KR102183163B1 - Cleanser including Grape Seeds Extract Nano Emulsion and Manufacturing Method - Google Patents

Abstract

The present invention relates to a cleanser comprising a grape seed extract nanoemulsion and a method for preparing the same, and more particularly, to obtain a grape seed extract by optimizing an extraction solvent for maximizing the extraction of active ingredients having antibacterial properties and antioxidant properties from grape seeds. , Using the obtained grape seed extract and additives, it contains a grape seed extract nanoemulsion prepared to have excellent dispersion stability, antibacterial activity, and antioxidant activity even when stored at room temperature for more than 6 months, and a cleanser with enhanced moisturizing power and skin stability and preparation thereof It's about how.
The method for preparing a cleanser containing the grape seed extract nanoemulsion of the present invention includes the step of obtaining a grape seed extract (S100) for obtaining a grape seed extract using an extraction solvent; and a nanoemulsion manufacturing step (S200) for preparing a nanoemulsion including the grape seed extract. ); and distilled water, emulsifier, grape seed extract nanoemulsion, foam stabilizer, glycerin, citric acid, ethanol, preservative, mixing step of mixing a cleaning composition (S300); includes. The cleanser prepared by adding the grape seed nanoemulsion as described above has excellent moisturizing power and skin safety.

Description

Cleanser including Grape Seeds Extract Nano Emulsion and Manufacturing Method {Cleanser including Grape Seeds Extract Nano Emulsion and Manufacturing Method}

The present invention relates to a cleanser comprising a grape seed extract nanoemulsion and a method for preparing the same, and more particularly, to obtain a grape seed extract by optimizing an extraction solvent for maximizing the extraction of active ingredients having antibacterial properties and antioxidant properties from grape seeds. , Using the obtained grape seed extract and additives, it contains a grape seed extract nanoemulsion prepared to have excellent dispersion stability, antibacterial activity, and antioxidant activity even when stored at room temperature for more than 6 months, and a cleanser with enhanced moisturizing power and skin stability and preparation thereof It's about how.

Grapes not only contain sugars, organic acids and unique fragrances, but also anticancer, antihypertensive and antioxidant polyphenol compounds such as anthocyanin pigments, phenolic acids, flavonoids (flavonols flavan-3-ols and flavanonols) and resveratrol. As it contains, it is steadily consumed as various processed foods such as grape juice, wine and grape vinegar.

On the other hand, grape seeds account for about 3 to 5% of the weight of grapes, and contain large amounts of dietary fiber (about 45%) such as fat (9 to 12%), protein (8 to 12%) and hemicellulose. In addition, it is known that it contains a high content of minerals such as Ca, Mg and P, contains a large amount of unsaturated fatty acids such as linoleic acid, and contains a large amount of vegetable sterols, tocopherols and anticancer, antihypertensive and antioxidant catechins.

However, despite the many advantages of grape seeds, most of them are not utilized and are being disposed of.

Therefore, for the purpose of utilizing conventional grape seeds, Korean Patent No. 10-0341933 proposes a method of fermenting black grape seeds, skin, and stems, which are black grape residues, to produce a fermented extract, and Korean Patent No. 10-1787406 In the issue, a cosmetic composition containing bromelain, butcherbroom, grape seed, arnica and lemon balm as active ingredients is proposed, and in Korean Patent No. 10-1086504, it has a function of improving blood circulation and expanding blood vessels including grape seed oil. A composition for application to the skin is presented.

The present inventors developed a grape seed extract nanoemulsion that utilizes the waste processed grape seeds, contains a large amount of active ingredients of grape seeds, and has excellent dispersion stability, antibacterial activity, and antioxidant activity even when stored at room temperature for 6 months or longer. As part of a study to be applied to the cleaning composition, an extraction solvent was derived to maximize the extraction of active ingredients of grape seeds, and the obtained grape seed extract and additives were combined to ensure dispersion stability when stored at room temperature for 6 months or longer. , A nanoemulsion having excellent antibacterial activity and antioxidant activity was prepared, and a skin cleanser with enhanced moisturizing power and skin safety was prepared using this, and the present invention was reached.

Domestic registered patent No. 10-0341933 Domestic registered patent No. 10-1787406 Domestic registered patent No. 10-1086504

An object of the present invention for solving the above problems is to obtain a grape seed extract by optimizing an extraction solvent for maximizing the extraction of an active ingredient having antibacterial and antioxidant properties from grape seeds, and using the obtained grape seed extract and additives It includes a grape seed extract nanoemulsion prepared to have excellent dispersion stability, antibacterial activity, and antioxidant activity even when stored at room temperature for more than 6 months, and to provide a cleanser with enhanced moisturizing power and skin stability, and a method of manufacturing the same.

The method for preparing a cleanser comprising the grape seed extract nanoemulsion of the present invention for solving the above problem is the step of obtaining a grape seed extract (S100) for obtaining a grape seed extract using an extraction solvent; and preparing a nanoemulsion including the grape seed extract. Nano-emulsion manufacturing step (S200); And distilled water, emulsifier, grape seed extract nano-emulsion, foam stabilizer, glycerin, citric acid, ethanol, preservatives, a cleaning composition mixing step (S300) of mixing a fragrance; includes.

In the step of obtaining the grape seed extract (S100), the grape seed powder was added to an extraction solvent containing any one of distilled water, an organic solvent, and a combination thereof, and stirred at 30 to 45°C for 12 to 36 hours, at 50 to 300 rpm, and then filtered. And concentrated.

