JPH0347835B2 - - Google Patents

Description

[Detailed description of the invention]

(Industrial Application Field) The present invention relates to a foam for culturing microorganisms that can effectively culture microorganisms. (Prior Art) In addition to the submerged culture method, microorganisms can be cultured using a solid culture method using rice bran or the like. Depending on the type of target product (e.g. bacterial cells, metabolic products, etc.)
Submerged culture methods and solid state culture methods are used alone or in combination. However, in the submerged culture method, pellets are formed during culture, which reduces the efficiency of culturing microorganisms. With solid state culture methods, it is difficult to separate products produced by microorganisms. In order to solve these drawbacks, a method has been proposed in which a foam is made to support a culture medium and culture is carried out. For example, Japanese Patent Publication No. 55-36313 discloses a method in which a foam such as a sponge is impregnated with a culture medium and then cultured stationary. However, with this method, it is difficult to impregnate the foam with the medium solution. Moreover, there is a limit to the amount of impregnation, with commercially available polyurethane foams having 30 to 50% impregnation, and even foams with good impregnation properties.
Up to 90%. Therefore, a sufficient medium cannot be supplied to the microorganisms, and the microorganisms cannot be cultured effectively. Such foams have poor water retention capacity and tend to dry out, making them unsuitable for culturing microorganisms. In the submerged culture method, a culture method in which a large number of foam pieces are added (disclosed in JP-A-60-214878) and a hydrophilic foam in which a peptide matrix is formed in the molecule (JP-B-53-11316) (disclosed in the official gazette) is also known. However, with these foams, the solid content and fats and oils in the medium components are adsorbed by the foam,
The medium does not penetrate inside the foam. Therefore,
Since microorganisms are cultured only on the surface of the foam, the microorganism culture efficiency is low. (Problems to be Solved by the Invention) The present invention solves the above conventional problems, and its purpose is to provide a foam for culturing microorganisms that can effectively culture microorganisms. It is in. Another object of the present invention is to provide a foam for culturing microorganisms that allows easy separation of products obtained by culturing microorganisms. (Means for Solving the Problems) Instead of the conventional method of impregnating a foam with a medium liquid, the present invention mixes a medium component into a foam composition and foams it, thereby impregnating a medium into a foam matrix. Since the components are incorporated and physically or chemically supported in the matrix, cultivation can be performed not only on the surface of the foam but also inside the foam, thereby allowing effective cultivation of microorganisms. It was completed based on the knowledge of the people involved. The foam for culturing microorganisms of the present invention is obtained by foaming a foam composition together with a medium component, and the medium component is supported within the foam matrix, thereby achieving the above object. The media components are primarily physically incorporated into the foam matrix during foaming of the foam composition. However, for example, if the medium component has an amino group or a carboxyl group and the foam composition has an isocyanate group, the medium component and the foam matrix will chemically react. As a result, the culture medium and other components are chemically supported within the foam matrix through urea bonds and acid amide bonds. The hydroxyl groups of the medium components react with the isocyanate groups to generate carbon dioxide gas and promote foaming. Foam matrices include, for example, polyurethane foam, polystyrene foam, vinyl chloride foam, polyethylene foam, and polyester foam. Particularly preferred is polyurethane foam. Polyurethane foam is obtained by reacting and foaming polyether or polyester, an isocyanate compound having two or more isocyanate groups in the molecule, and water or other blowing agent. As the isocyanate compound, common polyfunctional isocyanates are used, such as tolylene diisocyanate, diphenylmethane diisocyanate,
Diphenyl diisocyanate, naphthalene diisocyanate, xylene diisocyanate, butane diisocyanate, triphenylmethane-4,
There is 4′,4″-triisocyanate. Polyether or polyester reacts with an isocyanate compound to form a prepolymer. By reacting this prepolymer with water, carbon dioxide gas is generated and foams, forming polyurethane foam. Polystyrene foam is formed by adding a blowing agent (pentane, hexane, heptane, etc.) to polystyrene prepolymer and emulsion polymerization in water.
Vinyl chloride foam can be obtained by a pyrolysis method or a gas blowing method. A polystyrene foam is obtained by adding a foaming agent such as petroleum ether or gas Freon 12 to a polyethylene prepolymer, kneading the mixture, and foaming the mixture by heating. Regardless of which foam is used, by adding a medium component together with a blowing agent to the prepolymer before foaming and foaming, the medium component is incorporated into the foam matrix and is physically or chemically bonded to the matrix. carried. Media components include an assimilable carbon source, an assimilable nitrogen source, inorganic salts, and natural organic matter. Carbon sources include, for example, glucose, sutucarose, lactose, maltose, glycerin, starch, and molasses. Nitrogen sources include, for example, ammonium sulfate, ammonium chloride, and ammonium nitrate. Inorganic salts include, for example, phosphates such as potassium dihydrogen phosphate, magnesium nitrate,
Contains magnesium, potassium, and calcium. Natural organic substances include, for example, meat extract, fish extract,
Animal tissue extracts or milled products such as pupa flour; plant tissue extracts or milled products such as course steep liquor, soybean oil, malt extract, soybean flour; dried yeast,
There are microorganisms or their extracts such as yeast extract and polypeptone. If the foam matrix is a polyurethane foam, the prepolymer (a reaction product of polyether or polyester with an isocyanate compound) and water or other blowing agent are added to the medium components and subjected to reaction. Water-soluble medium components are mixed with the prepolymer in an aqueous solution. Water-insoluble medium components are dispersed in the prepolymer solution. The amount of water is preferably in the range of 10 to 100 parts by weight based on 100 parts by weight of the prepolymer. If it is less than 10 parts by weight, the foaming reaction will be delayed and a foam with the desired foam density will not be obtained.
