KR102561353B1 - Nanoparticles encapsulated with propolis extract, preparation method thereof and antioxidant cosmetic composition containing the same as an active ingredient - Google Patents

Abstract

The present invention relates to nanoparticles encapsulated with propolis extract, a method for producing the same, and a cosmetic composition for antioxidant containing the same as an active ingredient. In addition, nanoparticles have excellent long-term stability, small size, high skin permeability, and excellent antioxidant effect, so that they can delay or inhibit skin aging, so they can be usefully used as antioxidant cosmetic compositions.

Description

Nanoparticles encapsulated with propolis extract, preparation method thereof and antioxidant cosmetic composition containing the same as an active ingredient}

The present invention relates to a cosmetic composition, and specifically, to a cosmetic composition containing nanoparticles for stabilizing poorly soluble propolis extract as an active ingredient.

Skin aging occurs largely through two processes. Changes that occur with time in anyone without special environmental factors are called endogenous aging, and changes that appear on the face, neck, and hands due to long-term exposure to environmental factors such as ultraviolet radiation are It is called photo aging or extrinsic aging.

The main cause of endogenous aging is the accumulation of damage to components of our body by reactive active oxygen radicals created during the metabolism of the human body. On the other hand, extrinsic aging is different in that harmful active oxygen is produced by ultraviolet rays. After the active oxygen is created, when the active oxygen is not effectively removed by various protective devices in the body, a series of inflammatory reactions occur, resulting in skin damage, and a typical example of the skin damage is skin wrinkling.

The main mechanism related to skin wrinkle formation is that the skin is continuously exposed to oxidative stress from harmful environments such as air pollution, UV exposure, stress or disease, so that radicals in the body increase and collagen, a connective tissue of the dermis, It is explained that it can cause subsidence (wrinkles) in certain parts of the skin by destroying elastin, hyaluronic acid, and the like. In addition, these mechanisms oxidize the lipid portion of the cell membrane and cause cell destruction, which can cause diseases such as dermatitis, acne, or skin cancer. In addition, radicals are involved in the melanin formation process and cause spots, freckles, and wrinkles.

In other words, aging appears as a result of the integration of various processes. According to many studies, it is reported that skin aging caused by external factors such as UV (ultra violet, ultraviolet) accounts for 80% of total aging rather than natural aging. In particular, since the skin is always in contact with the external environment and exposed to ultraviolet rays, it is highly likely to be photooxidatively damaged due to reactive oxygen species (ROS).

A complex antioxidant defense system is developed in the skin, and the system acts to protect the skin against active oxygen. Specifically, non-enzymatic antioxidant substances such as antioxidant enzymes, α-tocopherol, ascorbic acid, and carotenoids protect the skin from being oxidized. However, excessive production of free radicals from continuous UV exposure weakens the skin's enzymatic and non-enzymatic antioxidant defenses, and this oxidative stress eventually causes damage to cellular components. Specifically, damage to biocomponents such as lipid peroxidation, protein oxidation, chain scission and abnormal cross-linking of collagen and elastin, which are elastic fibers, severing of hyaluronic acid chains, promotion of melanin production, and DNA oxidation occurs, resulting in loss of elasticity and wrinkles. And skin aging such as melasma, freckles, etc. is accelerated. Therefore, skin aging can be delayed and inhibited when antioxidants are continuously supplemented through foods or cosmetics containing antioxidants to strengthen the skin antioxidant defense network.

On the other hand, propolis is a substance made by mixing bee saliva and enzymes with substances extracted from various plants for survival and reproduction by bees. . Propolis has been widely used as a folk medicine in Europe and Brazil since around 300 BC, and has recently been widely used in health foods and cosmetics.

Propolis is known to have antioxidant, antibiotic, antifungal, antiprotozoal, antitumor, and anti-inflammatory effects, and its main components are polyphenols and flavonoids. Propolis has a distinctive scent and is dark yellow in color. Propolis is not soluble in water, so it is mainly extracted with ethanol and used.

Specifically, due to the poor solubility of propolis, the formulation of cosmetics becomes unstable, active ingredients are not effectively extracted, resulting in reduced functionality and productivity, and it is difficult to continuously maintain and control the activity of the extract. Patent Document 1 (Public Patent Publication 10-2016-0148174) discloses a cosmetic composition in which the content of the active ingredient contained in the propolis extract is increased, but there is no disclosure of nanoparticles in which the propolis extract is encapsulated.

