Chitosan-allyl isothiocyanate nano slow-release pesticide as well as preparation method and application thereof

Abstract

The invention belongs to the field of agricultural chemistry, and discloses a chitosan-allyl isothiocyanate nano slow-release pesticide and a preparation method and application thereof.The preparation method comprises the steps that S1, a hydrophobic phase is prepared, specifically, allyl isothiocyanate is dissolved in ethyl acetate and fully stirred, and the mixture serves as the hydrophobic phase; s2, preparing a hydrophilic phase: dissolving chitosan of which the deacetylation degree is greater than or equal to 85% into an acetic acid solution of which the concentration is 1%-3% to serve as the hydrophilic phase; s3, mixing: slowly dropwise adding the hydrophobic phase into the hydrophilic phase, dropwise adding a glutaraldehyde solution with the mass concentration of 2-5% into the mixed solution, and stirring after dropwise adding; and S4, after the reaction is finished, putting the mixed solution into a dialysis bag, dialyzing for 2-3 days by using deionized water, replacing the deionized water for 3-4 times every day during the dialysis, and performing freeze drying treatment on the dialyzed solution to finally obtain the chitosan-nano allyl isothiocyanate sustained-release agent. The composition has the effects of preventing and treating wheat scab, reducing toxin accumulation, controlling toxin and reducing toxin.

Description

Technical Field

[0001] The invention belongs to the technical field of agricultural chemistry, and particularly relates to a chitosan-allyl isothiocyanate nano slow-release pesticide, a preparation method thereof, and an application thereof. Background Art

[0002] Wheat is an important grain crop in my country. Head blight is a major disease that harms wheat production, severely impacting both yield and quality. Wheat head blight is caused by various Fusarium species, with Fusarium graminearum and Fusarium asiaticum being the primary causative fungi. Head blight can affect wheat from seedling stage to heading stage, with ear rot being the most severe, severely impacting wheat yield. Head blight fungi also produce a toxin, deoxynivalenol (DON). DON can contaminate not only crops but also grain products. It is highly toxic, second only to aflatoxin. DON is rapidly and efficiently absorbed by the body and is highly cytotoxic. Ingestion of DON-contaminated food by humans and animals can cause acute poisoning symptoms such as anorexia, vomiting, diarrhea, fever, unsteadiness, and slowed reflexes. In severe cases, it can damage the hematopoietic system and cause death.

[0003] While there are currently several fungicides available for wheat fusarium head blight control, most are ineffective in inhibiting the pathogen's production of the toxin DON, and some even stimulate the pathogen to synthesize more toxins. Therefore, it is crucial to identify new fungicides that can both prevent the disease and control and reduce the toxin. Summary of the Invention

[0004] The present invention aims to provide a chitosan-allyl isothiocyanate nano slow-release pesticide and its preparation method and application, which has the effects of preventing and treating wheat scab, reducing toxin accumulation, controlling toxicity and reducing toxicity.

[0005] The above technical objectives of the present invention are achieved through the following technical solutions: A method for preparing chitosan-allyl isothiocyanate nano-sustained release pesticide, comprising the following steps:

[0006] S1. Preparation of hydrophobic phase: dissolve allyl isothiocyanate in ethyl acetate and stir thoroughly to prepare the hydrophobic phase;

[0007] S2. Preparation of hydrophilic phase: dissolving chitosan with a deacetylation degree ≥85% in 1% to 3% acetic acid solution as the hydrophilic phase;

[0008] S3. Cross-linking: Slowly add the hydrophobic phase dropwise into the hydrophilic phase, add 2% to 5% glutaraldehyde solution dropwise to the mixed solution, and stir after the addition is complete;

[0009] S4. After the cross-linking is completed, the mixed solution is placed in a dialysis bag and dialyzed with deionized water for 2 to 3 days, during which the deionized water is replaced 3 to 4 times a day. The dialyzed solution is freeze-dried to finally obtain the chitosan-nano allyl isothiocyanate sustained-release agent.

[0010] The present invention is further configured as follows: the mass ratio of ethyl acetate to allyl isothiocyanate is 80-100:1, and the stirring time is 20 minutes.

[0011] The present invention is further configured as follows: in step S2, the mass ratio of chitosan to acetic acid solution is 1:100.

[0012] The present invention is further configured as follows: in step S3, the mass ratio of the hydrophilic phase to the hydrophobic phase is 3 to 5:1.

