Plant stomatal opening regulator

Abstract

Provided is a plant stomatal opening regulator. The plant stomatal opening regulator contains at least one selected from the group consisting of compounds represented by general formula (1), salts thereof, and solvates thereof.

Description

Plant stomatal opening regulator

[0001] The present invention relates to a plant stomatal opening regulator and the like.

[0002] Terrestrial higher plants regulate the amount of carbon dioxide taken up and transpiration required for photosynthesis through the regulation of stomata in the leaf epidermis. For example, when exposed to a water shortage, it is known that plants close their stomata and suppress transpiration in order to maintain moisture within the plant. It is also known that stomata open in response to light, promoting the uptake of carbon dioxide required for photosynthesis. Therefore, artificial regulation of stomatal opening can be expected to have effects such as promoting photosynthesis, stimulating growth, and improving drought tolerance.

[0003] An example of artificial regulation of stomatal opening is the plasma membrane H ＋ A method for overexpressing the α-ATPase (AHA2) has been reported (Patent Document 1, Non-Patent Document 1). This method promotes stomatal opening, which in turn promotes photosynthetic rate and plant growth. Another method for improving drought tolerance has also been reported (Non-Patent Document 2), in which the magnesium chelatase H subunit is overexpressed in guard cells to promote stomatal closure.

[0004] Because these methods rely on genetic engineering, 1. For species for which genetic engineering technology has not yet been established, it is necessary to establish and optimize the technology each time. 2. It takes time to create genetically modified organisms (GMOs). For example, it takes about six months to create a transgenic poplar, and more than six months to obtain the next generation of GMO seeds for rice. 3. Approval is required each time a GMO is created.

[0005] In contrast, if there were a substance that could regulate stomatal opening by applying it to already growing plants, it would be extremely useful, as it would not require the procedures of creating and approving GMOs for various plants.

[0006] Patent Document 2 reports that a compound called SCL1 has the effect of regulating stomatal opening.

[0007] International Publication No. 2014 / 142334 Japanese Patent Application Laid-Open No. 2020-055767

[0008] PNAS, January 7, 2014, vol.111, no.1, pp.533-538Front Plant Sci, October 30, 2013, vol.4, Article440

[0009] An object of the present invention is to provide a plant stomatal opening regulator.

[0010] As a result of intensive research in view of the above problems, the present inventors have found that each compound represented by general formula (1) has a stomatal opening regulating effect. The present invention has been completed as a result of further research based on this finding. That is, the present invention includes the following aspects.

[0011] Item 1. General formula (1):

[0012]

[0013] A plant stomatal opening regulator comprising at least one compound selected from the group consisting of a compound represented by the formula: [wherein A represents an optionally substituted ring; L is the same or different and represents a linker; and n represents an integer of 1 to 6], a salt thereof, and a solvate thereof.

[0014] Item 2. The plant stomatal opening regulator according to item 1, wherein A is a monocyclic 5- or 6-membered ring.

[0015] Item 3. The plant stomatal opening regulator according to claim 1 or 2, wherein A is an unsubstituted ring or a ring substituted with at least one selected from the group consisting of an optionally substituted alkyl group, an optionally substituted alkoxy group, an optionally substituted alkoxycarbonyl group, an optionally substituted aryl group, a cyano group, a halogen atom, a nitro group, an optionally substituted amino group, an optionally substituted amido group, an optionally substituted ketone group, an optionally substituted ammonio group, and an optionally substituted thio group.

[0016] Item 4. The plant stomatal opening regulator according to any one of items 1 to 3, wherein A has 0 to 3 substituents.

[0017] Item 5. The plant stomatal opening regulator according to any one of items 1 to 4, wherein L is an alkylene group optionally substituted with an alkyl group and / or an aryl group.

[0018] Item 6. The plant stomatal opening regulator according to any one of claims 1 to 5, wherein A is a monocyclic 5- or 6-membered ring, L is an alkylene group optionally substituted with an alkyl group and / or an aryl group, and when A is a benzene ring and n is 1, A is a benzene ring substituted with at least one selected from the group consisting of an optionally substituted aryl group, a cyano group, an iodine atom, and an optionally substituted alkoxycarbonyl group, and / or L is an alkylene group substituted with an alkyl group and / or an aryl group.

[0019] Item 7. The plant stomatal opening regulator according to any one of items 1 to 6, which is a plant stomatal opening inhibitor.

[0020] Item 8. A drought tolerance improver comprising the plant stomatal opening regulator according to any one of items 1 to 7.

[0021] Item 9. The drought tolerance improver according to claim 8, for use in suppressing wilting.

[0022] Item 10. A method for improving drought tolerance of a plant, comprising applying the plant stomatal opening regulator according to any one of items 1 to 7 to the plant.

[0023] Item 11. A method for improving drought tolerance according to item 10, which comprises contacting the plant stomatal opening regulator according to any one of items 1 to 7 with the stomata of a plant.

[0024] Item 12. General formula (1):

[0025]

[0026] [wherein A represents an optionally substituted ring, L represents a linker, and n represents an integer of 1 to 6], a salt thereof, and a solvate thereof. A cell membrane proton pump phosphorylation inhibitor comprising at least one compound selected from the group consisting of a compound represented by the formula:

[0027] Item 13. General formula (1):

[0028]

[0029] A plant covering or supporting material comprising at least one compound selected from the group consisting of a compound represented by the formula: [wherein A represents an optionally substituted ring, L represents a linker, and n represents an integer of 1 to 6], a salt thereof, and a solvate thereof.

[0030] Section 14. General formula (1X):

[0031]

[0032] [In the formula: A' represents an optionally substituted non-aromatic ring; L's are the same or different and represent a linker; and n' represents an integer of 2 to 6], a salt thereof, or a solvate thereof.

