CN112237187A - Application of phenolic compound as nematicide - Google Patents
Application of phenolic compound as nematicide Download PDFInfo
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- CN112237187A CN112237187A CN202010227637.XA CN202010227637A CN112237187A CN 112237187 A CN112237187 A CN 112237187A CN 202010227637 A CN202010227637 A CN 202010227637A CN 112237187 A CN112237187 A CN 112237187A
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N31/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic oxygen or sulfur compounds
- A01N31/08—Oxygen or sulfur directly attached to an aromatic ring system
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N37/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids
- A01N37/36—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids containing at least one carboxylic group or a thio analogue, or a derivative thereof, and a singly bound oxygen or sulfur atom attached to the same carbon skeleton, this oxygen or sulfur atom not being a member of a carboxylic group or of a thio analogue, or of a derivative thereof, e.g. hydroxy-carboxylic acids
- A01N37/38—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids containing at least one carboxylic group or a thio analogue, or a derivative thereof, and a singly bound oxygen or sulfur atom attached to the same carbon skeleton, this oxygen or sulfur atom not being a member of a carboxylic group or of a thio analogue, or of a derivative thereof, e.g. hydroxy-carboxylic acids having at least one oxygen or sulfur atom attached to an aromatic ring system
- A01N37/40—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids containing at least one carboxylic group or a thio analogue, or a derivative thereof, and a singly bound oxygen or sulfur atom attached to the same carbon skeleton, this oxygen or sulfur atom not being a member of a carboxylic group or of a thio analogue, or of a derivative thereof, e.g. hydroxy-carboxylic acids having at least one oxygen or sulfur atom attached to an aromatic ring system having at least one carboxylic group or a thio analogue, or a derivative thereof, and one oxygen or sulfur atom attached to the same aromatic ring system
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Abstract
The invention belongs to the field of pesticides, and particularly relates to application of a phenolic compound as a nematicide. The application of one or more phenolic compounds shown in a general formula I or salts thereof as nematicides; the general formula I is shown as follows; the compound is suitable for being used as a nematicide, particularly the phenol sodium salt with better activity is commonly used as a preservative, and 3-hydroxybenzoic acid and 4-hydroxybenzoic acid are antioxidants widely applied at present, and have the advantages of low toxicity, low price and easy obtainment.
Description
Technical Field
The invention belongs to the field of pesticides, and particularly relates to an application of a phenolic compound as a nematicide (especially Bursaphelenchus xylophilus).
Background
The nematodes belong to nematode animal phyla in classification status, are various in types and extremely wide in geographical distribution, plant nematode diseases are developed into important factors for restricting the health and sustainable development of agriculture and forestry globally, and the conservative estimation of economic loss caused each year is about 1570 hundred million dollars. Meloidogyne incognita, Meloidogyne javanica, M.janicuda, M.arenaria and M.hapla are the most common species of Meloidogyne incognita, and M.hapla, which are the dominant species in China. The nematode has small body type and the harmful part is usually positioned at the root system of a plant submerged in soil, the harm is not easy to be perceived in the early stage, although the activity of the nematode is very limited (the annual moving range of the nematode may not exceed 1m), the nematode can realize effective long-distance and short-distance transmission by depending on rainwater, surface and underground runoff, farm work operation, sick seedlings and the like, the stress resistance is very strong, and the nematode can survive for 3 years without food sources. Once introduced into the plant, nematodes are difficult to eradicate.
Pine wood nematode (Bursaphelenchus xylophilus) disease is the most dangerous forest disease in the world, mainly damaging the Pinaceae. The plant can die as soon as 40 days after the pine wood nematode disease is infected by a vector longicorn (Monochamus spp.), and only 3-5 years are needed from the disease to the destruction of the whole pine forest. The disease has wide spread path, high spread speed and great control difficulty, is classified as an international quarantine disease, and is the primary prevention and control object in forestry. The epidemic situation is firstly discovered in Nanjing Zhongshan Ling in China in 1982, and has spread to 18 provinces and 588 counties (urban areas) in China to date, the diseased and dead trees reach more than 1000 million, the occurrence area is nearly 1000 million mu, the survival and death of 9 million mu pine forest in China are threatened, and great threat is brought to the production and ecological safety of forestry in China. At present, the pine trees lost due to the pine wilt disease in China are accumulated to reach billions of plants, and the direct economic loss and the loss of the ecological service value are billions of yuan.
