JP2001103927A - Method of producing agaricus blazei murill extract having high antitumor activity - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method of efficiently producing Agaricus blazei Murill extract which is safe and has a high antitumor activity, and to obtain a composition allowing industrially useful use of the extract. SOLUTION: This method of efficiently producing Agaricus blazei Murill extract comprises the following steps: subjecting Agaricus blazei Murill to extraction under pressure, preferably at 1 to 5 atm in an aqueous solvent preferably water or a hydrous alcohol, further preferably under heating; and fractionating the thus obtained extract using 50% ethanol to collect the resultant precipitate. The other objective edible composition is obtained by formulating this Agaricus blazei Murill extract.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to a method for producing an Agaricus mushroom extract having high antitumor activity, and an edible composition obtained by blending the extract. More specifically, Agaricus blazei belonging to Agaricaceae
The present invention relates to a method for efficiently producing an extract having high antitumor activity from Agaricus mushrooms such as muril and the like, and further relates to an edible composition containing the extract.

[0002]

2. Description of the Related Art In recent years, the relationship between dietary patterns and so-called lifestyle-related diseases such as hypertension, diabetes, hyperlipidemia, and ischemic heart disease has been elucidated, and interest in food components has been increasing. On the other hand, cancer has become the leading cause of death due to illness, and the treatment and prevention of cancer has been made more intensive than ever. Among them, attention has been paid to functions such as physiological activity and pharmacological action of foods and food raw materials from the viewpoint of cancer prevention.

[0003] Mushrooms have been used for foods since ancient times and used as raw materials for herbal medicines and folk medicines. Many of the components contained in these mushrooms exhibit a special physiological action or pharmacological activity. Have been. As an antitumor active substance contained in mushrooms, β-glucan has received particular attention. For example, lentinan from shiitake mushroom, krestin from kawatake mushroom, schizophyllan from shirohirotake mushroom, and the like have already been approved and sold as pharmaceuticals in Japan. The mechanism of these physiological activities is that β-glucan binds to the surface of lymphocytes and specific serum proteins, and the activation of macrophages, NK cells, and the like suppresses the growth of cancer cells.

Further, as an antitumor active component contained in Agaricus brazeiimuril, a kind of Agaricus mushroom, an acid polysaccharide (Japanese Patent Application Laid-Open No. 64-671) is known from its fruiting bodies.
No. 94), neutral polysaccharides (JP-A-64-67195) and protein polysaccharides (JP-A-2-78630) and protein polysaccharides (JP-A-61-475) from mycelia.
No. 18), and protein polysaccharides (JP-A-61-47519) are further fractionated from the culture filtrate of mycelium, respectively, and the antitumor activity of each component is reported. It is said that these active ingredient polysaccharides also have a β-glucan structure.

As described above, most of the antitumor active components contained in mushrooms such as Agaricus mushrooms have polysaccharides of β-
It has a glucan structure, has various sugar compositions, forms a complex with a protein, and is water-soluble. Therefore, conventionally, the active ingredient was obtained by extracting mushrooms with water or hot water. However, since the amount of these active ingredients contained in mushrooms is extremely small, it has not been possible to extract a large amount of these active ingredients. In addition, since the active ingredient has a complicated structure and composition, the antitumor effect of the extract varies depending on the extraction method, and the quality is not constant.

[0006]

In view of this situation, the present invention has developed a method for efficiently producing an Agaricus mushroom extract having high antitumor activity, and the obtained extract can be used industrially. It was intended to provide a composition.

[0007]

Means for Solving the Problems In order to solve the above-mentioned problems, the present inventors have conducted intensive studies on a method for producing an Agaricus mushroom extract, and found that Agaricus mushrooms were extracted under high pressure using an aqueous solvent. The present inventors have found that an extract having high antitumor activity can be efficiently obtained, and have completed the present invention. That is, according to the present invention, there is provided a method for producing an Agaricus mushroom extract having high antitumor activity, comprising extracting Agaricus mushrooms under pressure and using an aqueous solvent, and fractionating the extract with 50% ethanol. Is done. Here, the pressure under pressure is desirably 1 to 5 atm, the aqueous solvent is preferably water or hydroalcohol, more preferably the aqueous solvent exhibits alkalinity. Below, more preferably 101-1
It is better to carry out at 50 ° C. The present invention also provides an edible composition containing the Agaricus mushroom extract obtained by the above-mentioned production method. A preferred embodiment of the composition is in the form of a powder, granules, tablets or capsules.

