JP5295607B2 - Liquid medium of basidiomycetous basidiomycetes - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a liquid culture medium which can dramatically improve the production of cells, when a Boletaceae basidiomycete is cultured in the liquid. <P>SOLUTION: A conventional liquid culture composition is compounded with soybean milk and a silicone, and then used to culture the Boletaceae basidiomycete. Thus, the production of the cells can dramatically be improved. <P>COPYRIGHT: (C)2010,JPO&amp;INPIT

Description

The present invention relates to a liquid culture medium for basidiomycetous basidiomycetes.

Boletaceae belongs to the order of Agaricales and forms a fruiting body with a tube hole. The basidiomycete family basidiomycete (Suillus) includes the following species: Suillus grevillei, Suillus luteus, Amitake (Suillus).
bovinus), Chichiawatake (Suillus granulatus), Shirolus laricinus, Numelizuiguchi (Suillus)
luteus) and the like belong. Some of the basidiomycetes belonging to the genus Numeriguchi show various physiological activity effects. For example, tyrosinase activity inhibitory action, vascular endothelial cell growth promoting action, α-glucosidase inhibitory action, COX-2 activity inhibitory action are known. Yes. (For example, see Patent Document 1, Patent Document 2, Patent Document 3, and Patent Document 4.)

However, the natural fruit body as its supply source is limited in the time of acquisition, and even after it is obtained, there are many problems in practical use such as fluctuations in usefulness due to individual differences of fruit bodies and mass production. Therefore, in recent years, a method for artificially culturing mycelium of ectomycorrhizal fungi containing basidiomycetous basidiomycetes has been proposed (for example, Patent Document 5). In this culturing method, after a liquid medium is contained in the porous carrier so that the water content is 30 to 40% by weight, the porous carrier is sterilized in a sealed state in a culture tank, and then an exogenous bacterium. A liquid medium containing root mycelium vegetative mycelium is added to the culture tank to adjust the water content of the porous carrier to 65 to 80% by weight. Thus, an ectomycorrhizal fungus solid culture method characterized by culturing ectomycorrhizal fungi.

Furthermore, Patent Document 6 describes a method of adding okara to an artificial culture medium in the artificial cultivation of Hatake shimeji. However, the culture medium described in the publication is a solid culture medium for generating fruit bodies of basidiomycetes, and is not the liquid culture medium of the present invention. In general, the nutrients necessary for the growth of basidiomycetes and the nutritional conditions necessary for the formation of fruiting bodies are greatly different. In addition, the components in the fruit body, the components in the mycelium obtained by liquid culture, and the components in the mycelium obtained by solid culture or fungal bed culture are often very different, That is, a nutrient component effective for a solid culture medium is not always effective for a liquid culture medium as it is (for example, Patent Document 7).

Similarly, Patent Document 7 describes a method for dramatically increasing the yield of Meshimakob mycelium by adding soy milk to a conventional liquid medium. However, this publication considers shaking culture and does not describe the effect of aeration culture applied when culturing in large quantities.

Japanese Patent No. 3678832 JP 2000-212059 A Japanese Patent No. 3511231 Japanese translation of PCT publication No. 2004-529079 JP 2005-27546 A Japanese Patent Laid-Open No. 2002-70 JP 2006-271339 A

The problem to be solved is that basidiomycetous basidiomycetes have slow growth of mycelia, and the yield of mycelium obtained by the conventional liquid culture method is small, and sufficient mycelium cannot be obtained. .

Means to solve the problem

As a result of intensive research, the inventors of the present invention cultivate basidiomycetes belonging to the genus Iridaceae by blending soymilk and silicone into the conventional liquid culture composition when performing liquid culture of basidiomycete genus Iridaceae. As a result, it was found that the production amount of mycelium was dramatically improved, and the present invention was completed. That is, the present invention drastically increases the growth of Mycidae mycorrhizal fungi mycelia by inoculating the mycobacteria basidiomycetes on a liquid medium to which soy milk and silicone are added, and subjecting them to liquid culture. .

