KR20050079789A - Anti-cancer composition containing mushroom cultivated in artemisa iwaomogi - Google Patents

Abstract

 The present invention relates to an anticancer composition comprising an extract of mushrooms cultured in Injin mugwort. In particular, the anticancer composition of the present invention is not only harmless to the human body but also exhibits significant anticancer activity against cervical cancer, colorectal cancer and leukemia cells, and thus can be used for the purpose of preventing and treating the same.

Description

Anticancer composition comprising extracts of mushrooms grown in Injin mugwort {ANTI-CANCER COMPOSITION CONTAINING MUSHROOM CULTIVATED IN ARTEMISA IWAOMOGI}

[TECHNICAL FIELD OF THE INVENTION]

The present invention relates to an anticancer composition comprising an extract of mushrooms cultured in Injin mugwort, and more particularly, to a locust fungus mushroom extract cultured in Injin mugwort which exhibits very good anticancer activity against rectal cancer, cervical cancer and leukemia cells.

[Private Technology]

Recently, functional dietary supplements are developing rapidly with the improvement of national income, and the demand for high quality functional dietary supplements will increase further due to social changes such as changes in dietary life, diversification of preferences, and pursuit of leisurely living. It is expected. In particular, the functional health drink sector is rapidly developing in Korea, and with the change of dietary habits, the demand level of consumers' preferences and quality is getting higher and higher in recent years. We are developing and developing new functional health drinks and functional health supplements.

Many food and pharmaceutical companies ship functional health drinks and functional supplements to the market in the form of simply extracting and mixing ingredients rich in bioactive substances. However, functional health drinks and functional dry food supplements contain various bioactive substances, while at the same time, the high activity of the bioactive substances is very important in terms of functionality. Therefore, studies on how to maximize the activity of physiologically active substances rather than a simple mixing method of medicinal foods are required.

On the other hand, while mushrooms are rich in carbohydrates, proteins, inorganic salts and vitamins, they are low in lipids and calories, and contain many pharmacologically active ingredients. In particular, the herbaceous fungus (hericicum erinaceus) is a biological resource for the development of new medicines and dietary supplements, attention is focused. However, so far, only the level of bioactive substances are separated from the scab, and studies on maximizing the metabolite activity of the scab are inadequate.

In order to solve the problems of the prior art, an object of the present invention is to provide an extract of the mushroom cultured by the mushroom culture method for enhancing the anticancer activity of the mushroom.

It is another object of the present invention to provide an anticancer composition having an excellent anticancer effect on rectal cancer, cervical cancer and leukemia.

In order to achieve the above object, the present invention provides an anticancer composition comprising an extract of mushrooms cultured in Injin mugwort.

Hereinafter, the present invention will be described in detail.

The inventors of the present invention, while studying methods for enhancing the anticancer activity of the mushroom, confirmed that the anticancer activity can be significantly increased by cultivating the mushrooms in Injin mugwort, thereby completing the present invention.

The present invention relates to an anticancer composition comprising the extract of the mushrooms cultured in Injin mugwort as an active ingredient.

Mushrooms cultured in jinjinjin in accordance with the present invention may be a common mushroom, for example, beetle mushrooms, situation mushrooms, ganoderma lucidum mushrooms, agaricus and matsutake mushrooms. Preferred is the roe deer mushroom. The mushroom may be a fruiting body and may be a mycelium or a mycelium liquid culture.

The mushroom cultured in the jinjin mugwort can be obtained by incubating the mushroom spawn in a common mushroom liquid medium and then inoculating and culturing it in jinjin wormwood. In the case of culturing roe deer mushrooms in Injin mugwort may be carried out by the method described in the Republic of Korea Patent Publication No. 2002-72878, briefly described as follows. However, specific culture methods and culture conditions can be easily changed by those skilled in the art.

Roe deer beetle cultured in Injin mugwort

(a) obtaining a seed of the roe deer mushroom;

(b) preparing a solid medium comprising phosphorus mugwort; And

(C) it can be prepared by the method comprising the step of inoculating and incubating the scavenger mushroom seedling in the solid medium to obtain a mycelium.

It may also further comprise the step of obtaining a fruiting body by further culturing the mycelium.

In the step (a), the seedling of the roe deer fungus is inoculated into the cultivation of the roe deer fungi, and cultured for 5 to 8 days at 23 to 26 ℃, pH 5 to 6, stirring speed 100 to 140 rpm It can be obtained by. At this time, the culture temperature, the pH of the medium, the stirring speed and the incubation period set in the above range is the maximum to obtain the worm mushroom mycelium, the liquid culture medium for the spawn YMPG, MCM, PDA or Soybean meal can be used.

