Plant environmental stress tolerance enhancer

Abstract

The present invention addresses the problem of providing a new means for imparting tolerance to environmental stress such as salt stress, particularly providing a means for imparting tolerance to environmental stress to plants by using a cyclic cyclodipeptide that is a 2,5-diketopiperazine derivative.　The present invention provides: a plant environmental stress tolerance enhancer composed of a cyclodipeptide that contains phenylalanine and / or leucine, and / or a salt of said cyclodipeptide; a composition for imparting tolerance to environmental stress to plants, the composition containing the aforementioned enhancer; and a seed containing the aforementioned composition on the surface and / or in the interior thereof.

Description

Plant environmental stress tolerance enhancer

[0001] The present invention relates to an agent for imparting plant environmental stress tolerance, a composition containing the same, and seeds containing the composition.

[0002] Environmental changes resulting from recent human activities and climate change have had a significant impact on agricultural production on a global scale. Therefore, there is an urgent need to develop technologies that will enable the harvesting of crops even in lands that have previously been considered unsuitable for agriculture, such as lands with high salinity, low or high temperatures, and dry lands. To address this issue, it is important to develop methods to confer stress tolerance to plants.

[0003] Plant stress can be broadly divided into biological stress such as plant disease and environmental stress such as salt concentration and temperature.

[0004] There have been several reports on the effectiveness of 2,5-diketopiperazine derivatives in imparting stress tolerance to plants.

[0005] Non-Patent Document 1 reports that a cyclic dipeptide composed of leucine and phenylalanine, a 2,5-diketopiperazine derivative derived from Bacillus subtilis KS1 strain, induces resistance to plant diseases caused by infection with oomycetes and confers resistance to downy mildew fungi. However, Non-Patent Document 1 does not disclose anything about imparting resistance to environmental stress.

[0006] Patent Document 1 reports that cyclic cyclodipeptides, cyclo(leucine-phenylalanine) and cyclo(proline-valine), can be used as agents for controlling plant diseases caused by oomycete infection and downy mildew. However, Patent Document 1 also reports on biological plant diseases and does not disclose any information on imparting resistance to environmental stress.

[0007] Patent Document 2 discloses that 2,5-diketopiperazine derivatives in which two amino acid molecules are cyclically bonded, and those having a specific structure, provide induced resistance to plant diseases and promote plant growth. Patent Document 2 discloses the application of 2,5-diketopiperazine derivatives to impart disease resistance to plants. However, Patent Document 2 does not disclose anything about imparting resistance to environmental stress.

[0008] Therefore, there have been no reports to date examining whether cyclic cyclodipeptides, which are 2,5-diketopiperazine derivatives, can confer environmental tolerance to plants.

[0009] Regarding environmental stress, Patent Document 3 discloses that branched amino acids such as valine, leucine, and α-ketoisovaleric acid confer high temperature stress tolerance to plants by altering the amino acid metabolism of the plant. However, Patent Document 3 only discloses the conferring of high temperature stress tolerance, and does not disclose the conferring of other stress tolerances such as salt stress tolerance.

[0010] International Publication WO2021 / 125272 Patent Publication No. 2013-531688 Patent Publication No. 2012-197249

[0011] Suzuki et al., Letters to Applied Microbiology, 2022. 0: 1-12.

[0012] An object of the present invention is to provide a new means for imparting tolerance to environmental stresses such as salt stress, and in particular to provide a new means for imparting tolerance to environmental stresses to plants using a cyclic cyclodipeptide that is a 2,5-diketopiperazine derivative.

[0013] In order to solve the above problems, the present inventors have conducted extensive research and have found that cyclodipeptides containing phenylalanine and / or leucine, and / or salts thereof, can confer tolerance to plants to various environmental stresses, including salt stress.

