Reagent for detecting methyl salicylate, methyl salicylate sensor, sensing method of methyl salicylate using reagent and sensor, and detection method of pathogen infection in plant

Abstract

To provide a reagent for detecting methyl salicylate that is a plant hormone released when pathogen infection has arisen in plant cultivation including farm crop, and further, a methyl salicylate sensor, and a sensing method of methyl salicylate using the reagent and the sensor, and to provide a method for detecting pathogen infection in plants using the sensing method.SOLUTION: A reagent for detecting methyl salicylate contains a terbium compound doped with a rare earth element other than terbium. A methyl salicylate sensor comprises: a capturing part of methyl salicylate including the reagent; and a detection part for detecting that methyl salicylate has been captured by the capturing part.SELECTED DRAWING: None

Description

Technical Field

[0001] The present invention relates to a reagent for detecting methyl salicylate, which is a plant hormone released when a plant is infected with a pathogenic bacterium, a methyl salicylate sensor, a sensing method for methyl salicylate using them, and a method for detecting pathogenic bacterium infection of a plant.

Background Art

[0002] When a plant is infected with a pathogenic bacterium, it is known that the plant synthesizes and releases a plant hormone as a signal substance, and by informing the surrounding plants of the pathogenic bacterium infection, the defense mechanism is promoted in advance. By quickly recognizing such a signal substance released by the plant, it becomes possible to detect damage by pests and diseases.

[0003] As a method for discovering pest damage to a target plant, for example, Patent Document 1 discloses a method in which a monitor plant having a luminescent protein gene is placed near the target plant, and the monitor plant senses a volatile substance released by the target plant in response to stress and emits light.

Prior Art Documents

Patent Documents

[0004]

Patent Document 1

Summary of the Invention

Problems to be Solved by the Invention

[0005] An object of the present invention is to provide a reagent for detecting methyl salicylate, which is a plant hormone released when a plant including a crop is infected with a pathogenic bacterium, a methyl salicylate sensor, a sensing method for methyl salicylate using them, and thereby provide a method for detecting pathogenic bacterium infection of a plant.

Means for Solving the Problems

[0006] The inventors of the present invention have conducted intensive research to solve the above problems. As a result, by using a reagent containing a terbium compound doped with a rare earth element other than terbium (hereinafter also referred to as "other rare earth elements") as a receptor for methyl salicylate, compared with the case of using a reagent containing an undoped terbium compound, the fluorescence emission derived from the complex formed by the reaction of the terbium compound and methyl salicylate increases, and it has been found that plant pathogen infection can be detected, thus completing the present invention.

[0007] One aspect of the present embodiment relates to a reagent for detecting methyl salicylate, which contains a terbium compound doped with a rare earth element other than terbium.

[0008] Also, one aspect of the present embodiment relates to a methyl salicylate sensor for detecting methyl salicylate, which includes a capturing part of methyl salicylate having a reagent containing a terbium compound doped with a rare earth element other than terbium, and a detecting part for detecting that methyl salicylate has been captured by the capturing part.

[0009] Furthermore, one aspect of the present embodiment relates to a sensing method for methyl salicylate using the above reagent to detect methyl salicylate, which includes: (i) a step of reacting a terbium compound doped with a rare earth element other than terbium with methyl salicylate to form a complex; (ii) a step of irradiating the complex with excitation light; (iii) a step of detecting the fluorescence emitted by the complex and relates to a sensing method for methyl salicylate.

[0010] Furthermore, one aspect of the present embodiment relates to a method for detecting plant pathogen infection by installing the above reagent near a plant and confirming the fluorescence emission derived from the complex formed by the reaction of a terbium compound doped with a rare earth element other than terbium and methyl salicylate.

Advantages of the Invention

[0011] According to the present invention, in a reagent for detecting methyl salicylate, by containing a terbium compound doped with a rare earth element other than terbium (other rare earth elements) as a receptor for methyl salicylate, compared with the case of containing an undoped terbium compound, the fluorescence emission intensity from the complex formed by the reaction between methyl salicylate, which is a volatile plant hormone released when a plant is infected with a pathogenic bacterium, and the terbium compound increases, and it becomes possible to detect the infection of the plant by the pathogenic bacterium.

Brief Description of Drawings

[0012]

Figure 1

Figure 2

Figure 3

Figure 4

Figure 5

Figure 6

Modes for Carrying Out the Invention

[0013] Hereinafter, modes for carrying out the present invention will be described with reference to the drawings and the like. However, although the embodiments described below have technically preferable limitations for carrying out the present invention, they do not limit the scope of the invention below.

[0014] [1] Reagent for Detecting Methyl Salicylate One embodiment of the present invention is a reagent for detecting methyl salicylate, which contains a terbium compound doped with a rare earth element other than terbium as a receptor for methyl salicylate. In the present invention, the term "reagent" is defined as a chemical substance used for detecting or quantifying a substance by a chemical method, conducting an experiment on the synthesis of a substance, or measuring physical properties.

[0015] In addition, the reagent preferably contains a non-volatile ionic liquid that effectively captures methyl salicylate in addition to the terbium compound doped with other rare earths. When the reagent contains an ionic liquid, the terbium compound doped with other rare earths exists dissolved in the non-volatile ionic liquid, but may also partially precipitate. Whether the terbium compound doped with other rare earths is dissolved or precipitated in the non-volatile ionic liquid, it can similarly function as a receptor for methyl salicylate.