[0033] The clauses specifying the above-mentioned agents can be replaced with "use of the active ingredient of the present invention for the manufacture of the agent," "use of the active ingredient of the present invention for the application of the agent," "active ingredient of the present invention for use in the application of the agent," etc.

[0034] According to the present invention, it is possible to provide a plant stomatal opening regulator and a compound that serves as an active ingredient of the regulator. It is also possible to provide a drought tolerance improver, a cell membrane proton pump phosphorylation inhibitor, etc.

[0035] The measurement results of Test Example 2 are shown. The vertical axis indicates stomatal aperture. On the horizontal axis, Dark indicates the dark-treated group, Light indicates the light-treated group, Ctl indicates the case where a test solution containing no test compound was used, ABA indicates the case where a test solution containing abscisic acid as the test compound was used, BITC indicates the case where a test solution containing BITC (compound 1) as the test compound was used, and +FC indicates the case where a test solution containing fusicoccin in addition to the test compound was used. The measurement results of Test Example 3 are shown. The vertical axis indicates the signal intensity of the phosphorylated cell membrane proton pump. On the horizontal axis, Ctrl indicates the case where no test compound was applied, BITC, m-Bis-BITC, and Tris-BITC indicate the case where the respective test compounds were applied, R indicates the case where treatment was performed with red light only, RB indicates the case where treatment was performed with red light and blue light, and FC indicates the case where fusicoccin was applied. The measurement results of Test Example 4 are shown. At the top of the photograph, Ctrl indicates the case without the test compound, BITC indicates the case with BITC (compound 1), and ABA indicates the case with abscisic acid. The detected proteins are shown on the right side of the photograph, with 61 kDa indicating an ABA-responsive kinase substrate and 14-3-3 indicating the 14-3-3 protein used as a loading control. The stomatal aperture measurement results after 3 hours of incubation in Test Example 5 are shown. The vertical axis indicates stomatal aperture. On the horizontal axis, Ctrl indicates the case where a test solution containing no test compound was used, while BITC and m-Bis-BITC indicate the case where a test solution containing the respective test compound was used. The numbers indicate the concentration of the test compound in the test solution (unit: μM). The stomatal aperture measurement results after 3 hours or 48 hours of incubation in Test Example 5 are shown. The vertical axis indicates stomatal aperture. On the horizontal axis, the numbers indicate the incubation time, and ABA, BITC, and m-Bis-BITC indicate the case where a test solution containing the respective test compound was used. The leftmost column shows the results when a test solution containing no test compound was used. The test compound concentrations in the test solution were 100 μM for ABA, 2500 μM for BITC, and 50 μM for m-Bis-BITC. Photographs of the external appearance of Test Example 6 are shown.Ctrl indicates the case where a test solution containing no test compound was used, while BITC and m-Bis-BITC indicate the case where a test solution containing the respective test compounds was used, and the numerical values ​​indicate the concentration of the test compound in the test solution. The upper row shows a photograph of the appearance at the start of incubation in a dehydrated state, and the lower row shows a photograph of the appearance at the end of incubation in a dehydrated state. Photographs of the appearance in Test Example 7 are shown. Ctrl indicates the case where a test solution containing no test compound was used, while m-Bis-BITC indicates the case where a test solution containing the respective test compound was used. The upper row shows a photograph of the appearance at the start of incubation in a water-depleted state, and the lower row shows a photograph of the appearance at the end of incubation in a water-depleted state. Compound 27 synthesized in Test Example 1-1-3. 1 The results of H NMR (500 MHz, CDCl3) are shown below.

[0036] In this specification, the expressions "contain" and "comprise" include the concepts of "contain", "include", "consist essentially of" and "consist only of".

[0037] In one aspect, the present invention provides a compound represented by general formula (1):

[0038]

[0039] The present invention relates to a plant stomatal opening regulator (sometimes referred to herein as "the plant stomatal opening regulator of the present invention") containing at least one compound (sometimes referred to herein as "the compound of the present invention") selected from the group consisting of a compound represented by the formula (sometimes referred to herein as "the compound of the present invention"), a salt thereof, and a solvate thereof. The compound of the present invention, the active ingredient of the present invention, and their uses are described below.

[0040] 1. Active ingredient A represents an optionally substituted ring. Note that "substituted" here does not include substitution by -L-N=C=S.

[0041] The ring is not particularly limited, and includes aromatic rings composed only of hydrocarbons, aromatic rings containing heteroatoms other than carbon and hydrogen (heterocycles), non-aromatic rings composed only of hydrocarbons (alicyclic hydrocarbons), and non-aromatic rings containing heteroatoms other than carbon and hydrogen (heterocycles). The heteroatoms contained in the heterocycle are not particularly limited, and examples thereof include sulfur atoms, nitrogen atoms, and oxygen atoms. The ring includes both monocyclic rings (e.g., 4- to 8-membered rings) and fused rings (e.g., bicyclic or tricyclic rings). Examples of aromatic rings include monocyclic rings such as benzene, thiophene, thiazole, furan, pyrrole, imidazole, pyrazole, oxazole, isoxazole, triazole, tetrazole, pyridine, pyrazine, pyrimidine, pyridazine, and triazine; bicyclic rings such as naphthalene, benzothiophene, benzofuran, indole, benzimidazole, indazole, benzoxazole, benzothiazole, isobenzofuran, isoindole, purine, quinoline, isoquinoline, quinoxaline, quinazoline, cinnoline, and phthalazine; and tricyclic rings such as anthracene and phenanthrene. Examples of non-aromatic rings include cycloalkanes such as cyclopropane, cyclobutane, cyclopentane, cyclohexane, cycloheptane, cyclooctane, cyclononane, cyclodecane, cycloundecane, and cyclododecane; cycloalkenes such as cyclopropene, cyclobutene, cyclopropene, cyclohexene, cycloheptene, and cyclooctene; bicyclic alkanes such as bicycloundecane and decahydronaphthalene; and bicyclic alkenes such as bicycloundecane and decahydronaphthalene.