[0014] The present invention is based on the above-mentioned new findings and is as follows: (1) An agent for imparting environmental stress tolerance to plants, comprising a cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof. (2) The agent according to (1), wherein the cyclodipeptide containing phenylalanine and / or leucine is cyclo(leucine-phenylalanine). (3) The agent according to (1) or (2), wherein the environmental stress is one or more selected from the group consisting of salt stress, low temperature stress, and drought stress. (4) A composition for imparting environmental stress tolerance to a plant, comprising the agent according to any one of (1) to (3). (5) The composition according to (4), which imparts environmental stress tolerance to the plant in the early growth stage. (6) The composition according to (4) or (5), which promotes the growth of the plant under environmental stress conditions compared to when the cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof is not applied. (7) A seed, the surface and / or interior of which contains the composition according to (4). (8) The seed according to (7), wherein at least a portion of the seed surface is coated with the composition according to (4). (9) The seed according to (7) or (8), wherein the growth of a plant derived from the seed under environmental stress conditions is promoted compared to the absence of the cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof. (10) The seed according to any one of (7) to (9), wherein the cyclodipeptide containing phenylalanine and / or leucine is cyclo(leucine-phenylalanine). (11) The seed according to any one of (7) to (10), wherein the environmental stress is one or more selected from the group consisting of salt stress, low temperature stress, and drought stress. (12) The seed according to any one of (7) to (10), wherein the amount of the cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof is 0.05 x 10 －７ ~50 x 10 －４parts by mass of a cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof. (13) A method for cultivating a plant under environmental stress conditions, comprising a step of applying the composition according to (4) to the plant. (14) The method according to (13), wherein the cyclodipeptide containing phenylalanine and / or leucine is cyclo(leucine-phenylalanine). (15) The method according to (13) or (14), wherein the environmental stress is one or more selected from the group consisting of salt stress, low temperature stress, and drought stress. (16) In the step of applying, the composition according to (4) is applied as a seed composition comprising seeds and the composition, and the seed composition is in an amount of 0.05 x 10 per 100 parts by mass of the seeds. －７ ~50 x 10 －４ parts by mass of a cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof. (17) A method for reducing environmental stress in a plant, comprising a step of applying the composition according to (4) to a plant. (18) The method according to (17), wherein the cyclodipeptide containing phenylalanine and / or leucine is cyclo(leucine-phenylalanine). (19) The method according to (17) or (18), wherein the environmental stress is one or more selected from the group consisting of salt stress, low temperature stress, and drought stress. (20) In the step of applying, the composition according to (4) is applied as a seed composition comprising seeds and the composition, and the seed composition is in an amount of 0.05 x 10 per 100 parts by mass of the seeds. －７ ~50 x 10 －４ The method according to any one of (17) to (19), comprising: parts by mass of a cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof. This specification incorporates the disclosure of Japanese Patent Application No. 2024-041512, from which the present application claims priority.

[0015] A cyclodipeptide containing phenylalanine and / or leucine, and / or a salt thereof, is useful as an agent for imparting tolerance to environmental stresses such as salt stress to plants. Furthermore, a composition containing the agent as an active ingredient is useful as a composition for imparting tolerance to environmental stresses such as salt stress to plants. Furthermore, plants derived from seeds containing the composition exhibit suppressed growth inhibition caused by environmental stresses such as salt stress, even under environmental stress conditions.

[0016] The present invention will be described in further detail below. 1. Agent for Imparting Environmental Stress Tolerance to Plants of the Present Invention 1-1 Overview A first aspect of the present invention is an agent for imparting environmental stress tolerance to plants, comprising a cyclodipeptide containing phenylalanine and / or leucine, and / or a salt thereof.

[0017] 1-2 Cyclodipeptide of the Present Invention Containing Phenylalanine and / or Leucine As used herein, the term "cyclodipeptide containing phenylalanine and / or leucine" refers to a dipeptide containing phenylalanine and / or leucine, in which the amino group and the carboxyl group of two peptides forming the dipeptide are bonded to each other to form a cyclic structure.

[0018] Herein, phenylalanine refers to an amino acid having the chemical structure commonly known in the art as "phenylalanine," and leucine refers to an amino acid having the chemical structure commonly known in the art as "leucine."

[0019] More specifically, the cyclodipeptides of the present invention are as follows: 1) A cyclodipeptide containing phenylalanine and leucine, in which the carboxyl group contained in the phenylalanine and the amino group contained in the leucine are bonded to each other to form a ring, 2) A cyclodipeptide containing phenylalanine and an arbitrary amino acid, in which the carboxyl group contained in the phenylalanine and the amino group contained in the arbitrary amino acid are bonded to each other to form a ring, and 3) A cyclodipeptide containing leucine and an arbitrary amino acid, in which the carboxyl group contained in the leucine and the amino group contained in the arbitrary amino acid are bonded to each other to form a ring.

[0020] The optical isomerism of the amino acids constituting the plant environmental stress tolerance imparting agent of the present invention is not particularly limited, and they may be either L- or D-isomers.

[0021] In the present invention, it is a preferred embodiment that the cyclodipeptide is a cyclodipeptide consisting of phenylalanine and leucine.

[0022] Furthermore, in the present invention, it is a particularly preferred embodiment that the cyclodipeptide is cyclo(leucine-phenylalanine) represented by the following formula:

[0023]

[0024] In this specification, the above cyclo(leucine-phenylalanine) may also be represented as cyclo(Leu-Phe) or cLF.

[0025] 1-3 Salts of the Cyclodipeptide of the Present Invention

[0033] In the present invention, the term "salt of a cyclodipeptide containing phenylalanine and / or leucine" is not particularly limited as long as it is a salt of the above-described cyclodipeptide that is acceptable for agricultural use. Examples of such salts include, but are not limited to, alkali metal salts such as sodium salt and potassium salt; alkaline earth metal salts such as calcium salt, magnesium salt, and barium salt; aluminum salt; salts with organic bases such as trimethylamine, triethylamine, pyridine, picoline, 2,6-lutidine, ethanolamine, diethanolamine, triethanolamine, cyclohexylamine, dicyclohexylamine, and N,N-dibenzylethylenediamine; salts with inorganic acids such as hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, and phosphoric acid; and organic salts with acids such as formic acid, acetic acid, trifluoroacetic acid, phthalic acid, fumaric acid, oxalic acid, tartaric acid, maleic acid, citric acid, succinic acid, methanesulfonic acid, benzenesulfonic acid, and p-toluenesulfonic acid.