BEST MODE FOR CARRYING OUT THE INVENTION

According to the liquid culture method of the Ibusidae genus Basidiomyces basidiomycetes according to the present invention, by culturing in the liquid medium containing soy milk and silicone, the yield of the mussels Iridae genus Basidiomycetes mycelium is dramatically reduced in a short period of time. It is possible to increase.

The soy milk in the present invention is a juice obtained by grinding and squeezing soybeans soaked in water. Since soy milk contains components such as proteins, lipids, and minerals, it is a good nitrogen source for basidiomycetes. The soy milk used for the liquid culture used in the present invention is not particularly limited as long as it is soy milk. In the examples, commercially available products manufactured by Kibun Food Chemifa Co., Ltd. were used. Further, the effective amount is not particularly limited as long as an effective amount is added, but preferably 0.1 to 20% by weight in the liquid medium.
More preferably, it is 0.5 to 10% by weight. When the blending amount is less than 0.1% by weight, it is not preferable because a sufficient effect cannot be exhibited. This is not preferable because it cannot be carried out easily or contamination easily occurs during ventilation.

Silicone in the present invention is a general term for organic compounds having a continuous siloxane bond (—Si—O—) as a skeleton. Structurally, dimethyl silicone, methylphenyl silicone, amino-modified silicone, polyether-modified silicone, alkyl-modified silicone, epoxy-modified silicone, fluorine-modified silicone, carboxy-modified silicone and the like can be exemplified. In the examples, trade name silicon KS-66 (component name: Simethicone) manufactured by Shin-Etsu Chemical Co., Ltd. was used. Further, the effective amount is not particularly limited as long as it is an effective amount, but preferably 0.0002 to 0.04% by weight in the liquid medium.
, More preferably 0.001-0. 02% by weight. When the blending amount is less than 0.0002% by weight, it is not preferable because a sufficient effect cannot be exhibited, and when it exceeds 0.04% by weight, the effect reaches equilibrium.

The basic medium used in the liquid culture of the present invention is preferably a potato dextrose medium, a modified Merin norclan (MMN) medium, or a modified Hamada medium that is used for culturing ectomycorrhizal fungi including iridid basidiomycetes. . A potato dextrose medium can be prepared, for example, by dissolving 2.4 g of Potato-Dextrose Broth in 100 mL of purified water. MMN medium is, for example, glucose 1.0
g, malt extract 0.3 g, calcium chloride dihydrate 0.0066 g, sodium chloride 0.0025 g, potassium dihydrogen phosphate 0.05 g, dihydrogen ammonium phosphate 0.025 g, magnesium sulfate heptahydrate It can be prepared by dissolving 015 g, 0.12 mL of a 1% iron (III) chloride solution and 0.01 mg of thiamine hydrochloride in 100 mL of purified water. The modified Hamada medium can be prepared, for example, by dissolving 2.0 g of glucose and 0.5 g of dry yeast in 100 mL of purified water. The basic medium contains nutrients necessary for growth when cultivating mycidae basidiomycetous mycelium such as carbon source and nitrogen source, and may further contain vitamins and minerals.

As the carbon source of the basic medium, monosaccharides such as glucose, xylose, mannose and galactose, disaccharides such as sucrose, maltose, lactose and trehalose, and polysaccharides such as dextrin, starch and cellulose can be used.

As the nitrogen source of the basic medium, organic substances such as yeast extract, malt extract, peptone, polypeptone, and casamino acid, and nitrogen-containing compounds such as calcium nitrate, ammonium sulfate, sodium nitrate, and ammonium chloride can be used.