In the step (b), the solid medium is prepared by sterilizing after adding water to the jinjinjin. Water may be added in a weight ratio of 1: 1 to 10 with respect to phosphorus mugwort, but is not limited thereto. The sterilization method can be carried out by a known method, preferably high temperature sterilization method.

In the step (c), seed spawn obtained in step (a) is inoculated in a solid medium at 0.1 to 30% by weight, and cultured at a culture temperature of 23 to 26 ° C. and a culture period of 40 to 50 days. Mycelia can be obtained by the primary growth as described above, the mycelium can be further grown in culture for 15 to 20 ℃, the secondary growth for 10-15 days in the culture period to further culture the fruiting body.

The method of cultivating roe deer mushroom in the above-mentioned jinjin wormwood is only one example and is not limited to the above. In addition, mushrooms, such as situation mushrooms, ganoderma lucidum mushrooms, agaricus or matsutake mushrooms, in addition to roe deer mushrooms, can also be cultured in jinjinjin by changing some of the above methods. Since the change is caused by the culture conditions vary depending on the type of mushroom, it can be easily implemented by those skilled in the art.

Mushrooms cultured in jinjin jinjin of the present invention can be prepared by extracting the mushrooms with water or an organic solvent. For example, the mushroom extract may be prepared by immersing the mushroom in one or more extracting solvents to obtain or fractionate the supernatant, and then may be powdered through spray drying or lyophilization. As the organic solvent, alcohols of 1 to 5 carbon atoms, hexane, chloroform or ethyl acetate may be used, and as alcohols, ethanol, methanol, butanol, propanol or isopropanol may be used. The alcohol may be used as a 100% stock solution, but 90 to 50% alcohol may be used. However, the extraction solvent is not limited to the solvents described above, and any known solvent can be used. The preferred extraction method is to immerse the mushroom in an extract solvent of 1: 1 to 20 by weight, and then filter it, and then depressurize, concentrate and spray-dry or lyophilize.

Extracts of mushrooms cultured from Injin mugwort according to the present invention exhibit anticancer activity against various kinds of cancers, in particular, extracts of Roeden beetles cultured from Injin mugwort exhibit very good anticancer activity against rectal cancer, cervical cancer and leukemia cells. The anticancer activity of the rot fungi cultured in the above-mentioned ginseng mugwort is significantly enhanced compared to the anticancer activity identified in ginseng worm or gingicolor fungi, and the fungus metabolizes ginseng worms to produce a bioactive substance with enhanced anticancer activity It can be seen.

The anticancer composition of the present invention may further include a carrier or an excipient which can be used pharmacologically according to the formulation and the method of use, in addition to the extract of the mushrooms cultured in the above-mentioned jinjin Mugwort. In this case, the content of the active ingredient in the anticancer composition may be 0.001 to 99% by weight. If the amount of the active ingredient in the composition is less than 0.001% by weight may require a large amount of administration for effective efficacy, if the amount exceeds 99% by weight may be constant compared to the amount may be uneconomical. However, it is preferable to properly adjust the content of the active ingredient according to the method of use and purpose of use of the anticancer composition.

The pharmacologically acceptable carrier or excipient may use a conventional material depending on the dosage form, and when formulated, fillers, extenders, wetting agents, disintegrating agents or surfactants may be used. Representative diluents or excipients include water, dextrin, calcium carbonate, lactose, propylene glycol, liquid paratin and physiological saline.

The anticancer composition may be administered orally or parenterally, and its formulation may vary depending on the method of use, so it is not limited to the following. Examples of formulations include PLASTERS, GRANULES, LOTIONS, POWDERS, SYRUPS, LIQUIDS AND SOLUTIONS, AEROSOLS, OINTMENTS, FLOWERS FLUIDEXTRACTS, EMULSIONS, SUSPENSIONS, INFUSIONS, TABLETS, INJECTIONS, CAPSULES and PILLS.

The dosage or amount of the anticancer composition should be determined in consideration of the form of administration, the age, sex, condition of the patient, the absorption of the active ingredient in the body, the inactivation rate, and the concomitant drug. And 0.001 mg / kg body weight to 500 mg / kg body weight.

The anticancer composition according to the present invention can be used as a food, a food additive, a beverage or a medicament, and can be easily utilized as a functional food or a beverage.

Hereinafter, examples of the present invention will be described. The following examples are only for illustrating the present invention and the present invention is not limited to the following examples.