The nano-emulsion manufacturing step (S200) is a blending step (S210) of blending the grape seed extract with any one of surfactants, oils, lecithin, polyhydric alcohol, and combinations thereof; and a stirring step of stirring the blend to form a mixture. (S220); and the ultrasonic treatment step (S230) by ultrasonicating the mixture at 10 to 30 khz for 5 to 20 minutes.

The cleaning composition mixing step (S300) is based on 100 parts by weight of distilled water, 50 to 85 parts by weight of emulsifier, 0.001 to 5 parts by weight of grape seed extract nanoemulsion, 0.5 to 5 parts by weight of foam stabilizer, 1 to 5 parts by weight of glycerin, 0.1 parts by weight of citric acid It is characterized by mixing to 3 parts by weight, 1 to 5 parts by weight of ethanol, 0.01 to 0.2 parts by weight of preservative, and 0.01 to 3 parts by weight of perfume.

Cleanser comprising the grape seed extract nanoemulsion of the present invention for solving the above problems is characterized in that it contains distilled water, emulsifier, grape seed extract nanoemulsion, foam stabilizer, glycerin, citric acid, ethanol, preservatives, fragrance.

The cleanser comprising the grape seed extract nanoemulsion of the present invention is based on 100 parts by weight of distilled water, 50 to 85 parts by weight of emulsifier, 0.001 to 5 parts by weight of grape seed extract nanoemulsion, 0.5 to 5 parts by weight of foam stabilizer, 1 to 5 parts by weight of glycerin It is characterized in that it comprises 0.1 to 3 parts by weight of citric acid, 1 to 5 parts by weight of ethanol, 0.01 to 0.2 parts by weight of preservative, and 0.01 to 3 parts by weight of flavor.

The grape seed extract nanoemulsion is formulated and stirred with any one of surfactants, oils, lecithin, polyhydric alcohol, and combinations thereof, and grape seed extract to form a mixture, and the mixture is mixed at 10 to 30 khz for 5 minutes to 20 minutes. It is characterized in that it is obtained by sonication.

The grape seed extract is obtained by adding grape seed powder to an extraction solvent containing any one of distilled water, an organic solvent, and a combination thereof, stirred at 30 to 45°C for 12 to 36 hours, at 50 to 300 rpm, and then filtered and concentrated. It features.

As described above, according to the cleanser including the grape seed extract nanoemulsion according to the present invention and the preparation method thereof, the extraction solvent for maximizing the extraction of active ingredients having antibacterial properties and antioxidant properties from grape seeds is optimized to obtain grape seed extract. And, using the obtained grape seed extract and additives, it contains a grape seed extract nanoemulsion prepared to have excellent dispersion stability, antibacterial activity, and antioxidant activity even when stored at room temperature for more than 6 months, and a cleanser with enhanced moisturizing power and skin stability, and its There is an effect of providing a manufacturing method.

Figure 1 is a flow chart showing a cleanser comprising a grape seed extract nanoemulsion according to the present invention.
Figure 2 is a photograph showing a state of concentrating the grape seed extract in the grape seed extract obtaining step (S100) of the method for producing a cleanser containing a grape seed extract nanoemulsion according to the present invention.
Figure 3 is a photograph showing the grape seed extract obtained using the extraction solvent in the grape seed extract obtaining step (S100) of the method for producing a cleanser containing a grape seed extract nanoemulsion according to the present invention.
Figure 4 is a photograph showing the antibacterial activity of the grape seed extract obtained by the grape seed extract obtaining step (S100) of the method for producing a cleanser containing a grape seed extract nanoemulsion according to the present invention.
5 is a graph showing the DPPH free radical scavenging ability of grape seed extract according to the extraction solvent and concentration.
6 is a graph showing the content of phenolic compounds in 100% ethanol extract.
7 is a graph showing the total flavonoid content of 100% ethanol extract.
8 is a photograph showing the dispersion stability of nanoemulsions prepared according to Examples 1 to 5.
9 is a result of dynamic light scattering (DLS) analysis conducted after storing Example 5 containing 70% ethanol extract at room temperature for 6 months or longer.
10 is a dynamic light scattering (DLS) analysis result performed after storing Example 5 containing 100% ethanol extract for 6 months or more at room temperature.
11 is a polydispersity index and zeta potential analysis results performed after storing Example 5 containing 70% ethanol extract at room temperature for 6 months or longer.
12 is a result of polydispersity index and zeta potential analysis conducted after storing Example 5 containing 100% ethanol extract at room temperature for 6 months or longer.
13 is a graph showing the free radical scavenging ability and IC50 according to the amount of the nanoemulsion formulation after being stored at room temperature for at least 6 months after preparation.
14 is a photograph showing the ability to inhibit bacteria against E. coli and B. cereus, which was carried out after storing Example 5 containing 100% ethanol extract at room temperature for 6 months or longer.
Figure 15 (a) shows the position for measuring the moisturizing power, (b) is a test result of the moisturizing effect of the grape seed extract nanoemulsion cleanser after hand washing and forced drying.
Figure 16 is a photograph showing a dry state after applying a 1% crystal violet solution on the skin.
FIG. 17 is a comparative graph showing the cleaning power of foam cleansing including no treatment (washing only with tap water), control (washing using Dr. Belmeur foam cleanser), and grape seed extract nanoemulsion.
18 is a 3D image analysis result of the dyed area before and after washing.