[0026] The "cyclodipeptide containing phenylalanine and / or leucine, and / or a salt thereof" constituting the agent for imparting plant environmental stress tolerance of the present invention may be chemically synthesized or may be derived from natural products such as microorganisms and plants.

[0027] In this specification, the "cyclodipeptide containing phenylalanine and / or leucine, and / or a salt thereof" may be simply referred to as the active ingredient.

[0028] 1-4 Environmental Stress In this specification, "environmental stress" refers to an environmental factor that imposes stress on a plant and thereby inhibits the growth of the plant. Note that "environmental stress" in this specification refers to abiotic stress related to environmental or physical factors, and does not include biological stress such as infection by pathogenic bacteria.

[0029] Specific examples of "environmental stress" include, but are not limited to, salt stress, low temperature stress, high temperature stress, drought stress, excessive humidity stress, and light stress.

[0030] As used herein, the term "salt stress" refers to, but is not limited to, stress that occurs under environmental conditions in which the salt concentration in the soil in which a plant is growing deviates from the optimum concentration for growth and shifts to a higher concentration.

[0031] As used herein, the term "low temperature stress" refers to, but is not limited to, stress that occurs under environmental conditions in which the temperature of the environment in which a plant is growing is lower than the optimum growth temperature or is below the low temperature limit.

[0032] As used herein, the term "high temperature stress" refers to, but is not limited to, stress that occurs under environmental conditions in which the temperature of the environment in which a plant is growing exceeds the upper limit of the optimum growth temperature.

[0033] As used herein, the term "drought stress" refers to, but is not limited to, stress that occurs when a plant is put into a water-deficient state due to drought.

[0034] As used herein, the term "excessive moisture stress" refers to stress that occurs when the moisture content in the soil in which a plant is growing exceeds the optimum amount for plant growth, but is not limited to this.

[0035] As used herein, the term "light stress" refers to, but is not limited to, stress that occurs when the light intensity in the environment in which a plant is growing exceeds the upper or lower limit of the light intensity optimal for growth.

[0036] 1-5 Plant Environmental Stress Inducing Agent As used herein, the term "plant environmental stress inducing agent" refers to an agent that can impart tolerance to the above-mentioned environmental stress to a plant. In other words, the term refers to an agent that can reduce the effects of the above-mentioned environmental stress on a plant (e.g., suppression of plant growth) even in the presence of the above-mentioned environmental stress.

[0037] In the examples shown below, it has been demonstrated that application of cyclo(leucine-phenylalanine) to a plant (corn) confers tolerance to salt stress, low temperature stress, and drought stress.

[0038] 1-6 Plants Targeted by the Agent for Imparting Plant Environmental Stress Tolerance of the Present Invention The term "plant" as used herein is not particularly limited, but is preferably a crop plant, and more preferably an edible plant. Examples of crop plants include corn (maize), wheat, barley, rye, oat, rice, soybean, canola (rapeseed), cotton, sunflower, sugar beet, potato, tobacco, broccoli, lettuce, cabbage, spinach, komatsuna (Japanese mustard spinach), cauliflower, coconut, tomato, cucumber, eggplant, melon, pumpkin, okra, bell pepper, watermelon, carrot, radish, onion, leek, fruit trees, ornamental plants, turf, and pasture grass, but are not limited thereto. In terms of increasing food production, it is preferable that the plant to which environmental stress tolerance is imparted by the agent of the present invention is a monocotyledonous cereal plant.

[0039] 2. Composition of the Present Invention for Imparting Environmental Stress Tolerance to Plants 2-1 Overview A second aspect of the present invention is a composition for imparting environmental stress tolerance to plants, which contains the plant environmental stress tolerance imparting agent described above in 1. By applying the composition of the present invention to a plant, it is possible to impart environmental stress tolerance to the plant.

[0040] 2-2 Constitution of the composition of the present invention As described above, the composition of the present invention is a composition containing "a cyclodipeptide containing phenylalanine and / or leucine, and / or a salt thereof" as an active ingredient. The "cyclodipeptide containing phenylalanine and / or leucine, and / or a salt thereof" of the present invention is as described in 1-3 above.

[0041] The composition of the present invention may contain other ingredients such as excipients in addition to the active ingredient "cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof." The composition of the present invention may also contain one or more of the above active ingredients.

[0042] In a preferred embodiment of the present invention, the cyclodipeptide containing phenylalanine and / or leucine is cyclo(leucine-phenylalanine) (cLF).

[0043] The composition of the present invention may be in the form of a liquid composition or a solid composition, and may be a composition suitable for application to the seeds, or above-ground parts such as leaves, stems, fruits, flowers, and roots, or the rhizosphere of a target plant.

