KR101337213B1 - Gynostemma pentaphyllum extract with anti-aging activity and process for preparing the same using supercritical carbon dioxide extraction - Google Patents

Abstract

The present invention relates to a method for producing an extradol extract using supercritical carbon dioxide and a cosmetic composition for preventing skin aging comprising an extradol extract extracted by a supercritical extraction method. Supercritical extrapolation method of the present invention is characterized by obtaining a high extrapolated extract of carotenoids as an active ingredient by adding supercritical carbon dioxide under the conditions of 40 to 90 ℃ and 300 to 700bar. Dodol supercritical extract of the present invention is effective in improving wrinkles due to skin aging by promoting collagen production and integrin expression of human skin cells.

Dol et al, supercritical, collagen, integrin

Description

Gynostemma pentaphyllum extract with anti-aging activity and process for preparing the same using supercritical carbon dioxide extraction}

The present invention relates to a method for producing an extradol extract using supercritical carbon dioxide and a cosmetic composition for preventing skin aging comprising an extradol extract extracted by a supercritical extraction method.

More specifically, the supercritical extrapolation method of the present invention is characterized by obtaining a high extrapolation extract of carotenoids as an active ingredient by adding supercritical carbon dioxide under conditions of 40 to 90 ° C. and 300 to 700 bar. Dodol supercritical extract of the present invention is effective in improving wrinkles due to skin aging by promoting collagen production and integrin expression of human skin cells.

Gynostemma pentaphyllum ) is a dicotyledonous plant of the genus Camellia japonica , a perennial vine plant. In Korea, it grows mainly in the mountains and wild forests of Jeju and Ulleungdo. Known as a medicinal plant with tonic gangjeong, diuretic, anti-inflammatory and dry stomach effects, after the spring of 1979, the Japanese Pharmacy Society announced that it contained more saponin than ginseng. Subsequently, many studies have been conducted to claim that extraversion is effective in anticancer, hypertension, and hepatocyte regeneration.

All of the research data published to date are extracted and utilized using organic solvents such as alcohol or water. However, these methods have disadvantages that are very undesirable in terms of their active utilization. That is, the extra rock extract extracted with an organic solvent or water does not have a high content of antioxidants. In this case, a separate purification process is required to increase the content of antioxidants. Recently, it has been found that many carotenoid components exist in addition to stones, and these components have strong antioxidant activity and are known to be effective in preventing aging. However, these carotenoid components were not extracted when extracted with an organic solvent or water, or contained a problem that the antioxidant content of the extract is not high.

Therefore, the necessity of developing an effective method for effectively extracting antioxidant useful substances such as carotenoids from the stone has emerged continuously, and at the same time, the demand for applying to cosmetics by confirming the anti-aging effect of the extracts containing these ingredients Was constantly.

Thus, the inventors of the present invention sought to develop an effective extraction method with high antioxidant content from stones, using supercritical carbon dioxide, and as a result, supercritical extracts with a high content of carotenoids were obtained. Through the study of the efficacy effect of confirming that there is an excellent effect on the anti-aging of the skin, to complete the present invention.

Therefore, the first object of the present invention is to provide an extract containing a lot of antioxidant components by increasing the content of antioxidant components such as carotenoids of the extra-extract by extracting using supercritical carbon dioxide.

A second object of the present invention is to provide a cosmetic composition for preventing skin aging comprising the extra rock extract obtained by the above method as an active ingredient.

The extradol extraction method using supercritical carbon dioxide of the present invention is characterized in that the extradol extract is obtained by adding supercritical carbon dioxide under conditions of 40 to 90 ° C. and 300 to 700 bar.

In order to extract the high-oxidation extrapolated extract using supercritical carbon dioxide, as described in European Patent No. 925,724, an extraction tank, a heat exchanger, a pump, a carbon dioxide storage tank, a chiller (cooling condenser), a pressure reducing separator, and a pressure reducing valve Use an extractor consisting of In the extraction device, the extraction tank serves to extract the raw material by the extra stone, the pump serves to apply pressure to the supercritical carbon dioxide, the heat exchanger serves to heat the supercritical carbon dioxide, pressure reducing valve Decompresses the supercritical carbon dioxide emitted from the extractor, and the decompressor completely decompresses the supercritical carbon dioxide to separate gaseous carbon dioxide, and the chiller converts the decompressed carbon dioxide into supercritical carbon dioxide. In addition, the carbon dioxide reservoir serves to store supercritical carbon dioxide.

In order to extract a specific component from the stone using the extracting device, first, the raw material is added to the extraction tank, the supercritical carbon dioxide passed through the pump and the heat exchanger is introduced to the bottom of the extraction tank, and then the mixture of the component extracted from the raw material and carbon dioxide After discharging to the top of the extraction tank, the discharged mixture is decompressed through a pressure reducing valve, the carbon dioxide is separated and collected in a gas phase in a pressure reduction separator, and the extracted components are collected. At this time, the separated carbon dioxide is converted to the liquid carbon dioxide through the chiller, stored in the carbon dioxide storage tank, and is then supplied to the extractor using a pump and a heat exchanger to extract a specific component in a circulating manner.

