CN114938803A

CN114938803A - Multiple stimulus response type pesticide microsphere and preparation method thereof

Info

Publication number: CN114938803A
Application number: CN202210532544.7A
Authority: CN
Inventors: 周红军; 赵明; 周新华; 郝丽; 陈铧耀
Original assignee: Zhongkai University of Agriculture and Engineering
Current assignee: Zhongkai University of Agriculture and Engineering
Priority date: 2022-05-12
Filing date: 2022-05-12
Publication date: 2022-08-26

Abstract

The invention relates to the technical field of air conditioners, in particular to a multiple stimulus response type pesticide microsphere and a preparation method thereof.

Description

Multiple stimulus response type pesticide microsphere and preparation method thereof

Technical Field

The invention relates to the technical field of pesticide insecticides, in particular to a multiple stimulus response type pesticide microsphere and a preparation method thereof.

Background

According to the prediction of united nations, the world population reaches 96 hundred million in 2050 in the future, the food demand is predicted to increase by 70-110%, and how to use limited land to live the rapidly growing population is an urgent need to solve a global problem. 1, the diseases and insect pests directly cause 40% of the global agricultural productivity loss, so that a large amount of pesticides are used in the agricultural production process to ensure the grain safety. About 400 million tons of insecticide are used worldwide each year, most of the pesticides are lost to the environment due to dust drift, rain erosion, photodegradation and rapid evaporation, and only 0.1% of the active ingredient is used to destroy the target organism. The inefficient use of pesticides severely damages the natural environment, such as resistance generation by target organisms, loss of biodiversity, water pollution and the like, and further dangerously, the direct exposure of pesticides and the accumulation of food chain damage human health, and the development of sustainable agriculture is hindered by the inefficient use of pesticides. New materials and methods are needed to improve the efficiency of pesticide use in order to achieve the sustainable development goals of united nations (zero hunger, clean water and sanitary facilities).

In recent years, the application of the controlled release technology in the field of pesticides provides a new platform for green agriculture. Controlled release pesticides can be targeted for responsive release through the use of carriers with controlled release and target selectivity, and these intelligent systems can provide active ingredients in a targeted, controlled and safe manner under specific microenvironment conditions. Wherein, the stimulation response system improves the pesticide controlled release characteristic and is a feasible scheme for promoting the pesticide to respond to biological or non-biological stimulation and intelligently release. Researchers have developed advanced controlled-release avermectin formulations based on inorganic materials (mesoporous silica, montmorillonite, boron nitride, etc.), synthetic polymeric materials (polysuccinimide, polylactic acid) and natural polymeric materials (cellulose, chitosan, sodium alginate, zein, etc.). However, the intelligent response degree of the current controlled-release pesticide is not enough, and the targeted controlled-release pertinence of the active ingredients is not strong enough.

The intelligent response carrier material can respond to the change of environmental stimulus, such as the change of stimulus of pH, temperature, enzyme, oxidation reduction, light, magnetic field, ionic strength and the like, and realizes the targeted controlled release of the effective components. Therefore, the development of new carriers with intelligent response to environmental stimuli is an important method for preparing controlled-release pesticide formulations. In order to improve the effective period and utilization rate of pesticide, however, people have not passed through the design

Diamondback moth is the most harmful pest to crucifers in the world and causes a worldwide economic loss of about $ 4 to $ 50 billion each year. . The temperature is an important factor influencing the growth of insects and microorganisms, and researches show that the growth and development speed of the diamondback moth is accelerated along with the temperature rise in the range of 17.5-32.5 ℃, so that a controlled release preparation which is quickly released along with the temperature rise in the range needs to be designed to cope with the high population density of the diamondback moth at high temperature. On the other hand, the microcapsule has longer service life and biosafety, but the slow release of the microsphere in a living body causes low bioactivity of the microcapsule, and is an important factor for limiting the use of the microcapsule. Wherein, the micro environment of the insects is used as a stimulus factor to control the rapid release of the pesticide in the insects, which is a high-efficiency pest killing mode. The plutella xylostella is rich in glutathione (an antioxidant mercaptan) and has a unique alkaline environment in the midgut, and the stimulation factors can be used as a key to open a switch on a controlled-release pesticide carrier. In consideration of the complex environment of the pesticide in practical application, an intelligent pesticide microcapsule with multiple responsiveness needs to be designed, so that the intelligent pesticide microcapsule has longer lasting effect after being sprayed, and can quickly release effective ingredients for controlling plant diseases and insect pests under the biotic and abiotic stresses of crops.

