Application of pyrazine compound in preparation of plant parasitic nematode attractant

Abstract

The invention discloses application of a pyrazine compound in preparation of a plant parasitic nematode attractant, and belongs to the technical field of plant parasitic nematode prevention and control. The structural formula of the pyrazine compound is shown in the figure 1, it is found for the first time that the pyrazine compound has high attraction activity on meloidogyne incognita and bursaphelenchus xylophilus, and the pyrazine compound can rapidly take effect at the low concentration; test data show that the chemotactic indexes of the pyrazine compound to meloidogyne incognita and pine wood nematodes are not lower than 20%, the chemotactic indexes of the pyrazine compound to the pine wood nematodes can reach 40% or above, and even the chemotactic indexes of the meloidogyne incognita can reach 80% or above; meanwhile, the pyrazine compound is matched with conventional nematicidal drugs, so that the chemotactic index of the nematodes can be increased, the nematicidal drugs are assisted to improve the nematicidal effect, and a new scheme is provided for preventing and treating plant parasitic nematodes.

Description

Technical Field

[0001] This invention belongs to the field of plant parasitic nematode control technology, specifically relating to the application of a pyrazine compound in the preparation of plant parasitic nematode attractants. Background Technology

[0002] Plant parasitic nematodes pose a significant threat to global agricultural production, especially the southern root-knot nematode (Simple Root-Knot Nematode). Meloidogyne incognita ), pine wood nematode ( Bursaphelenchus xylophilus Southern root-knot nematodes infect plant roots, forming root knots that hinder water and nutrient absorption, leading to stunted growth and sharp yield reductions. Pine wilt nematodes cause pine wilt disease, resulting in significant losses of forest resources. Statistics show that global agricultural economic losses due to root-knot nematodes exceed US$100 billion annually, seriously threatening food security and ecological balance. Currently, plant parasitic nematode control mainly relies on chemical nematicides, such as carbamates and organophosphates. However, long-term use of chemical pesticides leads to soil pollution, pesticide residues, and increased nematode resistance, while also harming non-target organisms and human health. In recent years, the use of plant-derived or microbial compounds to regulate nematode behavior has become a research hotspot. For example, volatile organic compounds (VOCs) secreted by plant roots can attract or repel nematodes, providing a new approach for green nematode control. Therefore, developing highly efficient, low-toxicity, and environmentally friendly plant parasitic nematode attractants to induce rapid aggregation of plant parasitic nematodes is highly beneficial for concentrated extermination or synergistic trapping with other active ingredients. Summary of the Invention

[0003] The first objective of this invention is to provide an application of pyrazine compounds in the preparation of plant parasitic nematode attractants, and the second objective is to provide a plant parasitic nematode attractant.

[0004] The first objective of this invention is achieved through the application of the pyrazine compounds in the preparation of attractants for plant parasitic nematodes, wherein the chemical structural formula of the pyrazine compounds is as follows: ; R1, R2, R3, and R4 are derived from methyl, methoxy, ethyl, ethoxy, isobutyl, isopropyl, sec-butyl, amino, amide, ester, acetyl, or carboxyl groups; the plant parasitic nematode is the southern root-knot nematode or the pine wood nematode, and the concentration of the pyrazine compound in the attractant is 0.1 ppm to 10 ppm.

[0005] The aforementioned pyrazine compounds can be 2-ethyl-3-methoxypyrazine, 2-ethoxy-3-methylpyrazine, 2-isobutyl-3-methoxypyrazine, 3-hydroxypyrazine-2-amide, methyl 3-aminopyrazine-2-carboxylate, 2-amino-6-methoxypyrazine, 2-amino-3-methoxypyrazine, 3-methylpyrazine-2-carboxylic acid, 5-methoxypyrazine-2-carboxylic acid, 2-amino-3-ethoxypyrazine, (S)-2,5-dihydro-3,6-dimethoxy-isobutylpyrazine, 2-isobutylpyrazine, or 6-methoxy-2-pyrazine carboxylic acid, with structural formulas as shown in Formulas 1-1, 1-2, 1-3, 1-13, 1-17, 1-18, 1-24, 1-25, 1-26, 1-29, 1-30, 1-31, and 1-32, respectively. .