Chemical control is the main measure for controlling nematodes at present. As 12 months in 2019, 312 nematicidal products are registered in China, the active ingredients mainly concentrate on abamectin, fosthiazate, methyl isoxathion, fenamiphos, methyl bromide, aldicarb, amino-oligosaccharin, dazomet, cartap, metam, carbofuran, carbosulfan and the like, most of the products are organic phosphorus products, and the toxicity is high. With the development of low toxicity and environmental protection of raw medicines and dosage forms, the traditional highly toxic and highly toxic raw medicines such as carbamate pesticides and organophosphorus pesticides are gradually limited or forbidden, and meanwhile, a plurality of dosage forms such as fumigants are difficult to exert effects on nematodes in an open-air non-closed environment and face the embarrassment of quitting the stage; further, nematodes on crops such as soybean, corn, cotton, fruits and vegetables are getting more and more serious, and new active substances for killing nematodes are urgently needed for forestry epidemic-detection pine wood nematodes. At present, only the curative cutting can be carried out on pines in pine nematode affected areas, the affected trees subjected to the curative cutting need to be ground or burnt in the mountain areas, the burnt affected trees need to be photographed and filed in the whole process, the ground affected trees can be used only in the local areas, the cross-provincial and political areas are forbidden to be used, and the mountain areas are forbidden to process plates by the affected trees. The cost for preventive and preventive felling of each epidemic wood is approximately 150-200 yuan according to the initial rough estimation.
Disclosure of Invention
The invention aims to solve the problems in the prior art and provides application of phenolic compounds as nematicides.
In order to achieve the purpose, the invention adopts the technical scheme that:
use of a phenolic compound as a nematicide characterized in that: the application of one or more phenolic compounds shown in a general formula I or salts thereof as nematicides;
the general formula I is as follows:
in the formula (I), the compound is shown in the specification,
R1is selected from C1-C4Alkyl, hydrogen or hydroxy of (a); r2、R3Are identical or different and are each selected from C1-C4Alkyl, hydrogen or carboxyl of (a); r4、R5May be the same or different and are each selected from hydrogen or C1-C4Alkyl groups of (a); when the alkyl is butyl, the alkyl is not sec-butyl;
and when R is2、R4And R5When it is hydrogen, R1、R3Is not tert-butyl at the same time; when in useR2、R3And R4When it is hydrogen, R1、R5Not tert-butyl at the same time.
Preferably, in the formula I, R1H, OH or butyl; r2、R3Can be the same or different and is respectively H, methyl, butyl or COOH; r4、R5Can be the same or different and is respectively H or butyl;
or, salts of the compounds of formula I.
The salt of the compound shown in the general formula I is formed by replacing hydrogen in hydroxyl or carboxyl in the compound shown in the general formula I by metal ions. Furthermore, in the compound shown in the general formula I, hydrogen in hydroxyl or carboxyl is replaced by metal ions such as Na, K and the like to form corresponding salt.
The application of one or more phenolic compounds shown as the general formula I or salts thereof in protecting plants or plant propagation materials against damage caused by pine wood nematodes (Bursaphelenchus xylophilus), retransmission caused by transportation or re-harm nematicides.
The active ingredient of the nematicide contains one or more of phenolic compounds shown as the general formula I or salts thereof.
The nematicide is an active ingredient and is mixed with an agriculturally acceptable carrier; wherein the active ingredient accounts for 0.01-99% of the weight of the nematicide.
A nematicidal composition preparation comprises an active component A and an active component B, wherein the active component A and the active component B are mixed according to the weight part ratio of 1:100-100: 1; the active component A is one or more of phenolic compounds shown in a general formula I or salts thereof in claim 1; the active component B is one or more of other nematocides, insecticides, acaricides and bactericides; preferably: such as abamectin, matrine, fosthiazate, pyraclostrobin and amino-oligosaccharin.
The preparation is injection, spray, fumigant, coating agent or nanocrystallization preparation. Wherein, the nano preparation can be prepared into a compound nano preparation by adopting a wet medium grinding method, and the nano particle size is 50-500nm, preferably 300-500 nm.
A method for protecting a plant or its propagation material against damage caused by nematodes, said method comprising applying to the plant or its locus a formulation of said phenolic compound of formula i or one or more of its salts, said formulation or said composition. Wherein, the place is the place which is polluted by the nematodes and is needed to be treated in order to meet the reproduction requirement, such as agricultural or forestry land.
A method of preventing the re-spread and re-hazard of nematodes following infestation of plant transport, said method comprising applying to the plant material following nematode infestation a formulation of said phenolic compound of formula i or one or more of its salts, said formulation or said composition formulation for four weeks.
