KR20230139637A - Cosmetics composition with cudrania tricuspidata extract and manufacturing method thereof - Google Patents

Abstract

The present invention relates to a cosmetic for preventing or improving white hair containing a Cudrania extract. The Cudrania extract according to an embodiment of the present invention has no cytotoxicity and at the same time prevents gray hair due to the effect of increasing melanin production and melanin-producing enzyme activity. It can be prevented or improved.

Description

Cosmetic composition containing cudrania tricuspidata extract and manufacturing method thereof}

The present invention relates to a cosmetic composition for preventing or improving white hair containing an extract of Kujippong and a method for producing the same.

The pigment melanin determines skin color and plays a role in protecting the skin by absorbing ultraviolet rays. Melanocytes, which produce melanin pigment, are derived from melanoblasts. When melanoblasts are stimulated and differentiated into melanocytes, tyrosinase is activated and converts tyrosine contained in the cells into DOPA (DOPA). 3,4-dihydroxy-L-phenyl-alanine), and then DOPA is converted into a substance called dopaquinone through the action of an enzyme called dopaoxidase, and then becomes melanin through a series of chemical reactions. . After making melanin, melanocytes move to the epidermal tissue of the upper layer of the skin and deliver melanin to keratinocytes present in the epidermal tissue. During the cell migration process, melanin pigment is stored and transported in melanosomes. At the beginning of production, there is no color, but as it gradually moves to the upper layer, it is filled with black pigment over 4-5 stages, becoming fully pigmented mature melanin granules. .

White hair is a graying/whitening of hair caused by a decrease in melanin produced by melanocytes in the hair such as hair, eyebrows, and eyelashes. According to what is known to date, the mechanism of occurrence of white hair is known to be due to a decrease in the number of melanocytes that form melanin pigment in hair follicles, and the inability to move melanin to surrounding keratinocytes, resulting in pigment deficiency.

Currently, attempts are being made around the world to manufacture cosmetics that are both natural and functional. Cosmetic materials extracted from natural products, especially plants, are attracting attention because the extraction cost is low and the raw materials are easy to secure.

Cudrania tricuspidate is a short-leafed tree belonging to the genus Cudrania of the Moraceae family. It grows on the foothills of mountains or fields in the south-central part of Hwanghae-do, Korea. Unlike ordinary mulberry trees, the Cudrania mulberry tree has long thorns, the edges of the leaves are smooth, and when the leaves are cut, a milky white liquid comes out, and the same liquid comes out from the fruit. Cudrania contains physiologically active substances such as planoids, GABA, rutin, and aspartic acid, and is known to have anti-cancer, analgesic, antibacterial, blood clot removal, diabetes, high blood pressure, cerebral hemorrhage, menstrual pain, alleviation of coldness, atopy, and anti-inflammatory properties. there is. However, there have been no reports on the prevention or improvement of white hair disease using cucurbita.

Accordingly, the present invention was completed by confirming that the Cudrania tricuspidata extract has no cytotoxicity and is effective in increasing melanin production and tyrosinase activity.

The problem to be solved by the present invention is to provide a cosmetic for preventing or improving gray hair containing an extract of Kujippong.

In order to achieve the above technical problem, in one aspect of the present invention, a cosmetic composition for preventing or improving white hair containing a Cudrania extract is provided.

In one embodiment, the Cudrania extract may include Maclurin of the structural formula 1 below.

[Structural Formula 1]

In one embodiment, the kkujippong extract may be an ethanol extract of 60 to 80% by volume.

In one embodiment, the kkujippong extract may be extracted by adding ethanol to kkujippong at room temperature for 10 to 24 hours.

In one embodiment, the kkujippong extract may be included in an amount of 0.05% by weight to 10% by weight in the composition.

In another aspect of the present invention, there is provided a method for producing a cosmetic composition for preventing or improving white hair, comprising the step of obtaining a Cudrania extract by adding ethanol to Cudrania, wherein the Cudrania extract includes Maclurin of the following structural formula 1: .

[Structural Formula 1]

In one embodiment, the step of obtaining the kkujippong extract may be performed at room temperature for 10 to 24 hours.