Disclosure of Invention

One of the purposes of the invention is to provide a multiple stimulus response type pesticide microsphere which can automatically release pesticide and high-efficiency target controlled release pesticide according to needs and has the advantages of long validity period, high utilization rate and environmental friendliness.

The second purpose of the invention is to provide a preparation method of the pesticide microsphere with multiple stimulus responses.

In order to achieve one of the above purposes, the invention provides the following technical scheme:

provided is a multi-stimulus response type pesticide microsphere, which comprises modified carboxymethyl cellulose, an insecticidal component, isophorone diisocyanate and n-hexadecane, wherein the modified carboxymethyl cellulose comprises carboxymethyl cellulose and cystamine dihydrochloride.

In some embodiments, the insecticidal composition is one or more of abamectin, ivermectin, chlorpyrifos, 2, 4-dichlorophenoxyacetic acid, emamectin benzoate, chlorantraniliprole, lambda-cyhalothrin.

The multiple stimulus response type pesticide microsphere has the beneficial effects that:

(1) the pesticide microsphere comprises an oil phase component and a water phase component, wherein the oil phase component and the water phase component can form interfacial polymerization, and the interfacial polymerization microsphere can efficiently encapsulate insecticidal components, so that the ultraviolet light stability of the insecticidal components in the microsphere is greatly improved, the stability of the insecticidal components under ultraviolet light is better, and the longer-acting insecticidal effect is achieved.

(2) According to the multi-stimulus response type pesticide microsphere, the modified carboxymethyl cellulose is cystamine dihydrochloride modified carboxymethyl cellulose, so that amino, hydroxyl and urea bonds exist on the shell of the pesticide microsphere, the amino, hydroxyl and urea bonds form hydrogen bonds with fatty acid, fatty alcohol and fatty aldehyde on a wax layer, and acting force between the amino, hydroxyl and urea bonds and leaf surfaces can effectively prevent microcapsules on the leaf surfaces from being washed away by rainwater, and the durability of the pesticide microsphere is improved.

(3) The pesticide microsphere of the multiple stimulus response type pesticide microsphere contains n-hexadecane, and the n-hexadecane is converted from a solid state to a liquid state at the temperature of higher than 20 ℃, so that the pesticide microsphere can accelerate the release of the pesticide in the microsphere at high temperature, further more pesticide can be released at high temperature, and the problem that plant diseases and insect pests are easy to break out at high temperature can be better prevented and treated; on the contrary, in a low-temperature environment, n-hexadecane is not easy to be converted into a liquid state, the pesticide released by the pesticide microspheres is less, and pests are less at the moment, so that the pesticide dosage can be saved, the targeted controlled release of the pesticide is realized, and the effective period, the utilization rate and the environmental friendliness of the pesticide microspheres are improved.

(4) According to the multi-stimulus response type pesticide microsphere, the modified carboxymethyl cellulose is cystamine dihydrochloride modified carboxymethyl cellulose, so that disulfide bonds are contained on a spherical shell of the pesticide microsphere, when insects eat the pesticide microsphere, rich glutathione in the insect body can be reduced and destroyed by the disulfide bonds on the pesticide microsphere, the pesticide microsphere is disintegrated and the effective components are quickly released, the quick-pest killing effect is achieved, the pesticide microsphere has higher pest killing activity by stimulus release of microenvironment in the insect body, and the effect of releasing according to needs is achieved.

(5) The multi-stimulus response type pesticide microsphere provided by the invention takes the environmental temperature and glutathione in insects as controlled release signals, can better control insects, and improves the intelligence of the pesticide microsphere, thereby improving the utilization rate.