[0008]

BEST MODE FOR CARRYING OUT THE INVENTION Agaricus mushrooms as raw materials belong to the genus Agaricus, and include Agaricus blazei Murill,
Examples include agaric mushrooms and mushrooms. Agaricus blazei muril is a mushroom belonging to the agaricaceae native to Brazil, and it was initially difficult to cultivate artificially because it requires high temperature and high humidity conditions. However, nowadays, cultivation methods have been developed and mass production systems are available not only in Japan but also in Taiwan, China, etc., so they can be easily obtained. Agaric mushrooms and mushrooms are provided for normal consumption.

In the present invention, fruit bodies and / or mycelia of Agaricus blazeimuril are used. The fruiting body may be either raw or dried, but a dried fruiting body is preferred in terms of handleability, preservability, extraction efficiency and the like. As the mycelium, a raw or dried mycelium obtained by culturing an inoculum in a medium containing a carbon source and a nitrogen source can be used. Dried ones are simple. Similarly, in the case of agaric mushrooms, mushrooms, etc., it is preferable to use a dried body.

When extracting the Agaricus mushroom extract of the present invention, it is important to carry out extraction under pressure using an aqueous solvent. Here, examples of the aqueous solvent include water and hydrous alcohol. The alcohol is preferably a lower monohydric alcohol such as methanol, ethanol, propanol, isopropanol and butanol. When the agaricus mushroom extract of the present invention is used for food, water or hydrated ethanol is used. However, when it is used for medical use, other alcohols may be used. The water content of the water-containing alcohol varies depending on the type of alcohol, and is approximately 50% (v /
v) or more are preferred. If the water content is lower than this, the yield of the desired product and the antitumor activity decrease. In addition,
When an alcohol other than ethanol is used, it is preferable to increase the water content as the number of carbon atoms increases. This preferred guideline is that the water content for methanol is about 50% (v / v) or more, similarly for ethanol is about 70% (v / v) or more, and for isopropanol is about 80% (v / v) or more. .

It is desirable that the aqueous solvent exhibits alkalinity. This makes it possible to obtain the Agaricus mushroom extract having a high antitumor activity, which is the object of the present invention, in a larger amount. The alkalinity is preferably weak to moderate alkalinity, and a preferable pH is up to about 10,
More preferably, it is 7.5 to 9. To make the aqueous solvent alkaline, a conventional method may be used, and for example, a hydroxide or phosphorus oxide of potassium or sodium may be used.

The hydrous alcohol or water is used in an amount of 1 to 20 times, more preferably 3 to 15 times the volume of the raw material.
Extraction is performed under pressure in a pressure-resistant container using double volume.
Here, the raw material is appropriately crushed, shredded or crushed by freezing treatment, impact destruction treatment, enzyme treatment, etc., and the pressure under pressure in the extraction treatment is 1 to 5 atm, more preferably 1.1 to 5 atm.
3 atm. When the pressure is out of the above range, the yield and the antitumor activity of the target compound of the present invention decrease. The extraction treatment of the Agaricus mushroom extract of the present invention is preferably performed at the time of heating the above-mentioned raw material and the aqueous solvent, and in particular, 101 to 1
It is carried out at the time of heating at 50 ° C., more preferably 110 to 130 ° C., and under the above-mentioned pressure, with appropriate stirring. The Agaricus mushroom extract obtained under these conditions has an extremely high antitumor activity and a high yield.

The Agaricus mushroom extract of the present invention can be produced by the following operation. That is, the mixture of the above Agaricus mushrooms and the aqueous solvent is subjected to extraction under the above-mentioned conditions for about 10 minutes to 5 hours, more preferably 30 minutes to 2 hours while appropriately stirring, and the residue is filtered to obtain an extract. . The aqueous solution is added to the residue and the residue is treated in the same manner to obtain an extract. This operation is further repeated about 2 to 10 times to obtain an extract. These extracts are combined, concentrated under reduced pressure, subjected to a treatment such as spray-drying or freeze-drying to obtain an extract. The extract is further fractionated with 50% ethanol, and the precipitate is collected. The agaricus mushroom extract of the present invention can be produced.

