KR20200066910A - Anticancer composition comprising herbal extract - Google Patents

Abstract

The present invention relates to an anticancer composition. According to the anticancer composition containing an herbal medicine extract as an active component, the anticancer composition has an anticancer effect that specifically inhibits proliferation of cancer cells without affecting the growth of normal cells, and also can inhibit cancer cell metastasis. More specifically, the present invention relates to a lung cancer therapeutic agent containing the herbal medicine extract which can exhibit an excellent effect on preventing and treating lung cancer by using the herbal medicine extract selected from a group consisting of Astragalus propinquus, Codonopsis lanceolate, Mori cortex, and a mixture thereof.

Description

Anticancer composition comprising herbal extract as an active ingredient

The present invention relates to an anti-cancer composition comprising an herbal extract as an active ingredient, and has an anti-cancer effect that specifically inhibits proliferation, but not cancer cell metastasis, without affecting normal cell growth. It relates to a composition for anti-cancer that has an excellent effect, which can be useful for the prevention or treatment of cancer, and to assist in anti-cancer treatment.

Cancer is one of the incurable diseases that mankind needs to solve, and a huge amount of capital is being invested in the development to cure it worldwide. In Korea, the disease is the number one cause of death, and more than 100,000 people are diagnosed annually. More than 60,000 people are dying.

Carcinogens that cause these cancers include smoking, ultraviolet rays, chemicals, food, and other environmental factors, but the causes of this are diverse, making it difficult to develop therapeutic agents, and the effectiveness of therapeutic agents varies depending on the site where they occur. Currently, substances used as therapeutic agents have considerable toxicity, and cannot selectively remove only cancer cells, so it is urgently needed to develop effective anti-cancer drugs with low toxicity to prevent the development of cancer as well as treatment after cancer. Do.

Chemotherapy, radiation therapy, surgical therapy, immunotherapy, etc. are being used in Western medicine for cancer treatment. Due to its strong toxicity to normal tissues and organs of the human body as well as cancer cells, digestive disorders, bone marrow function suppression, and immune function decrease in patients. There are many disadvantages that cause side effects such as inflammatory reactions and body weakness, and recently, the development of a therapeutic agent that exhibits anti-cancer effects without side effects has been studied in Chinese medicine.

The treatment of cancer in oriental medicine includes dry swelling, dry blood consonants, positive swelling, bosin hot spring, dry swelling, etc. The law of injustice, such as cheongyeol detoxification, live blood language, hwadamso, and kigisojong , It is summarized as an injustice method (扶正祛邪法) which is a combination of an injustice method and a prosaic method. Among the cheating methods, dry-wing technique is a method that can be used for digestive disorders such as loss of appetite, nausea, nausea, vomiting, diarrhea, and abdominal swelling, which are often seen when using clinical symptoms of cancer, anticancer drugs, and chemotherapy drugs, and lower immune function. Representative prescriptions such as Sagunja-tang, quadrang-gun, and Bojung-ikgi-tang are experimentally effective in improving side effects of cancer treatment or chemo and radiation therapy, and clinically. It has been reported that it can be used for digestive cancer, liver cancer, and lung cancer.

Utilizing the above-mentioned oriental medicine ripening method (益氣養陰法) and geodam method (祛痰法), a therapeutic agent capable of exhibiting an anti-cancer effect without side effects has been developed, and a therapeutic agent that can be used as a single anti-cancer agent or as an anti-cancer adjuvant Development is necessary.

(Patent Document 1) KR 10-2003-0023243 (03.19.2003)

An object of the present invention is to provide a composition for anti-cancer containing the herbal extract as an active ingredient.

Another object of the present invention is to provide a composition for anti-cancer that has an anti-cancer effect that specifically inhibits proliferation and inhibits cancer cell metastasis without affecting normal cell growth.

Another object of the present invention is to provide a therapeutic agent for lung cancer using herbal extracts selected from the group consisting of astragalus, ginseng, agar, and mixtures thereof.

In order to achieve the above object, the composition for anticancer according to an embodiment of the present invention is from the group consisting of Astragalus mongholicus Bunge, Adenophoratriphylla var. japonica Hara, Mori Cortex Radicis and mixtures thereof The selected herbal extract or fractions thereof may be included as an active ingredient.

The extract is extracted with water, C1 to C4 alcohol, ether, chloroform, ethyl acetate, methylene chloride, or a mixture thereof.

The fraction is to fractionate any one selected from the group consisting of hexane, chloroform, ethyl acetate and butanol.

The anti-cancer composition is excellent in preventing or treating any one or more cancers selected from the group consisting of prostate cancer, uterine cancer, liver cancer, rectal cancer, lung cancer, breast cancer, and ovarian cancer.

The pharmaceutical composition according to another embodiment of the present invention is to include the composition for anticancer.

The pharmaceutical composition for preventing or treating lung cancer according to another embodiment of the present invention is to include the composition for anticancer.

Functional food composition according to another embodiment of the present invention is to include the composition for anti-cancer.

Hereinafter, the present invention will be described in more detail.

The composition for anti-cancer according to an embodiment of the present invention is a herbal extract selected from the group consisting of astragalus mongholicus bunge, adenophoratriphylla var. japonica hara, mori cortex radicis and mixtures thereof Fractions are included as active ingredients.

More specifically, each of the extracts of astragalus, ginseng and epidermis is used as a composition for anti-cancer, or a mixture of astragalus and epithelium extracts, a mixture of astragalus and ginseng extract, a mixture of apricot and ginseng extract, or a mixture of astragalus, sasam and epithelium extracts is anti-cancer It can be used as a composition.

The Astragalus mongholicus Bunge is a perennial herb belonging to the legume (Leguminosae), and is peeled and dried to harvest in the fall of the second year after sowing. It is also used. At the root of astragalus, there are more than 40 types of polysaccharides such as tritepene glycoside (saponin), astrgaram I, II, flavonoids such as camphor, and quentin, and more than 20 types of free amino acids, iron, magnesium, etc. It is known to contain trace elements of.

The ginseng (Adenophoratriphylla var. japonica Hara) is one of five ginseng together with ginseng (人蔘), hyeonsam (玄蔘), dansam (丹蔘), and gosam (苦蔘). In Korea, it refers to the roots of the stems belonging to the Campanulaceae family. The perennial plant is a perennial herb that grows in Korea, Japan, and China around 50 to 100 cm in height, with dense hairs and thick roots all over the ground, and flowers bloom in July to September in purple bells.