The reaction between the prepolymer and the medium components and the support of the medium components in the foam matrix are also insufficient. When the amount exceeds 100 parts by weight, the reaction between water and prepolymer takes precedence, making it difficult for medium components to be incorporated into the foam matrix. The medium component is used in an amount of 20 to 500 parts by weight, preferably 50 to 200 parts by weight, based on 100 parts by weight of the prepolymer. Below 20 parts by weight, the medium components are not sufficiently contained within the foam matrix. Amounts of medium components exceeding 500 parts by weight cannot be carried within the foam matrix. Peptides such as soluble collagen, gelatin, albumin, etc. may be added to the foam of the present invention in order to form urea bonds or acid amide bonds in the foam matrix and obtain a tough peptide matrix. The foam matrix preferably contains a hydrophilic polymer to increase its water retention capacity. Due to the inclusion of hydrophilic polymers, there is little need to replenish the foam matrix with water. Hydrophilic polymers include, for example, agar, polyvinyl alcohol, and polyacrylamide. The microorganism culture foam thus obtained is sterilized in an autoclave (120° C., 1.2 atm) after containing an appropriate amount of water, and then subjected to static culture. After the foam is crushed, it may be subjected to sub-liquid culture together with medium components and water. It is also possible to inoculate the foam with insecticidal microorganisms, culture them within the foam, and use the microorganisms as an insecticide. For example, Beauveria tenella, which is an insecticidal microorganism that kills longhorn beetles and scarab beetle larvae, is cultivated and used to exterminate longhorn beetles and scarab beetle larvae. This foam can be stored aseptically for a long period of time if the moisture in the foam is removed by drying at 110° C. for 1 hour or more. It may be hydrated when used. (Example) The present invention will be described below with reference to Examples. Example 1 100 parts by weight of polyether isocyanate (Sofranate, manufactured by Toyo Tire & Rubber Co., Ltd.) were mixed at room temperature with 15 parts by weight of glucose, 30 parts by weight of pupa powder, and 11 parts by weight of agar as culture medium components, and further mixed with gelatin. 20 parts by weight of a 5% aqueous solution were mixed and reacted for several minutes at room temperature. The reaction resulted in a microbial culture foam containing medium components within the foam matrix. The resulting foam was autoclaved (120℃,
1.2 atm) for 20 minutes. The filamentous fungus (Beauveria tenella) is
Using 100 ml of culture medium extracted from 20 g of pupa powder and 40 g of pupa powder, preculture was carried out in a 300 ml Erlenmeyer flask while shaking with a rotary shaker. Transfer 3 ml of this culture solution to the above sterilized foam (30 cm ² × 1
cm) and cultured statically. After culturing at 25°C for one week, the foam was observed and found that the entire surface of the foam was covered with filamentous fungal spores. Add 5ml or 10ml to the foam before contacting the culture solution.
When similar static culture was carried out by impregnating the foam with water, spores were present on the entire surface of the foam. Comparative Example 1 Example 1 except that no culture medium components were added.
A foam for culturing microorganisms was obtained in the same manner as described above.
Three foams (30 cm ² × 1 cm) each were impregnated with 2.5 ml, 5 ml, and 10 ml of the liquid medium of Example 1, and similarly inoculated with 3 ml of culture solution and cultured. After culturing at 25°C for one week, we observed the foam and found that more than two-thirds of the foam was covered with filamentous fungal spores in the foam with an impregnated amount of 10 ml. Mycelia did not grow in areas with little liquid, and uneven culture was observed. Example 2 150 parts by weight of lactose was added to 100 parts by weight of polyether isocyanate (Sofranate, manufactured by Toyo Rubber Co., Ltd.)
Parts by weight, sodium nitrate 11.25 parts by weight, potassium dihydrogen phosphate 18.75 parts by weight, magnesium nitrate
9.40 parts by weight were added and mixed at room temperature. Then, 75 parts by weight of corn steep liquor was added to the above isocyanate mixture and reacted for several minutes at room temperature to obtain a foam containing medium components. The obtained foam was shredded into 5 mm square pieces. 4.9 to 9.7 g of this foam and 100 ml of distilled water were placed in a 300 ml Erlenmeyer flask, and one platinum loop of Pen. chrysogenum (spores) was inoculated and cultured in a rotary shaker. Mycelium grew uniformly within the foam and produced penicillin, which was released outside the foam. The amount of penicillin obtained was comparable to that produced using the foam culture method of JP-A-60-214878. The culture results on the 7th day of culture are shown in Table 1. In this example, only foam and distilled water are used for culturing the bacterial cells, and no ordinary liquid medium is used.
Therefore, the produced penicillin can be easily isolated without the need for a step of separating it from medium components such as lactose. Example 3 100 parts by weight of polyether isocyanate (Sofranate, manufactured by Toyo Rubber Co., Ltd.), 0 lactose
Parts by weight, sodium nitrate 11.25 parts by weight, potassium dihydrogen phosphate 18.75 parts by weight, magnesium sulfate
9.40 parts by weight were added and mixed at room temperature. Then, 75 parts by weight of corn steep liquor was added to the above isocyanate mixture and reacted for several minutes at room temperature to obtain a foam containing medium components. The obtained foam was shredded into 5 mm square pieces. 3-6 g of this foam and 100 ml of 4% lactose solution by weight
Put it in a 300ml Erlenmeyer flask and put it in a Pen.
One platinum loop of chrysogenum (spores) was inoculated and cultured in a rotary shaker. Mycelia grew within the foam and produced penicillin, which was released outside the foam. The amount of penicillin obtained was
The amount of penicillin produced using the foam culture method of 214878 was comparable. The culture results on the 7th day of culture are shown in Table 2. Comparative Example 2 Pen. with normal medium composition without using foam.
chrysogenum (spores) were cultured. The culture method is
The procedure was the same as in Example 2, and the results are shown in Table 3.