The agaricus mushroom extract of the present invention contains about 8% by weight or more of a complex of polysaccharide and protein (hereinafter sometimes referred to as protein polysaccharide) in the extract. Here, the protein polysaccharide is a mixed composition obtained by adding an equal volume of ethanol to a 4% by weight aqueous solution of the extract, and precipitating after cooling with ice. That is, the protein polysaccharide that is the Agaricus mushroom extract of the present invention has β-glucan whose main constituent sugar is glucose as a main chain, has a plurality of branched chains of sugars, and further has a protein bound to this polysaccharide. A composition comprising a substance having a molecular weight of 10,000 or more, more preferably about 10,000 to about 100,000 as a main component, mainly containing mannose as a monosaccharide in addition to the protein polysaccharide, and also containing amino acids and peptides. It is a complex composition. The agaricus mushroom extract of the present invention exhibits remarkable antitumor activity as described later. Also,
It also has the effect of preventing P. aeruginosa infection.

According to the production method of the present invention, an Agaricus mushroom extract which is a composition having a high antitumor activity as described above is provided, and a composition further comprising the same is also provided. As an embodiment of this composition, an edible composition is suitable.

The Agaricus mushroom extract of the present invention can be used as it is in a liquid, gel or solid food such as juice, soft drink, tea, soup, jelly, yogurt,
Add to pudding, sprinkle, cake mix, powdered or liquid dairy products, bread, cookies, etc., or, if necessary, form pellets, tablets, granules, etc. with excipients such as dextrin, lactose, starch, etc. It can be processed, or coated with gelatin or the like, formed into capsules, and used as health foods or dietary supplements. The amount of the Agaricus mushroom extract of the present invention in these foods or edible compositions cannot be uniformly defined depending on the type or state of the food or edible composition, but is preferably about 0.01 to 50% by weight, Preferably it is 0.1 to 30% by weight. The amount is 0.01
If the amount is less than 5% by weight, the desired effect of ingestion is small, and
If it exceeds 0% by weight, the flavor may be lost or the food may not be prepared depending on the type of food. The agaricus mushroom extract of the present invention may be used for food as it is.

[0017]

[Example] Example 1 Agaricus in a pressure-resistant extraction pot made of stainless steel (hereinafter the same)
1. 150 g of dried fruit body of brazei muril and pure water
76 L was added, the mixture was subjected to an extraction treatment under the conditions of 1.2 atm and 125 ° C. for 1 hour, and the content was filtered under reduced pressure to separate the extract from the residue. 1.41 L of pure water was added to the residue, and pressure extraction was performed in the same manner. The two extracts were combined, concentrated under reduced pressure, and freeze-dried to obtain 85.5 g of an extract. This extract was converted to an aqueous solution having a concentration of 4% by weight, an equal volume of ethanol was added thereto, and the mixture was ice-cooled for 15 minutes, and the resulting precipitate was collected by centrifugation. This was washed with ethanol and then with ether and air-dried to obtain 9.83 g of the Agaricus mushroom extract of the present invention.

Example 2 1.76 L of pure water was added to 150 g of dried fruit body of Agaricus brazeiimuril, extracted under the conditions of 2.0 atm and 60 ° C. for 1 hour, and filtered under reduced pressure to obtain an extract. On the other hand, 1.41 L of pure water was added to the residue, and pressure extraction was performed in the same manner. The two extracts were combined, concentrated under reduced pressure, and subjected to freeze-drying to obtain 78.4 g of an extract. This extract was fractionated by 50% ethanol in the same manner as in Example 1 to obtain 6.27 g of the Agaricus mushroom extract of the present invention.

EXAMPLE 3 1.76 L of pure water was added to 150 g of a viable mycelium obtained by culturing an inoculum of Agaricus brazeimuryl in a medium containing glucose and peptone at 25 to 30 ° C. for 72 hours.
The mixture was extracted at 1.5 atm and 95 ° C. for 1 hour, and filtered under reduced pressure to obtain an extract. Meanwhile, 1.41 L of pure water was added to the residue, and extraction was performed in the same manner. The two extracts were combined, concentrated under reduced pressure, and subjected to a spray drying treatment to obtain 77.6 g of an extract.
This extract was fractionated by 50% ethanol in the same manner as in Example 1 to obtain 3.55 g of the Agaricus mushroom extract of the present invention.