Sasam is described as the root of the Corollaopsis lanceolata in the North Korean Pharmacopoeia. According to the Food Ingredients Base of the Ministry of Food and Drug Safety, Ginseng is the name of the herbal medicine of the zodiac, and it is called Bekju, Namsasam, Jimo, Dancer, Yangyu, Ambassador, Ginseng, Possam, or Japanese Lady Bell.

It is a perennial herb of the Campanulaceae family. It is used as a detoxifying agent and expectorant for eating as an herb and distributed in Korea, Japan, and China. As a major component, adenofolate methyl ester, beta sitosterol, docosterol, lupeon, and tripyrrole are known, and pharmacological actions have been reported of expectorant, strong heart, immunomodulatory and antifungal and antioxidant activities.

The Mori Cortex Radicis is the root of the mulberry plant, a deciduous tree belonging to the family Mulaceae, the root bark of the mulberry is 1 to 4 mm thick, the outer surface is white or pale yellowish white, relatively flat, yellow or yellowish brown scales There is a pattern. Since ancient times, epithelial skin has antipyretic, anticonvulsant, anti-allergic, anti-diabetic, and anti-inflammatory effects, and has recently been shown to have a protective effect on the liver and hair growth effect against alopecia, whitening, antioxidant, diuretic stimulation, and suppression of an inflammatory inflammatory reaction. Known (Hikino et al., 1985; Kim et al., 1999,; Kim et al., 1995).

The astragalus extract may be extracted with a solvent selected from the group consisting of water, alcohol having 1 to 6 carbon atoms, alcohol having 1 to 6 carbon atoms, ether, chloroform, ethyl acetate, methylene chloride, and mixtures thereof. The'Astragalus extract' means an extract obtained by extracting from astragalus.

The astragalus extract is a mixture of a pulverized dry astragalus pulverized water, a polar solvent such as alcohol having C ₁ to C ₆ carbon atoms such as ethanol, methanol, or a mixed solvent having a mixing ratio of 1:0.1 to 1:10 of alcohol and water. It can be eluted, and more specifically, ethanol can be used as an extraction solvent. At this time, the extraction temperature may be 10 ℃ to 100 ℃, preferably an extract extracted at room temperature.

The extraction solvent may be used 2 to 50 times based on the weight of the sample, preferably 2 to 20 times. For extraction, the sample can be left for 1 to 72 hours for leaching in the extraction solvent, and in particular for 1 to 24 hours.

The filtered extract may be a product obtained by concentrating under reduced pressure with a vacuum rotary concentrator, but is not limited thereto, as long as it is an astragalus extract capable of exhibiting the anti-cancer effect of the present invention, and is obtained by drying an extract, a diluent or a concentrate of the extract, and drying the extract Dry matter, or a crude or purified product thereof.

The ginseng extract and the epidermis extract can also be prepared using the same method as the hwanggi extract and the ginseng extract and the epithelium extract.

Preferably, the astragalus extract may use astragalus fraction, and the astragalus fraction may more specifically use the ethyl acetate fraction of astragalus extract.

The ginseng extract may use a ginseng fraction, and more specifically, the ginseng fraction may use a hexane fraction of a ginseng extract or an ethyl acetate fraction of a ginseng extract.

As the above extract, the methylene chloride extract using methylene chloride as an extraction solvent may be used.

Preferably, an herbal extract selected from the group consisting of ethyl acetate fraction of astragalus extract, hexane fraction of ginseng extract, ethyl acetate fraction of ginseng extract, methylene chloride extract of epithelium and mixtures thereof can be used as a composition for anticancer.

When using the hwanggi extract, ginseng extract and epithelium extract as a composition for anticancer, due to the effect of reducing the activity of a protein cloned from the v-akt oncogene of retrovirus AKT8 (AKT) caused by ROS (Reactive Oxygen Species), It is effective against cancer cell lines.

In addition, it has an excellent effect of inhibiting interaction between stromal cells and cancer cells, and an excellent inhibitory effect on M2 macrophage differentiation, and thus can exhibit excellent tumor microenvironmental control effects.

In addition, it inhibits the migration of fibroblast and macrophage to cancer cells, inhibits cancer cell exacerbation factor secretion, reduces the interaction between cancer cells and stromal cells, and inhibits M2 macrophage differentiation, which adversely affects the tumor microenvironment. Can be adjusted.

The hwanggi extract, ginseng extract, and epithelium extract, respectively, when used alone, as described above, the effect of reducing AKT activity caused by excellent ROS, the inhibition effect of the interaction between matrix cells and cancer cells (M2 Macrophage differentiation inhibitory effect, Although it can exhibit the effect of inhibiting the interaction between cancer cells and stromal cells, when using a mixture of astragalus extract, ginseng extract and epithelium extract, it can exhibit a superior anti-cancer effect.

Preferably, the astragalus extract, ginseng extract and epithelium extract are used by mixing in a weight ratio of 1:1:1 to 1:1:0.5, the effect of reducing AKT activity by causing excellent ROS within the weight range, substrate Interaction between cells and cancer cells (Interaction) inhibitory effect, M2 Macrophage differentiation inhibitory effect, and may exhibit an effect of inhibiting the interaction between cancer cells and stromal cells, when the range value is less than or exceeded, such an anti-cancer effect does not occur.

More preferably, the herbal extract containing the ethyl acetate fraction of the astragalus extract, the hexane fraction of the extract of ginseng and the extract of methylene chloride from the astragalus extract, or the herbal extract of the ethyl acetate fraction of the extract of astragalus extract, the ethyl acetate fraction of the extract of ginseng extract and the herbal extract of the mixture of methylene chloride extract When used, the anti-cancer effect is increased due to the synergistic effect of the mixed use between each component.

That is, according to the mixing ratio between the components, it will be said that it can exhibit an excellent anti-cancer effect compared to the case of using the herbal extract alone as a synergistic effect of the mixing use between the components.

The anticancer composition is excellent in preventing or treating any one or more cancers selected from the group consisting of prostate cancer, uterine cancer, liver cancer, rectal cancer, lung cancer, breast cancer, and ovarian cancer, and preferably has excellent prevention or treatment effect of lung cancer It is to provide a composition for anticancer.

The pharmaceutical composition according to another embodiment of the present invention includes the anti-cancer composition.

The pharmaceutical composition for preventing and treating lung cancer according to another embodiment of the present invention includes the anti-cancer composition.