[0044] That is, the composition of the present invention may contain, in addition to the active ingredient, any other ingredients necessary for formulation, so long as the ingredients do not impair the effect of imparting environmental stress tolerance to plants. Such ingredients include solvents, carriers, pH adjusters, thickeners, spreaders, surfactants, antioxidants, extenders, dispersants, humectants, colorants, antifoaming agents, UV protectants, antifreeze agents, preservatives, biological control agents or biocides, emulsifiers, sequestrants, plasticizers, phospholipids, flow agents, coalescing agents, waxes, preservatives, and / or elements necessary for plant growth (e.g., one or more selected from the group consisting of Mo, Co, B, Fe, Cu, Zn, Mn, S, Mg, Ca, N, P, and K). However, the ingredients are not limited to those commonly used in agricultural compositions.

[0045] The form of the composition containing the active ingredient may be a liquid composition or a solid composition, but from the viewpoint of ease of measurement and handling, and ease of mixing with other ingredients, a liquid composition is preferable. The dosage form may be any of liquid, granule, emulsion, wettable powder, oil, aerosol, etc., and is not particularly limited.

[0046] 2-3 Effects of the Composition of the Present Invention By applying the composition of the present invention to plants, plant growth can be promoted under environmental stress conditions compared to when the phenylalanine-containing cyclodipeptide and / or a salt thereof is not applied. In other words, environmental stress tolerance can be imparted to plants. Here, "imparting environmental stress tolerance to plants" means that the effects of environmental stress on plants (e.g., plant growth inhibition) are reduced even in the presence of environmental stress.

[0047] The "environmental stress" of the present invention is as described in 1-4 above, and is preferably salt stress, low temperature stress, or drought stress.

[0048] The plants to which the composition of the present invention is applied are those for which it is desired to reduce the above-mentioned environmental stress conditions, and preferably the plants described in 1-6 above.

[0049] The composition of the present invention confers environmental stress tolerance to plants, preferably to plants at the early growth stage, but may also confers environmental stress tolerance not only to the early growth stage but throughout the entire growth period of the plant.

[0050] Here, "a plant in an early growth stage" means a plant at a stage from sowing seeds of a plant to which stress tolerance is intended to be imparted until germination, and a plant at a stage where the seeds have germinated and several leaves, for example, about 3 to 7 leaves, are produced, but is not limited to this range of stages.

[0051] For example, in the following examples, the effect of the composition of the present invention on imparting environmental stress tolerance was investigated using corn. That is, seeds were coated with the composition of the present invention, sown in soil, and cultivated under various environmental stresses including salt stress. 21 days after sowing, the growth status of corn seedlings derived from the seeds was evaluated to determine whether the composition of the present invention had imparted environmental stress tolerance.

[0052] In addition, it is a preferred embodiment to apply the composition of the present invention to the seeds of plants to which environmental stress resistance is to be imparted.

[0053] 2-4 Method for Producing the Composition of the Present Invention The method for producing the composition of the present invention is not particularly limited, and the composition can be produced by mixing the components by an appropriate means. A liquid composition can be produced by stirring, dispersing, or the like in a liquid medium such as water or alcohol, as needed. A solid composition can be produced by pulverizing, granulating, drying, or the like, as needed.

[0054] 3. Seeds containing on their surface and / or inside a composition for imparting environmental stress tolerance to plants 3-1 Overview A third aspect of the present invention is a seed containing on its surface and / or inside the composition described in 2 above.

[0055] 3-2 Configuration of the Seed of the Present Invention The seed of the present invention contains on its surface and / or inside a composition containing "a cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof," thereby enabling a plant derived from or developed from the seed of the present invention to have environmental stress resistance. The "cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof" of the present invention is as described in 1-3 above.

[0056] In a preferred embodiment of the present invention, the cyclodipeptide containing phenylalanine and / or leucine is cyclo(leucine-phenylalanine) (cLF).

[0057] The "environmental stress" of the present invention is as described in 1-4 above, and is preferably salt stress, low temperature stress, or drought stress.

[0058] The seeds of the present invention are seeds of plants for which reduction of the above-mentioned environmental stress is desired, and are preferably seeds of the plants described in 1-6 above.

[0059] The growth of plants derived from the seeds of the present invention is promoted under environmental stress conditions, as compared with plants derived from seeds that do not contain the active ingredient phenylalanine-containing cyclodipeptide and / or its salt, and this effect is demonstrated by the experimental data in the Examples described below.

[0060] 3-3 Method for Producing Seeds of the Present Invention The surface of a plant seed to which tolerance to environmental stress is to be imparted can be coated with the composition described in 2 above. For example, the surface of the seed can be coated by subjecting the seed to a coating treatment. While it is sufficient that at least a portion of the surface of the seed is coated, preferably the entire surface of the seed is coated.

[0061] The composition containing the active ingredient for coating plant seeds with the composition of the present invention is not particularly limited. However, in order to ensure reliable coating, it is preferable that the composition for coating seeds has a certain level of viscosity or higher, and it is a preferred embodiment of the present invention that the composition contains a thickener.