Specific features and advantages of the present invention will be described in detail below with reference to the accompanying drawings. Prior to this, when it is determined that a detailed description of functions and configurations related to the present invention may unnecessarily obscure the subject matter of the present invention, a detailed description will be omitted.

The present invention relates to a cleanser comprising a grape seed extract nanoemulsion and a method for preparing the same, and more particularly, to obtain a grape seed extract by optimizing an extraction solvent for maximizing the extraction of active ingredients having antibacterial properties and antioxidant properties from grape seeds. , Using the obtained grape seed extract and additives, it contains a grape seed extract nanoemulsion prepared to have excellent dispersion stability, antibacterial activity, and antioxidant activity even when stored at room temperature for more than 6 months, and a cleanser with enhanced moisturizing power and skin stability and preparation thereof It's about how.

1 is a flow chart showing a method of manufacturing a cleanser comprising a grape seed extract nanoemulsion according to the present invention.

The method for preparing a cleanser containing the grape seed extract nanoemulsion of the present invention includes a grape seed extract obtaining step (S100) for obtaining a grape seed extract using an extraction solvent and a nanoemulsion preparation step (S200) for preparing a nano emulsion including the grape seed extract. And distilled water, emulsifier, grape seed extract nanoemulsion, foam stabilizer, glycerin, citric acid, ethanol, preservative, and a cleaning composition mixing step (S300) of mixing.

Grape seed extract obtaining step (S100) is a step of obtaining a grape seed extract using an extraction solvent, by adding grape seed powder to an extraction solvent containing any one of organic solvents such as distilled water and ethanol, and a combination thereof at 30 to 45°C. After stirring at 12 to 36 hours, 50 to 300 rpm, it is filtered and concentrated to prepare a grape seed extract.

The average particle diameter of the grape seed powder is not limited, but preferably, those having an average particle diameter of 5 to 100 μm are used.

As an extraction solvent for extracting secondary metabolites having antimicrobial activity and antioxidant properties from grape seed powder, any one of organic solvents such as distilled water and ethanol, and combinations thereof may be used.

The organic solvent is preferably at least one selected from the group consisting of a lower alcohol having 1 to 4 carbon atoms, hexane, ethyl acetate, dichloromethane, ether, chloroform and acetone. The lower alcohol having 1 to 4 carbon atoms is preferably at least one selected from the group consisting of ethanol, methanol, propanol, isopropanol, butanol, and n-butanol, and the lower alcohol having 1 to 4 carbon atoms may include both anhydrous or hydrous alcohols. I can.

Preferably, ethanol may be used, more preferably, 70 to 100% (v/v) ethanol may be used.

Grape seed powder is added to the extraction solvent so that it becomes 20-50% (w/v), and when an organic solvent is used as an extraction solvent, 30-45℃, in the case of using distilled water, 12-36 hours at 70-120℃, 50-300rpm Stir in. After completing the stirring, it is filtered using filter paper or the like, and the filtrate is concentrated to have a moisture content of 5 to 30 wt% using a rotary evaporator.

In the nano-emulsion manufacturing step (S200), the blending step (S210) of blending the grape seed extract with any one of surfactants, oils, lecithin, polyhydric alcohol, and combinations thereof, and a stirring step of stirring the blend to form a mixture (S220) ) And the mixture by ultrasonication at 10 to 30 khz for 5 to 20 minutes, and an ultrasonic treatment step (S230).

In the blending step (S210), the grape seed extract is blended with any one of surfactants, oils, lecithin, polyhydric alcohol, and combinations thereof.

Surfactants include any one of polysorbate 80 (Tween 80), polysorbate 20 (Tween 20), polysorbate 40 (Tween 40), polysorbate 60 (Tween 60), and combinations thereof. It can be, but is not limited thereto. The surfactant may be blended in an amount of 80 to 250 parts by volume based on 100 parts by volume of the grape seed extract.

Oils include MCT (Medium Chain Triglyceride), Carrot Oil, Peanut Oil, Almond Oil, Jojoba Oil, Apricot Seed Oil, Avocado Oil, Canola Oil, Dharmaji Flower Seed Oil, Grape Seed Oil, Olive Oil, Rice Bran Oil, Rosehip Oil, Sesame Oil , Soybean oil, sunflower seed oil, coconut oil, and combinations thereof may be included, but are not limited thereto. Oils may be blended in 80 to 150 parts by volume based on 100 parts by volume of the grape seed extract.

Lecithin is at least one lecithin selected from the group consisting of hydrogenated lecithin, hydrogenated phosphatidylcholine, phospholipid, hydrogenated lysophosphatidylcholine, hydrogenated lysolecithin, hydrooxylated lecithin, and unsaturated lecithin, and their Any one of the combinations may be included, but the present invention is not limited thereto. Lecithin may be blended with 10 to 50 parts by volume based on 100 parts by volume of the grape seed extract.

The polyhydric alcohol may include any one of glycerol, propylene glycol, dipropylene glycol, 1,3-butylene glycol, ethylene glycol, sorbitol, and combinations thereof, but is not limited thereto. Polyhydric alcohol may be blended in 80 to 150 parts by volume based on 100 parts by volume of the grape seed extract.

In the stirring step (S220), the mixture is stirred to form a mixture, and the mixture is uniformly mixed by stirring at 50 to 400 rpm for 3 to 20 minutes.

In the ultrasonic treatment step (S230), the mixture is sonicated for 5 to 20 minutes at 10 to 30 khz. At this time, it is preferable to process in an ice bucket so that heat denaturation and oxidation do not occur in the mixture due to heat generated during ultrasonic treatment.

