ROS-targeted chlorantraniliprole nanocrystal as well as preparation method and application thereof

Abstract

The invention discloses an ROS-targeted chlorantraniliprole nanocrystal which is prepared by taking an intermediate formed by assembling a metal polyphenol network based on tannic acid-iron on the surface of a chlorantraniliprole crystal nucleus as a core and driving MPN, BSA and PEG to be assembled on the surface of the nanocrystal through a coordination reaction of surface tannic acid and Fe < 3 + >. The invention also discloses a preparation method and application thereof. The ROS-targeted chlorantraniliprole nanocrystal CAP (at) TF-PEG synthesized by the invention has the advantages of efficient CAP carrying capacity, stable ROS responsive release capacity, paddy rice injury site targeting capacity, ideal microscopic size, better stability and high biocompatibility. The ROS-targeted chlorantraniliprole nanocrystal CAP (at) TF-PEG has relatively strong superiority in the aspects of carrying and targeted delivery of CAP, and a theoretical basis is provided for the development of a subsequent related pesticide responsive slow release technology.

Description

Technical Field

[0001] This invention relates to a chlorantraniliprole nanocrystal targeting ROS, its preparation method and application, belonging to the field of biomedical materials technology. Background Technology

[0002] Pesticides are crucial for the control of plant pests and diseases, as well as for food security. Traditional pesticide formulations are inefficient, easily leading to overuse and abuse, further exacerbating environmental pollution. Extending pesticide shelf life and improving pesticide effectiveness on crops helps reduce pesticide waste and environmental pollution. Research on responsive pesticide release is of practical significance in addressing the shortcomings of traditional pesticides. Responsive-release pesticides exhibit precise release of specific substances. Plant defense systems have response mechanisms to various types of damage, including pathogen invasion, drought and flood stress, physical damage, and pests and diseases. Oxidative stress is a rapid response process accompanied by the accumulation of large amounts of reactive oxygen species (ROS), and it is one of the important early signals in plant defense. ROS mainly include superoxide anion radicals, hydrogen peroxide, hydroxyl radicals, and singlet oxygen. ROS-responsive release systems can overcome the shortcomings of environmental responses. A typical approach is to use cleavable ROS groups as carriers to prepare slow-release systems. These materials are inexpensive and safe, but have not yet been used in agriculture.

[0003] Metal-Phenolic Networks (MPNs) are supramolecular structures that combine metal ions with plant polyphenols via coordination bonds. In recent years, biomacromolecule assembly systems based on polyethylene glycol and MPNs have been reported for the encapsulation and delivery of macromolecules such as proteins and nucleic acids. Summary of the Invention

[0004] The purpose of this invention is to provide chlorantraniliprole nanocrystals that target ROS.

[0005] The technical solution adopted in this invention is as follows: An intermediate for ROS-targeting chlorantraniliprole nanocrystals, CAP@TA-Fe III It is assembled on the surface of chlorantraniliprole (CAP) crystal nuclei by a tannic acid (TA)-iron-based metal polyphenol network. The metal polyphenol network anchors the hydrophobic surface and forms a dense film around the CAP crystal nuclei, thereby inhibiting Ostwald ripening and limiting the size of the nanocrystals.

[0006] The molecular formula of TA is C 76 H 52 O 46 The chemical structural formula is shown in formula (I): (I) Preferably, the material contains 1.29% Fe by mass, with the Fe element bonded in the form of Fe3+, a surface Fe atomic ratio of 0.7%, and chlorantraniliprole content of 29.22-31.58%.

[0007] This invention also discloses a ROS-targeting chlorantraniliprole nanocrystal, using the aforementioned intermediate as a core, through surface tannins and Fe... 3+ The coordination reaction drives MPN, BSA, and PEG to assemble into a ROS-responsive gating system on the surface of nanocrystals, encapsulating the nanocrystals and enabling ROS-responsive release.

[0008] Preferably, the hydrated particle size distribution of the chlorantraniliprole nanocrystals is 192.23-228.59 nm, the CAP content is 7.16-7.59%, the Fe element mass ratio in the nanocrystals is 1.29%, and the Fe atomic ratio on the surface of the nanocrystals is 0.7%.

[0009] Preferably, the chlorantraniliprole nanocrystals are made of... 89 Zr mark.

[0010] The present invention also discloses a method for preparing the above-mentioned intermediate, wherein a DMSO solution of CAP is added to pure water in an ultrasonic atmosphere, followed by the addition of TA and FeCl3. After ultrasonic reaction, the precipitate is collected by centrifugation, which is the intermediate of chlorantraniliprole nanocrystals.

[0011] Preferably, 100 μL of 5.7 mg / mL CAP in DMSO solution is added to 20 mL of pure water under an ultrasonic atmosphere, followed by the addition of 100 μL of 40 mg / mL TA and 50 μL of 6 mg / mL FeCl3. The mixture is ultrasonicated for 5 min, centrifuged at 12000 rpm and the supernatant is discarded. The precipitate is washed three times with pure water to obtain an intermediate of chlorantraniliprole nanocrystals.

[0012] The present invention also discloses a method for preparing the above-mentioned chlorantraniliprole nanocrystals, wherein the intermediate obtained above is dissolved in pure water, and polyethylene glycol, bovine serum albumin, tannic acid and FeCl3 are added in sequence, the mixture is stirred and reacted, and the precipitate is collected by centrifugation to obtain chlorantraniliprole nanocrystals.

[0013] Preferably, specifically, CAP@TA-Fe III Dissolve 2 mg in 6 mL of pure water, then add PEG4000 (20 mg / mL, 900 μL), BSA (10 mg / mL, 600 μL), TA (8 mg / mL, 600 μL), and FeCl3 (3 mg / mL, 180 μL) sequentially, and stir for 10 h. Centrifuge at 12000 rpm and wash three times with pure water.

[0014] The prepared chlorantraniliprole nanocrystals exhibit ROS-responsive sustained-release properties.

[0015] Preferably, the FeCl3 is mixed with 89 Zr, specifically, FeCl3 solution and 89 Zr (700 μCi) was mixed and added to the reaction system in place of the non-radioactive FeCl3 solution. (Labeling) 89 Zr's CAP@TF-PEG enables monitoring of metabolic distribution in rice based on PET signals.

[0016] The present invention also discloses the application of the above-mentioned chlorantraniliprole nanocrystals in the damaged tissues of targeted plants.

[0017] This invention provides a ROS-targeted chlorantraniliprole nanocrystal CAP@TF-PEG. A tannic acid-iron (TA-Fe)-based metal polyphenol network (MPN) is bound to the surface of chlorantraniliprole (CAP) crystals to restrict growth, resulting in CAP nanocrystals (CAP@TA-Fe). III Polyethylene glycol (PEG-4000), bovine serum albumin (BSA), tannic acid (TA), iron ions (Fe) 3+ ) in CAP@TA-Fe III Surface self-assembly forms a reactive oxygen species (ROS) responsive shell. TA-Fe III The phenolic hydroxyl groups in the structure undergo a quinone transformation in a ROS environment, weakening their affinity for Fe. 3+ The chelating ability and supramolecular structure degradation of the core CAP@TA-FeIII enable ROS-responsive sustained release. CAP@TF-PEG can be administered to rice via foliar spraying and root soaking, exhibiting targeted accumulation and responsive drug delivery capabilities in damaged rice tissues. CAP@TF-PEG, through... 89 Zr labeling followed by PET-CT scanning enables real-time monitoring of material distribution within plants, providing a reference for the design of related drug targeted delivery materials.

[0018] The ROS-targeting chlorantraniliprole nanocrystals CAP@TF-PEG synthesized in this invention possess highly efficient CAP carrying capacity, stable ROS-responsive release capability, targeting ability to rice damage sites, ideal microstructure, good stability, and high biocompatibility. These ROS-targeting chlorantraniliprole nanocrystals CAP@TF-PEG exhibit significant advantages in CAP carrying and targeted delivery, providing a theoretical basis for the subsequent development of related pesticide responsive sustained-release technologies.

[0019] Compared with the prior art, the present invention has the following advantages: (1) This invention utilizes TA and Fe 3+The stable coordination of the CAP crystals restricts their formation in water-based solvents, enabling nano-sized CAP crystals. A ROS-responsive degradation shell is constructed using TA-FeIII-based MPN combined with hydrophilic macromolecules, giving the system ROS-responsive release capability. This design method is simple and easy to operate, and can efficiently synthesize targeted delivery materials.

[0020] (2) The ROS-targeting chlorantraniliprole nanocrystals CAP@TF-PEG synthesized in this invention exhibit stable ROS-responsive degradation ability and targeted delivery capability to damaged rice sites. They are safe and reliable, and have broad application prospects in the field of pesticide targeted delivery.

[0021] (3) The ROS-targeting chlorantraniliprole nanocrystals CAP@TF-PEG of this invention are the first to simultaneously achieve the functions of CAP nanocrystal loading and targeted delivery to damaged rice tissue, which has not been reported in domestic or foreign literature. It can be seen that the ROS-targeting chlorantraniliprole nanocrystals CAP@TF-PEG obtained by the method of this invention are novel in design and have potential application value.

[0022] (4) The ROS-targeting chlorantraniliprole nanocrystals CAP@TF-PEG described in this invention can achieve real-time monitoring of drug distribution in rice through 89Zr labeling, providing a new research idea for pesticide delivery systems. Attached Figure Description

[0023] Figure 1 This is a schematic diagram of the synthetic route for CAP@TF-PEG.

