JP7624320B2 - Algae-killing agent and algae-killing method - Google Patents

Description

The present invention relates to an algaecide and an algae-removal method that are applied to algae that live in aquatic environments.

Aquatic algae such as Spirogyra and Polytrichum commune are generally known as aquatic plants that inhabit aquatic environments such as rice paddies, aquariums, and ponds. However, when aquatic algae overgrow, especially in rice paddies in early summer, they can entangle rice plants and knock them over, or the water temperature can not rise, causing growth to deteriorate, and they can rob rice of fertilizer nutrients, inhibiting rice growth by suppressing tillering, among other problems. Methods for removing aquatic algae include direct manual removal, mechanical removal, and spraying of algaecides. However, direct manual removal and mechanical removal are not easily adopted due to the labor and cost involved.

Conventionally, biocides having an algae killing effect are known, as disclosed in Patent Document 1. Patent Document 1 discloses organic antifouling biocides such as aminochloronaphthoquinone.

Special table 2012-507637 publication

However, conventional biocides require careful use because it is necessary to consider their impact on organisms other than the target organism and on the aquatic environment. For this reason, there is a demand for an algaecide and method that is easy to use and has excellent algae-killing effects.

The present invention is based on the discovery that a polysaccharide-degrading enzyme that degrades the cell walls of algae that live in aquatic environments has an excellent algae-killing effect.
In order to achieve the above object, an algicide according to one embodiment of the present invention is characterized in that it contains a polysaccharide-degrading enzyme that degrades the cell walls of algae that live in aquatic environments.

In the algicide, the polysaccharide-degrading enzyme may be an endo-type or exo-type polysaccharide-degrading enzyme.
In the algicide, the polysaccharide-degrading enzyme may have cellulolytic activity.

In the algaecide, the polysaccharide-degrading enzyme may contain cellulase derived from the genus Cellulomonas and cellulase derived from the genus Trichoderma.
In the algaecide agent, the algae may be at least one selected from green algae, zygophytes, glaucophytes, red algae, diatoms, brown algae, pinguiophytes, euphthalmophytes, apicomplexa, dinoflagellates, chlorarachniophytes, haptophytes, cryptophytes, euglena, and cyanobacteria.

Another embodiment of the algae-removal method of the present invention is characterized in that the algae-removal agent is administered to an aquatic environment in which algae live.
In the algae removal method, the aquatic environment may be a paddy field, an aquarium, a pond, a lake, the sea, a waterway, or a pool.

The present invention is easy to use and provides excellent algae-removal effects.

Photographs showing the appearance of each example of Amphibian in Test Example 1. 3 is a microscopic photograph of each example of Spirogyra in Test Example 1.

First Embodiment
Hereinafter, a first embodiment of the algaecide of the present invention will be described.
The algaecide of the present embodiment contains, as an algaecide component, a polysaccharide-degrading enzyme that decomposes the cell walls of algae that live in aquatic environments.

(Polysaccharide degrading enzyme)
The polysaccharide decomposing enzyme may be a polysaccharide decomposing enzyme that decomposes the cell walls of algae that live in an aquatic environment. Specific examples of the polysaccharide decomposing enzyme include cellulase, xylanase, pectinase, glucanase, mannanase, β-glucosidase, etc. These polysaccharide decomposing enzymes may be used alone or in combination of two or more.

The origin (source) of these polysaccharide decomposing enzymes is not particularly limited, and enzymes derived from animals, plants, and microorganisms can be used. Specific examples of microorganisms include the genera Cellulomonas, Trichoderma, Aspergillus, Bacillus, and Penicillium. Among these, from the viewpoint of improving the algae killing action, it is preferable that the polysaccharide decomposing enzyme has cellulose decomposing activity such as cellulase. Examples of enzymes having cellulose decomposing activity include exo-type cellobiohydrolase (EC 3.2.1.91), endo-type endoglucanase (EC 3.2.1.4), and β-glucosidase (EC 3.2.1.21). It is preferable to use at least two of these enzymes in combination. From the viewpoint of a more excellent algae killing action, it is preferable to use a mixture of cellulase and β-glucosidase derived from the genus Trichoderma. It is more preferable to use a mixture of cellulase derived from the genus Cellulomonas and cellulase and β-glucosidase derived from the genus Trichoderma in combination.

When using a mixture of cellulase derived from the genus Cellulomonas and cellulase derived from the genus Trichoderma with β-glucosidase in combination, the mass ratio is set appropriately, but is preferably 1:1 to 1000, and more preferably 1:5 to 500.

An example of a polysaccharide-degrading enzyme derived from the genus Cellulomonas is cellulase derived from Cellulomonas sp. K32A. An example of Cellulomonas sp. K32A is a strain with the accession number FERM BP-6766. An example of a polysaccharide-degrading enzyme derived from the genus Trichoderma is cellulase derived from Trichoderma reesei, and Sucrase C (manufactured by Mitsubishi Chemical Foods Corporation) derived from Trichoderma longibrachiatum. A composition containing an exo- or endo-type cellulase derived from Cellulomonas sp. K32A, and a mixture of an exo- or endo-type cellulase derived from Trichoderma reesei and β-glucosidase may also be applied.

The polysaccharide-degrading enzyme may be a publicly known product, a commercially available product, or the like as described above, or may be crudely extracted or purified from animals, plants, microorganisms, or culture supernatants of microorganisms using a publicly known method. In addition, the polysaccharide-degrading enzyme may be obtained by a publicly known method from DNA encoding the polysaccharide-degrading enzyme, for example, by using an in vitro protein synthesis system in which expression is achieved by introducing an expression vector such as a plasmid into a microorganism such as E. coli or yeast.

(Algae)
The algae-killing agent of this embodiment is applied to algae that live in aquatic environments. Specific examples of algae include, for example, green algae, gray algae, red algae, etc. as Archaeplastida, zygophyte (Zygoscelis), diatoms, brown algae, pinguiophytes, and true eyespot algae, etc. as Stramenopiles, Apicomplexa and Dinoflagellates, etc. as Alveolata, Chlorarachniophyte, etc. as Rhizaria, Haptophyte, Cryptophyte, Euglena, etc. as Excavata, and blue-green algae. Specific examples of green algae include, for example, Acrogyria and Fushimadara. Specific examples of zygophyte include, for example, Spirogyra, etc. Specific examples of blue-green algae include, for example, Blue-green algae. In particular, the agent exerts an excellent algae-killing effect on green algae and zygophyte that live in rice paddies.

Second Embodiment
A second embodiment of the algae removal method of the present invention will now be described.
The algae removal method of the present invention is a method of administering the algae-removing agent of the first embodiment to an aquatic environment that algae live in. In the algae removal method, the aquatic environment is not particularly limited as long as it is an aquatic environment in which algae live, and examples thereof include rice paddies, aquariums, ponds, lakes, the sea, waterways, and pools.

The concentration of the polysaccharide-degrading enzyme when the algaecide is administered to the aquatic environment is set as appropriate, but is preferably 0.1 μg/L or more from the viewpoint of efficient efficacy. The algaecide can be administered in a temperature environment in which algae can survive, for example, at 0 to 50°C, but it is preferable to administer the algaecide in a temperature environment in which the polysaccharide-degrading enzyme acts efficiently, for example, at 20 to 35°C.

The action and effect of the algae-killing agent and algae-killing method of the above embodiment will be described.
(1) In the above embodiment, the algae-killing component contains a polysaccharide-degrading enzyme that decomposes the cell walls of algae that live in aquatic environments. Therefore, the composition is easy to use and can exert an excellent algae-killing effect.

(2) Furthermore, by using polysaccharide-degrading enzymes derived from natural ingredients, safety can be further improved. Furthermore, the impact on the natural environment can be further reduced. Furthermore, the accumulation of residual substances can be suppressed due to the natural decomposition of the enzymes (proteins).

(3) The above embodiment has a mucilaginous layer in the cell wall and exerts an excellent algae-killing effect against algae that do not contain a cuticle. Even if the algae-killing agent is applied to the stems and leaves of other plants, such as agricultural crops such as rice, it does not affect their growth, and does not adversely affect the aquatic environment or the activity of organisms other than the target organism. Therefore, it has excellent selectivity for the algae-killing action.

(4) The algaecide of the above embodiment can reduce the hindrance to seedling growth, the exploitation of fertilizer nutrients, the obstruction of sunlight, the obstruction of water flow, etc., caused by the excessive proliferation of aquatic algae that occurs particularly in rice paddies in early summer.
The above embodiment can be modified as follows: The above embodiment and the following modifications can be combined with each other as long as they are not technically inconsistent.

- The algaecide of the above embodiment may be mixed with known algaecides as long as the effects of the present invention are not impaired. However, from the perspective of further improving safety, it is preferable that the algaecide does not contain a chemically synthesized algaecide.

- The algaecide of the above embodiment is applied to algae that live in aquatic environments, but it can be applied directly to growing algae, or it can be sprayed in advance on algae breeding sites to prevent the growth of algae.