[0006] The aforementioned pyrazine compounds may be 2-methoxy-3,5-dimethoxypyrazine, 2-methoxy-3-sec-butylpyrazine, 2-isobutyl-3-methylpyrazine, 3-isopropyl-4-methoxyaniline, 2-methoxy-3-methylpyrazine, 3-methoxy-2,5-dimethylpyrazine, 2-methoxy-6-methylpyrazine, 2-amino-3-methoxy-5-methylpyrazine, methyl 2-pyrazine carboxylate, and ethyl 5-methoxypyrazine-2-carboxylate. 5-Methoxypyrazine-2-carboxylic acid, 2-pyrazine carboxylic acid, 2-acetylpyrazine, methyl 3-methylpyrazine-2-carboxylic acid, 2-hydroxy-3-methoxypyrazine, 6-methoxypyrazine-2-carboxamide, 3-methoxypyrazine-2-carboxylic acid, methyl 5-methoxypyrazine-2-carboxylic acid, pyrazinamide, 2-methoxy-3-isopropylpyrazine, 2-methoxypyrazine or (R)-2,5-dihydro-1,6-dimethoxy-2-isopropylpyrazine.

[0007] The second objective of this invention is achieved by the fact that the plant parasitic nematode attractant has the above-mentioned pyrazine compounds as the main active ingredient or one of the active ingredients.

[0008] This invention is the first to discover that the above-mentioned pyrazine compounds have strong attraction activity against southern root-knot nematodes and pine wood nematodes, and also against the non-plant pathogenic nematode *C. elegans* (…). Caenorhabditis elegansThe presence of no attractant activity indicates that the above-mentioned compounds have significant specificity in attracting plant parasitic nematodes. Experiments showed that when the effective concentration of the pyrazine compounds in the attractant was 0.1 ppm to 10 ppm, the chemotactic index for *Hemiberlesia sapiens* and *Pinus thunbergii* was no less than 20%, reaching over 40% for *Pinus thunbergii* and even over 80% for *Hemiberlesia sapiens*. Furthermore, the pyrazine compounds, when combined with conventional nematicides, can enhance the chemotactic index of nematodes, thereby improving the nematicidal effect of the nematicides. This provides new lead compounds for the development of pesticides to control plant parasitic nematodes, especially *Hemiberlesia sapiens* and *Pinus thunbergii*. Attached Figure Description

[0009] Figure 1 The chemical structural formula of the pyrazine compounds described in this invention is shown below. Figure 2 The preferred chemical structural formula of the pyrazine compounds described in this invention is shown below. Figure 3 The nematode chemotaxis test was conducted using a water agar plate with a diameter of 6 cm. The center point was the nematode implantation site, and the left-side points were the drug solution implantation sites for the blank control group. Figure 3 The middle part is a 20% ethanol solution (or other solutions), and the right-hand dot is the implantation point for the test compound (i.e., the experimental group drug solution or the blank control drug solution). Figure 4 The chemotactic index of 2-ethyl-3-methoxypyrazine on southern root-knot nematodes in Experiment Example 1; Figure 5 The chemotactic index of 2-ethoxy-3-methylpyrazine on southern root-knot nematodes in Experiment Example 1; Figure 6 The chemotactic index of 2-isobutyl-3-methoxypyrazine on southern root-knot nematodes in Experiment Example 1; Figure 7 The chemotactic index of 3-hydroxypyrazine-2-amide on southern root-knot nematodes in Experiment Example 1; Figure 8 The chemotactic index of methyl 3-aminopyrazine-2-carboxylate on southern root-knot nematodes in Experiment Example 1; Figure 9 The chemotactic index of 2-amino-6-methoxypyrazine on southern root-knot nematodes in Experiment Example 1; Figure 10 The chemotactic index of 2-amino-3-methoxypyrazine on southern root-knot nematodes in Experiment Example 1; Figure 11 The chemotactic index of 3-methylpyrazine-2-carboxylic acid on southern root-knot nematodes in Experiment Example 1; Figure 12 The chemotactic index of 5-methoxypyrazine-2-carboxylic acid on southern root-knot nematodes in Experiment Example 1; Figure 13 The chemotactic index of 2-amino-3-ethoxypyrazine on southern root-knot nematodes in Experiment Example 1; Figure 14 The chemotactic index of (S)-2,5-dihydro-3,6-dimethoxy-isobutylpyrazine on southern root-knot nematodes in Experiment Example 1; Figure 15 The chemotactic index of 2-isobutylpyrazine on southern root-knot nematodes in Experiment Example 1; Figure 16 The chemotactic index of 6-methoxy-2-pyrazincarboxylic acid on southern root-knot nematodes in Experiment Example 1; Figure 17 The chemotactic index of 2-ethyl-3-methoxypyrazine on pine wood nematode in Experiment Example 2; Figure 18 The chemotactic index of 2-ethoxy-3-methylpyrazine on pine wood nematode in Experiment Example 2; Figure 19 The chemotactic index of 2-isobutyl-3-methoxypyrazine on pine wood nematode in Experiment Example 2; Figure 20 The chemotactic index of 2-ethoxy-3-methylpyrazine in combination with abamectin (i.e., the complex) and abamectin (i.e., the compound) on southern root-knot nematodes in Experiment Example 3; Figure 21 The mortality rate of southern root-knot nematodes was determined by the combination of 2-ethoxy-3-methylpyrazine and abamectin (i.e., the complex) and abamectin (i.e., the compound) in Experiment Example 3. Detailed Implementation