[0062] The composition containing the active ingredient can be used as a coating solution to coat the seeds of plants intended to confer resistance to environmental stress with the composition using a coating device. To coat the seeds with the composition containing the active ingredient, the seeds and the composition can be mechanically blended to ensure uniform coating. An example of a coating device that can be used for this purpose is the CONCEPT ML2000 manufactured by SATEC, which was used in the following examples. Alternatively, the seeds can be coated with the composition containing the active ingredient by spraying it on the seeds after sowing.

[0063] Alternatively, the seeds of a plant to which tolerance to environmental stress is to be imparted may be soaked in the composition in solution to incorporate the composition into the seeds, which may then be washed to remove excess active ingredient.

[0064] When the seeds of the present invention are not sown in soil immediately after coating treatment, the coated seeds may be dried once, but drying the seeds is not necessarily required.

[0065] 3-4 Amount of active ingredient to be coated on the seeds of the present invention The cyclodipeptide containing phenylalanine and / or leucine and / or salts thereof to be coated on the seeds of the present invention is not particularly limited as long as it can impart environmental stress resistance to plants. For example, 0.05 × 10 －9 ~50 x 10 －３ Part by mass, 0.1×10 －9 ~100 x 10 －３ Part by mass, 0.05×10 －７ ~50 x 10 －４ Part by mass, 0.1×10 －７ ~100 x 10 －４ Part by mass, 0.05×10 －6 ~50 x 10 －5 Parts by mass, or 0.1 x 10 －6 ~100 x 10 －5It can be contained in the range of 0.05 × 10 parts by mass for 100 parts by mass of seeds. －７ ~50 x 10 －４ In a preferred embodiment of the present invention, the active ingredient is a cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof, and the amount of the active ingredient to coat the seeds can be adjusted appropriately depending on the type of plant and the amount of the solution normally applied for pest control.

[0066] In the examples below, when a cyclodipeptide containing phenylalanine and / or leucine was used as cLF, significant promotion of plant growth was observed when 0.1 mg to 0.2 g of cLF was applied to 1 ton of corn seeds in the presence of salt stress of 6 g per 1 L of soil, compared to a control where no cLF was applied.

[0067] 4. Method for cultivating plants under environmental stress conditions 4-1 Overview A fourth aspect of the present invention is a method for cultivating plants under environmental stress conditions, which comprises the step of applying the composition described in 2 above to the plants.

[0068] 4-2 Configuration of the plant cultivation method of the present invention The method of cultivating plants under environmental stress conditions of the present invention enables plants to be cultivated even under environmental stress conditions by applying to the plants a composition containing "a cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof."

[0069] The "cyclodipeptide containing phenylalanine and / or leucine, and / or a salt thereof" of the present invention is as described above in 1-3. In a preferred embodiment of the present invention, the cyclodipeptide containing phenylalanine and / or leucine as an active ingredient is cyclo(leucine-phenylalanine) (cLF).

[0070] The cultivation method of the present invention only needs to include the step of applying the composition described in 2 above, and other steps in plant cultivation may be conventional, such as sowing seeds in soil and growing the plants in the soil, etc. The cultivation method of the present invention only needs to include the step of applying the composition described in 2 above, and other steps are optional.

[0071] The "environmental stress" of the present invention is as described in 1-4 above, and is preferably salt stress, low temperature stress, or drought stress.

[0072] In the cultivation method of the present invention, the plants to which the composition described in 2 above is applied are plants for which reduction of the above environmental stress is desired, and preferably the plants described in 1-6 above.

[0073] Examples of application of the composition described in 2 above to target plants include, but are not limited to, application to the seeds, or above-ground parts such as leaves, stems, fruits, flowers, and roots, or the rhizosphere of the target plants. A preferred embodiment of the method of the present invention is application to seeds. Furthermore, application to seeds may be combined with application to the rhizosphere or above-ground parts such as leaves and stems.

[0074] In the application step, the seeds may be coated with the composition described in 2 above, and then the seeds can be sown in soil. The method for coating seeds with the composition described in 2 above is as described in 3-3 above.

[0075] In the application step, the seeds may be immersed in the solution of the composition described in 2 above, and then the seeds can be sown in soil.

[0076] In the application step of the present invention, the cyclodipeptide containing phenylalanine and / or leucine and / or salts thereof when the composition is applied to seeds is not particularly limited. For example, 0.05 × 10 －9 ~50 x 10 －３ Part by mass, 0.1×10 －9 ~100 x 10 －３ Part by mass, 0.05×10 －７ ~50 x 10 －４ Part by mass, 0.1×10 －７ ~100 x 10 －４ Part by mass, 0.05×10 －6 ~50 x 10 －5 Parts by mass, or 0.1 x 10 －6 ~100 x 10－5 It can be contained in the range of 0.05 × 10 parts by mass for 100 parts by mass of seeds. －７ ~50 x 10 －４ In a preferred embodiment of the present invention, the active ingredient is a cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof, and the amount of the active ingredient to coat the seeds can be adjusted appropriately depending on the type of plant and the amount of the solution normally applied for pest control.