The compound is used as a nematicide, not only can effectively control the harm of the pine wood nematodes, but also can greatly reduce the cost of the nematicide by being easy to obtain, and can be combined with the nematicide, the insecticide, the acaricide, the bactericide and the like for use, such as abamectin, matrine, fosthiazate, pyraclostrobin and amino-oligosaccharin, so that the activity can be obviously improved; the preparation of the invention is beneficial to subsequent market popularization and application.
Drawings
FIG. 1 is a graph of a nematode corrected mortality fit 2h after treatment with the agent as provided in example 1 of the present invention.
FIG. 2 is a graph of a fitted nematode corrected mortality curve 2h after treatment with the agent as provided in example 2 of the present invention.
FIG. 3 is a fitted graph of nematode corrected mortality after 24h treatment with the agents provided in example 3 of the present invention.
FIG. 4 is a fitted graph of nematode corrected mortality after 2h treatment with the agents provided in example 5 of the present invention.
FIG. 5 is a fitted graph of nematode corrected mortality after 24h treatment with the agents provided in example 5 of the present invention.
FIG. 6 is a fitted graph of nematode corrected mortality after 2h treatment with the agents provided in example 6 of the present invention.
FIG. 7 is a fitted graph of nematode corrected mortality after 24h treatment with the agents provided in example 6 of the present invention.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below, and the described embodiments are only a part of the present invention, and not all of the present invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
In the following experimental materials of the examples:
reagent to be tested: the phenolic compounds shown in the general formula I are known compounds, and are low in cost and easy to obtain.
Example 1, poisoning Activity of 2, 6-Di-tert-butyl-p-cresol on Bursaphelenchus xylophilus and LC50 determination
Preparing a medicament:
the following experimental group medicament is prepared by preparing 2, 6-di-tert-butyl-p-cresol into 2, 6-di-tert-butyl-p-cresol aqueous solutions with different concentrations through aqueous solutions containing DMSO and Tween 80; wherein, the DMSO content in the aqueous solution is 20%, and the Tween 80 content is 10%.
Experimental groups:
1) 100mL of 2, 6-di-tert-butyl-p-cresol aqueous solution with the concentration of 10 g/L;
2) 100mL of 2, 6-di-tert-butyl-p-cresol aqueous solution with the concentration of 20 g/L;
3) 100mL of an aqueous solution of 30 g/L2, 6-di-tert-butyl-p-cresol.
Control group: the corresponding system solvent of each experimental group is used as a blank control of each experimental group.
Preparation of nematode suspension: suspending the pine wood nematodes on the surface of a 9cm PDA culture dish in a water phase by using a proper amount of sterile water, separating by a Bellman funnel method to obtain a live pine wood nematode suspension, centrifuging the nematode suspension for 2min at 2000r/min, removing supernatant, retaining precipitates, adding a proper amount of sterile water, performing microscopic examination to observe the age and activity of the nematodes and count the number of the nematodes, and finally quantifying the nematode suspension to 1000 nematodes/mL by using the sterile water, wherein most nematodes are in the J3 stage, the few are in the J2 stage and the J4 stage, and the temperature is 4 ℃ for later use.
Medicament treatment: taking 0.5mL of the prepared nematode suspension, centrifuging for 2min at 2000r/min, removing supernatant, adding 0.5mL of different reagent to be tested into the precipitate, blowing uniformly by using a pipette, covering the centrifuge tube, culturing in a climatic chamber with the temperature of 25 ℃, the relative humidity of 60% and no light, observing and recording the total number of nematodes and the number of dead nematodes under a 40-time Olympus CX33 optical microscope after 2 h.
TABLE 12 poisoning effect of 6, 6-di-tert-butyl-p-cresol on Bursaphelenchus xylophilus at different working concentrations
The experimental results are as follows: as can be seen from Table 1 and FIG. 1, 2, 6-di-tert-butyl-p-cresol was used to treat the Bursaphelenchus xylophilus for 2h by the method of immersion and the fitting curve obtained from the data was that y is 3.5x-12.5, R is20.8942, the 2, 6-di-tert-butyl-p-cresol was obtained at 17.86g/L against pine wood nematode LC50, and the mortality of pine wood nematodes exceeded 95% when the working concentration reached 30g/L, and it was seen that 2, 6-di-tert-butyl-p-cresol had potent pine wood nematode killing activity.
Example 2 poisoning Activity of phenol (phenol) against Bursaphelenchus xylophilus and determination of LC50
Preparing a medicament: phenol was formulated with sterile water to 9 concentrations of 0, 0.1, 1, 2, 3,4, 5, 15 and 30g/L, with 0g/L being a control.