The cleaning composition mixing step (S300) is a step of mixing distilled water, emulsifier, grape seed extract nanoemulsion, foam stabilizer, glycerin, citric acid, ethanol, preservative, fragrance, specifically, based on 100 parts by weight of distilled water, emulsifier 50 to 85 Part by weight, grape seed extract nanoemulsion 0.001 to 5 parts by weight, foam stabilizer 0.5 to 5 parts by weight, glycerin 1 to 5 parts by weight, citric acid 0.1 to 3 parts by weight, ethanol 1 to 5 parts by weight, preservative 0.01 to 0.2 parts by weight, fragrance 0.01 to 3 parts by weight are mixed. Mixing is performed at room temperature (20~25℃), and stirring is performed at a stirring speed of 50-300 rpm for 5 to 60 minutes until a homogeneous state is achieved.

The cleanser comprising the grape seed extract nanoemulsion according to the present invention is prepared by the method of manufacturing a cleanser comprising the grape seed extract nanoemulsion described above.

Cleanser containing grape seed extract nanoemulsion according to the present invention includes distilled water, emulsifier, grape seed extract nanoemulsion, foam stabilizer, glycerin, citric acid, ethanol, preservatives, fragrances, and specifically, based on 100 parts by weight of distilled water, emulsifier 50 to 85 parts by weight, grape seed extract nanoemulsion 0.001 to 5 parts by weight, foam stabilizer 0.5 to 5 parts by weight, glycerin 1 to 5 parts by weight, citric acid 0.1 to 3 parts by weight, ethanol 1 to 5 parts by weight, preservatives 0.01 to 0.2 parts by weight Parts and 0.01 to 3 parts by weight of perfume.

The grape seed extract nanoemulsion is formulated and stirred with any one of surfactants, oils, lecithin, polyhydric alcohol, and combinations thereof, and grape seed extract to form a mixture, and the mixture is mixed at 10 to 30 khz for 5 minutes to 20 minutes. Obtained by sonication.

The grape seed extract is obtained by adding grape seed powder to an extraction solvent containing any one of organic solvents such as distilled water and ethanol, and a combination thereof, stirred at 30 to 45°C for 12 to 36 hours, at 50 to 300 rpm, and then filtered and concentrated. Obtained by

Hereinafter, the present invention will be described in detail with reference to a preferred embodiment. However, the following examples are intended to specifically illustrate the present invention, and are not limited thereto.

1. Experimental method and analysis method

1-1. Obtaining grape seed extract

Dried grape seeds were purchased and pulverized from Pacific Products, located in Gyeongsan, Gyeongsangbuk-do. High temperature distilled water (90 ~ 100 ℃) and 30, 50, 70 and 100% ethanol were prepared as an extraction solvent. 140 g of grape seed powder was added to 420 ml of extraction solvent in a 500 ml sterile triangular flask, and then wrapped with aluminum foil and stirred at 37° C. for 24 hours at 100 rpm. After stirring, the mixture was filtered using Whatman filter paper (Grade 1: 11 μm, Sigma Aldrich), and the filtrate was concentrated using a rotary evaporator. Figure 2 shows a state of concentrating the filtered grape seed extract.

1-2. Reaction reagent

Ethanol, 2,2-diphenyl-1-picrylhydrazyl (DPPH), Folin's reagent, sodium carbonate, aluminum chloride, sodium acetate, quercetin, and gallic acid were used as reaction reagents for the experiment, and the reagents were from Merck or Sigma. Purchased, all reagents were used for analysis grade.

1-3. Antimicrobial analysis

Antimicrobial activity of grape seed extract against Escherichia coli (ATCC 43889) and Bacillus cereus (ATCC 13061) was confirmed using an Agar well diffusion assay. E. coli (ATCC 43889) and B. cereus (ATCC 13061) were cultured overnight in Luria-Bertani (LB) liquid medium, and 10 ⁹ cells when the growth of bacteria reached a stop (OD600∼2). /mL 100 μl was plated on LB medium.

A well of 8 mm diameter was formed in the inoculated medium, 100 μl of grape seed extract (1 mg/ml) was added to a labeled well, and the medium was cultured at 37° C. for 12 hours. The broth micro-dilution technique was used to measure the minimum inhibitory concentration (MIC) of grape seed extract against E. coli (ATCC 43889) and B. cereus (ATCC 13061). According to CLSI (Clinical and Laboratory Standards Institute), it was carried out using untreated polystyrene microfilter plate CC7672-7596; (CytoOne). As a result of observation after culturing at 37° C. for 24 hours, it was shown that the growth of bacteria can be inhibited visually.

1-4. DPPH Radical Scavenging ability

Free radicals increase the risk of causing oxidative stress and many diseases in the human body, and antioxidants are known to be effective in reducing the production of free radicals. Antioxidants directly or indirectly react with active radicals to inhibit the reaction of enzymes that generate free radicals. In addition, antioxidants enhance the body's defense mechanisms by promoting the expression of other antioxidant enzymes.

DPPH radical scavenging ability is a function of inducing stabilization by providing proton ions with a reducing function to unstable free radicals, and plays a role in stabilizing unstable and harmful free radicals occurring in vivo. Therefore, it is an index used when evaluating the antioxidant ability of an unknown specific substance to remove hydroxyl radicals or superoxide radicals generated by physiological action or oxidation of a living body. The higher the value, the better the antioxidant ability.