Example 1 Preparation of Extract of Roebuck Mushroom

YMPG (KH ₂ PO ₄ 2.0 g / L, MgSO ₄ · 7H ₂ O 1.0 g / L, glucose 10.0 g / L, thiamine-HCl 1.0 g / L, adjusted to pH 5.5 for Hericium erinaceus fungi DL-asparagine 1.0 g / L, peptone 2.0 g / L, malt extract 10.0 g / L, yeast extract 2.0 g / L, agar 20.0 g / L) or MCM (K ₂ HPO ₄ 1.0 g / L, KH ₂ PO ₄ 0.46 g / L, MgSO ₄ .7H ₂ O 0.5 g / L, glucose 20.0 g / L, peptone 2.0 g / L, yeast extract 2.0 g / L, agar 20.0 g / L) The inoculum was incubated by shaking culture for 7 days at 120 rpm with agitation speed.

Water corresponding to twice the total weight of the phosphorus mugwort was added to the mugwort, which was sterilized at 120 ° C. for 2-4 hours to prepare a solid medium.

The inoculum was inoculated at 10% by weight based on the weight of the solid medium, and mycelia were first grown at 25 ° C. for 40 to 50 days. Thereafter, fruiting bodies were grown by performing secondary growth at a culture temperature of 15 to 20 ° C. for 10 to 15 days.

The mycelium obtained above was dried at room temperature for 24 hours, and then 3 L of 30% ethanol was added to 145 g of the dried product and extracted at room temperature for 2 weeks. The ethanol extract was filtered to give an ethanol extract, which was concentrated at 80 ° C. and then lyophilized to give 20.5 g of dried product. Hereinafter, the ethanol extract, that is, lyophilisate is named "Arinacol".

Example 2: Roebuck Mushroom Extract

After crushing 75 g of roe deer mushroom dried product, 3 L of 30% ethanol was added and extracted at room temperature for 2 weeks. The extract was filtered to give the filtrate, which was concentrated at 80 ° C. and lyophilized to give 41.9 g of dried product.

Example 3: Injin mugwort extract

After crushing 145 g of Artemisa iwayomogi dry matter finely, 3 L of 30% ethanol was added and extracted at room temperature for 2 weeks. The extract was filtered to give the filtrate, which was concentrated at 80 ° C. and lyophilized to give 19 g of dry matter.

Example 4: Agaricus Mushroom Extract

145 g of agaricus dry matter was finely crushed, and 3 L of 30% ethanol was added thereto, followed by extraction at room temperature for 2 weeks. The extract was filtered to give the filtrate, which was concentrated at 80 ° C. and lyophilized to give 78.3 g of dry matter.

Experimental Example: Cancer Cell Proliferation Inhibition Experiment

MTT (3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyl tetrazolium bromide) assay was used to investigate the effects of cancer cell proliferation.

1. Cancer cell line

This experiment was carried out using a total of six cancer cell lines. Where to buy each cancer cell, culture medium and culture conditions are as follows.

HCT116 cell line: Cell line derived from human rectal carcinoma, which was distributed by Rockville MD. RPMI 1640 was used as a culture medium, and cultured in a 37 ° C., 5% CO ₂ incubator.

-Jurkat cell line: T cell leukemia (T lymphocyte) -derived cell line. Cells were distributed by Rockville MD. RPMI 1640 was used as a culture medium, and cultured in a 37 ° C., 5% CO ₂ incubator.

HepG2 cell line: Hepatocarcinoma-derived cell line. Cells were distributed by Rockville MD. RPMI 1640 was used as a culture medium, and cultured in a 37 ° C., 5% CO ₂ incubator.

-CaSki cell line: A cervix carcinoma-derived cell line, which was distributed by Rockville MD. RPMI 1640 was used as a culture medium, and cultured in a 37 ° C., 5% CO ₂ incubator.

-SK-HEP-1 cell line: Human liver cancer (Hepatocarcinoma) cell line, cells were distributed by Rockville MD. RPMI 1640 was used as a culture medium, and cultured in a 37 ° C., 5% CO ₂ incubator.

SK-MES-1 cell line: Human lung cancer-derived cell line, cells were distributed by Rockville MD. RPMI 1640 was used as a culture medium, and cultured in a 37 ° C., 5% CO ₂ incubator.

2. Sample Preparation

Each of the Arinacol, locust beetle extract, erythmus extract and Agaricus extract of Examples 1 to 4 was dissolved in distilled water at 1 mg / ml, and then serially diluted by 1/5, and 1000 µg / ml, 200 µg / ml, and 40 µg. / Ml, 8 µg / ml, 1.6 µg / ml, 0.32 µg / ml, 0.064 µg / ml, 0.0128 µg / ml and 0 µg / ml. In addition, doxorubicin, which is used as an anticancer agent as a positive control group, was dissolved in distilled water, respectively, and 5 μg / ml, 1 μg / ml, 0.2 μg / ml, 0.04 μg / ml, 0.008 μg / ml, 0.0016 μg / ml, and 0.00032 μg / ml, respectively. , 0.000064 μg / ml and 0 μg / ml.