In order to achieve the second purpose, the invention provides the following technical scheme:

provides a preparation method of the pesticide microsphere with multiple stimulus responses, which comprises the following steps,

step one, preparing modified carboxymethyl cellulose, comprising the following steps:

weighing carboxymethyl cellulose with a formula amount, and fully mixing and dissolving the carboxymethyl cellulose with a PBS solution to obtain a first solution; wherein the PBS solution has a pH of 6;

heating the first solution to 30-50 ℃, then adding a catalyst into the first solution, wherein the catalyst consists of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide, and continuously stirring for 0.5-1.5 h. Activating the carboxyl groups on the carboxymethyl cellulose;

weighing cystamine dihydrochloride according to the formula ratio, and mixing and dissolving the cystamine dihydrochloride with another PBS solution to obtain a second solution;

mixing the second solution with the activated carboxymethyl cellulose, and stirring and reacting for 15-30 h at 30-50 ℃ to obtain a mixed material;

purifying the material, removing the catalyst and unreacted substances, collecting the purified material, and freeze-drying to obtain modified carboxymethyl cellulose, wherein the modified carboxymethyl cellulose is cystamine dihydrochloride modified carboxymethyl cellulose;

step two, preparing pesticide microspheres, comprising the following steps:

fully mixing n-hexadecane, an insecticidal component, isophorone diisocyanate and dibutyltin dilaurate according to the formula ratio to obtain an oil phase;

mixing the modified carboxymethyl cellulose prepared in the step one with deionized water, and mixing and dissolving the mixture with tween to obtain a water phase;

fully homogenizing the oil phase and the water phase at room temperature to obtain emulsion;

and (2) stirring the emulsion at 60-80 ℃ and reacting for 3-6 h to ensure that isophorone diisocyanate and modified carboxymethyl cellulose fully react to obtain a pesticide microsphere solution, and freeze-drying the pesticide microspheres to obtain the pesticide microspheres.

In some embodiments, in step one, the weight ratio of the carboxymethylcellulose, the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride, the N-hydroxysuccinimide, and the cystamine dihydrochloride is 1 (0.1-0.3) to (0.1-0.2) to (1-3).

In some embodiments, the weight ratio of the modified carboxymethyl cellulose, the pesticidal component, the isophorone diisocyanate, the n-hexadecane, and the tween is (0.2-0.6): (0.5-0.7): (1-3): (0.1-0.3).

In some embodiments, in step one, the material is purified by dialysis using a dialysis bag.

In some embodiments, the dialysis bag is a Mw5000 dialysis bag.

In some embodiments, the time for dialysis purification is 40h to 50 h.

In some embodiments, in the second step, the oil phase and the water phase are homogenized for 2min to 6min at 5000 to 15000rpm at room temperature.

In some embodiments, the tween is tween 80.

The preparation method of the multiple stimulus response type pesticide microsphere has the following beneficial effects:

(1) according to the preparation method of the multi-stimulus response type pesticide microsphere, a catalyst is used for activating the carboxyl of carboxymethyl cellulose, and then cystamine dihydrochloride reacts with the activated carboxymethyl cellulose, so that the cystamine dihydrochloride modifies the carboxymethyl cellulose to prepare the redox response type modified carboxymethyl cellulose, and disulfide bonds and amino groups are added to the modified carboxymethyl cellulose, so that a foundation is provided for subsequent targeted controlled release.

(2) According to the preparation method of the multiple stimulus response type pesticide microsphere, the oil phase and the water phase are subjected to interfacial polymerization, so that the easily photolyzed insecticidal component is encapsulated in the pesticide microsphere, the photostability of the pesticide microsphere is improved, and the effectiveness of the pesticide is further improved.

(3) According to the preparation method of the multiple stimulus response type pesticide microsphere, the isophorone diisocyanate is catalyzed by the catalyst, and the catalyzed isophorone diisocyanate can fully react with the amino and the hydroxyl of the modified carboxymethyl cellulose to prepare the pesticide microsphere with a stable structure.

Drawings

FIG. 1 is the reaction process of cystamine dihydrochloride modified carboxymethyl cellulose of the example.

FIG. 2 is the encapsulation and drug loading ratios for different AVM @ CM-SS-PUs of the examples.

FIG. 3 is a Zeta potential diagram for different AVMs @ CM-SS-PUs of the example.

FIG. 4 is a graph of the particle size change after 45 days of storage for various AVM @ CM-SS-PU samples of the examples.

FIG. 5 is the CMC, CYS and CMC-SS-NH of the examples ₂ Respectively correspond to ¹ HNMR picture.

FIG. 6 is CMC and CMC-SS-NH of examples ₂ GPC curve of (1).