Republic of Korea Patent Publication 10-2016-0148174

An object of the present invention is to provide nanoparticles encapsulated with propolis extract having excellent long-term stability, skin permeability, and antioxidant activity.

Another object of the present invention is to provide a method for preparing the nanoparticles.

Another object of the present invention is to provide a cosmetic composition for antioxidant containing the nanoparticles as an active ingredient.

Another object of the present invention is to provide a cosmetic composition for improving wrinkles containing the nanoparticles as an active ingredient.

Another object of the present invention is to provide a cosmetic composition for improving whitening containing the nanoparticles as an active ingredient.

Another object of the present invention is to provide a health functional food composition for antioxidant containing the nanoparticles as an active ingredient.

To achieve the above purpose,

One aspect of the present invention is propolis extract 1 part by weight;

1 to 20 parts by weight of a solubilizing agent; and

[Polyethylene oxide (PEO)-polypropylene oxide (PPO)-polyethylene oxide (PEO)] 5 to 50 parts by weight of a triblock copolymer in the form; provides nanoparticles containing a propolis extract encapsulated.

Another aspect of the present invention includes the step (step 1) of stirring by adding a triblock copolymer in the form of a solubilizer and [polyethylene oxide (PEO)-polypropylene oxide (PPO)-polyethylene oxide (PEO)] to the propolis extract. It provides a method for producing the nanoparticles to do.

Another aspect of the present invention provides a cosmetic composition for antioxidant containing the nanoparticles as an active ingredient.

Another aspect of the present invention provides a cosmetic composition for improving wrinkles containing the nanoparticles as an active ingredient.

Another aspect of the present invention provides a cosmetic composition for improving whitening containing the nanoparticles as an active ingredient.

Another aspect of the present invention provides a health functional food composition for antioxidant containing the nanoparticles as an active ingredient.

The nanoparticles encapsulated with the propolis extract according to the present invention have high solubility in water by micellarizing propolis, a poorly soluble substance, as well as excellent long-term stability of the nanoparticles, high skin permeability due to their small size, and antioxidant Since it has excellent efficacy and can delay or inhibit skin aging, it can be usefully used as an antioxidant cosmetic composition.

1 is a graph showing the results of measuring the particle size after diluting the nanoparticles of Example 1 in water.
Figure 2 is a graph showing the results of measuring the particle size after diluting the nanoparticles of Comparative Example 1 in water.
3 is a photograph of a solution obtained by diluting the nanoparticles of Example 1 in water.
4 is a photograph of a solution obtained by diluting the nanoparticles of Comparative Example 1 in water.
5 is a photograph of a solution obtained by diluting the nanoparticles of Example 1 in water after 6 months at room temperature (25° C.).
6 is a photograph of a solution obtained by diluting the nanoparticles of Comparative Example 1 in water after 6 months at room temperature (25° C.).
Figure 7 is a graph showing the results of measuring the DPPH radical scavenging ability of the control group, Comparative Example 1 and Example 1.
8 is a graph showing the results of measuring the cell viability of Comparative Example 1 and Example 1.
9 is a graph showing the results of measuring the cell viability of TGF ß1 used as a positive control.
10 is a graph showing the results of measuring the amount of intracellular collagen (Procollagen Type 1 C-peptide, PIP) produced in Comparative Example 1 and Example 1.
11 is a graph showing the results of measuring the amount of intracellular collagen (Procollagen Type 1 C-peptide, PIP) produced by TGF ß1 used as a positive control.
Figure 12 is a graph showing the results of measuring the intracellular collagenase (Matrix matalloproteinase-1, MMP-1) activity inhibition ability of Comparative Example 1 and Example 1.
13 is a graph showing the results of measuring the intracellular collagenase (Matrix matalloproteinase-1, MMP-1) activity inhibitory ability of TGF ß1 used as a positive control.

Hereinafter, the present invention will be described in detail.

One aspect of the present invention is propolis extract 1 part by weight;

1 to 20 parts by weight of a solubilizing agent; and

[Polyethylene oxide (PEO)-polypropylene oxide (PPO)-polyethylene oxide (PEO)] 5 to 50 parts by weight of a triblock copolymer in the form; provides nanoparticles containing a propolis extract encapsulated.