In one embodiment, the step of concentrating the extract of Kujippong and then freeze-drying it may be further included.

In one embodiment, the concentration may be rotationally concentrated under reduced pressure at 30 ℃ to 50 ℃ at 30 rpm to 75 rpm.

In one embodiment, the freeze-drying may be performed at a temperature of -110°C to -60°C for 65 to 80 hours.

The present invention relates to a cosmetic for preventing or improving white hair containing a Cudrania extract. The Cudrania extract according to an embodiment of the present invention has no cytotoxicity and at the same time prevents gray hair due to the effect of increasing melanin production and melanin-producing enzyme activity. It can be prevented or improved.

The effects of the present invention are not limited to the effects described above, and should be understood to include all effects that can be inferred from the configuration of the invention described in the description or claims of the present invention.

Figure 1 shows the extract securing process using the 70% ethanol extraction method.
Figure 2 shows the extract securing process using hot water extraction.
Figure 3 shows the results of an experiment measuring melanin content in B16F10 cells treated with 100㎍/mL Cudrania extract (hot water extraction and room temperature extraction).
Figure 4 shows the results of an experiment measuring melanin content in B16F10 cells treated with Cudrania root extract (hot water extraction and ethanol extraction) at a concentration of 200 μg/mL.
Figure 5 is a repeated measurement of melanin content in B16F10 cells treated with Cudrania root extract (hot water extraction and ethanol extraction) at a concentration of 100 μg/mL and in B16F10 cells treated with Cudrania root extract (hot water extraction and ethanol extraction) at a concentration of 200 μg/mL. This shows the results of the experiment.
Figure 6 shows the results of repeated experiments measuring melanin content in B16F10 cells treated with 200㎍/mL Cudrania extract (hot water extraction and ethanol extraction).
Figure 7 shows the results of a cytotoxicity test in B16F10 cells treated with Cudrania extract (hot water extraction and ethanol extraction).
Figure 8 shows a graph of Tyrosinase activity measurement of absorbance over time including standard deviation.
Figure 9 shows a graph of Tyrosinase activity measured by absorbance over time, excluding standard deviation.
Figure 10 shows a graph of Tyrosinase activity by group at the incubation time point of 30 minutes.
Figure 11 shows the expression of melanin producing enzyme according to cudra extract treatment.
Figure 12 shows kkujippong raw material manufacturing instructions and process records.

Hereinafter, the present invention will be described in detail.

According to one aspect of the present invention, a cosmetic composition for preventing or improving white hair, comprising an extract of Cudrania chinensis, is provided.

The Cudrania tricuspidata can be obtained from Cudrania tricuspidata, which is a deciduous tree or shrub belonging to the Moraceae family and is also called Gutthorn tree. Cudrania tree leaves are mainly used as a substitute for mulberry leaves, the fruit is mainly used to make jam or alcohol, and the bark and roots of the tree are used for medicinal purposes or as raw materials for paper. In the present invention, Cudrania may include without limitation all parts included in the Cudrania tree, such as leaves, fruits, bark, and roots, and may preferably be the fruit.

The Cudrania extract may contain Maclurin of the structural formula 1 below.

[Structural Formula 1]

The Cudrania extract contains Maclurin, increases melanin production, and exhibits melanin-producing enzyme activity, so it is effective in preventing or improving white hair.

The kkujippong extract uses the extract itself and the extract, such as an extract obtained by extraction treatment of the kkujippong, a diluted or concentrated solution of the extract, a dried product obtained by drying the extract, a crude product or purified product of the extract, or a mixture thereof. It can include extracts of all formulations that can be formed.

The extract may be extracted using known methods and conditions known in the art, and may be, for example, ethanol extract, hot water extract, cold soak extract, reflux extract, steam distillation extract, or ultrasonic extract. There is no particular limitation as long as the active ingredient in kkujippong can be included in an appropriate amount to exert an effect of improving white hair.

Specifically, it may be an ethanol extract, and the ethanol may be, for example, 60 to 80% by volume or 65 to 75% by volume of ethanol.