Preparation of nematode suspension: suspending the pine wood nematodes on the surface of a 9cm PDA culture dish in a water phase by using a proper amount of sterile water, separating by a Bellman funnel method to obtain a live pine wood nematode suspension, centrifuging the nematode suspension for 2min at 2000r/min, removing supernatant, retaining precipitates, adding a proper amount of sterile water, performing microscopic examination to observe the age and activity of the nematodes and count the number of the nematodes, and finally quantifying the nematode suspension to 1000 nematodes/mL by using the sterile water, wherein most nematodes are in the J3 stage, the few are in the J2 stage and the J4 stage, and the temperature is 4 ℃ for later use.
Medicament treatment: taking 0.5mL of the prepared nematode suspension, centrifuging for 2min at 2000r/min, removing supernatant, adding 0.5mL of different reagent to be tested into the precipitate, blowing uniformly by using a pipette, covering the centrifuge tube, culturing in a climatic chamber with the temperature of 25 ℃, the relative humidity of 60% and no light, observing and recording the total number of nematodes and the number of dead nematodes under a 40-time Olympus CX33 optical microscope after 2 h.
TABLE 2 poisoning effect of phenol on Bursaphelenchus xylophilus at different working concentrations
As can be seen from table 2 and fig. 2 above, after phenol treatment for 2h, the fitted curve obtained from the data was y-0.2733 x, R2-0.9551, thus obtaining phenol at 1.83g/L against pine wood nematode LC 50; further, it can be seen that phenol has a potent activity against bursaphelenchus xylophilus.
Example 3 poisoning Activity of sodium phenolate against Bursaphelenchus xylophilus and LC50 assay
Preparing a medicament: sodium phenolate was formulated with sterile water to 5 concentrations of 0, 1, 2, 3 and 4g/L, with 0g/L being the control.
Preparation of nematode suspension: suspending the pine wood nematodes on the surface of a 9cm PDA culture dish in a water phase by using a proper amount of sterile water, separating by a Bellman funnel method to obtain a live pine wood nematode suspension, centrifuging the nematode suspension for 2min at 2000r/min, removing supernatant, retaining precipitates, adding a proper amount of sterile water, performing microscopic examination to observe the age and activity of the nematodes and count the number of the nematodes, and finally quantifying the nematode suspension to 1000 nematodes/mL by using the sterile water, wherein most nematodes are in the J3 stage, the few are in the J2 stage and the J4 stage, and the temperature is 4 ℃ for later use.
Medicament treatment: taking 0.5mL of the prepared nematode suspension, centrifuging for 2min at 2000r/min, removing supernatant, adding 0.5mL of different reagent to be tested into the precipitate, blowing uniformly by using a pipette, covering the centrifuge tube, culturing in a climatic chamber with the temperature of 25 ℃, the relative humidity of 60% and no light, observing and recording the total number and the dead number of the nematodes under a 40-time Olympus CX33 optical microscope after 24 h.
TABLE 3 poisoning effect of sodium phenolate on Bursaphelenchus xylophilus at different working concentrations
As can be seen from Table 3 and FIG. 3, sodium phenolate was used to treat Bursaphelenchus xyfolus for 24h by the method of immersion, and the fitting curve obtained from the data was that y is 46.25x +8.6833, R20.9044, sodium phenate was thus obtained at 0.89g/L against pine wood nematode LC 50; further, sodium phenolate was found to have potent activity against pine wood nematodes.
EXAMPLE 4 determination of the poisoning Activity of 3-hydroxybenzoic acid (3-Hydroxybenzoic acid) against Bursaphelenchus xylophilus
Preparing a medicament: 3-hydroxybenzoic acid was formulated with 4% DMSO + 15% Tween in water to 5 concentrations of 0, 1, 10, 20 and 30g/L, with 0g/L being the control.
Preparation of nematode suspension: suspending the pine wood nematodes on the surface of a 9cm PDA culture dish in a water phase by using a proper amount of sterile water, separating by a Bellman funnel method to obtain a live pine wood nematode suspension, centrifuging the nematode suspension for 2min at 2000r/min, removing supernatant, retaining precipitates, adding a proper amount of sterile water, performing microscopic examination to observe the age and activity of the nematodes and count the number of the nematodes, and finally quantifying the nematode suspension to 1000 nematodes/mL by using the sterile water, wherein most nematodes are in the J3 stage, the few are in the J2 stage and the J4 stage, and the temperature is 4 ℃ for later use.
Medicament treatment: taking 0.5mL of the prepared nematode suspension, centrifuging for 2min at 2000r/min, removing supernatant, adding 0.5mL of different reagent to be tested into the precipitate, blowing uniformly by using a pipette, covering the centrifuge tube, culturing in a climatic chamber with the temperature of 25 ℃, the relative humidity of 60% and no light, observing and recording the total number and the dead number of the nematodes under a 40-time Olympus CX33 optical microscope after 24 h.
