A foliar adhesion-type nano-pesticide based on self-assembly technology, its preparation method and application

Abstract

The present invention discloses a foliar adhesion-type nano-pesticide based on self-assembly technology, its preparation method and application. The drug-loaded foliar adhesion-type nano-pesticide is prepared by self-assembly of a pesticide with an amino group and chlorogenic acid. In the present invention, natural chlorogenic acid molecules are used as carriers, and through a self-assembly process with pesticide molecules, nano-pesticides are prepared under low-speed stirring. It has good biocompatibility, good adhesion to the leaf surface, improves the bioavailability of pesticides, reduces the application cost, alleviates environmental pollution, and improves the economic value of pesticides.

Description

Technical Field

[0001] The present invention belongs to the technical field of pesticide formulations, and particularly relates to a foliar adhesion-type nano-pesticide based on self-assembly technology, a preparation method thereof, and an application thereof. Background Art

[0002] Pesticides play a crucial role in controlling the spread of plant diseases, pests, and weeds and increasing crop yields. However, the overuse of pesticides has led to the resistance of pests and diseases, and at the same time caused serious environmental problems, such as high residue levels, ecological imbalance, and decline in biodiversity. During agricultural application, more than about 90% of pesticide molecules will be lost or dispersed into the surrounding environment through ways such as bouncing, leaching, photodegradation, and transpiration during spraying. Pesticide molecules in the environment pose risks to human health and the survival of animals and plants, and reducing the loss pathways and improving the bioavailability can significantly improve the pesticide application efficiency. Therefore, enhancing the deposition of pesticides on crop leaves and prolonging the action time of pesticides are very important.

[0003] As a new type of pesticide formulation, nano-pesticides have great advantages in improving the bioavailability of pesticides, droplet coverage efficiency, drug deposition amount, and dose transmission efficiency due to their small size, large specific surface area, etc.

[0004] Foliar adhesion force is the key factor determining the retention ability of the pesticide sustained-release system. Increasing the foliar adhesion force of pesticides can significantly improve the retention rate of pesticides on plant leaves and improve the pesticide utilization rate. Research on traditional pesticides shows that due to characteristics such as low adhesion force and poor hydrophilicity, less than 0.1% of pesticides act on harmful biological targets. Nano-pesticides have small particle sizes and strong targeting properties, and are expected to be an important method to solve these problems.

[0005] Foliar adhesion-type nano-pesticides can interact with the micro-nano structure of the leaf surface, causing the liquid medicine to adhere to the leaf surface, avoiding the loss of the liquid medicine caused by rain washing, and increasing the retention amount of pesticides on the leaf surface. Therefore, enhancing the deposition and adhesion of nano-pesticides on the crop surface is particularly crucial for improving the effective utilization rate of pesticides, and this is also an important strategy for realizing the sustainable and green development of pesticides.

[0006] However, the existing preparation processes of foliar adhesion-type nano-pesticides have the following defects. During the actual application process, they still face many challenges, making it difficult for them to achieve mass production.

[0007] 1. The preparation process is complex. Sometimes it requires multiple chemical reactions and a large amount of toxic solvents, causing environmental pollution.

[0008] 2. In order to improve the foliar permeability and adhesion of pesticides, various auxiliaries and surfactants are often added in the aqueous phase during the preparation process, which is likely to cause adverse effects on the environment.

[0009] 3. The carrier in the preparation process is an inorganic material or a non-degradable organic material, resulting in incomplete release of pesticides, low pesticide use efficiency, and large amounts of pesticide residues. Summary of the Invention

[0010] In order to solve the above technical problems, improve the bioavailability of traditional pesticides, reduce the residual pesticides in the environment, and lower the production cost of agriculture. The present invention provides the following technical solutions.

[0011] In a first aspect, the present invention provides a foliar adhesion-type nano-pesticide based on self-assembly technology, and the drug-loaded particles of the foliar adhesion-type nano-pesticide are prepared by self-assembly of a pesticide carrying an amino group and chlorogenic acid.

[0012] Preferably, the pesticide includes, but is not limited to, any one or a combination of two or more of emamectin benzoate, spinosad, spinetoram, matrine, or veratrine.