More specifically, the kkujippong extract may be, for example, extracted by adding ethanol to kkujippong at room temperature for 5 to 30 hours, preferably for 10 to 24 hours. The extraction time is not limited to this, but when extracted within the above range, melanin production by the Cudrania extract and melanin production enzyme activity can be maximized.

In the above-mentioned white hair disease, the number of melanocytes in the hair roots (hair bulbs) is significantly reduced, the activity of producing melanocytes is reduced, melanin in the hair matrix and fur cortex is lost, and the hair turns white, resulting in white hair (gray hair). It may include diseases that appear in the form of: The location of the disease is mainly hair, eyebrows, eyelashes, beards, etc. The cause of the disease may be a natural aging process as the body ages, but may also include genetic factors, hyperthermia, mental shock, drugs, diabetes, etc. Thyroid disease, etc. may be the cause. Because the phenomenon of graying hair due to aging progresses gradually, it can appear in various intermediate colors between black and white, and white hair in the present invention may also include these phenomena, hair discoloration, gray hair, etc.

The cosmetic composition can be prepared in any conventionally prepared formulation, for example, solution, emulsion, suspension, paste, cream, lotion, gel, powder, spray, surfactant-containing cleansing, oil, soap, liquid. It can be formulated as a cleanser, bath additive, foundation, makeup base, essence, lotion, foam, pack, softening water, sunscreen cream, or sun oil, but is not limited to this.

The cosmetic composition of the present invention may further include commonly used auxiliaries and carriers such as antioxidants, stabilizers, solubilizers, vitamins, pigments, fragrances, etc. For example, the cosmetic composition may further include auxiliary ingredients such as glycerin, butylene glycol, polyoxyethylene hydrogenated castor oil, tocopheryl acetate, citric acid, panthenol, squalane, sodium citrate, and allantoin.

According to another aspect of the present invention, a method for producing a cosmetic composition for preventing or improving white hair comprising the step of obtaining a Cudrania extract by adding ethanol to Cudrania, wherein the extract of Cudrania contains Maclurin of the following structural formula 1: provides.

[Structural Formula 1]

The Cudrania root, Cudrania extract, and ethanol extraction method included in the manufacturing method of the cosmetic composition for preventing or improving gray hair of the present invention are as described above.

The present invention may further include the step of concentrating and then freeze-drying the kkujippong extract, if necessary.

The concentration may be performed using known means using known methods and conditions. For example, it may be performed using a vacuum concentrator. Specifically, the reduced-pressure concentration may be performed using a reduced-pressure concentrator after passing the extract of Kujippong through filter paper to remove impurities, more specifically, at 30 ℃ to 50 ℃ or 35 ℃ to 45 ℃ at 30 rpm to 75 rpm Alternatively, it may be performed at 40 rpm to 50 rpm, but is not limited thereto.

The freeze-drying may be performed using known means and under known methods and conditions. For example, the freeze-drying may be performed at a temperature of -110°C to -60°C or -90°C to -70°C for 65 to 80 hours or 70 to 75 hours.

Hereinafter, the present invention will be described in more detail through examples and experimental examples. However, the following production examples and test examples are for illustrating the present invention, and the scope of the present invention is not limited thereto.

Manufacturing example. Preparation of Cudrania extract

Cudrania extract was prepared using a 70% ethanol extraction method (Example, Figure 1) and a hot water extraction method (Comparative Example 1, Figure 2).

Experimental Example 1. Confirmation of ability to increase melanin production

Mouse B16F10 melanoma (B16F10) was inoculated into a 6-well plate using DMEM medium containing 10% FBS (fetal bovine serum) at 1 × 10 ⁵ per well, and then incubated at 5% CO ₂ and 37°C. Cells were cultured until approximately 80% or more adhered to the bottom of the well plate.

