KR20200109592A - The composition with antioxidant and anti-inflammatory activity containing Lycium chinense leaf extract of Chengyang - Google Patents

Abstract

A composition having antioxidant and anti-inflammatory effects and including a Cheongyang Lycium chinense leaf extract as an active ingredient according to the present invention is obtained by extracting Cheongyang Lycium chinense leaves by water, C1-C4 alcohol, or a mixed solvent thereof, or by acetone, ether, benzene, or a mixed solvent thereof so as to obtain an extract obtained by removing chlorophyll from Cheongyang Lycium chinense leaves. The composition having antioxidant and anti-inflammatory effects and including a Cheongyang Lycium chinense leaf extract as an active ingredient is obtained by efficiently and simply extracting an extract having antioxidant and anti-inflammatory effects from Cheongyang Lycium chinense leaves. Accordingly, a composition having antioxidant and anti-inflammatory effects and differentiated from conventional products can be provided.

Description

The composition with antioxidant and anti-inflammatory activity containing Lycium chinense leaf extract of Chengyang}

The present invention relates to a composition having antioxidant and anti-inflammatory effects using Cheongyang gugi leaf extract as an active ingredient, and more particularly, extracting gugi leaf with water, alcohol having 1 to 4 carbon atoms or a mixed solvent thereof, or from gugi leaf In order to obtain an extract from which chlorophyll has been removed, the present invention relates to a composition having antioxidant and anti-inflammatory effects, using as an active ingredient the Cheongyang gugi leaf extract extracted with acetone, ether, benzene, or a mixed solvent thereof.

Lycium Chinensis Miller tree is a deciduous shrub belonging to the eggplant family. It is native to Asia Minor. In Korea, Cheongyang-gun, Chungnam and Jindo-gun, Jeonnam are the main production areas.

Lycii fructus refers to the fruit (Lycii fructus) of the lychee tree, and the bark of the root is phalanx, the leaves are goji leaves, and the young shoots are called pure grass. According to Donguibogam, goji is effective in nourishment, tonic, blood, and jigal. According to herbal medicine, goji is not toxic. When taken, goji is not toxic. When taken, it does not cause heat and cold, and inflammation of the chest and diabetes are accompanied by thirst. It is good for diseases where nerves are paralyzed, and it has been recorded that it improves vision by promoting lung and kidney function.

Gugija and Goji leaf contain various functional ingredients such as betain, cholin, physalien, rutin, β-sitosterol, and zeazanthin, and have various antioxidant effects, antibacterial and anticancer effects, immunity enhancing effects, liver function improvement effects, blood cholesterol lowering effects, etc. It is known to exhibit physiological activity.

As a prior art, a cosmetic composition comprising a fermented wolfberry extract of Korean Patent Publication No. 10-2017-0092293 and a method for manufacturing the same (hereinafter,'Patent Document') are disclosed. The cosmetic composition comprising the fermented wolfberry extract disclosed in the above patent document is inoculated with 1 to 20% of the yeast culture solution based on the weight of the wolfberry powder in the wolfberry powder obtained by drying and pulverizing the wolfberry leaves, fruits, and roots, and 72 at 25 to 35°C. After fermentation for ~96 hours, containing fermented extract obtained by repeating the low-temperature extraction method for 12 to 24 hours at an extraction temperature of 20 to 60°C using water of 1 to 10 times the weight of fermented goji berry powder as a solvent. It has a characteristic to do.

However, the above patent document provides a fermentation extract obtained by inoculating a yeast culture from Gugija, and by extracting extracts having antioxidant and anti-inflammatory effects efficiently and simply from Goji leaves, it has differentiated antioxidant and anti-inflammatory effects from existing products. There is a need for technology development of the composition.

KR 10-2017-0092293 A (2017. 08. 11.)

The present invention has been devised to solve the problems of the prior art, and the object to be solved in the present invention is to provide a composition having antioxidant and anti-inflammatory effects from goji lobe.

In the composition having antioxidant and anti-inflammatory effects using the Cheongyang Goji leaf extract according to the present invention for achieving the above object, the Cheongyang Goji leaf extract is extracted with water, alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof. It features.

In addition, the Cheongyang gugi leaf extract is characterized in that chlorophyll is removed by extraction with acetone, ether, benzene, or a mixed solvent thereof.