[0013] Preferably, the molar ratio of the pesticide to the chlorogenic acid is in the range of 1:1 to 10, for example: 1:1, 1:2.3, 1:25, 1:2.8, 1:3.3, 1:3.4, 1:4.6, 1:6.9, 1:8, 1:10.

[0014] Preferably, the particle size of the nano-pesticide is 40 - 120 nm, for example: 40 nm, 50 nm, 60 nm, 70 nm, 80 nm, 90 nm, 101 nm, 104 nm, 105 nm, 108 nm, 112 nm, 115 nm, 120 nm.

[0015] Further, the particle size of the nano-pesticide is 70 - 120 nm.

[0016] In a second aspect, the present invention provides a preparation method of the foliar adhesion-type nano-pesticide described in the first aspect, which includes the following steps:

[0017] (1) Dissolve the pesticide in ethanol to obtain a pesticide ethanol solution;

[0018] (2) Dissolve chlorogenic acid in water to obtain a chlorogenic acid solution;

[0019] (3) Slowly add the pesticide ethanol solution to the chlorogenic acid solution, and obtain the foliar adhesion-type nano-pesticide through stirring, dialysis, and drying.

[0020] Preferably, in step (1), the mass percentage concentration of the pesticide ethanol solution is 2 - 10%, for example: 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%.

[0021] Preferably, in step (2), the mass percentage concentration of the chlorogenic acid solution is 4-12%, for example: 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%.

[0022] Preferably, in step (2), the pH of the chlorogenic acid solution is 6.2-6.8, for example: 6.2, 6.4, 6.5, 6.6, 6.8.

[0023] Preferably, in step (3), the rotation speed of the stirring is 400-1000 rpm, for example: 400 rpm, 500 rpm, 600 rpm, 700 rpm, 800 rpm, 900 rpm, 1000 rpm.

[0024] Preferably, in step (3), the stirring time is 2-4 h, for example 2 h, 2.5 h, 3 h, 3.5 h, 4 h.

[0025] Preferably, the temperature conditions in steps (1), (2) and (3) are 50-60 °C, for example 50 °C, 52 °C, 55 °C, 58 °C, 60 °C.

[0026] In a third aspect, the present invention provides the use of the foliar adhesion type nano-pesticide described in the first aspect in the prevention and control of crop diseases and pests.

[0027] Preferably, the diseases and pests include but are not limited to diamondback moth, aphids, Plutella xylostella or thrips, and are further preferably aphids.

[0028] Advantages of the present invention:

[0029] 1. The present invention uses chlorogenic acid and amino-group-containing pesticides as raw materials. The two molecules self-assemble into nanoparticles through electrostatic, hydrophobic and hydrogen bond interactions, forming nanoparticles with a hydrophobic core and a hydrophilic outer layer. The self-assembly process is simple, and the foliar adhesion type nano-pesticide can be prepared only under low-speed stirring.

[0030] 2. The present invention makes full use of the characteristics of chlorogenic acid as a natural adhesion molecule, reduces various auxiliaries added to improve leaf permeability and adhesion during the use of traditional pesticides, and reduces production and application costs.

[0031] 3. The foliar adhesion type nano-pesticide of the present invention can be completely degraded under natural conditions, is environmentally friendly, has low pesticide residues, high efficacy and high bioavailability, and provides a new research and development idea and technical basis for the development of new nano-pesticide formulations. Description of the Drawings

[0032] Figure 1 Shown is the hydrated particle size diagram of the foliar adhesion type emamectin benzoate nano-pesticide;

[0033] Figure 2 Shown is the scanning electron microscope image of foliar adhesion type emamectin benzoate nano - pesticide;

[0034] Figure 3 Shown is the transmission electron microscope image of foliar adhesion type emamectin benzoate nano - pesticide;

[0035] Figure 4 Shown is the sustained - release curve graph of foliar adhesion type emamectin benzoate nano - pesticide and commercially available suspension;

[0036] Figure 5 Shown is the photolysis curve graph of foliar adhesion type emamectin benzoate nano - pesticide and commercially available suspension;

[0037] Figure 6 Shown is the change graph of foliar retention rate of foliar adhesion type emamectin benzoate nano - pesticide and commercially available suspension after simulated rainwater scouring;

[0038] Figure 7 Shown is the change graph of foliar retention rate of foliar adhesion type emamectin benzoate nano - pesticide after scouring with 2.5% NaCl and 2.5% urea solution. Detailed implementation manners

[0039] The technical solutions of the present invention will be clearly and completely described below in conjunction with the embodiments and the accompanying drawings. The described embodiments are only a part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, other embodiments obtained by those of ordinary skill in the art without creative efforts fall within the scope of protection of the present invention.