In this case, the particle size of the nanoparticles may be 5 to 150 nm, 20 to 130 nm, 35 to 110 nm, 50 to 90 nm, or 55 to 75 nm.

In a specific embodiment of the present invention, nanoparticles containing propolis extract, polyethylene glycol 400 as a solubilizer and F127 (poloxamer 407) as a triblock copolymer were prepared, and the particle size of the nanoparticles was 71.3 nm. , It can be seen that the nanoparticles according to the present invention have high skin permeability and can be usefully used as a cosmetic composition for antioxidant (see Example 1 and Experimental Example 1).

In addition, the nanoparticles prepared in a specific embodiment of the present invention have little change in particle size and no precipitation even after 6 months, so the nanoparticles according to the present invention have excellent long-term stability and the antioxidant effect of propolis is sustained. Since it is maintained, it can be seen that it can be usefully used as a cosmetic composition for antioxidant (see Experimental Example 2).

The propolis extract may be prepared according to a known method or purchased and used on the market, and the propolis extract may be used in either liquid or solid form.

Typically, propolis may use any one selected from the group consisting of water and C ₁ to C ₄ lower alcohol or a mixed solution thereof as an extraction solvent, and specifically, the C ₁ to C ₄ lower alcohol is ethanol. may, but is not limited thereto.

In addition, the dissolving agent serves to dissolve the propolis extract.

Polyethylene glycol 200, polyethylene glycol 400, propylene glycol, tetraglycol, butylene glycol, etc. may be used as the glycol solubilizer, and polysorbate 20, polysorbate 80, polysorbate 60 as polysorbate-based solubilizers. , Polysorbate 40, polysorbate 65, etc. can be used, but these are only examples, and are not limited thereto, and any solubilizer capable of dissolving propolis extract can be used without limitation, and glycols commonly used by those skilled in the art A liquid solubilizer or a polysorbate-based solubilizer may be used.

Here, the dissolving agent may be used in an amount of 1 to 20 parts by weight, preferably 1 to 10 parts by weight, and most preferably 1 to 5 parts by weight, based on 1 part by weight of the propolis extract. If the dissolving agent is used in an amount of less than 1 part by weight based on 1 part by weight of the propolis extract, there is a problem in that the dispersing of the propolis extract is incomplete, making it difficult to release the propolis extract when used later, and 1 part by weight of the propolis extract When used in excess of 20 parts by weight, there is a problem of waste of reagents due to insufficient improvement in the stability of the propolis extract.

In addition, the [polyethylene oxide (PEO)-polypropylene oxide (PPO)-polyethylene oxide (PEO)] type triblock copolymer serves to form micelles and includes poloxamer 184, poloxamer 185, poloxamer 188, poloxamer 124, poloxamer 237, poloxamer 338, poloxamer 407, etc. can be used, but these are only examples, and are not limited thereto, and can be used without limitation as long as they can form micelles, and those skilled in the art can usually A triblock copolymer in the form of [polyethylene oxide (PEO)-polypropylene oxide (PPO)-polyethylene oxide (PEO)] may be used.

Here, the triblock copolymer is preferably used in an amount of 5 to 50 parts by weight, more preferably in an amount of 5 to 30 parts by weight, and most preferably in an amount of 5 to 10 parts by weight, based on 1 part by weight of the propolis extract. . If the triblock copolymer is used in less than 5 parts by weight with respect to 1 part by weight of the propolis extract, there is a problem of precipitation of the propolis extract, and in excess of 50 parts by weight with respect to 1 part by weight of the propolis extract When used, there is a problem in that the particle size increases and the stability decreases.

The nanoparticles can increase intracellular collagen production. At this time, the type of intracellular collagen is not particularly limited. For example, it may be procollagen Type 1 to procollagen Type 5, and through the examples, procollagen type It was confirmed that the production of 1 C-peptide (Procollagen Type 1 C-peptide) could be increased.

The nanoparticles can inhibit intracellular collagenase activity. At this time, the type of intracellular collagenase is not particularly limited, for example, MMP-1 (Matrix matalloproteinase-1), MMP-3 (Matrix matalloproteinase -3) or MMP-9 (Matrix matalloproteinase-9).