The pharmaceutical composition of the present invention may include a pharmaceutically acceptable carrier. The composition comprising a pharmaceutically acceptable carrier may be various oral or parenteral formulations. In the case of formulation, it may be prepared using diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, surfactants, etc., which are usually used. Solid preparations for oral administration may include tablets, powders, granules, capsules, etc. These solid preparations include at least one excipient in one or more compounds such as starch, calcium carbonate, sucrose or lactose ( lactose), gelatin, and the like. In addition, lubricants such as magnesium stearate, talc, etc. may be used in addition to simple excipients. Liquid preparations for oral administration include suspending agents, intravenous solutions, emulsions, syrups, etc. In addition to water and liquid paraffin, which are commonly used, various excipients, such as wetting agents, sweeteners, fragrances, and preservatives, can be included. have. Formulations for parenteral administration may include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, freeze-dried preparations, and suppositories. Non-aqueous solvents, suspension solvents may include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate. As a base for suppositories, witepsol, macrogol, tween 61, cacao butter, laurin butter, and glycerogelatin may be used.

In addition, the pharmaceutical composition of the present invention is from the group consisting of tablets, pills, powders, granules, capsules, suspensions, intravenous solutions, emulsions, syrups, sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilizers and suppositories. It can have any one formulation selected.

The extracts of hwanggi, ginseng extract and musketeers from the present invention have been used for edible and medicinal purposes since ancient times, and there are no particular restrictions on their dosage, body absorption, body weight, patient's age, sex, health status, diet, administration time , Administration method, excretion rate, disease severity, etc. may be changed. In general, the root extract can be administered in detail about 10 to 1000 mg per kg of body weight, and more specifically, about 50 to 500 mg per kg of body weight.

The pharmaceutical composition comprising the astragalus extract, ginseng extract and musk pit extract of the present invention as an active ingredient is prepared in consideration of an effective amount range, and the unit dosage form formulation so formulated is an expert for monitoring or observing the administration of the medicament as necessary. Depending on the judgment of the individual and the needs of the individual, a specialized dosing method may be used or may be administered several times at regular time intervals.

Health functional food according to another embodiment of the present invention includes the composition for anti-cancer.

Preferably, the composition for anti-cancer for use as a health functional food may include a mixture of astragalus extract, ginseng extract and epidermis extract, Kirilow indigo extract, and Pachysandra terminalis extract.

When preparing a composition for anticancer using only the extracts of hwanggi, ginseng and baekbaekpi, there is a problem in that the preference for the composition for anticancer is poor due to the bitter taste peculiar to hwanggi, ginseng and baekbaekpi.

Thus, when adding a small amount of ground fern extract and suhocho extract, to prepare a composite composition, the anti-cancer effect is maintained, and the preference can be increased.

More preferably, the composition may include 5 to 10 parts by weight of Kirilow indigo extract and 5 to 10 parts by weight of Pachysandra terminalis extract, based on 100 parts by weight of the complex mixture of astragalus extract, ginseng extract, and musk extract. have. When the extract is further included, the thick and bitter tastes of hwanggi, ginseng, and baekbaekpi are neutralized, and the flavor is improved by mixing with other extracts, thereby providing a more palatable food composition.

There are no particular restrictions on the type of health functional food of the present invention. Examples of health functional foods to which the extract mixture can be added are meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gums, dairy products including ice cream, various soups, beverages, tea , Drinks, alcoholic beverages and vitamin complexes, and may include all health functional foods in a general sense, and may include foods used as feed for animals. In addition to the above, the health functional food composition of the present invention includes various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH adjusting agents, stabilizers, preservatives, glycerin , Alcohol, carbonic acid used in carbonated beverages, and the like. In addition, it may contain flesh for the production of natural fruit juices, fruit juice drinks and vegetable drinks.

According to an anti-cancer composition comprising the herbal extract of the present invention as an active ingredient, it has an anti-cancer effect that specifically inhibits proliferation, but not cancer cell metastasis, without affecting normal cell growth. have.

More specifically, the present invention relates to a treatment for lung cancer, which includes a biomedical extract that can exhibit excellent effects in the prevention and treatment of lung cancer as an active ingredient by utilizing a herbal extract selected from the group consisting of astragalus, ginseng, acupuncture and mixtures thereof.

1 is a flow cytometry result for ROS according to the treatment of a complex herbal extract according to an embodiment of the present invention.
2 is a result of MTT analysis and dye exclusion ability analysis (Trypan blue exclusion assay) according to the treatment of the complex herbal extract according to an embodiment of the present invention.
Figure 3 is a flow cytometry analysis results for the sub-G1 phase and nnexin V-positive cell population according to the treatment of the complex herbal extracts according to an embodiment of the present invention.
4 is a Western blot result for the expression of p-AKT according to the treatment of the complex herbal extract according to an embodiment of the present invention.
5 is a result of measuring the cell viability according to the treatment of the complex herbal extract according to an embodiment of the present invention.
Figure 6 is a result of measuring the anti-cancer effect in human lung cancer cell lines according to the treatment of a complex herbal extract according to an embodiment of the present invention.
7 is a result of inhibiting cell proliferation of H1299 cells according to the treatment of a complex herbal extract according to an embodiment of the present invention.
8 is a result of MTT analysis, pigment exclusion ability analysis and soft agar analysis for cell viability and colony formation of H1299 cells according to treatment of a complex herbal extract according to an embodiment of the present invention.
9 is a Western blot result for the expression of p-EGFR, p-STAT3, p-SRC according to the treatment of the complex herbal extract according to an embodiment of the present invention.
Figure 10 is a Western blot result for the expression of p-EGFR, p-STAT3, p-SRC according to the treatment of the complex herbal extracts according to an embodiment of the present invention.
11 is a result of Transwell analysis of the interaction between fibroblast and cancer cells according to the treatment of the complex herbal extract according to an embodiment of the present invention.
12 is a result of inhibiting the interaction between the fibroblast and cancer cells according to the treatment of the complex herbal extract according to an embodiment of the present invention.
13 is an analysis result of factors affecting M2 macrophage differentiation according to treatment of a complex herbal extract according to an embodiment of the present invention.
14 is a result of transwell analysis of the interaction between vascular endothelial cells and cancer cells according to the treatment of a complex herbal extract according to an embodiment of the present invention.
15 is a result of cell adhesion analysis according to the treatment of a complex herbal extract according to an embodiment of the present invention.
16 is a result of Transwell analysis according to the treatment of a complex herbal extract according to an embodiment of the present invention.
17 is a result of confirming whether or not it affects various factors secreted by cancer cells according to the treatment of the complex herbal extract according to an embodiment of the present invention.
18 is a result of transwell analysis on whether to inhibit recruitment of HP-1 macrophage according to treatment of a complex herbal extract according to an embodiment of the present invention.
19 is a result of transwell analysis on whether to inhibit recruitment of HP-1 macrophage according to treatment of a complex herbal extract according to an embodiment of the present invention.
20 is a result of Transwell analysis on whether to suppress recruitment of WI38 fibroblast according to treatment of a complex herbal extract according to an embodiment of the present invention.
21 is a result of transwell analysis on whether to suppress recruitment of WI38 fibroblast according to treatment of a complex herbal extract according to an embodiment of the present invention.
22 is a result of confirming the expression level of p-p65, p-STAT3, p-SRC, p-AKT and p-ERK of H1299 induced by PMA according to the treatment of the complex herbal extract according to an embodiment of the present invention .
23 is a result of confirming the expression level of mRNA of plasminogen activator inhibitor-1 (PAI-1) according to the treatment of a complex herbal extract according to an embodiment of the present invention.
24 is a result of analyzing the cytokine array according to the treatment of the complex herbal extract according to an embodiment of the present invention.
25 is a result of Transwell analysis for confirming the migration of stromal cells to lung cancer cells in WI38 fibroblast and Raw264.7 macrophage according to treatment of a complex herbal extract according to an embodiment of the present invention.
26 is an analysis result of mRNA expression levels of arginase-1, MRC-1, and IL-10 according to treatment of a complex herbal extract according to an embodiment of the present invention.
27 is a result of transwell analysis for confirming the effect on migration of Lewis lung carcinoma (LLC) cells according to treatment of a complex herbal extract according to an embodiment of the present invention.
28 is a result of cell adhesion analysis according to the treatment of a complex herbal extract according to an embodiment of the present invention.