[0040] It should be noted that the experimental methods used in the following embodiments are all conventional methods in the art unless otherwise specified.

[0041] Example 1 Preparation of pesticide solution

[0042] Weigh a certain amount of pesticide (emamectin benzoate, spinosad, spinetoram, matrine, veratrine) powder, and dissolve it in a certain amount of ethanol solution at 50 - 60 °C to prepare a 1.0 L pesticide solution with a mass percentage concentration of 2 - 10%.

[0043] Example 2 Preparation of chlorogenic acid solution

[0044] Weigh a certain amount of chlorogenic acid, and dissolve it in a certain amount of aqueous solution at 50 - 60 °C to prepare a 1.0 L aqueous solution with a mass percentage concentration of 4 - 12%.

[0045] Example 3 Preparation of foliar adhesion type emamectin benzoate nano - pesticide sample 1

[0046] In Example 1, 50 mL of a 5% emamectin benzoate solution was slowly added dropwise to 50 mL of a 4% chlorogenic acid aqueous solution prepared in Example 2. The molar ratio of emamectin benzoate to chlorogenic acid was 1:2.3. The temperature was maintained at 50 - 60 °C, the pH was adjusted to 6.5, and it was stirred at a speed of 500 rmp for 2 h. After filtration, dialysis, and drying, the foliar adhesion-type emamectin nano-pesticide sample 1 was obtained. Its particle size was measured by a laser particle size analyzer to be approximately 100 nm.

[0047] Example 4 Preparation of foliar adhesion-type emamectin nano-pesticide sample 2

[0048] In Example 1, 50 mL of a 5% emamectin benzoate solution was slowly added dropwise to 100 mL of a 4% chlorogenic acid solution prepared in Example 2. The molar ratio of emamectin benzoate to chlorogenic acid was 1:4.6. The temperature was maintained at 50 - 60 °C, the pH was adjusted to 6.5, and it was stirred at a speed of 500 rmp for 2 h. After filtration, dialysis, and drying, the foliar adhesion-type emamectin nano-pesticide sample 2 was obtained. Its particle size was measured by a laser particle size analyzer to be approximately 105 nm, and after dilution, samples for scanning and transmission electron microscopy were prepared.

[0049] Example 5 Preparation of foliar adhesion-type emamectin nano-pesticide sample 3

[0050] In Example 1, 50 mL of a 5% emamectin benzoate solution was slowly added dropwise to 150 mL of a 4% chlorogenic acid solution prepared in Example 2. The molar ratio of emamectin benzoate to chlorogenic acid was 1:6.9. The temperature was maintained at 50 - 60 °C, the pH was adjusted to 6.5, and it was stirred at a speed of 500 rmp for 2 h. After filtration, dialysis, and drying, the foliar adhesion-type emamectin nano-pesticide sample 3 was obtained. Its particle size was measured by a laser particle size analyzer to be approximately 112 nm.

[0051] Example 6 Preparation of foliar adhesion-type spinosad nano-pesticide sample

[0052] In Example 1, 50 mL of a 5% spinosad solution was slowly added dropwise to 100 mL of a 5% chlorogenic acid solution prepared in Example 2. The molar ratio of spinosad to chlorogenic acid was 1:3.3. The temperature was maintained at 50 - 60 °C, the pH was adjusted to 6.5, and it was stirred at a speed of 500 rmp for 2 h. After filtration, dialysis, and drying, the foliar adhesion-type spinosad nano-pesticide sample 1 was obtained. Its particle size was measured by a laser particle size analyzer to be approximately 104 nm.