In addition, it features a composition having antioxidant and anti-inflammatory effects comprising 0.1 to 10% by weight of the Cheongyang gugi leaf extract as an active ingredient.

In addition, the composition, characterized in that it has any one formulation selected from a flexible lotion, astringent lotion, nutrition lotion, nutrition cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray and powder To do.

In addition, the composition is characterized in that it has any one formulation selected from the group consisting of capsules, pills, and granules.

In addition, the composition is characterized in that it further comprises a food pharmaceutically acceptable carrier, excipient or diluent.

The composition having antioxidant and anti-inflammatory effects using Cheongyang Goji leaf extract according to the present invention as an active ingredient has antioxidant and anti-inflammatory effects differentiated from existing products by extracting extracts having antioxidant and anti-inflammatory effects efficiently and simply from Goji leaf. There is an effect of providing a composition.

1 is a view showing the cytotoxicity evaluation of Cheongyang gugi leaf extract (LLE) and Cheongyang gugi leaf extract (LLE with CR) from which chlorophyll was removed according to the present invention.
Figure 2 is a view measuring the amount of Nitric oxide (NO) production of Cheongyang gugi leaf extract (LLE) according to the present invention.
Figure 3 is a view measuring the amount of Nitric oxide (NO) production of Cheongyang gugi leaf extract (LLE with CR) from which chlorophyll has been removed according to the present invention.
Figure 4 is a view measuring the amount of inflammatory cytokine (TNF-α) production of Cheongyang gugi leaf extract (LLE) according to the present invention.
5 is a view measuring the amount of inflammatory cytokine (TNF-α) production of Cheongyang gugi leaf extract (LLE with CR) from which chlorophyll has been removed according to the present invention.
6 is a view measuring the amount of inflammatory cytokine (IL-6) production of Cheongyang gugileaf extract (LLE) according to the present invention.
7 is a view measuring the amount of inflammatory cytokine (IL-6) production of Cheongyang gugi leaf extract (LLE with CR) from which chlorophyll has been removed according to the present invention.

Hereinafter, preferred embodiments of the present invention will be described in detail with reference to the accompanying drawings.

The present invention relates to a composition having antioxidant and anti-inflammatory effects using Cheongyang gugi leaf extract as an active ingredient.

Goji leaf contains betaine, rutin, and beta-sitosterol, as well as vitamin C, which is good for skin care, and contains physiologically active substances such as antioxidant, anti-inflammatory, and moisturizing. .

Cheongyang gugi leaf extract is extracted with water, alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof.

In addition, the Cheongyang gugi leaf extract may be extracted with any one of acetone, ether, and benzene to remove chlorophyll.

First, a method of preparing a goji leaf extract is not particularly limited, and may be performed by a conventional method known in the art. For example, goji leaf is washed, dried and refined, and then extracted with an organic solvent, filtered through a filter paper, extracted with a separatory funnel, and the separated extract is concentrated under reduced pressure to prepare a goji leaf extract.

The organic solvent used in the present invention may be selected by mixing water, ethanol, methanol, acetone, ether, benzene or these organic solvents, preferably 70% ethanol, and added so as to be 8 times the weight of bulbous leaves. And extract.

And, in order to remove chlorophyll from goji leaves, it is used as acetone, ether, benzene, or a mixed solvent thereof, and is extracted by adding so as to be 8 times the weight of goji leaves.

At this time, the extraction temperature is 20 ~ 25 ℃, it is preferable to extract for 3 ~ 4 hours, and outside the extraction temperature and extraction time, extraction efficiency may decrease or a change in components may occur.

It will be described in more detail with respect to the manufacturing method of Goji leaf extract.

Goji leaf extract was washed, dried, and 800 parts by weight of 70% ethanol was added to 100 parts by weight of granulated goji leaf, and extracted with stirring for 3-4 hours at a temperature of 20-25℃, filtered through a filter paper, and extracted with a separatory funnel. The separated extract is prepared by concentrating with a rotary evaporator.

In addition, a method for preparing a chlorophyll-removed goji leaf extract will be described in more detail.

Goji leaf extract from which chlorophyll has been removed was washed, dried, and 800 parts by weight of acetone, ether, benzene or a mixed solvent thereof were added to 100 parts by weight of Goji leaf, followed by stirring and extracting at a temperature of 20 to 25°C for 3 to 4 hours. , Filtered with filter paper, extracted with a separatory funnel, and concentrated with a rotary evaporator to prepare the separated extract.