3. MTT Analysis

 The MTT assay is a method of measuring viable cell concentration by measuring the amount of dark blue formazan produced by the dehydrogenase enzyme present in mitochondria of living cells by reducing MTT.

Each of the six cancer cell lines in culture was adjusted to a concentration of 3 × 10 ³ cells / ml and placed in 100 μl per well in a 96-well microplate (Falcon, USA). Here, the samples prepared above were added to each well by concentration and incubated for 48 hours at 37 ℃, 5% CO ₂ incubator. After treatment with 20 μl of PBS containing 5 mg / ml of MTT reagent, the cancer cell lines were incubated for 4 hours, 100 μl of the supernatant was removed, and then 100 μl of 0.04 N HCl-isopropanol was further incubated overnight. Then, the absorbance was measured at 560 nm using an automatic ELIZA reader, and cell viability was calculated according to Equation 1.

(Equation 1)

% Inhibition of cancer cell proliferation = absorbance of sample / absorbance of control x 100

The results are shown in the graph of Figures 1-6. Figure 1 shows the proliferation inhibition rate for the rectal cancer cell line HCT116, Figure 2 shows the proliferation inhibition rate for CaSki, cervical cancer cell line, Figure 3 shows the growth inhibition rate for Jurkat, leukemia cell line, Figure 4 is liver cancer It shows the growth inhibition rate for the cell line HepG2, Figure 5 shows the growth inhibition rate for liver cancer cell line SK-HEP-1, Figure 6 shows the growth inhibition rate for lung cancer cell line SK-MES-1.

In addition, the effective concentration (EC ₅₀ ) of the sample at which the average inhibition rate of the cancer cells is 50% was obtained and shown in Table 1 below.

EC ₅₀ Arina Cole Roe deer Injin mugwort Agaricus Doxorubicin HepG2 > 1000 > 1000 400 > 1000 2.89 CaSki 334 > 1000 378 > 1000 1.41 SK-HEP-1 > 1000 > 1000 490.6 > 1000 0.154 Jurkat 364.9 > 1000 453.18 185.1 0.07 SK-mes-1 > 1000 > 1000 904 > 1000 0.15 HCT116 66.82 896 167.7 241.2 0.582

As a result of the experiment, the extract of Roeden beetle showed very low anticancer activity, and the Agaricus extract showed excellent anticancer activity against leukemia cells and rectal cancer cells, and Injin mugwort extract showed a certain level of anticancer activity against all six cancer cells. Indicated. Arinacol showed very high anticancer activity in colorectal cancer cells and excellent anticancer activity in cervical cancer cells and leukemia cells. Particularly, Arinacol was found to have a very good anticancer activity against rectal cancer cells compared to the extracts of Roebendung and Roots of the Roots. Could know.

As described above, the extract of the mushroom cultured in Injin mugwort of the present invention is not only harmless to the human body but also shows significant anti-cancer activity against cancer cells, especially cervical cancer, rectal cancer and leukemia cells, and thus can be used for the purpose of preventing and treating the same. Can be.

1 is a graph showing the proliferation inhibition rate for HCT116, a rectal cancer cell line.

Figure 2 is a graph showing the proliferation inhibition rate for CaSki, a cervical cancer cell line.

Figure 3 is a graph showing the proliferation inhibition rate for the leukemia cell line Jurkat.

Figure 4 is a graph showing the proliferation inhibition rate for HepG2, a liver cancer cell line.

Figure 5 is a graph showing the growth inhibition rate for liver cancer cell line SK-HEP-1.

Figure 6 is a graph showing the proliferation inhibition rate for lung cancer cell line SK-MES-1.

Claims (7)

Anticancer composition comprising the extract of the mushrooms cultured in Injin mugwort as an active ingredient. The composition of claim 1, wherein the mushroom is a roe deer mushroom. The composition of claim 2, wherein the mushroom is a mushroom mycelium or fruiting body. The composition of claim 1, wherein the extract is prepared using ethanol as an extractant.  According to claim 1, wherein the mushroom cultured in jinjin (a) inoculating the mushrooms in the seedling liquid culture medium, and shaking them for 5 to 8 days at 23 to 26 ° C., pH 5 to 6, and agitation speed at 100 to 140 rpm to obtain seed material; (b) water 1: 1 Adding to a weight ratio to 10 to sterilize to prepare a solid medium; And (c) inoculating the seed medium obtained in step (a) at a solid medium of 0.1 to 30% by weight, and incubating the same at a culture temperature of 23 to 26 ° C. and a culture period of 40 to 50 days to obtain a mycelia; Composition prepared by the method comprising a. The composition of claim 1, wherein the anticancer composition is used for cervical cancer, rectal cancer or leukemia. The composition of claim 1, wherein the composition is provided as a food, food additive, beverage or medicament.