FIG. 7 is CMC, CYS, CMC-SS-NH of examples ₂ And infrared spectrograms of CM-SS-PU, AVM and AVM @ CM-SS-PU.

FIG. 8 is a graph of the UV degradation curves of the aqueous AVM dispersion of the examples, a commercially available AVM emulsifiable concentrate and various AVM @ CM-SS-PU; (b) and (5) fitting a degradation curve graph by first-order dynamics.

Figure 9 is a graph of the response release of the example AVM @ CM-SS-PU to stimulation by (a) temperature, (b) glutathione, (c) pH, (d) cellulase, (e) urease.

FIG. 10 shows insecticidal activity of AVM technical, AVM commercial emulsifiable concentrate and AVM @ CM-SS-PU against plutella xylostella in the examples.

FIG. 11 is a schematic diagram showing insecticidal activities of AVM technical, AVM commercial emulsifiable concentrate, AVM @ CM and AVM @ CM-SS-PU of examples after (a)24h and (b)48h at different temperatures, and (c) intelligent control of pests by pesticide microcapsules at different temperatures.

Detailed Description

Preferred embodiments of the present invention will be described in more detail below with reference to the accompanying drawings. While the preferred embodiments of the present invention are shown in the drawings, it should be understood that the present invention may be embodied in various forms and should not be limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete, and will fully convey the scope of the invention to those skilled in the art.

The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. As used in this disclosure and the appended claims, the singular forms "a", "an", and "the" are intended to include the plural forms as well, unless the context clearly indicates otherwise. It should also be understood that the term "and/or" as used herein refers to and encompasses any and all possible combinations of one or more of the associated listed items.

It is to be understood that, although the terms first, second, third, etc. may be used herein to describe various information, such information should not be limited by these terms. These terms are only used to distinguish one type of information from another. For example, first information may also be referred to as second information, and similarly, second information may also be referred to as first information, without departing from the scope of the present invention. Thus, a feature defined as "first" or "second" may explicitly or implicitly include one or more of that feature. In the description of the present invention, "a plurality" means two or more unless specifically defined otherwise.

Example 1

Diamondback moth is the most harmful pest to crucifers in the world and causes a worldwide economic loss of about $ 4 to $ 50 billion each year. The temperature is an important factor influencing the growth of insects and microorganisms, and researches show that the growth and development speed of the diamondback moth is accelerated along with the temperature rise in the range of 17.5-32.5 ℃, so that a controlled release preparation which is quickly released along with the temperature rise in the range needs to be designed to cope with the higher population density of the diamondback moth at high temperature. On the other hand, the microcapsule has longer service life and biosafety, but the slow release of the microsphere in a living body causes low bioactivity of the microcapsule, and is an important factor for limiting the use of the microcapsule. Wherein, the micro environment of the insects is used as a stimulus factor to control the rapid release of the pesticide in the insects, which is a high-efficiency pest killing mode. The plutella xylostella is rich in glutathione (an antioxidant mercaptan) and has a unique alkaline environment in the midgut, and the stimulation factors can be used as a key to open a switch on a controlled-release pesticide carrier. In consideration of the complex environment of the pesticide in practical application, an intelligent pesticide microcapsule with multiple responsiveness needs to be designed, so that the intelligent pesticide microcapsule has longer lasting effect after being sprayed, and can quickly release effective ingredients for controlling plant diseases and insect pests under the biotic and abiotic stresses of crops. The present embodiment is merely an example in which the processing object is a diamond back moth.

The multi-stimulus response type pesticide microsphere disclosed in this example comprises modified carboxymethyl cellulose, an insecticidal component, isophorone diisocyanate, and n-hexadecane, wherein the modified carboxymethyl cellulose comprises carboxymethyl cellulose and cystamine dihydrochloride.