The composition having antioxidant and anti-inflammatory properties of the present invention may be prepared in any formulation, and solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing, oil, powder foundation, It can be formulated as an emulsion foundation, wax foundation, and spray, and more specifically, flexible lotion, astringent lotion, nutrition lotion, nutrition cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, It is formulated in packs, sprays and powders.

In addition, it is preferable to include 0.1 to 10% by weight of gugi leaf extract as an active ingredient with respect to the total composition.

It is preferable that the content of the composition be more than the minimum value so as to achieve the minimum antioxidant and anti-inflammatory effects, and the content of the composition is preferably less than the maximum value in consideration of the reduction in feeling of use and applicability to various formulations due to excessive addition. At this time, the content of the composition is preferably properly adjusted within the above range according to the content of the ingredients contained in the formulation or cosmetic composition.

Hereinafter, the present invention will be described in more detail through examples. However, the following examples are for illustrative purposes only, and are not intended to limit the scope of the present invention. That is, simple modifications or changes of the present invention can be easily implemented by a person skilled in the art, and all such modifications or changes can be considered to be included in the scope of the present invention.

[Experiment 1] Antioxidant experiment

(1) Measurement of total polyphenol content

The total polyphenol content was measured by the Folin-Denis method. 10μL of a mixture of 2N Folin-ciocalteu's phenol reagent (Sigma-Aldrich) and distilled water in a ratio of 1:2 by weight and 10μL of a sample were mixed and left in the dark for 3 minutes, followed by 20% sodium carbonate (Na2CO3, Junsei Chemical, Chuo- ku, Japan) 150 μL was added and reacted in the dark for 1 hour, and then absorbance was measured at 765 nm using an ELISA reader (Microplate Absorbance Spectrophotometer, Bio-rad, Hercules, CA, USA). The total polyphenol content was calculated from a standard curve prepared using gallic acid (Sigma-Aldrich) as a standard material.

Polyphenols that are closely correlated with the efficacy of antioxidant activity against Lycium'leaf extracts (LLE) and Lycium'leaf extracts with chlorophyll removal (LLE with CR)) It was confirmed whether there is a correlation by measuring the content of the.

(2) Measurement of total flavonoid content

The total flavonoid content was measured using the method of Zhishen et al. After adding 100 μL of distilled water and 7.5 μL of a 5% NaNO ₂ (sodium nitrite; Thermo Fisher Scientific, Waltham, MA, USA) solution to 25 μL of the sample, let stand for 5 minutes, 10% AlCl ₃ 6H ₂ O (aluminium chloride; Junsei Chemical , Tokyo, Japan) 15 μL of solution was added and reacted for 6 minutes, 50 μL of 1M NaOH (Samchun Pure Chemical, Pyeongtaek, Korea) was added and reacted at room temperature for 10 minutes, and the absorbance was measured at 510 nm using an ELISA reader. After preparing a standard curve using Catechin (Sigma-Aldrich) as a standard material, the total flavonoid content of the sample was calculated.

For the Cheongyang Goji leaf extract (LLE) and Cheongyang Goji leaf extract (LLE with CR) from which the chlorophyll was removed, the content of flavonoids, which had a close correlation with the efficacy of antioxidant activity, was measured to confirm whether there was a correlation.

<Total polyphenol content and total flavonoid content> sample Total polyphenol content
(mg GAE/g) ¹⁾ Total flavonoid content
(mg CE/g)
Goji leaf extract (LLE)

19.83±0.51

6.20±0.68
Goji leaf extract with chlorophyll removed
(LLE with CR)

1.39±0.38
7.33±0.37
¹⁾ GAE: gallic acid equivalents, CE: Catechin equivalents.
²⁾ Values represent mean±SD(n=3).

As a result of measuring the total polyphenol content, as shown in Table 1, it can be seen that the polyphenol content is high, and it can be confirmed that the antioxidant effect according to the polyphenol content is correlated.

In addition, as a result of measuring the total flavonoid content, as shown in Table 1, it can be seen that the flavonoid content is high, and it can be seen that the antioxidant effect according to the flavonoid content is correlated.