[0016] <Receptor for methyl salicylate: Terbium compound doped with other rare earths> As the terbium compound doped with other rare earths that can be used to recognize methyl salicylate in the present invention, a compound represented by the following general formula (1) is used. Ln c TbX a Y b (1) In the formula, Ln represents at least one rare earth element other than terbium, X and Y each independently represent at least one selected from the group consisting of acetate ion, propionate ion, butyrate ion, pivalate ion, 2-methylbutyrate ion, trifluoroacetate ion, benzoate ion, chloride ion, and nitrate ion, c represents 0.015 to 0.05, and a and b each independently represent a number satisfying 0 ≦ a ≦ 3.15, 0 ≦ b ≦ 3.15, and a + b = 3 + 3 × c.

[0017] Even when doping with two or more rare earth elements other than terbium, the total value of c is 0.015 to 0.05. When c is within this range, the effect of increasing the fluorescence emission intensity derived from the complex formed by the reaction with methyl salicylate is fully exerted.

[0018] Specific examples of other rare earth elements include scandium, yttrium, lanthanum, cerium, praseodymium, neodymium, promethium, samarium, europium, gadolinium, dysprosium, holmium, erbium, thulium, ytterbium, and lutetium.

[0019] In addition, specific examples of terbium compounds doped with other rare earth elements include Ce 0.02 Tb(CH 3 COO) 0.81 (t-C 4 H 9 COO) 2.25 、Eu 0.02 Tb(CH 3 COO) 0.81 (t-C 4 H 9 COO) 2.25 、Sm 0.02 Tb(CH 3 COO) 0.81 (t-C 4 H 9 COO) 2.25 、Dy 0.02 Tb(CH 3 COO) 0.81 (t-C 4 H 9 COO) 2.25 、Gd 0.02 Tb(CH 3 COO) 0.81 (t-C 4 H 9 COO) 2.25 、Yb 0.02 Tb(CH 3 COO) 0.81 (t-C 4 H 9 COO) 2.25 、Ce 0.01 Tb(CH 3 COO) 0.78 (t-C4 H 9 COO) 2.25 、Ce 0.015 Tb(CH 3 COO) 0.795 (t-C 4 H 9 COO) 2.25 、Ce 0.025 Tb(CH 3 COO) 0.825 (t-C 4 H 9 COO) 2.25 、Ce 0.04 Tb(CH 3 COO) 0.87 (t-C 4 H 9 COO) 2.25 、Ce 0.05 Tb(CH 3 COO) 0.9 (t-C 4 H 9 COO) 2.25 、Ce 0.06 Tb(CH 3 COO) 0.93 (t-C 4 H 9 COO) 2.25 、Ce 0.02 Tb(CH 3 COO) 3.06、 Ce 0.04 Tb(CH 3 COO) 3.12 、Yb 0.15 Tb(CH 3 COO) 0.795 (t-C 4 H 9 COO) 2.25 、Yb 0.025 Tb(CH 3 COO) 0.825 (t-C 4 H 9 COO) 2.25 、Yb 0.04 Tb(CH 3 COO) 0.87 (t-C 4 H 9 COO) 2.25 、Ce 0.02 Tb(NO 3 ) 3.06 、Yb 0.02Tb(NO 3 ) 3.06 etc. can be mentioned, but it is not limited to only these.

[0020] Terbium compounds doped with other rare earth elements, for example, terbium acetate-pivalate complex salt doped with cerium (Ce c Tb(CH 3 COO) a (t-C 4 H 9 COO) b ) can be synthesized by dissolving terbium acetate, terbium pivalate, and an arbitrary number of moles of cerium acetate with respect to terbium (Tb) in methanol and subjecting them to a heating reaction. Also, by changing the ratio of the two terbium salts (terbium acetate and terbium pivalate) and the molar ratio of cerium acetate during synthesis, the ratio (a, b) of carboxylic acid ions (X, Y) and the doping amount (c) of cerium can be changed.

[0021] And the terbium compound doped with other rare earth elements can selectively recognize methyl salicylate by reacting with methyl salicylate to form a complex. The terbium compound doped with other rare earth elements has an increased fluorescence emission intensity derived from the complex formed by the reaction with methyl salicylate compared to the terbium compound not doped with other rare earth elements, which facilitates the sensing of methyl salicylate and enables the detection of infection by plant pathogens.

[0022] And the generated complex emits fluorescence specific to the complex when excited by UV. Only the terbium compound doped with other rare earth elements does not show observable fluorescence emission intensity even when irradiated with UV light. Also, since the terbium compound does not react with or recognize other plant hormones such as methyl jasmonate other than methyl salicylate, it can selectively recognize methyl salicylate.

[0023] <Non-volatile ionic liquid> In order to effectively capture methyl salicylate, the reagent of the present invention can contain a non-volatile ionic liquid in addition to a terbium compound doped with other rare earth elements. Examples of the non-volatile ionic liquids that can be used in the present invention include imidazolium salts, phosphonium salts, pyridinium salts, ammonium salts, piperidinium salts, pyrrolidinium salts, and the like. Specifically, 1-ethyl-3-methylimidazolium acetate, 1-butyl-3-methylimidazolium acetate, 1-ethyl-3-methylimidazolium tetrafluoroborate, 1-butyl-1-methylpyrrolidinium bis(fluorosulfonyl)imide, 1-butyl-3-methylimidazolium dibutyl phosphate, tetrabutylammonium acetate, 1-butylpyridinium bis(trifluoromethanesulfonyl)imide, 1-methyl-1-propylpiperidinium bis(fluorosulfonyl)imide, 1-ethyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide, 1-butyl-3-methylimidazolium dicyanamide, etc. can be mentioned, but it is not limited thereto.

