KR20090091400A - A composition for anti-cancer containing agrocybe chaxingu extracts - Google Patents

Abstract

A composition containing an Agrocybe chaxingu Huang which has anticancer activity is provided to have the effect of suppressing cell lines of lung cancer, colon cancer, prostate cancer, and cervical cancer. A composition for anticancner comprises Agrocybe chaxingu Huang extract which is extracted at high pressure and with hot water. The Agrocybe chaxingu Huang extract is a hexane fraction, dichloromethane fraction or ethylacetate fraction using hexane, dichloromethane, ethylacetate, and butanol. The composition has suppression effect to solid cancer. The solid cancer is lung cancer, colon cancer, cervical cancer or prostate cancer.

Description

A composition for anti-cancer containing chashingo mushroom extract {A composition for anti-cancer containing Agrocybe chaxingu extracts}

The present invention relates to an anticancer composition comprising an extract of chashingo mushroom, and more particularly, to an anticancer composition comprising a high pressure hydrothermal extract or methanol extract having anticancer activity obtained from chashingo mushroom fruiting body.

In addition to the recent rapid industrialization, environmental pollution has significantly increased the incidence of harmful oxygen-related refractory diseases with the development of new diseases, and these refractory diseases require continuous and long-term treatment after the onset, resulting in reduced economic activity of these patients. The loss is huge and the cost of treatment is up to trillions of dollars annually worldwide.

Mushroom has long been used as a health supplement to prevent diseases as well as to cure diseases, and it has been proved through various studies that it has an immune enhancing, anti-cancer and anti-aging effect.

Studies on the functionality of mushrooms have been focused on glycoprotein components in mushrooms (Pak et al. 1998, Korean J. Food Sci.Technol, 30 (5): 1236-1242; Jin et al. 1999, Journal of Pharmacy, 43 (5): 635-641; Jung et al. 2002, Korean Patent 10-0340663; Cho et al. 2004, Korean Journal of Microbiology, 40 (3): 217-220). It has been reported to have various anticancer and antioxidant activities (Lin, JY and Chou, TB 1984, J. Biochem. 6: 35-40; Kawagishi, H. et . Al ., 1990, Biochim. Biophys. Acta, 1034: 247 -252; Yu, LG et . Al ., 1993, Cancer Res. 53: 4627-4632; Ye, SF et . Al ., 2003, Life Sciences. 74: 593-602), proteins having anticancer activity from various mushrooms Polysaccharides have been identified, especially Retinan extracted from shiitake mushrooms, Schizophyllan from squirrel mushrooms and Crestine (PSK) from cloud mushrooms. It is used in clinical trials as an anticancer adjuvant (Zhu D. 1987, Abstr. Chin. Med. 1: 251-286; Hayakawa K. et . al ., 1993, Anticancer Res. 13: 1815-1820; Nakajato H. et . al ., 1994, Cancer. Lancet 343: 1122-1226; Hobbs C. 1995, Fenichel RL, Chirigos MA (eds) Immunomodulation agents and their mechanisms. Dekker, New York, 409-436).

Agrocybe chaxingu Huang is a wood fungus that occurs in the tea tree, which belongs to the folds of the fungi, dung mushroom, and Agrocybe , which help diuresis, spleen and stomach, and brighten the eyes. It is known to have high medicinal value.

Chashingo mushroom is Agrocybe Among the mushrooms belonging to the genus, Agrocybe aegerita is known to be relatively close genetically (Gonzalez, P. and J. Labarre, 1998, Appl.Environ.Microbiol., 64 (11): 4149-4160) The functionality of mushrooms has been reported several times (Hyun, JW et . Al ., 1996, Arch. Pharm. Res., 19 (3): 207-212; Kim, BK et . Al ., 1997, Arch. Pharm Res., 20 (2): 128-137; Kim et al., 2000, Korean Journal of Mycology, 36 (1): 64-68; Kim et al., Korean Journal of Mycology, 30 (2): 124-130) There was no report on the functionality of the mushrooms.