TABLE 43 poisoning Effect of hydroxybenzoic acids on Bursaphelenchus Xyla at different working concentrations
The data in the table show that the 3-hydroxybenzoic acid has certain activity of killing pine wood nematodes.
Example 5 poisoning Activity of 4-hydroxybenzoic acid (4-Hydroxybenzoic acid) against Bursaphelenchus xyfolus and LC50 assay
Preparing a medicament: 4-hydroxybenzoic acid was dissolved in sterile water to make up the drug at concentrations of 0, 1, 3, and 5g/L4, with 0g/L being the control.
Preparation of nematode suspension: suspending the pine wood nematodes on the surface of a 9cm PDA culture dish in a water phase by using a proper amount of sterile water, separating by a Bellman funnel method to obtain a live pine wood nematode suspension, centrifuging the nematode suspension for 2min at 2000r/min, removing supernatant, retaining precipitates, adding a proper amount of sterile water, performing microscopic examination to observe the age and activity of the nematodes and count the number of the nematodes, and finally quantifying the nematode suspension to 1000 nematodes/mL by using the sterile water, wherein most nematodes are in the J3 stage, the few are in the J2 stage and the J4 stage, and the temperature is 4 ℃ for later use.
Medicament treatment: taking 0.5mL of the prepared nematode suspension, centrifuging for 2min at 2000r/min, removing supernatant, adding 0.5mL of different reagent to be tested into the precipitate, blowing uniformly by using a pipette, covering the centrifuge tube, culturing in a dark artificial climate box at 25 ℃, with relative humidity of 60%, and observing and recording the total number of nematodes and the number of dead nematodes under a 40-time Olympus CX33 optical microscope after 2h and 24 h.
TABLE 54 poisoning effect of hydroxybenzoic acids on Bursaphelenchus xyfolus at different working concentrations
As can be seen from table 5 above and fig. 4 and 5, the data after 2h treatment with 4-hydroxybenzoic acid gave a fitted curve with y ═ 0.0442x-0.0262, R20.9253, 11.9g/L for Bursaphelenchus xylophilus LC 50; after processing for 2h at 24h, a fitted curve is obtained according to data, wherein y is 0.066x-0.0094, and R is20.9163, 7.7g/L for bursaphelenchus xylophilus LC 50; increasing the concentration of 4-hydroxybenzoic acidThe poisoning activity of the compound on the pine wood nematodes is in a positive correlation, and the half-lethal concentration of the 4-hydroxybenzoic acid on the pine wood nematodes is in a descending trend along with the prolonging of the treatment time, so that the 4-hydroxybenzoic acid has the activity of killing the pine wood nematodes.
Example 6 poisoning Activity of 3,4-Dihydroxybenzoic acid (3, 4-dihydrobenzoic acid) against Bursaphelenchus xyfolus and LC50 assay
Preparing a medicament: 3,4-dihydroxy benzoic acid was dissolved in sterile water to prepare 5 concentrations of 0, 5, 10, 15, and 20g/L, wherein 0g/L was the control.
Preparation of nematode suspension: suspending the pine wood nematodes on the surface of a 9cm PDA culture dish in a water phase by using a proper amount of sterile water, separating by a Bellman funnel method to obtain a live pine wood nematode suspension, centrifuging the nematode suspension for 2min at 2000r/min, removing supernatant, retaining precipitates, adding a proper amount of sterile water, performing microscopic examination to observe the age and activity of the nematodes and count the number of the nematodes, and finally quantifying the nematode suspension to 1000 nematodes/mL by using the sterile water, wherein most nematodes are in the J3 stage, the few are in the J2 stage and the J4 stage, and the temperature is 4 ℃ for later use.
Medicament treatment: taking 0.5mL of the prepared nematode suspension, centrifuging for 2min at 2000r/min, removing supernatant, adding 0.5mL of different reagent to be tested into the precipitate, blowing uniformly by using a pipette, covering the centrifuge tube, culturing in a dark artificial climate box at 25 ℃, with relative humidity of 60%, and observing and recording the total number of nematodes and the number of dead nematodes under a 40-time Olympus CX33 optical microscope after 2h and 24 h.
TABLE 63 poisoning Effect of 4-Dihydroxybenzoic acid on Bursaphelenchus xyfolus at different working concentrations
As can be seen from Table 6 and FIGS. 6 and 7, the roots of 3,4-dihydroxybenzoic acid after 2 hours of treatmentObtaining a fitting curve with y being 0.0174x-0.0138 according to the data, and R20.8982, 29.5g/L for bursaphelenchus xylophilus LC 50; after 24h treatment, the fitted curve obtained from the data was y-0.0297 x-0.007, R20.9296, 17.1g/L for bursaphelenchus xylophilus LC 50; it can be seen that 3,4-dihydroxybenzoic acid has activity against Bursaphelenchus xylophilus.