[0010] The present invention will be further described in detail below with reference to experimental examples and embodiments, but this does not limit the present invention in any way. Any modifications or improvements made based on the present invention shall fall within the protection scope of the present invention.

[0011] The application of the pyrazine compounds of this invention in the preparation of plant parasitic nematode attractants, wherein the chemical structural formula of the pyrazine compounds is as follows: ; Among them, R1, R2, R3, and R4 are derived from methyl, methoxy, ethyl, ethoxy, isobutyl, isopropyl, sec-butyl, amino, amide, ester, acetyl, or carboxyl groups. The plant parasitic nematode is the southern root-knot nematode or the pine wood nematode, and the concentration of the pyrazine compound in the attractant is 0.1 ppm to 10 ppm.

[0012] Further, the pyrazine compounds are 2-ethyl-3-methoxypyrazine, 2-ethoxy-3-methylpyrazine, 2-isobutyl-3-methoxypyrazine, 3-hydroxypyrazine-2-amide, methyl 3-aminopyrazine-2-carboxylate, 2-amino-6-methoxypyrazine, 2-amino-3-methoxypyrazine, 3-methylpyrazine-2-carboxylic acid, 5-methoxypyrazine-2-carboxylic acid, 2-amino-3-ethoxypyrazine, (S)-2,5-dihydro-3,6-dimethoxy-isobutylpyrazine, 2-isobutylpyrazine, or 6-methoxy-2-pyrazine carboxylic acid, with structural formulas as shown in Formulas 1-1, 1-2, 1-3, 1-13, 1-17, 1-18, 1-24, 1-25, 1-26, 1-29, 1-30, 1-31, and 1-32, respectively. .

[0013] Further, the pyrazine compounds are 2-methoxy-3,5-dimethoxypyrazine (1-4), 2-methoxy-3-sec-butylpyrazine (1-5), 2-isobutyl-3-methylpyrazine (1-6), 3-isopropyl-4-methoxyaniline (1-7), 2-methoxy-3-methylpyrazine (1-8), 3-methoxy-2,5-dimethylpyrazine (1-9), 2-methoxy-6-methylpyrazine (1-10), 2-amino-3-methoxy-5-methylpyrazine (1-11), methyl 2-pyrazinecarboxylate (1-12), ethyl 5-methoxypyrazine-2-carboxylate (1-14), 5-methoxypyrazine-2-carboxylic acid (...). 1-15), 2-pyrazinic acid (1-16), 2-acetylpyrazine (1-19), methyl 3-methylpyrazine-2-carboxylate (1-20), 2-hydroxy-3-methoxypyrazine (1-21), 6-methoxypyrazine-2-carboxamide (1-22), 3-methoxypyrazine-2-carboxylic acid (1-23), methyl 5-methoxypyrazine-2-carboxylate (1-27), pyrazinamide (1-28), 2-methoxy-3-isopropylpyrazine (1-33), 2-methoxypyrazine (1-34) or (R)-2,5-dihydro-1,6-dimethoxy-2-isopropylpyrazine (1-35); Note: The numbers in parentheses above are... Figure 2 The corresponding serial numbers of the chemical structural formulas of the above compounds.

[0014] The plant parasitic nematode attractant of the present invention uses the above-mentioned pyrazine compounds as the main active ingredient or one of the active ingredients.

[0015] Furthermore, the effective concentration of the pyrazine compound in the attractant is 0.1 ppm to 10 ppm.

[0016] Furthermore, the attractant is in aqueous form.

[0017] Furthermore, the solvent in the aqueous solution is an ethanol aqueous solution with a concentration of ≤20% or pure water.

