PH-responsive hexaconazole controlled-release nanoparticles as well as preparation method and application thereof

Abstract

The invention discloses pH response type hexaconazole controlled-release nanoparticles as well as a preparation method and application thereof, and belongs to the technical field of agricultural pesticide preparations and nano materials. The problems that existing hexaconazole is poor in water solubility, fast in photolysis, low in leaf surface adhesion rate, insufficient in utilization rate and the like are solved. According to the invention, a molybdenum-based polyacid nanocage (MO) is used as a carrier, and a bactericide hexaconazole (Hex) is loaded by an impregnation method, so that the HexatMO nanoparticles are prepared. The prepared HexatMO nano-particles are small in particle size and good in dispersity, can significantly improve the water solubility and thermal stability of Hex, have excellent leaf surface spreadability, are beneficial to reduction of pesticide application drift loss, have unique pH response release characteristics, can intelligently release drugs in an acidic microenvironment generated by pathogen infection, realize targeted delivery, and have broad application prospects. Therefore, the sterilization efficiency is improved, the sterilization activity is enhanced, the influence on non-target organisms is reduced, and the environmental risk is reduced.

Description

Technical Field

[0001] This invention relates to a pH-responsive hexaconazole controlled-release nanoparticle, its preparation method, and its application, belonging to the field of agricultural pesticide formulation and nanomaterials technology. Background Technology

[0002] Rice sheath blight is caused by Rhizoctonia solani (… Rhizoctonia solani This fungal disease, caused by [unspecified fungus], seriously threatens rice yield and food security. Currently, control mainly relies on chemical fungicides, such as hexazole (Hex), but in practical applications, it suffers from poor water solubility, rapid photodegradation, low leaf adhesion, and insufficient utilization, leading to frequent application, environmental pollution, and high pesticide residue risks.

[0003] Nanoparticle-based drug delivery technology offers a new approach to improving pesticide utilization and achieving intelligent controlled release. Polyoxometalates (polyacids) are a class of inorganic metal-oxygen clusters composed of pre-transition metals, possessing characteristics such as tunable structure, large specific surface area, good biocompatibility, and excellent thermal properties, making them ideal pesticide carrier materials. However, current research on the application of polyoxometalate nanocages in the delivery of hexaconazole pesticides is limited, especially regarding intelligent controlled release systems for rice sheath blight, which have yet to be systematically reported. Summary of the Invention

[0004] To address the problems of poor water solubility, rapid photolysis, low leaf adhesion rate, and insufficient utilization of existing hexaconazole, this invention provides a pH-responsive hexaconazole controlled-release nanoparticle, its preparation method, and its application.

[0005] The technical solution of this invention: One objective of this invention is to provide a pH-responsive hexaconazole controlled-release nanoparticle, which comprises a polyacid nanocage carrier and hexaconazole loaded by an impregnation method.

[0006] The second objective of this invention is to provide a method for preparing the above-mentioned pH-responsive hexaconazole controlled-release nanoparticles, the method comprising the following steps: (1) Add ammonium acetate and hydrazine sulfate to ammonium molybdate solution, stir evenly, slowly add aqueous acetic acid solution, let stand at room temperature, filter, wash, dry, and obtain polyacid nanocage carrier; (2) Hexaconazole was dissolved in methanol, polyacid nanocage carrier was dissolved in water, and then the methanol solution of hexaconazole was slowly added dropwise to the polyacid nanocage carrier solution. The mixture was stirred at room temperature, centrifuged, washed, and dried to obtain pH-responsive hexaconazole controlled-release nanoparticles, which were named Hex@MO nanoparticles.

[0007] Further specifying, the molar ratio of ammonium molybdate, ammonium acetate and hydrazine sulfate in (1) is 1:36:1.36.

[0008] Further specifying, the volume concentration of the acetic acid aqueous solution in (1) is 50%.

[0009] Further specifying, the settling time in (1) is 7 days.

[0010] Further specified, in (1) washing is performed using 90% ethanol and anhydrous ethanol in sequence.

[0011] Further specifying, the stirring time at room temperature in (2) is 24h.

[0012] Further specifying, (2) uses methanol for washing.

[0013] Further specifying, in (2), drying is natural drying at room temperature.

[0014] The third objective of this invention is to provide an application of the above-mentioned pH-responsive hexaconazole controlled-release nanoparticles, specifically as a controlled-release nanopesticide for the prevention and control of rice sheath blight.