Hereinafter, embodiments of the present invention will be described in detail so that those skilled in the art to which the present invention pertains can easily practice. However, the present invention can be implemented in many different forms and is not limited to the embodiments described herein.

[Production Example 1: Preparation of natural extracts]

The Hwanggi, Sasam, and Sangbaekpi used in this experiment were purchased from Nuri Herb Co., Ltd. and were tested for purity and tested for standard products based on the Korean Pharmacopoeia in Hwasun Hanjae Pharmaceutical Distribution. To obtain ethanol extract of Astragali membranaceus (EAM) and ethanol extract of Adenophora triphilla var. japonica, EAT, 1.5 L of 80% ethanol was added to 100 g of dried powder of Hwanggi and Ginseng each. After that, it was extracted 5 times for 30 minutes in an ultrasonic grinder to separate only the supernatant through a filter. The supernatant obtained by repeating this process twice was concentrated under reduced pressure and lyophilized to dissolve the powder obtained in dimethylsulfoxide (DMSO) at a concentration of 200 mg/ml.

To obtain methylene chloride extract (Methylenechloride extract of Mori Cortex Radicis, MEMC), except for using methylene chloride as an extraction solvent, MEMC was prepared using the same production method as that of EAM and EAT.

The EAM, EAT, and MEMC are suspended in 10 L of water, and sequentially extracted three times with 10 L of butanol, hexane and ethyl acetate, respectively, as a solvent, butanol fractions (BEAM, BEAT and BMEMC), and hexane fractions (HEAM, HEAT and HMEMC) and ethyl acetate fractions (EEAM, EEAT and EMEMC) were prepared.

RPMI 1640, fatal bovine serum (FBS), penicillin-streptomycin (PS), phosphate-buffered saline (PBS) and Trypsin-EDTA used for cell culture were purchased from WelGENE (Seoul, Korea). All primary and secondary antibodies for protein analysis are Santa Cruz Biotechnology Inc. (Santa Cruz, CA, USA), and all reagents not specified were purchased from Sigma (St. Louis, MO).

[Experimental Example 1: Experiment of anticancer effect on lung cancer cell line]

1. Experimental method

Cell culture

The H1299 human lung cancer cell line used in this study was provided by Prof. Ho-Young Lee, College of Pharmacy, Seoul National University. For the cultivation of cells, 10% FBS and 1% PS were added to 90% RPMI 1640, and the cells were cultured under 37°C and 5% CO2 conditions.

MTT assay

After seeding H1299 cells in a 96 well plate at a concentration of 2.5×10 ³ cells/well, EAM, EAT, and EMC were treated at the suggested concentrations, respectively. After 72h incubation, tetrazolium bromide salt (MTT) was dispensed to a concentration of 0.5 µg/ml and incubated for 2 h. After that, the supernatant was removed and 100 μl of DMSO was added to dissolve the formazin of each well, and absorbance was measured at 540 nm in a multiplate reader (Molecular Devices, Sunnyvale, CA, USA).

Trypan blue exclusion assay

After seeding H1299 cells in a 12 well plate at a concentration of 3×104 cells/well, EAM (500 µg/ml), EAT (500 µg/ml) and EMC (500 µg/ml) were each treated for the indicated time. After that, the cells were collected, suspended in a 0.1% trypan blue solution, and the cell number was counted. Cells dyed in blue were judged as dead cells and excluded from counting. Only unstained cells were counted as live cells.

Soft agar assay

4% low-melting agarose (Lonza, Rockland, ME) was prepared by dissolving in advance in PBS, and then warm media was added to make a 1% bottom agar, and 1 ml was dispensed into a 24 well plate to harden at room temperature. Thereafter, H1299 cells were suspended in 0.4% top agar at a concentration of 5×10 ² cells/0.5 mL, and dispensed 0.5 mL per well, and then solidified at room temperature. After the Agar was hardened, EAM and EAT were treated in 500 µl of the medium, and then dispensed on top agar. The supernatant was replaced once every 3 days, and cultured for 2 weeks after seeding. After that, the supernatant was treated with 0.5 mg/ml of MTT solution for 2 h incubation, and the dyed colony was photographed with a digital camera (EOS750D, Canon, Japan). Colony numbers were counted through Image J software.

DAPI staining

H1299 cells treated with EAM (500 µg/ml), EAT (500 µg/ml) and EMC (500 µg/ml) were collected and fixed with 3.7% paraformaldehyde for 30 minutes at room temperature. Then, after washing twice with cold PBS, cells were attached to slide glass with cytospin, washed alternately with PBS and DW, and dried. The dried cells were stained in a dark room for about 30 minutes with DAPI solution diluted to 2.5 mg/ml, alternately washed and dried in PBS and DW, and then mounted and mounted through a fluorescence microscope (Carl Zeiss, Germany) at a magnification of 400 times. The shape of the was observed.