(3) DPPH radical scavenging ability measurement

DPPH radical scavenging ability was measured using DPPH (1,1-diphenyl-2-picrylhydrazyl, Sigma-Aldrich) by partially modifying the Blois method to check the radical scavenging ability of the sample. Mix 190 μL of 0.15 mM DPPH solution to 10 μL of samples diluted by concentration (0.125, 0.25, 0.5, 1, 2, 4, 8 mg/mL) and react in a dark room for 30 minutes, and then use an ELISA reader to measure the absorbance at 517 nm. Measured. The IC50 was calculated after calculating the DPPH radical scavenging ability by concentration using the following equation.

DPPH radical scavenging ability (%) = [1-(absorbance of experimental group/absorbance of control group)] x 100

(4) ABTS radical scavenging ability measurement

ABTS radical scavenging ability was measured by Fellegrini et al. 7mM ABTS[2,2'-azinobis(3-ethylenbenzothiazoline-6-sulfonate), Sigma-Aldrich] and 140mM K2S2O8 (Sigma-Aldrich) solution were well mixed and left in the dark for more than 12 hours. This was mixed with absolute ethanol at a weight-to-weight ratio of 1:88, and the ABTS solution adjusted so that the absorbance value of the control group at 734 nm was 0.7±0.02 was used. Mix 10 μL of the sample diluted by concentration (0.0625, 0.125, 0.25, 0.5, 1, 2, 4 mg/mL) and 200 μL of ABTS solution, react in the dark for 2 minutes and 30 seconds, and then measure the absorbance at 734 nm using an ELISA reader. Measured. After calculating the ABTS radical scavenging ability for each concentration, the IC50 was calculated.

ABTS radical scavenging ability (%) = [1-(absorbance of experimental group/absorbance of control)] x 100

(5) FRAP (ferric reducing antioxidant power) value measurement

The reducing power test by FRAP of the extract was measured by the Benzie and Strain method. 300mM sodium acetate buffer (pH 3.5), 10mM TPTZ (2,4,6-tris(2-pyridyl)-s-triazine), 20mM FeCl ₃ ·6H ₂ O, respectively, 10:1:1 (v/v/v) ) To prepare a FRAP reagent. After 10 μL of sample and 300 μL of FRAP reagent were mixed and reacted for 10 minutes at 37° C., absorbance was measured at 593 nm using an ELISA reader. FRAP value was obtained using FeSO ₄ ·7H ₂ O (iron(II) sulfate heptahydrate) as a standard material.

<DPPH radical scavenging ability measurement, ABTS radical scavenging ability measurement, FRAP (ferric reducing antioxidant power) value measurement>

Sample IC50 ¹⁾ (mg/mL)
FRAP value
(mg ISHE/g) ²⁾
DPPH radical
scavenging activity

ABTS radical
scavenging activity
Goji leaf extract (LLE)

6.17±0.40
2.63±0.03
47.74±0.71
Goji leaf extract with chlorophyll removed
(LLE with CR)

5.70±0.15
2.47±0.07
54.84±0.64
¹⁾ The concentration in mg/mL required for 50% reduction of DPPH and ABTS radical.
²⁾ ISHE: iron(Ⅱ) sulfate hepatahydrate equicalents (FeSO₄·7H₂O).
³⁾ Values represent mean±SD(n=3).

DPPH radical scavenging ability measurement, ABTS radical scavenging ability measurement, and FRAP (ferric reducing antioxidant power) value measurement results, as shown in Table 2, it can be confirmed that both the gugi leaf extract and the gugi leaf extract from which chlorophyll was removed have excellent antioxidant effects. .

[Experiment 2] Anti-inflammatory experiment

(1) Cytotoxicity evaluation (WST assay)

Cell proliferation was measured using water soluble tetrazolium salts (WST) in order to investigate the cytotoxicity of Goji leaf extract and Goji leaf extract from which chlorophyll was removed. 100 μL of RAW264.7 cells were dispensed into a 96-well plate at a concentration of 1×105 cells/mL, cultured for 24 hours, and then extracts (31.25, 62.5, 125, 250, 500, 1,000 μg/mL) and 1 μg/mL concentration (Sigma-Aldrich) was treated and cultured for 24 hours. After incubation, 10 μL of EZ-Cytox WST assay reagent (Dogenbio, Seoul, Korea) was added, and after 3 hours, absorbance was measured at 450 nm using an ELISA reader. The survival rate of the experimental group was expressed in% compared to that of the control group, and the results are shown in FIG.

As shown in Figure 1, it can be confirmed that the cytotoxicity does not appear.

(2) Nitric oxide (NO) production measurement

RAW264.7 cells were dispensed at a concentration of 4X105cells/mL into a 48-well plate at a concentration of 500 μL, cultured for 24 hours, and then extracted (31.25, 62.5, 125, 250, 500, 1,000 μg/mL) and LPS at a concentration of 1 μg/mL. And cultured for 24 hours. After taking a certain amount of cell culture solution and mixing it with the same amount of Griess reagent (Promega, Madison, WI, USA), the absorbance was measured at 550 nm using an ELISA reader. The content of NO generated from the cells was calculated using a standard curve prepared using sodium nitrite (NaNO2), and the results are shown in FIGS. 2 and 3.

As shown in Figures 2 and 3, it can be seen that the extract from the gugi leaf extract and the chlorophyll removed according to the present invention has a high anti-inflammatory effect.

(3) Measurement of inflammatory cytokines (TNF-α, IL-6) production

RAW264.7 cells were dispensed into 48 well plates at a concentration of 4×105 cells/mL in 500 μL each, cultured for 24 hours, and then extracted (31.25, 62.5, 125, 250, 500, 1,000 μg/mL) and LPS (1 μg/mL) by concentration. ) And cultured for 24 hours. The amount of TNF-α and IL-6 produced in the cell culture solution was treated according to the suggested method using a mouse ELISA kit (BD Biosciences, San Diego, CA, USA), and then the absorbance was measured at 450 nm with an ELISA reader and then each standard The production amount of inflammatory cytokines was calculated based on the curve, and the results are shown in FIGS. 4 to 7.

As shown in FIGS. 4 to 7, it can be seen that the extract from Goji leaf and the chlorophyll-removed extract according to the present invention has a high anti-inflammatory effect.

Claims (12)

A composition having antioxidant and anti-inflammatory effects using Cheongyang gugileaf extract as an active ingredient.
The method of claim 1,
The Cheongyang Goji leaf extract is a composition having antioxidant and anti-inflammatory effects, characterized in that extracted with water, alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof.
The method of claim 1,
A composition having antioxidant and anti-inflammatory effects using 0.1 to 10% by weight of Cheongyang gugi leaf extract as an active ingredient.
The method according to any one of claims 1 to 3,
The composition,
Antioxidant and anti-inflammatory effect, characterized by having any one formulation selected from flexible lotion, astringent lotion, nutrition lotion, nutrition cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray and powder A composition having.
The method according to any one of claims 1 to 3,
The composition,
A composition having antioxidant and anti-inflammatory properties, characterized in that it has any one formulation selected from the group consisting of capsules, pills, and granules.
The method according to any one of claims 1 to 3,
The composition,
A composition having antioxidant and anti-inflammatory properties, characterized in that it further comprises a food pharmaceutically acceptable carrier, excipient or diluent.
A composition having antioxidant and anti-inflammatory effects using Cheongyang gugileaf extract from which chlorophyll has been removed as an active ingredient.
The method of claim 7,
The cheongyang gugi leaf extract from which the chlorophyll has been removed is extracted with acetone, ether, benzene, or a mixed solvent thereof. A composition having antioxidant and anti-inflammatory effects.
The method of claim 7,
A composition having antioxidant and anti-inflammatory effects using 0.1 to 10% by weight of Cheongyang gugileaf extract from which the chlorophyll has been removed as an active ingredient.
The method according to any one of claims 7 to 9,
The composition,
Antioxidant and anti-inflammatory effect, characterized by having any one formulation selected from flexible lotion, astringent lotion, nutrition lotion, nutrition cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray and powder A composition having.
The method according to any one of claims 7 to 9,
The composition,
A composition having antioxidant and anti-inflammatory effects, characterized in that it has any one formulation selected from the group consisting of capsules, pills, and granules.
The method according to any one of claims 7 to 9,
The composition,
A composition having antioxidant and anti-inflammatory properties, characterized in that it further comprises a food-acceptable carrier, excipient, or diluent.

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