In addition, another aspect of the present invention is the step of stirring by adding a triblock copolymer in the form of a solubilizer and [polyethylene oxide (PEO)-polypropylene oxide (PPO)-polyethylene oxide (PEO)] to the propolis extract (step 1) It provides a method for producing the nanoparticles comprising a.

Hereinafter, the method for producing nanoparticles encapsulated with the propolis extract according to the present invention will be described step by step in detail.

In the method for producing nanoparticles encapsulated with propolis extract according to the present invention, step 1 is a solubilizer and [polyethylene oxide (PEO)-polypropylene oxide (PPO)-polyethylene oxide (PEO)] form of propolis extract This is a step of stirring by adding a triblock copolymer.

At this time, the stirring temperature may be 20 ℃ to 100 ℃, preferably 40 ℃ to 80 ℃. In addition, the stirring may be performed through ultrasonic treatment.

Furthermore, another aspect of the present invention provides a cosmetic composition for antioxidant containing the nanoparticles as an active ingredient.

And, another aspect of the present invention provides a cosmetic composition for improving wrinkles containing the nanoparticles as an active ingredient.

In addition, another aspect of the present invention provides a cosmetic composition for improving whitening containing the nanoparticles as an active ingredient.

In the cosmetic composition, the specific description of the nanoparticles is the same as the specific description of the nanoparticles in which the propolis extract is encapsulated.

Furthermore, the nanoparticles according to the present invention have been shown to have better DPPH radical scavenging ability than the antioxidant ascorbic acid, and the nanoparticles according to the present invention have excellent antioxidant efficacy and can be usefully used as a cosmetic composition for antioxidant It can be seen (see Experimental Example 3).

In addition, after active oxygen is created in the body, if the active oxygen is not effectively removed by various protective devices in the body, a series of inflammatory reactions occur, resulting in skin damage, which can lead to the formation of skin wrinkles and promotes the melanin production reaction. It can be. Therefore, since the nanoparticles according to the present invention have excellent antioxidant efficacy, can effectively remove active oxygen in the body, and inhibit skin wrinkle formation and melanin production reaction caused by the skin damage, a cosmetic composition for improving wrinkles or a cosmetic for improving whitening It can be usefully used as a composition.

In preparing the cosmetic composition, specifically, 1 to 10% by weight of propolis extract, preferably 1 to 5% by weight, based on the total weight of the cosmetic composition; 1 to 20% by weight of a solubilizing agent, preferably 2 to 15% by weight; [Polyethylene oxide (PEO)-polypropylene oxide (PPO)-polyethylene oxide (PEO)] type triblock copolymer 5 to 50% by weight, preferably 5 to 30% by weight; may be contained.

In the cosmetic composition of the present invention, in addition to the nanoparticles of the present invention, a fatty substance, an organic solvent, a solubilizing agent, a thickening agent and a gelling agent, a softening agent, a suspending agent, and an optical brightening agent. Stabilizers, pH regulators, antioxidants, foaming agents, fragrances, surfactants, water, ionic or non-ionic emulsifiers, fillers, sequestering and chelating agents, preservatives, vitamins, blocking agents, wetting agents, essential It may contain adjuvants commonly used in the field of dermatology, such as oils, dyes, pigments, hydrophilic or lipophilic actives, lipid vesicles or any other ingredient commonly used in compositions for improving skin beauty.

In addition, the components may be introduced in an amount generally used in the field of skin science.

Furthermore, the cosmetic composition according to the present invention is a solution, external ointment, cream, foam, nutrient lotion, softening lotion, pack, softening water, emulsion, makeup base, essence, soap, liquid cleanser, bath additive, sunscreen cream, sun oil, Suspensions, emulsions, pastes, gels, lotions, powders, soaps, surfactant-containing cleansers, oils, powder foundations, emulsion foundations, wax foundations, patches and sprays may be prepared in formulations selected from the group consisting of It is not limited.

In addition, the cosmetic composition of the present invention may further include one or more cosmetically acceptable carriers formulated in general skin cosmetics, and as typical ingredients, for example, oil, water, surfactants, moisturizers, lower alcohols, A thickener, a chelating agent, a colorant, a preservative, a flavoring agent, and the like may be suitably blended, but are not limited thereto.