After adding 3 mL of 0.1 mM DPPH solution to 1 mL of the sample solution, mixing and reacting for 30 minutes in a dark room at room temperature, absorbance was measured at 517 nm using a spectrophotometer (UV-2120 Optizen, Mecasys, South Korea). Was calculated using

Erasing ability% = [1-(At /A0)]×100

Here, At and A0 are the absorbances of the sample solution added group and the non-added group, respectively. Gallic acid was used as a positive control, and the result was repeated three times to show the average value.

1-5. Determination of total phenol content

The total phenol content of grape seed extract was measured using the Folin-Ciocalteau method. Galic acid and grape seed extract were diluted with water as a standard, and a folin reagent (0.5ml) was added and reacted at 22°C for 5 minutes. Thereafter, 4 mL of sodium carbonate solution (20 g/100 mL) was added, and distilled water was added to 10 mL to form a mixture. Incubated at room temperature (37 °C) for 90 minutes, and the absorbance was measured at 725 nm using a spectrophotometer (UV-2120 Optizen, Mecasys, South Korea), and compared with the standard curve of Galic acid. Total phenol content was calculated using the standard curve of Galic acid.

1-6. gun Polaronoid Measurement of content

The content of total flavonoids was quantified using the colorimetric method of aluminum chloride.Quercetin and grape seed extract as standard substances were mixed with 0.1 mL of 10% aluminum chloride, 0.1 mL of 1M potassium acetate, and 2.8 mL of distilled water, and then allowed to stand at room temperature for 30 minutes. Then, the absorbance was measured at 415 nm using a spectrophotometer (UV-2120 Optizen, Mecasys, South Korea). The total flavonoid content was expressed through a standard calibration curve prepared using quercetin as a standard material.

1-7. Statistics processing

Unless otherwise noted, all samples were analyzed 3 times and expressed as Mean±SD.

1-8. Nano Emulsion Produce

The nanoemulsion containing grape seed extract was formulated as shown in Table 1 below.

In Examples 1 to 4, grape seed extract extracted from 100% ethanol was used, and in Example 5, grape seed extract extracted from 70% and 100% ethanol was used. MCT was prepared as an oil, polysorbate 80 as a surfactant, and glycerol as a polyhydric alcohol. Examples 1 and 4 were 1% (w/v) grape seed extract, Examples 2 and 3 were 2% (w/v) grape seed extract, and Example 5 was 0.5% (w/v) ) Of grape seed extract was used.

Grape seed extract and reagents were prepared and added at the blending ratio according to Table 1, and distilled water was added until it became 50 ml. After the mixture was stirred for 5 minutes, it was treated for 10 minutes in 25 khz ultrasound (Ultrasonic Processor, GEX 750, Newtown, CT, 750 W). In order to prevent denaturation of the mixture due to heat generated during ultrasonic treatment, the test tube containing the mixture was immersed in an ice bucket and treated.

The droplet size and zeta potential of the grape seed extract nanoemulsion were analyzed by dynamic light scattering (DLS), and a zeta potential and particle size analyzer (ELSZ-2000, Otsuka Electronics Co., Ltd. Japan) was used.

2. Experiment result

2-1. Yield of grape seed extract

3 is a photograph showing the obtained grape seed extract.

Table 2 shows the yield of grape seed extract according to the type of solvent.

The 100, 70, 50, and 30% ethanol extracts showed yields of 5.17, 5.57, 3.98 and 4.67% (w/w), respectively, and the hot water extracts showed a yield of 6.1%.

2-2. Antimicrobial Activity of Grape Seed Extract

Figure 4 is a photograph showing the antibacterial activity of grape seed extract. Among the bacteria, E. coli showed better antibacterial activity than B. cereus.

Table 3 shows the antibacterial activity of grape seed extract.

In particular, grape seed extract extracted in 100% ethanol solvent (hereinafter, described abbreviated as 100% ethanol extract.) showed higher inhibitory activity compared to other extraction solvents (70, 50 and 30% ethanol extract). In E. coli medium, the inhibition area of 100% ethanol extract was 28±1.1 mm, and in the case of B. cereus, it was 13±0.7 mm. In addition, the hot-water extract did not show antibacterial activity against all strains used in the experiment despite the high 50 mg/ml. For the experimental strain, the MIC of the 100% ethanol extract was lower than that of the 70, 50 and 30% ethanol extracts, so the antimicrobial activity was relatively high. The MIC of 100% ethanol extract was 0.3 mg/ml and 0.8 mg/ml for E. coli and B. cereus, respectively. The hot water extract did not show any antibacterial activity, so it was excluded from subsequent experiments.

2-3. DPPH Free radical Scavenging ability

There was a large deviation in DPPH activity depending on the extraction solvent, and it was confirmed that the 100% ethanol extract had high antioxidant activity. Through this, it was possible to hypothesize that the dissolution capacity of secondary metabolites of plants differs depending on the concentration of ethanol. 5 and Table 4 show the IC50 for DPPH free radical scavenging ability of grape seed extract according to the extraction solvent and concentration.

The IC50 of the 100% ethanol extract was found to be 102.1 ug/ml, and the IC50 of the 70, 50 and 30% ethanol extracts were 106.1, 197.1, and 259.3 ug/ml, respectively. The IC50 of galic acid used as a positive control was found to be 53.78 ug/ml. Through the above results, it was determined that the 100% ethanol extract was an appropriate solvent for extracting the secondary metabolites of grape seeds, and then the biochemical properties were studied using the 100% ethanol extract.