[0015] Beneficial effects: This invention utilizes molybdenum-based polyoxocyanate nanocages (MO) as a carrier to load the fungicide hexazole (Hex) using in-situ encapsulation technology, thus preparing Hex@MO nanoparticles. The prepared Hex@MO nanoparticles have small particle size and good dispersibility, significantly improving the water solubility and thermal stability of Hex, and exhibit excellent leaf spreadability, which helps reduce application drift loss. They also possess unique pH-responsive release characteristics, enabling intelligent drug release targeting the acidic microenvironment created by pathogen infection, achieving targeted delivery, thereby improving bactericidal efficiency, enhancing bactericidal activity, reducing the impact on non-target organisms, and lowering environmental risks.

[0016] Furthermore, the carrier MO is safe for rice and can be bidirectionally transported within the plant and pathogens, further improving the bioavailability of pesticides. This provides a new approach for developing green, efficient, and intelligent nanopesticides, meeting the needs of sustainable agricultural development. Attached Figure Description

[0017] Figure 1 This is a schematic diagram illustrating the synthesis of Hex@MO nanoparticles according to the present invention; Figure 2 Transmission electron microscopy (TEM) image of Hex@MO nanoparticles; Figure 3 A comparison of the infrared spectra (FTIR) of Hex, MO, and Hex@MO; Figure 4 A comparison chart of thermogravimetric analysis (TGA) curves for Hex, MO, and Hex@MO; Figure 5 X-ray diffraction (XRD) patterns of Hex, MO, and Hex@MO; Figure 6 The in vitro cumulative release curves of Hex@MO in different pH buffer media; Figure 7 Hex@MO and Hex commercial suspension (SC) for the treatment of rice sheath blight pathogen ( R. solani Comparison of in vitro antibacterial effects of ) Figure 8 A comparison of the rain resistance and retention capacity of Hex@MO and Hex commercial suspending agent (SC) on rice leaves. Detailed Implementation

[0018] To make the above-mentioned objects, features and advantages of the present invention more apparent and understandable, the specific embodiments of the present invention will be described in detail below with reference to the examples in the specification.

[0019] Many specific details are set forth in the following description in order to provide a full understanding of the invention. However, the invention may also be practiced in other ways different from those described herein, and those skilled in the art may make similar extensions without departing from the spirit of the invention. Therefore, the invention is not limited to the specific embodiments disclosed below.

[0020] Secondly, the term "one embodiment" or "embodiment" as used herein refers to a specific feature, structure, or characteristic that may be included in at least one implementation of the present invention. The phrase "in one embodiment" appearing in different places in this specification does not necessarily refer to the same embodiment, nor is it a single or selective embodiment that is mutually exclusive with other embodiments.

[0021] Unless otherwise specified, the experimental methods used in the following examples are conventional methods. Unless otherwise specified, the materials, reagents, methods, and instruments used are all conventional materials, reagents, methods, and instruments in the art, and can be obtained commercially by those skilled in the art.

[0022] Example 1: The process for preparing Hex@MO nanoparticles is as follows: Figure 1 As shown, it includes the following steps: (1) Preparation of MO nanocages: 5.6 g (4.5 mmol) of ammonium molybdate was dissolved in 250 mL of distilled water. Then, 12.5 g (162.2 mmol) of ammonium acetate and 0.8 g (6.1 mmol) of hydrazine sulfate were added sequentially, and the mixture was stirred for 10 min until the solution turned blue-green. 100 mL of a 50% (v / v) aqueous acetic acid solution was slowly added dropwise to the above solution. The mixture was stored at room temperature for 7 days, during which reddish-brown crystals precipitated on the inner wall of the beaker. The solution was filtered, washed sequentially with 90% ethanol and anhydrous ethanol, and dried in air to obtain MO nanocages.

[0023] (2) Preparation of Hex@MO: 0.4 g of hexaconazole (Hex) was completely dissolved in 20 mL of methanol to obtain solution A. MO prepared according to the above steps was dissolved in 20 mL of water to obtain solution B. Solution A was slowly added dropwise to solution B, stirred at room temperature, centrifuged, washed with methanol, and dried at room temperature to obtain Hex@MO nanoparticles.

[0024] Example of results: (1) The microstructure of the MO nanocages and Hex@MO nanoparticles prepared in Example 1 was characterized. Specifically, appropriate amounts of the MO nanocages and Hex@MO nanoparticles prepared in Example 1 were dispersed in ethanol, sonicated for 30 min, and then dropped onto a copper grid. The morphology was observed using an H-7650 transmission electron microscope. The particle size of 100 particles was randomly measured using Nano Measure software, and the average particle size of the Hex@MO nanoparticles was measured to be (5.84±1.15) nm. The TEM image of the MO nanocage is shown below. Figure 2 As shown in (a) and (b), Figure 2 (c) shows the Tyndall effect plot of Hex. The TEM image of Hex@MO is shown below. Figure 2 As shown in (d) and (e), the particles are spherical, uniformly distributed, and have smooth surfaces. The Tyndall effect diagram is as follows. Figure 2 As shown in (f).