A carotenoid compound is included as a representative effective ingredient of the supercritical fluid extracellular extract of the present invention, which is effective in preventing skin aging. Carotenoid compounds included in the extract obtained from the stone by the extraction method using the supercritical carbon dioxide include lutein, neochrome, neoxanthin, alpha and beta carotene, o Auroxanthin, violaxanthin, and luteoxanthin. The carotenoid compound of the dodol supercritical fluid extract prevents skin aging by promoting the synthesis of proteins necessary for the structure formation and function of skin cells, and has the effect of enhancing the expression of collagen and integrin.

Wrinkle formation due to skin aging may occur when the production and conversion of collagen protein, which forms an extra-cellular matrix structure inside the dermal layer, is not performed properly. Another protein that affects wrinkles and reduced skin elasticity is integrin. Integrin is a membrane protein present in skin cells, which promotes adhesion between skin cells and interactions between cells and proteins, and greatly affects skin elasticity. The level of integrin expression is closely related to the elasticity of the skin. The present inventors confirmed that the extradolescent supercritical extract containing carotenoids as an active ingredient promotes collagen production of human skin cells, increases the expression of integrin, and is effective in improving wrinkles by skin aging.

Dodol supercritical extract of the present invention can be used as a raw material for cosmetics, food, etc., and provides a cosmetic composition for the purpose of preventing skin aging.

Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention in more detail, it will be apparent to those skilled in the art that the scope of the present invention is not limited by these examples in accordance with the gist of the present invention. .

Example 1 Preparation of Extra-Extracts and Identification of Carotenoids Components

The carotenoids in the extracellular extract of the present invention were isolated and identified by the following method.

10 g of the dried stone was placed in an extraction tank, and the flow rate of supercritical carbon dioxide was 10 ml / min, the temperature of the extraction tank was 60 ° C., and the pressure of the supercritical carbon dioxide was maintained at 400 bar. The carotenoid component of the extract was analyzed using HPLC. Lutein and carotene standards were purchased from Sigma, and NMR, neochrome, neooxanthin, auroxanthin and violaxanthin were isolated after pure separation. The structure was identified and identified as a standard product. As an analytical column, a C30 reverse phase column was used, and the mobile phase was subjected to a gradient composition. The methanol-isopropanol mixture (99: 1, v / v) and methylene chloride were 50 to 100: 0 to 70:30 (v / v). Changed over minutes. As a detector, a photodiode-array UV detector was used for component verification.

As a result of analyzing the carotenoid composition of the dodol extract of the present invention by the HPLC analysis, lutein (62%), neochrome (12%), neoxanthin (8%), alpha carotene (carotene) 4%, beta carotene 1%, auroxanthin (7%) and violaxanthin (violaxanthin) 6%.

Example 2 Supercritical Carbon Dioxide Condition for Enhancing Carotenoid Content

In order to determine the conditions of supercritical carbon dioxide to increase the carotenoid content of the extra rock extract, the conditions were changed as follows.

The temperature of the extraction tank is changed in the range of 40-90 ℃, and extracted for 1 hour by changing the pressure of supercritical carbon dioxide to 200, 300, 400, 500, 600 or 700 bar, respectively, and then extracted by measuring the carotenoid content in the extra-extraction extract The efficiency was determined. Tables 1 and 2 show changes in carotenoid content of extracts according to supercritical extraction conditions. The carotenoid content is expressed as the total carotenoid weight extracted per unit weight of the extradol sample, as determined by the HPLC analysis of Example 1. At this time, the carotenoid content in the extract using the organic solvent methanol was applied as a control to compare the extraction efficiency. As a control, an extract obtained by stirring and extracting a dry dodol sample with methanol three times at room temperature for 24 hours, and then drying the reduced liquid at 40 ° C. under filtration was used.

 Changes in the carotenoid content (mg / g dry sample) of extra-extracted extracts according to extraction pressure (extraction temperature: fixed at 60 ℃) Content / Extraction Condition Supercritical CO2 Pressure Conditions (bar) Control group 200 300 400 500 600 700 carotenoids
(mg / g sample) 0.6 1.0 1.2 1.4 1.6 1.7 0.4

  Changes in Carotenoid Contents (mg / g Dry Sample) of Extra-Dolty Extracts According to Extraction Temperature Content / Extraction Condition Supercritical Carbon Dioxide Temperature Condition (℃) Control group 40 50 60 70 80 90 carotenoids
(mg / g sample) 1.1 1.2 1.2 1.3 1.4 1.2 0.4

As shown in Table 1, the total carotenoid content extracted when the pressure is more than 300bar showed a sharp increase, and when the pressure is more than 500bar it can be seen that the increase in the extracted content is gentle. Therefore, it was found that the supercritical carbon dioxide pressure is more than 300 bar for carotenoid extraction. As shown in Table 2, the effect of temperature did not significantly affect the extraction efficiency above 40 ℃. Rather, the extraction efficiency decreases above 90 ° C., which indicates that the stability of various components may have decreased with increasing temperature.