Japanese Patent Application Laid-Open No. 2006-025725 (February 02, 2006) "Multipurpose Japanese Sauce and Manufacturing Method thereof" discloses a Japanese sauce made from chashin mushroom extract, Japanese soy sauce, kelp, bonito, and the like. In addition, Chinese Patent Publication No. 1349767 (2002.05.22.) "Flavour Chicken Recipe Using Shingo Mushroom" discloses the use of Chashingo mushroom for the purpose of adding flavor to chicken dishes.

As such, chashingo mushroom has been used only for cooking or for some folk remedies, but its anticancer efficacy is unknown.

It is an object of the present invention to provide a composition for anticancer derived from natural products that is safe for human body and inhibits the growth of cancer cells.

In order to achieve the above object, the present inventors have completed the present invention by obtaining an anticancer activity of known carcinogenic mushroom extracts and then testing the anticancer activity of the extract of carcinogenic mushrooms against cancer cell lines to show excellent anticancer activity. Reached.

The present invention provides an anticancer composition comprising an extract of chashingo mushroom, wherein the extract of high-pressure hot water extract of Chashingo mushroom fruit body exhibits high growth inhibitory activity against lung cancer, colorectal cancer, prostate cancer and cervical cancer cell lines, and methanol extract It was confirmed that all fractions except the aqueous layer of the organic solvent fractions fractionated by polarity also showed excellent anticancer activity.

Therefore, chashingo mushroom extract of the present invention may be used as an active ingredient of anti-cancer treatment.

In order to achieve the above object, the present invention provides an anticancer composition comprising the extract of tea Shingo mushroom.

Chashingo mushroom extract in the above is preferably extracted from the fruiting body.

The anticancer composition may include an extract extracted from Chashingo mushroom with high-pressure hot water or 0.001 ~ 10% (w / v) of the fraction obtained by extracting Chashingo mushroom with methanol and then again fractionated with hexane, dichloromethane and ethyl acetate. It is desirable to.

Hereinafter, the present invention will be described in detail.

The present invention relates to a composition for anticancer comprising chashingo mushroom extract.

The inventors of the present invention examined the anticancer effect on the cancer cells cultured with respect to the extract of chashingogi fruit body with hot water, organic solvents, etc., it was confirmed that the anticancer activity. Natural extract derived from shishingo mushroom is widely used in cooking and is used for folk remedies.

The term "anticancer" in the present invention means the ability to inhibit or kill the proliferation of cancer cells. When administration of chashingo mushroom extract was confirmed that cell growth is inhibited and cell death increased in cancer cell lines.

Cancers that can be prevented or treated with the compositions of the present invention are solid cancers. Solid cancers include, but are not limited to, colorectal cancer, prostate cancer, small intestine cancer, rectal cancer, anal cancer, esophageal cancer, pancreatic cancer, stomach cancer, kidney cancer, cervical cancer, breast cancer, lung cancer, ovarian cancer, and the like. In particular, it is effective in the treatment of cervical cancer, lung cancer, prostate cancer and colon cancer.

The composition may comprise a pharmaceutically acceptable carrier and be formulated for human or animal use.

Pharmaceutical compositions for oral administration include, for example, capsules or tablets, powders or granules, solvents, syrups or suspensions.

Suitable excipients for tablets or hard gelatin capsules are lactose, corn starch or derivatives thereof, stearic acid or salts thereof. Suitable excipients for use in soft gelatin capsules are, for example, vegetable oils, waxes, fats, semi-solids or liquid polyols. Excipients that can be used to prepare solvents and syrups are, for example, water, polyols and sugars. Oils, such as vegetable oils, can be used to prepare suspensions to provide oil-in-water or water-in-oil suspensions. Pharmaceutical compositions for transdermal administration may be presented as separate patches to enable intimate contact with the epidermis of the recipient for a long time.