2-4. Total phenol content analysis

Phenolic compounds directly or indirectly provide antioxidant effects by strengthening the body's immune system and suppressing inflammatory reactions. The phenolic compound contained in the fruit is bound to the cell wall and is present in a soluble state.In the present invention, the extraction content of the phenolic compound of grape seeds according to the extraction solvent was measured, and the content of the phenolic compound in the 100% ethanol extract was 58.6 mg It showed GAE/50 mg (shown in Figure 6). 6 shows the total phenol content measured in 100% ethanol extract and Galic acid used as a control.

2-5. Total flavonoid content analysis

As a result of comparing the 100% ethanol extract with the positive control, the 100% ethanol extract showed 5.944 mg QE/50 mg in quercetin equivalent, while the total flavonoid content of 40 mg quercetin was 6.322 mg. Through this result, it was confirmed that the total flavonoid content of the 100% ethanol extract was smaller than that of quercetin, but the concentration of ethanol had an effect on the total flavonoid content. 7 is a comparison of total flavonoid content.

2-6. Grape Seed Extract Nano Emulsion Dispersion stability

Nanoemulsions have various advantages over general emulsions. In order to control the optical transparency of the nanoemulsion, stability against phase separation, particle agglomeration, and bioavailability, control of various variables is required. The addition of stabilizers and adjuvants affects the interaction between molecules and the rheological behavior, and thus the stabilization and physical properties of the nanoemulsion. Thus, in the present invention, a nanoemulsion was formed using ethanol extracts of grape seeds having different protocols, and Table 5 below shows the dispersion stability of the nanoemulsion. The nanoemulsion formulation prepared by the method of Example 5 stably maintained the formulation even after 6 months at room temperature, and particle size and zeta potential were measured to confirm the detailed characteristics of Example 5. 8A to 8E show nanoemulsions according to Examples 1 to 5.

Example 1 Layer separation occurs after 24 hours Example 2 Layer separation occurs after 42 hours Example 3 Layer separation occurs after 1 week Example 4 Layer separation occurs after 2 weeks Example 5 Maintains the formulation stably even after 6 months at room temperature

2-7. Formed nano Emulsion characteristic

A nanoemulsion formulation containing grape seed extract extracted with 70% and 100% ethanol using dynamic light scattering (DLS) was stored at room temperature for 6 months, and then the droplet size of Example 5 was measured, and FIG. 9 shows a 70% ethanol extract. It is the droplet size of Example 5 containing, Figure 10 shows the droplet size of Example 5 containing 100% ethanol extract. The droplet size of the nanoemulsion was 181.3 nm and 221.5 nm, respectively, of grape seed extract extracted with 70% and 100% ethanol. Zeta potentials were 62.73 mV and 49.92 mV in grape seed extracts extracted with 70% and 100% ethanol, respectively. 11 is a polydispersity index and zeta potential of a nanoemulsion (70% ethanol extract) measured through dynamic light scattering (DLS) analysis, and FIG. 12 is a nanoemulsion 100 measured through dynamic light scattering (DLS) analysis. % Ethanol extract) and the zeta potential.

In order to confirm the biological properties of the formed nanoemulsion, the DPPH free radical scavenging ability of the 100% ethanol extract was confirmed after storage at room temperature for 6 months in a nanoemulsion formulation. 13 shows the free radical scavenging ability and IC50 according to the input amount. As a result, the I50 of the 100% ethanol extract was 97.72 ug/ml.

Example 5 showed a MIC of 0.25 mg/ml for antimicrobial activity after 6 months storage at room temperature in a nanoemulsion formulation. Before treatment with nanoemulsion, grape seed extract showed a lower value than that of E. coli and B. cereus 0.3mg/ml and 0.8mg/ml. Compared to the simple extract of grape seed, after storage at room temperature for 6 months It can be seen that the antibacterial activity was better in the nanoemulsion formulation.

14 shows the antibacterial activity of the nanoemulsion against E. coli and B. cereus.

Table 6 shows the microbial minimum inhibitory concentration of the nanoemulsion.

In the present invention, it was confirmed that the ethanol extract of grape seed contains phenol and flavonoids, showing excellent antibacterial activity. It was suggested that the total phenol content and flavonoid content of grape seeds identified in this experiment can increase the utility value of grape seeds.

In addition, in the present invention, grape seed extract is obtained by optimizing the extraction solvent for maximizing the extraction of secondary metabolites having antibacterial properties and antioxidant properties from grape seeds, and nanoemulsion having excellent dispersion stability using the obtained grape seed extract and additives Was prepared. When comparing the antimicrobial and antioxidant properties of the nanoemulsion using grape seed extract and grape seed extract, the antibacterial and antioxidant properties of the nanoemulsion were more excellent.Through this, the interaction between grape seed extract and additives was found to have antimicrobial and antioxidant properties. It suggested that it can be further improved.

In order to apply the grape seed extract emulsion according to the present invention to a skin cleanser, it was prepared as follows.

3. Preparation of a cleanser containing grape seed extract nanoemulsion

Distilled water 51.985wt%, ASCO 141 (anionic emulsifier) 5.72wt%, ASCO 24-3/28 (anionic emulsifier) 35.71wt%, ASCO CDE (foam stabilizer) 1.00wt%, glycerin 2.00wt%, citric acid 0.50wt% , Ethanol 3.00wt%, K-CG (preservative) 0.03wt%, grape seed extract nanoemulsion 0.005wt%, lemon flavor was mixed with 0.05wt% to prepare a cleanser.