After culturing, B16F10 cells were treated with the Kujippong extract of the above preparation example and then cultured for about 1 day under conditions of 5% CO ₂ and 37°C. The melanin content was measured using the melanin content test method, and for comparison, the melanin content of cells in the untreated group (control) was also measured. In the melanin content test, the cells were dissolved using 1N NaOH and the melanin content was confirmed by measuring the absorbance of melanin in an aqueous solution. Then, the relative number of cells was measured using the CCK-8 (Cell Counting Kit-8) assay method for normalization. The final melanin content was normalized by dividing by the number of cells in each treatment group. The CCK-8 is prepared using a water-soluble tetrazolium salt, WST-8 [2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium]. As a result, colorless WST-8 is reduced by dehydrogenase within cells and produces formazan (WST-8 formazan), which turns yellow. Because dehydrogenase is produced only in living cells, the degree of formazan produced is proportional to the number of living cells. As a result of measuring the normalized melanin content, it was confirmed that the melanin content significantly increased in the group treated with the extract concentration in B16F10.

The results of an experiment measuring melanin content in B16F10 cells treated with 100㎍/mL Cudrania extract (hot water extraction and room temperature extraction) are shown in Figure 3.

As a result, it was confirmed that the melanin production effect of Cudrania extract extracted at room temperature was higher than that obtained by hot water extraction.

The results of an experiment measuring the melanin content in B16F10 cells treated with Cudrania root extract (hot water extraction and ethanol extraction) at a concentration of 200 μg/mL are shown in Figure 4.

As a result, it was confirmed that Cudrania extract extracted from 70% ethanol had a higher melanin production effect.

In addition, repeated experiments measuring melanin content in B16F10 cells treated with Cudrania root extract (hot water extraction and ethanol extraction) at a concentration of 100 μg/mL and B16F10 cells treated with Cudrania root extract (hot water extraction and ethanol extraction) at a concentration of 200 μg/mL The results are shown in Figure 5.

As a result, it was confirmed that the melanin production effect of Cudrania extract at a concentration of 200 ㎍/mL extracted in 70% ethanol was the highest.

In addition, the results of repeated experiments measuring melanin content in B16F10 cells treated with Cudrania root extract (hot water extraction and ethanol extraction) at a concentration of 200 μg/mL are shown in Figure 6.

As a result, it was confirmed that the melanin production effect of Cudrania extract extracted in 70% ethanol for 12 hours was higher.

Normalization was performed by cell-viability through CCK-8 assay, and the results of the cytotoxicity test in B16F10 cells treated with Cudrania extract (hot water extraction and ethanol extraction) are shown in Figure 7.

As a result, no significant cytotoxicity was confirmed up to a concentration of 200㎍/mL.

Experimental Example 2. Confirmation of skin melanin production enzyme activity

Mouse B16F10 melanoma (B16F10) was inoculated into a 6-well plate using DMEM medium containing 10% FBS (fetal bovine serum) at 1 × 10 ⁵ per well, and then incubated at 5% CO ₂ and 37°C. Cells were cultured until approximately 80% or more adhered to the bottom of the well plate. After culturing, B16F10 cells were treated with the extract at a concentration of 200 ㎍/mL and then cultured for about 1 day under conditions of 5% CO ₂ and 37°C.

Next, the cells from which the medium was removed were washed with ice-cold PBS (phosphated buffer saline), and the cells were recovered using a scraper. The melanin producing enzyme activity was confirmed using the recovered cells. The recovered cells were dissolved in 200㎕ of tyrosinase activity lysis buffer (pH 6.8, 50mM sodium phosphate, 1% Triton X-100) and then lysed using an ultrasonicator. The centrifuge was spun at 15000 rpm for 15 minutes and the supernatant was used. Protein was normalized by measuring the protein absorbance value at 560 nm using the BCA protein assay. Melanin-producing enzyme activity was confirmed using Reaction buffer (pH 6.8, 50mM sodium phosphate, 2Mm L-DOPA) containing L-DOPA, a melanin substrate. The concentration of the protein was calculated, and 100 ㎍ of protein was inoculated per well of a 96-well plate, followed by 200 ㎕ of reaction buffer, and the degree of melanin production was estimated by measuring the absorbance value over time. As a result of measuring the normalized melanin-producing enzyme activity, it was confirmed that melanin-producing enzyme activity significantly increased in the group treated with the extract concentration in B16F10.