[0042] The ring is preferably a monocyclic ring from the viewpoint of regulating plant stomatal opening. The ring is preferably a 5- or 6-membered ring from the viewpoint of regulating plant stomatal opening. When the ring is a heterocyclic ring, the heteroatom contained in the heterocyclic ring preferably contains a sulfur atom, more preferably only sulfur atoms, from the viewpoint of regulating plant stomatal opening. The ring is preferably an aromatic ring from the viewpoint of regulating plant stomatal opening.

[0043] The ring is an unsubstituted ring or a ring substituted with a hydrogen atom.The substituent of the ring is not particularly limited, but for example, an optionally substituted alkyl group, an optionally substituted alkoxy group, an optionally substituted alkoxycarbonyl group, an optionally substituted aryl group, a cyano group, a halogen atom, a nitro group, an optionally substituted amino group, an optionally substituted amide group, an optionally substituted ketone group, an optionally substituted ammonio group, an optionally substituted thio group, etc. are mentioned.Among these, an optionally substituted alkyl group, an optionally substituted alkoxy group, an optionally substituted alkoxycarbonyl group, an optionally substituted aryl group, a cyano group, and a halogen atom are more preferred, and an optionally substituted aryl group, a cyano group, an iodine atom, and an optionally substituted alkoxycarbonyl group are particularly preferred.

[0044] The alkyl group includes any of linear, branched, and cyclic alkyl groups. From the viewpoint of the plant stomatal opening regulation effect, the alkyl group is preferably linear or branched, and more preferably linear. The number of carbon atoms in the alkyl group is not particularly limited and may be, for example, 1 to 8. From the viewpoint of the plant stomatal opening regulation effect, the number is preferably 1 to 6, more preferably 1 to 4, even more preferably 1 to 2, and particularly preferably 1. Specific examples of the alkyl group include a methyl group, an ethyl group, an n-propyl group, an isopropyl group, an n-butyl group, an isobutyl group, a tert-butyl group, a sec-butyl group, an n-pentyl group, a neopentyl group, an n-hexyl group, and a 3-methylpentyl group. The alkyl group may be substituted with a halogen atom such as a fluorine atom, a chlorine atom, a bromine atom, or an iodine atom.

[0045] The alkoxy group includes both linear and branched groups. From the viewpoint of the plant stomatal opening regulation effect, the alkoxy group is preferably linear. The number of carbon atoms in the alkoxy group is not particularly limited and may be, for example, 1 to 8. From the viewpoint of the plant stomatal opening regulation effect, the number of carbon atoms is preferably 1 to 6, more preferably 1 to 4, even more preferably 1 to 2, and particularly preferably 1. Specific examples of the alkoxy group include a methoxy group, an ethoxy group, an n-propoxy group, an isopropoxy group, an n-butoxy group, an isobutoxy group, a sec-butoxy group, and a tert-butoxy group. The alkoxy group may be substituted with a halogen atom such as a fluorine atom, a chlorine atom, a bromine atom, or an iodine atom.

[0046] The alkoxycarbonyl group is a group formed by linking an alkoxy group and a carbonyl group. The alkoxy group is as described above.

[0047] The aryl group is not particularly limited, but from the viewpoint of plant stomatal opening regulation, it preferably has 6 to 12 carbon atoms, and more preferably 6 to 8 carbon atoms. The aryl group may be either monocyclic or polycyclic (e.g., bicyclic, tricyclic, etc.), but from the viewpoint of plant stomatal opening regulation, it is preferably monocyclic. Specific examples of the aryl group include phenyl, naphthyl, biphenyl, pentalenyl, indenyl, anthranyl, tetracenyl, pentacenyl, pyrenyl, perylenyl, fluorenyl, and phenanthryl groups, with phenyl being preferred. The aryl group may be substituted with a halogen atom such as a fluorine atom, a chlorine atom, a bromine atom, or an iodine atom.

[0048] The halogen atom is not particularly limited, and examples thereof include a fluorine atom, a chlorine atom, a bromine atom, and an iodine atom.

[0049] The hydrogen atoms of the amino group (-NH2), amide group (-CO-NH2), ketone group (-CO-H), ammonio group (-NH3), and thio group (-SH) may be substituted with hydrocarbon groups such as alkyl groups and aryl groups. The alkyl groups and aryl groups are as explained above.

[0050] As the substituent on the ring, an optionally substituted aryl group is particularly preferred from the viewpoint of the plant stomatal opening regulating action.

[0051] The number of substituents on the ring (substituents other than -L-N=C=S) is not particularly limited, but is, for example, 0 to 6. From the viewpoint of the plant stomatal opening regulating effect, the number of substituents is preferably 0 to 3, more preferably 0 to 2, and particularly preferably 0 to 1.

[0052] In the compound of the present invention, the optionally substituted ring represented by A has n (1 to 6) hydrogen atoms on the ring substituted with -L-N=C=S, thereby enabling the compound of the present invention to exert an excellent effect of regulating plant stomatal opening.

[0053] L may be the same or different and represents a linker.