Cosmetically acceptable carriers included in the cosmetic composition of the present invention vary depending on the formulation. When the formulation of the present invention is an ointment, paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tracanth, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide or Mixtures of these may be used.

Another aspect of the present invention provides a health functional food composition for antioxidant containing the nanoparticles as an active ingredient.

In the health functional food composition, the specific description of the nanoparticles is the same as the specific description of the nanoparticles in which the propolis extract is encapsulated.

Furthermore, the nanoparticles according to the present invention have been shown to have better DPPH radical scavenging ability than the antioxidant ascorbic acid, and the nanoparticles according to the present invention have excellent antioxidant efficacy and can be usefully used as health functional food compositions for antioxidants. It can be seen that (see Experimental Example 3).

In addition, the nanoparticles according to the present invention can be added to health functional food such as food and beverage as a health functional food composition for antioxidant.

Specifically, the nanoparticles according to the present invention can be added to food as it is or used together with other foods or food ingredients, and can be appropriately used according to conventional methods. The mixing amount of the active ingredient may be appropriately determined depending on the purpose of use (for prevention or improvement). Generally, the amount of the nanoparticles in the health food may be added to 0.1 to 90 parts by weight of the total food weight. However, in the case of long-term intake for the purpose of health and hygiene or health control, the amount may be less than the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount above the above range.

In addition, the health functional beverage composition of the present invention is not particularly limited in other components except for containing the nanoparticles as essential components in the indicated ratio, and may contain various flavoring agents or natural carbohydrates as additional components like conventional beverages. can Examples of the aforementioned natural carbohydrates include monosaccharides such as glucose, fructose, and the like; disaccharides such as maltose, sucrose and the like; and polysaccharides such as conventional sugars such as dextrins, cyclodextrins, and the like, and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those mentioned above, natural flavoring agents (thaumatin, stevia extract (eg rebaudioside A, glycyrrhizin, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can advantageously be used. The proportion of the natural carbohydrate is generally about 1 to 20 g, preferably about 5 to 12 g per 100 g of the composition of the present invention.

Furthermore, in addition to the above, the nanoparticles according to the present invention are various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, colorants and enhancers (cheese, chocolate, etc.), pectic acid and salts thereof , alginic acid and its salts, organic acids, protective colloidal thickeners, pH regulators, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, and the like. In addition, the nanoparticles of the present invention may contain fruit flesh for the production of natural fruit juice, fruit juice beverages, and vegetable beverages.

Hereinafter, the present invention will be described in detail through Examples and Experimental Examples.

However, the Examples and Experimental Examples to be described below are only to specifically illustrate the present invention in one aspect, but the present invention is not limited thereto.

<Preparation Example 1> Preparation of propolis extract

In order to prepare the nanoparticles encapsulated with the propolis extract, the propolis extract was prepared according to the following method.

After adding 10 L of ethanol to 500 g of bong-gyo and sufficiently stirring to settle the solid, the supernatant is collected and filtered through a filter paper (Watman 185 mmΦ × 100). After putting 300 mL of the filtered ethanol extract into a 1000 mL round flask and evaporating all the ethanol with a rotary evaporator, 300 mL of ethanol was added to the remaining extract to dissolve it. The dissolved ethanol extract was filtered through a filter paper (Watman 185 mmΦ × 100). All of the filtered ethanol extract was placed in a 1000 mL round flask and all of the ethanol was evaporated using a rotary evaporator to prepare a propolis extract.

< Example 1> Preparation of nanoparticles containing propolis extract, solubilizer, and triblock copolymer

Using the propolis extract prepared in Preparation Example 1, polyethylene glycol 400 (PEG 400, polyethyleneglycol 400) as a solubilizer, and F127 (poloxamer 407, Poloxamer 407) as a triblock copolymer, nanoparticles were prepared. manufactured.

Specifically, after adding and mixing 3 g of polyethylene glycol 400 (PEG400) to 1 g of the propolis extract of Preparation Example 1, 6 g of triblock copolymer F127 (poloxamer 407) was added and melted at a temperature of 60 ° C. then dissolved in water.