4. Evaluation

4-1. Moisturizing power

In order to evaluate the moisturizing power, the part of the body where the moisturizing power appears constant was determined. Figure 15 (a) shows the location for measuring the moisturizing power, and (b) shows the test results of the moisturizing effect upon forced drying after hand washing of the grape seed extract nanoemulsion cleanser.

According to the above results, the moisture content was most stably measured regardless of the number of rounds in the area 2 from the back of the hand to the arm, and the future moisturizing power test was measured in the area 2.

Table 7 shows the measured values and average values of the moisture content for each cycle in a defined body part.

Moisturizing power was measured by the following method. Grape seed extract nanoemulsion foam cleansing prototype samples were treated on each site 2, washed hands with tap water, and then measured three times for 1 hour at intervals of 5 minutes while forcibly blowing with cold air to determine the moisture content as an average value. In the above method, the test product was not treated as a control and was compared after washing hands with tap water.

FIG. 15 shows the results of a test result of a moisturizing effect upon forced drying after hand washing of a grape seed extract nanoemulsion cleanser. Here, the red line is the moisturizing power when washing the grape seed extract nanoemulsion cleanser, and the blue line is the moisturizing power when washing tap water.

As a result, it was confirmed that the cleanser prototype to which the grape seed extract nanoemulsion material was added has a high moisturizing power (moisture content) of about 3 to 5% compared to the control.

4-2. Cleaning power

The cleaning power of the cleanser containing grape seed extract emulsion according to the present invention was confirmed by requesting the Daegu TP Oriental Medicine Industry Support Center. As a control sample, a commercially available liquid foam cleanser of a similar formulation was used, and the contaminant was a commercially available cream-formed lip color cosmetics.

All ingredients of the control product (J Company Essential Facial Foaming Cleanser) are as follows.

Purified water, triethanolamine, glycerin, myristic acid, lauric acid, lauryl phosphate, phenoxyethanol, cocamidopropylbetaine, cocamide MAE, sodium lauroamphoacetate, triclosan, fragrance, methylparaben, pantasodium tri Phosphate, propyl paraben, ethyl paraben, BHT, Red No. 504, Yellow No. 203 (Appearance: Orange liquid)

All contaminants (plum mellow glow lip lacquer) are listed below.

Purified water, octyldodecanol, trimethylsiloxyphenyldimethicone, ethanol, glyceryl stearate SE, ethylcellulose, sorbitan stearate, glycerin, PEG-40 stearate, phytosteryl/behenyl/octyldodecila Uroyl glutamate, diphenylmethylsiloxyphenylmethicone/phenylsilsesquioxane crosspolymer, acrylate copolymer, pentylene glycol, hydroxyethylacrylate/sodium acryloyldimethyltaurate copolymer, ethylhexyl Glycerin, trimethylpentaphenyltrisiloxane, tetraphenyldimethyldisiloxane, fragrance, plum seed oil, soribitan isostearate, polysorbate 60, barium sulfate, titanium dioxide, red iron oxide, red No. 226, Red No. 227, Black Iron Oxide (Appearance: Cream with viscosity)

The test piece has a weight of 7.75±0.17mg, and a filter paper disc (Smart Practice, 3400 E McDowell Rd, Phoenix, AZ 85008, USA) having a diameter of 8mm was used.

The pollutant capacity was 10ul/disc, the drying conditions were room temperature 22~24℃, humidity 40~60%, and air-dried indoors for 4 hours without air circulation. The ratio of the contaminant and the cleaning agent was 1:40 (w/v), and the sealed container containing the sample and the contaminant was allowed to stand at 40° C. for 18 hours and washed.

After washing, the test piece was placed on a paper towel to remove excess moisture, and then naturally dried for 4 hours. For the cleaning power, the pollutant removal efficiency was calculated from the weight of the initial pollutant and the weight of the removed pollutant, and the removal efficiency of the test sample and the control sample was compared.

The weight of contaminants before and after cleaning was measured using a chemical precision balance (TB215D, Satorious-Denver) after completely drying the test piece, excluding the weight of the test piece without contaminants. The cleaning power evaluation was performed using 3 test pieces for each cleaning agent.

The pollutant removal efficiency (Removal Efficiency, RE, %) was calculated according to Equation 1 below.

As a result of the test, the pollutant removal efficiency of the test sample, grape seed-containing foam cleanser was measured to be 72.5±3.28, and the commercial product as a control sample was 68.7±2.92, and the removal efficiency of the test sample was 5.5% contaminant removal efficiency compared to the control sample. Appeared to be high.

Table 8 below shows the results of the cleaning power test of the grape seed cleanser.

4-3. 3D analysis before and after cleaning contaminated skin areas

Before applying a 1% crystal violet solution, the hands were washed with tap water, dried at room temperature, and then a 100 μl solution of 1% crystal violet was applied to the skin and dried for 10 minutes. 16 is a photograph showing a dry state after applying a 1% crystal violet solution on the skin of subjects A and B.

After drying, wash until the dyed part disappears using foam cleansing containing no treatment (washing only with tap water), control (washing with Dr. Belmeur foam cleanser), grape seed extract nanoemulsion to check the cleaning power. I did. FIG. 17 is a comparative graph showing the cleaning power of foam cleansing including no treatment (washing only with tap water), control (washing using Dr. Belmeur foam cleanser), and grape seed extract nanoemulsion.

As a result, the control group showed 40% detergency, while the grape seed foam showed 60% detergency. When washing only with tap water, the pigment hardly disappeared.