According to the multiple stimulus response type pesticide microsphere, the pesticide microsphere consists of the oil phase component and the water phase component, the oil phase component and the water phase component can form interfacial polymerization, the interfacial polymerization microsphere can efficiently encapsulate the insecticidal component, the ultraviolet light stability of the insecticidal component in the microsphere is greatly improved, the stability of the insecticidal component under ultraviolet light is better, and the longer-acting insecticidal effect is achieved. The modified carboxymethyl cellulose is cystamine dihydrochloride modified carboxymethyl cellulose, so that amino, hydroxyl and urea bonds exist on the shell of the pesticide microsphere, the groups form hydrogen bonds with fatty acid, fatty alcohol and fatty aldehyde of a wax layer, and acting force between the groups and the leaf surface can effectively prevent microcapsules on the leaf surface from being washed by rainwater, and the durability of the pesticide microsphere is improved. The pesticide microspheres contain n-hexadecane, and the n-hexadecane is converted from a solid state to a liquid state at the temperature of higher than 20 ℃, so that the pesticide microspheres can accelerate the release of pesticides in the microspheres at high temperature, further more pesticides can be released at high temperature, and the problem that pests and diseases are easy to outbreak at high temperature can be better prevented and treated; on the contrary, in a low-temperature environment, n-hexadecane is not easy to be converted into a liquid state, the pesticide released by the pesticide microspheres is less, and pests are less at the moment, so that the pesticide dosage can be saved, the targeted controlled release of the pesticide is realized, and the effective period, the utilization rate and the environmental friendliness of the pesticide microspheres are improved. The modified carboxymethyl cellulose is prepared by modifying carboxymethyl cellulose with cystamine dihydrochloride, so that disulfide bonds are contained on a spherical shell of the pesticide microsphere, after the insect eats the pesticide microsphere, the disulfide bonds on the pesticide microsphere can be reduced and destroyed by rich glutathione in the insect body, so that the pesticide microsphere is disintegrated and the effective components are quickly released, the quick insect killing effect is achieved, the pesticide microsphere has higher insecticidal activity due to the stimulated release of microenvironment in the insect body, and the effect of releasing according to needs is achieved. The environment temperature and glutathione in the insects are used as controlled release signals, so that the insect control can be better aimed at controlling the insects, the intelligence of the pesticide microspheres is improved, and the utilization rate is improved.

In this embodiment, the insecticidal component is one or more of abamectin, ivermectin, chlorpyrifos, 2, 4-dichlorophenoxyacetic acid, emamectin benzoate, chlorantraniliprole and lambda-cyhalothrin. In practical application, other insecticidal components can be adopted according to actual needs, and are not limited herein.

The preparation method of the multi-stimulus response type pesticide microsphere comprises the following steps,

heating the first solution to 30-50 ℃, then adding a catalyst to the first solution, wherein the catalyst consists of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide, and continuously stirring for 0.5 h. Activating the carboxyl groups on the carboxymethyl cellulose;

mixing the second solution with the activated carboxymethyl cellulose, and stirring and reacting for 15 hours at the temperature of 30 ℃ to obtain a mixed material;

step two, preparing pesticide microspheres, which comprises the following steps:

and (2) reacting the emulsion for 3 hours at 60 ℃ while stirring, so that isophorone diisocyanate and modified carboxymethyl cellulose fully react to obtain a pesticide microsphere solution, and freeze-drying the pesticide microspheres to obtain the pesticide microspheres.

According to the preparation method of the multi-stimulus response type pesticide microsphere, the catalyst is used for activating the carboxyl of the carboxymethyl cellulose, and then the cystamine dihydrochloride reacts with the activated carboxymethyl cellulose, so that the cystamine dihydrochloride modifies the carboxymethyl cellulose to prepare the redox response type modified carboxymethyl cellulose, and the modified carboxymethyl cellulose increases disulfide bonds and amino groups, so that a foundation is provided for subsequent targeted controlled release. The method adopts the interfacial polymerization of the oil phase and the water phase, so that the insecticidal components which are easy to photolyze are encapsulated in the pesticide microspheres, the photostability of the pesticide microspheres is improved, and the effectiveness of the pesticide is further improved. The method adopts a catalyst to catalyze isophorone diisocyanate, and the catalyzed isophorone diisocyanate can fully react with amino and hydroxyl of modified carboxymethyl cellulose to prepare the pesticide microsphere with a stable structure.

In this example, in the first step, the weight ratio of the carboxymethyl cellulose to the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride to the N-hydroxysuccinimide to the cystamine dihydrochloride is 1:0.1:0.1: 1. The weight ratio can be adjusted according to the actual requirement, and is not limited herein.