<Comparative Example 1> Preparation of nanoparticles containing propolis extract and solubilizing agent

Nanoparticles were prepared using the propolis extract of Preparation Example 1 and polyethylene glycol 400 as a dissolving agent.

Specifically, 3 g of polyethylene glycol 400 (PEG400) was added to 1 g of the propolis extract of Preparation Example 1, mixed at a temperature of 60 ° C, and then dissolved in water.

<Experimental Example 1> Evaluation of particle size of nanoparticles

Immediately after preparing the nanoparticles of Example 1 and Comparative Example 1 according to the present invention, they were diluted in water, and the particle size of the nanoparticles was evaluated using an ELS (ELS-Z, Otsuka, Japan) device, and the results are shown below. It is shown in Table 1, FIGS. 1 and 2, and the results of visually observing the solution in which the nanoparticles were diluted in water are shown in FIGS. 3 and 4.

Example 1 Comparative Example 1 Particle size (nm) 71.3 331.1

As shown in Table 1 above,

The nanoparticles prepared in Example 1 containing the triblock copolymer showed a small particle size of 71.3 nm, but the size of the nanoparticles prepared in Comparative Example 1 not containing the triblock copolymer was 331.1 nm. was (see FIGS. 1 and 2).

In addition, when the nanoparticles of Example 1 were diluted in water, a pale yellow transparent solution was observed, but when the nanoparticles of Comparative Example 1 were diluted in water, a white opaque solution was observed (see FIGS. 3 and 4).

The above results show that when nanoparticles are prepared including the triblock copolymer, the propolis extract is well soluble in water even though the particle size is small, compared to when nanoparticles are prepared without the triblock copolymer. shows high skin permeability and improved efficiency in expressing the antioxidant effect of propolis.

From this, it can be seen that the nanoparticles containing the propolis extract, solubilizer, and triblock copolymer according to the present invention can be usefully used as a cosmetic composition for antioxidant.

<Experimental Example 2> Measurement of particle size of nanoparticles for long-term stability evaluation

In order to evaluate the long-term stability of the nanoparticles prepared in Example 1 and Comparative Example 1 according to the present invention, the nanoparticles diluted in water were placed at room temperature (25 ° C) and 40 ° C, respectively, and the particle size according to time was measured by ELS ( ELS-Z, Otsuka, Japan) was measured using an instrument, and the results are shown in Table 2 below. In addition, after 6 months at room temperature (25 ° C.), the results of visually observing the solution in which the nanoparticles were diluted in water are shown in FIGS. 5 and 6 .

Particle size unit (nm) Room temperature (25℃) 40℃ Example 1 0 months 71.3 71.3 6 months 72.5 60.2 Comparative Example 1 0 months 331.1 331.1 6 months 1408.2 3321.1

As shown in Table 2 above,

The nanoparticles prepared in Example 1 containing the triblock copolymer showed little change in particle size even after 6 months. However, the particle size of Comparative Example 1, which did not contain the triblock copolymer, increased after 6 months. Specifically, at room temperature (25°C), the particle size increased from 331.1 nm to 1408.2 nm, about four times, and at 40°C, from 331.1 nm to 1408.2 nm. It increased to 3321.1 nm, about 10 times.

In addition, at room temperature (25 ° C), the solution in which the nanoparticles of Example 1 were diluted in water did not show a significant change after 6 months, but the solution in which the nanoparticles of Comparative Example 1 were diluted in water was 6 months old. After that, the color became darker, and a precipitate was formed, and the nanoparticles settled to the bottom or remained on the wall of the container showed changes (see FIGS. 5 and 6).

When evaluating long-term stability, if the solution in which the nanoparticles are diluted in water is transparent, has no precipitate, and does not change color, the stability of the nanoparticles is excellent. there is.

Therefore, the above results indicate that the long-term stability of the nanoparticles is better when the nanoparticles are prepared by including the triblock copolymer than when the nanoparticles are prepared without the triblock copolymer, and also the antioxidant of propolis. indicates that the efficacy is more sustained. From this, it can be seen that the nanoparticles containing the propolis extract, solubilizer, and triblock copolymer according to the present invention can be usefully used as a cosmetic composition for antioxidant.