[0018] All pyrazine compounds described in this invention were purchased from MedChem Express, and their attraction activity against southern root-knot nematodes and pine wood nematodes was verified through relevant experiments. Due to space limitations, only the following compounds are shown: 2-ethyl-3-methoxypyrazine (1-1), 2-ethoxy-3-methylpyrazine (1-2), 2-isobutyl-3-methoxypyrazine (1-3), 3-hydroxypyrazine-2-amide (1-13), methyl 3-aminopyrazine-2-carboxylate (1-17), and 2-amino-6-methoxypyrazine (1-18). The test contents, using 2-amino-3-methoxypyrazine (1-24), 3-methylpyrazine-2-carboxylic acid (1-25), 5-methoxypyrazine-2-carboxylic acid (1-26), 2-amino-3-ethoxypyrazine (1-29), (S)-2,5-dihydro-3,6-dimethoxy-isobutylpyrazine (1-30), 2-isobutylpyrazine (1-31), and 6-methoxy-2-pyrazine carboxylic acid (1-32) as test materials, involve a total of 13 pyrazine compounds: 2-Ethyl-3-methoxypyrazine has CAS number 25680-58-4, molecular weight 138.17, and molecular formula C7H. 10 N2O; 2-Ethoxy-3-methylpyrazine has CAS number 32737-14-7, molecular weight 138.17, and molecular formula C7H. 10 N2O; 2-Isobutyl-3-methoxypyrazine has CAS number 24683-00-9, molecular weight 166.22, and molecular formula C9H. 14 N2O; 3-Hydroxypyrazine-2-amide has CAS number 55321-99-8, molecular weight 139.112, and molecular formula C5H5N3O2; 3-Aminopyrazine-2-carboxylic acid methyl ester has the CAS number 16298-03-6, molecular weight 153.139, and molecular formula C6H7N3O2; 2-Amino-6-methoxypyrazine has CAS number 6905-47-1, molecular weight 125.129, and molecular formula C5H7N3O; 2-Amino-3-methoxypyrazine has CAS number 4774-10-1, molecular weight 125.129, and molecular formula C5H7N3O; 3-Methylpyrazine-2-carboxylic acid has the CAS number 41110-28-5, molecular weight 138.124, and molecular formula C6H6N2O2. 5-Methoxypyrazine-2-carboxylic acid has the CAS number 40155-42-8, a molecular weight of 154.123, and the molecular formula C6H6N2O3. 2-Amino-3-ethoxypyrazine has CAS number 89464-86-8, molecular weight 139.15500, and molecular formula C6H9N3O; (S)-2,5-dihydro-3,6-dimethoxy-isopropylpyrazine has CAS number 78342-42-4, molecular weight 184.236, and molecular formula C9H. 16 N2O2; 2-Isobutylpyrazine has CAS number 29460-92-2, molecular weight 136.194, and molecular formula C8H. 12 N2; 6-Methoxy-2-pyrazinic acid has the CAS number 24005-61-6, a molecular weight of 154.123, and the molecular formula C6H6N2O3.

[0019] Experimental Example 1: Attractant Activity Test of Pyrazine Compounds on Southern Root-Knot Nematodes 1. Preparation of test solution 1-1. Preparation of experimental drug solutions: Take 2-ethyl-3-methoxypyrazine (1-1), 2-ethoxy-3-methylpyrazine (1-2), 2-isobutyl-3-methoxypyrazine (1-3), 3-hydroxypyrazine-2-amide (1-13), methyl 3-aminopyrazine-2-carboxylate (1-17), 2-amino-6-methoxypyrazine (1-18), 2-amino-3-methoxypyrazine (1-24), 3-methylpyrazine-2-carboxylic acid (1-25), and 5-methoxypyrazine-2-carboxylic acid (1-26), respectively. 1 mg each of 2-amino-3-ethoxypyrazine (1-29), (S)-2,5-dihydro-3,6-dimethoxy-isobutylpyrazine (1-30), 2-isobutylpyrazine (1-31), and 6-methoxy-2-pyrazine carboxylic acid (1-32) were dissolved in 1 mL of 20% (w / w) aqueous ethanol solution to obtain working solutions of the above compounds; the working solutions of each compound were diluted with 20% (w / w) aqueous ethanol solution to prepare test group solutions with concentrations of 10 ppm, 1 ppm, and 0.1 ppm, respectively. 1-2. Blank control group solution: 20% ethanol aqueous solution.