Experimental Example 1 : Wrinkle improvement effect of the extra stone extract

Wrinkle formation due to skin aging may occur when the production and conversion of collagen protein, which forms an extra-cellular matrix structure inside the dermal layer, is not performed properly. Therefore, in order to measure the anti-wrinkle effect of the doldol supercritical extract according to the skin aging, the supercritical extract was administered to human skin cells to measure the collagen production enhancement effect. Human skin fibroblasts (CCD986-SK, Korea Cell Line Bank) were inoculated to ⁵ × 10 ⁵ cells in microwell plates and incubated for 36 hours in DMEM medium in a 37 ° C. CO ₂ incubator. Subsequently, to replace DMEM medium, DMEM medium containing 0, 10, 20, 50, 100, and 200 ppm concentrations of the extradolone carotenoids was applied. After incubation for 24 hours after the medium exchange, the collagen concentration was analyzed using a PICP collagen assay kit by collecting only the solutions except the cells in each well. The collagen concentration measured the amount of c-peptide (PICP) of the collagen type 1 synthesized during the cultivation, shown in Table 3 in ㎍ / ml unit concentration.

  Enhancement of Collagen Formation of Dodol Extracritical Extracts Effect / condition Dodol carotenoid concentration (ppm) 0 10 20 50 100 200 Collagen Production
(Μg / ml medium) 0.18 0.45 0.62 0.60 0.61 0.23

As shown in Table 3, by increasing the amount of collagen production in human skin cells to which the extradolescent supercritical extract of the present invention was administered, it was possible to confirm the collagen production promoting effect. There was no enhancement effect at concentrations above 200 ppm, which is considered to be cytotoxicity.

Another protein that affects wrinkles and decreases skin elasticity is integrin. Integrin is a membrane protein present in skin cells, which promotes adhesion between skin cells and interactions between cells and proteins, and greatly affects skin elasticity. The degree of integrin expression is known to be closely related to the elasticity of the skin. Therefore, in order to measure the skin elasticity enhancing effect of the extradolary supercritical extract, the supercritical extract was administered to human skin cells to measure the integrin expression enhancing effect.

The experiment was performed in the same manner as the human skin fibroblast culture method, but only the cells cultured in each well were recovered by centrifugation and crushed, and then the concentration of integrin beta 1 present in the cell extract was measured. The concentration of integrin beta 1 in using an ELISA assay by ELISA, and measured using the integrin ELISA kit (TAKARA, Japan) integrin concentrations expressed OD ₅₃₀ absorbance values as the control group absorbance and relative proportions (ratio) value 4 is shown.

  Integrin Expression Enhancement Effect of Dodol Extracritical Extracts Effect / condition Dodol carotenoid concentration (ppm) 0 10 20 50 100 200 Absorbance ratio
(ratio) One 1.1 1.2 1.4 1.5 0.9

As shown in Table 4, integrin beta 1 expression enhancement effect was confirmed in the human skin cells to which the dodol supercritical extract of the present invention was administered, the cells did not show an enhancement effect at the concentration of 200 ppm or more as in the electrical collagen experiment It is considered to be cytotoxicity.

 The present invention provides a method for preparing an extract having a high content of carotenoids from a dodol, using supercritical carbon dioxide, and an extradol extract having a high carotenoid content prepared by the method. The high carotenoids extract according to the present invention has excellent anti-wrinkle effect due to skin aging and includes various useful ingredients, so that it can be widely used as a raw material for functional cosmetics or foods.

Claims (4)

The dried dodol is placed in an extraction tank and supercritical carbon dioxide, which has passed through a pump and a heat exchanger at a temperature of 50 to 80 ° C. and a carbon dioxide pressure of 300 to 600 bar, is introduced into the bottom of the extraction tank to extract the dodol, and then extracted from raw materials. Extracting dodol by applying supercritical carbon dioxide, which discharges a mixture of components and carbon dioxide to the top of the extraction tank: And separating the carbon dioxide from the mixture of the extradolous extract and the supercritical carbon dioxide in a gas phase and collecting the extrapolated extract having an carotenoid compound, which is an antioxidant component, in an amount of 0.1 wt% or more. delete delete A cosmetic composition for improving skin wrinkles containing as an active ingredient an extract of an carotenoid which is an antioxidant component according to claim 1, wherein the carotenoid compound is 0.1% by weight or more as an active ingredient.