Examples of vitamins that can be used include biotin, thiamine (vitamin B1), pyridoxine (vitamin B6), pantothenic acid, inositol, nicotinic acid, and the like. Examples of the mineral include phosphorus, nitrogen, potassium, calcium, magnesium, sulfur, iron, manganese, zinc, boron, copper, molybdenum, chlorine, sodium, iodine, cobalt and the like. These components include, for example, potassium nitrate, sodium nitrate. , Calcium nitrate, potassium chloride, potassium hydrogen phosphate, potassium dihydrogen phosphate, calcium chloride, magnesium sulfate, sodium sulfate, ferrous sulfate, ferric sulfate, manganese sulfate, zinc sulfate, boric acid, copper sulfate, It can be used as a compound such as sodium molybdate, molybdenum trioxide, potassium iodide, cobalt chloride or the like.

Of course, any medium other than the above can be applied to the present invention as long as it is suitable for culturing mycomycetes basidiomycetes mycelium.

According to the present invention, the liquid culture method of the genus Nigiriidae basidiomycetes inoculates the above-mentioned soymilk and silicone-added liquid culture medium with the inoculum of Physiidae basidiomycetes, and is performed under aerobic conditions at room temperature of about 20 to 30 ° C. It is desirable to culture for 10 to 20 days.

Hereinafter, the present invention will be described in more detail with reference to examples, but the present invention is not limited thereto. In addition, culture and operation methods shall be performed aseptically.

<Example 1>
Glucose 10.0 g, malt extract 3.0 g, calcium chloride dihydrate 0.066 g, sodium chloride 0.025 g, potassium dihydrogen phosphate 0.5 g, diammonium hydrogen phosphate 0.25 g, magnesium sulfate 7 water A basic liquid medium was prepared by adding 0.15 g of the Japanese product, 1.2 mL of a 1% iron (III) chloride solution and 0.1 mg of thiamine hydrochloride to 1000 mL of water. Further, soymilk and silicone as additives were added in the amounts shown in Table 1 to prepare a liquid medium, and then autoclaved using an autoclave.

After the above-mentioned liquid medium was allowed to cool to room temperature, it was pre-cultured with 2.4% th (manufactured by Difco) (shaking culture at 24 ° C. and 150 rpm for 14 days). Purchased, NBRC No. 32786) was taken from Potato-Dextrose Bro-Niguchi culture, and homogenated inoculated into 10 mL flasks. After inoculation, aeration culture was performed at 24 ° C.

About 12 days after the start of culture, mycelium grew on the liquid medium. Therefore, the grown mycelium was collected, washed well with water and then freeze-dried.

The growth ratio of the collected mycelium is shown in Table 1. The growth ratio was calculated by calculating the ratio with respect to the yield when cultured in the basic medium of Comparative Example 1 as 100.

According to Table 1, when soymilk or silicone alone is mixed, the growth rate in the basic medium is hardly changed, but when soymilk and silicone are used in combination, the growth rate is dramatically increased. . In addition, soy milk is 0.5 to 10% by weight of the total amount of the medium.
And 0.001 to 0 silicone. It was recognized that the growth rate increased explosively when 02 wt% was added.

As for other basidiomycetes belonging to the genus Numeriiguchi, Numeriiguchi, Amitake, Titiawatake, Shirunumeiguchi, and Numerizbuiguchi, remarkable growth promotion was similarly recognized by the combined use of soy milk and silicone. From the above results, it has been clarified that blending soy milk and silicone has a great influence on the growth of basidiomycetous basidiomycetes.

According to the liquid culture method of the mycorrhizal genus basidiomycetous mycelium according to the present invention, by culturing it in a liquid medium containing soy milk and silicone, the yield of mysterious mycorrhizal mycelium is drastically reduced in a short period of time. It can be increased and can be widely applied in the fields of cosmetics, pharmaceuticals, and foods.

Claims (2)

0.001 to 0 5 to 10 wt% and a silicone medium entire amount of soy milk. A method for producing Mytilus fungus body using a liquid culture medium characterized by comprising 02 wt% . The basidiomycetes belonging to the genus Suillus are Suillus grevillei, Suillus luteus, Suillus boulinus, Suilus sulph. The production method according to claim 1 , wherein: