Plant growth promoter

Abstract

Provided are a method and a material which are for promoting the growth of leguminous plants that are pulses.　This growth promoter for leguminous plants that are pulses is obtained by combining (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol, wherein: the (B) antioxidant is at least one substance selected from among tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole; and the ratio (mass ratio) of the (A) ascorbic acid or a salt thereof is 10-3,000,000 when the amount of the component (B) is 1.

Description

Plant growth promoter

[0001] The present invention relates to methods and materials for promoting the growth of legumes, including pulses.

[0002] Approximately one-third of the Earth's land area is arid, and future global warming is expected to further increase this aridity. To address the serious food shortages caused by population growth, there is an urgent need to develop technologies to improve, maintain, and increase yields of soybeans, rice, wheat, corn, and other grains in arid, saline, high-temperature, and low-temperature regions—regions where growth is difficult or where growth is impaired and yields are reduced. Soybeans, in particular, are an important grain and are widely consumed around the world, including in Japan. Unlike other grains, soybeans have high protein and lipid content and are highly nutritious. Therefore, they are also important as feed and a source of oils and fats, and technologies to increase yields are being developed.

[0003] Oxygen is an essential substance for the survival of many plants and animals, but at the same time, oxygen generates highly reactive oxygen species within the cells of plants and animals, which cause serious damage to living organisms by damaging genes and deactivating enzymes. For this reason, plants store many antioxidants, including L-ascorbic acid, and a complex enzyme system works in various parts of the cell to use these antioxidants to eliminate reactive oxygen species.

[0004] From this perspective, antioxidants containing ascorbic acid are often incorporated into materials applied to plants to promote their growth. For example, Non-Patent Document 1 discloses the growth-promoting and yield-increasing effects of ascorbic acid application to olives, sugarcane, wheat, and other crops. Non-Patent Document 2 discloses that a maximum yield increase of 30% can be achieved by foliar spraying a fixed concentration of ascorbic acid dissolved in distilled water onto legume crops three times during the vegetative or reproductive growth phase. Patent Document 1 also discloses the promotion of plant growth using an aqueous composition containing an iron (II) compound and L-ascorbic acid, and Patent Document 2 discloses the promotion of plant growth using an aqueous solution containing glycyrrhizin and L-ascorbic acid.

[0005] [Patent Document 1] Japanese Patent Application Laid-Open No. 60-202805 [Patent Document 2] Japanese Patent Application Laid-Open No. 08-143406

[0006] [Unauthorized document 1] Akram et al., (2017) Ascorbic Acid-A Potential Oxidant Scavenger and Its Role in Plant Development and Abiotic Stress Tolerance. Frontiers in Plant Science, 8:613 [Unauthorized document 2] Zarghamnejad et al., (2014) (Chickpea response to ascorbic acid foliar application at vegetative and reproductive stages. International Journal of Biosciences, 5: 166-170

[0007] The present invention relates to the following 1) to 9): 1) A growth promoter for leguminous plants included in pulses, comprising a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol, wherein the (B) antioxidant is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000, when the component (B) is taken as 1. 2) A root nodule activity promoter, comprising a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol, wherein the (B) antioxidant is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000, when the component (B) is taken as 1. 3) A yield-enhancing agent for legumes included in pulses, comprising a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol, wherein the (B) antioxidant is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000, when component (B) is taken as 1. 4) A growth-promoting method for legumes included in pulses, comprising a step of combining (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol with water, and applying the resulting combination to soil or plants, wherein the (B) antioxidant is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000, when component (B) is taken as 1. 5) A method for promoting nodule activity, comprising the step of applying a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol with water to soil or a plant, wherein the (B) antioxidant is at least one selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the component (B) is taken as 1.6) A method for increasing the yield of legumes included in pulses, comprising a step of applying to soil or plants a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol, with water, wherein the antioxidant (B) is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000, when component (B) is taken as 1. 7) Use of a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol, for producing a growth promoter for legumes included in pulses, wherein the antioxidant (B) is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000, when component (B) is taken as 1. 8) Use of a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol for producing a root nodule activity promoter, wherein the (B) antioxidant is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the component (B) is taken as 1. 9) Use of a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol for producing a yield-enhancing agent for leguminous plants included in pulses, wherein the (B) antioxidant is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the component (B) is taken as 1.

[0008] Yield-increasing effect on kidney beans: Plot 1: control, Plot 2: ascorbic acid (tap water), Plot 3: ascorbic acid (ion-exchanged water), Plot 4: ascorbic acid + BHT + glycerin + sorbitan monolaurate + silica (tap water). Effect on improving nodule activity on soybeans: Plot 1: control, Plot 2: ascorbic acid (tap water), Plot 3: ascorbic acid + BHT + diethylene glycol + sorbitan monolaurate (tap water). Yield-increasing effect on soybeans (green soybeans): Plot 1: control, Plot 2: ascorbic acid (well water), Plot 3: ascorbic acid + BHT + glycerin + sorbitan monolaurate + silica (well water). Yield-increasing effect on lotus grass: Plot 1: control, Plot 2: ascorbic acid (tap water), Plot 3: ascorbic acid + BHT + glycerin + sorbitan monolaurate + silica (tap water). Growth-promoting effect on soybeans. Compartment 1: control, compartment 2: ascorbic acid (tap water), compartment 3: ascorbic acid + BHT + glycerin (tap water), compartment 4: ascorbic acid + BHT + propylene glycol (tap water), compartment 5: sodium ascorbate + tocopherol + glycerin (tap water). Detailed Description of the Invention

[0009] The present invention relates to providing methods and materials for promoting the growth of legumes, including pulses.

[0010] The present inventors have investigated the use of ascorbic acid to promote plant growth and have found that when an ascorbic acid solution dissolved in water used in agricultural fields, such as tap water or well water, is sprayed on legumes, including pulses, the growth-promoting effect of ascorbic acid is hardly obtained. They have also found that the use of ascorbic acid in combination with a polyol and a specific antioxidant can exert the effects of promoting nodule activity and growth.

[0011] According to the method of the present invention, the growth of legumes included in pulses can be increased. Furthermore, nodule activity is promoted in legumes included in pulses that have formed nodules, improving the ureide-producing ability of the plant. That is, according to the present invention, the yield of fruits, seeds, or grains of legumes included in pulses can be increased.

[0012] In the context of the growth promoter for legumes included in pulses of the present invention, "growth promotion" means increasing the growth amount (fresh weight, elongation, etc.) of legumes included in pulses and increasing the yield of fruits, seeds, or grains. When the growth promoter for legumes included in pulses of the present invention is intended solely to increase grain yield, it is referred to as a "yield-increasing agent for legumes included in pulses." In the present invention, legumes (Fabaceae) included in pulses include soybeans (including edamame), adzuki beans, chickpeas, lotus root, kidney beans, peanuts, broad beans, peas, scarlet beans, lima beans, mung beans, cowpeas, black-eye beans, hyacinth beans, jack beans, lentils, winged beans, and other so-called "pulses" whose seeds are harvested and used. As legumes included in pulses, soybean, adzuki bean, kidney bean, pea, broad bean, chickpea, and lotus grass are preferred, and soybean is more preferred.

[0013] In the context of the nodule activity promoter of the present invention, "promotion of nodule activity" means promoting nodule activity in nodule-forming plants, i.e., the nitrogen fixation function in a host plant exerted by nodules. Here, "nodule" refers to a nodule that forms on the root of a plant through symbiosis with bacteria (rhizobia). Rhizobia reduce atmospheric nitrogen in the nodules, converting it into ammonia nitrogen, which is then supplied to the host, thereby performing so-called symbiotic nitrogen fixation. "Nodule-forming plant" refers to a host plant on which nodules have formed, and in the present invention, preferred examples include legumes, which are included in pulses.

[0014] The legume growth promoter or nodule activity promoter encompassed by the pulses of the present invention comprises a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol. The combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol is achieved by pre-mixing components (A), (B), and (C) in an appropriate mass ratio or by mixing them at the time of use. In one embodiment, components (A), (B), and (C) are combined to form a single formulation (single-component formulation (composition)). In another embodiment, for example, a formulation containing component (A) and a formulation containing components (B) and (C) are prepared separately and then combined at the time of use to form a two-component formulation (kit).

[0015] In the present invention, the IUPAC systematic name of ascorbic acid, which is component (A), is (R)-3,4-dihydroxy-5-((S)-1,2-dihydroxyethyl)furan-2(5H)-one. Ascorbic acid may be in any of the D-, L-, and DL-forms, but the L-form (so-called L-ascorbic acid) is preferred. Commercially available ascorbic acid of various grades can be used. Examples of salts of ascorbic acid include sodium salts, potassium salts, calcium salts, magnesium salts, barium salts, ammonium salts, and salts with nitrogen-containing organic bases such as pyridine, trimethylamine, triethylamine, tributylamine, and diethylamine.

[0016] When applying the legume growth promoter or root nodule activity promoter of the present invention to plants, the concentration of (A) ascorbic acid or a salt thereof in the composition combining the various components can be adjusted appropriately depending on the supply method, for example, within a range of 100 ppm by mass or more and 300,000 ppm by mass or less. For example, when sprayed using a sprayer (e.g., a boom sprayer), the concentration in the spray solution is preferably 100 ppm by mass or more, more preferably 300 ppm by mass or more, even more preferably 500 ppm by mass or more, and preferably 20,000 ppm by mass or less, more preferably 10,000 ppm by mass or less, and even more preferably 4,500 ppm by mass or less. Also, 100 to 20,000 ppm by mass is preferred, more preferably 300 to 10,000 ppm by mass, and even more preferably 500 to 4,500 ppm by mass. In the case of aerial spraying, the concentration in the spray solution is preferably 20,000 mass ppm or more, more preferably 80,000 mass ppm or more, and even more preferably 150,000 mass ppm or more, and is preferably 300,000 mass ppm or less, more preferably 250,000 mass ppm or less, and even more preferably 200,000 mass ppm or less. Also, 20,000 to 300,000 mass ppm is preferred, more preferably 80,000 to 250,000 mass ppm, and even more preferably 150,000 to 200,000 mass ppm is preferred.

[0017] In the present invention, the antioxidant, component (B), is specifically at least one selected from tocopherol (vitamin E), dibutylhydroxytoluene (BHT), and butylhydroxyanisole (BHA). Among these, the phenolic antioxidants BHT and BHA are preferred, and BHT is more preferred. In the present invention, antioxidants other than tocopherol, BHT, and BHA, such as sodium erythorbate, propyl gallate, sodium sulfite, potassium sulfite, potassium pyrosulfite, chlorogenic acid, catechin, glutathione, uric acid, etc., preferably one or more selected from sodium sulfite, potassium sulfite, potassium pyrosulfite, chlorogenic acid, catechin, glutathione, and uric acid, can also be used in combination. Therefore, a preferred embodiment is to use, as the antioxidant of component (B), one or more phenolic antioxidants selected from BHT and BHA in combination with one or more antioxidants selected from sodium sulfite, potassium sulfite, glutathione, and uric acid.

[0018] When ascorbic acid is dissolved in water containing metal ions, such as tap water, the metal ions react with ascorbic acid to generate hydrogen peroxide, which then undergoes a Fenton reaction with the metal ions to generate hydroxyl radicals. When the hydroxyl radicals enter the plant, they react with lipids present in the cell membrane, triggering a series of lipid peroxidation reactions, generating lipid radicals and lipid peroxy radicals. The use of antioxidants can sequester these radicals.

[0019] In the present invention, the antioxidant (B) is used in the form of a solution dissolved in the polyol (C), thereby effectively exerting the plant growth-promoting or root nodule activity-promoting effect of ascorbic acid. The legume growth promoter or root nodule activity promoter encompassed by the pulses of the present invention may be either a single- or two-component formulation, as described above. It may also be used in a two-component formulation (kit) in which a formulation containing component (A) (first formulation) and a formulation containing components (B) and (C) (second formulation) are separately prepared and combined, and the two formulations are mixed at the time of use. The two-component formulation can suppress discoloration that occurs during storage of a formulation containing all of components (A), (B), and (C). In this case, the first formulation containing component (A) may contain one or more antioxidants other than tocopherol, BHT, and BHA, such as sodium sulfite, potassium sulfite, potassium pyrosulfite, chlorogenic acid, catechin, glutathione, and uric acid.

[0020] When the legume growth promoter or nodule activity promoter of the present invention is applied to a plant, the concentration of the antioxidant (B) in the composition obtained by combining the components is preferably 0.001 ppm by mass or more, more preferably 0.01 ppm by mass or more, and even more preferably 0.1 ppm by mass or more, and is preferably 100 ppm by mass or less, more preferably 20 ppm by mass or less, and even more preferably 5 ppm by mass or less, and is preferably 0.001 to 100 ppm by mass, more preferably 0.01 to 20 ppm by mass, and even more preferably 0.1 to 5 ppm by mass.

[0021] The polyol, which is component (C), is a solvent used to dissolve the antioxidant (B). Specific examples include dihydric alcohols such as ethylene glycol, diethylene glycol, hexylene glycol, polyethylene glycol, propylene glycol, isoprene glycol, 1,3-propanediol, dipropylene glycol, and polypropylene glycol; trihydric or higher alcohols such as glycerin, diglycerin, and triglycerin; and sugars or sugar alcohols such as erythritol, pentaerythritol, maltitol, xylitol, sorbitan, and sorbitol. Preferred are ethylene glycol, diethylene glycol, propylene glycol, dipropylene glycol, and glycerin, and more preferred are diethylene glycol, propylene glycol, and glycerin.

[0022] When one or more selected from BHT and BHA are used as the antioxidant (B), it is preferable to use diethylene glycol, propylene glycol, glycerin, or a mixture thereof as the polyol (C).

[0023] When applying the legume growth promoter or root nodule activity promoter of the present invention to plants, the concentration of the polyol (C) in the composition combining the various components can be adjusted appropriately depending on the supply method within a range of 1 ppm by mass or more and 100,000 ppm by mass or less. For example, when spraying using a sprayer (e.g., a boom sprayer), the concentration in the spray solution is preferably 1 ppm by mass or more, more preferably 10 ppm by mass or more, even more preferably 50 ppm by mass or more, and preferably 10,000 ppm by mass or less, more preferably 5,000 ppm by mass or less, and even more preferably 1,000 ppm by mass or less. Also, the concentration is preferably 1 to 10,000 ppm by mass, more preferably 10 to 5,000 ppm by mass, and even more preferably 50 to 1,000 ppm by mass. In the case of aerial spraying, the concentration in the spray solution is preferably 100 ppm by mass or more, more preferably 1,000 ppm by mass or more, and even more preferably 5,000 ppm by mass or more, and is preferably 100,000 ppm by mass or less, more preferably 50,000 ppm by mass or less, and even more preferably 10,000 ppm by mass or less. Also, the concentration is preferably 100 to 100,000 ppm by mass, more preferably 1,000 to 50,000 ppm by mass, and even more preferably 5,000 to 10,000 ppm by mass.

[0024] The ratio (mass ratio) of the combination of (A) ascorbic acid or a salt thereof, (B) antioxidant, and (C) polyol, when component (B) is taken as 1, is preferably 10 or more, more preferably 100 or more, even more preferably 500 or more, and is preferably 3,000,000 or less, more preferably 2,000,000 or less, even more preferably 200,000 or less. Also, it is preferably 10 to 3,000,000, more preferably 100 to 2,000,000, even more preferably 500 to 200,000. Of these, when spraying with a sprayer, 500 to 45,000 is preferred, when spraying by aerial spraying, 150,000 to 2,000,000 is preferred, and further, 500 to 4,500 is more preferred when spraying with a sprayer, and 150,000 to 200,000 is preferred when spraying by aerial spraying. Furthermore, when component (B) is taken as 1, component (C) is preferably 4 or more, more preferably 10 or more, even more preferably 20 or more, even more preferably 50 or more, and preferably 100,000 or less, more preferably 50,000 or less, even more preferably 20,000 or less, even more preferably 10,000 or less. Furthermore, it is preferably 4 to 100,000, more preferably 10 to 50,000, even more preferably 20 to 20,000, even more preferably 50 to 10,000. Among these, in the case of spraying by a sprayer, 20 to 10,000 is preferable, and in the case of spraying by aerial spraying, 2,000 to 100,000 is preferable. Furthermore, in the case of spraying by a sprayer, 50 to 1,000 is more preferable, and in the case of spraying by aerial spraying, 5,000 to 10,000 is more preferable.

[0025] In the present invention, (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol can be used in combination with (D) a surfactant. The use of a surfactant improves the wettability, adhesion, and penetration of (A) ascorbic acid or a salt thereof onto the plant surface, thereby enhancing the effects of (A) ascorbic acid or a salt thereof or enabling the effects to be more efficiently exerted. As described above, when the legume growth promoter or root nodule activity promoter of the present invention is formulated as a two-component formulation (kit) combining a formulation containing component (A) (first agent) and a formulation containing components (B) and (C) (second agent), it is preferable that the surfactant (D) be contained in the second agent containing components (B) and (C). Examples of surfactants include nonionic surfactants, ionic surfactants, and amphoteric surfactants, and these can be used singly or in combination of two or more.Examples of the nonionic surfactant include one or more selected from sorbitan fatty acid esters, glycerin fatty acid esters, polyoxyalkylene sorbitan fatty acid esters, polyoxyethylene fatty acid esters, polyglycerin fatty acid esters, polyalkylene glycol fatty acid esters, polyoxyethylene resin acid esters, polyoxyethylene alkyl ethers, polyoxyethylene alkylaryl ethers, polyoxyalkylene aryl ethers, polyoxyethylene alkenyl ethers, alkyl polyglycosides, polyoxyalkylene alkyl polyglycosides, and sucrose fatty acid esters, Examples of the anionic surfactant include one or more selected from alkyl sulfates, alkylbenzene sulfonates, polyoxyalkylene alkyl ether sulfates, polyoxyalkylene alkenyl ether sulfates, polyoxyalkylene alkylaryl ether sulfates, fatty acid salts, pyrophosphates, lauryl phosphoric acid, polycarboxylic acid polymers, polyoxyethylene alkylene alkyl acetates, aromatic sulfonate formalin condensates, polyoxyethylene distyrenated ether sulfates, alkyl diphenyl ether disulfonates, dialkyl sulfosuccinates, and alkyl naphthalene sulfonates. Examples of the amphoteric surfactant include one or more selected from 2-alkyl-N-carboxymethyl-N-hydroxyethyl imidazolinium betaine, alkyl dimethylaminoacetic acid betaine, and alkyl acid amidopropyl betaine.Among these, from the viewpoint of preventing phytotoxicity to plants when excessively applied, the nonionic surfactant is preferably one or more selected from sorbitan fatty acid esters, glycerin fatty acid esters, polyoxyalkylene sorbitan fatty acid esters, polyalkylene glycol fatty acid esters, polyoxyethylene resin acid esters, polyoxyethylene alkyl ethers, alkyl polyglycosides, and sucrose fatty acid esters, and more preferably one or more selected from sorbitan fatty acid esters, glycerin fatty acid esters, polyalkylene glycol fatty acid esters, polyoxyethylene resin acid esters, and polyoxyethylene alkyl ethers; the anionic surfactant is preferably one or more selected from alkyl sulfates (e.g., sodium lauryl sulfate, ammonium lauryl sulfate, triethanolamine lauryl sulfate, etc.), aromatic sulfonate formalin condensates, and fatty acid salts; and the amphoteric surfactant is preferably 2-alkyl-N-carboxymethyl-N-hydroxyethylimidazolinium betaine.

[0026] When applying the legume plant growth promoter or root nodule activity promoter included in the pulses of the present invention to plants, the concentration of the surfactant (D) in the composition combining the respective components can be adjusted appropriately depending on the supply method within a range of 10 ppm by mass or more and 30,000 ppm by mass or less. For example, when spraying using a sprayer (e.g., a boom sprayer), the concentration in the spray solution is preferably 10 ppm by mass or more, more preferably 100 ppm by mass or more, even more preferably 200 ppm by mass or more, and preferably 5,000 ppm by mass or less, more preferably 1,000 ppm by mass or less, and even more preferably 500 ppm by mass or less. Also, 10 to 5,000 ppm by mass is preferred, more preferably 10 to 1,000 ppm by mass, and even more preferably 100 to 500 ppm by mass. When sprayed aerially, the concentration in the spray solution is preferably 100 ppm by mass or more, more preferably 500 ppm by mass or more, and even more preferably 1,000 ppm by mass or more, and is preferably 10,000 ppm by mass or less, more preferably 8,000 ppm by mass or less, and even more preferably 5,000 ppm by mass or less. Also, the concentration is preferably 100 to 10,000 ppm by mass, more preferably 500 to 8,000 ppm by mass, and even more preferably 1,000 to 5,000 ppm by mass. When surfactant (D) is combined, the ratio (mass ratio) of component (D) is preferably 10 or more, more preferably 50 or more, even more preferably 100 or more, and is preferably 300,000 or less, more preferably 100,000 or less, and even more preferably 5,000 or less, relative to component (B) being 1. Also, it is preferably 10 to 300,000, more preferably 50 to 100,000, and even more preferably 100 to 5,000. Among these, in the case of spraying with a sprayer, 100 to 5,000 is preferred, and in the case of spraying by aerial spraying, 1,000 to 50,000 is preferred. Furthermore, in the case of spraying with a sprayer, 100 to 500 is more preferred, and in the case of spraying by aerial spraying, 1,000 to 5,000 is more preferred.

[0027] In addition, in the present invention, (E) a chelating agent can be used in combination with (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol. The use of a chelating agent can improve the stability of (A) ascorbic acid or a salt thereof, thereby stabilizing the effects of ascorbic acid or a salt thereof. As described above, when the legume growth promoter or root nodule activity promoter encompassed by the pulses of the present invention is formulated as a two-component formulation (kit) combining a formulation (first agent) containing component (A) with a formulation (second agent) containing components (B) and (C), the chelating agent (E) is preferably contained in the first agent containing component (A). Examples of chelating agents include aminocarboxylic acid chelating agents, phosphonic acid chelating agents, hydroxycarboxylic acid chelating agents, and polycarboxylic acid chelating agents. Examples of aminocarboxylic acid chelating agents include ethylenediaminetetraacetic acid (EDTA), nitrilotriacetic acid (NTA), hydroxyethylethylenediaminetriacetic acid (HEDTA), diethylenetriaminepentaacetic acid (DTPA), methylglycine diacetic acid (MGDA), triethylenetetraminehexaacetic acid (TTHA), glutamic acid diacetic acid (GLDA), hydroxyethyliminodiacetic acid (HIDA), dihydroxyethylglycine (DHEG), aspartic acid diacetic acid (ASDA), ethylenediaminesuccinic acid (EDDS), and salts thereof. Examples of phosphonic acid chelating agents include hydroxyethylidene diphosphonic acid (HEDP), nitrilotrismethylenephosphonic acid (NTMP), phosphonobutanetricarboxylic acid (PBTC), ethylenediaminetetramethylenephosphonic acid (EDTMP), and salts thereof. Examples of hydroxycarboxylic acid chelating agents include citric acid, malic acid, tartaric acid, gluconic acid, lactic acid, and salts thereof. Examples of polycarboxylic acid chelating agents include succinic acid, oxalic acid, glutaric acid, adipic acid, fumaric acid, malonic acid, and salts thereof.

[0028] When the legume growth promoter or root nodule activity promoter of the present invention is applied to a plant, the concentration of the chelating agent (E) in the composition obtained by combining the components is preferably 0.01 ppm by mass or more, more preferably 0.1 ppm by mass or more, and even more preferably 1 ppm by mass or more, and is preferably 100 ppm by mass or less, more preferably 50 ppm by mass or less, and even more preferably 10 ppm by mass or less. Also, it is preferably 0.01 to 100 ppm by mass, more preferably 0.1 to 50 ppm by mass, and even more preferably 1 to 10 ppm by mass. When the chelating agent (E) is combined, the ratio (mass ratio) of component (E) to component (B) is preferably 0.01 or more, more preferably 0.1 or more, even more preferably 1 or more, and is preferably 100 or less, more preferably 50 or less, and even more preferably 10 or less. Further, it is preferably 0.01 to 100, more preferably 0.1 to 100, more preferably 1 to 100, and even more preferably 1 to 10.

[0029] Suitable combinations of (B) antioxidant, (C) polyol, and (D) surfactant used together with (A) ascorbic acid include, for example, the following: a combination in which component (B) is an antioxidant containing one or more selected from BHT and BHA, component (C) is diethylene glycol or glycerin, and component (D) is a surfactant containing one or more selected from sorbitan fatty acid esters, glycerin fatty acid esters, polyalkylene glycol fatty acid esters, polyoxyethylene resin acid esters, and polyoxyethylene alkyl ethers, and optionally further containing an alkyl sulfate; a combination in which component (B) is an antioxidant containing BHT, sodium sulfite, glutathione, and uric acid, component (C) is diethylene glycol or glycerin, and component (D) is a surfactant containing one or more selected from sorbitan fatty acid esters, glycerin fatty acid esters, polyalkylene glycol fatty acid esters, polyoxyethylene resin acid esters, and polyoxyethylene alkyl ethers, and optionally further containing an alkyl sulfate.

[0030] As shown in the examples below, when soybean seeds, a nodule-forming plant, are sown and then inoculated with rhizobia for cultivation, spraying ascorbic acid in combination with an antioxidant (BHT) and a polyol increases nodule weight and nodule activity, promotes the production of ureides in the nodules, and promotes the growth of the host plant. Furthermore, spraying ascorbic acid in combination with an antioxidant (BHT) and a polyol during the cultivation of kidney beans results in increased yield. Therefore, a combination of ascorbic acid with a specific antioxidant and a polyol can serve as a growth promoter or nodule activity promoter for legumes, which can be used to promote the growth or nodule activity of legumes, and can also be used to produce a growth promoter or nodule activity promoter for legumes, which can be used. It has been reported that the amount of ureide derived from nodules correlates with grain yield (Diagnosing method for total basal nitrogen application to soybeans in rotational field, Ibaraki Prefectural Agricultural Center, Agricultural Research Institute, Major Results of 2006; http: / / www.pref.ibaraki.jp / nourinsuisan / noken / seika / h18pdf / documents / 27.pdf). Therefore, it is thought that such nodule activity promoters are useful for increasing grain yield (increasing yield).

[0031] Since the nitrogen fixation function in a host plant can be evaluated as the ureide production ability or amide production ability, the nodule activity of the present invention can be specifically evaluated as the amount of ureide or amide produced per wet nodule weight. The amount of ureide or amide produced can be calculated by measuring the amount of ureide or amide in exudate collected when the plant body is cut (for example, by cutting the above-ground part just below the cotyledons). Here, examples of ureides include allantoin, allantoic acid, and citrulline, and examples of amides include asparagine and glutamine, but it is preferable to measure the amount of allantoic acid or asparagine.

[0032] The growth promoter or nodule activity promoter for legumes included in pulses can be a composition (e.g., various agricultural or horticultural materials) for promoting the growth, increased yield, or nodule activity of legumes included in pulses, or a material (single substance) or formulation to be added to or incorporated into cultivation substrates for cultivating plants, such as soil, culture medium, or hydroponic solution. The composition may be in the form of a liquid or gel composition, or may be in the form of a solid (block, powder, granule, etc.). Here, the composition may be a composition in which components (A), (B), and (C) are mixed in advance, or a composition in which a formulation containing component (A) and a formulation containing components (B) and (C) are separately prepared and then mixed together at the time of use.

[0033] The composition may contain any optional components in addition to the components (A) to (C) and further (D) and (E) used in the present invention. Such components include solvents (e.g., water, buffer solutions, culture media, solutions for hydroponics, etc.), carriers (e.g., zeolite, silica, bentonite, Glauber's salt, diatomaceous earth, vermiculite, perlite, peat moss, activated carbon, humus, talc, clay, carbon black, pulp, straw, soybean meal, kaolin, montmorillonite, alumina, etc.), pH adjusters for promoting dissolution of the compounds, spreading agents for increasing the spreading ability of the compounds on plants or soil, fertilizer components for increasing fertilizer efficacy, pesticide components, binders, and bulking agents. Examples of suitable compounds include plant growth-promoting microorganisms such as rhizobia and mycorrhizal fungi, essential plant nutrients, flavonoids, organic acids, amino acids, peptides, nucleosides, nucleotides, nucleic acid bases, sugars, monohydric alcohols, food additives, microbial extracts, plant hormones, NOD factors, i.e., lipo-chitooligosaccharides, synthetic lipo-chitooligosaccharides, chitooligosaccharides, chitinous compounds, linoleic acid or derivatives thereof, linolenic acid or derivatives thereof, karrikin, acyl-homoserine lactone derivatives, betaine compounds, and phenolic compounds.

[0034] When the legume growth promoter or root nodule activity promoter of the present invention is used as a one-component formulation, it is preferable to incorporate one or more carriers selected from zeolite, silica, bentonite, and sodium sulfate in order to improve the storage stability or inhibit browning of the formulation, and it is more preferable to incorporate silica in order to improve storage stability and inhibit browning. In this case, the concentration of the carrier (component (F)) in the composition is preferably 50 ppm by mass or more, more preferably 100 ppm by mass or more, and even more preferably 250 ppm by mass or more, and preferably 1,000 ppm by mass or less, more preferably 750 ppm by mass or less, and even more preferably 650 ppm by mass or less. It is also preferably 50 to 1,000 ppm by mass, more preferably 100 to 750 ppm by mass, and even more preferably 250 to 650 ppm by mass. When the (F) carrier is combined, the mass ratio of component (F) is preferably 50 or more, more preferably 100 or more, even more preferably 250 or more, and is preferably 1,000 or less, more preferably 750 or less, even more preferably 650 or less, when component (B) is taken as 1. The mass ratio is also preferably 50 to 1,000, more preferably 100 to 750, even more preferably 250 to 650.

[0035] Compositions for aerial application may also contain emulsifiers such as polyoxyethylene fatty acid esters and oils such as decyl alcohol.

[0036] Examples of the composition include, but are not limited to, cultivation substrates (e.g., agricultural or horticultural soil, culture medium, culture solution for hydroponics, water, etc.) containing at least components (A) to (C) of the present invention, fertilizers, water for watering, microbial materials such as rhizobia materials, soil conditioners, pesticides, sowing materials, and plant supplements (e.g., activators, nutrients, etc.).

[0037] The fertilizers, microbial materials, soil conditioners, sowing materials, and plant supplements are preferred because they contribute to improving the soil in which plants are grown. The fertilizers, microbial materials, soil conditioners, sowing materials, and plant supplements may be solid or liquid, and if solid, may be in the form of blocks, powders, granules, etc., but are preferably powders or granules. In addition to containing components (A) to (C) as active ingredients, the fertilizers, microbial materials, soil conditioners, sowing materials, and plant supplements may also contain components of fertilizers, microbial materials, soil conditioners, sowing materials, and plant supplements that are typically used in plant cultivation.

[0038] The cultivation substrate, fertilizer, microbial materials such as rhizobia materials, soil conditioners, pesticides, sowing materials, and plant supplements may be prepared by adding components (A) to (C) of the present invention to ordinary cultivation substrates (e.g., agricultural or horticultural soil, culture medium, culture medium, solution for hydroponics, water, etc.), fertilizer, microbial materials such as rhizobia materials, soil conditioners, pesticides, sowing materials, plant supplements (e.g., activators, nutrients, etc.), and the like.

[0039] The composition may be one in which a formulation containing component (A) and a formulation containing components (B) and (C) are prepared separately and then mixed together at the time of use. In the case of a two-component formulation in which a formulation containing component (A) (first agent) and a formulation containing components (B) and (C) (second agent) are mixed together at the time of use, the weight ratio of the first agent, relative to the weight ratio of the second agent, is 1 to 100, preferably 1 to 50, and more preferably 1 to 25. When the first agent and the second agent are prepared by dissolving them in water, respectively, the concentration of the first agent in the aqueous solution is preferably 100 ppm by mass or more and 300,000 ppm by mass or less, and the concentration of the second agent in the aqueous solution is preferably 10 ppm by mass or more and 100,000 ppm by mass or less.

[0040] The method of supplying the legume growth promoter or root nodule activity promoter included in the pulses of the present invention is not particularly limited, as long as it is applied to plants so as to achieve the effects of the present invention. That is, as long as the composition contacts or is delivered to the plant body of the legume included in the pulses or the soil in the rhizosphere of the plant, there are no particular limitations. Examples include surface spraying on soil, irrigation, plowing, foliar spraying on plants, application by mixing with fertilizer, addition to hydroponic solution, or application or smearing on seeds before sowing (e.g., seed dressing), etc., but it is preferable to apply the components of the present invention in the form of a spray solution diluted with water, and foliar spraying is particularly preferred. The spray solution may be prepared at the time of application, and the dilution water used in this case may be any of agricultural water, well water, groundwater, river water, lake water, tap water, etc.

[0041] The application method is not particularly limited, and examples thereof include a spray method, i.e., a method of spraying the application liquid in the form of a mist by spraying. According to such a method, the legume plant growth promoter or root nodule activity promoter included in the pulses of the present invention adheres to the plant and then exhibits good spreadability on the plant. Specific examples of spraying methods include manual spraying using a mister, sprayer, sprayer (e.g., boom sprayer), etc., and aerial spraying using an airplane, helicopter, drone, etc.

[0042] The application amount of the legume growth promoter or root nodule activity promoter included in the pulses of the present invention depends on the concentrations of components (A) to (C) contained in the composition at the time of application. For example, when the concentration of component (A) contained in the spray solution is 100 to 300,000 mass ppm, the amount of component (A) used per plant in the composition is preferably 1 mg or more, more preferably 5 mg or more, more preferably 10 mg or more, and preferably 150 mg or less, more preferably 100 mg or less, more preferably 50 mg or less. Also, the amount is preferably 1 to 150 mg, more preferably 5 to 100 mg, more preferably 10 to 50 mg. The legume growth promoter or root nodule activity promoter included in the pulses of the present invention may be applied in an amount within the above range at once or in multiple applications.

[0043] The timing and frequency of application may vary depending on the type of legume included in the beans, but in the case of soybeans, when application is made by surface spraying, irrigation, plowing into a cultivation substrate such as soil, or seed dressing, it is usually preferable to apply the compound once or 1 to 3 times before or simultaneously with sowing, and when application is made after sowing, it is preferable to apply the compound between the early stage of vegetative growth before the start of the reproductive growth period and the grain filling period after the start of the reproductive growth period.

[0044] In relation to the above-mentioned embodiments, the present invention further discloses the following aspects: <1> A growth promoter for legumes included in pulses, comprising a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol, wherein the (B) antioxidant is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000, when the component (B) is taken as 1. <2> A root nodule activity promoter, comprising a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol, wherein the (B) antioxidant is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000, when the component (B) is taken as 1. <3> A yield-enhancing agent for legumes, including pulses, comprising a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol, wherein the (B) antioxidant is at least one selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000, when the component (B) is taken as 1. <4> In <1> to <3>, the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is more preferably 100 to 2,000,000, and even more preferably 500 to 200,000, when the component (B) is taken as 1. <5> The agent according to any one of <1> to <3>, wherein the (C) polyol is one or more selected from diethylene glycol, glycerin, and propylene glycol. <6> The agent according to any one of <1> to <3>, wherein the proportion (mass ratio) of the polyol (C) is 4 to 100,000, when the proportion of the component (B) is 1. <7> In <6>, the proportion (mass ratio) of the polyol (C) is more preferably 10 to 50,000, even more preferably 20 to 20,000, and even more preferably 50 to 10,000, when the proportion of the component (B) is 1. <8> The agent according to any one of <1> to <7>, further comprising a surfactant (D) in combination.<9> The agent according to <8>, wherein the ratio (mass ratio) of the surfactant (D) is 10 to 300,000, when the ratio of component (B) is 1. <10> In <9>, the ratio (mass ratio) of the surfactant (D) is more preferably 50 to 100,000, and even more preferably 100 to 5,000, when the ratio of component (B) is 1. <11> The agent according to any one of <8> to <10>, wherein the surfactant (D) comprises one or more selected from the group consisting of nonionic surfactants, anionic surfactants, and amphoteric surfactants. <12> The agent according to <11>, wherein the (D) nonionic surfactant is one or more selected from sorbitan fatty acid esters, glycerin fatty acid esters, polyalkylene glycol fatty acid esters, polyoxyethylene resin acid esters, and polyoxyethylene alkyl ethers, the anionic surfactant is one or more selected from fatty acid salts, aromatic sulfonate-formalin condensates, and alkyl sulfates, and the amphoteric surfactant is 2-alkyl-N-carboxymethyl-N-hydroxyethylimidazolinium betaine. <13> The agent according to any one of <1> to <12>, further comprising a carrier (F). <14> The agent according to <13>, wherein the ratio (mass ratio) of the carrier (F) is 50 to 1,000, relative to the ratio of component (B) being 1. <15> In <14>, the ratio (mass ratio) of the carrier (F) is more preferably 100 to 750, and even more preferably 250 to 650, relative to the ratio of component (B) being 1. <16> The agent according to any one of <13> to <15>, wherein (F) the carrier is one or more selected from zeolite, silica, bentonite, and sodium sulfate. <17> The agent according to any one of <1> to <16>, further comprising (E) a chelating agent in combination.

[0045] <18> A method for promoting the growth of leguminous plants included in pulses, comprising a step of combining (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol with water and applying the resulting mixture to soil or plants, wherein the (B) antioxidant is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the proportion (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the proportion of component (B) is 1. <19> A method for promoting root nodule activity, comprising a step of combining (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol with water and applying the resulting combination to soil or a plant, wherein the (B) antioxidant is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the component (B) is taken as 1. <20> A method for increasing the yield of leguminous plants included in pulses, comprising a step of combining (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol with water and applying the resulting mixture to soil or plants, wherein the (B) antioxidant is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the component (B) is taken as 1.

[0046] <21> Use of a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol for producing a growth promoter for legumes included in pulses, wherein the (B) antioxidant is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the component (B) is taken as 1. <22> Use of a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol for producing a root nodule activity promoter, wherein the (B) antioxidant is at least one or more selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the component (B) is taken as 1. <23> Use of a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol for producing a yield-enhancing agent for legumes, wherein the (B) antioxidant is at least one selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000, relative to the ratio of component (B) being 1. <24> In <21> to <23>, the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is more preferably 100 to 2,000,000, and even more preferably 500 to 200,000, relative to the ratio of component (B) being 1. <25> The use according to any one of <21> to <23>, wherein the (C) polyol is one or more selected from diethylene glycol, glycerin, and propylene glycol. <26> The use according to any one of <21> to <23>, wherein the proportion (mass ratio) of the polyol (C) is 4 to 100,000, when the proportion of the component (B) is 1. <27> In <26>, the proportion (mass ratio) of the polyol (C) is more preferably 10 to 50,000, even more preferably 20 to 20,000, and even more preferably 50 to 10,000, when the proportion of the component (B) is 1. <28> The use according to any one of <21> to <27>, further comprising a surfactant (D) in combination.<29> The use according to <28>, wherein the ratio (mass ratio) of the surfactant (D) is 10 to 300,000, when the ratio of component (B) is 1. <30> In <29>, the ratio (mass ratio) of the surfactant (D) is more preferably 50 to 100,000, and even more preferably 100 to 5,000, when the ratio of component (B) is 1. <31> The use according to any one of <28> to <30>, wherein the surfactant (D) comprises one or more selected from the group consisting of nonionic surfactants, anionic surfactants, and amphoteric surfactants. <32> The use according to <31>, wherein the (D) nonionic surfactant is one or more selected from sorbitan fatty acid esters, glycerin fatty acid esters, polyalkylene glycol fatty acid esters, polyoxyethylene resin acid esters, and polyoxyethylene alkyl ethers, the anionic surfactant is one or more selected from fatty acid salts, aromatic sulfonate-formalin condensates, and alkyl sulfates, and the amphoteric surfactant is 2-alkyl-N-carboxymethyl-N-hydroxyethylimidazolinium betaine. <33> The use according to any one of <21> to <32>, further comprising (F) a carrier. <34> The use according to <33>, wherein the ratio (mass ratio) of the (F) carrier is 50 to 1,000, relative to the ratio of component (B) being 1. <35> In <34>, the ratio (mass ratio) of the (F) carrier is more preferably 100 to 750, and even more preferably 250 to 650, relative to the ratio of component (B) being 1. <36> The use according to any one of <33> to <35>, wherein (F) the carrier is one or more selected from zeolite, silica, bentonite, and sodium sulfate. <37> The use according to any one of <21> to <35>, further comprising (E) a chelating agent in combination.

[0047] Example 1 Effect on Green Bean Yield (1) Soil Preparation and Sowing Medium-term fertilizer release soil (Takii Hydrated Cell Culture Medium-term Fertilizer Release, Takii Seed Co., Ltd.) and vermiculite granules (Akagi Engei Co., Ltd.) were mixed at a volume ratio of 1:1, and the soil was filled into polypots (10.5 cm diameter, 9 cm height). After adding 250 mL of tap water per pot, green bean seeds (variety: Satsuki Midori No. 2) were coated with 1.3 g of rhizobial seed dressing material "Mamezo" for green beans (Tokachi Agricultural Cooperative Association) per 100 mL of seeds. Two seeds were then sown per pot, approximately 1 cm below the soil surface. After germination, the seeds were thinned to one plant per pot.

[0048] (2) Cultivation conditions Cultivation was carried out indoors for 12 days after sowing, under the following conditions: 16 hours of light, 25°C, LED light source, light intensity 400 to 440 μmol / m ２ Watering was performed by adding tap water to the tray placed under the pot after the water had run out, so that the bottom 5 cm of the pot was covered.

[0049] (3) Transplanting into planters Planters (width 65.3 cm, depth 24.5 cm, height 18.5 cm) were filled with 2 L of potting soil, and then filled with 10 L of medium-term fertilizer-effect soil (Takii Hydrated Cell Culture Soil Medium-Term Fertilizer-Effect, Takii Seed Co., Ltd.). 2 L of tap water was added to each planter, and then kidney bean seedlings were transplanted 12 days after sowing. Subsequent cultivation was carried out outdoors, and watering was performed by adding the amount of water that flowed out from the bottom of the planter when the soil surface dried, with the remainder being from rainfall. Each test plot was repeated 3 times (n = 3).

[0050] (4) Foliar Spray Treatment, Harvest The spray solution shown below was prepared, and 6.7 mL was sprayed per plant using a spray bottle on the 30th day after planting of green bean seedlings. Foliar spray was performed only once, and four types of test plots were examined. The green bean seedlings were harvested from the 40th to 54th day after planting, and the weight of fresh green bean pods per plant was measured. <Spray Solution> - Plot 1: Control (no application) - Plot 2: Foliar spray of 1,500 ppm by mass of ascorbic acid (dissolved in tap water) - Plot 3: Foliar spray of 1,500 ppm by mass of ascorbic acid (dissolved in ion-exchanged water) - Plot 4: Foliar spray of 1,500 ppm by mass of ascorbic acid + 1 ppm by mass of BHT + 50 ppm by mass of glycerin + 350 ppm by mass of sorbitan monolaurate + 300 ppm by mass of silica (dissolved in tap water)

[0051] The spray solution for test area 4 was prepared as follows. First, BHT (component (B)) and glycerin (component (C)) were mixed in advance, and then sorbitan monolaurate (component (D)) was added and heated to 60°C and mixed. Ascorbic acid (component (A)) and silica gel (component (F)) were then added and mixed to prepare a powdered formulation. This formulation was dispersed in tap water to prepare a spray solution.

[0052] (5) Results Figure 1 shows the results of measuring the fresh pod weight per plant. The graph in the figure represents the mean ± standard deviation. Experimental plots 2, 3, and 4 showed average yield increases of approximately 0.9%, 44.5%, and 34.9%, respectively, compared to the control. A comparison of experimental plots 2 and 3 showed that when the same concentration of ascorbic acid was applied to green beans as foliar sprays, the effect on pod weight differed between when tap water and deionized water were used for dissolution, indicating that the effect on yield differs depending on the water quality in which ascorbic acid is dissolved. Furthermore, a comparison of experimental plots 2 and 4 showed that adding components (B), (C), and (D) to ascorbic acid resulted in increased yield even when tap water was used for dissolution.

[0053] Example 2 Evaluation of soybean nodule activity (1) Soil preparation and sowing Medium-term fertilizer release soil (Takii Hydrated Cell Soil Medium-term Fertilizer Release, Takii Seed Co., Ltd.) and vermiculite granules (Akagi Engei Co., Ltd.) were mixed at a volume ratio of 1:1, and the soil was filled into polypots (diameter 10.5 cm, height 9 cm). Soybean seeds used were "Enrei" (purchased from Nikko Seed Co., Ltd.). After adding 250 mL of tap water per pot, two seeds were sown in each pot, one seed at a depth of approximately 1 to 2 cm from the soil surface. Each test plot was repeated 6 times (n = 6).

[0054] (2) Inoculation of rhizobia Yeast-Mannitol (YM) medium (K ２ HPO ４ 0.5g MgSO ４ ・7H ２ A solid medium was prepared by adding 1.5% agar (Fujifilm Wako Pure Chemical Industries, Ltd.) to 1 L of distilled water (pH 6.8) containing 0.2 g of acetic acid, 0.1 g of NaCl, 0.4 g of yeast extract, 10 g of mannitol, and 1 L of distilled water. The Bradyrhizobium japonicum NBRC14783T strain was grown on the solid medium. A loopful of the grown rhizobia was inoculated into 50 mL of YM liquid medium in a 500 mL Sakaguchi flask and cultured with shaking at 30°C for approximately 36 hours. A rhizobia culture solution with a bacterial turbidity (OD600) of approximately 0.3 was prepared. After sowing, 1 mL of the rhizobia culture solution was dropwise inoculated onto the seeds using a micropipette.

[0055] (3) Cultivation conditions: From sowing to thinning, cultivation was carried out in an artificial climate chamber (LPH-411SP, Nippon Medical and Chemical Instruments Manufacturing Co., Ltd.), and light conditions were light (light intensity 440 to 480 μmol / m ２ The incubation period was 16 hours light / 8 hours dark, with a temperature of 30°C light / 25°C dark, and humidity of 50%. Seven days after sowing, the plants were thinned out to one per pot. Watering was performed by adding tap water to the pot after the water in the tray placed under the pot had run out, enough to cover the bottom 5 cm of the pot.

[0056] (4) Foliar spray treatment A spray solution was prepared, and on the 14th day after sowing, 6.7 mL was sprayed per plant using a spray bottle. Foliar spray was performed only once, and three types of test plots were examined. The test plots 1 to 3 evaluated are as follows. <Spray solution> - Plot 1: Control (no application) - Plot 2: Foliar spray of 1,500 ppm by mass of ascorbic acid (dissolved in tap water) - Plot 3: Foliar spray of 1,500 ppm by mass of ascorbic acid + 1 ppm by mass of BHT + 200 ppm by mass of diethylene glycol + 350 ppm by mass of sorbitan monolaurate (dissolved in tap water)

[0057] (5) Method for collecting exudate to measure ureido nitrogen content, an index of nodule activity. After 21 days of cultivation, soybean stems were cut at the cotyledonary node using pruning shears, and a 1.5 mL microtube (Eppendorf, Inc.) filled with a #10 cotton ball (Iwatsuki Co., Ltd.) was placed over the stem. Exudate exudation from the cut surface was collected for 2 hours. After exudate collection, the cotton ball was stored in a -80°C freezer. The volume of exudate was calculated by comparing the weight of the cotton ball before and after collection.

[0058] (6) Extraction of exudate components collected on cotton balls. The exudate components contained in the cotton balls were eluted with ultrapure water and filtered using a Micro Biospin Chromatography column (Bio-Rad Laboratories, Inc.). The filtrate remaining in the column was collected by centrifugation using a centrifuge (CR15RN, Hitachi, Ltd.) operated at 10,000 rpm for 1 minute. The filtrate was diluted 3,000 times and quantified.

[0059] (7) Apparatus and measurement method used for quantification of ureido nitrogen content: The HPLC system and mass spectrometer used were an Agilent 1260 Infinity LC system (Agilent Technologies, Inc.) and an AB SCIEX TripleQuad 4500 system (AB SCIEX Corporation), respectively. The column used was a Scherzo SS-C18 (100 mm x 2 mm, 3 μm) (Intact Corporation), and the oven temperature was 40°C. Five microliters of an appropriately diluted sample was injected, and the flow rate was 0.5 mL / min. The eluents were a 0.1% aqueous formic acid solution (eluent A) and a 50 mM ammonium acetate / methanol solution (eluent B). After equilibration at a ratio of eluent A:eluent B of 95:5, the sample was applied, and ureido nitrogen was eluted using a linear gradient such that after 5 minutes the ratio became eluent A:eluent B of 80:20.

[0060] (8) Standards Used and Quantification The amount of ureido nitrogen in exudate was analyzed by LC-MS. Allantoic acid (Toronto Research Chemicals Inc.) was used as the standard. The standard was analyzed by LC-MS, and a calibration curve was created in the range of 10-1000 ppb. Allantoic acid was identified for each sample based on the retention time, exact mass, and MS / MS spectrum agreement with each reagent. Furthermore, allantoic acid in the sample was quantified from the calibration curve, and the total amount per plant was calculated.

[0061] (9) Results The measurement results of nodule activity are shown in Figure 2. The graphs in each figure show the mean ± standard deviation. A comparison of test plots 2 and 3 showed that adding components (B), (C), and (D) to ascorbic acid resulted in a greater improvement in nodule activity than when ascorbic acid was dissolved in tap water and sprayed alone.

[0062] Example 3: Effect on soybean (green soybean) yield in the field (1) Cultivation conditions: Cultivation was carried out in a field in Ibaraki Prefecture, and the soybean (green soybean) cultivar "Yuagari Musume" (Kaneko Seed Co., Ltd.) was used. 12 days before sowing, 2.0 kg of AG Max (AiAgri Co., Ltd., N:P:K = 10:10:10), 5.2 kg of Yorin (17.5%), and 1.5 kg of potassium chloride (60%) were applied to the soil as basal fertilizer to achieve an N:P:K = 8:10:5 (kg / 10a). The planting density was 0.15 m between plants and 0.70 m between rows. Two seeds were sown per location, and the plants were thinned to one plant 15 days after sowing.

[0063] (2) Foliar spray treatment and harvesting Foliar spray was carried out 58 days after sowing, at a growth stage corresponding to the pod-setting stage. Foliar spray treatment was carried out in a plot of the field, with each plot having an area of ​​1.68 square meters, measuring 1.20 m on the short side and 1.40 m on the long side. Three plots were tested per test plot. Each plot had 16 plants. Foliar spray was carried out only once, and three types of test plots were examined. Test plots 1 to 3 evaluated are as follows. The spray solution for test plot 3 was prepared according to the method described in Example 1. <Spray solution> - Zone 1: Control (no application) - Zone 2: Foliar spray of 1,500 ppm by mass of ascorbic acid (dissolved in well water) - Zone 3: Foliar spray of 1,500 ppm by mass of ascorbic acid + 1 ppm by mass of BHT + 50 ppm by mass of glycerin + 350 ppm by mass of sorbitan monolaurate + 300 ppm by mass of silica (dissolved in well water)

[0064] The spray solution was prepared and sprayed using a battery sprayer (ADB150Li, Maruyama Seisakusho Co., Ltd.) at a rate of 100 L / 10 a, at 168 mL per plot, covering the entire soybean plant. Harvesting was carried out 76 days after sowing. Ten plants were harvested from each plot. After harvest, all pods were collected from each individual plant, and the fresh pod weight was measured as yield data.

[0065] (3) Results The results of measuring the fresh pod weight per plant are shown in Figure 3. The graph in the figure shows the average value ± standard deviation. When ascorbic acid was dissolved in well water and sprayed on the leaves, no yield-increasing effect was confirmed. However, by adding components (B), (C), and (D) to ascorbic acid, a yield-increasing effect of approximately 2.3% was observed compared to the control, demonstrating that a yield-increasing effect can also be obtained when well water is used.

[0066] Example 4 Effect on seed yield in Lotus japonicus (1) Soil preparation Medium-term fertilizer release soil (Takii hydrous cell soil, medium-term fertilizer release type, Takii Seed Co., Ltd.) and vermiculite granules (Akagi Engei Co., Ltd.) were mixed in a volume ratio of 1:1, and the soil was filled into a polypot (diameter 6 cm, height 5.5 cm).

[0067] (2) Germination treatment and sowing Lotus japonicus seeds were Miyakojima MG-20 strain. Approximately 5 mL of seeds were placed in a 50 mL centrifuge tube (Iwaki Corporation), to which 2 mL of concentrated sulfuric acid (Fujifilm Wako Pure Chemical Industries, Ltd.) was added, and the mixture was left to stand for 10 minutes. After rinsing with tap water five times, the mixture was submerged for 2 hours. After adding 100 mL of tap water per pot, the seeds were sown at a depth of approximately 1 cm from the soil surface in a tray filled with medium-release soil to a height of 3 cm, with the seeds not overlapping. One week after sowing, 1 cm deep planting holes were made in the soil of the polypot, and the germinated seedlings on the tray were replanted. The number of replicates for each test plot was 8 (n = 8).

[0068] (3) Inoculation of Rhizobium A solid medium was prepared by adding 1.5% agar (Fujifilm Wako Pure Chemical Industries, Ltd.) to yeast-mannitol (YM) medium (0.5 g K2HPO4, 0.2 g MgSO4 7H2O, 0.1 g NaCl, 0.4 g Yeast Extract, 10 g Mannitol, and 1 L of distilled water (pH 6.8)). Mesorhizobium loti MAFF303099 (ML GUS) strain was grown on the solid medium. A loopful of the grown rhizobium was inoculated into 50 mL of YM liquid medium in a 500 mL Sakaguchi flask and cultured at 30 °C for approximately 36 hours with shaking. A rhizobial culture solution was prepared with a bacterial cell turbidity OD600 value of about 0.3. After sowing, 1 mL of the rhizobial culture solution was dropwise inoculated onto the seeds using a micropipette.

[0069] (4) Cultivation conditions: Cultivation was performed indoors under the conditions of 16 hours light / 8 hours dark, 25°C, LED light source (Ogetsu Corporation; model number: VGL-1200W), and light intensity of 400 to 440 μmol / m ２ Watering was performed by adding tap water to the tray placed under the pot after the water had run out, enough to cover the bottom 5 cm of the pot. Starting three weeks after sowing, a 1,000-fold dilution of High Grade Flowering Promoter (Hyponex Japan Co., Ltd.) was added to the water once a week.

[0070] (5) Foliar spray treatment and harvest The following three types of spray solutions were prepared, and 6 weeks after sowing, 6.7 mL per plant was sprayed using a spray bottle. The spray solution for test group 3 was prepared according to the method described in Example 1. <Spray solutions> Test group 1: Control (no application) Test group 2: 1,500 ppm by mass of ascorbic acid, foliar spray (dissolved in tap water) Test group 3: 1,500 ppm by mass of ascorbic acid + 1 ppm by mass of BHT + 50 ppm by mass of glycerin + 350 ppm by mass of sorbitan monolaurate + 300 ppm by mass of silica, foliar spray (dissolved in tap water)

[0071] Mature pods were harvested sequentially from about 8 weeks after sowing, and cultivation was terminated 12 weeks after sowing. The pods were air-dried, and the seeds were collected and the weight of the seeds obtained from each plant was measured.

[0072] (6) Results The results of measuring seed weight are shown in Figure 4. The graph in the figure shows the average value ± standard deviation. Compared to the control group, no yield-increasing effect was confirmed when ascorbic acid was dissolved in well water and sprayed on the leaves. However, adding component (B), component (C), and component (D) to ascorbic acid showed a yield-increasing effect of 16.7% compared to the control, demonstrating that a yield-increasing effect can also be obtained when tap water is sprayed.

[0073] Example 5 Effect on soybean growth (1) Soil preparation and sowing Medium-term fertilizer release soil (Takii Hydrated Cell Soil Medium-term Fertilizer Release, Takii Seed Co., Ltd.) and vermiculite granules (Akagi Engei Co., Ltd.) were mixed at a volume ratio of 1:1, and the soil was filled into polypots (diameter 10.5 cm, height 9 cm). Soybean seeds used were "Enrei" (purchased from Nikko Seed Co., Ltd.). After adding 250 mL of tap water per pot, two seeds were sown in each pot, one seed at a depth of approximately 1-2 cm from the soil surface. Each test plot was repeated 6 times (n = 6).

[0074] (2) Inoculation of Rhizobium A solid medium was prepared by adding 1.5% agar (Fujifilm Wako Pure Chemical Industries, Ltd.) to yeast-mannitol (YM) medium (0.5 g KHPO, 0.2 g MgSO.7H2O, 0.1 g NaCl, 0.4 g yeast extract, 10 g mannitol, and 1 L distilled water (pH 6.8)). Bradyrhizobium japonicum NBRC14783T strain was grown on the solid medium. A loopful of the grown rhizobium was inoculated into 50 mL of YM liquid medium in a 500 mL Sakaguchi flask and cultured with shaking at 30°C for approximately 36 hours. A rhizobial culture solution was prepared with a bacterial cell turbidity OD600 value of about 0.3. After sowing, 1 mL of the rhizobial culture solution was dropwise inoculated onto the seeds using a micropipette.

[0075] (3) Cultivation conditions Cultivation was carried out in an artificial climate chamber (LPH-411SP, Nippon Medical and Chemical Instruments Manufacturing Co., Ltd.), and light conditions were light (light intensity 440 to 480 μmol / m ２The temperature was 30°C light / 25°C dark, and the humidity was 50%. After germination, the plants were thinned out to one per pot. After the water in the tray placed under the pot had run out, fresh tap water was added to the tray, enough to cover the bottom 5 cm of the pot.

[0076] (4) Foliar spray treatment A spray solution was prepared, and on the seventh day after sowing, 6.7 mL was sprayed per plant using a spray bottle. Foliar spray was performed only once, and five types of test plots were examined. The test plots 1 to 5 evaluated are as follows. <Spray solution> - Plot 1: Control (no application) - Plot 2: Foliar spray of 1,500 ppm by mass of ascorbic acid (dissolved in tap water) - Plot 3: 1,500 ppm by mass of ascorbic acid + 1 ppm by mass of BHT + 50 ppm by mass of glycerin (dissolved in tap water) - Plot 4: 1,500 ppm by mass of ascorbic acid + 1 ppm by mass of BHT + 50 ppm by mass of propylene glycol (dissolved in tap water) - Plot 5: 1,500 ppm by mass of sodium ascorbate + 10 ppm by mass of tocopherol + 50 ppm by mass of glycerin (dissolved in tap water)

[0077] (5) Results On the 9th day after sowing, the plants were dried at 100°C for 7 hours and then the dry weight of the aboveground parts of the plants was measured. The results of measuring the dry weight of the aboveground parts are shown in Figure 5. The graphs in each figure represent the mean ± standard deviation. A comparison of test plots 1 to 5 showed that when ascorbic acid was dissolved in tap water and sprayed on the leaves, no growth-promoting effect was observed. However, adding component (B) and component (C) to ascorbic acid or sodium ascorbate resulted in a growth-promoting effect even when tap water was used as the dissolving water. Compared to the control, the dry weight of the aboveground parts increased by an average of approximately 3.7% in test plot 3, approximately 9.1% in test plot 4, and approximately 2.1% in test plot 5.

Claims

1. A growth promoter for leguminous plants included in beans, comprising a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol, wherein the antioxidant (B) is at least one selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the component (B) is taken as 1.

2. A root nodule activity promoter comprising a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol, wherein the antioxidant (B) is at least one selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the component (B) is taken as 1.

3. A yield-enhancing agent for leguminous plants, including beans, comprising a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol, wherein the antioxidant (B) is at least one selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the component (B) is taken as 1.

4. The agent according to any one of claims 1 to 3, wherein the polyol (C) is one or more selected from diethylene glycol, glycerin and propylene glycol.

5. The agent according to any one of claims 1 to 3, wherein the ratio (mass ratio) of the polyol (C) is 4 to 100,000 when the ratio of the component (B) is 1.

6. The agent according to any one of claims 1 to 3, further comprising (D) a surfactant in combination.

7. The agent according to claim 6, wherein the ratio (mass ratio) of the surfactant (D) is 10 to 300,000 when the ratio of the component (B) is 1.

8. The agent according to claim 6 or 7, wherein the surfactant (D) comprises one or more selected from nonionic surfactants, anionic surfactants and amphoteric surfactants.

9. The agent according to claim 8, wherein (D) the nonionic surfactant is one or more selected from sorbitan fatty acid esters, glycerin fatty acid esters, polyalkylene glycol fatty acid esters, polyoxyethylene resin acid esters, and polyoxyethylene alkyl ethers, the anionic surfactant is one or more selected from fatty acid salts, aromatic sulfonate-formaldehyde condensates, and alkyl sulfates, and the amphoteric surfactant is 2-alkyl-N-carboxymethyl-N-hydroxyethylimidazolinium betaine.

10. The agent according to any one of claims 1 to 3 and 6, further comprising (F) a carrier in combination.

11. The agent according to claim 10, wherein the ratio (mass ratio) of the carrier (F) is 50 to 1,000 when the ratio of the component (B) is 1.

12. The agent according to claim 10 or 11, wherein the carrier (F) is one or more selected from zeolite, silica, bentonite and mirabilite.

13. A method for promoting the growth of legumes, which comprises the step of combining (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol with water and applying the resulting mixture to soil or a plant, wherein the (B) antioxidant is at least one selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the ratio of component (B) is 1.

14. A method for promoting nodule activity, comprising the step of combining (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol with water and applying the resulting mixture to soil or a plant, wherein the (B) antioxidant is at least one selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the component (B) is taken as 1.

15. A method for increasing yield of legumes, including beans, which comprises a step of combining (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol with water and applying the resulting mixture to soil or a plant, wherein the (B) antioxidant is at least one selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the ratio of component (B) is 1.

16. Use of a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol for producing a growth promoter for legumes, wherein the antioxidant (B) is at least one selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the component (B) is taken as 1.

17. Use of a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol for producing a root nodule activity promoter, wherein the antioxidant (B) is at least one selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof is 10 to 3,000,000 when the ratio of component (B) is 1.

18. Use of a combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol for producing a yield-enhancing agent for legumes, the use comprising the combination of (A) ascorbic acid or a salt thereof, (B) an antioxidant, and (C) a polyol, the antioxidant (B) being at least one selected from tocopherol, dibutylhydroxytoluene, and butylhydroxyanisole, and the ratio (mass ratio) of (A) ascorbic acid or a salt thereof being 10 to 3,000,000 when the component (B) is taken as 1.

19. The use according to any one of claims 16 to 18, wherein the polyol (C) is one or more selected from diethylene glycol, glycerin and propylene glycol.

20. The use according to any one of claims 16 to 18, wherein the proportion (mass ratio) of the polyol (C) is 4 to 100,000 when the proportion of the component (B) is 1.

21. The use according to any one of claims 16 to 18, further comprising in combination (D) a surfactant.

22. The use according to claim 21, wherein the ratio (mass ratio) of the surfactant (D) is 10 to 300,000 relative to the ratio of component (B) being 1.

23. The use according to claim 21 or 22, wherein the surfactant (D) comprises one or more selected from nonionic surfactants, anionic surfactants and amphoteric surfactants.

24. (D) The use according to claim 23, wherein the nonionic surfactant is one or more selected from sorbitan fatty acid esters, glycerin fatty acid esters, polyalkylene glycol fatty acid esters, polyoxyethylene resin acid esters, and polyoxyethylene alkyl ethers, the anionic surfactant is one or more selected from fatty acid salts, aromatic sulfonate-formaldehyde condensates, and alkyl sulfates, and the amphoteric surfactant is 2-alkyl-N-carboxymethyl-N-hydroxyethylimidazolinium betaine.

25. The use according to any one of claims 16 to 18 and 21, further comprising (F) a carrier in combination.

26. The use according to claim 25, wherein the ratio (mass ratio) of the carrier (F) is 50 to 1,000 relative to the ratio of the component (B) being 1.

27. The use according to claim 25 or 26, wherein the carrier (F) is one or more selected from zeolite, silica, bentonite and mirabilite.

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