[0053] Example 7 Preparation of foliar adhesion-type spinetoram nano-pesticide sample

[0054] In Example 1, 50 mL of a 5% spinetoram solution was slowly added dropwise to 100 mL of a 5% chlorogenic acid solution prepared in Example 2. The molar ratio of spinetoram to chlorogenic acid was 1:3.4. The temperature was maintained at 50 - 60 °C, the pH was adjusted to 6.5, and it was stirred at a speed of 500 rmp for 2 h. After filtration, dialysis, and drying, the foliar adhesion type spinetoram nano-pesticide sample 1 was obtained. The particle size measured by a laser particle size analyzer was about 108 nm.

[0055] Example 8 Preparation of foliar adhesion type matrine nano-pesticide sample

[0056] In Example 1, 50 mL of a 5% matrine solution was slowly added dropwise to 180 mL of a 5% chlorogenic acid solution prepared in Example 2. The molar ratio of matrine to chlorogenic acid was 1:2.5. The temperature was maintained at 50 - 60 °C, the pH was adjusted to 6.5, and it was stirred at a speed of 500 rmp for 2 h. After filtration, dialysis, and drying, the foliar adhesion type matrine nano-pesticide sample 1 was obtained. The particle size measured by a laser particle size analyzer was about 101 nm.

[0057] Example 9 Preparation of foliar adhesion type veratridine nano-pesticide sample

[0058] In Example 1, 50 mL of a 5% veratridine solution was slowly added dropwise to 120 mL of a 5% chlorogenic acid solution prepared in Example 2. The molar ratio of matrine to chlorogenic acid was 1:2.8. The temperature was maintained at 50 - 60 °C, the pH was adjusted to 6.5, and it was stirred at a speed of 500 rmp for 2 h. After filtration, dialysis, and drying, the foliar adhesion type veratridine nano-pesticide sample 1 was obtained. The particle size measured by a laser particle size analyzer was about 101 nm.

[0059] Example 10 Performance test of foliar adhesion type emamectin benzoate nano-pesticide sample 2

[0060] 10.1 Particle size and morphology

[0061] Weigh 2 mg of the foliar adhesion type emamectin benzoate nano-pesticide sample 2, dissolve it in 5.0 mL of water, sonicate for 5 min, take 1.0 mL and inject it into the dynamic light scattering sample cell, and the measured hydrodynamic diameter is 105 nm (see Figure 1 ).

[0062] Absorb 50 μL of the foliar adhesion type emamectin benzoate nano-pesticide sample 2 prepared in Example 4, add it to 5.0 mL of deionized water, sonicate and disperse it evenly, then take 5.0 μL and spot it on a silicon wafer. After natural drying, observe its morphological characteristics with a scanning electron microscope. The results are shown in Figure 2 .

[0063] Absorb 50 μL of the foliar-adhesive emamectin benzoate nano-pesticide sample 2, add it to 5.0 mL of deionized water, ultrasonically disperse it evenly, and then pipette 5.0 μL for spotting on a copper grid. After natural drying, observe its morphological characteristics with a transmission electron microscope. The results are shown in Figure 3 .

[0064] It can be seen from the results of Figure 2 and Figure 3 that the foliar-adhesive emamectin benzoate nano-pesticide is spherical in shape, with a particle size of about 100 nm.

[0065] 10.2 Sustained-release effect of the foliar-adhesive emamectin benzoate nano-pesticide sample 2

[0066] Place 5 mL of the foliar-adhesive emamectin benzoate nano-pesticide sample 2 solution (pesticide content: 5 mg / mL) in a dialysis bag (MWCO: 3500 Da), and immerse it in 10 mL of 30% acetonitrile aqueous solution. Shake and release at 25 °C for 48 h. Pipette 1 mL of the release medium at 0 h, 0.5 h, 1 h, 2 h, 3 h, 5 h, 7 h, 9 h, 11 h, 15 h, 19 h, 24 h, 30 h, 36 h, 42 h, and 48 h respectively, and quickly supplement the corresponding volume of the release medium to the dialysis system. Use high-performance liquid chromatography to measure the concentration of emamectin benzoate in the samples taken at each time point. At the same time, use the emamectin benzoate suspending agent solution (content: 5 mg / mL) as a control, and calculate the comparison chart of the cumulative release curves of the two. The results are shown in Figure 4 .