In this example, the weight ratio of the modified carboxymethyl cellulose, the pesticidal component, the isophorone diisocyanate, the n-hexadecane, and the tween was 0.2:0.2:0.5:1: 0.1. The weight ratio can be adjusted according to the actual requirement, and is not limited herein.

In this embodiment, in the first step, a dialysis bag is used to perform dialysis and purification on the material. This dialysis purification procedure removed the catalyst and unreacted cystamine dihydrochloride.

In this embodiment, the dialysis bag is a Mw5000 dialysis bag.

In this example, the dialysis purification time was 40 hours. The dialysis purification time can be selected according to the actual situation and is not limited herein.

In the second step of the present example, the oil phase and the aqueous phase were homogenized at 5000rpm for 2min at room temperature. This operation enables the emulsion to be produced quickly.

In this example, the tween is tween 80.

Example 2

The multi-stimulus response type pesticide microsphere disclosed in this example includes modified carboxymethyl cellulose, an insecticidal component, isophorone diisocyanate, and n-hexadecane, where the modified carboxymethyl cellulose includes carboxymethyl cellulose and cystamine dihydrochloride.

According to the multi-stimulus response type pesticide microsphere, the pesticide microsphere consists of the oil phase component and the water phase component, the oil phase component and the water phase component can form interfacial polymerization, the interfacial polymerization microsphere can efficiently encapsulate the insecticidal component, the ultraviolet light stability of the insecticidal component in the microsphere is greatly improved, the stability of the insecticidal component under ultraviolet light is better, and the longer-acting insecticidal effect is achieved. The modified carboxymethyl cellulose is cystamine dihydrochloride modified carboxymethyl cellulose, so that amino, hydroxyl and urea bonds exist on the shell of the pesticide microsphere, the groups form hydrogen bonds with fatty acid, fatty alcohol and fatty aldehyde of a wax layer, and acting force between the groups and the leaf surface can effectively prevent microcapsules on the leaf surface from being washed by rainwater, and the durability of the pesticide microsphere is improved. The pesticide microspheres contain n-hexadecane, and the n-hexadecane is converted from a solid state to a liquid state at the temperature of higher than 20 ℃, so that the pesticide microspheres can accelerate the release of pesticides in the microspheres at high temperature, further more pesticides can be released at high temperature, and the problem that pests and diseases are easy to outbreak at high temperature can be better prevented and treated; on the contrary, in a low-temperature environment, n-hexadecane is not easy to be converted into a liquid state, the pesticide released by the pesticide microspheres is less, and pests are less at the moment, so that the pesticide dosage can be saved, the targeted controlled release of the pesticide is realized, and the effective period, the utilization rate and the environmental friendliness of the pesticide microspheres are improved. The modified carboxymethyl cellulose is prepared by modifying carboxymethyl cellulose with cystamine dihydrochloride, so that disulfide bonds are contained on a spherical shell of the pesticide microsphere, after the insect eats the pesticide microsphere, the disulfide bonds on the pesticide microsphere can be reduced and destroyed by rich glutathione in the insect body, so that the pesticide microsphere is disintegrated and the effective components are quickly released, the quick insect killing effect is achieved, the pesticide microsphere has higher insecticidal activity due to the stimulated release of microenvironment in the insect body, and the effect of releasing according to needs is achieved. The environmental temperature and glutathione in the insects are used as controlled release signals, so that the insect control can be better aimed at preventing and controlling the insects, the intelligence of the pesticide microspheres is improved, and the utilization rate is improved.

the first solution was heated to 50 ℃ and then a catalyst consisting of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide was added to the first solution with continuous stirring for 1.5 h. Activating the carboxyl groups on the carboxymethyl cellulose;

mixing the second solution with the activated carboxymethyl cellulose, and stirring and reacting for 30 hours at 50 ℃ to obtain a mixed material;

step two, preparing pesticide microspheres, comprising the following steps:

and (2) reacting the emulsion for 6 hours at 80 ℃ while stirring, so that isophorone diisocyanate and modified carboxymethyl cellulose fully react to obtain a pesticide microsphere solution, and freeze-drying the pesticide microspheres to obtain the pesticide microspheres.

In this example, in step one, the weight ratio of the carboxymethyl cellulose to the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride to the N-hydroxysuccinimide to the cystamine dihydrochloride is 1:0.3:0.2: 3. The weight ratio can be adjusted according to practical requirements, and is not limited herein.