[0054] The linker includes a chain structure. The chain structure is not particularly limited, but includes, for example, a chain structure having 1 to 20 atoms constituting the main chain. From the viewpoint of plant stomatal opening regulation, the number of atoms constituting the main chain is preferably 1 to 15, more preferably 1 to 12, even more preferably 1 to 10, even more preferably 1 to 8, particularly preferably 1 to 6, and especially preferably 1 to 2. Examples of atoms constituting the main chain include carbon atoms, oxygen atoms, sulfur atoms, and nitrogen atoms. Part or all of the chain structure can be an alkylene group or a heteroalkylene group. The chain structure can be either linear or branched, but from the viewpoint of plant stomatal opening regulation, a linear structure is preferred. Furthermore, the chain structure can include, on the main chain, a partial structure such as -O-, -C(=O)-O-, -CO-NH-, -C(=O)-, -NH-, or -S(=O)2-. The chain structure may be substituted with an alkyl group and / or an aryl group. The alkyl and alkylene groups are the same as those described above.

[0055] From the viewpoint of the plant stomatal opening regulation effect, the linker is preferably an alkylene group which may be substituted with an alkyl group and / or an aryl group, and particularly preferably an alkylene group substituted with an alkyl group and / or an aryl group.

[0056] n represents an integer of 1 to 6.

[0057] From the viewpoint of the plant stomatal opening regulating effect, n is particularly preferably 2 or more. The upper limit of n is, for example, 5, 4, or 3. In one embodiment of the present invention, from the viewpoint of the plant stomatal opening regulating effect, n is preferably 2 to 5, more preferably 2 to 4, and even more preferably 2 to 3. From the viewpoint of the plant stomatal opening regulating effect when applied to intact leaves, n is particularly preferably 2.

[0058] In one embodiment of the present invention, preferably, A is a monocyclic 5- to 6-membered ring, L is an alkylene group which may be substituted with an alkyl group and / or an aryl group, and when A is a benzene ring and n is 1, A is a benzene ring which may be substituted with at least one selected from the group consisting of an aryl group, a cyano group, an iodine atom, and an alkoxycarbonyl group which may be substituted, and / or L is an alkylene group substituted with an alkyl group and / or an aryl group.

[0059] In one embodiment of the present invention, among the general formula (1), the general formula (1X) is preferably:

[0060]

[0061] Examples include:

[0062] A' represents an optionally substituted non-aromatic ring; L's may be the same or different and represent a linker; and n' represents an integer of 2 to 6.

[0063] In one embodiment of the present invention, among the general formula (1), the general formula (1Y):

[0064]

[0065] In one embodiment of the present invention, among the compounds represented by general formula (1), compounds represented by general formula (1A):

[0066]

[0067] Examples include:

[0068] A" is a benzene ring or cyclohexane. In one embodiment, A" is a benzene ring, and in another embodiment, it is cyclohexane.

[0069] L 1 are the same or different and represent an alkylene group which may be substituted with an alkyl group and / or an aryl group.

[0070] R 1 are the same or different and represent an optionally substituted alkyl group, an optionally substituted alkoxy group, an optionally substituted alkoxycarbonyl group, an optionally substituted aryl group, a cyano group, a halogen atom, a nitro group, an optionally substituted amino group, an optionally substituted amide group, an optionally substituted ketone group, an optionally substituted ammonio group, or an optionally substituted thio group. Among these (in one embodiment, when A" is a benzene ring, or when A" is a benzene ring and n is 1), an optionally substituted aryl group, a cyano group, an iodine atom, and an optionally substituted alkoxycarbonyl group are particularly preferred, and an optionally substituted aryl group is particularly preferred.

[0071] L 1 , R 1 The group represented by is as explained above.

[0072] n is the same as above.

[0073] m is a number that satisfies 0≦m≦6−n, and m is preferably 0 to 3, more preferably 0 to 2, and particularly preferably 0 to 1, from the viewpoint of the plant stomatal aperture regulating effect.

[0074] In one embodiment of the present invention, among the general formula (1), the general formula (1Z):

[0075]

[0076] In one embodiment of the present invention, among the compounds represented by general formula (1), compounds represented by general formula (1B):

[0077]

[0078] Examples include:

[0079] A" is a benzene ring or cyclohexane. In one embodiment, A" is a benzene ring, and in another embodiment, it is cyclohexane.

[0080] R 11 , R 12 , R 13 , R 14 , and R 15 are the same or different and represent a hydrogen atom, -L-N=C=S, an optionally substituted alkyl group, an optionally substituted alkoxy group, an optionally substituted alkoxycarbonyl group, an optionally substituted aryl group, a cyano group, a halogen atom, a nitro group, an optionally substituted amino group, an optionally substituted amido group, an optionally substituted ketone group, an optionally substituted ammonio group, or an optionally substituted thio group. Among these, an optionally substituted aryl group, a cyano group, an iodine atom, and an optionally substituted alkoxycarbonyl group are particularly preferred, and an optionally substituted aryl group is particularly preferred.

[0081] L may be the same or different and represents a linker.

[0082] L 1 , R 11 , R 12 , R 13 , R 14 , and R 15 The group represented by is as explained above.

[0083] In one embodiment (embodiment 1) of the present invention, R 12 and R 13 At least one of is -L-N=C=S, and R 13 If -L-N=C=S, then R 11 , R 12 , R 14 , and R 15 In embodiment 1, at least one of R is a group other than a hydrogen atom. 14 and R 15 is a hydrogen atom. In embodiment 1, preferably R 12 and R 13 In embodiment 1, one of R is -L-N=C=S and the other is a hydrogen atom. 12When -L-N=C=S, preferably R 11 is a hydrogen atom. 13 When -L-N=C=S, preferably R 11 is an alkyl group or -L-N=C=S.

[0084] The compound represented by general formula (1) may include stereoisomers and optical isomers, but these are not particularly limited.