[0067] As Figure 4 shown, the release rate of the foliar-adhesive emamectin benzoate nano-pesticide sample 2 gradually slows down over time, following a slow diffusion pattern. The cumulative release at 48 h is about 62.3%, showing good sustained-release performance. While the cumulative release rate of the control suspending agent sample exceeds 98.4% within 24 h, and its sustained-release performance is poor. This shows that the foliar-adhesive emamectin benzoate nano-pesticide of the present invention has good long-lasting performance.

[0068] 10.3 Photodegradation experiment

[0069] Respectively take 50 mL samples of the emamectin benzoate nano-pesticide sample 2 and the commercially available emamectin benzoate suspending agent (pesticide content: 25 mg / mL), place them in a xenon arc lamp illumination incubator at 25 °C and 250 W for continuous illumination. Take 2.0 mL samples at 0 h, 12 h, 24 h, 36 h, and 48 h respectively, completely dissolve them in methanol, ultrasonicate for 15 min to completely release the pesticide, and use a high-performance liquid chromatograph to measure the content of the original drug in the filtrate obtained after filtration. The results are shown inFigure 5 。

[0070] From Figure 5 It can be seen that after 48 h of light exposure, less than 40% of emamectin benzoate nano-pesticide was degraded, while more than 80% of the commercially available suspending agent was degraded. This indicates that the nano-pesticide formed by self-assembly can significantly reduce the photodegradation rate of emamectin benzoate and improve its photo-stability.

[0071] 10.4 Evaluation of the adhesion performance of emamectin benzoate nano-pesticide sample 2 for foliar adhesion

[0072] Take 500 μL of emamectin benzoate nano-pesticide sample 2 for foliar adhesion and emamectin benzoate suspending agent solution (content: 5 mg / mL), and spray them evenly on cucumber leaves with a high-pressure spray gun. After natural drying, carefully divide the leaf surface into two parts. One part is rinsed with 50 mL of deionized water to simulate rain resistance, and then these parts are quickly frozen in liquid nitrogen, ground into powder, and emamectin benzoate is extracted with 5.0 mL of acetonitrile. The content of emamectin benzoate is quantified by high performance liquid chromatography (HPLC), and the retention rate of emamectin benzoate is determined by comparing the changes before and after rinsing, so as to evaluate the foliar adhesion performance. The results are shown in Figure 6 。

[0073] In order to study the foliar adhesion mechanism of emamectin benzoate nano-pesticide sample, the other part of the leaves is rinsed with 2.5% NaCl and 2.5% urea solution instead of deionized water to destroy the electrostatic interaction or hydrogen bond respectively. The results are shown in Figure 7 。

[0074] From Figure 6 It can be seen that the emamectin benzoate nano-pesticide for foliar adhesion has strong rain resistance. After simulated rain washing, the retention rate of the pesticide on the leaves exceeds 80%, while for the suspending agent sample as a control, the retention rate after washing is less than 30%. These results indicate that after introducing the adhesive molecule chlorogenic acid by self-assembly, the foliar adhesion of emamectin benzoate nano-pesticide is significantly enhanced.

[0075] As Figure 7 shown, electrostatic and hydrogen bond interactions mainly participate in the adhesion mechanism between the nano-pesticide and the leaf surface.

[0076] 10.5 Indoor toxicity determination against Aphis gossypii

[0077] The indoor toxicity of foliar adhesion type emamectin nano-pesticide and commercially available emamectin benzoate suspension agent to Aphis gossypii on cucumber leaves was compared by spray method. The foliar adhesion type emamectin nano-pesticide, commercially available suspension agent and emulsifiable concentrate were formulated into 7 series of concentration gradients of medicament concentrations (ppm): 500, 250, 125, 67.5, 34, 17, 8.5 for standby. The cucumber leaves that had not been exposed to the medicament were cut into round pieces with a diameter of 7.5 cm, sprayed by a spray tower, taken out and air-dried naturally indoors. About 2 mm of agar solution was poured into a petri dish. After cooling, the back of the leaf was attached to the inside of the petri dish. Cucumber seedlings with a large number of insects were collected in the field, and wingless adult aphids were picked up with a writing brush and placed on the leaves, 30 aphids per dish. It was sealed with plastic wrap, and 15 - 20 small holes were pricked on the film with an insect needle. The experiment was set with 4 replicates. The treated test insects were placed in an insect rearing room for normal feeding. The feeding conditions were L / D = 14:10, T = (25 ± 2) °C, RH = (75 ± 5)%. Three biological replicates were established. After 48 h, the number of dead and live insects was checked, the mortality rate was calculated, and LC was calculated using SPSS software. 50 , and the results are shown in Table 1.