[0025] (2) FTIR analysis was performed on the MO nanocages and Hex@MO nanoparticles prepared in Example 1. Specifically, MO and Hex@MO were mixed and ground with potassium bromide at a mass ratio of 1:100 (sample: potassium bromide), dried, pressed into tablets, and scanned using an infrared spectrometer with a scanning range of 4000-500 cm⁻¹. -1 The test results are as follows: Figure 3 As shown, the infrared spectrum of Hex@MO also shows the characteristic absorption peak of Hex (such as 3131 cm⁻¹). -1 The triazole ring CH stretching vibration at 1608 and 1509 cm⁻¹ -1 The C=N and C=C skeletal vibrations at the point of origin and the characteristic absorption peaks of MO (such as 1543 and 1421 cm⁻¹) -1 (The acetic acid ligand vibration at the loading point). The characteristic peak position shifted slightly after loading, indicating that there is an interaction (such as hydrogen bonding) between Hex and the MO nanocage, but no new metal coordination bonds were formed, proving that Hex was successfully encapsulated in the MO cavity.

[0026] (3) The MO nanocages, Hex@MO nanoparticles, and Hex prepared in Example 1 were analyzed by TGA. Specifically, a TGA-1000 thermogravimetric analyzer was used, and the temperature was increased from 30°C to 800°C at a rate of 10°C / min under a nitrogen atmosphere. The test results are as follows: Figure 4As shown in the figure, Hex decomposes rapidly in the range of 251-353℃, almost completely losing weight. The weight loss of MO nanocages mainly consists of two stages: loss of water of crystallization and coordination water at 50-320℃, and framework collapse above 400℃. Hex@MO shows three weight loss stages, with the second stage (240-400℃) corresponding to the decomposition of the encapsulated Hex. The weight loss temperature range in this stage is wider than that of pure Hex, indicating that the encapsulation of MO nanocages significantly improves the thermal stability of Hex.

[0027] (4) The crystal structure of the MO nanocages, Hex@MO nanoparticles, and Hex prepared in Example 1 was determined using an X-ray diffractometer. Specifically, the scanning speed was 2° / min, the scanning angle was 5°-80°, and the test voltage and current were set to 40kV and 30mA, respectively. The test results are as follows: Figure 5 As shown, the MO nanocage exhibits good crystallinity, and its characteristic diffraction peaks are consistent with those reported in the literature. After loading Hex, the intensity of the characteristic diffraction peak of Hex@MO around 5.4° decreased significantly, but the peak position did not shift significantly. This indicates that Hex molecules enter the MO cavity and affect the lattice strength of MO through intermolecular forces, but do not destroy its basic crystal structure.

[0028] (5) The release behavior of Hex@MO nanoparticles prepared in Example 1 under different pH conditions was determined by the dialysis bag method. Specifically, a 30% methanol-water solution was prepared, and the pH of the solution was adjusted with dilute hydrochloric acid and sodium hydroxide to prepare release media under different pH conditions (pH=4.0, 5.0, 7.0, 9.0). 30 mg of sample was dispersed in 5 mL of release media and placed in a dialysis bag (molecular weight cutoff: 8000 Da). The dialysis bag was sealed with a dialysis clamp and placed in a 100 mL centrifuge tube, and 60 mL of release media was added to the tube. The centrifuge tube was placed in a constant temperature shaker and shaken at a speed of 100 r / min. The supernatant was taken at specific time intervals (15 min, 30 min, 1 h, 2 h…), and an equal volume of fresh release media was added immediately after each sampling. The release of hexaconazole was determined by HPLC. The HPLC parameters were: C18 reversed-phase column (5 μm × 4.6 mm × 150 mm); column temperature: 30℃; mobile phase: methanol / water (V / V) = 70:30; flow rate: 1 mL / min; detection wavelength: 275 nm. The test results are as follows: Figure 6As shown, under a simulated weakly acidic environment (pH=5.0) of bacterial infection, the cumulative release rate reached 73.21% after 6 days, significantly higher than that under neutral (42.83%) and alkaline (38.42%) conditions, demonstrating its excellent pH-responsive release characteristics. Fitting the release data using the Ritger-Peppas model, the release index n=0.306 (<0.45) at pH=5.0, indicating that the release mechanism of Hex is mainly Fickian diffusion, i.e., the drug diffuses from the carrier through a concentration gradient.

[0029] (6) The mycelial growth rate method was used to investigate the effect of Hex@MO nanoparticles prepared in Example 1 and Hex commercial suspension (HexSC) on the pathogen of rice sheath blight (… Rhizoctonia solani, R. solani The inhibitory effect of Hex@MO, Hex commercial suspension (Hex SC), and blank control were characterized by adding different concentrations of Hex to PDA medium and inoculating the culture. R. solani Mycelial pellets were cultured at 27℃. Colony diameters were measured on days 3 and 5, and the mycelial growth inhibition rate was calculated. Results are as follows: Figure 6 As shown, at the same Hex concentration, the antibacterial rate of Hex@MO was consistently higher than that of Hex SC. The half-maximal effective concentration (EC50) was calculated. 50 The results showed that after 5 days of culture, Hex@MO EC... 50 The concentration was 4.795 mg / L, lower than Hex SC's 5.763 mg / L, indicating that it has superior and longer-lasting bactericidal activity.

[0030] (7) The rain erosion resistance of Hex@MO nanoparticles and Hex SC prepared in Example 1 on the surface of rice leaves was evaluated using a laboratory-scale washing method. To facilitate observation of Hex@MO and Hex SC, fluorescein isothiocyanate (FITC) was used for staining, with both formulations stained with the same concentration of FITC, and designated as FITC@MO@Hex and FITC@Hex SC, respectively. Rice leaves were fixed to a glass slide with tape to prevent curling. The sample was evenly applied to a 2cm × 2cm section of the leaf and then air-dried under ambient conditions. The slide was then placed on a platform tilted at 40°, and the treated leaf surface was rinsed with deionized water at a flow rate of 30 mL / min. Fluorescence retention was evaluated by comparing fluorescence images taken immediately after application (0 min) and 6 min after rinsing. The fluorescence imaging results are shown below. Figure 7As shown, before washing, the FITC@Hex SC group exhibited numerous discrete fluorescent bright spots, while the FITC@MO@Hex group showed a more uniform fluorescence distribution. After simulating heavy rain, the fluorescence signal of the FITC@Hex SC group was significantly weakened, while the FITC@MO@Hex group maintained a clearer fluorescence distribution and higher signal intensity, indicating that it has stronger resistance to rain erosion and better leaf adhesion.

[0031] The above description is only a preferred embodiment of the present invention. Given that those skilled in the art can make appropriate changes and modifications to the above embodiments, the present invention is not limited to the specific embodiments described above, and some modifications and changes to the present invention should also fall within the protection scope of the claims of the present invention.

Claims

1. A pH-responsive hexaconazole controlled-release nanoparticle, characterized in that, It includes polyacid nanocage carriers and hexaconazole loaded by impregnation.

2. A method for preparing pH-responsive hexaconazole controlled-release nanoparticles as described in claim 1, characterized in that, include: (1) Add ammonium acetate and hydrazine sulfate to ammonium molybdate solution, stir evenly, slowly add aqueous acetic acid solution, let stand at room temperature, filter, wash, dry, and obtain polyacid nanocage carrier; (2) Hexaconazole was dissolved in methanol, polyacid nanocage carrier was dissolved in water, and then the methanol solution of hexaconazole was slowly added dropwise to the polyacid nanocage carrier solution. The mixture was stirred at room temperature, centrifuged, washed, and dried to obtain pH-responsive hexaconazole controlled-release nanoparticles, which were named Hex@MO nanoparticles.

3. The preparation method according to claim 2, characterized in that, (1) The molar ratio of ammonium molybdate, ammonium acetate and hydrazine sulfate is 1:36:1.

36.

4. The preparation method according to claim 2, characterized in that, (1) The volume concentration of the acetic acid aqueous solution is 50%.

5. The preparation method according to claim 2, characterized in that, (1) The settling time is 7 days.

6. The preparation method according to claim 2, characterized in that, (1) Washing is performed sequentially with 90% ethanol and anhydrous ethanol.

7. The preparation method according to claim 2, characterized in that, (2) The stirring time at room temperature is 24h.

8. The preparation method according to claim 2, characterized in that, (2) Methanol is used for washing.

9. The preparation method according to claim 2, characterized in that, (2) Drying is done naturally at room temperature.

10. An application of the pH-responsive hexaconazole controlled-release nanoparticles according to claim 1, characterized in that, It is used as a controlled-release nano-pesticide for the prevention and control of rice sheath blight.

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