In the preceding Example 1, the effect of the Cudrania extract at a concentration of 200 μg/mL was confirmed, and experiments were conducted on the ethanol extract for 12h and 24h.

Tyrosinase activity and absorbance measurement results over time, including standard deviation, are shown in Figure 8.

Tyrosinase activity time-dependent absorbance measurement results without standard deviation are shown in Figure 9.

Tyrosinase activity measurement results for each group at the incubation time point of 30 minutes are shown in Figure 10.

As a result, it was confirmed that the 200㎍/mL Cudrania extract extracted in 70% ethanol for 12 hours had a higher effect on increasing Tyrosinase activity.

Experimental Example 3. Confirmation of skin melanin production enzyme (Tyrosinase) expression level by extract treatment

Mouse B16F10 melanoma (B16F10) was inoculated into a 6-well plate using DMEM medium containing 10% FBS (fetal bovine serum) at 1 × 10 ⁵ per well, and then incubated at 5% CO ₂ and 37°C. The cells were cultured until more than 80% of the cells adhered to the bottom of the well plate.

After culturing, B16F10 cells were treated with the extract at a concentration of 200 ㎍/mL and then cultured for about 1 day under conditions of 5% CO ₂ and 37°C. Next, the cells from which the medium was removed were washed with ice-cold PBS (phosphated buffer saline), and the cells were recovered using a scraper. Western blot analysis was performed using the recovered cells.

The recovered cells were lysed by incubation at 0°C for about 30 minutes using 1X RIPA (Radioimmunoprecipitation assay) cell lysis buffer, and then centrifuged to obtain the protein of the supernatant. The obtained proteins were separated by size using 8% sodium dodecyl sulfate polyacrylamide gel electrophoresis and then transferred to a polyvinylidene fluoride membrane activated with methanol. Afterwards, the membrane was blocked for 1 hour using BSA solution (TBS (tris-buffered saline) containing 3% Bovine Serum Albumin + Tween20), and incubated with primary antibody against melanogenesis-related protein at 4°C for 16 hours. During incubation. The membrane was washed with TBS (tris-buffered saline) containing Tween20, then incubated with secondary antibody at room temperature for 1 hour, and the protein of the finally obtained membrane was detected by enhanced chemiluminescence (ECL)-ECL substrate reaction.

As a result, it was found that the expression level of melanin producing enzyme was increased in B16F10 cells following treatment with Cudrania extract extracted from ethanol (Figure 11).

Claims (10)

A cosmetic composition for preventing or improving white hair, comprising an extract from Cudrania.
According to paragraph 1,
A cosmetic composition for preventing or improving white hair, wherein the Cudrania extract contains Maclurin of the structural formula 1 below.
[Structural Formula 1]

According to paragraph 1,
The kkujippong extract is a 60-80% by volume ethanol extract in a cosmetic composition for preventing or improving white hair.
According to paragraph 1,
The kkujippong extract is a cosmetic composition for preventing or improving white hair, which is obtained by adding ethanol to kkujippong and extracting it at room temperature for 10 to 24 hours.
According to paragraph 1,
A cosmetic composition for preventing or improving white hair, wherein the Cudrania extract is contained in an amount of 0.05% by weight to 10% by weight in the composition.
It includes adding ethanol to kkujippong to obtain a kkujippong extract,
A method for producing a cosmetic composition for preventing or improving white hair, wherein the Cudrania extract contains Maclurin of the structural formula 1 below.
[Structural Formula 1]

According to clause 6,
A method of producing a cosmetic composition for preventing or improving white hair, wherein the step of obtaining the Cudrania extract is performed at room temperature for 10 to 24 hours.
According to clause 6,
A method for producing a cosmetic composition for preventing or improving white hair, further comprising the step of concentrating the Cudrania extract and then freeze-drying it.
According to clause 8,
The method of producing a cosmetic composition for preventing or improving white hair, wherein the concentration is concentrated under reduced pressure by rotation at 30 ℃ to 50 ℃ at 30 rpm to 75 rpm.
According to clause 8,
A method of producing a cosmetic composition, wherein the freeze-drying is performed at a temperature of -110 ° C to -60 ° C for 65 to 80 hours.