Multi-response type nano-pesticide for preventing and treating root-knot nematode and preparation method of multi-response type nano-pesticide

Abstract

The invention provides a multi-response type nano-pesticide for preventing and treating root-knot nematode and a preparation method of the multi-response type nano-pesticide. The pesticide composition is prepared from the following raw materials in parts by weight: 2.5%-10% of an abamectin pesticide, 5%-20% of a phase change material, 23%-35% of a stabilizer and 45%-62% of zein. The nano pesticide capable of realizing three response controlled release functions on temperature, pH and protease is constructed by using an environment-friendly carrier and a simple preparation process, and compared with a traditional missible oil preparation, the nano pesticide shows a better control effect on root-knot nematode and has lower toxicity on non-target organisms, namely zebra fish and earthworm. And the obtained preparation is efficient, safe and environment-friendly.

Description

Technical Field

[0001] This invention belongs to the field of pesticides, specifically relating to a multi-responsive nanopesticide for controlling root-knot nematodes and its preparation method. Background Technology

[0002] Root-knot nematodes are important plant parasitic nematodes globally, characterized by their wide distribution, diverse host species, and difficulty in control, posing a significant challenge to pest and disease management. Root-knot nematode infestation not only leads to a substantial reduction in crop yield and quality but also makes plants more susceptible to bacterial and fungal infections. Although various methods exist for controlling root-knot nematodes, including agricultural, physical, and biological control, these methods often fail to achieve significant results in actual agricultural production, resulting in relatively low economic benefits and hindering widespread application. Against this backdrop, chemical control, particularly abamectin B2 family pesticides, has gained widespread attention and favor in the industry due to its superior control efficacy. However, the main formulation of abamectin B2 is currently emulsifiable concentrate, and the large amounts of organic solvents and adjuvants contained in these formulations can lead to significant environmental and ecological problems. Furthermore, the high adsorption capacity of this formulation in soil significantly reduces the actual utilization rate of the pesticide's active ingredient, forcing farmers to increase the dosage in pursuit of control efficacy. This practice not only exacerbates the waste of pesticide resources but also promotes the development of pesticide resistance in pests, further worsening soil pollution and posing a long-term threat to sustainable agricultural development. Therefore, exploring more environmentally friendly, efficient, and economically viable alternative formulations is of great significance for optimizing root-knot nematode control strategies and promoting green agricultural development.

[0003] The application of nanotechnology in agriculture has provided new avenues for the creation of novel formulations. Nanospheres are a typical type of carrier-encapsulated nano-formulation, encapsulating pesticide active ingredients within a spherical carrier. On one hand, they can effectively mitigate pesticide degradation and loss caused by environmental factors such as light, heat, rain, soil, microorganisms, and other chemicals, thereby improving formulation stability and reducing volatility, odor, and contact toxicity. On the other hand, utilizing the small size, large surface area, and modifiability of nanomaterials, controlled release and environmentally responsive intelligent release can be achieved, thereby reducing the number of applications and application costs, and improving the utilization rate of active ingredients.

[0004] Zeatin is an ideal carrier for drug encapsulation due to its safety, environmental friendliness, and self-assembly properties. However, there are few reports on environmentally responsive zeatin nanopesticide systems with high targeting and high utilization efficiency. The few reports that exist mainly use chemical synthesis methods such as grafting and modification to achieve a response to a single external stimulus. The process is complex, cumbersome, and costly, making it unsuitable for large-scale production and application, and it does not have the controlled release function of multiple responses.

[0005] Using zein as a carrier and employing a simple and low-cost physical encapsulation method to replace chemical synthesis, and introducing a stimulus-responsive "switch," constructing multi-release abamectin-based nanopesticides is a scientific and technological approach to improve pesticide utilization efficiency, extend the duration of action, and enhance the control of root-knot nematodes. The development of related preparation technologies and processes is also an urgent problem to be solved by those skilled in the art. Summary of the Invention

[0006] To overcome the shortcomings of traditional pesticide formulations, such as low utilization rate, environmentally unfriendly composition, poor controlled-release performance, and short duration of effect, this invention provides a multi-responsive nanopesticide, its preparation method, and its application in controlling root-knot nematodes. The nanopesticide of this invention uses zein as a carrier and achieves controlled-release functions in response to temperature, pH, and protease through the physical encapsulation of phase change materials and avermectin-like drugs. This allows the formulation to intelligently regulate the pesticide release rate based on the population occurrence patterns, living environment, and physiological characteristics of root-knot nematodes, thereby improving the pesticide's targeting, effectiveness, and duration of effect.