[0077] 5. Method for Reducing Environmental Stress in Plants 5-1 Overview A fifth aspect of the present invention is a method for reducing environmental stress in plants, which comprises the step of applying the composition described in 2 above to the plants.

[0078] 5-2 Configuration of the method for reducing environmental stress in plants of the present invention The method for reducing environmental stress of the present invention comprises the step of applying to plants a composition containing "a cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof," thereby reducing environmental stress in plants.

[0079] The "cyclodipeptide containing phenylalanine and / or leucine, and / or a salt thereof" of the present invention is as described above in 1-3. In a preferred embodiment of the present invention, the cyclodipeptide containing phenylalanine and / or leucine as an active ingredient is cyclo(leucine-phenylalanine) (cLF).

[0080] The method for reducing environmental stress of the present invention only needs to include the step of applying the composition described in 2 above, and other steps are optional.

[0081] The "environmental stress" of the present invention is as described in 1-4 above, and is preferably salt stress, low temperature stress, or drought stress.

[0082] In the method for reducing environmental stress in plants of the present invention, the plant to which the composition described in 2 above is applied is a plant for which reduction of the above environmental stress is desired, preferably a plant described in 1-6 above.

[0083] Examples of application of the composition described in 2 above to target plants include, but are not limited to, application to the seeds, or above-ground parts such as leaves, stems, fruits, flowers, and roots, or the rhizosphere of the target plants. A preferred embodiment of the method of the present invention is application to seeds. Furthermore, application to seeds may be combined with application to the rhizosphere or above-ground parts such as leaves and stems.

[0084] In the application step, the seeds may be coated with the composition described in 2 above, and then the seeds can be sown in soil. The method for coating seeds with the composition described in 2 above is as described in 3-3 above.

[0085] In the application step, the seeds may be immersed in the solution of the composition described in 2 above, and then the seeds can be sown in soil.

[0086] In the application step of the present invention, the cyclodipeptide containing phenylalanine and / or leucine and / or salts thereof when the composition is applied to seeds is not particularly limited. For example, 0.05 × 10 －9 ~50 x 10 －３ Part by mass, 0.1×10 －9 ~100 x 10 －３ Part by mass, 0.05×10 －７ ~50 x 10 －４ Part by mass, 0.1×10 －７ ~100 x 10 －４ Part by mass, 0.05×10 －6 ~50 x 10 －5 Parts by mass, or 0.1 x 10 －6 ~100 x 10 －5 It can be contained in the range of 0.05 × 10 parts by mass for 100 parts by mass of seeds. －７ ~50 x 10 －４ In a preferred embodiment of the present invention, the active ingredient is a cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof, and the amount of the active ingredient to coat the seeds can be adjusted appropriately depending on the type of plant and the amount of the solution normally applied for pest control.

[0087] The present invention will be described in more detail with reference to the following examples, but the present invention is not limited to these examples.

[0088] In the following Examples and Comparative Examples, cyclodipeptides are abbreviated as follows: cyclo(leucine-phenylalanyl): cLF cyclo(phenylalanine-proline): cFP cyclo(proline-valine): cPV

[0089] Example 1: Cultivation of corn in the absence of environmental stress (cLF concentration: 50 g) <Method> 100 g of corn seeds (variety: Snowdent Otoha) were coated with 800 μL of a coating solution containing cyclo(leucine-phenylalanine) (cLF) or cyclo(proline-valine) (cPV) as a cyclodipeptide and a thickener in water, and the corn seeds were coated with the coating solution using a coating device (CONCEPT ML2000, manufactured by SATEC Corporation). The coating treatment was carried out so that the amount of both cyclodipeptides per ton of seeds was 50 g.

[0090] After filling 7.5 cm pots with soil (Tsuchitarou), the coated corn seeds were sown at one seed per pot. After sowing, the seeds were kept in a closed greenhouse set at 24°C / 14 hours during the day and 18°C / 10 hours at night, and an investigation was carried out 21 days after sowing. The investigation involved measuring the dry weight of the aboveground and underground parts. The test was repeated eight times, and the average value was calculated.

[0091] <Results> As shown in Table 1, under normal conditions, no significant difference was observed in the T-test between cyclo(leucine-phenylalanine) (cLF) and cyclo(proline-valine) (cPV), and no effect of promoting early growth was confirmed.

[0092]

[0093] Example 2: Maize cultivation under salt stress (cLF concentration: 50 g) <Method> 100 g of corn seeds (variety: Snowdent Otoha) were coated with 800 μL of a coating solution containing cyclo(leucine-phenylalanine) (cLF) or cyclo(proline-valine) (cPV) as a cyclodipeptide and a thickener in water, and the corn seeds were coated with the coating solution using a coating device (CONCEPT ML2000, manufactured by SATEC Corporation). The coating treatment was carried out so that the amount of both cyclodipeptides per ton of seeds was 50 g.