[0085] The salt of the compound represented by formula (1) is not particularly limited as long as it is agriculturally acceptable. Both acidic and basic salts can be used. Examples of acidic salts include inorganic acid salts such as hydrochloride, hydrobromide, sulfate, nitrate, and phosphate; organic acid salts such as acetate, propionate, tartrate, fumarate, maleate, malate, citrate, methanesulfonate, and paratoluenesulfonate. Examples of basic salts include alkali metal salts such as sodium salt and potassium salt; alkaline earth metal salts such as calcium salt and magnesium salt; salts with ammonia; and salts with organic amines such as morpholine, piperidine, pyrrolidine, monoalkylamines, dialkylamines, trialkylamines, mono(hydroxyalkyl)amines, di(hydroxyalkyl)amines, and tri(hydroxyalkyl)amines.

[0086] The compound represented by formula (1) can also be converted into a hydrate or solvate. Examples of the solvent include agriculturally acceptable organic solvents (e.g., ethanol, glycerol, acetic acid, etc.).

[0087] As the compound represented by general formula (1), for example, a known compound (e.g., a commercially available product) can be used. Examples of known compounds include the compounds shown in Tables 1 to 3. In the tables, -NCS represents an isothiocyanate group (-N=C=S).

[0088]

[0089]

[0090]

[0091] The compound represented by general formula (1) can be, for example, a compound represented by general formula (a):

[0092]

[0093] [wherein A, L, and n are as defined above, and X represents a halogen atom], and a method comprising reacting a compound represented by the formula (compound a) with a thiocyanate. Alternatively, when X is replaced with an amino group, the compound can also be produced by reacting the compound with a compound containing thione-type sulfur, such as di(1H-imidazol-1-yl)methanethione.

[0094] Compound a can be commercially available or synthesized according to a known method.

[0095] Examples of thiocyanates include potassium thiocyanate and sodium thiocyanate, with potassium thiocyanate being preferred.

[0096] In view of yield, ease of synthesis, etc., the amount of thiocyanate used is usually preferably 3 to 30 moles, more preferably 7 to 15 moles, per mole of compound a.

[0097] This reaction is usually carried out in the presence of a reaction solvent. The reaction solvent is not particularly limited, but examples thereof include dimethylformamide, dichloromethane, acetonitrile, tetrahydrofuran, acetone, toluene, etc., and preferably dimethylformamide, etc. The solvent may be used alone or in combination.

[0098] The reaction can be carried out under heating, at room temperature, or under cooling, and is usually carried out at a temperature of 0 to 120° C. The reaction temperature is particularly preferably 70 to 100° C. The reaction time is not particularly limited, and is usually 30 minutes to 60 hours.

[0099] The progress of the reaction can be monitored by conventional methods such as chromatography. After the reaction is completed, the solvent is removed by distillation, and the product can be isolated and purified by conventional methods such as chromatography and recrystallization. The structure of the product can also be confirmed by elemental analysis, MS (ESI-MS), IR analysis, etc. 1 H-NMR, 13 It can be identified by C-NMR or the like.

[0100] 2. Uses The active ingredient of the present invention has the effect of regulating plant stomatal opening. Furthermore, regulation of stomatal opening can also regulate photosynthesis and plant growth. Therefore, at least one compound selected from the group consisting of compounds represented by general formula (1), salts thereof, and solvates thereof can be used as an active ingredient in photosynthesis regulators, plant growth regulators, etc.

[0101] In particular, compounds represented by general formula (1) have the effect of inhibiting plant stomatal opening (particularly stomatal opening in response to light or chemicals (such as fusicoccin)). It is also known that the inhibition of plant stomatal opening inhibits transpiration and maintains the water content within the plant. Therefore, at least one compound selected from the group consisting of compounds represented by general formula (1), salts thereof, and solvates thereof can be used as an active ingredient in plant stomatal opening inhibitors, drought resistance improvers, wilting inhibitors, freshness-preserving agents, and the like. It is also known that pathogenic microorganisms enter plants through the openings of their stomata. Therefore, at least one compound selected from the group consisting of compounds represented by general formula (1), salts thereof, and solvates thereof can be used as an active ingredient in disease resistance improvers, and the like.

[0102] Furthermore, it is believed that the inhibition of phosphorylation of the plasma membrane proton pump is part of the mechanism of action of the compound represented by general formula (1) to regulate stomatal opening. Therefore, at least one compound selected from the group consisting of the compound represented by general formula (1), its salt, and their solvates can be used as an active ingredient of a plasma membrane proton pump phosphorylation inhibitor. The target plasma membrane proton pump is not particularly limited as long as it is expressed in guard cells. For example, in the case of Arabidopsis, these include AHA1 (AT2G18960), AHA2 (AT4G30190), AHA3 (AT5G57350), AHA4 (AT3G47950), AHA5 (AT2G24520), AHA6 (AT2G07560), AHA7 (AT3G60330), AHA8 (AT3G42640), AHA9 (AT1G80660), AHA10 (AT1G17260), and AHA11 (AT5G62670). In the case of rice, these include OSA1 (LOC_Os03g48310), OSA2 (LOC_Os07g09340), OSA3 (LOC_Os12g44150), OSA4 (LOC_Os05g25550), OSA5 (LOC_Os08g14360), OSA6 (LOC_Os02g55400), OSA7 (LOC_Os04g56160), OSA8 (LOC_Os03g01120), OSA9 (LOC_Os03g08560), and OSA10 (LOC_Os06g08310).