[0007] The multi-responsive nanopesticide provided by this invention is made from raw materials comprising the following components in parts by weight:

[0008]

[0009] The abamectin pesticides include abamectin B1 and abamectin B2.

[0010] The phase change material is at least one of octadecane, butyl stearate, dodecylamine, and dodecyl alcohol, specifically butyl stearate, dodecyl alcohol, and dodecylamine.

[0011] The stabilizer is at least one of sodium caseinate, sodium alginate, and chitosan, specifically sodium caseinate or sodium alginate.

[0012] The aforementioned multi-responsive nanopesticide possesses controlled-release functions in response to temperature, pH, and protease. This multi-responsive nanopesticide is prepared via a method comprising the following steps:

[0013] 1) Dissolve the stabilizer in water to obtain an aqueous phase;

[0014] 2) Dissolve zein in a 90% (volume concentration) aqueous ethanol solution, add abamectin-type pesticides and phase change materials, and obtain an oil phase after dissolution;

[0015] 3) Under high-speed stirring conditions, the oil phase is added dropwise to the aqueous phase to obtain a mixed solution;

[0016] 4) The mixed solution was heated to 35℃-50℃ and the ethanol was removed by evaporation under vacuum to obtain a zein nano-suspension loaded with abamectin pesticides.

[0017] 5) Remove the water from the nano suspension to obtain zein nanoparticle solid powder loaded with abamectin pesticide.

[0018] Furthermore, in the aqueous phase of step 1), the concentration of the stabilizer is 2 mg / mL to 15 mg / mL, preferably 7.5 mg / mL.

[0019] Furthermore, in the oil phase of step 2), the concentration of zein is 10-40 mg / mL, preferably 30 mg / mL; the concentration of abamectin pesticide is 1 mg / mL-6 mg / mL, preferably 1.3 mg / mL; and the concentration of phase change material is 2.5 mg / mL-12 mg / mL, preferably 4.5 mg / mL.

[0020] The effective particle size of the obtained zein nanoparticles loaded with abamectin pesticides is 50-500 nm.

[0021] The application of the aforementioned multi-responsive nanopesticides in the prevention and control of plant diseases and pests also falls within the scope of protection of this invention.

[0022] The pests and diseases mentioned include: root-knot nematodes, grubs, cutworms, mole crickets, root maggots, and root aphids.

[0023] Experimental results show that the multi-responsive nano-pesticide of this invention exhibits better control effect against root-knot nematodes compared with traditional emulsifiable concentrate formulations, while having lower toxicity to non-target organisms such as zebrafish and earthworms.

[0024] This invention utilizes an environmentally friendly carrier and a simple preparation process to construct a nanopesticide capable of controlled-release in response to temperature, pH, and protease. Compared to traditional emulsifiable concentrate formulations, it exhibits better control efficacy against root-knot nematodes, while also showing lower toxicity to non-target organisms such as zebrafish and earthworms. The resulting formulation is highly efficient, safe, and environmentally friendly.

[0025] Compared with existing technologies, the advantages of this invention are as follows: Based on the temperature range (25-30℃) that facilitates the outbreak and reproduction of root-knot nematodes, this invention selects a phase change material with a phase change temperature within this range as a gating "switch." This allows the active ingredient to be released with little or no pesticide release when root-knot nematodes are few (i.e., under low-temperature conditions), thus protecting the effective ingredient from premature consumption. Conversely, when the root-knot nematode population density is high (in the temperature environment of outbreaks), the active substance is released rapidly, achieving effective pest control. Furthermore, root-knot nematodes easily migrate to weakly acidic and weakly alkaline soils, and the nano-delivery system in this invention readily disintegrates under acidic and alkaline conditions, rapidly releasing the active ingredient. Additionally, the proteases produced in the root-knot nematode's intestines accelerate the hydrolysis of the nano-pesticide carrier, enabling the active ingredient to quickly and efficiently kill the target pest. In summary, the nano-pesticide formulation of this invention can respond to environmental factors according to the characteristics of root-knot nematodes, thereby achieving on-demand release of the active ingredient, improving pesticide utilization, and reducing the dosage of nematicides. Attached Figure Description

[0026] Figure 1 This is an electron microscope image of the nanopesticide prepared in Example 1 of the present invention.