[0094] After filling 7.5 cm pots with soil (Tsuchitarou), an aqueous sodium chloride solution was allowed to absorb from the bottom, resulting in 6 g of sodium chloride per liter of soil. The coated corn seeds were sown at one kernel per pot. After sowing, the plants were kept in a closed greenhouse set at 24°C / 14 hours during the day and 18°C / 10 hours at night, and an investigation was carried out 21 days after sowing. The investigation involved measuring the dry weight of the aboveground and underground parts. The test was repeated eight times, and the average value was calculated.

[0095] <Results> As shown in Table 2, when seeds were coated with cyclo(leucine-phenylalanine) (cLF) at a dose of 50 g per ton of seeds, a significant effect of promoting early growth was observed in both above-ground and below-ground parts. On the other hand, when the same dose of cyclo(proline-valine) (cPV) was used, no effect of promoting early growth was observed. These results demonstrate the growth-promoting effect of cyclodipeptides containing phenylalanine under salt stress conditions.

[0096]

[0097] Example 3: Cultivation of corn in the absence of environmental stress (cLF concentration: 2 mg) <Method> 100 g of corn seeds (variety: Snowdent Otoha) were coated with 800 μL of a coating solution containing cyclo(leucine-phenylalanine) (cLF) or cyclo(phenylalanine-proline) (cFP) as a cyclodipeptide and a thickener in water. The corn seeds were then coated with the coating solution using a coating device (CONCEPT ML2000, manufactured by SATEC Corporation). The coating treatment was carried out so that the amount of cyclo(leucine-phenylalanine) (cLF) per ton of seeds was 2 mg and the amount of cyclo(phenylalanine-proline) (cFP) per ton of seeds was 50 g.

[0098] After filling 7.5 cm pots with soil (Tsuchitarou), the coated corn seeds were sown at one seed per pot. After sowing, the seeds were kept in a closed greenhouse set at 24°C / 14 hours during the day and 18°C / 10 hours at night, and an investigation was carried out 21 days after sowing. The investigation involved measuring the dry weight of the aboveground and underground parts. The test was repeated 12 times, and the average value was calculated.

[0099] <Results> As can be seen from Table 3, under normal conditions, no significant difference was observed in the T-test compared to the untreated group for either cyclo(leucine-phenylalanine) (cLF) or cyclo(phenylalanine-proline) (cFP), and no effect of promoting early growth could be confirmed.

[0100]

[0101] Example 4: Maize cultivation under salt stress (cLF concentration: 2 mg) <Method> 100 g of corn seeds (variety: Snowdent Otoha) were coated with 800 μL of a coating solution containing cyclo(leucine-phenylalanine) (cLF) or cyclo(phenylalanine-proline) (cFP) as a cyclodipeptide and a thickener in water. The corn seeds were then coated with the coating solution using a coating device (CONCEPT ML2000, manufactured by SATEC Corporation). The coating treatment was carried out so that the amount of cyclo(leucine-phenylalanine) (cLF) per ton of seeds was 2 mg and the amount of cyclo(phenylalanine-proline) (cFP) per ton of seeds was 50 g.

[0102] After filling 7.5 cm pots with soil (Tsuchitarou), an aqueous sodium chloride solution was allowed to absorb into the bottom, resulting in 6 g of sodium chloride per liter of soil. The coated corn seeds were sown at one kernel per pot. After sowing, the seeds were kept in a closed greenhouse set at 24°C / 14 hours during the day and 18°C / 10 hours at night, and an investigation was carried out 21 days after sowing. The dry weights of the aboveground and underground parts were measured. The test was repeated 12 times and the average value was calculated.

[0103] <Results> As shown in Table 4, when seeds were coated with cyclo(leucine-phenylalanine) (cLF) at a dose of 2 mg per ton of seeds, a significant effect of promoting early growth was observed in the aboveground parts. When seeds were coated with cyclo(phenylalanine-proline) (cFP) at a dose of 50 g, no statistically significant difference was observed, but an effect of promoting early growth was observed in the aboveground parts. These results confirmed that cyclodipeptides containing phenylalanine exhibit growth-promoting effects under salt stress conditions.

[0104]

[0105] Example 5: Effect of the concentration of cyclo(leucine-phenylalanine) used to coat seeds <Method> 100 g of corn seeds (variety: Snowdent Otoha) were coated with 800 μL of a coating solution containing cyclo(leucine-phenylalanine) (cLF) and a thickener in water, and the corn seeds were coated with the coating solution using a coating device (CONCEPT ML2000, manufactured by SATEC Corporation). The coating process was carried out so that the amount of cyclo(leucine-phenylalanine) (cLF) per ton of seeds was 0.05 mg to 10 g.

[0106] After filling 7.5 cm pots with soil (Tsuchitarou), an aqueous sodium chloride solution was allowed to absorb into the bottom, resulting in 6 g of sodium chloride per liter of soil. One coated corn seed was sown per pot. After sowing, the seeds were kept in a closed greenhouse set at 24°C / 14 hours during the day and 18°C / 10 hours at night, and an investigation was carried out 21 days after sowing. The dry weight of the aboveground and underground parts was measured. The test was repeated 12 times, and the average value was calculated.