<Experimental Example 3> Measurement of DPPH radical scavenging activity for evaluation of antioxidant activity

In order to evaluate the antioxidant activity of the nanoparticles prepared in Example 1 and Comparative Example 1 according to the present invention, the DPPH radical scavenging activity was measured in the following manner, and the results are shown in FIG. 7 .

Measurement of DPPH radical scavenging ability is well known as an experiment to measure the antioxidant efficacy of a target substance. The deep purple DPPH (Diphenylpicrylhydrazyl, diphenylpicrylhydrazyl) radical is converted to diphenylpicrylhydrazine by antioxidants. Since it uses the fact that it is reduced and turns to pale yellow, the closer it is to pale yellow, the stronger the antioxidant power of the antioxidant.

A 0.2 mM DPPH solution was prepared and prepared in 150 μl each in a 96 well plate, and a 1% solution of ascorbic acid as an antioxidant was prepared as a control. 50 μl each of the control group, Example 1, and Comparative Example 1 were treated in a 96-well plate prepared with DPPH solution, and the 96-well plate was placed in an incubator at 25° C. for 30 minutes, followed by a microplate reader (microplate reader). ) was used to measure the DPPH radical scavenging ability at an absorbance of 492 nm.

The DPPH radical scavenging ability of the 1% solution of ascorbic acid as a control was 87.5%, the DPPH radical scavenging ability of Comparative Example 1 without the triblock copolymer was 70.6%, and the DPPH radical scavenging ability of Example 1 containing the triblock copolymer was 70.6%. It was 88.2% (see FIG. 7).

The above results show that compared to the antioxidant ascorbic acid, the DPPH radical scavenging ability of Comparative Example 1 is low, and the DPPH radical scavenging ability of Example 1 is excellent. It indicates that the antioxidant efficacy of the nanoparticles is better.

From this, it can be seen that the nanoparticles containing the propolis extract, solubilizer, and triblock copolymer according to the present invention can be usefully used as a cosmetic composition for antioxidant.

< Experimental example 4> Assessment of cell viability

In order to evaluate the cell viability of the nanoparticles according to Example 1, the experiment was performed as follows. Specifically, human dermal fibroblasts (HDFs) were selected and used based on 「Guidelines for Efficacy Evaluation of Functional Cosmetics」, and HDFs ATCC are primary cells isolated from adult skin. Those in which Mycoplasma, Hepatitis B, Hepatitis C, HIV 1, bacteria, yeast, and other fungi were not detected were used. HDFs were inoculated on the bottom of a culture dish and cultured in a 3:1 DMEM/F12 mixed medium containing 1% Antibiotic Antimycotic and 10% FBS in a 37 °C, 5% CO ₂ incubator (HERA cell 150i, Thermo). did

Cell viability was analyzed using the principle of MTT. First, HDFs were inoculated in a 24-well plate at a concentration of 5Х10 ⁴ cells/well, cultured for 24 hours, and then transferred to FBS-free medium containing diluted test substances for each concentration. After exchange, it was cultured for 48 hours. After culturing for 48 hours, the presence or absence of cell shape change by treatment with each concentration was observed under a microscope, and then MTT solution was added to each well so that the final treatment concentration was 0.05% and cultured for 4 hours. After removing the culture medium, add 1,000 μL of DMSO and shake for 10 minutes, then use a 540 nm Microplate reader; Absorbance was measured at Epoch 2, BioTek, USA. The results are shown in Figures 8 and 9. 8 shows a cell viability graph of Comparative Example 1 and Example 1, and FIG. 9 shows a cell viability graph of TGF ß1 as a positive control.

8 and 9, Comparative Example 1 and Example 1 confirmed that there was no change in cell shape when compared to the solvent control group at concentrations of 0.31% or less and TGF ß1 at all concentrations, and no cytotoxicity was confirmed. .

< Experimental example 5> Analysis of intracellular collagen (PIP) production

In order to analyze the intracellular collagen (Procollagen Type 1 C-peptide, PIP) production of the nanoparticles according to Example 1, the following experiment was performed. Specifically, human dermal fibroblasts (HDFs) were inoculated in a 24-well plate at 5Х10 ⁴ cells/well, cultured for 24 hours, and then exchanged with FBS-free medium containing diluted test substances at each concentration. and then incubated for 24 hours. After 24 hours, each cultured supernatant was centrifuged at 10,000 rpm for 10 minutes, and the amount of procollagen in the supernatant from which debris was removed was measured using the Procollagen Type IC Peptide (PIP) ELIS A Kit. Absorbance was measured at 450 nm. The degree of collagen production was evaluated by correcting with the total protein amount. The results are shown in Figures 10 and 11. Figure 10 shows a graph of intracellular collagen (PIP) production of Comparative Example 1 and Example 1, and Figure 11 shows a graph of intracellular collagen (PIP) production of TGF ß1 as a positive control.