In addition, the stained area before washing, the control group, and the stained area after washing with grape seed nanoemulsion foam cleansing were analyzed using 3D plot ImageJ software (version 1.45), and FIG. 18 shows the results of 3D image analysis of the stained area of subjects A and B. Show.

4-4. Skin stability (patch test)

The safety of the product was evaluated by confirming the presence or absence of primary irritation to the human skin of the foam cleanser (10 times diluted solution and 100 times diluted solution)' with grape seed extract added to the Korea Institute of Chemical Convergence Testing (KTR).

Thirty-one adult males and females were selected as test subjects, and the test site was selected on the back of the test subject where no skin abnormalities were observed.

After washing the test site with purified water, it was naturally dried for about 5 minutes, and 20 μL of the test sample was loaded onto the patch, and then the closed patch was applied for 24 hours. The patch was removed after 24 hours, and skin reactions of each test site were evaluated after 30 minutes, 24 hours, and 48 hours after the patch removal.

Visual evaluation of the test site is as follows It was conducted in accordance with the International Contact Dermatitis Research Group (ICDRG) criteria of Table 9. Visual evaluation was conducted by two or more testers, and if the results of the judgment were different between the two testers, a higher grade was selected.

After visual judgment, the average skin reactivity for each test site was calculated using the formula below, and the results of the human patch test were determined based on the skin reactivity as shown in Table 9.

Equation 2 below shows an equation for calculating the average skin reactivity.

Table 10 shows the judgment table of the results of the human patch test.

As a result of conducting a human patch test for safety evaluation over a total of 4 days, the skin reactivity was 0.01 for both the 10 times diluted solution of the foam cleanser added with grape seed extract and the 100 times diluted solution. It was judged as non-irritating according to the criterion for determining that 0.00 to 0.75 was non-irritating.

As described above, the present invention has been described based on preferred embodiments with reference to the accompanying drawings, but those of ordinary skill in the technical field to which the present invention pertains within the scope not departing from the technical spirit and scope described in the claims of the present invention. In various modifications or variations of the present invention can be implemented. Accordingly, the scope of the invention should be construed by the claims set forth to include examples of such many variations.

Claims (8)

In the method for producing a cleanser comprising a grape seed extract nanoemulsion having both antioxidant and antibacterial activity,
Grape seed powder having an average particle diameter of 5 to 100 μm was added to ethanol having a concentration of 70 to 100 (v/v)% as an extraction solvent, stirred at 30 to 45°C for 12 to 36 hours, at 50 to 300 rpm, and then the moisture content 5 Obtaining a grape seed extract to obtain a grape seed extract by filtration and concentration to have a to 30 wt% (S100); And
A nanoemulsion preparation step (S200) of preparing a nanoemulsion by stirring and ultrasonicating the mixture of the obtained grape seed extract and medium chain triglyceride (S200); and
Based on 100 parts by weight of distilled water, 50 to 85 parts by weight of emulsifier, 0.001 to 5 parts by weight of grape seed extract nanoemulsion, 0.5 to 5 parts by weight of foam stabilizer, 1 to 5 parts by weight of glycerin, 0.1 to 3 parts by weight of citric acid, 1 to 5 parts by weight of ethanol It characterized in that it comprises a cleaning composition mixing step (S300) of mixing parts by weight, 0.01 to 0.2 parts by weight of preservative, and 0.01 to 3 parts by weight of fragrance
Method for producing a cleanser comprising grape seed extract nanoemulsion.
delete The method of claim 1,
The nanoemulsion manufacturing step (S200)
Mixing step (S210) of mixing 80 to 150 parts by volume of medium chain triglyceride with respect to 100 parts by volume of grape seed extract and grape seed extract; and
Stirring step of stirring the blend to form a mixture (S220); And
Characterized in that it comprises; ultrasonic treatment step (S230) by ultrasonicating the mixture at 10 to 30 khz for 5 minutes to 20 minutes
Method for producing a cleanser comprising grape seed extract nanoemulsion.
delete In a cleanser comprising a grape seed extract nanoemulsion having both antioxidant and antibacterial activity,
Cleanser comprising the grape seed extract nanoemulsion
Based on 100 parts by weight of distilled water, 50 to 85 parts by weight of emulsifier, 0.001 to 5 parts by weight of grape seed extract nanoemulsion, 0.5 to 5 parts by weight of foam stabilizer, 1 to 5 parts by weight of glycerin, 0.1 to 3 parts by weight of citric acid, 1 to 5 parts by weight of ethanol Including parts by weight, 0.01 to 0.2 parts by weight of preservative, 0.01 to 3 parts by weight of fragrance,
The grape seed extract nanoemulsion is
Grape seed powder having an average particle diameter of 5 to 100 μm was added to ethanol having a concentration of 70 to 100 (v/v)% as an extraction solvent, stirred at 30 to 45°C for 12 to 36 hours, at 50 to 300 rpm, and then the moisture content 5 To 30 wt% of a grape seed extract obtained by filtration and concentration; and a mixture of medium chain triglyceride is obtained by stirring and sonicating
Cleanser containing grape seed extract nanoemulsion.
delete The method of claim 5,
The grape seed extract nanoemulsion is
A mixture was formed by mixing and stirring 80 to 150 parts by volume of a medium chain triglyceride with respect to 100 parts by volume of the grape seed extract and grape seed extract, and the mixture was sonicated at 10 to 30 khz for 5 to 20 minutes. doing
Cleanser containing grape seed extract nanoemulsion.

delete

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