[0103] The target plants of the plant stomatal opening regulator of the present invention are not particularly limited as long as they have stomata. For example, it can be widely applied to plants such as angiosperms (dicotyledons, monocotyledons, etc.), gymnosperms, ferns, etc. Specific examples include eggplants such as tomatoes, bell peppers, chili peppers, eggplants, etc.; gourds such as cucumbers, pumpkins, melons, watermelons, etc.; vegetables such as cabbage, broccoli, Chinese cabbage, etc.; fresh or spicy vegetables such as celery, parsley, lettuce, etc.; alliums such as leeks, onions, garlic, etc.; beans such as soybeans, peanuts, kidney beans, peas, adzuki beans, etc.; other fruit vegetables such as strawberries; taproots such as radishes, turnips, carrots, burdock, etc.; tubers such as taro, cassava, potato, sweet potato, Chinese yam, etc.; asparagus, spinach, mitsuba, etc. soft vegetables, flowering plants such as lisianthus, stock, carnation, chrysanthemum, etc.; cereals such as rice, wheat, barley, oats, corn, etc.; turf grasses such as bentgrass and Zoysiagrass, oil crops such as rapeseed and peanut, sugar crops such as sugarcane and sugar beet, fiber crops such as cotton and rush, forage crops such as clover, sorghum, dent corn, deciduous fruit trees such as apples, pears, grapes, peaches, etc.; citrus fruits such as Satsuma mandarins, lemons, and grapefruit, and woody plants such as azalea, azalea, and cedar.

[0104] The plant stomatal opening regulator of the present invention may consist solely of the above-mentioned agent, or may contain various additives in addition to the above-mentioned agent depending on the dosage form, application mode, etc., as described below. The content of the above-mentioned agent in the plant stomatal opening regulator can be determined appropriately depending on the dosage form, application mode, etc., as described below, and can be, for example, in the range of 0.0001 to 100 mass%. More specifically, when the plant stomatal opening regulator of the present invention is in the form of a liquid formulation and is brought into contact with stomata, the content of the above-mentioned agent is, for example, about 1 to 5000 μM, preferably 5 to 3000 μM, more preferably 10 to 1000 μM, even more preferably 20 to 500 μM, and even more preferably about 20 to 200 μM.

[0105] The formulation of the plant stomatal opening regulator of the present invention is not particularly limited as long as it is an agriculturally acceptable formulation, and examples thereof include liquid formulations, solid formulations, dust formulations, granules, granules, wettable powders, flowable formulations, emulsifiable concentrates, pastes, and dispersions.

[0106] The additives are not particularly limited as long as they are agriculturally acceptable additives, and examples thereof include carriers, surfactants, spreading agents, spray adjuvants, thickeners, extenders, binders, vitamins, antioxidants, pH adjusters, evaporation inhibitors, and pigments.

[0107] Among these, plant water repellency inhibitors such as silicone surfactants are particularly preferred. The content of the silicone surfactant may vary depending on the formulation, but in the case of a liquid formulation, for example, it is 0.01 to 0.1% by mass, preferably 0.02 to 0.05% by mass, relative to 100% by mass of the plant stomatal opening regulator of the present invention.

[0108] The application mode of the plant stomatal opening regulator of the present invention is not particularly limited, as long as it is a known application mode for pesticides (or a mode to be developed in the future). Examples include spraying, dripping, coating, mixing or dissolving in the plant growth environment (soil, water, solid medium, liquid medium, etc.). Since the target of action of the plant stomatal opening regulator of the present invention is stomata, it is preferably applied by contacting the plant stomata with the plant.

[0109] The active ingredient of the present invention can be used by being contained in a plant covering or supporting material. From this viewpoint, in one aspect, the present invention relates to a plant covering or supporting material containing the active ingredient of the present invention.

[0110] Plant covering materials are materials for covering part or all of a plant, and are not particularly limited as long as they are. The shape of the plant covering material is not particularly limited, but is preferably in sheet form. The material of the plant covering material is not particularly limited, and can be paper, plastic, etc. Specific examples of plant covering materials include plant packaging sheets, agricultural sheets (sheets for agricultural greenhouses), and sheets for preserving the freshness of food (for example, sheets used inside containers (e.g., lunch boxes) in which food containing plants can be placed).

[0111] Plant support materials are materials that support plants so that they do not fall over, and are not particularly limited thereto. Plant support materials are typically used by inserting or supporting plant stems or roots. The material of the plant support material is not particularly limited, and can be paper, plastic, or the like. Examples of plant support materials include porous materials such as sponges and fiber aggregates, and more specific examples include solid culture media (e.g., rockwool pots) and absorbent sponges that are mainly used in flower arranging and other floricultural purposes.

[0112] The active ingredient of the present invention may constitute a surface layer of the coating or supporting material, or may be contained throughout the coating or supporting material.

[0113] The active ingredient of the present invention in the plant coating or support material can exert its effect on plants by contacting the plant with the coating or support material, or by volatilizing from the coating or support material and coming into contact with the plant.

[0114] The present invention will be described in detail below based on examples, but the present invention is not limited to these examples.

[0115] Test Example 1. Measurement of stomatal opening regulation activity Test 1 Compounds 1 to 27 (Tables 4 to 10) were prepared and their stomatal opening regulation activity was measured. Compounds 3, 5, and 27 were synthesized as follows. At least compound 27 is a new compound. The other compounds were synthesized according to or similar to previously reported methods, or were purchased commercially.

[0116] Test Example 1-1. Synthesis of Compounds Test Example 1-1-1. Synthesis of Compound 3 Di(1H-imidazol-1-yl)methanethione (10 mmol, 5 equiv.) and DMF (15 mL) were added to a dry round-bottom flask equipped with a stir bar. A solution of EtN (4.8 mmol, 2.4 equiv.) and 1,4-phenylenedimethanamine (2.0 mmol, 1.0 equiv.) in DMF (5 mL) was added to the flask, and the mixture was stirred at 50 °C overnight. The reaction mixture was quenched with HO (20 mL) and extracted with hexane / EtOAc = 1:1 (20 mL × 3). The organic layer was washed with 1 M aqueous HCl and water (3 times). The organic layer was then dried over NaSO, filtered, and concentrated. The crude product was purified by chromatography on silica gel to give compound 3 (221 mg, 1.00 mmol, 50%) as a pale yellow solid. 1 H NMR (500 MHz, CDCl3) δ 4.73 (s, 4H), 7.35 (s, 4H); 13 C NMR (126 MHz, CDCl3) δ 48.37, 127.52, 132.83, 134.59; LRMS (GC-MS, EI): m / z = 220 calcd for C 10 H8N2S2220 [M + ].

