Application of a pyrazole compound in the control of plant root-knot nematodes

Abstract

This invention discloses the application of a pyrazole compound in the control of plant root-knot nematodes. This invention is the first to discover... N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H (-pyrazol-1-yl)-6-morpholinopyrimidine-4-amine exhibits strong toxic activity against various plant root-knot nematodes. Its 24-hour median lethal dose (LC50) is [not specified in the original text]. 50 The concentrations were 12.70 μg / ml, 24.35 μg / ml, and 20.57 μg / ml, respectively; the median lethal dose (LC50) at 48 hours was [not specified in the original text]. 50 The concentrations were 12.50 μg / ml, 23.46 μg / ml and 19.47 μg / ml, respectively, and the drug showed no toxic activity against non-plant pathogenic nematodes such as Caenorhabditis elegans. It is safe and efficient, providing new ideas and approaches for the development of new root-knot nematode insecticides.

Description

Technical Field

[0001] This invention belongs to the field of pest and disease control technology, specifically relating to the application of a pyrazole compound in the control of plant root-knot nematodes. Background Technology

[0002] Plant parasitic nematodes cause serious damage to the ecological environment and agricultural production, resulting in economic losses. Among them, root-knot nematodes, which are obligate parasites of plant roots (… Meloidogyne spp.) Root-knot nematodes are widespread and highly damaging parasitic nematodes. They infect crop roots, affecting nutrient and water absorption, hindering normal plant growth, and in severe cases, causing plant death, thus significantly reducing crop yield and quality. Furthermore, crops infected with root-knot nematodes can become transmission routes for other pathogens, causing even greater damage, making it a major challenge in global plant protection. In addition, the sensitivity of root-knot nematodes to chemical control agents varies across different regions, increasing the difficulty of control. Traditional commercial chemical pesticides are gradually being phased out due to their high toxicity and environmental pollution. Therefore, in-depth research into highly effective and low-toxicity compounds that kill root-knot nematodes, and using these as lead compounds for the creation of new pesticides, has become an important aspect of research on root-knot nematode disease control.

[0003] Pyrazole compounds are widely used in the development of agricultural insecticides due to their advantages such as safety, high efficiency, unique mechanism of action, and lack of cross-resistance. Pyrazole insecticides include fipronil (5-amino-3-acrylonitrile-1-(2,6-dichloro-4-trifluoromethylphenyl)-4-trifluoromethylsulfinylpyrazole), cyclopropamide, abamectin, and etoxazole. Although many pyrazole insecticides have been developed, no... N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H Publicly available literature reports on the nematicidal activity of pyrazol-1-yl)-6-morpholinopyrimidine-4-amine and its application in the preparation of nematicidal formulations. Summary of the Invention

[0004] The purpose of this invention is to provide pyrazole compounds. N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H Application of pyrazole-1-yl)-6-morpholinopyrimidine-4-amine in the control of plant root-knot nematodes. N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H The structural formula of (-pyrazol-1-yl)-6-morpholinopyrimidine-4-amine is as follows: Figure 1As shown. This compound was purchased from MCE, part number HY-109160, CAS number 2167246-24-2, with a relative molecular mass of 378.42 and mass-to-charge ratios (m / z) of 378.20 (100.0%), 379.20 (19.5%), and 379.20 (2.2%).

[0005] This invention is the first discovery N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H (-pyrazol-1-yl)-6-morpholinopyrimidine-4-amine exhibits strong toxic activity against various plant root-knot nematodes, but shows no toxic activity against non-plant pathogenic nematodes such as Caenorhabditis elegans. It is safe and highly effective, providing new ideas and approaches for the development of new root-knot nematode insecticides. Attached Figure Description

[0006] Figure 1 The pyrazole compound of this invention N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H The structural formula of (-pyrazol-1-yl)-6-morpholinopyrimidine-4-amine;

[0007] Figure 2 Example 1 N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H Calculation results of toxicity determination of pyrazol-1-yl)-6-morpholinopyrimidine-4-amine against southern root-knot worm;

[0008] Figure 3 Example 1 N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H Calculation results of toxicity determination of pyrazol-1-yl)-6-morpholinopyrimidine-4-amine against Javan root-knot nematode;

[0009] Figure 4 Example 1 N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H The toxicity test results of pyrazole-1-yl)-6-morpholinopyrimidine-4-amine on the root-knot nematode of the target bean. Detailed Implementation

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

[0011] This invention provides pyrazole compounds N -(4,4-difluorocyclohexyl)-2-(3-methyl-1 HApplication of pyrazol-1-yl)-6-morpholinopyrimidine-4-amine and / or its pesticide-acceptable salts in the control of plant root-knot nematodes.