[0027] Figure 2 The release curves of the nano-pesticide prepared in Example 1 of this invention at different temperatures are shown.

[0028] Figure 3 The release curves of the nano-pesticide prepared in Example 1 of this invention at different pH values ​​are shown.

[0029] Figure 4 The release curves of the nano-pesticide prepared in Example 1 of this invention are shown in the presence and absence of protease.

[0030] Figure 5 The toxicity of commercially available emulsifiable concentrate formulations and the nano-pesticide prepared in Example 1 of this invention to root-knot nematodes.

[0031] Figure 6 The toxicity of commercially available emulsifiable concentrate formulations and the nano-pesticide prepared in Example 1 of this invention to earthworms. Detailed Implementation

[0032] The present invention will now be described in further detail with reference to specific embodiments. The given embodiments are merely illustrative of the invention and not intended to limit its scope. The embodiments provided below can serve as a guide for further improvements by those skilled in the art and do not constitute a limitation on the invention in any way.

[0033] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, performed according to the techniques or conditions described in the literature in this field or according to the product instructions. Unless otherwise specified, the materials and reagents used in the following examples are commercially available.

[0034] Example 1

[0035] Multi-responsive nanopesticides were prepared using an antisolvent precipitation method. The specific preparation steps are as follows:

[0036] 1) Dissolve 0.3g of sodium caseinate in water to obtain an aqueous phase with a sodium caseinate concentration of 7.5mg / mL.

[0037] 2) Dissolve 0.6g of zein in a 90% (v / v) aqueous ethanol solution, and add 0.026g of abamectin B2 and 0.09g of butyl stearate to obtain the oil phase (the concentration of zein is 30mg / mL; the concentration of abamectin B2 is 1.3mg / mL; and the concentration of butyl stearate is 4.5mg / mL).

[0038] 3) The oil phase is added dropwise to the aqueous phase under high-speed stirring to obtain a mixed solution;

[0039] 4) The mixed solution was heated to 45°C and the organic solvent was removed by evaporation under vacuum to obtain zein nano suspension;

[0040] 5) The nano suspension was freeze-dried to obtain zein nanopesticide loaded with abamectin B2.

[0041] Figure 1 Electron micrograph of the prepared nanopesticide. Figure 1 It is known that the particle size of nano-pesticides is 234 nm.

[0042] Example 2

[0043] 1) Dissolve 0.36g of sodium caseinate in water to obtain an aqueous phase with a sodium caseinate concentration of 9mg / mL.

[0044] 2) Dissolve 0.54g of zein in a 90% aqueous ethanol solution, and add 0.12g of abamectin B1 and 0.18g of dodecanol to obtain the oil phase (the concentration of zein is 27mg / mL; the concentration of abamectin B1 is 6mg / mL; and the concentration of dodecanol is 9mg / mL).

[0045] 3) The oil phase is added dropwise to the aqueous phase under high-speed stirring to obtain a mixed solution;

[0046] 4) The mixed solution was heated to 45°C and the organic solvent was removed by evaporation under vacuum to obtain zein nano suspension;

[0047] 5) The nano suspension was freeze-dried to obtain zein nanopesticide loaded with abamectin B1.

[0048] Example 3

[0049] 1) Dissolve 0.28g of sodium alginate in water to obtain an aqueous phase, the concentration of sodium alginate in the aqueous phase is 7mg / mL;

[0050] 2) Dissolve 0.74g of zein in a 90% aqueous ethanol solution, and add 0.12g of abamectin B2 and 0.06g of butyl stearate to obtain the oil phase (the concentration of zein is 37mg / mL; the concentration of abamectin B2 is 6mg / mL; and the concentration of butyl stearate is 3mg / mL).

[0051] 3) The oil phase is added dropwise to the aqueous phase under high-speed stirring to obtain a mixed solution;

[0052] 4) The mixed solution was heated to 45°C and the organic solvent was removed by evaporation under vacuum to obtain zein nano suspension;

[0053] 5) The nano suspension was freeze-dried to obtain zein nanopesticide loaded with abamectin B2.

[0054] Example 4

[0055] 1) Dissolve 0.39g of sodium caseinate in water to obtain an aqueous phase with a sodium caseinate concentration of 9.8mg / mL.

[0056] 2) Dissolve 0.54g of zein in a 90% aqueous ethanol solution, and add 0.03g of abamectin B1 and 0.24g of dodecylamine to obtain the oil phase (the concentration of zein is 27mg / mL; the concentration of abamectin B1 is 1.5mg / mL; and the concentration of dodecylamine is 12mg / mL).

[0057] 3) The oil phase is added dropwise to the aqueous phase under high-speed stirring to obtain a mixed solution;

[0058] 4) The mixed solution was heated to 45°C and the organic solvent was removed by evaporation under vacuum to obtain zein nano suspension;

[0059] 5) The nano suspension was freeze-dried to obtain zein nanopesticide loaded with abamectin B1.

[0060] Example 5: Temperature Response Release Test

[0061] The nano-pesticide (0.2 g) prepared in Example 1 was placed in a dialysis bag and released at 15°C, 25°C, and 35°C using 100 mL of 30% (v / v) ethanol aqueous solution as the release medium. At specified time intervals, 1 mL of the sustained-release solution was collected and replenished with an equal amount of fresh sustained-release medium. The pesticide content in the sustained-release solution was determined using high-performance liquid chromatography (HPLC), the cumulative release rate was calculated, and release curves of the nano-pesticide at different temperatures were plotted.

[0062] Depend on Figure 2 It is evident that, after 24 hours of release, the cumulative release rate of the nano-pesticide at 35°C is nearly 60% higher than that at 15°C. This result indicates that the nano-pesticide prepared in this invention has significant temperature-responsive characteristics; the higher the temperature, the faster the drug release rate. This characteristic allows the agent to intelligently respond to ambient temperature and release active ingredients, thereby achieving better control effects during root-knot nematode outbreaks.

[0063] Example 6: pH Response Release Test

[0064] The nano-pesticide (0.2 g) prepared in Example 1 was placed in a dialysis bag and released using 100 mL of 30% (v / v) ethanol aqueous solution at pH 5, 7, and 9 as the release medium. At specified time intervals, 1 mL of the released solution was collected, and an equal amount of fresh slow-release medium was added. The pesticide content in the slow-release solution was determined by high-performance liquid chromatography (HPLC), and release curves at different pH values ​​were plotted.

[0065] Depend on Figure 3 It is evident that the release rate of the nano-pesticide under both weakly acidic and weakly alkaline conditions is higher than that under neutral conditions. After 120 hours of continuous release, the cumulative release rates at pH 5 and 9 were 93.4% and 93.9%, respectively, while the cumulative release rate at pH 7 was 71.1%. These results indicate that the nano-pesticide prepared in this invention exhibits release characteristics responsive to both weakly acidic and weakly alkaline conditions.

[0066] Example 7: Enzyme Response Release Test

[0067] The nano-pesticide (0.2 g) prepared in Example 1 was incubated with 5 mg of protease (200 U / mg) for 24 h, and then placed in a dialysis bag. It was released using 30% (v / v) ethanol aqueous solution as the release medium. At specified time intervals, 1 mL of the release solution was collected and an equal amount of fresh sustained-release medium was added. The pesticide content in the sustained-release solution was determined by high performance liquid chromatography, and the release curve under the condition of no added protease was plotted.

[0068] Depend on Figure 4It is evident that the release rate of the active ingredient in the nanopesticide is faster and the cumulative release rate is higher when protease is added. After 120 hours, the cumulative release rate was 96.6%, while the cumulative release rate without protease addition was only 68.2%. This result demonstrates that the nanopesticide prepared in this invention exhibits significant enzyme-responsive characteristics.

[0069] The nano-pesticide and control emulsifiable concentrate prepared in Example 1 were diluted with pure water to form aqueous dispersions of different concentrations. The pesticide dispersions were then transferred to 24-well culture plates, and a suspension containing 50 second-instar root-knot nematodes was added to each well. The plates were then placed in incubators at 15°C, 25°C, and 35°C for culture. Pure water was used as a control to observe the survival of the root-knot nematodes.

[0070] Figure 5 The toxicity of commercially available emulsifiable concentrate formulations (Xingbakexian) and nano-pesticides prepared in Example 1 of this invention to root-knot nematodes.