[0107] <Results> As shown in Table 5, the early growth-promoting effect was observed regardless of the amount of cyclo(leucine-phenylalanine) (cLF) coated per ton of seeds, from 0.05 mg to 10 g. In particular, a significant effect was confirmed when the amount of cyclo(leucine-phenylalanine) (cLF) coated per ton of seeds was 0.1 mg and 0.2 g. This, combined with the experimental data from Examples 2 and 4, confirmed that coating seeds with cyclo(leucine-phenylalanine) (cLF) under salt stress conditions exhibited a growth-promoting effect at the early developmental stage of the plants derived from those seeds.

[0108]

[0109] Example 6: Effect of cyclo(leucine-phenylalanine) on low temperature stress <Method> 100 g of corn seeds (variety: Snowdent Otoha) were coated with 800 μL of a coating solution containing cyclo(leucine-phenylalanine) (cLF) and a thickener in water, using a coating device (CONCEPT ML2000 manufactured by SATEC Corporation) to coat the corn seeds with the coating solution. The coating process was carried out so that the amount of cyclo(leucine-phenylalanine) (cLF) per ton of seeds was 200 mg.

[0110] After filling 7.5 cm pots with soil (Tsuchitarou), the coated corn seeds were sown at one seed per pot. Until four days after sowing, the plants were kept in a closed greenhouse set at 24°C / 14 hours during the day and 18°C / 10 hours at night. From five days after sowing, the plants were kept in an incubator set at 17°C / 14 hours during the day and 12°C / 10 hours at night. 22 days after sowing, the plants were examined. The dry weights of the aboveground and underground parts were measured. The test was repeated 12 times and the average was calculated.

[0111] <Results> From Table 6, it was confirmed that seeds coated with cyclo(leucine-phenylalanine) exhibited an effect of promoting early growth even under low temperature stress.

[0112]

[0113] Example 7: Effect of cyclo(leucine-phenylalanine) on drought stress <Method> 100 g of corn seeds (variety: Snowdent Otoha) were coated with 800 μL of a coating solution containing cyclo(leucine-phenylalanine) (cLF) and a thickener in water, using a coating device (CONCEPT ML2000 manufactured by SATEC Corporation) to coat the corn seeds with the coating solution. The coating process was carried out so that the amount of cyclo(leucine-phenylalanine) (cLF) per ton of seeds was 200 mg.

[0114] After filling 12 cm pots with soil (Tsuchitarou), seven coated corn seeds were sown per pot. Until 6 days after sowing, the plants were kept in a closed greenhouse set at 24°C / 14 hours during the day and 18°C / 10 hours at night, and from 7 to 17 days after sowing, they were kept in an incubator set at 24°C / 14 hours during the day and 18°C / 10 hours at night without irrigation. From 18 to 21 days after sowing, the plants were again moved to a closed greenhouse and kept under normal conditions, and 21 days after sowing, an investigation was carried out. The dry weights of the aboveground and underground parts were measured. The experiment was repeated four times, and the average values ​​were calculated.

[0115] <Results> From Table 7, it was confirmed that seeds coated with cyclo(leucine-phenylalanine) (cLF) exhibited a significant effect of promoting early growth even under drought stress.

[0116]

[0117] The experimental data shown in Examples 1 to 7 above demonstrate that cyclo(leucine-phenylalanine) (cLF) confers tolerance to salt stress, low temperature stress, or drought stress to plants.

[0118] The agent for imparting plant environmental stress tolerance of the present invention, which comprises a cyclodipeptide containing phenylalanine and / or leucine and / or a salt thereof, enables plants to grow even under various environmental stresses, including salt stress. Therefore, the agent for imparting plant environmental stress tolerance of the present invention is useful in agriculture, such as for increasing food production. All publications, patents, and patent applications cited herein are incorporated herein by reference in their entirety.

Claims

1. A plant environmental stress tolerance imparting agent comprising a cyclodipeptide containing phenylalanine and / or leucine, and / or a salt thereof.

2. The agent according to claim 1, wherein the environmental stress is one or more selected from the group consisting of salt stress, low temperature stress, and drought stress.

3. A composition for imparting resistance to environmental stress to plants, comprising the agent according to claim 1 or 2.

4. The composition of claim 3, which confers resistance to environmental stress to the plant at an early growth stage.

5. Seeds containing the composition according to claim 3 on the surface and / or inside thereof.

6. A method for cultivating plants under environmental stress conditions, comprising the step of applying to the plants the composition of claim 3.

7. The method according to claim 6, wherein the environmental stress is one or more selected from the group consisting of salt stress, low temperature stress, and drought stress.

8. A method for reducing environmental stress in plants, comprising the step of applying the composition of claim 3 to the plants.

9. The method according to claim 8, wherein the environmental stress is one or more selected from the group consisting of salt stress, low temperature stress, and drought stress.

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