[0012] The plant root-knot nematodes include, but are not limited to, *Southern root-knot nematodes*. M.incongnita Northern root-knot nematodes M.hapla Java root-knot nematodes M.javanica Peanut root-knot nematodes M.arenaria Or bean root nematode M.enterolobii .

[0013] This invention also provides a formulation for controlling plant root-knot nematodes, the formulation being... N -(4,4-difluorocyclohexyl)-2-(3-methyl-1 H -pyrazol-1-yl)-6-morpholinopyrimidine-4-amine and / or its pesticide-acceptable salts are the main active ingredient or one of the active ingredients.

[0014] The formulation N -(4,4-difluorocyclohexyl)-2-(3-methyl-1 H The concentration of pyrazol-1-yl)-6-morpholinopyrimidine-4-amine is 3-50 μg / ml.

[0015] The formulation is an aqueous solution, an emulsion, a wettable powder, a soluble powder, a soluble granule, a water-dispersible granule, a suspension, an oil suspension, a tablet, or an effervescent granule.

[0016] The solvent for the aqueous solution or water emulsion is a fatty alcohol aqueous solution or a dimethyl sulfoxide aqueous solution with a concentration ≤10%.

[0017] The fatty alcohol is methanol, ethanol, or propanol.

[0018] Example 1 N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H Detection of the root-knot nematode activity of pyrazol-1-yl)-6-morpholinopyrimidine-4-amine

[0019] 1. Sample preparation

[0020] Take 0.5-1.5mg N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H(-pyrazol-1-yl)-6-morpholinopyrimidine-4-amine was dissolved in DMSO and diluted with 4% DMSO solution to prepare the test solution. The concentrations of each solution were: 0.03 μg / ml, 0.05 μg / ml, 0.1 μg / ml, 0.3 μg / ml, 0.5 μg / ml, 1 μg / ml, 3 μg / ml, 5 μg / ml, 10 μg / ml, 30 μg / ml, and 50 μg / ml, with a DMSO concentration of 0.4% in each solution. Water containing 0.4% DMSO was used as a blank control.

[0021] 2. Preparation of experimental nematodes

[0022] Tomato roots infected with southern root-knot nematodes were washed with water. Oocysts of the nematodes were collected using a dissecting needle and placed in 0.4 M KCl solution. The cysts were then washed and sterilized sequentially in 5 ml centrifuge tubes with 4 ml sterile water (three times), 1 ml sterile penicillin-streptomycin solution, and 3 ml 0.1% NaClO solution. Each step involved brief centrifugation (8000 rpm for 30 seconds) to remove the supernatant. The washed eggs were incubated in 0.4 M ZnCl2 solution for 48 hours. Second-instar larvae were collected in 1.5 ml centrifuge tubes using a pipette and enriched by brief centrifugation (5000 rpm for 30 seconds) for later use. In addition, this experiment also prepared experimental nematodes such as *Gnaphalium affine*, *C. elegans*, and *Caenorhabditis elegans* using conventional methods.

[0023] 3. Test methods

[0024] Take 100 μl of each test solution prepared in step 1 into a 20-well plate, add 50-100 live nematodes prepared in step 2, and place at 25℃. Examine the nematodes under an optical microscope at 24, 48, and 72 hours to calculate the mortality rate. The method for identifying mortality is as follows: add 1-5 drops of 5% NaCl solution to the wells of the treatment plate, observe after 2 minutes. Dead nematodes will be stiff and show no reaction to needle stimulation, while live nematodes will curl or twist.

[0025] Using the test solution without the sample as a control, the entire experiment was repeated three times, and the average value of the three tests was taken to calculate the corrected mortality rate.

[0026] Mortality rate % = (Number of dead nematodes / (Number of dead nematodes + Number of surviving nematodes)) × 100%

[0027] Corrected mortality rate % = Sample mortality rate – Control mortality rate

[0028] 4. Experimental Results

[0029] 4.1 N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1HThe toxic effects of pyrazol-1-yl)-6-morpholinopyrimidine-4-amine on southern root-knot nematodes are shown in Table 1:

[0030] Table 1 N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H Results of pyrazol-1-yl)-6-morpholinopyrimidine-4-amine on the toxicity of southern root-knot nematodes

[0031]

[0032] 4.2 N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H The toxic effects of pyrazol-1-yl)-6-morpholinopyrimidine-4-amine on Javan root-knot nematodes are shown in Table 2.