[0024] The ratio of the ionic liquid to the terbium compound doped with other rare earth elements is preferably 2 to 40 times, more preferably 3 to 30 times, and particularly preferably 4 to 20 times by weight ratio. If the weight ratio of the ionic liquid to the terbium compound doped with other rare earth elements is less than 2 times, the effect of improving the detection sensitivity of methyl salicylate may not be obtained. If the weight ratio of the ionic liquid to the terbium compound doped with other rare earth elements exceeds 40 times, the terbium compound doped with other rare earth elements may be less likely to form a complex with methyl salicylate.

[0025] The reagent for detecting methyl salicylate of the present invention may optionally contain other solvents within a range that does not impair the effects of the present invention. As the solvent, dimethyl sulfoxide, methanol, ethanol, water, N,N-dimethylformamide, tetrahydrofuran, acetone, acetonitrile, 1,4-dioxane, etc. can be used, but it is not limited thereto.

[0026] [2] Methyl Salicylate Sensor One embodiment of the present invention relates to a methyl salicylate sensor for detecting methyl salicylate, comprising a capturing part for methyl salicylate having a reagent containing a terbium compound doped with a rare earth element other than terbium, and a detecting part for detecting that methyl salicylate has been captured by the capturing part.

[0027] (1) Capturing part The capturing part of the methyl salicylate sensor of the present invention has a reagent containing a terbium compound doped with another rare earth element, which is a receptor that selectively captures methyl salicylate. In the capturing part, it is preferable that the reagent is contained in a medium.

[0028] <Medium> Examples of the medium for containing the terbium compound doped with another rare earth element of the present invention include paper or glass fiber, resins (e.g., polymethyl methacrylate, polyethylene, polypropylene, polyvinyl chloride, polystyrene, nylon resin, polyamide, polycarbonate, polyethylene terephthalate, polybutylene terephthalate, polyphenylene oxide), water-soluble polymers (cellulose-based, agarose, starch-based, sodium alginate, acrylic acid-based, acrylamide-based, polyvinyl alcohol, polyethylene oxide, polyvinyl pyrrolidone, etc.), but are not limited thereto.

[0029] For example, when using paper as the medium, the terbium compound doped with another rare earth element is dissolved in a solvent, and the obtained solution is impregnated into paper (e.g., filter paper), and then dried at room temperature to 60 ° C to remove the solvent, thereby obtaining a medium containing the terbium compound doped with another rare earth element. After drying, the solvent evaporates and is removed, but may also remain. Using a solvent is preferable because it facilitates impregnating the medium with the terbium compound doped with another rare earth element and also facilitates adjusting the concentration of the terbium compound doped with another rare earth element.

[0030] As the solvent for dissolving terbium compounds doped with other rare earth elements, dimethyl sulfoxide, methanol, ethanol, water, N,N-dimethylformamide, tetrahydrofuran, acetone, acetonitrile, 1,4-dioxane, etc. can be used, but are not limited thereto.

[0031] In order to effectively capture methyl salicylate, it is preferable to add the above-mentioned non-volatile ionic liquid to the solvent. When using a non-volatile ionic liquid, after removing the solvent, the terbium compound doped with other rare earth elements exists dissolved in the non-volatile ionic liquid, but may also partially precipitate. Whether the terbium compound doped with other rare earth elements is dissolved or precipitated in the non-volatile ionic liquid, it can similarly function as a receptor for methyl salicylate.

[0032] In addition, the ratio of the ionic liquid to the solvent can be appropriately set with respect to the medium to be impregnated. If the ratio of the ionic liquid to the solvent is low, the amount of the ionic liquid in the dried medium may decrease, and the effect of improving the detection sensitivity may be reduced. On the other hand, if the ratio of the ionic liquid to the solvent increases, there may be a demerit that it becomes difficult to impregnate the medium because the ionic liquid has a high viscosity. Therefore, the ratio of the ionic liquid to the solvent is appropriately set with respect to the medium. For example, when impregnating filter paper, the ratio of the ionic liquid to the solvent is preferably 5 to 50% by weight, and more preferably 10 to 30% by weight.

[0033] (2) Detection unit The detection unit of the methyl salicylate sensor of the present invention is configured to optically detect that methyl salicylate has been captured by the capture unit. The detection unit may be configured as a separate device rather than a device integrated with the capture unit. In one aspect of the present invention, in the optical detection unit, in order to detect the fluorescence emission of the complex formed by a terbium compound doped with another rare earth element and methyl salicylate, it includes an excitation light source (light emitting unit) and a detection element (fluorescence receiving unit), and based on the change in the observed fluorescence intensity, the detection and / or concentration measurement of methyl salicylate can be performed.

[0034] In one aspect of the present invention, the detection unit may include a computer that executes a program for processing the detection and / or concentration measurement of methyl salicylate. Such a program may be a program that causes the computer to perform (i) the step of receiving a signal from an optical detection element, (ii) the step of analyzing the received signal to determine the presence and / or concentration of methyl salicylate, and (iii) the step of outputting the analysis result.

[0035] In one aspect of the present invention, the analysis of the received signal may include, for example, determining the presence and / or concentration of methyl salicylate by comparing the received signal with a predetermined reference value. Also, in one aspect of the present invention, the analysis result may be output to, for example, a display device connected to the sensor or other devices connected via a network.

[0036] In one aspect of the present invention, the methyl salicylate sensor of the present invention senses methyl salicylate, a plant hormone released when a crop is infected with a pathogenic bacterium. Therefore, the methyl salicylate sensor of the present invention can be used as a sensor for detecting pathogenic bacterium infection in plants including crops.

[0037] [3] Sensing method of methyl salicylate One embodiment of the present invention relates to a method for sensing methyl salicylate by using the fluorescence emission phenomenon from a complex obtained by reacting a terbium compound doped with another rare earth element with methyl salicylate using the reagent or the methyl salicylate sensor, (i) a step of reacting a terbium compound doped with another rare earth element with methyl salicylate to form a complex, (ii) a step of irradiating the complex with excitation light, (iii) a method for sensing methyl salicylate, comprising a step of detecting the fluorescence emitted by the complex.

[0038] While a terbium compound doped with another rare earth element alone shows almost no fluorescence emission, a complex formed by the reaction of a terbium compound doped with another rare earth element with methyl salicylate newly shows fluorescence emission. By utilizing this phenomenon, it becomes possible to detect methyl salicylate.

[0039] In one aspect of the present invention, an appropriate wavelength within the range of 300 to 400 nm is selected as the excitation wavelength. Further, in one aspect of the present invention, a step of determining the concentration of methyl salicylate by comparing the intensity of the detected fluorescence with a predetermined reference value may also be performed.

[0040] [4] Method for detecting plant pathogen infection One embodiment of the present invention relates to a method for detecting plant pathogen infection by installing the reagent or the methyl salicylate sensor near a plant and confirming the fluorescence emission derived from a complex formed in association with the reaction of a terbium compound doped with another rare earth element with methyl salicylate.

[0041] Examples of plants that can be monitored include, but are not limited to, cucumber, watermelon, tomato, eggplant, pepper, paprika, shishito, melon, Chinese cabbage, cabbage, radish, lettuce, green onion, broccoli, onion, garlic, yam, asparagus, carrot, potato, celery, tobacco, rice, and strawberry.

[0042] Examples of detectable diseases include, for example, ring rot, white spot, brown ring spot, leaf mold, blight, root rot blight, half blight, brown root rot, gray blight, root rot, black dot root rot, southern blight, damping-off, brown spot, powdery mildew, rust, gray mold, anthracnose, scab, sclerotinia, vine wilt, leaf spot, blight, mosaic, yellow dwarf, yellow leaf curl, bacterial wilt, soft rot, seaweed disease, stem dwarf bacteria disease, black spot bacteria disease, spot bacteria disease, etc., but are not limited thereto. Further, examples of detectable pathogenic bacteria infections include infections caused by the causative bacteria of the above diseases, but are not limited thereto.

[0043] In the context of the present disclosure, when it is said to "install near a plant", examples of the term "nearby" include, for example, a distance within 2 m, within 1 m, within 75 cm, within 50 cm, within 40 cm, within 30 cm, within 20 cm, within 10 cm, or within 5 cm from the plant to be monitored, but are not limited thereto, and an appropriate distance is appropriately selected in consideration of various factors. A person skilled in the art would be able to appropriately set the position where the sensor is installed considering various conditions.

[0044] Furthermore, one embodiment of the present invention relates to the use of a reagent or a methyl salicylate sensor in the detection of pathogenic bacteria infection of plants. Also, one embodiment of the present invention relates to the use of a terbium compound doped with other rare earth elements in the production of a reagent or a methyl salicylate sensor.

Examples

[0045] Hereinafter, the present invention will be described more specifically with reference to examples, but the present invention is not limited to these examples.

[0046] (Synthesis Example 1) Terbium acetate doped with 2% cerium (Ce) (Tb(CH 3 COO) 3 :TbA) and terbium pivalate (Tb(t-C 4 H 9 COO) 3 :TbPv) composite salt [Ce0.02 Tb(CH 3 COO) 0.81 (C 4 H 9 COO) 2.25 :0.02CeA-0.25TbA-0.75TbPv] Dissolve 0.2 g of terbium acetate tetrahydrate, 0.6797 g of terbium pivalate, and 0.0131 g of cerium acetate monohydrate in 120 ml of methanol, and heat under reflux for 5 hours. After cooling, distill off the methanol, and vacuum-dry the precipitated white crystals to obtain 0.733 g of the target terbium compound doped with 2% cerium.

[0047] (Synthesis Example 2) Composite salt of terbium acetate (TbA) doped with 2% europium (Eu) and terbium pivalate (TbPv) [Eu 0.02 Tb(CH 3 COO) 0.81 (C 4 H 9 COO) 2.25 :0.02EuA-0.25TbA-0.75TbPv] Synthesized in the same manner as in Synthesis Example 1, except that 0.01573 g of europium acetate tetrahydrate was used instead of 0.0131 g of cerium acetate monohydrate.

[0048] (Synthesis Example 3) Composite salt of terbium acetate (TbA) doped with 2% gadolinium (Gd) and terbium pivalate (TbPv) [Gd 0.02 Tb(CH 3 COO) 0.81 (C 4 H 9 COO) 2.25 :0.02GdA-0.25TbA-0.75TbPv] Synthesized in the same manner as in Synthesis Example 1, except that 0.01593 g of gadolinium acetate tetrahydrate was used instead of 0.0131 g of cerium acetate monohydrate.

[0049] (Synthesis Example 4) Complex salt of terbium acetate (TbA) doped with 2% dysprosium (Dy) and terbium pivalate (TbPv) [Dy 0.02 Tb(CH 3 COO) 0.81 (C 4 H 9 COO) 2.25 :0.02DyA - 0.25TbA - 0.75TbPv] Synthesized in the same manner as in Synthesis Example 1, except that 0.01614 g of dysprosium acetate (DyA) tetrahydrate was used instead of 0.0131 g of cerium acetate monohydrate.