[0071] Depend on Figure 5 It can be seen that the control effect of the nano-pesticide prepared in Example 1 against root-knot nematodes increases with increasing temperature. The nematicidal activity at 35℃ is 5.5 times that at 15℃, and 2.3 times that of the emulsifiable concentrate formulation. These results indicate that the release characteristics of the nano-pesticide prepared in this invention at different temperatures are consistent with the population growth trend of root-knot nematodes, which is beneficial for achieving precise pest control, improving pesticide utilization, reducing dosage, and delaying pesticide resistance development.

[0072] The nano-pesticide prepared in Example 1 and the control emulsifiable concentrate were added to the soil and mixed thoroughly. The soil moisture was then adjusted to 60% water holding capacity using deionized water. The soil containing the pesticide was transferred to 1L beakers, and 10 earthworms were added to each beaker. The beakers were then cultured in a climate chamber (20°C, 70%-90% humidity). Earthworm survival was monitored and recorded on days 7 and 14.

[0073] Figure 6 The toxicity of nano-pesticides to earthworms was prepared using commercially available emulsifiable concentrate formulations (Xingbake line) and in Example 1 of this invention.

[0074] Depend on Figure 6 It was observed that the survival rate of earthworms decreased with increasing pesticide concentration and time in both treatment groups. By day 14, the emulsifiable concentrate formulation showed 3.2 times the toxicity to earthworms compared to the nano-pesticide prepared in Example 1, indicating that the nano-formulation has higher biocompatibility with earthworms, which are not the target organism. This is because the encapsulation effect of the carrier in the nano-formulation reduces the direct contact between earthworms and the active ingredients of the pesticide, thereby reducing toxicity.

[0075] The present invention has been described in detail above. Those skilled in the art will recognize that the invention can be practiced in a wide range of ways with equivalent parameters, concentrations, and conditions without departing from its spirit and scope, and without requiring unnecessary experiments. While specific embodiments have been provided, it should be understood that further modifications can be made to the invention. In summary, according to the principles of the invention, this application is intended to include any changes, uses, or improvements to the invention, including changes made using conventional techniques known in the art that depart from the scope disclosed herein.

Claims

1. A multi-responsive nanopesticide, made from raw materials comprising the following components in parts by weight:

2. The nano-pesticide according to claim 1, characterized in that, The pesticides mentioned are abamectin pesticides, including abamectin B1 and abamectin B2; The phase change material is at least one of octadecane, butyl stearate, dodecylamine, and dodecanol; The stabilizer is at least one of sodium caseinate, sodium alginate, and chitosan.

3. The nano-pesticide according to claim 1, characterized in that, The multi-responsive nanopesticide has three controlled-release functions: temperature, pH, and protease.

4. A method for preparing the multi-responsive nanopesticide according to any one of claims 1-3, comprising the following steps: 1) Dissolve the stabilizer in water to obtain an aqueous phase; 2) Dissolve zein in a 90% (volume concentration) aqueous ethanol solution, add abamectin-type pesticides and phase change materials, and obtain an oil phase after dissolution; 3) Under high-speed stirring conditions, the oil phase is added dropwise to the aqueous phase to obtain a mixed solution; 4) The mixed solution was heated to 35℃-50℃ and the ethanol was removed by evaporation under vacuum to obtain a zein nano-suspension loaded with abamectin pesticides. 5) Remove the water from the nano suspension to obtain zein nanoparticle solid powder loaded with abamectin pesticide.

5. The method according to claim 4, characterized in that, In step 1), the concentration of the stabilizer in the aqueous phase is 2 mg / mL to 15 mg / mL.

6. The method according to claim 4, characterized in that, In step 2), the concentration of zein in the oil phase is 10 mg / mL to 40 mg / mL; the concentration of abamectin pesticide is 1 mg / mL to 6 mg / mL; and the concentration of phase change material is 2.5 mg / mL to 12 mg / mL.

7. The method according to claim 4, characterized in that, The effective particle size of the obtained zein nanoparticles loaded with abamectin pesticides is 50-500 nm.

8. The multi-responsive nanopesticide according to any one of claims 1-3 is used for the prevention and control of plant diseases and pests.

9. The application according to claim 8, characterized in that, The pests and diseases mentioned include: root-knot nematodes, grubs, cutworms, mole crickets, root maggots, and root aphids.

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