[0033] Table 2 N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H Results of pyrazol-1-yl)-6-morpholinopyrimidine-4-amine on Javan root-knot nematodes

[0034]

[0035] 4.3 N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H The toxic effects of pyrazol-1-yl)-6-morpholinopyrimidine-4-amine on *Auricularia auricula-judae* root-knot nematode are shown in Table 3.

[0036] Table 3 N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H The toxicity of pyrazol-1-yl)-6-morpholinopyrimidine-4-amine to *Heliotropium indicum* root-knot nematodes.

[0037]

[0038] 4.4 N -(4,4-difluorocyclohexyl)-2-(3-methyl- 1H (-pyrazol-1-yl)-6-morpholinopyrimidine-4-amine showed no toxic activity against the non-plant pathogenic nematode Caenorhabditis elegans within 24 hours.

[0039] Results Analysis: Tables 1-3 show that N-(4,4-difluorocyclohexyl)-2-(3-methyl-1H-pyrazol-1-yl)-6-morpholinopyrimidine-4-amine exhibits strong toxic activity against plant root-knot nematodes. Its 24-hour median lethal dose (LC50) against *Symplocos septemlobus*, *Cytomalacia javanica*, and *Eriocaulon buergerianum* was [data missing]. 50The concentrations were 12.70 μg / ml, 24.35 μg / ml, and 20.57 μg / ml, respectively; the median lethal dose (LC50) at 48 hours was [not specified in the original text]. 50 The concentrations were 12.50 μg / ml, 23.46 μg / ml, and 19.47 μg / ml, respectively. Figure 2-4 ).

[0040] Example 2

[0041] An insecticide for controlling plant root-knot nematodes, wherein the insecticide uses a 6% ethanol solution as a solvent and N-(4,4-difluorocyclohexyl)-2-(3-methyl-1H-pyrazol-1-yl)-6-morpholinopyrimidine-4-amine at a concentration of 30 μg / ml as the active ingredient.

[0042] Example 3

[0043] An insecticide for controlling plant root-knot nematodes, wherein the insecticide uses a 4% methanol solution as a solvent and N-(4,4-difluorocyclohexyl)-2-(3-methyl-1H-pyrazol-1-yl)-6-morpholinopyrimidine-4-amine at a concentration of 20 μg / ml as the active ingredient.

[0044] Example 4

[0045] An insecticide for controlling plant root-knot nematodes, wherein the insecticide uses a 10% propanol solution as a solvent and N-(4,4-difluorocyclohexyl)-2-(3-methyl-1H-pyrazol-1-yl)-6-morpholinopyrimidine-4-amine at a concentration of 50 μg / ml as the active ingredient.

[0046] Example 5

[0047] An insecticide for controlling plant root-knot nematodes, wherein the insecticide uses a 5% dimethyl sulfoxide aqueous solution as a solvent and N-(4,4-difluorocyclohexyl)-2-(3-methyl-1H-pyrazol-1-yl)-6-morpholinopyrimidine-4-amine at a concentration of 10 μg / ml as the active ingredient.

[0048] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any changes, modifications, substitutions, combinations, or simplifications made without departing from the spirit and principle of the present invention shall be considered equivalent substitutions and shall be included within the protection scope of the present invention.

Claims

1. Pyrazole compounds N -(4,4-difluorocyclohexyl)-2-(3-methyl-1 H The application of pyrazol-1-yl)-6-morpholinopyrimidine-4-amine and / or its pesticide-acceptable salts in the control of plant root-knot nematodes at concentrations of 3–50 μg / mL.

2. The application according to claim 1, characterized in that, The root-knot nematodes are Southern Root-Knot Nematodes, Java Root-Knot Nematodes, or Elephant Bean Root-Knot Nematodes.

3. The application according to claim 1, characterized in that, Using the above N -(4,4-difluorocyclohexyl)-2-(3-methyl-1 H Preparations for controlling plant root-knot nematodes are made from pyrazol-1-yl)-6-morpholinopyrimidine-4-amine and / or its phytochemically acceptable salts.

4. The application according to claim 3, characterized in that, The formulations for controlling plant root-knot nematodes are aqueous solutions, emulsions, wettable powders, soluble powders, soluble granules, water-dispersible granules, suspensions, tablets, or effervescent granules.

5. The application according to claim 4, characterized in that, The solvent for the aqueous solution or water emulsion is a fatty alcohol aqueous solution or a dimethyl sulfoxide aqueous solution with a concentration ≤5%.

6. The application according to claim 5, characterized in that, The fatty alcohol is methanol, ethanol, or propanol.

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