As can be seen through Figures 10 and 11, Comparative Example 1 is 8.36% at a concentration of 0.008% compared to the solvent control group, Example 1 is 6.40% to 33.06% at a concentration of 0.002% to 0.031%, TGF ß1 is 0.000001% to 0.00001% At concentrations of 25.31% to 34.39%, the amount of intracellular collagen production was significantly increased.

< Experimental Example 6> intracellular collagenase ( MMPs -1) Active inhibition analyze

In order to analyze the intracellular collagenase (Matrix matalloproteinase-1, MMP-1) production of the nanoparticles according to Example 1, the experiment was performed as follows. Specifically, human dermal fibroblasts (Human Dermal Fibroblasts, HDFs) were inoculated in a 24-well plate at 5Х10 ⁴ cells/well and cultured for 24 hours. After culturing, it was cultured for 48 hours after exchanging with FBS-free medium containing diluted test materials for each concentration. After 48 hours of culture, the supernatant was centrifuged at 10,000 rpm for 10 minutes, and the absorbance was measured at 450 nm using the MMP-1 Human ELISA Kit with the supernatant from which debris was removed. MMP-1 activity was evaluated by correcting for the total protein amount. The results are shown in Figures 12 and 13. Figure 12 shows a graph of intracellular collagenase (MMP-1) activity inhibition ability of Comparative Example 1 and Example 1, and Figure 13 is a graph of intracellular collagenase (MMP-1) activity inhibition activity of TGF ß1 as a positive control am.

As can be seen through Figures 12 and 13, Comparative Example 1 did not significantly reduce intracellular collagenase activity compared to the solvent control group, and Example 1 was 5.67% to 13.45% at a concentration of 0.008% to 0.031% compared to the solvent control group. , TGF ß1 significantly reduced intracellular collagenase activity from 25.56% to 38.06% at concentrations of 0.000001% to 0.00001%.

Claims (14)

1 part by weight of propolis extract;
1 to 20 parts by weight of polyethylene glycol 400 based on 1 part by weight of propolis extract; and
5 to 50 parts by weight of a triblock copolymer in the form of [polyethylene oxide (PEO)-polypropylene oxide (PPO)-polyethylene oxide (PEO)] based on 1 part by weight of propolis extract; nano-encapsulated propolis extract consisting of Nanoparticles, characterized in that they increase the amount of intracellular collagen production as particles.
According to claim 1,
Characterized in that the propolis extract is extracted using ethanol as a solvent, nanoparticles.
delete delete According to claim 1,
The [polyethylene oxide (PEO)-polypropylene oxide (PPO)-polyethylene oxide (PEO)] type triblock copolymer is poloxamer 184, poloxamer 185, poloxamer 188, poloxamer 124, poloxamer 237, poloxamer Characterized in that at least one selected from the group consisting of 338 and poloxamer 407, nanoparticles.
delete According to claim 1,
The intracellular collagen is characterized in that the procollagen Type 1 C-peptide (Procollagen Type 1 C-peptide), nanoparticles.
According to claim 1,
Characterized in that, the nanoparticle inhibits intracellular collagenase activity.
According to claim 8,
Characterized in that the intracellular collagenase is at least one selected from the group consisting of MMP-1 (Matrix matalloproteinase-1), MMP-3 (Matrix matalloproteinase-3) and MMP-9 (Matrix matalloproteinase-9) , nanoparticles.
The nanoparticles of claim 1 comprising adding a solubilizer and a triblock copolymer in the form of [polyethylene oxide (PEO)-polypropylene oxide (PPO)-polyethylene oxide (PEO)] to the propolis extract and stirring (step 1) Manufacturing method of.
delete A cosmetic composition for improving wrinkles, characterized in that it contains the nanoparticles of claim 1 as an active ingredient.


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