Next, the above embodiment will be described more specifically with reference to examples and comparative examples.
<Test Example 1: Test on algae killing effect of algae killing agent>
The algae-killing effect of polysaccharide-degrading enzymes on algae living in aquatic environments was tested.

(Polysaccharide degrading enzyme)
As the polysaccharide decomposition enzyme, composition A containing endo- or exo-type cellulase derived from Cellulomonas sp. K32A and a mixture of endo- or exo-type cellulase and β-glucosidase derived from Trichoderma reesei in a mass ratio of 1:200 to 250 was used. Also, composition B containing endo- or exo-type cellulase derived from Cellulomonas sp. K32A and a mixture of endo- or exo-type cellulase and β-glucosidase derived from Trichoderma reesei in a mass ratio of 1:6 to 9 was used.

(Algae)
Spirogyra and Polytrichum commune were collected from rice paddies in June and kept in water at a temperature of 22-24°C.

(test)
In Example 1, 4 μL of composition A was added to 200 mL of water to prepare an enzyme solution with an enzyme concentration of 0.77 μg/L.

In Example 2, 40 μL of composition A was added to 200 mL of water to prepare an enzyme solution with an enzyme concentration of 7.7 μg/L.
In Example 3, 400 μL of composition A was added to 200 mL of water to prepare an enzyme solution with an enzyme concentration of 77 μg/L.

In Example 4, composition B was added to 200 mL of water to prepare an enzyme solution with an enzyme concentration of 570 μg/L.
In Example 5, a mixture of endo- or exo-cellulase derived from Trichoderma reesei and β-glucosidase was added to 200 mL of water to prepare an enzyme solution with an enzyme concentration of 250 μg/L.

The control group was water without addition of enzyme.
First, about 200 mL of each of the enzyme solutions was added to a beaker of a given volume. Spirogyra or Polytrichum commune in good condition was placed in the solution, and the water temperature was set to 22-24°C. The hours of sunlight were not controlled. The observation period was 10 days for Spirogyra and 8 days for Polytrichum commune.

(Algae removal evaluation)
The appearance of each test group was visually observed every day. The appearance of Spirogyra changed significantly, so it was magnified and observed under a microscope. Each example group was evaluated in comparison with the control group according to the following criteria. The evaluation method was as follows: ○ if the color turned brown or most of the plants were necrotic, △ if there was poor growth compared to the control, and × if there was no change compared to the control. The results are shown in Table 1. Figure 1 shows a photograph of the appearance of a part of the enzyme-treated Spirogyra transferred to a petri dish. Figure 2 shows a microscopic photograph of the enzyme-treated Spirogyra magnified 150 times.

As shown in Table 1 and Figures 1 and 2, in each Example, it was confirmed that the cellulolytic activity caused decolorization or necrosis of most of the algae living in the aquatic environment.

Furthermore, it has been confirmed that when the polysaccharide-degrading enzyme used in each example group is administered to rice seedlings, no changes are observed in the growth rate, size, etc., and that it has no effect on normal growth.
Next, the technical ideas that can be understood from the above-described embodiment and other examples will be described below together with their effects.

(A) The algaecide does not contain a chemically synthesized algaecide. Therefore, according to the invention described in (A), the burden on the natural environment can be further reduced and safety can be further improved.
(ii) The algaecide does not contain starch decomposing enzymes, but contains cellulase derived from the genus Cellulomonas and cellulase derived from the genus Trichoderma. Therefore, according to the invention described in (ii), the algaecide can efficiently exert an algaecidal effect using only the specific cellulase.

Claims (4)

It contains polysaccharide-degrading enzymes that break down the cell walls of algae that live in aquatic environments.
The algaecide (excluding application as an anti-algae paint composition) is characterized in that the polysaccharide-degrading enzyme contains at least one selected from cellulase derived from the genus Cellulomonas, cellulase derived from the genus Trichoderma, and β-glucosidase derived from the genus Trichoderma, and the aquatic environment is a rice paddy, or a pond, lake, or waterway leading to the rice paddy. The algaecide according to claim 1, characterized in that the polysaccharide degrading enzyme is an endo-type or exo-type polysaccharide degrading enzyme. The algaecide according to claim 1 or 2, wherein the algae is at least one selected from the group consisting of green algae, zygophytes, glaucophytes, red algae, diatoms, brown algae, pinguiophytes, euphthalmophytes, apicomplexa, dinoflagellates, chlorarachniophytes, haptophytes, cryptophytes, euglenids, and cyanobacteria. An algae removal method comprising administering the algae-removing agent according to any one of claims 1 to 3 to an aquatic environment in which algae live, the aquatic environment being a paddy field, or a pond, lake, or waterway leading to the paddy field.