[0020] 2. Sample preparation for the Southern Root-knot Nematode experiment According to known techniques, the southern root-knot nematode infects plant roots at the second instar larval (J2) stage. Therefore, second instar larvae of the southern root-knot nematode were selected as the experimental sample. Tomato roots infected with the southern root-knot nematode were washed with water, and then egg sacs were collected using a dissecting needle and placed in 0.4M KCl solution. The egg sacs were then washed sequentially in 5mL centrifuge tubes with 4mL sterile water (three washes), 1mL sterile penicillin-streptomycin solution, and 3mL 0.1% NaClO solution (simultaneous sterilization). After each wash, the sacs were centrifuged at 4400rpm for 60s to remove the supernatant. The washed egg sacs were incubated in 0.4M ZnCl2 solution for 48h. Then, the second instar larvae were collected using a pipette into 5mL centrifuge tubes, and the viable second instar larvae (J2) of the southern root-knot nematode were enriched by centrifugation at 4400rpm for 60s for later use.

[0021] 3. Test methods 3-1. Prepare 6cm diameter water agar plates for each of the 13 pyrazine compounds, using three concentrations of each experimental group of the drug solution, as per the instructions. Figure 3 150-200 second-instar larvae of *Symplocos spp.* were added to the central implantation point of each culture plate, 5 μL of the blank control solution was added to the left implantation point, and 5 μL of the experimental group solution of one concentration of a pyrazine compound was added to the right implantation point. The central implantation point and the left and right implantation points were located on the diameter axis of the culture plate, and the distance between the central implantation point and the left and right implantation points was 2 cm. After the culture plate was set up, it was placed in an environment of 25℃. After 3 hours, the number of second-instar larvae of *Symplocos spp.* within a 1 cm diameter range (i.e., the blank control area and the compound area) extended outward from the left and right implantation points was counted.

[0022] 3-2. Repeat step 3-1 five times, and take the average of the five parallel treatments. Calculate the chemotactic index of the 13 pyrazine compounds at three concentrations for each region against *Strombus heterophylla*. The formula for calculating the chemotactic index is as follows: Chemotaxis index (%) = (Number of nematodes in the compound region - Number of nematodes in the blank control region) / Number of nematodes on the main axis × 100% The number of nematodes on the bus refers to the number of effective nematodes that have crawled more than 1 cm away from the central implantation point, not the total number of nematodes added.

[0023] 4. Results of the experiment after 3 hours Based on the nematode situation in the blank control area, it can be determined that the drug solution in the blank control group (i.e., 20% ethanol aqueous solution) has no attraction effect on southern root-knot nematodes.

[0024] refer to Figure 4 The chemotactic indices of 2-ethyl-3-methoxypyrazine were 49.4% (10 ppm), 58.7% (1 ppm), and 46.6% (0.1 ppm), respectively. refer to Figure 5The chemotactic indices of 2-ethoxy-3-methylpyrazine were 57.2% (10 ppm), 55.4% (1 ppm), and 59.5% (0.1 ppm), respectively. refer to Figure 6 The chemotactic indices of 2-isobutyl-3-methoxypyrazine were 77.6% (10 ppm), 60.3% (1 ppm), and 69.7% (0.1 ppm), respectively. refer to Figure 7 The chemotactic indices of 3-hydroxypyrazine-2-amide were 62.5% (10 ppm), 39.2% (1 ppm), and 62.9% (0.1 ppm), respectively. refer to Figure 8 The chemotactic indices of methyl 3-aminopyrazine-2-carboxylate were 44.2% (10 ppm), 49.3% (1 ppm), and 22.6% (0.1 ppm), respectively. refer to Figure 9 The chemotactic indices of 2-amino-6-methoxypyrazine were 21.2% (10 ppm), 21.7% (1 ppm), and 28.6% (0.1 ppm), respectively. refer to Figure 10 The chemotactic indices of 2-amino-3-methoxypyrazine were 46.3% (10 ppm), 31.1% (1 ppm), and 41.7% (0.1 ppm), respectively. refer to Figure 11 The chemotactic indices of 3-methylpyrazine-2-carboxylic acid were 40% (10 ppm), 51.9% (1 ppm), and 39.3% (0.1 ppm), respectively. refer to Figure 12 The chemotactic indices of 5-methoxypyrazine-2-carboxylic acid were 48.9% (10 ppm), 51.2% (1 ppm), and 35.5% (0.1 ppm), respectively. refer to Figure 13 The chemotactic indices of 2-amino-3-ethoxypyrazine were 49.1% (10 ppm), 61.8% (1 ppm), and 61.8% (0.1 ppm), respectively. refer to Figure 14 The chemotactic indices of (S)-2,5-dihydro-3,6-dimethoxy-isobutylpyrazine were 51.1% (10 ppm), 60.2% (1 ppm), and 48.8% (0.1 ppm), respectively. refer to Figure 15 The chemotactic indices of 2-isobutylpyrazine were 71% (10 ppm), 82.7% (1 ppm), and 66.7% (0.1 ppm), respectively. refer to Figure 16The chemotactic indices of 6-methoxy-2-pyrazinic acid were 20.9% (10 ppm), 50.9% (1 ppm), and 41.6% (0.1 ppm), respectively.

