of the control and UV control groups, and showed a free radical scavenging ability twice as excellent as that of Comparative Example 4 containing only red grape leaf extract .

[Test Example 4] Suppression of photoaging using skin template

In order to investigate the effect of the composition of the present invention on photoaging symptoms, an experiment was conducted using the animal model used in Test Example 3. From 2 weeks to 5 weeks after the oral administration, the control, UV control, Comparative Example 4 and Example 2 were irradiated with UV light similar to sunlight three times a week. At this time, total UV irradiation amount was 600 mJ / ㎠ during the experiment. To determine the effect of wrinkle improvement, a skin polymer replica was taken from the dorsal side of a hairless mouse before the autopsy. To compare the degree of wrinkles of the skin, a skin visiometer was used And the image of the skin surface was filed. This is shown in Fig.

As can be seen from the results of FIG. 2, the bending degree of the wrinkles on the skin surface of the hairless mouse of Example 2 was remarkably alleviated as compared with the UV control and Comparative Example 4, It was confirmed that it is effective in suppressing the photoaging phenomenon by improving the wrinkles of the skin by the skin.

[Test Example 5]

Forty adult women aged 25 to 45 years were divided into two groups. One group of the grape leaf extract of Reference Example 1, 0.2 g of the berry mixture of Reference Example 2, 0.2 g of selenium, 1 g of lactose, 1.1 g of glycerin, And 0.5 g of xylitol were mixed in a usual manner to prepare 4 g per one ring. The control group was administered with the active ingredients (red grape leaf extract, berry mixture and selenium ) Was replaced with 1.4 g of glucose, and the ring was prepared by the same method. After the test, a questionnaire about the skin condition was conducted, and the results are shown in Table 6 below.

Survey item Experimental group Control group 20's
(4 people) 30s
(10 people) 40s
(6 people) Sum
(20 people) 20's
(6 people) 30s
(8 people) 40s
(6 people) Sum
(20 people) Your skin becomes moist 3 8 5 16 (80%) 2 One 2 5 (25%) The skin becomes taut 2 8 5 15 (75%) One 2 One 4 (20%) Reduced fine lines 2 8 4 14 (70%) One 3 2 6 (30%) Makeup is well received 3 9 5 17 (85%) 2 2 3 7 (35%) Overall skin improved 3 7 4 14 (70%) 2 3 One 6 (30%)

As can be seen from the results in Table 6, the experimental group showed a higher rate of feeling a moisturizing or elastic feeling of the skin than the control group, and many people felt that the fine lines had decreased and the makeup was well received. It was found that the use of a composition containing a mixed antioxidant containing red grape leaf extract can improve the skin condition as a whole.

The composition for oral skin beauty improvement provided by the present invention can be applied to various formulations as described below, but the present invention is not limited thereto.

[Formulation Example 1] Soft capsule

The mixture was mixed with 200 mg of the antioxidant of Example 4, 50 mg of soybean extract, 180 mg of soybean oil, 50 mg of red ginseng extract, 2 mg of palm oil, 8 mg of palm kernel oil, 4 mg of yellow pearls and 6 mg of lecithin, 400 mg per well was filled to prepare a soft capsule.

[Formulation Example 2] Tablets

After granules were formed by mixing 200 mg of the antioxidant of Example 4, 50 mg of soybean extract, 100 mg of glucose, 50 mg of red ginseng extract, 96 mg of starch and 4 mg of magnesium stearate, and 40 mg of 30% ethanol, Dried at 60 ° C and tableted using a tablet machine. The final weight of the contents was 400 mg.

[Formulation Example 3]

200 mg of the antioxidant of Example 4, 50 mg of soybean extract, 100 mg of glucose, 50 mg of red ginseng extract and 600 mg of starch were mixed and 100 mg of 30% ethanol was added to form granules, And then filling the pores. The final weight of the contents was 1 g.

[Formulation Example 4] Drinking agent

200 mg of the antioxidant of Example 4, 50 mg of soybean extract, 10 g of glucose, 50 mg of red ginseng extract, 2 g of citric acid and 188 g of purified water were mixed and filled in a bottle. The final weight of the contents was 100 ml.

Fig. 1 shows a standard curve of absorbance inhibition rate according to the change in trolox concentration described in Test Example 3 of the present invention.

2 is a photograph of skin molds for the control, UV control, Comparative Examples 4 and 2.

Claims (6)

Red grape leaf extract, berry mixture and selenium as active ingredients, Wherein the berry mixture is composed of pomegranate, raspberry, blackberry, acai, cranberry and blueberry. The antioxidative food composition according to claim 1, wherein the grape leaf extract, the berry mixture and the selenium are mixed in a weight ratio of 1: 1: 1 to 10: 1: 1. delete The antioxidative food composition according to claim 1, wherein the composition is for removing active oxygen in the body. The antioxidative food composition according to claim 1, wherein the composition is for preventing the reduction of the activity of antioxidant enzymes in the body. The antioxidative food composition according to claim 1, wherein the composition is for preventing aging.

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