Example 7 synergistic experiment of Compound pesticide 4-hydroxybenzoic acid on poisoning Activity of Bursaphelenchus xylophilus
Pharmaceutical formulation
The following solutions were prepared with sterilized water as test agents, respectively:
1) 4-hydroxybenzoic acid is prepared into 5g/L aqueous solution by sterilized water;
2) preparing the matrine into 0.6mg/L aqueous solution by using sterilized water;
3) preparing the abamectin into 18mg/L aqueous solution by using sterilized water;
4) the compound preparation 1 is prepared by adding 4-hydroxybenzoic acid and matrine into sterilized water, wherein the final concentration of 4-hydroxybenzoic acid is 5g/L, and the final concentration of matrine is 0.6 mg/L;
5) the compound medicament 2 is prepared by adding 4-hydroxybenzoic acid and abamectin into sterilized water, wherein the final concentration of the 4-hydroxybenzoic acid is 5g/L, and the final concentration of the abamectin is 0.6 mg/L;
6) sterile water was used as a control group;
preparation of nematode suspension: suspending the pine wood nematodes on the surface of a 9cm PDA culture dish in a water phase by using a proper amount of sterile water, separating by a Bellman funnel method to obtain a live pine wood nematode suspension, centrifuging the nematode suspension for 2min at 2000r/min, removing supernatant, retaining precipitates, adding a proper amount of sterile water, performing microscopic examination to observe the age and activity of the nematodes and count the number of the nematodes, and finally quantifying the nematode suspension to 1000 nematodes/mL by using the sterile water, wherein most nematodes are in the J3 stage, the few are in the J2 stage and the J4 stage, and the temperature is 4 ℃ for later use.
Medicament treatment: taking 0.5mL of the prepared nematode suspension, centrifuging for 2min at 2000r/min, removing supernatant, adding 0.5mL of different reagent to be tested into the precipitate, blowing uniformly by using a pipette, covering the centrifuge tube, culturing in a climatic chamber with the temperature of 25 ℃, the relative humidity of 60% and no light, observing and recording the total number and the dead number of the nematodes under a 40-time Olympus CX33 optical microscope after 24 h.
Synergistic effect of compound pesticide of table 74-hydroxybenzoic acid on poisoning activity of pine wood nematodes
Synergistic effect of surface 84-hydroxybenzoic acid compound pesticide on poisoning activity of pine wood nematodes
And (4) conclusion: from tables 7 and 8, it can be seen that the poisoning activity of 4-hydroxybenzoic acid compounded with abamectin and matrine on the pine wood nematode is greatly improved compared with abamectin and matrine which are not compounded with 4-hydroxybenzoic acid, and the synergistic effects are 57.43% and 74.39% respectively; compared with 4-hydroxybenzoic acid, the abamectin and matrine compound preparation respectively has 35.13 percent and 62.04 percent of synergistic effect. The 4-hydroxybenzoic acid compound pesticide can greatly improve the line killing activity of the pesticide.
Example 8 synergistic experiment of Compound-Compound against Bursaphelenchus xylophilus poisoning Activity
Preparing a medicament:
the following solutions were prepared with sterile water and used as the group of formulas:
1) 4-hydroxybenzoic acid is prepared into a 2-ethyl hydroxybenzoate water solution with the concentration of 20g/L by using sterilized water;
2)3, 4-dihydroxy benzoic acid is prepared into a 22g/L ethyl 3-hydroxybenzoate water solution by using sterilized water;
3) compounding a medicament: adding 4-hydroxybenzoic acid and 3,4-dihydroxybenzoic acid to sterilized water; wherein the final concentration of the 4-hydroxybenzoic acid is 5g/L, and the final concentration of the 3,4-dihydroxybenzoic acid is 10 g/L;
4) sterile water is used as a blank control;
preparation of nematode suspension: suspending the pine wood nematodes on the surface of a 9cm PDA culture dish in a water phase by using a proper amount of sterile water, separating by a Bellman funnel method to obtain a live pine wood nematode suspension, centrifuging the nematode suspension for 2min at 2000r/min, removing supernatant, retaining precipitates, adding a proper amount of sterile water, performing microscopic examination to observe the age and activity of the nematodes and count the number of the nematodes, and finally quantifying the nematode suspension to 1000 nematodes/mL by using the sterile water, wherein most nematodes are in the J3 stage, the few are in the J2 stage and the J4 stage, and the temperature is 4 ℃ for later use.