[0117] Test Example 1-1-2. Synthesis of Compound 5

[0118]

[0119] To a dry 100 mL round-bottom flask equipped with a stir bar were added KSCN (1.36 g, 14 mmol, 10 equiv.), NaI (900 mg, 6 mmol, 4.3 equiv.), and 1,3,5-tris(bromomethyl)benzene (498 mg, 1.4 mmol, 1.0 equiv.). Dimethylformamide (10 mL) was then added to the flask, and the mixture was stirred at 90 °C for 24 h. The reaction mixture was diluted with HO (20 mL) and extracted with diethyl ether (3 × 20 mL). The organic layer was dried over MgSO and then concentrated under reduced pressure. The crude product was purified by flash column chromatography on silica gel (hexane / EtOAc = 100:0 to 70:30) and preparative recycling gel permeation chromatography to give 1,3,5-tris(isothiocyanatomethyl)benzene (137 mg, 0.47 mmol, 34%). 1 H NMR (500 MHz, CDCl3) δ 4.79 (s, 6H), 7.26 (s, 3H); 13 C NMR (126 MHz, CDCl3) δ 48.38, 125.29, 133.89, 136.39; LRMS(GC-MS): m / z= 291 calcd for 291 [M + ].

[0120] Experimental Example 1-1-3. Synthesis of Compound 27. Di(1H-imidazol-1-yl)methanethione (1.78 g, 10 mmol, 5 equiv.) and DMF (15 mL) were added to a dry 100 mL round-bottom flask equipped with a stir bar. Next, a solution of triethylamine (0.55 mL, 9.6 mmol, 4.8 equiv.) and trans-1,4-bis(aminomethyl)cyclohexane (280 mg, 2.0 mmol, 1.0 equiv.) in DMF (5 mL) was added to the flask, and the mixture was stirred at 50 °C for 16 h. The reaction mixture was diluted with HO (20 mL) and extracted with ethyl acetate (3 × 20 mL). The organic layer was dried over sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash column chromatography on silica gel to give trans-1,4-bis(isothiocyanatomethyl)cyclohexane (315 mg, 1.39 mmol, 70%) as a white solid.

[0121] 1 The results of H NMR (500 MHz, CDCl3) are shown in Figure 8. Chemical Formula: C 10 H 14 N2S2, Exact Mass: 226.06, Molecular Weight: 226.36.

[0122] Test Example 1-2: Measurement test As test solutions, stomatal aperture measurement solutions (5 mM MES-BTP [pH 6.5], 50 mM KCl, 0.1 mM CaCl, 0.5% DMSO) were prepared by dissolving each of the test compounds (compounds 1 to 27) at various concentrations.

[0123] A portion of a stem from Commelina benghalensis was inserted into a planter filled with a mixture of vermiculite and peat moss and grown for 2-4 weeks in a room with natural light (room temperature 25±3°C). The planter was then placed in a dark room and darkened overnight. After darkening, a 4mm diameter leaf disk was excised from a fully expanded leaf using a hole punch (Biospy Punch, Kai Medical). ​

[0124] Each test solution was dispensed into five wells of a 96-well plate at 50 μl / well, and the leaf disks were immersed in the solution. -2 s -1 The mixture was incubated under illumination with blue light for 3 hours. After incubation, stomatal closure was evaluated under observation images (600x) using a stereofluorescence microscope (Leica M205FA). The minor diameters of approximately 20 stomata per leaf disc (hereinafter referred to as "stomatal aperture") were measured. Based on the measured values ​​of stomatal aperture at each concentration for each test compound, the 50% inhibitory concentration of stomatal aperture (IC 50 ) was calculated.

[0125] The results are shown in Tables 4 to 9. In the tables, -NCS represents an isothiocyanate group (-N=C=S), -Me represents a methyl group, and -Ph represents a phenyl group.

[0126]

[0127]

[0128]

[0129]

[0130]

[0131]

[0132]

[0133] Test Example 2. Measurement test 2 of stomatal opening regulation activity Compound 1 (BITC: benzyl isothiocyanate) (final concentration in the test solution: 100 μM) and abscisic acid (final concentration in the test solution: 20 μM) were used as test compounds. A test solution containing fusicoccin (final concentration: 10 μM) was also prepared. In the measurement test, light (150 μmol m -2 s -1 Red light and 50 μmol m -2 s -1The test was conducted in the same manner as in Test Example 1, except that in addition to a group incubated under blue light irradiation for 3 hours (light irradiation group), a group incubated in a dark room for 3 hours instead of light irradiation (dark treatment group) was also established. The mean and standard deviation (SD) were calculated based on the measured values.

[0134] The results are shown in Figure 1. It was found that Compound 1 exhibited a more potent inhibitory effect on stomatal opening than abscisic acid, and also exhibited an inhibitory effect on fusicoccin-induced stomatal opening.

[0135] Test Example 3. Analysis of the mechanism of stomatal opening regulation 1 Blue light- and fusicoccin-induced phosphorylation of plasma membrane proton pumps in guard cells derived from the epidermis of Arabidopsis thaliana was measured using the same method as previously reported (Hayashi et al. (2011) Immunohistochemical detection of blue light-induced phosphorylation of the plasma membrane H + β-ATPase in stomatal guard cells. Plant Cell Physiol. 52: 1238-1248.) The test compounds used were Compound 1 (BITC), Compound 4 (m-Bis-BITC), and Compound 5 (Tris-BITC). The concentrations of Compound 1, Compound 4, Compound 5, and Fusicoccin were 100 μM, 20 μM, and 20 μM, respectively.