Experimental Example 4 1.76 L of weak alkaline water adjusted to pH 8.0 with sodium hydroxide was added to 150 g of dried fruit body of Agaricus brazeimuryl, and the mixture was added under a condition of 1.2 atm and 125 ° C.
The mixture was extracted for a time and filtered under reduced pressure to obtain an extract. On the other hand, 1.41 L of the extraction solvent was added to the residue, and pressure extraction was performed in the same manner. The two extracts were combined, adjusted to pH 6.9 with acetic acid, concentrated under reduced pressure, and subjected to freeze-drying to obtain 72.5 g of an extract. This extract was fractionated with ethanol in the same manner as in Example 1 to obtain 7.83 g of the Agaricus mushroom extract of the present invention.

The antitumor activity of each Agaricus mushroom extract prepared in the above Examples was tested and evaluated by the method described below. Test Example 1 Six-week-old ICR male mice (purchased from CLEA Japan, Inc.) were preliminarily reared for one week, and then healthy mice were used for the experiment. Four mice were composed of 10 mice per group, and 1.0 × 10 ⁶ cells / mouse of 180 sarcoma cells were implanted subcutaneously in the abdomen. Agaricus mushroom extract (800 mg / kg mouse body weight) was orally administered for 14 days from the day after the tumor was implanted, and the change in body weight and tumor volume (major axis × minor axis ² /
2 calculated). On the 15th day, the mice in each group were sacrificed by cervical dislocation under ether anesthesia, the spleen and thymus were excised, and their weight was measured. Table 1 shows the weight of the spleen and thymus of each group, and the inhibition rate of the tumor volume of the test group with respect to the control (sarcoma 180 tumor-bearing mouse) at each day ｛(tumor volume of control group-tumor volume of test group) Quantity) ×
Table 2 shows the tumor volume １００ of 100 / control, respectively.

[0022]

[Table 1]

In Table 1, the numerical values indicate the average value ± standard error. From this data, no significant difference was observed between the tumor-bearing mouse group and each of the Agaricus mushroom extract administration groups. From this, it can be determined that no side effect was observed by oral administration of Agaricus mushroom extract.

[0024]

[Table 2]

The tumor volume of the tumor-bearing mice increased day by day, whereas the tumor volume of the agaricus mushroom extract-administered groups obtained in Examples 1 to 4 decreased relatively,
In particular, the extract extracted under high pressure and high temperature conditions showed a remarkable tumor-suppressing effect as compared with other extracts.

Example 5 2.34 L of 25% ethanol was added to 200 g of dried fruit body of Agaricus brazeimuryl, and the mixture was extracted at 1.3 atm under reflux for 1 hour and filtered under reduced pressure to obtain an extract. Further, 1.88 L of 30% ethanol was added to the residue, and an extraction treatment was performed in the same manner. The two extracts were combined, concentrated under reduced pressure, and subjected to freeze-drying to obtain 60.2 g of a paste-like extract. This extract showed almost the same antitumor activity as the extract prepared in Example 3.

COMPARATIVE EXAMPLE 1 1.76 L of pure water was added to 150 g of dried fruit body of Agaricus brazeiimuril, and extracted at 60 ° C. for 1 hour at normal pressure.
The extract was obtained by filtration under reduced pressure. On the other hand, pure water 1.4 was added to the residue.
1. L was added and extraction was performed similarly. The two extracts were combined, concentrated under reduced pressure, and subjected to freeze-drying to obtain 19.5 g of an extract. This extract was used as in Example 1 for 5
Agaricus mushroom extract 0.3
9 g were obtained. The antitumor activity of this extract was tested and evaluated using the method described in Test Example 1 (see Table 2).

The molecular weight distribution of the Agaricus mushroom extract prepared in the above Examples and Comparative Examples was measured by the following method. Test Example 2 A sample solution prepared by dissolving 30 mg of Agaricus mushroom extract in 2 ml of water was analyzed by high performance liquid chromatography based on a gel filtration method. At this time, dextran was used as a standard, and the molecular weight distribution was calculated by the ratio of each peak area / total sugar mass (total 100%). Table 3 shows the results.