Medicament treatment: taking 0.5mL of the prepared nematode suspension, centrifuging for 2min at 2000r/min, removing supernatant, adding 0.5mL of different reagent to be tested into the precipitate, blowing uniformly by using a pipette, covering the centrifuge tube, culturing in a dark artificial climate box at 25 ℃, with relative humidity of 60%, and observing and recording the total number of nematodes and the number of dead nematodes under a 40-time Olympus CX33 optical microscope after 24 h.
TABLE 9 poisoning Activity of combinations of Compounds on Bursaphelenchus xylophilus
Table 10 synergistic combination of the compounds with poisoning activity against Bursaphelenchus xylophilus
| Compound compound | Has a synergistic effect range compared with that of 4-hydroxybenzoic acid | Compared with the 3,4-dihydroxy benzoic acid, the synergistic range |
| 4-hydroxybenzoic acid and 3,4-dihydroxybenzoic acid | 52.91％ | 40.10％ |
And (4) conclusion: as shown in tables 9 and 10, the two compounds of 4-hydroxybenzoic acid and 3,4-dihydroxybenzoic acid have obvious synergistic effect, and the two-component compound has at least 40.10 percent synergistic effect compared with the single compound.
Example 94-method for nanoscopic preparation of hydroxybenzoic acid and determination of the poisoning activity against Bursaphelenchus xylophilus 1) method for preparation of nanoscopic preparation of 4-hydroxybenzoic acid:
0.3g of Tween 80 and 0.3g of PVP K30 were added to 100ml of deionized water, and dissolved in a magnetic stirrer at 500r/min to form an aqueous phase. 0.5g of 4-hydroxybenzoic acid is weighed and added into the solution, the mixture is transferred to a WG-0.3L medium grinder after being mixed evenly, zirconia beads with the diameter of 0.3mm are selected as grinding media, the filling amount of the media is 75 percent of the cavity of the grinder, the mixture is ground for 20min at the speed of 2000r/min, the ground nanosuspension is divided into 4 parts, and the parts are respectively placed at the temperature of 52 ℃ for 0.5h, 2d, 6d and 12d, and then the particle size, the polydispersity and the decomposition rate of the 4-hydroxybenzoic acid of the nanosuspension are measured and counted (Table 11).
Table 11 nanosuspension the various indices are shown in the following table:
after being ground by the wet medium, the 4-hydroxybenzoic acid has average particle size of less than 500nm and polydispersity PDI of less than 0.25 when stored for 12 days at 25 ℃, which shows that the solution particles not only reach the nano level, but also the 4-hydroxybenzoic acid has narrow distribution in the nano suspension and better system uniformity. In the wet medium grinding process, due to the fact that a certain decomposition rate exists in the effective components due to repeated collision of the grinding medium, the decomposition rates of the 4-hydroxybenzoic acid in the nanometer suspension are less than 5% through high performance liquid chromatography quantitative analysis, the GB/T19136-containing 2003 'determination method for pesticide thermal storage stability' stipulates, the pesticide passes a thermal storage chemical stability test, and the medicine is qualified when the decomposition rate of the effective components is less than 5%. In conclusion, the average particle size of the active ingredients, the dispersibility of the active ingredients in the system, the stability of the active ingredients and the like of the 4-hydroxybenzoic acid nanosuspension prepared by the medium grinding method meet the requirements, so that the qualified 4-hydroxybenzoic acid nanosuspension can be prepared by using the method.
2) The poisoning activity of the 4-hydroxybenzoic acid nano preparation on the pine wood nematodes is determined:
preparing a medicament: taking the prepared 4-hydroxybenzoic acid nanometer suspension with the effective component concentration of 5g/L, preparing 4-hydroxybenzoic acid solution with the concentration of 5g/L and without nano treatment, and preparing 0.3 percent of Tween 80+ PVP K30 aqueous solution as a contrast agent.
Preparation of nematode suspension: suspending the pine wood nematodes on the surface of a 9cm PDA culture dish in a water phase by using a proper amount of sterile water, separating by a Bellman funnel method to obtain a live pine wood nematode suspension, centrifuging the nematode suspension for 2min at 2000r/min, removing supernatant, retaining precipitates, adding a proper amount of sterile water, performing microscopic examination to observe the age and activity of the nematodes and count the number of the nematodes, and finally quantifying the nematode suspension to 1000 nematodes/mL by using the sterile water, wherein most nematodes are in the J3 stage, the few are in the J2 stage and the J4 stage, and the temperature is 4 ℃ for later use.