[0050] (Synthesis Example 5) Complex salt of terbium acetate (TbA) doped with 2% ytterbium (Yb) and terbium pivalate (TbPv) [Dy 0.02 Tb(CH 3 COO) 0.81 (C 4 H 9 COO) 2.25 :0.02DyA - 0.25TbA - 0.75TbPv] Synthesized in the same manner as in Synthesis Example 1, except that 0.01655 g of ytterbium acetate (YbA) tetrahydrate was used instead of 0.0131 g of cerium acetate monohydrate.

[0051] (Synthesis Example 6) Complex salt of terbium acetate (TbA) doped with 1% cerium (Ce) and terbium pivalate (TbPv) [Ce 0.01 Tb(CH 3 COO) 0.78 (C 4 H 9 COO) 2.25 :0.01CeA - 0.25TbA - 0.75TbPv] Synthesized in the same manner as in Synthesis Example 1, except that 0.0066 g of cerium acetate monohydrate was used.

[0052] (Synthesis Example 7) Complex salt of terbium acetate (TbA) doped with 1.5% cerium (Ce) and terbium pivalate (TbPv) [Ce 0.015 Tb(CH 3 COO)0.795 (C 4 H 9 COO) 2.25 :0.015CeA-0.25TbA-0.75TbPv] In the same manner as in Synthesis Example 1, except that 0.0098 g of cerium acetate monohydrate was used for the synthesis.

[0053] (Synthesis Example 8) Composite salt of terbium acetate (TbA) and terbium pivalate (TbPv) doped with 2.5% cerium (Ce) [Ce 0.025 Tb(CH 3 COO) 0.825 (C 4 H 9 COO) 2.25 :0.025CeA-0.25TbA-0.75TbPv] In the same manner as in Synthesis Example 1, except that 0.0164 g of cerium acetate monohydrate was used for the synthesis.

[0054] (Synthesis Example 9) Composite salt of terbium acetate (TbA) and terbium pivalate (TbPv) doped with 4% cerium (Ce) [Ce 0.04 Tb(CH 3 COO) 0.87 (C 4 H 9 COO) 2.25 :0.04CeA-0.25TbA-0.75TbPv] In the same manner as in Synthesis Example 1, except that 0.0262 g of cerium acetate monohydrate was used for the synthesis.

[0055] (Synthesis Example 10) Composite salt of terbium acetate (TbA) and terbium pivalate (TbPv) doped with 5% cerium (Ce) [Ce 0.05 Tb(CH 3 COO) 0.9 (C 4 H 9 COO) 2.25 :0.05CeA-0.25TbA-0.75TbPv] In the same manner as in Synthesis Example 1, except that 0.0328 g of cerium acetate monohydrate was used for the synthesis.

[0056] (Synthesis Example 11) Complex salt of terbium acetate (TbA) doped with 6% cerium (Ce) and terbium pivalate (TbPv) [Ce 0.06 Tb(CH 3 COO) 0.93 (C 4 H 9 COO) 2.25 :0.06CeA-0.25TbA-0.75TbPv] Synthesized in the same manner as in Synthesis Example 1, except that 0.0393 g of cerium acetate monohydrate was used.

[0057] (Synthesis Example 12) Complex salt of terbium acetate (TbA) doped with 2.5% ytterbium (Yb) and terbium pivalate (TbPv) [Yb 0.025 Tb(CH 3 COO) 0.825 (C 4 H 9 COO) 2.25 :0.025YbA-0.25TbA-0.75TbPv] Synthesized in the same manner as in Synthesis Example 5, except that 0.02069 g of ytterbium acetate (YbA) tetrahydrate was used.

[0058] (Synthesis Example 13) Complex salt of terbium acetate (TbA) doped with 4% ytterbium (Yb) and terbium pivalate (TbPv) [Yb 0.04 Tb(CH 3 COO) 0.87 (C 4 H 9 COO) 2.25 :0.04YbA-0.25TbA-0.75TbPv] Synthesized in the same manner as in Synthesis Example 5, except that 0.0331 g of ytterbium acetate (YbA) tetrahydrate was used.

[0059] (Synthesis Example 14) Terbium acetate (TbA) doped with 2% cerium (Ce) [Ce 0.02 Tb(CH 3 COO) 3.06: 0.02 CeA-TbA Dissolve 1 g of terbium acetate tetrahydrate and 0.0164 g of cerium acetate monohydrate in 100 ml of methanol, and heat under reflux for 5 hours. After allowing to cool, the precipitated white crystals are dried under vacuum to obtain 0.411 g of terbium acetate doped with 2% cerium.

[0060] (Synthesis Example 15) Terbium acetate (TbA) doped with 4% cerium (Ce) [Ce 0.04 Tb(CH 3 COO) 3.12 : 0.04 CeA-TbA Synthesized in the same manner as in Synthesis Example 14, except that 0.0328 g of cerium acetate monohydrate was used.

[0061] (Synthesis Example 16) Terbium nitrate (TbN) doped with 2% cerium (Ce) [Ce 0.02 Tb(NO 3 ) 3.06 : 0.02 CeA-TbN Dissolve 1 g of terbium nitrate hexahydrate and 0.0192 g of cerium nitrate hexahydrate in 50 ml of methanol, and heat under reflux for 5 hours. After allowing to cool, the methanol is distilled off, and further dried under vacuum at 60 °C to quantitatively obtain terbium nitrate doped with 2% cerium.