Pharmaceutical compositions for parenteral administration include aqueous and non-aqueous sterile injectable solutions which may contain antioxidants, buffers, bacteriostats and solutes that are substantially isotonic with the blood of the recipient; And aqueous and non-aqueous sterile suspending agents which may include suspending agents and thickening agents. Excipients that can be used in the preparation of injectables include, for example, water, alcohols, polyols, glycerin and vegetable oils. Such compositions may be stored in unit-dose (single) or multi-dose (several) containers, such as sealed ampoules and vials, with the addition of a sterile liquid carrier, such as water for injection, just prior to use. Can be stored under freeze-drying conditions for use.

Instant injection solutions and suspensions can be prepared from sterile powders, granules and tablets.

The route of administration of the compositions of the invention can be administered via a common route that can reach the target tissue. Intraperitoneal administration, intravenous administration, intramuscular administration, subcutaneous administration, intradermal administration, oral administration, topical administration, intranasal administration, pulmonary administration, rectal administration, but is not limited to the above listed methods of administration. Pharmaceutical compositions can also be administered by any device capable of moving the active substance to the target cell.

The composition of the present invention is administered in a pharmaceutically effective amount. As used herein, "pharmaceutically effective amount" means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment, and the effective dose level refers to the type and severity, severity, age, sex, and drug of the patient. Can be determined according to the activity of the drug, the sensitivity to the drug, the time of administration, the route of administration and the rate of release, the duration of treatment, factors including the drug used concurrently and other factors well known in the medical field. The compositions of the present invention may be administered in a single or multiple doses. In consideration of all the above factors, it is important to administer an amount that can obtain the maximum effect in a minimum amount without side effects, and a pharmaceutically effective amount can be easily determined by those skilled in the art.

Pharmaceutical formulations comprising the composition of the present invention can be applied orally, parenterally, ie, subcutaneously, intramuscularly, or topically, as desired. The dosage is a daily dosage of 0.001 μg˜10 mg / kg. It may be administered in several divided doses.

The present invention relates to a composition having anticancer activity extracted from Chashingo mushroom, Chashingo mushroom extract has excellent anticancer activity and can be usefully used as a preventive and therapeutic agent for various solid cancers such as lung cancer, colorectal cancer, prostate cancer and cervical cancer. .

The anticancer composition of the present invention includes chashingo mushroom extract, which is obtained by extracting and extracting chashingo mushroom with one or more selected solvents from methanol, hexane, dichloromethane, ethyl acetate, butanol and water.

The present invention to obtain the extract by extracting the fruiting body part of chashingo mushroom under high pressure, hot water or methanol, hexane, dichloromethane, ethyl acetate, butanol and water and lung cancer, colon of the extract Identifying the anticancer effects on four cell lines: cancer, prostate cancer, cervical cancer.

Chashingo mushroom high-pressure hot water extract of the present invention is obtained by drying or crushing Chashingo mushroom fruiting bodies in the dry or dried at 40-80 ℃ for 12-72 hours, concentrated by depressurizing and lyophilizing the extract extracted under hot water under lung cancer. It has excellent anticancer activity against colorectal cancer, prostate cancer and cervical cancer.

In addition, chashingo mushroom organic solvent extract of the present invention is dried or pulverized Chashingo mushroom fruiting body or dried for 12-72 hours at 40 ~ 80 ℃, extracted with stirring by adding methanol to extract the concentrated under reduced pressure and freeze-dried Extract obtained by re-fractionation with hexane, dichloromethane, ethyl acetate, butanol and water, shows excellent anticancer activity against lung cancer, colorectal cancer, prostate cancer.

For example, methanol extraction of chashingo mushroom is preferably carried out by dipping in 1-2 liters of methanol (methanol) per 100g of dried sample of chashingo mushroom.

When extracting the tea Shingo mushroom in the above 80 to 120rpm shaker 20 to 30 times it is preferable to extract repeatedly 2-3 times. In order to remove the residue of the methanol fraction obtained by repeated extraction in the above, it is passed through a funnel with a filter paper and concentrated completely using a vacuum concentrator.

It is preferable that the concentrate is dissolved well with 50-100 ml of distilled water and then dried using a lyophilizer.

In the composition of the present invention, the freeze-dried tea Shingo mushroom methanol extract was sequentially partitioned into hexane, dichloromethane, ethyl acetate and butanol according to the polarity of the solvent, and the remaining residue was partitioned into an aqueous layer, and each fraction was concentrated and frozen. After use, it is preferable to dissolve it in DMSO (Dimethyl Sulfoxide) while using it in the freezer after drying.

According to a preferred embodiment of the present invention, the high-pressure hot water extract of Chashingo mushroom exhibits a high growth inhibitory effect of 46.1-50.8% against four types of cancer cell lines. Cancer cell line growth inhibitory activity was shown.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, these examples are intended to describe the present invention in more detail, and the scope of the present invention is not limited by the description of the embodiments.

< Example  1> Chashingo Mushroom  Methanol extract manufacturer

100 g of dried fruiting body of Chashingo mushroom was placed in a 5 l Erlenmeyer flask containing 2 l of methanol (methanol) and extracted twice with a 100 rpm shaker for 24 hours. In order to remove the residue of the methanol fraction, the filter paper was passed through a funnel, completely concentrated using a vacuum condenser, dissolved in 50 ml of distilled water, and then dried using a lyophilizer.

< Example  2> Chashingo Mushroom  Of methanol extract Fractional layer  Produce

10 g of the methanol extract of chashingo mushroom of Example 1 was dissolved in 100 ml of distilled water, placed in a silica gel column, and fractionated using a solvent mixed with hexane and water at 7: 3 to form a hexane layer. Using this solvent fractionation method, five fractions such as hexane layer, dichloromethane layer, ethyl acetate layer, butanol layer, and water layer were obtained, and each fraction was concentrated in vacuo to extract hexane, dichloromethane, and ethyl acetate. , Butanol extract and the aqueous layer was prepared.

< Example  3> Chashingo Mushroom  High pressure Hydrothermal  Extract Preparation Method

2 liters of distilled water was added to 100 g of the dried fruiting body of Chashingo mushroom, followed by extraction at 121 ° C. for 2 to 4 hours, and the high-pressure hot water extract was centrifuged at 6,000 rpm for 30 to 60 minutes. After centrifugation, 2-4 times the volume of ethanol was added to the supernatant and allowed to stand at 4 ° C. for 24 to 48 hours, followed by centrifugation at 6,000 rpm for 40 minutes to recover the pellet layer. The recovered pellet layer was sufficiently dissolved in distilled water, dried using a lyophilizer, and used while storing the lyophilized high pressure hot water extract sample under refrigerated dark conditions.

< Example  4> Chashingo Mushroom  Antitumor Activity of Extracts

The solvent-specific fractions of the Chashingo mushroom methanol extract of Example 2 and the Chashingo mushroom high-pressure hot water extract of Example 3 were used as anticancer activity assay samples for cancer cell lines, respectively.

Culture of Cancer Cell Lines

Four types of cancer cell lines, including prostate cancer cell line (DU145), cervical cancer cell line (HeLa), colorectal cancer cell line (HT29), and lung cancer cell line (H1299), were distributed from the American Type Culture Collection (ATCC), and each cancer cell line was 100 mm. The dishes were incubated in a 37 ° C., 5% CO ₂ incubator. The medium for culturing each cell line was DMEM (Dulbeco's Modified Eagle's Medium) medium added with 2 mg / ml sodium bicarbonate, 10% Fetal Bovine Serum and 1% penicillin-streptomycin.

Cancer cell line  Growth inhibitory activity assay

Each cultured cancer cell line was incubated in a 24-well plate and treated with 0.1 mg / ml and 1 mg / ml concentrations of high-pressure hot water extracts of methanol and extracts of methanol extract, respectively, at 37 ° C and 5%. Incubated for 48 hours in a CO ₂ incubator. After incubation with 20 μl of MTT [3- (4,5-dimethylthiazol-2-yl) -2,5, -diphenyltetrazolium bromide] reagent and incubated for 4 hours, the absorbance was measured at 490 nm of ELISA reader. Compared with the absorbance of the control group, the cell proliferation rate was converted into percentage.

< Experimental Example  1> Chashingo Mushroom  High pressure Hydrothermal  Anticancer Activity of Extracts

Figure 1 shows the results of growth inhibitory activity against four cancer cell lines by concentrations of the high-pressure hot water extract of tea Shingo mushroom by the method of Example 4.

When treated with 0.1% (w / v) hot water extract of Chashingo mushroom, 40.8% growth of cervical cancer cell line (HeLa), 45.9% growth of lung cancer cell line (H1299), and 46.1 growth of colon cancer cell line (HT29) %, 50.8% inhibited the growth of prostate cancer cell line (DU145).

< Experimental Example  2> Chashingo Mushroom  Organic solvent Fraction  Anticancer activity

Table 1 shows the survival rate of the cancer cell line after treatment with the fractions of the four different organic carcinoma organic solvent in a concentration of 0.01% (w / v) in four cancer cell lines in the same manner as in Experimental Example 1.

 Cancer cell line  Cancer Cell Line Survival Rate by Type of Fraction (%) No treatment Dichloromethane layer Ethyl acetate layer Hexane layer Butanol layer Water layer  HeLa  100  104.8  121.5  121.3  100.3  113.1  H1299  100  80.9  76.2  67.4  97.4  103.4  HT29  100  71.9  75.3  60.4  89.4  83.4  DU145  100  82.2  71.8  57.1  114.1  85.9

As shown in Table 1, when ethyl acetate layer was treated with cancer cell lines at 0.01% (w / v) concentration, growth of H1299 cancer cell line, HT29 cancer cell line, and DU145 cancer cell line was inhibited by 24.7%, 24.7% and 28.2%, respectively. The layer inhibited the growth of H1299 cancer cell line, HT29 cancer cell line, and DU145 cancer cell line by 32.6%, 39.6% and 42.9%, respectively, when treated at 0.01% (w / v).

Table 2 shows the survival rate of the cancer cell line when the fractions of the organic solvents of Chashingo mushroom were treated with 0.1% (w / v) in four cancer cell lines.

 Cancer cell line  Cancer Cell Line Survival Rate by Type of Fraction (%)  No treatment  Dichloromethane layer  Ethyl acetate layer  Hexane layer  Butanol layer  Water layer  HeLa  100  76.0  78.1  82.1  73.9  98.4  H1299  100  64.5  49.4  59.1  59.5  109.4  HT29  100  54.9  69.6  62.2  67.2  90.5  DU145  100  42.1  65.3  68.2  61.7  86.4

All fractions except the aqueous layer had growth inhibition of cancer cell lines. Especially, when dichloromethane layer was treated with 0.1% (w / v) concentration, growth of H1299 cancer cell line, HT29 cancer cell line and DU145 cancer cell line was 35.5%, respectively. 45.1%, 57.9% inhibited the high activity was confirmed.

Figure 1 shows the anticancer activity of the high-pressure hot water extract of tea Shingo mushrooms by concentration.

Claims (6)

Anticancer composition comprising an extract of high-pressure hot water extracted mushrooms. To the methanol extract prepared by adding methanol to Chashingo mushroom, hexane, dichloromethane, ethyl acetate and butanol were added sequentially. Anticancer composition. The method according to claim 1 or 2, The anticancer composition is an anticancer composition, characterized in that exhibits an inhibitory effect on solid cancer. The method of claim 3, The solid term is anticancer composition, characterized in that one of lung cancer, colorectal cancer, cervical cancer and prostate cancer. The method according to claim 1 or 2, The chashingo mushroom extract is anticancer composition, characterized in that Chashingo mushroom fruiting body extract. The method according to claim 1 or 2, The tea Shingo mushroom extract is contained 0.001 ~ 10% (w / v), anticancer composition, characterized in that it comprises a pharmaceutically possible carrier.

Images