[0136] The results are shown in Figure 2. Compounds 1, 4, and 5 were found to have inhibitory effects on blue light-induced and fusicoccin-induced phosphorylation of the plasma membrane proton pump.

[0137] Test Example 4. Analysis of the mechanism of stomatal opening regulation 2. Abscisic acid (ABA)-induced phosphorylation of ABA-responsive kinase substrate (AKS) in guard cells derived from the epidermis of broad bean (Vicia faba) was measured by immunoblot analysis as previously reported (Takahashi et al. (2007) Protein Phosphorylation and Binding of a 14-3-3 Protein in Vicia Guard Cells in Response to ABA. Plant and Cell Physiology. 48: 1182-1191.). Compound 1 (BITC) and abscisic acid were used as test compounds. The concentration of compound 1 during application was 100 μM, and the concentration of abscisic acid during application was 20 μM.

[0138] The results are shown in Figure 3. Compound 1 did not induce phosphorylation of ABA-responsive kinase substrates.

[0139] Further analysis revealed that compound 1 did not affect ABA-related responses in Arabidopsis (inhibition of seed germination, induction of ABA-responsive genes (RAB18 and RD29B)).

[0140] Test Example 5. Measurement of stomatal opening regulation activity Test 3 A bouquet of chrysanthemum (Crysanthemum) flowers was purchased and exposed to white light (100 μmol m ) at 24°C for 16 hours. -2 s -1 The leaves were cultured in a water bottle under a photoperiod of 8 h / 8 h darkness for 2 days. After the dark period ended, the leaves were immersed in a test solution (0.033% Makupika (spreader, Ishihara Biosciences)) containing the test compound (compound 1 (BITC), compound 4 (m-Bis-BITC), or abscisic acid (ABA)) and exposed to white light (100 μmol m -2 s -1 After incubation, the stomatal aperture was measured in the same manner as in Test Example 1.

[0141] The results are shown in Figures 4 and 5. Compounds 1 and 4 were found to have an inhibitory effect on stomatal opening even in intact leaves. Compound 4 was also found to have an excellent durability of the inhibitory effect on stomatal opening.

[0142] Test Example 6. Desiccation Tolerance Test 1 After 3 hours of incubation as in Test Example 5, the bouquet was dehydrated and 10 μmol m -2 s -1 The plates were incubated at 21°C for 90 minutes under white light and 35-40% relative humidity.

[0143] Photographs of the appearance at the start and end of incubation under dehydration conditions are shown in Figure 6. It was found that compounds 1 and 4 could improve drought tolerance and suppress wilting.

[0144] Test Example 7. Drought Tolerance Test 2 A test solution containing compound 4 (m-Bis-BITC) (0.033% Makupika (spreader, Ishihara Biosciences)) was sprayed onto 4-5 week-old Chinese cabbage (Brassica rapa) growing in 110 ml of soil. The concentration of compound 4 in the test solution was 50 μM. The water was then drained, and the plants were cultured for 24 hours in a greenhouse at 25°C and 40-50% relative humidity during a sunny day.

[0145] Photographs of the appearance at the start and end of cultivation under water-deprived conditions are shown in Figure 7. It was found that Compound 4 improved drought tolerance and was able to suppress wilting for a long period of time.

Claims

1. General formula (1): 【Chemistry 1】 [In the formula, A represents a monocyclic 5- or 6-membered ring which may be substituted; L may be the same or different and represents an alkylene group which may be substituted with an alkyl group and / or an aryl group; and n represents an integer of 1 to 6.] The compound contains at least one selected from the group consisting of a compound represented by the formula: When A is a benzene ring and n is 1, A is a benzene ring substituted with at least one selected from the group consisting of an optionally substituted aryl group, a cyano group, an iodine atom, and an optionally substituted alkoxycarbonyl group, and / or L is an alkylene group substituted with an alkyl group and / or an aryl group. , a plant stomatal opening regulator.

2. (delete)

3. The plant stomatal opening regulator according to claim 1, wherein A is an unsubstituted ring or a ring substituted with at least one selected from the group consisting of an optionally substituted alkyl group, an optionally substituted alkoxy group, an optionally substituted alkoxycarbonyl group, an optionally substituted aryl group, a cyano group, a halogen atom, a nitro group, an optionally substituted amino group, an optionally substituted amide group, an optionally substituted ketone group, an optionally substituted ammonio group, and an optionally substituted thio group.

4. The plant stomatal opening regulator according to claim 1 or 3, wherein A has 0 to 3 substituents.

5. (delete)

6. (delete)

7. The plant stomatal opening regulator according to claim 1 , 3 or 4 , which is a plant stomatal opening inhibitor.

8. A drought tolerance improving agent comprising the plant stomatal opening regulator according to claim 1 , 3 , 4 or 7 .

9. The drought resistance improver according to claim 8 for use in suppressing wilting.

10. A method for improving drought tolerance, comprising applying the plant stomatal opening regulator according to claim 1 to a plant.

11. 11. A method for improving drought tolerance according to claim 10, comprising contacting the plant stomatal opening regulator according to claim 1, 3, 4 or 7 with the stomata of a plant.

12. General formula (1): 【Chemistry 2】 [In the formula, A represents an optionally substituted ring, L represents a linker, and n represents an integer of 1 to 6.] A cell membrane proton pump phosphorylation inhibitor comprising at least one member selected from the group consisting of a compound represented by the formula:

13. General formula (1): 【Chemistry 3】 [In the formula, A represents an optionally substituted ring, L represents a linker, and n represents an integer of 1 to 6.] A plant covering or supporting material comprising at least one member selected from the group consisting of a compound represented by the formula (I):

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