[0062] (Synthesis Example 17) Terbium nitrate (TbN) doped with 2% ytterbium (Yb) [Yb 0.02 Tb(NO 3 ) 3.06 : 0.02 YbA-TbN Dissolve 1 g of terbium nitrate hexahydrate and 0.0198 g of ytterbium nitrate pentahydrate in 50 ml of methanol, and heat under reflux for 5 hours. After allowing to cool, the methanol is distilled off, and further dried under vacuum at 60 °C to quantitatively obtain terbium nitrate doped with 2% ytterbium.

[0063] (Comparative Synthesis Example 1) Composite salt [Tb(CH 3 COO) 0.75 (C 4 H 9 COO) 2.25 :0.25TbA - 0.75TbPv] of terbium acetate (TbA) and terbium pivalate (TbPv) without doping with other rare earth elements 0.2 g of terbium acetate tetrahydrate and 0.6797 g of terbium pivalate (the molar ratio of TbA to TbPv is 1:3) are heated under reflux in 65 ml of methanol for 5 hours. After cooling, the methanol is distilled off, and the precipitated white crystals are dried in vacuo to obtain 0.74 g of the target composite salt of terbium acetate and terbium pivalate.

[0064] (Example 1 and Comparative Example 1) [Fluorescence emission behavior associated with the reaction with methyl salicylate (MSA)] 0.9 ml of a DMSO solution (concentration 1.73 mM) of a composite salt (0.02CeA - 0.25TbA - 0.75TbPv) of terbium acetate (TbA) and terbium pivalate (TbPv) doped with 2% cerium obtained in Synthesis Example 1 and 0.1 ml of a DMSO solution (concentration 1.5 mM) of methyl salicylate (MSA) are mixed. After 10 minutes, the mixture is diluted 20-fold, and 3 ml of the solution is placed in a quartz cell, and the fluorescence spectrum is measured at an excitation wavelength of 365 nm (Example 1). Also, 0.9 ml of a DMSO solution (concentration 1.73 mM) of 0.25TbA - 0.75TbPv obtained in Comparative Synthesis Example 1 and 0.1 ml of DMSO are mixed, further diluted 20-fold, and 3 ml of the solution is placed in a quartz cell, and the fluorescence spectrum is measured at an excitation wavelength of 365 nm (Comparative Example 1). The obtained fluorescence spectrum curves are shown in Fig. 1. The solid line represents the fluorescence spectrum of 0.02CeA - 0.25TbA - 0.75TbPv + MSA (Example 1), and the dashed line represents the fluorescence spectrum of 0.25TbA - 0.75TbPv (Comparative Example 1). In the obtained fluorescence spectrum, it was found that the fluorescence intensity at a wavelength of 546 nm increased by 23% by doping with cerium.

[0065] (Examples 2 - 5) Similar to Example 1, except that the terbium compounds obtained in Synthesis Examples 2 to 5 were used instead of the terbium compound obtained in Synthesis Example 1, and the fluorescence spectrum was measured. In the obtained fluorescence spectrum, the fluorescence intensity at a wavelength of 546 nm was summarized in Table 1. From these results, it was found that the fluorescence intensity increased by doping the terbium compound with other rare earth elements.

[0066]

Table 1

[0067] (Examples 6 to 9 and Comparative Examples 2 and 3) Similar to Example 1, except that the terbium compounds obtained in Synthesis Examples 6 to 11 were used instead of the terbium compound obtained in Synthesis Example 1, and the fluorescence spectrum was measured. In the obtained fluorescence spectrum, the fluorescence intensity at a wavelength of 546 nm was summarized in Table 2. In Examples 1, 6 to 9, the fluorescence intensity increased compared to Comparative Example 1 in which no other rare earth element (cerium) was doped. However, in Comparative Example 2 where the doping amount of cerium was too small and Comparative Example 3 where it was too large, the fluorescence intensity decreased compared to Comparative Example 1 where no cerium was doped. From these results, it was found that the doping amount of other rare earth elements of 1.5 to 5% (c is 0.015 to 0.05 in formula (1)) is effective.

[0068]

Table 2

[0069] (Example 10) 0.0437 g of terbium acetate-terbium pivalate composite salt doped with 2% cerium (0.02CeA-0.25TbA-0.75TbPv) obtained in Synthesis Example 1 was dissolved in 1.6 ml of dimethyl sulfoxide (DMSO). To this, 0.4 ml of 1-ethyl-1-methylimidazolium acetate (EMImAc) was added (DMSO / EMImAc = 8 / 2). 0.2 ml of this solution was dropped onto a circular filter paper (Φ40 mm) and dried to volatilize DMSO, obtaining a filter paper containing 0.02CeA-0.25TbA-0.75TbPv and EMImAc. The obtained filter paper was placed in a desiccator with a volume of 800 ml. Using a permeator PD-1B-2 (manufactured by Gastec Corporation), nitrogen was used as the carrier gas, and methyl salicylate with a concentration of 5 ppb was introduced. After exposure for 1 hour, the filter paper was taken out, and the fluorescence spectrum at an excitation wavelength of 365 nm was measured. In the obtained fluorescence spectrum shown in Figure 2, the fluorescence intensity at a wavelength of 544 nm was about 11 times the fluorescence intensity at a wavelength of 544 nm in the unexposed fluorescence spectrum, indicating that methyl salicylate in the gas phase with a concentration of 5 ppb can be sensed.