[0013] The present invention is further configured as follows: Step S3 is to dissolve zein in 75% by mass ethanol to prepare a zein solution with a mass concentration of 10 to 20 g / L, slowly add the hydrophilic phase to the zein solution, stir once, then slowly drop the hydrophobic phase into the stirred solution, and then add dropwise a glutaraldehyde solution with a mass concentration of 2% to 5%, and stir a second time.

[0014] The present invention is further configured as follows: the volume ratio of the zein solution to the hydrophilic phase is 1 to 2:1.

[0015] The present invention is further configured as follows: in step S3, the volume of the glutaraldehyde solution is 5% to 10% of the volume of the hydrophilic phase, the stirring time is 2 to 4 hours, and the reaction temperature is maintained at 30 to 40° C. during the dropwise addition and stirring process.

[0016] A chitosan-allyl isothiocyanate nano slow-release pesticide is prepared by the above-mentioned preparation method.

[0017] The invention discloses an application of chitosan-allyl isothiocyanate nano slow-release pesticide, wherein the chitosan-allyl isothiocyanate nano slow-release pesticide prepared by the above method is diluted to a concentration of 0.5 mg / mL for preventing and controlling wheat scab.

[0018] The invention is further configured as follows: applying pesticide to wheat stems and leaves once during the flowering period, and applying pesticide a second time 7-10 days later.

[0019] The beneficial effects of the present invention are:

[0020] 1. In this application, allyl isothiocyanate (AITC) is used as an antibacterial agent, which can significantly inhibit the pathogenic bacteria of wheat fusarium and prevent the continued synthesis of toxins. The nano sustained-release agent prepared by using chitosan as a nanocarrier and loading allyl isothiocyanate can deliver the antibacterial agent to the diseased site in a targeted manner, increase the drug concentration in the diseased area, and enhance the prevention and control effect of wheat fusarium; chitosan is used to wrap the antibacterial agent. On the one hand, the amino group of chitosan can form a dynamic covalent bond with the isothiocyanate group of AITC, reducing the decomposition of the pesticide and maintaining its activity. On the other hand, due to the wrapping effect of chitosan in the outer layer, the prepared pesticide has excellent sustained-release properties, which can effectively delay the release rate of allyl isothiocyanate. After the pesticide is applied, allyl isothiocyanate can be continuously released within 7 to 10 days, maintaining continuous inhibition of pathogens, reducing the number of applications, and reducing the pollution of pesticides to the environment.

[0021] 2. The isothiocyanate group of AITC in this application is highly reactive and can undergo nucleophilic reactions with the epoxy group or hydroxyl group of DON, destroying its toxic structure and reducing its bioavailability, thereby reducing the toxicity of the toxins already produced. Chitosan can not only adsorb DON toxins through hydrogen bonds and electrostatic effects through the amino and hydroxyl groups in its molecules, thereby reducing the content of some toxins and further reducing the toxins in wheat, but also produce a synergistic effect on the nucleophilic reaction, promoting the reduction of toxin toxicity, thereby achieving the effect of poison control.

[0022] 3. The chitosan compound in this application can not only load allyl isothiocyanate, but also stimulate the wheat plant's own defense mechanism, enhance the wheat's resistance to ergot, and promote the growth of beneficial microorganisms in the soil, thereby improving the soil ecological environment. The results of field tests show that the wheat treated with the present invention has significantly increased defense enzyme activity (such as peroxidase), indicating that the wheat's stress resistance is significantly improved.

[0023] 4. In this application, zein and chitosan are cross-linked through hydrophobic interactions, hydrogen bonds, etc. to form denser nanoparticles, which have improved resistance to mechanical shearing, are not easily broken during foliar spraying, and can also resist the damage of ultraviolet rays to pesticides. DETAILED DESCRIPTION

[0024] The technical solutions of the present invention will be described clearly and completely below with reference to specific embodiments. Obviously, the embodiments described are only some of the embodiments of the present invention, not all of them. All other embodiments derived by persons of ordinary skill in the art based on the embodiments of the present invention without inventive effort are also within the scope of protection of the present invention.

[0025] Preparation of chitosan-allyl isothiocyanate nano-sustained release pesticide

[0026] Example 1

[0027] S1. Dissolve 100 mg of allyl isothiocyanate in 10 mL of ethyl acetate and stir for 20 min to prepare a hydrophobic phase; S2. Dissolve 0.35 g of chitosan with a deacetylation degree of 90% in 50 mL of acetic acid solution with a mass concentration of 1% to 3%, and stir with a magnetic stirrer to prepare a hydrophilic phase; S3. Slowly add the hydrophobic phase dropwise to the hydrophilic phase with a mass ratio of hydrophilic phase to hydrophobic phase of 4:1, and add a glutaraldehyde solution with a mass concentration of 2% to 5% dropwise to the mixed solution. The volume of the glutaraldehyde solution is 8% of the volume of the hydrophilic phase. After the dropwise addition is completed, the mixture is stirred for 2 to 4 hours. The reaction temperature is maintained at 30 to 40° C. during the dropwise addition and stirring process to obtain a uniform mixed solution. S4. After the reaction is completed, the mixed solution is placed in a dialysis bag with a molecular weight cutoff of 3500 to 5000 Da and dialyzed with deionized water for 2 to 3 days. The deionized water is replaced 3 to 4 times a day during the dialyzed solution. The dialyzed solution is freeze-dried to finally obtain a chitosan-nano allyl isothiocyanate sustained-release agent.

[0028] Example 2

[0029] The preparation method is the same as that in Example 1, except that step S3 is to dissolve zein in 75% by mass ethanol to prepare a zein solution with a mass concentration of 5 to 20 g / L, slowly add the hydrophilic phase to the zein solution and stir once, then slowly dropwise add the hydrophobic phase to the stirred solution, and then dropwise add a glutaraldehyde solution with a mass concentration of 2% to 5% and stir a second time, the volume ratio of the hydrophilic phase to the hydrophobic phase is 5:1, and the volume ratio of the zein solution to the hydrophilic phase is 1 to 2:1.

[0030] Example 3

[0031] The preparation method was the same as that in Example 1, except that in S1, 112.75 mg of allyl isothiocyanate was dissolved in 10 mL of ethyl acetate and stirred for 20 min to prepare a hydrophobic phase; S2, 0.25 g of chitosan with a deacetylation degree of 90% was dissolved in 50 mL of acetic acid solution with a mass concentration of 1% to 3%; in S3, the mass ratio of the hydrophilic phase to the hydrophobic phase was 3:1, and the volume of the added glutaraldehyde solution was 5% of the volume of the hydrophilic phase.

[0032] Example 4

[0033] The preparation method was the same as that in Example 1, except that in S1, 75.16 mg of allyl isothiocyanate was dissolved in 10 mL of ethyl acetate and stirred for 20 min to prepare a hydrophobic phase; in S2, 0.5 g of chitosan with a deacetylation degree of 90% was dissolved in 50 mL of acetic acid solution with a mass concentration of 1% to 3%; in S3, the mass ratio of the hydrophilic phase to the hydrophobic phase was 5:1, and the volume of the added glutaraldehyde solution was 10% of the volume of the hydrophilic phase.

[0034] Comparative Example 1 Preparation of allyl isothiocyanate nano-sustained-release pesticide without chitosan coating.

[0035] The preparation method is the same as that of Example 1, except that chitosan is not added in step S2.

[0036] Active Cases

[0037] Example 5

[0038] Wheat fields where wheat fusarium head blight occurs all year round were selected and divided into 18 test plots. Each pesticide was repeated in 3 plots, and the plots were arranged in random blocks with an interval of 50m between adjacent test plots.

[0039] Treatment method: The sustained-release agent prepared in Examples 1 to 4 and Comparative Example 1 was prepared into a pesticide liquid with a concentration of 0.5 mg / mL, and sprayed evenly on the stems and leaves of wheat during the flowering period. A blank control group was set up, and an equal amount of water was sprayed in the blank control group. The second application was carried out 7 days later, and the application amount and method were the same as the first application.

[0040] Data processing and analysis: One month after the first spraying, the diseased ear rate of each test area was counted, diseased ear rate = (number of diseased ears ÷ total number of surveyed ears) × 100%; during the wheat maturity period, the peroxidase activity of wheat plants was measured using peroxidase activity assay reagent and colorimetric method respectively; during the harvest period, 500 g of wheat grain samples were randomly selected from each test area and the husks were removed. The DON toxin content of the wheat grains was determined in the laboratory according to the People's Republic of China Entry-Exit Inspection and Quarantine Industry Standard SN / T3137-2012, and the DON toxin control reduction rate was calculated according to the following formula: wheat grain DON toxin control reduction rate = [(DON toxin content of wheat grains in the control area - DON toxin content of wheat grains in the test area) ÷ DON toxin content of wheat grains in the control area] × 100%.