[0078] Table 1 Indoor toxicity test results of emamectin nano-pesticide and commercially available suspension agent

[0079]

[0080] It can be seen from the results in Table 1 that the toxicity of the foliar adhesion type emamectin nano-pesticide prepared by the method of the present invention to aphids is stronger than that of the commercially available suspension agent.

[0081] 10.6 Zebrafish toxicity determination

[0082] The static method was used to evaluate the toxicity of foliar adhesion type emamectin nano-pesticide sample 2 to zebrafish, and the commercially available emamectin benzoate suspension agent was used as the control group. The two pesticide preparations were diluted with deionized water into liquid medicines with different concentrations. 20 zebrafish (with a length of 2 - 3 cm) were placed in a container with a certain volume. Deionized water was used as the blank control. Three biological replicates were set. The mortality rate of zebrafish after 96 h in the liquid medicine was recorded, and LC was calculated using SPSS software. 50 , and the results are shown in Table 2.

[0083] Table 2 Toxicity test results of emamectin nano-pesticide and commercially available emulsifiable concentrate to zebrafish

[0084]

[0085] It can be seen from Table 2 that after 96 hours of pesticide incubation, the LC of emamectin nano-pesticide and commercially available suspension agent to zebrafish 50They are 2.08 μg / mL and 1.14 μg / mL respectively, indicating that the toxicity of emamectin benzoate is significantly reduced after self-assembly, the safety for non-target organisms is significantly improved, and the environmental friendliness is enhanced.

[0086] The above are only the embodiments of the present invention, and do not limit the patent scope of the present invention accordingly. Any equivalent structural transformation made by using the content of the specification of the present invention, or directly or indirectly applied in other related technical fields, shall be included in the patent protection scope of the present invention by the same token.

Claims

1. A foliar adhesion type nano-pesticide, characterized in that, The foliar adhesion type nano-pesticide is prepared from emamectin benzoate and chlorogenic acid by the following method; The molar ratio of the emamectin benzoate to the chlorogenic acid is 1:1 to 10; The method comprises the following steps: (1) Dissolve emamectin benzoate in ethanol to obtain a pesticide ethanol solution; (2) Dissolve chlorogenic acid in water to obtain a chlorogenic acid solution with a mass percentage concentration of 4% to 12%; (3) Slowly add the pesticide ethanol solution to the chlorogenic acid solution, adjust the pH to 6.2 to 6.8, then stir, dialyze, and dry to obtain the foliar adhesion type nano-pesticide.

2. The foliar adhesion type nano-pesticide according to claim 1, wherein The particle size of the nano-pesticide is 40 to 120 nm.

3. The preparation method of the foliar adhesion-type nano-pesticide according to claim 1, characterized in that, Comprises the following steps: (1) Dissolve emamectin benzoate in ethanol to obtain a pesticide ethanol solution; (2) Dissolve chlorogenic acid in water to obtain a chlorogenic acid solution with a mass percentage concentration of 4% to 12%; (3) Slowly add the pesticide ethanol solution to the chlorogenic acid solution, adjust the pH to 6.2 to 6.8, then stir, dialyze, and dry to obtain the foliar adhesion type nano-pesticide.

4. The preparation method according to claim 3, characterized in that, In step (1), the mass percentage concentration of the pesticide ethanol solution is 2% to 10%.

5. The preparation method according to claim 3, wherein In step (3), the rotation speed of the stirring is 400 to 1000 rpm, and the time is 2 to 4 h.

6. The preparation method according to claim 3, characterized in that, The temperature conditions in steps (1), (2), and (3) are 50 to 60 °C.

7. Use of the foliar adhesion type nano-pesticide according to any one of claims 1 to 2 in the prevention and control of crop diseases and pests.

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