Cell cycle analysis through flow cytometry

H1299 cells treated with EAM (500 µg/ml), EAT (500 µg/ml) and EMC (500 µg/ml) were collected for 72 h, fixed with 80% ethanol at 4°C for 1 h, and centrifuged to give 500 µl of propidium iodide solution The cells were suspended in (450 μl PBS, 50 μl 0.5 mg/ml propidium iodide, 1.5 μl 10 mg/ml RNase A) and stained for 30 minutes. Thereafter, the supernatant was removed by centrifugation, and then suspenstion in 500 μl PBS to analyze the cell cycle by histogram using a DNA flow cytometer (BD FACSCalibur) and CellQuest software.

Annexin V/PI double staining assay

H1299 cells treated with EAM (500 µg/ml), EAT (500 µg/ml) and EMC (500 µg/ml) for 72h were collected and stained according to the instructions of the Annexin VFITC apoptosis detection kit (PharMingen, San Diego, California) Did. After staining the cells in the dark for 15 minutes, Annexin Vpositive apoptotic cells were analyzed using a DNA flow cytometer and CellQuest software.

Western blotting

H1299 cells treated with EAM (500 µg/ml), EAT (500 µg/ml) and EMC (500 µg/ml) for 72 h were collected and lysis buffer (50 mM Tris-HCl (pH 7.4), 150 mM NaCl, 1% NP -40, 1% Triton X-100, 0.25% sodium deoxycholate, 1 mM EDTA, 10% Glycerol, a complete protease inhibitor cocktail tablet (Roche diagnostics, Mannheim, Germany), phosphatase inhibitors (1 Mm Na3VO4, 100 mM NaF, and 10 mM NaPP)) after 1 h lysis and quantified according to the Bio-rad protein quantification reagent and its usage. Thereafter, 20 μg of protein was loaded onto SDS-polyacrylamide gel, electrophoresed, transferred to PVDF membrane (Millipore Corporation, Bedford, MA) for 2 h through electroblotting, and then 3% bovine serum albumin (BSA, MP Biomedicals Europe, Illkirch) , France) for 1 h and attached the primary antibody, and incubated overnight at 4℃. After that, wash with TBST, attach the secondary antibody for 1 h at room temperature, then wash again, and spray the Enhanced Chemilunimoecence (ECL) solution (Amersham Life Science Corp., Arlington Height, IL, USA) in a dark room with X-ray film (Agfa- Gevaert, Melbourne, Australia).

Measurement of reactive oxygen species production

After seeding 5×10 ⁴ H1299 cells in a 6 well plate, stabilizing overnight, treating EAM (500 µg/ml), EAT (500 µg/ml) and EMC (500 µg/ml) hourly, 5-(and 6)-carboxy-2'7'-dichlorodihydrofluorescein diacetate (H2DCFDA) was treated with 10 μM and incubated at 37°C and 5% CO2 for 20 minutes. The stained cells were collected and suspended in 500 µl of PBS (pH 6.9) to observe relative changes in reactive oxygen species production using a DNA flow cytometer (BD FACSCalibur) and CellQuest software.

Cell cycle analysis

After seeding 1×105 H1299 cells in a 60Φ dish, stabilize them overnight, EAM (500 µg/ml), EAT (500 µg/ml), EMC (500 µg/ml), NAC (Co-treatment of N-acetyl) -L-cysteine, 5 mM). After 72 hours, cells were collected and fixed with 80% ethanol at 4°C for 1 h. The centrifuged cells were suspended in 500 μl of propidium iodide solution (450 μl PBS, 50 μl 0.5 mg/ml propidium iodide, 1.5 μl 10 mg/ml RNase A), and the nuclei were stained for 30 minutes. Thereafter, the supernatant was removed by centrifugation, and then suspended in 500 µl PBS (pH 6.9) to analyze the cell cycle by histogram using a DNA flow cytometer and CellQuest software.

Annexin V/PI double staining assay

After seeding 5×10 ⁴ H1299 cells in a 6-well plate, stabilize them overnight and treat with EAM (500 µg/ml), EAT (500 µg/ml), EMC (500 µg/ml), and NAC (5 mM). Did. After 72 hours, H1299 cells were collected and stained according to the manufacturer's manual using an Annexin V-FITC apoptosis detection kit (PharMingen, San Diego, California). After staining the cells in the dark for 15 minutes, fluorescence was analyzed using a DNA flow cytometer and CellQuest software. Annexin V-positive cells were analyzed as apoptotic cells to analyze the numerical values.

Statistical analysis

The results of all experiments were expressed as mean±SD, and statistical significance was judged to be P<0.05 using Student's t-test.

2. Experimental results

Effect of EAM-EAT combination treatment on the production of reactive oxygen species

To determine whether the anti-cancer synergistic effects of EAM and EAT are related to the occurrence of reactive oxygen species, the amount of free radicals produced was investigated by flow cytometry after treatment with EAM and EAT alone or in combination at a concentration of 500 µg/ml, where the anti-cancer synergistic effect appears. .

As a result, compared to EAM and EAT alone treatment, the generation of reactive oxygen species increased rapidly during the combination treatment, and the reactive oxygen species increased in a time-dependent manner, showing a tendency to increase more 6 h after 2 h after the combined treatment with EAM and EAT (Fig. 1A). . Compared to EAM and EAT alone treatment, it showed that the anti-cancer synergy effect of EAM and EAT was related to the occurrence of reactive oxygen species in association with the result of a significant increase in apoptosis when combined.

In addition, when EAM and EAT were treated in combination with each hour, reactive oxygen species were continuously generated from 6h to 48h, and it was predicted that the signaling pathway by the active oxygen species would have been activated for a long time (FIG. 1B).

Effects of free radical species on cell viability reduction by EAM-EAT combination treatment

In order to investigate the effect of free radicals on cell viability reduction by EAM-EAT combination treatment, NAC, an antioxidant, was treated with EAM-EAT. First, as a result of observing the cell density under a microscope, it was confirmed that the cell density, which was significantly reduced by the EAM-EAT combination treatment, increased again by NAC treatment (FIG. 2A ).

In addition, as a result of measuring the cell viability with the MTT assay and trypan blue exclusion assay, it was observed that the cell viability, which was rapidly decreased by the combination of EAM-EAT, inhibited the production of reactive oxygen species with NAC, and recovered a significant portion (FIG. 2B and FIG. 2C). ). These results indicate that reactive oxygen species play a key role in reducing cell viability by EAM-EAT combination treatment.