[0029]

[Table 3]

The agaricus mushroom extract of the present invention contains a large number of fractions having a molecular weight of 10,000 or more, which are presumed to show high tumor activity. It is easier to extract. This tendency is particularly remarkable due to the extraction conditions of high pressure and high temperature. On the other hand, in the extraction process under normal pressure, the amount of low molecular weight components increases even in the heated state. It was also found that high molecular weight substances were extracted by hydroalcoholic extraction, but low molecular weight substances tended to be extracted more easily than pressure extraction treatment using only water.

Comparative Example 2 1.76 L of pure water was added to 150 g of dried fruit body of Pleurotus oyster mushroom, and the mixture was extracted under the conditions of 1.2 atm and 125 ° C. for 1 hour, and filtered under reduced pressure to obtain an extract. In addition, 1.41 L of pure water was added to the residue.
And pressurized extraction was performed in the same manner. The two extracts were combined, concentrated under reduced pressure, and subjected to a freeze-drying treatment to obtain 65.3 g of an extract. This extract was used for 50 times as in Example 1.
% Ethanol fractionation to obtain 5.24 g of Oyster mushroom extract.

Comparative Example 3 Oyster mushroom extract 63.3 was treated in the same manner as in Comparative Example 2 except that the pressure at the time of extraction was 1 atm and the temperature was 90 ° C.
g, Oyster mushroom extract 3.35 g was obtained.

When the results of Comparative Examples 2 and 3 are compared, there is no difference in yield between oyster mushrooms due to different extraction conditions. The antitumor activity of both oyster mushroom extracts in Test Example 1 was extremely small as compared with the case of Agaricus mushroom extract. From this, it became clear that the pressure extraction method of the present invention is effective for Agaricus mushrooms.

Example 8 Each raw material was sufficiently mixed at a ratio of 10 mg of Agaricus mushroom extract obtained in Example 1, 150 mg of propolis, 50 mg of beeswax and 40 mg of corn oil while heating to 50 ° C. to obtain a uniform liquid composition. Prepared. This was supplied to a capsule filling machine to prepare a gelatin-coated capsule having an inner volume of 250 mg per grain.

Example 9 150 g of butter, 250 g of shortening, 80 g of milk
90 g of sugar and 90 g of hen's egg were added while thoroughly stirring with a household whipper, and the mixture was thoroughly stirred.
g, baking powder 1.5 g, and agaricus mushroom extract 10 g obtained in Example 2 were added and further mixed well. Then, after letting it stand for 30 minutes,
They were divided into pieces and baked in an oven to produce a butter cookie.

[0036]

According to the present invention, an extract having high antitumor activity and an Agaricus mushroom extract can be efficiently produced using Agaricus mushroom as a raw material. This agaricus mushroom extract significantly reduces the volume of tumor by oral ingestion, has no side effects, and is useful for treating and preventing cancer. Such an effect is more remarkably exhibited as the amount of the protein polysaccharide increases. Further, the present invention provides an edible composition containing the Agaricus mushroom extract, and the composition is used as a food for preventing or treating cancer diseases.

Claims (8)

[Claims] 1. A method for producing an agaricus mushroom extract having high antitumor activity, comprising extracting an agaricus mushroom under pressure using an aqueous solvent, and separating the extract with 50% ethanol. 2. The method for producing an extract according to claim 1, wherein the pressure under pressure is 1 to 5 atm. 3. The method according to claim 1, wherein the aqueous solvent is water or a hydroalcoholic. 4. The method for producing an extract according to claim 1, wherein the aqueous solvent exhibits alkalinity. 5. The method according to claim 1, wherein the substance is extracted during heating.
5. The method for producing the extract according to any one of the items 4 to 4. 6. The method for producing an extract according to claim 5, wherein the temperature at the time of heating is 101 to 150 ° C. 7. An edible composition comprising an agaricus mushroom extract obtained by the production method according to any one of claims 1 to 6. 8. The edible composition according to claim 7, which is in the form of a powder, granules, tablets or capsules.