[0025] 5. Results Analysis Based on the above experimental data, it can be seen that when the concentration range of the pyrazine compounds is 0.1ppm to 10ppm, after 3 hours of application of the above 13 pyrazine compounds, the chemotaxis index of the southern root-knot nematode is not less than 20% (a chemotaxis index greater than 20% is considered effective induction), and most of the chemotaxis indices can reach more than 50%, or even more than 80%. This indicates that the pyrazine compounds of the present invention have a strong effect on attracting the southern root-knot nematode and can be used as agents for the control of the southern root-knot nematode.

[0026] Experimental Example 2: Attractant Activity Test of Pyrazine Compounds on Pine Wood Nematode 1. Preparation of test solutions: Prepare working solutions of 2-ethyl-3-methoxypyrazine, 2-ethoxy-3-methylpyrazine, and 2-isobutyl-3-methoxypyrazine according to step 1-1 of Experimental Example 1, and dilute them to concentrations of 10 ppm, 1 ppm, and 0.1 ppm respectively. The blank control solution is prepared according to step 1-2 of Experimental Example 1.

[0027] 2. Sample preparation for pine wood nematode experiment Cultured in Botrytis cinerea ( Botrytis cinerea Pine wood nematodes were collected in 1.5 mL centrifuge tubes using the Bellman method. The live pine wood nematode test samples were enriched by centrifugation at 4400 rpm for 30 s and then used for further processing.

[0028] 3. Test methods 3-1. Prepare 9cm diameter water agar plates for the experimental groups of three concentrations of each of the three pyrazine compounds according to step 1 of this experimental example, referring to... Figure 3 One hundred live pine wood nematodes were added to the central implantation point of each culture plate, 5 μL of blank control solution was added to the left implantation point, and 5 μL of experimental solution of one concentration of a pyrazine compound prepared in step 1 of this experiment was added to the right implantation point. The central implantation point and the left and right implantation points were located on the diameter axis of the culture plate, and the distance between the central implantation point and the left and right implantation points was 2 cm. After the culture plate was set up, it was placed in an environment of 25℃. After 1 hour, the number of pine wood nematodes in the 1 cm diameter range (i.e., the blank control area and the compound area) extended outward from the left and right implantation points was counted.

[0029] 3-2. Repeat step 3-1 five times and take the average value of the five parallel treatments. Calculate the chemotaxis index of the three pyrazine compounds for each region at the three concentrations. The formula for calculating the chemotaxis index is the same as that in Experiment 1.

[0030] 4. Test results after 1 hour Based on the nematode situation in the blank control area, it can be determined that the drug solution in the blank control group (i.e., 20% ethanol aqueous solution) has no attraction effect on pine wood nematodes.

[0031] refer to Figure 17 The chemotactic indices of 2-ethyl-3-methoxypyrazine were 25.6% (10 ppm), 25.2% (1 ppm), and 26.6% (0.1 ppm), respectively. refer to Figure 18 The chemotactic indices of 2-ethoxy-3-methylpyrazine were 28.1% (10 ppm), 33.2% (1 ppm), and 24.6% (0.1 ppm), respectively. refer to Figure 19 The chemotactic indices of 2-isobutyl-3-methoxypyrazine were 34.6% (10 ppm), 32.9% (1 ppm), and 46.3% (0.1 ppm), respectively.

[0032] 5. Results Analysis Based on the above experimental data, it can be seen that when the concentration range of the pyrazine compounds is 0.1ppm to 10ppm, the chemotactic index of 2-ethyl-3-methoxypyrazine, 2-ethoxy-3-methylpyrazine, and 2-isobutyl-3-methoxypyrazine can all exceed 20% after 1 hour of application to pine wood nematodes. This indicates that the pyrazine compounds of the present invention have a strong effect on attracting pine wood nematodes and can be used as agents for the control of pine wood nematodes.

[0033] Experiments 1 and 2 show that when the concentration of the pyrazine compound in the attractant is 0.1 ppm to 10 ppm, it can achieve the desired effect of attracting southern root-knot nematodes and pine wood nematodes.