Effects of reactive oxygen species on apoptosis induction by EAM-EAT combination treatment

Next, the effect of free radical species on the induction of apoptosis by EAM-EAT combination treatment was investigated. First, after annexin V/PI double staining, annexin V-positive cells were analyzed by flow cytometry. In the early stage of apoptosis, phosphatidylserine (PS) located inside the cell membrane moves to the outside of the cell membrane and binds to annexin V, so that cells in the annexin V+/PI- state increase, and in the late apoptosis, the nucleus is stained by PI as pores form in the cell membrane Therefore, annexin V+/PI+ cells are increased. Therefore, annexin V is positive in cells with apoptosis regardless of stage. As a result of analyzing the data based on this, it was confirmed that the apoptotic cell population significantly increased by EAM-EAT combination treatment decreased to the control level by NAC treatment (FIG. 3A). Cell cycle analysis results also showed the same pattern, and the sub-G1 phase cell population, which means apoptotic cells, increased by EAM-EAT combination treatment and decreased by NAC treatment (FIG. 3B). These results show that the induction of apoptosis by the combined treatment of EAM-EAT is due to the production of reactive oxygen species.

Effect of free radical species production on AKT signaling pathway by EAM-EAT combination treatment

AKT is a signal molecule that suppresses apoptosis of cancer cells and contributes to cell survival, and is overactivated in various carcinomas. Therefore, drugs that can inhibit kinase activity by controlling the ATP-binding site or regulatory domain of AKT have been developed. As a result of performing Western blot to confirm that the anti-cancer synergistic effect by the EAM-EAT combination treatment is related to the regulation of the AKT signaling pathway, it was confirmed that the phosphorylation of AKT was significantly reduced when combined with EAM and EAT alone (Fig. 4A). Phosphorylation of AKT decreased by EAM-EAT combination treatment increased again when NAC was blocked to generate reactive oxygen species (FIG. 4B ). Through this, it was found that the decrease in the activity of AKT by the combined treatment of EAM-EAT was due to the occurrence of reactive oxygen species.

Influence of anti-cancer synergy by inhibiting AKT phosphorylation by EAM-EAT combination treatment

Cell cycle analysis was performed after treatment with LY294002 by concentration to confirm whether cell proliferation inhibition and apoptosis were induced in the H1299 cell line by inhibiting AKT phosphorylation alone. As a result, by confirming that the sub-G1 phase cell population of the H1299 cell line increased in a concentration-dependent manner, it was found that AKT phosphorylation inhibition induced apoptosis and inhibited cell proliferation (FIG. 5A).

Next, to investigate the effect of phosphorylation of AKT inhibited by EAM-EAT combination treatment on anti-cancer synergy, cell viability was measured by MTT assay after treatment with AKT inhibitor LY294002. As a result, it was confirmed that cell viability was significantly reduced when LY294002 was treated together in the 100 μg/ml and 250 μg/ml treatment groups, respectively, at concentrations at which anti-cancer synergy was not significantly observed (FIG. 5B). As a result of performing cell cycle analysis to confirm that the cell proliferation inhibition was caused by apoptosis, the sub-G1 phase cell population was significantly increased by the combination treatment of LY294002 and EAM-EAT (FIG. 5C), and apoptosis marker protein It was confirmed that the expression of phosphorus cleaved-PARP increased (FIG. 5D).

These results showed that the expression of p-AKT, which was weakly reduced by LY294002 alone, was significantly reduced by the combination treatment of LY294002 and EAM-EAT, and the phosphorylation level of AKT was closely related to the induction of apoptosis of the H1299 cell line. It was shown (Fig. 5E). Therefore, it was found that the inhibition of phosphorylation of AKT plays an important role in the combination of EAM-EAT treatment with anti-cancer synergy through induction of apoptosis.

Next, to confirm once again whether the inhibition of phosphorylation of AKT by EAM-EAT combination treatment is related to the generation of free radicals and apoptosis, NTT and LY294002 were treated together and then subjected to MTT assay. As a result, it was confirmed that the cell viability, which was significantly reduced by the EAM-EAT combination treatment, was restored by the NAC treatment, which again decreased to the level of the EAM-EAT combination treatment group by the LY294002 treatment (FIG. 5B). The decrease in the cell viability raised by the inhibition of reactive oxygen species production again by the inhibition of AKT activity means that the anti-cancer synergistic action of EAM and EAT by the reactive oxygen species production is mediated by the inhibition of AKT activity.

Analysis of cancer cell killing effect and mechanism of active ingredients of astragalus and ginseng

Among the extract fractions of astragalus and ginseng, the ethyl acetate layer and the hexane layer, respectively, showed the most significant anticancer effect, so AGIV and formononetin (FMN) (above astragalus effective) were found to have anticancer activity in other carcinomas among the active ingredients known to be extracted from these fractions. Ingredients) and lupeol (active ginseng active ingredient) were assumed as candidates, and the apoptosis effect in the lung cancer cell line and its mechanism were analyzed.

As a result of treating AG12, FMN, and lupeol on H1299 cells, it showed the most prominent cell proliferation inhibitory effect in lupeol (FIG. 7).

It was confirmed by MTT assay, trypan blue exclusion assay, and soft agar assay that Lupeol decreased the cell viability and colony formation of H1299 cells in a concentration-dependent manner.

This proliferation inhibitory effect was confirmed by inducing apoptosis through chromatin condensation through DAPI staining and subpopulation of sub-G1 phase cell population through apoptotic body formation and flow cytometry (FIG. 8).

Lupeol decreased the expression of p-EGFR, p-STAT3, and p-SRC in a concentration-dependent manner. After overactivating the most markedly reduced STAT3 through STAT3 Y705D transfection, there was no significant difference from the control vector as a result of confirming susceptibility to lupeol (FIGS. 9 and 10 ).

Tumor microenvironment regulation

EAM and EAT are divided into hexane layer (HEAM, HEAT), ethyl acetate layer (EEAM, EEAT) and butanol layer (BEAM, BEAT), respectively, and a total of 6 types of fractions are processed into WI38 fibroblast and Raw264.7 macrophage. Therefore, the experiment was conducted after the non-toxic conditions (cell survival rate of 90% or more) were obtained.

As a result of investigating whether the extract fraction influenced the interaction between fibroblast and cancer cells through a transwell assay, the interaction between WI38 fibroblast and H1299 cells was most significantly suppressed in the HEAT 100 μg/ml treatment group (FIG. 11 ).

In order to check whether HEAT affects various factors secreted by cancer cells, H1299 cells stimulated with PMA are treated with HEAT, received conditioned media (CM), placed in the down chamber of the transwell, and loaded with WI38 into the upper well to improve migration ability. As a result, increased migration during PMA treatment decreased to control level by HEAT treatment.

This will mean that HEAT inhibits the secretion of factors that help the interaction between fibroblast and cancer cells from cancer cells (FIG. 12).

As a result of examining the mRNA expression of IL-10, a key marker of M2 macrophage, to confirm the effect of Hwangki and Ginseng extract fractions on M2 macrophage differentiation, it was most significantly suppressed in the EEAT and BEAT treatment groups.

Accordingly, after treatment of EEAT and BEAT by concentration, mRNA expressions of arginase-1, MRC-1, and IL-10 were additionally investigated. As a result, expression was only decreased in the EEAT treatment group. P-STAT6, which regulates their expression by mediating the IL-4 signaling pathway, also decreased in a concentration-dependent manner in the EEAT-treated group, and did not show a significant decrease in the BEAT-treated group (FIG. 13).

As a result of investigating whether the extract fraction affects the interaction between vascular endothelial cells and cancer cells through a transwell assay, the interaction between HUVEC human vascular endothelial cells and H1299 cells was most significantly inhibited in the HEAT-treated group as in WI38 fibroblast (FIG. 14).

The ginseng extract fraction showed more remarkable tube formation suppression effect than the astragalus extract fraction, of which HEAT and EEAT showed the most excellent suppression ability.

As a result of comprehensive analysis, the effect of inhibiting the interaction between stromal cells and lung cancer cells is remarkable in HEAT, and the effect of inhibiting M2 macrophage differentiation is EEAT most prominent.

Tumor microenvironment control of epidermal extract

After treating the methylene chloride extract (MEMC) of the epithelium with stromal cells WI38 fibroblast and THP-1 cells differentiated with Raw264.7 macrophage and macrophage (THP-1 macrophage), the cells were treated under non-toxic conditions (over 90% cell viability). The experiment was conducted.

It was confirmed through a transwell assay that MEMC inhibited the interaction between Raw264.7 and H1299 in a concentration-dependent manner. In addition, it was observed that the expression of p-EGFR and p-p38 is significantly reduced by MEMC treatment (FIG. 16).

In order to confirm that MEMC affects various factors secreted by cancer cells, MEMC is treated with H1299 cells stimulated with PMA, receives conditioned media (CM), is placed in the down chamber of the transwell, and WI38 is loaded into the upper well to improve migration ability. As a result, increased migration during PMA treatment was decreased in a concentration-dependent manner by MEMC treatment (FIG. 17).

It was confirmed by a transwell assay that MEMC inhibited the interaction between THP-1 macrophage and H1299 cells, and inhibited recruitment of THP-1 macrophage by controlling factors secreted from H1299. In addition, MEMC reduced the expression of p-SRC and p-JNK in THP-1 macrophage (FIGS. 18 and 19 ).

It was confirmed by a transwell assay that MEMC inhibited the interaction between WI38 fibroblast and H1299 cells, and inhibited recruitment of WI38 fibroblast by controlling factors secreted from H1299. In addition, it was confirmed that MEMC decreases the expression of p-EGFR, p-STAT3, p-SRC, p-FAK, p-AKT and p-JNK of WI38 (FIGS. 20 and 21 ).

It was confirmed that the increase of p-p65, p-STAT3, p-SRC, p-AKT and p-ERK expression of H1299 induced by PMA decreases by MEMC treatment (FIG. 22 ).

It was confirmed that the expression of NF-kB increased and mRNA of plasminogen activator inhibitor-1 (PAI-1), which is known to exacerbate cancer, increased by PMA and then decreased by MEMC treatment. In addition, it was observed that the interaction between the H1299 lung cancer cell line inhibited by MEMC treatment and WI38 and THP-1 macrophage increased again by recombinant PAI-1 treatment. This will mean that MEMC inhibits the recruitment of stromal cells by inhibiting PAI-1 secretion of H1299 (FIG. 23).

It was confirmed that CXCL12 secretion was reduced by MEMC treatment as a result of cytokine array with CM obtained after PMA treatment with H1299 cells alone or with MEMC treatment (FIG. 24).

Overall, MEMC inhibits the migration of fibroblast and macrophage to lung cancer cells, reduces the interaction between lung cancer cells and stromal cells by inhibiting the secretion of cancer exacerbation factors of lung cancer cells, and inhibits M2 macrophage differentiation, thereby reducing the tumor microenvironment to cancer. It was confirmed that the adjustment was disadvantageous.

Control of tumor microenvironment of active ingredients of astragalus and ginseng

The effect of MN, Lupeol and AGIV on the control of the tumor microenvironment was investigated using WI38, Raw264.7 and THP-1 macrophage. All experiments were conducted under conditions of cell viability of 90% or more.

As a result of confirming the migration of stromal cells to lung cancer cells through a transwell assay, FMN, Lupeol, and AGIV were significantly decreased in WI38 fibroblast and Raw264.7 macrophage, and only FMN inhibited migration in THP-1 macrophage (Fig. 25).

As a result of examining mRNA expression of arginase-1, MRC-1, and IL-10, which are key markers of M2 macrophage, to determine how Hwanggi and Ginseng active components affect M2 macrophage differentiation, concentration-dependently in the Lupeol and AGIV treatment groups Expression decreased. P-STAT3, which regulates M2 macrophage differentiation through IL-10, was also decreased in a concentration-dependent manner in the Lupeol and AGIV treatment groups, but p-STAT6 expression was relatively insignificantly reduced (FIG. 26).

As a result of confirming the effect of inhibition of M2 macrophage differentiation of lupeol and AGIV on the migration of mouse lung cancer cell line, Lewis lung carcinoma (LLC) cells, transwell assay showed that both drugs showed significant concentration-dependent migration of LLC cell migration by IL-4 treatment. Suppressed. In addition, it was confirmed that both Lupeol and AGIV showed toxicity to LLC cells at concentrations that did not show toxicity to Raw264.7 cells, and thus had cancer cell specific toxicity (FIG. 27 ).

FMN, Lupeol, and AGIV did not affect HUVEC and cancer cell interaction or tube formation (FIG. 28).

Taken together, FMN commonly inhibited the interaction between lung cancer cells and fibroblast and macrophage, and Lupeol and AGIV inhibited M2 macrophage differentiation.

Effect of EAM-EAT-MEMC combination treatment on anticancer synergy

To confirm the anti-cancer synergistic effect by the combined treatment of EAM, EAT and MEMC, the weight ratio of EAM, EAT and MEMC is 1:1:0.1, 1:1:0.5, 1:1:1, 1:1:1.5. After treating the mixed extract mixed with, the amount of free radicals produced was investigated by flow cytometry.

For relative comparison, after 6 h of EAM and EAT combined treatment, after 6 h, the increase in the amount of free radicals was set to index 5, and the increase in the amount of free radicals in the complex extract was evaluated as an index. The index is evaluated from 1 to 10, and the higher the value, the better the effect.

DD1 DD2 DD3 DD4 DD5 Reactive oxygen species 5 3 6 7 4

(DD1 for EAM and EAT 1:1, DD2 for EAM, EAT and MEMC 1:1:0.1, DD3 for EAM, EAT and MEMC 1:1:0.5, DD4 for EAM, EAT and MEMC 1: 1:1, DD5 is a treatment of EAM, EAT and MEMC in 1:1:1.5) According to Table 1 above, compared to the case where EAM and EAT are combined, EAM, EAT and MEMC are treated. , It was confirmed that apoptosis resulted in a marked increase.

Effect of EEAM-HEAT-MEMC combination treatment on anti-cancer synergy

In order to confirm the anti-cancer synergy effect by the combination treatment of EEAM, HEAT and MEMC, the weight ratio of EEAM, HEAT and MEMC is 1:1:0.1, 1:1:0.5, 1:1:1, 1:1:1.5. After treating the mixed extract mixed with, the amount of free radicals produced was investigated by flow cytometry.

For relative comparison, when EAM and EAT were treated in a 1:1 ratio (DD1), index 5 was set, and the increase in the amount of free radicals in the complex extract was evaluated by index. The index is evaluated from 1 to 10, and the higher the value, the better the effect.

DD1 DD4 DDT1 DDT2 DDT3 DDT4 Reactive oxygen species 5 7 5 7 9 4

(DD1 for EAM and EAT 1:1, DD4 for EAM, EAT and MEMC 1:1:1, DDT1 for EEAM, HEAT and MEMC 1:1:0.1, DDT2 for EEAM, HEAT and MEMC 1: 1:0.5, DDT3 is EEAM, HEAT and MEMC 1:1:1, DDT4 is EEAM, HEAT and MEMC 1:1:1.5) According to Table 2 above, EAM, EAT and MEMC It was confirmed that apoptosis was significantly increased when EEAM, HEAT, and MEMC were treated compared to the combination treatment.

Whether the complex extract fraction affects the interaction between fibroblast and cancer cells was investigated by transwell assay. The HEAT treatment group showing the most remarkable inhibitory effect was set to an index of 5, and EEAM, HEAT and MEMC were mixed at a weight ratio of 1:1:0.1, 1:1:0.5, 1:1:1, and 1:1:1.5. The result of transwell assay for the treatment group treated with the complex extract was evaluated by an index.

HEAT DDT1 DDT2 DDT3 DDT4 transwell assay 5 4 8 9 4

As shown in Table 3 above, compared to the HEAT treatment group, the DDT1 treatment group had little inhibitory effect, but it was confirmed that the DDT2 and DDT3 treatment groups exhibited significant repellent effects. Preparation of composite composition comprising EAM-EAT-MEMC

Using the same method as the method for preparing the ethanol extract of hwanggi and ginseng of ethanol extract, ethanol extract (KAM) and ethanol extract of suchocho (PAM) are prepared, and an anticancer composition is prepared in the weight range as shown in Table 4 below. Did.

DDM1 DDM2 DDM3 DDM4 EAM-EAT-MEMC 100 100 100 100 KAM One 5 10 15 PAM One 5 10 15

(Unit parts by weight) Functionality experiment

After preparing the composite compositions of DDM1 to DDM4 as teas, preference was given to 15 adult males and females.

The sensory evaluation was evaluated by an index of 1 to 10 according to palatability based on aroma and taste, and classified as an average index. The results are shown in Table 5.

On the other hand, the index below is the higher the number, the higher the preference.

DDT3 DDM1 DDM2 DDM3 DDM4 incense 3 5 8 9 5 flavor One 4 7 9 4 Palatability 2 4 8 9 5

As shown in Table 5 above, in the case of DDT3, it was confirmed that the extract and the suchocho extract were not included, so that the aroma, taste, and preference were low.

Looking in light of the results of Table 5, when using the ginseng fern extract and suchocho extract within the scope of the present invention, such as DDM 2 and 3, the flavor, stubborn taste and bitter taste are neutralized, depending on the mutual combination of each extract It was confirmed that the flavor and taste of tea beverages are greatly enhanced.

As a result, when only the extracts of hwanggi, ginseng and baekpi skin were used as a health functional food, there was a problem that the preference was lower than that of the anticancer effect. As a result, it can be widely used as a functional food with a higher palatability or as a functional tea.

Although the preferred embodiments of the present invention have been described in detail above, the scope of the present invention is not limited thereto, and various modifications and improvements of those skilled in the art using the basic concept of the present invention defined in the following claims are also provided. It belongs to the scope of rights.

Claims (7)

Herbal extract selected from the group consisting of Astragalus mongholicus Bunge, Adenophoratriphylla var.
Anticancer composition. According to claim 1,
The extract is to extract water, C1 to C4 alcohol, ether, chloroform, ethyl acetate, methylene chloride or a mixture thereof as a solvent.
Anticancer composition. According to claim 1,
The fraction is to fractionate any one selected from the group consisting of hexane, chloroform, ethyl acetate, and butanol from herbal extracts selected from the group consisting of astragalus, ginseng, acupuncture and mixtures thereof.
Anticancer composition. According to claim 1,
The anti-cancer composition is excellent in the prevention or treatment effect of any one or more cancers selected from the group consisting of prostate cancer, uterine cancer, liver cancer, rectal cancer, lung cancer, breast cancer, and ovarian cancer.
Anticancer composition. Claim 1 to claim 5 comprising the composition for anticancer according to any one of
Pharmaceutical composition. Claim 1 to claim 5 comprising the composition for anti-cancer according to any one of
Pharmaceutical composition for prevention and treatment of lung cancer. Claim 1 to claim 5 comprising the composition for anti-cancer according to any one of
Functional food composition.

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