[0070] (Example 11) In the same manner as in Example 10, except that the terbium acetate-terbium pivalate composite salt doped with 2% gadolinium (0.02GdA-0.25TbA-0.75TbPv) obtained in Synthesis Example 3 was used instead of the terbium acetate-terbium pivalate composite salt doped with 2% cerium (0.02CeA-0.25TbA-0.75TbPv) obtained in Synthesis Example 1 for evaluation. In the obtained fluorescence spectrum shown in Figure 3, the fluorescence intensity at a wavelength of 544 nm was about 3 times the fluorescence intensity at a wavelength of 544 nm in the unexposed fluorescence spectrum, indicating that methyl salicylate in the gas phase with a concentration of 5 ppb can be sensed.

[0071] (Example 12) 0.0438 g of terbium acetate-terbium pivalate composite salt (0.02YbA-0.25TbA-0.75TbPv) doped with 2% ytterbium obtained in Synthesis Example 5 was dissolved in 1.6 ml of dimethyl sulfoxide (DMSO), and 0.4 ml of 1-ethyl-1-methylimidazolium acetate (EMImAc) was added thereto (DMSO / EMImAc = 8 / 2). 0.2 ml of the solution was dropped onto a circular filter paper (Φ40 mm) and dried to volatilize DMSO, obtaining a filter paper containing 0.02YbA-0.25TbA-0.75TbPv and EMImAc. The obtained filter paper was placed in a desiccator with a volume of 800 ml, and therein, using a permeator PD-1B-2, nitrogen was used as a carrier gas, and methyl salicylate with a concentration of 2 ppb was introduced. After exposure for 1 hour, the filter paper was taken out, and the fluorescence spectrum at an excitation wavelength of 365 nm was measured. In the obtained fluorescence spectrum shown in Fig. 4, the fluorescence intensity at a wavelength of 544 nm is about twice the fluorescence intensity at a wavelength of 544 nm in the unexposed fluorescence spectrum, revealing that methyl salicylate in the gas phase with a concentration of 2 ppb can be sensed.

[0072] (Example 13 and Comparative Example 4) In the same manner as in Example 1, except that the terbium compound obtained in Synthesis Example 14 was used instead of the terbium compound obtained in Synthesis Example 1, the fluorescence spectrum was measured. Also, as Comparative Example 4, the fluorescence spectrum was measured in the same manner using terbium acetate tetrahydrate. The obtained fluorescence spectrum curves are shown in Fig. 5. The solid line represents the fluorescence spectrum of the terbium compound of Synthesis Example 14 + MSA (Example 13), and the dashed line represents the fluorescence spectrum of terbium acetate + MSA (Comparative Example 4). In the obtained fluorescence spectrum, it was found that the fluorescence intensity at a wavelength of 546 nm increased by 72% by doping with cerium.

[0073] (Example 14) In the same manner as in Example 13, except that the terbium compound obtained in Synthesis Example 15 was used instead of the terbium compound obtained in Synthesis Example 14, the fluorescence spectrum was measured. In the obtained fluorescence spectrum, it was found that the fluorescence intensity at a wavelength of 546 nm increased by 88% by doping with 4% cerium.

[0074] (Examples 15 and 16 and Comparative Example 5) Similar to Example 1, except that the terbium compound obtained in Synthesis Example 16 was used instead of the terbium compound obtained in Synthesis Example 1, and the fluorescence spectrum was measured. Further, the fluorescence spectrum of the terbium compound obtained in Synthesis Example 17 was also measured in the same manner. Also, as Comparative Example 5, the fluorescence spectrum was measured in the same manner using terbium nitrate hexahydrate. The obtained fluorescence spectrum curves are shown in Fig. 6. The solid line represents the fluorescence spectrum of the terbium compound of Synthesis Example 16 + MSA (Example 15), the dashed line represents the fluorescence spectrum of the terbium compound of Synthesis Example 17 + MSA (Example 16), and the dotted-dashed line represents the fluorescence spectrum of terbium nitrate + MSA (Comparative Example 5). In the obtained fluorescence spectrum, it was found that the fluorescence intensity at a wavelength of 545 nm increased 2.9-fold by doping with 2% cerium and 3.4-fold by doping with 2% ytterbium.

Industrial Applicability

[0075] The methyl salicylate sensor for detecting methyl salicylate, which is a plant hormone according to the present invention, is provided with a capture part having a medium containing a terbium compound doped with other rare earth elements, so that methyl salicylate can be efficiently captured, a complex is formed, and fluorescence emission is exhibited. Therefore, it is possible to selectively detect methyl salicylate, which is a plant hormone released by plants when infected with pathogenic bacteria. By using the methyl salicylate sensor, it is possible to detect the infection of pathogenic bacteria in plants, and specifically, it can be used as a new sensor for agricultural ICT in protected horticulture such as greenhouses as a sensor for detecting the infection of pathogenic bacteria in agricultural crops.

[0076] As described above, the present invention has been described with reference to the embodiments and examples, but the present invention is not limited to the above embodiments and examples. Various changes that can be understood by those skilled in the art can be made to the configuration and details of the present invention within the scope of the present invention.