[0034] Experimental Example 3: Nematode Control Effect Test of the Combination of Pyrazine Compounds with Commercially Available Insecticides The pyrazine compounds mentioned are 2-ethoxy-3-methylpyrazine as an example, commercially available insecticides include abamectin as an example, and nematodes include the southern root-knot nematode as an example.

[0035] 1. Preparation of test solution 1-1. Dissolve 2-ethoxy-3-methylpyrazine in pure DMSO of abamectin, and then prepare different test group solutions with abamectin concentrations of 10 ppm, 50 ppm and 100 ppm. The final concentration of 2-ethoxy-3-methylpyrazine in the test group solutions is 1 ppm. 1-2. Mix abamectin with pure DMSO, and then dilute with pure DMSO to prepare drug control solutions with abamectin concentrations of 10 ppm, 50 ppm, and 100 ppm. 1-3. The blank control group's drug solution was pure DMSO.

[0036] 2. Preparation of nematode test samples: Prepare second-instar larvae of Southern Root-knot Nematodes with good activity, referring to Experiment Example 1.

[0037] 3. Test methods: 3-1. Experimental Group: Prepare 6cm diameter water agar plates according to the abamectin concentration of the experimental group solution, referring to... Figure 3 One hundred second-instar larvae of Southern Root-knot Nematode were added to the central implantation point of each culture plate. 5 μL of the blank control group solution from steps 1-3 of this experiment was added to the left implantation point. 5 μL of the experimental group solution at an abamectin concentration was added to the right implantation point. Other procedures were the same as in Experiment 1.

[0038] 3-2. Drug Control Group: Prepare 6cm diameter water agar plates according to the avermectin concentration in the drug control group solution, and refer to... Figure 3 One hundred second-instar larvae of Southern Root-knot Nematode were added to the central implantation point of each culture plate. 5 μL of the blank control group solution from steps 1-3 of this experiment was added to the left implantation point. 5 μL of the experimental group solution at an abamectin concentration was added to the right implantation point. Other procedures were the same as in Experiment 1.

[0039] 3-3. After the culture plates in steps 3-1 and 3-2 are set up, they are placed in an environment of 25℃. After 12 hours, the number of Southern Root-knot Nematodes and the number of dead nematodes are counted within a 1cm diameter area extending outward from the implantation points on both sides (i.e., the blank control area and the compound / control drug area). Each group is repeated 5 times, and the average value of the 5 parallel treatments is taken. The chemotaxis index and mortality rate in each area are calculated respectively. The chemotaxis index is calculated according to Experiment Example 1, and the mortality rate is calculated using the following formula: Mortality rate (%) = Number of dead insects / Number of insects in the area × 100%.

[0040] 4. Test Results: Based on the nematode situation in the blank control area of ​​the experimental group and the drug control group, it can be determined that the drug solution (i.e., pure DMSO) in the blank control group has no attraction effect on southern root-knot nematodes.

[0041] Experimental group (see Figure 20 , 21 ): At 10 ppm of abamectin, the chemotaxis index of southern root-knot nematodes in the compound area was 50%, and the mortality rate was 60%. At 50 ppm of abamectin, the chemotaxis index of southern root-knot nematodes in the compound area was 62%, and the mortality rate was 62%. At 100 ppm of abamectin, the chemotaxis index of southern root-knot nematodes in the compound area was 78%, and the mortality rate was 63%. Drug control group (see Figure 20 , 21 ): When abamectin was applied at 10 ppm, the chemotactic index of southern root-knot nematodes in the control drug area was 14%, and the mortality rate was 23%. At 50 ppm of abamectin, the chemotactic index of southern root-knot nematodes in the control drug area was 17%, and the mortality rate was 45%. At 100 ppm of abamectin, the chemotactic index of southern root-knot nematodes in the control drug area was 13%, and the mortality rate was 34%.

[0042] 5. Results Analysis When 5-1,2-ethoxy-3-methylpyrazine is used in combination with abamectin, the chemotactic index and mortality rate of southern root-knot nematodes are significantly better than when abamectin is used alone. This indicates that the pyrazine compounds have a good attraction effect on southern root-knot nematodes, prompting the nematodes to enter the toxic range of the insecticide, thereby improving the killing effect. 5-2. When 2-ethoxy-3-methylpyrazine is used in combination with abamectin, the concentration of 2-ethoxy-3-methylpyrazine is 1 ppm. As the concentration of abamectin increases, the chemotactic index of southern root-knot nematodes increases accordingly, indicating that abamectin can synergistically enhance the attraction effect of 2-ethoxy-3-methylpyrazine on southern root-knot nematodes. Therefore, the pyrazine compounds described in this invention are suitable as attractants for plant root-knot nematodes and can enhance the killing effect of commercially available insecticides on plant parasitic nematodes.