Medicament treatment: taking 0.5mL of the prepared nematode suspension, centrifuging for 2min at 2000r/min, removing supernatant, adding 0.5mL of different reagent to be tested into the precipitate, blowing uniformly by using a pipette, covering the centrifuge tube, culturing in a climatic chamber with the temperature of 25 ℃, the relative humidity of 60% and no light, observing and recording the total number and the dead number of the nematodes under a 40-time Olympus CX33 optical microscope after 24 h.
TABLE 124-hydroxybenzoic acid Mitigo-and Prenematicidal Activity
And (4) conclusion: after the 4-hydroxybenzoic acid is subjected to nanocrystallization, the insecticidal activity of the 4-hydroxybenzoic acid on the pine wood nematodes is greatly improved, and the improvement range is 172.44%.
In conclusion, the phenolic compounds and the salts thereof described in the general formula of the invention have obvious pine wood nematode killing activity, and the mixing and matching of different phenolic compounds and salts thereof in the general formula or the mixing and matching of the phenolic compounds and salts thereof with other insecticides and nematicides have synergistic effect on killing nematodes with poison, the compound with a structure similar to that of the general formula has no thread killing effect, and the general formula shows obvious biological characteristics on killing nematodes with poison.
Claims (9)
1. Use of a phenolic compound as a nematicide characterized in that: the application of one or more phenolic compounds shown in a general formula I or salts thereof as nematicides;
the general formula I is as follows:
in the formula (I), the compound is shown in the specification,
R1is selected from C1-C4Alkyl, hydrogen or hydroxy of (a); r2、R3Are identical or different and are each selected from C1-C4Alkyl, hydrogen or carboxyl of (a); r4、R5May be the same or different and are each selected from hydrogen or C1-C4Alkyl groups of (a); when the alkyl is butyl, the alkyl is not sec-butyl;
and when R is2、R4And R5When it is hydrogen, R1、R3Is not tert-butyl at the same time; when R is2、R3And R4When it is hydrogen, R1、R5Not tert-butyl at the same time.
2. Use according to claim 1, characterized in that: in the general formula I, R1H, OH or butyl; r2、R3Can be the same or different and is respectively H, methyl, butyl or COOH; r4、R5Can be the same or different and is respectively H or butyl;
or, salts of the compounds of formula I.
3. Use according to claim 1 or 2, characterized in that: the application of one or more phenolic compounds shown as the general formula I or salts thereof in serving as nematicides for protecting plants or plant propagation materials against damage caused by pine wood nematodes (Bursaphelenchus xylophilus), retransmission caused by transportation or re-harm.
4. A nematicide characterized by: the active ingredients of the nematicide contain one or more of phenolic compounds shown in the general formula I or salts thereof in claim 1.
5. The nematicide of claim 4, wherein: the nematicide is an active ingredient and is mixed with an agriculturally acceptable carrier; wherein the active ingredient accounts for 0.01-99% of the weight of the nematicide.
6. The nematicidal composition preparation is characterized in that the composition comprises an active component A and an active component B, wherein the active component A and the active component B are mixed according to the weight portion ratio of 1:100-100: 1; the active component A is one or more of phenolic compounds shown in a general formula I or salts thereof in claim 1; the active component B is one or more of other nematocides, insecticides, acaricides and bactericides.
7. The formulation of claim 4 or 6, wherein: the preparation is injection, spray, fumigant, coating agent or nanocrystallization preparation.
8. A method of protecting a plant or propagation material thereof against damage caused by nematodes, characterised in that: said method comprising applying to the plant or locus thereof the phenolic compound of formula I or one or more of its salts according to claim 1, or a formulation according to claim 4 or 6.
9. A method for preventing nematode from being transmitted and harmed again along with the transportation of infected plants is characterized in that: the method comprises applying the phenolic compound represented by the general formula I or one or more of salts thereof according to claim 1, the preparation according to claim 4 or 6 to the plant material after nematode infestation for four weeks.
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| CN113115799A (en) * | 2021-04-01 | 2021-07-16 | 沈阳恩柽研究院有限公司 | Application of turmeric volatile oil |
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| JP2001172111A (en) * | 1999-12-21 | 2001-06-26 | Sds Biotech:Kk | Method for selecting symbiotic bacterium of nematode and method for culturing nematode |
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| CN113115799A (en) * | 2021-04-01 | 2021-07-16 | 沈阳恩柽研究院有限公司 | Application of turmeric volatile oil |
| CN113115799B (en) * | 2021-04-01 | 2022-01-25 | 沈阳恩柽研究院有限公司 | Application of turmeric volatile oil |
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