[0077] (Supplementary Note 1) A reagent for detecting methyl salicylate, comprising a terbium compound doped with a rare earth element other than terbium. (Appendix 2) The reagent according to Appendix 1, wherein the terbium compound doped with a rare earth element other than terbium is represented by the following general formula (1). Ln c TbX a Y b (1) (In the formula, Ln represents at least one rare earth element other than terbium, X and Y each independently represent at least one selected from the group consisting of acetate ion, propionate ion, butyrate ion, pivalate ion, 2-methylbutyrate ion, trifluoroacetate ion, benzoate ion, chloride ion, and nitrate ion, c represents 0.015 to 0.05, and a and b each independently represent a number satisfying 0 ≦ a ≦ 3.15, 0 ≦ b ≦ 3.15, and a + b = 3 + 3 × c) (Appendix 3) The reagent according to Appendix 1 or 2, wherein the rare earth element other than terbium is at least one selected from the group consisting of scandium, yttrium, lanthanum, cerium, praseodymium, neodymium, promethium, samarium, europium, gadolinium, dysprosium, holmium, erbium, thulium, ytterbium, and lutetium. (Appendix 4) The reagent according to any one of the preceding appendices, further comprising a non-volatile ionic liquid. (Appendix 5) The reagent according to Appendix 4, wherein the non-volatile ionic liquid is at least one selected from the group consisting of 1-ethyl-3-methylimidazolium acetate, 1-butyl-3-methylimidazolium acetate, 1-ethyl-3-methylimidazolium tetrafluoroborate, 1-butyl-1-methylpyrrolidinium bis(fluorosulfonyl)imide, 1-butylpyridinium bis(trifluoromethanesulfonyl)imide, 1-methyl-1-propylpiperidinium bis(fluorosulfonyl)imide, 1-ethyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide, and 1-butyl-3-methylimidazolium methyl dicyanamide. (Appendix 6) A methyl salicylate sensor for detecting methyl salicylate, comprising a capturing part of methyl salicylate having a reagent containing a terbium compound doped with a rare earth element other than terbium, and a detecting part for detecting that methyl salicylate has been captured by the capturing part. A methyl salicylate sensor comprising the above. (Appendix 7) The methyl salicylate sensor according to Appendix 6, wherein the capturing part includes a medium containing the reagent. (Appendix 8) The methyl salicylate sensor according to Appendix 7, wherein the medium is paper, glass fiber, resin, or a water-soluble polymer. (Appendix 9) A sensing method for methyl salicylate for recognizing methyl salicylate using the reagent according to any one of Appendices 1 to 5, (i) a step of reacting a terbium compound doped with a rare earth element other than terbium with methyl salicylate to form a complex, (ii) a step of irradiating the complex with excitation light, (iii) a step of detecting the fluorescence emitted by the complex A sensing method for methyl salicylate including the above. (Appendix 10) A method for detecting a pathogen infection of a plant by installing the reagent according to any one of Appendices 1 to 5 near the plant and confirming the fluorescence emission derived from the complex formed by the reaction between the terbium compound doped with a rare earth element other than terbium and methyl salicylate.

Claims

1. A reagent for detecting methyl salicylate, comprising a terbium compound doped with a rare earth element other than terbium.

2. The reagent according to claim 1, wherein the terbium compound doped with a rare earth element other than terbium is represented by the following general formula (1). Ln ｃ TbX ａ Y ｂ (1) (In the formula, Ln represents at least one rare earth element other than terbium, X and Y each independently represent at least one selected from the group consisting of acetate ion, propionate ion, butyrate ion, pivalate ion, 2-methylbutyrate ion, trifluoroacetate ion, benzoate ion, chloride ion, and nitrate ion, c represents 0.015 to 0.05, and a and b each independently represent a number satisfying 0 ≦ a ≦ 3.15, 0 ≦ b ≦ 3.15, and a + b = 3 + 3×c)

3. The reagent according to claim 1 or 2, wherein the rare earth element other than terbium is at least one selected from the group consisting of scandium, yttrium, lanthanum, cerium, praseodymium, neodymium, promethium, samarium, europium, gadolinium, dysprosium, holmium, erbium, thulium, ytterbium, and lutetium.

4. The reagent according to claim 1 or 2, further comprising a non-volatile ionic liquid.

5. The reagent according to claim 4, wherein the non-volatile ionic liquid is at least one selected from the group consisting of 1-ethyl-3-methylimidazolium acetate, 1-butyl-3-methylimidazolium acetate, 1-ethyl-3-methylimidazolium tetrafluoroborate, 1-butyl-1-methylpyrrolidinium bis(fluorosulfonyl)imide, 1-butylpyridinium bis(trifluoromethanesulfonyl)imide, 1-methyl-1-propylpiperidinium bis(fluorosulfonyl)imide, 1-ethyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide, and 1-butyl-3-methylimidazolium methyl dicyanamide.

6. A methyl salicylate sensor for detecting methyl salicylate, comprising: A capture part of methyl salicylate having a reagent containing a terbium compound doped with a rare earth element other than terbium; A detection part for detecting that methyl salicylate has been captured by the capture part. The methyl salicylate sensor comprising the above.

7. The methyl salicylate sensor according to claim 6, wherein the capturing unit includes a medium containing the reagent.

8. The methyl salicylate sensor according to claim 7, wherein the medium is paper, glass fiber, resin, or a water-soluble polymer.

9. A sensing method for methyl salicylate that recognizes methyl salicylate using the reagent according to claim 1 or 2, comprising: (i) reacting a terbium compound doped with a rare earth element other than terbium with methyl salicylate to form a complex; (ii) irradiating the complex with excitation light; (iii) detecting fluorescence emitted by the complex The sensing method for methyl salicylate includes.

10. A method for detecting a pathogen infection of a plant by installing the reagent according to claim 1 or 2 near the plant and confirming fluorescence emission derived from a complex formed by the reaction of a terbium compound doped with a rare earth element other than terbium and methyl salicylate.

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