[0043] Examples 1-3: Preparation of attractants for the control of plant parasitic nematodes Example 1 The solvent is pure water, and the active ingredients are 2-isobutyl-3-methoxypyrazine at concentrations of 10 ppm, 1 ppm, and 0.1 ppm, respectively.

[0044] Example 2 The solvent is an aqueous ethanol solution with a concentration ≤20%, and the active ingredients are 2-ethoxy-3-methylpyrazine with concentrations of 10 ppm, 1 ppm, and 0.1 ppm, respectively.

[0045] Example 3 The solvent is pure DMSO, and the active ingredient is 2-ethyl-3-methoxypyrazine at a concentration of 1 ppm.

[0046] Other compounds in the pyrazine class described in this application were prepared as attractants for controlling plant parasitic nematodes, referring to Examples 1-3.

Claims

1. The application of a pyrazine compound in the preparation of a plant parasitic nematode attractant, characterized in that, The chemical structural formulas of the pyrazine compounds are as follows: ； R1, R2, R3, and R4 are derived from methyl, methoxy, ethyl, ethoxy, isobutyl, isopropyl, sec-butyl, amino, amide, ester, acetyl, or carboxyl groups; the plant parasitic nematode is the southern root-knot nematode or the pine wood nematode, and the concentration of the pyrazine compound in the attractant is 0.1 ppm to 10 ppm.

2. The application according to claim 1, characterized in that, The pyrazine compounds are 2-ethyl-3-methoxypyrazine, 2-ethoxy-3-methylpyrazine, 2-isobutyl-3-methoxypyrazine, 3-hydroxypyrazine-2-amide, methyl 3-aminopyrazine-2-carboxylate, 2-amino-6-methoxypyrazine, 2-amino-3-methoxypyrazine, 3-methylpyrazine-2-carboxylic acid, 5-methoxypyrazine-2-carboxylic acid, 2-amino-3-ethoxypyrazine, (S)-2,5-dihydro-3,6-dimethoxy-isobutylpyrazine, 2-isobutylpyrazine, or 6-methoxy-2-pyrazine carboxylic acid, with structural formulas as shown in Formulas 1-1, 1-2, 1-3, 1-13, 1-17, 1-18, 1-24, 1-25, 1-26, 1-29, 1-30, 1-31, and 1-32, respectively. 。 3. The application according to claim 1, characterized in that, The pyrazine compounds are 2-methoxy-3,5-dimethoxypyrazine, 2-methoxy-3-sec-butylpyrazine, 2-isobutyl-3-methylpyrazine, 3-isopropyl-4-methoxyaniline, 2-methoxy-3-methylpyrazine, 3-methoxy-2,5-dimethylpyrazine, 2-methoxy-6-methylpyrazine, 2-amino-3-methoxy-5-methylpyrazine, methyl 2-pyrazinecarboxylate, ethyl 5-methoxypyrazine-2-carboxylate, 5 -Methoxypyrazine-2-carboxylic acid, 2-pyrazine carboxylic acid, 2-acetylpyrazine, methyl 3-methylpyrazine-2-carboxylic acid, 2-hydroxy-3-methoxypyrazine, 6-methoxypyrazine-2-carboxamide, 3-methoxypyrazine-2-carboxylic acid, methyl 5-methoxypyrazine-2-carboxylic acid, pyrazinamide, 2-methoxy-3-isopropylpyrazine, 2-methoxypyrazine or (R)-2,5-dihydro-1,6-dimethoxy-2-isopropylpyrazine.

4. A plant parasitic nematode attractant, characterized in that, The attractant uses the pyrazine compound described in claim 1, 2 or 3 as the main active ingredient or one of the active ingredients.

5. The plant parasitic nematode attractant according to claim 4, characterized in that, The effective concentration of the pyrazine compound in the attractant is 0.1 ppm to 10 ppm.

6. The plant parasitic nematode attractant according to claim 4, characterized in that, The attractant is in aqueous form.

7. The plant parasitic nematode attractant according to claim 6, characterized in that, The solvent in the aqueous solution is an ethanol aqueous solution with a concentration of ≤20% or pure water.

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