【table】

【table】

[Table] (Effects of the invention) The foam for culturing microorganisms of the present invention is thus:
Since the medium components are supported within the foam matrix, microorganisms can be effectively cultured not only on the surface of the foam but also inside the foam. The medium components are supported within the foam matrix and are less likely to be mixed into the product obtained by culture. Therefore, separation of the products is facilitated. If static culture is performed using this foam, there will be less outflow of medium components, and in submerged culture, there is no risk of solids and fats and oils in the medium being adsorbed to the foam.
The culture efficiency of microorganisms increases. As a result, the foam for culturing microorganisms of the present invention can be effectively used for culturing various microorganisms.

Claims (1)

[Scope of Claims] 1. A foam for culturing microorganisms, which is obtained by foaming a foam composition together with culture medium components, and in which culture medium components are supported within a foam matrix. 2. The foam for culturing microorganisms according to claim 1, wherein the foam matrix is a polyurethane foam. 3. The foam for culturing microorganisms according to claim 1, wherein the foam matrix contains a hydrophilic polymer to increase water retention capacity. 4. The foam for culturing microorganisms according to claim 1, wherein the foam has a peptide matrix. 5. The foam for culturing microorganisms according to claim 1, wherein the foam composition contains a compound having an isocyanate group and a hydrophilic polymer capable of reacting with the isocyanate group.