[0024] Figure 2 The hydrated particle size of the CAP@TF-PEG of the present invention was determined using dynamic light scattering (DLS).

[0025] Figure 3 The intermediate CAP@TA-Fe of the present invention as photographed using a scanning electron microscope (SEM) III Material morphology diagram of CAP@TF-PEG.

[0026] Figure 4 The intermediate CAP@TA-Fe of this invention III The relative proportion of important elements in CAP@TF-PEG.

[0027] Figure 5 The image shows the X-ray photoelectron spectroscopy (XPS) spectrum of the CAP@TF-PEG of this invention.

[0028] Figure 6 This refers to the ROS-responsive release of CAP@TF-PEG according to the present invention.

[0029] Figure 7For the CAP@TF-PEG of the present invention in 89 Differences in tissue and temporal distribution between healthy rice and damaged rice models after Zr labeling. Detailed Implementation

[0030] The present invention will be further described below with reference to the embodiments, but the description of the embodiments does not limit the scope of protection of the present invention in any way.

[0031] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. The terminology used herein in the description of the invention is for the purpose of describing particular embodiments only and is not intended to limit the invention. Furthermore, while this document may provide examples of parameters containing specific values, it should be understood that the parameters need not be exactly equal to the corresponding values, but may approximate the corresponding values ​​within acceptable error tolerances or design constraints.

[0032] Unless otherwise specified, all substances or instruments used in the following examples can be obtained from conventional commercial sources.

[0033] Example 1: Synthesis of CAP@TF-PEG (1) such as Figure 1 As shown, a DMSO solution of CAP (5.7 mg / mL, 100 μL) was added to 20 mL of pure water under an ultrasonic atmosphere, followed by an aqueous solution of TA (40 mg / mL, 100 μL) and an aqueous solution of FeCl3 (6 mg / mL, 50 μL). The mixture was ultrasonicated for 5 min. After centrifugation at 12000 rpm, the supernatant was discarded, and the precipitate was washed three times with pure water to obtain the intermediate CAP@TA-Fe. III The CAP content was determined by HPLC (C18 reversed column, mobile phase 65% acetonitrile, water 35%, detection wavelength 245 nm), and the results are as follows. Figure 4 As shown. The obtained CAP@TA-Fe III Electron micrographs of the material, such as Figure 3 As shown.

[0034] (2) The CAP@TA-Fe obtained in step 1 III Dissolve in 6 mL of pure water, then add PEG4000 (20 mg / mL, 900 μL), BSA (10 mg / mL, 600 μL), TA (8 mg / mL, 600 μL), and FeCl3 (3 mg / mL, 180 μL) sequentially, and stir for 10 h. Centrifuge at 12000 rpm and wash three times with pure water. CAP content is determined by HPLC (C18 reverse-phase column, mobile phase 65% acetonitrile, 35% water, detection wavelength 245 nm). The results are shown below. Figure 4As shown. The hydrated particle size of CAP@TF-PEG was determined using dynamic light scattering (DLS), and the results are as follows. Figure 2 As shown. Electron micrographs of the obtained material are shown below. Figure 3 As shown. X-ray photoelectron spectroscopy (XPS) analysis was performed on the material, and the spectrum is shown below. Figure 5 As shown.

[0035] In aqueous solution, the intermediate CAP@TA-Fe III Dynamic light scattering (DLS) of CAP@TF-PEG showed its main particle size to be 153.71–202.59 nm; DLS of CAP@TF-PEG showed its main particle size to be 192.23–228.59 nm. CAP@TF-PEG was stably dispersed in aqueous solution. The intermediate CAP@TA-Fe was observed under SEM. III The microstructure shows that the particles are nearly cuboid; the microstructure of CAP@TF-PEG under SEM shows that the particles are nearly spherical.

[0036] Example 2: ROS-responsive release of CAP@TF-PEG The material CAP@TF-PEG obtained according to the steps of Example 1 was used to test its ROS-responsive release capability.

[0037] (1) Preparation of release medium: Prepare hydrogen peroxide solutions of different concentrations (0 mM, 10 mM, 50 mM, 100 mM), add 0.5% Tween-80 to obtain the release medium.

[0038] (2) Each CAP@TF-PEG group contained 27 mg of CAP, dissolved in pure water and added to a dialysis bag (3500 da). 900 mL of release medium was added externally, and the mixture was stirred at room temperature. Dialysis fluid was collected at different time points for HPLC analysis of the CAP concentration (C18 reverse-phase column, mobile phase 65% acetonitrile, 35% water, detection wavelength 245 nm). The total volume was then replenished with release medium. Results are as follows: Figure 6 As shown in the figure, CAP@TF-PEG exhibits ROS-responsive release properties.

[0039] Example 3: PET-CT Imaging Analysis of CAP@TF-PEG on a Rice Injury Model (1) Establishment of rice damage model: Rice seedlings were planted in the laboratory environment for 40 days and mechanical damage was caused by puncturing the middle of their stems.

[0040] (2) 89 Synthesis of Zr-labeled CAP@TF-PEG: In the synthesis of CAP@TF-PEG, FeCl3 solution was mixed with... 89Zr (700 μCi) was mixed and added to the reaction system instead of the non-radioactive FeCl3 solution. The product was centrifuged at 12,000 rpm and washed three times with pure water to obtain... 89 Zr-labeled CAP@TF-PEG. Radioactivity of starting materials and final products was measured using a radioactivity counter (RM-905A).

[0041] (3) PET-CT imaging analysis of damaged rice models: PET-CT scans of damaged rice models were performed using a Super Nova® PET / CT scanner (PingSeng Scientific). Leaves of healthy and damaged rice were immersed in the PET / CT scanner. 89 Zr-labeled CAP@TF-PEG (350 μCi, 20 mL) was administered via PET-CT imaging at 8, 24, and 48 hours post-administration. Images were reconstructed after scanning. Results are as follows: Figure 7 As shown, Figure 7 The left image shows the ROI statistics of the stem in PET-CT imaging between injured and normal rice (Ctrl), while the right image shows the ROI statistics of the root in PET-CT imaging between injured and normal rice (Ctrl). It is evident that CAP@TF-PEG has significant targeting ability to the injury sites in injured rice.

[0042] The above embodiments are only for illustrating the technical concept and features of the present invention. Their purpose is to enable those skilled in the art to understand the content of the present invention and implement it accordingly. They should not be used to limit the scope of protection of the present invention. All equivalent changes or modifications made in accordance with the spirit and essence of the present invention should be covered within the scope of protection of the present invention.

Claims

1. An intermediate for a ROS-targeted spinetoram nanocrystal, characterized in that, The nanocrystals have a size of 153.71-202.59 nm, which are assembled by a tannin acid-iron based metal polyphenol network on the surface of chlorantraniliprole crystal nucleus.

2. The intermediate of claim 1, wherein, The mass ratio of Fe element in the material is 1.29%, the Fe element is combined in the form of Fe 3+ The surface Fe atom ratio is 0.7%, and the content of chlorantraniliprole in the material is 29.22-31.58%.

3. A chlorantraniliprole nanocrystal targeting ROS, characterized in that, The intermediate of claim 1 or 2 as the core, through the coordination reaction of surface tannic acid and Fe 3+ MPN, BSA, PEG are assembled on the surface of nanocrystals driven by the coordination reaction.

4. The chlorantraniliprole nanocrystal of claim 3, wherein the chlorantraniliprole nanocrystal has a particle size of about 50 nm to about 200 nm. The hydrated particle size distribution of the chlorantraniliprole nanocrystals is 192.23-228.59 nm, the content of chlorantraniliprole is 7.16-7.59 %, and the mass ratio of Fe element in the nanocrystals is 1.29 %, and the ratio of Fe atoms on the surface of the nanocrystals is 0.7 %.

5. The chlorantraniliprole nanocrystal of claim 3, wherein the chlorantraniliprole nanocrystal has a particle size of about 50 nm to about 200 nm. The chlorantraniliprole nanocrystals are prepared by 89 Zr labeling.

6. Process for the preparation of the intermediates according to claim 1 or 2, characterized in that, The DMSO solution of chlorantraniliprole is added to pure water in an ultrasonic atmosphere, and then tannic acid and FeCl3 are added, and after ultrasonic reaction, the precipitate obtained by centrifugation is the intermediate of chlorantraniliprole nanocrystals.

7. The production method according to claim 6, wherein Specifically, 100 μL of 5.7 mg / mL DMSO solution of chlorantraniliprole is added to 20 mL of pure water in an ultrasonic atmosphere, and then 100 μL of 40 mg / mL tannic acid and 50 μL of 6 mg / mL FeCl3 are added, ultrasonic reaction is performed for 5 min, the supernatant is discarded by centrifugation at 12000 rpm, and the precipitate is washed with pure water for 3 times to obtain the intermediate of chlorantraniliprole nanocrystals.

8. The method of preparing a nanocrystal of chlorantraniliprole according to claim 3 or 4, characterized in that, The intermediate prepared in claim 6 or 7 is dissolved in pure water, and polyethylene glycol, bovine serum albumin, tannic acid and FeCl3 are added in sequence, and the mixture is stirred and reacted, and the precipitate obtained by centrifugation is the chlorantraniliprole nanocrystals.

9. The production method according to claim 8, characterized by, The FeCl3is mixed with 89 Zr.

10. The chlorantraniliprole nanocrystals according to any one of claims 3-5 for use in targeting damaged tissues of plants.

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