[0041] The test results are shown in Table 1 below.

[0042] Table 1

[0043]

[0044] It can be seen from the above table that the chitosan-allyl isothiocyanate nano slow-release pesticide prepared by the method of the present application can effectively prevent and control wheat fusarium scabra, and at the same time can also reduce the DON toxin in wheat grains and reduce the toxicity of wheat grains. Compared with Comparative Example 1, Example 1 has a significantly reduced diseased ear rate. It can be seen that the combination of chitosan and allyl isothiocyanate can enhance the prevention and control effect of wheat fusarium scabra, delay the release rate of allyl isothiocyanate, maintain continuous inhibition of pathogens, and reduce the number of applications; compared with Comparative Example 1, Example 1 has a significantly increased peroxidase activity. It can be seen that chitosan can increase the activity of defensive enzymes and improve the stress resistance of wheat.

Claims

1. A method for preparing chitosan-allyl isothiocyanate nano-slow-release pesticide, characterized in that: The following steps are involved: S1. Preparation of hydrophobic phase: dissolve allyl isothiocyanate in ethyl acetate and stir thoroughly to prepare the hydrophobic phase; S2. Preparation of hydrophilic phase: dissolving chitosan with a deacetylation degree ≥85% in 1% to 3% acetic acid solution as the hydrophilic phase; S3. Cross-linking: Slowly add the hydrophobic phase dropwise into the hydrophilic phase, add 2% to 5% glutaraldehyde solution dropwise to the mixed solution, and stir after the addition is complete; S4. After the cross-linking is completed, the mixed solution is placed in a dialysis bag and dialyzed with deionized water for 2 to 3 days, during which the deionized water is replaced 3 to 4 times a day. The dialyzed solution is freeze-dried to finally obtain the chitosan-nano allyl isothiocyanate sustained-release agent.

2. The method for preparing a chitosan-allyl isothiocyanate nano-sustained-release pesticide according to claim 1, characterized in that: In step S1, the mass ratio of ethyl acetate to allyl isothiocyanate is 80-100:1, and the stirring time is 20 minutes.

3. The method for preparing a chitosan-allyl isothiocyanate nano-sustained-release pesticide according to claim 1, characterized in that: In step S2, the mass ratio of chitosan to acetic acid solution is 0.5 to 1:

100.

4. The method for preparing a chitosan-allyl isothiocyanate nano-sustained-release pesticide according to claim 1, characterized in that: In step S3, the mass ratio of the hydrophilic phase to the hydrophobic phase is 3 to 5:

1.

5. The method for preparing a chitosan-allyl isothiocyanate nano-sustained-release pesticide according to claim 1, characterized in that: The step S3 comprises dissolving zein in 75% by mass ethanol to prepare a zein solution with a mass concentration of 10 to 20 g / L, slowly adding the hydrophilic phase to the zein solution, stirring once, then slowly dropping the hydrophobic phase into the stirred solution, and then gradually adding a glutaraldehyde solution with a mass concentration of 2% to 5%, stirring a second time.

6. The method for preparing a chitosan-allyl isothiocyanate nano-sustained-release pesticide according to claim 5, characterized in that: The volume ratio of the zein solution to the hydrophilic phase is 1 to 2:

1.

7. The method for preparing a chitosan-allyl isothiocyanate nano-sustained-release pesticide according to claim 1 or 5, characterized in that: In step S3, the volume of the glutaraldehyde solution is 5% to 10% of the volume of the hydrophilic phase, the stirring time is 2 to 4 hours, and the reaction temperature is maintained at 30 to 40° C. during the dropwise addition and stirring process.

8. A chitosan-allyl isothiocyanate nano-sustained-release pesticide, characterized by: The preparation method according to any one of claims 1 to 6 is used.

9. The use of a chitosan-allyl isothiocyanate nano-sustained-release pesticide according to any one of claims 1 to 6, characterized in that: The chitosan-allyl isothiocyanate nano slow-release pesticide is diluted to a concentration of 0.5 mg / mL for preventing and controlling wheat fusarium head blight.

10. The use of a chitosan-allyl isothiocyanate nano slow-release pesticide according to claim 8, characterized in that: Apply the pesticide to the wheat stems and leaves once during the flowering period and apply it a second time 7 to 10 days later.

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