In this example, the weight ratio of the modified carboxymethyl cellulose, the pesticidal component, the isophorone diisocyanate, the n-hexadecane, and the tween was 0.6:0.6:0.7:3: 0.3. The weight ratio can be adjusted according to the actual requirement, and is not limited herein.

In this embodiment, in the first step, a dialysis bag is used to dialyze and purify the material. This dialysis purification procedure removed the catalyst and unreacted cystamine dihydrochloride.

In this embodiment, the dialysis bag is a Mw5000 dialysis bag.

In this example, the dialysis purification time was 50 hours. The dialysis purification time can be selected according to the actual situation and is not limited herein.

In the present example, in the second step, the oil phase and the aqueous phase were homogenized at 15000rpm for 6min at room temperature. This operation enables the emulsion to be produced quickly.

In this example, the tween is tween 80.

Example 3

According to the multi-stimulus response type pesticide microsphere, the pesticide microsphere consists of the oil phase component and the water phase component, the oil phase component and the water phase component can form interfacial polymerization, the interfacial polymerization microsphere can efficiently encapsulate the insecticidal component, the ultraviolet light stability of the insecticidal component in the microsphere is greatly improved, the stability of the insecticidal component under ultraviolet light is better, and the longer-acting insecticidal effect is achieved. The modified carboxymethyl cellulose is cystamine dihydrochloride modified carboxymethyl cellulose, so that amino, hydroxyl and urea bonds exist on the shell of the pesticide microsphere, the groups form hydrogen bonds with fatty acid, fatty alcohol and fatty aldehyde of a wax layer, and acting force between the groups and the leaf surface can effectively prevent microcapsules on the leaf surface from being washed by rainwater, and the durability of the pesticide microsphere is improved. The pesticide microspheres contain n-hexadecane, and the n-hexadecane is converted from a solid state to a liquid state at the temperature higher than 20 ℃, so that the pesticide microspheres can accelerate the release of pesticides in the microspheres at high temperature, further more pesticides can be released at high temperature, and the problem that plant diseases and insect pests are easy to explode at high temperature can be better prevented; on the contrary, in a low-temperature environment, n-hexadecane is not easy to be converted into a liquid state, the pesticide released by the pesticide microspheres is less, and pests are less at the moment, so that the pesticide dosage can be saved, the targeted controlled release of the pesticide is realized, and the effective period, the utilization rate and the environmental friendliness of the pesticide microspheres are improved. The modified carboxymethyl cellulose is prepared by modifying carboxymethyl cellulose with cystamine dihydrochloride, so that disulfide bonds are contained on a spherical shell of the pesticide microsphere, after the insect eats the pesticide microsphere, the disulfide bonds on the pesticide microsphere can be reduced and destroyed by rich glutathione in the insect body, so that the pesticide microsphere is disintegrated and the effective components are quickly released, the quick insect killing effect is achieved, the pesticide microsphere has higher insecticidal activity due to the stimulated release of microenvironment in the insect body, and the effect of releasing according to needs is achieved. The environmental temperature and glutathione in the insects are used as controlled release signals, so that the insect control can be better aimed at preventing and controlling the insects, the intelligence of the pesticide microspheres is improved, and the utilization rate is improved.

heating the first solution to 30-50 ℃, then adding a catalyst to the first solution, wherein the catalyst consists of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide, and continuously stirring for 1 h. Activating the carboxyl groups on the carboxymethyl cellulose;

mixing the second solution with the activated carboxymethyl cellulose, and stirring and reacting for 20 hours at 40 ℃ to obtain a mixed material;

step two, preparing pesticide microspheres, comprising the following steps:

and (3) stirring the emulsion at 70 ℃ and reacting for 4h to ensure that isophorone diisocyanate and modified carboxymethyl cellulose fully react to obtain a pesticide microsphere solution, and freeze-drying the pesticide microspheres to obtain the pesticide microspheres.

In this example, in the first step, the weight ratio of the carboxymethyl cellulose to the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride to the N-hydroxysuccinimide to the cystamine dihydrochloride is 1:0.2:0.15: 2. The weight ratio can be adjusted according to the actual requirement, and is not limited herein.

In this example, the weight ratio of the modified carboxymethyl cellulose, the pesticidal component, the isophorone diisocyanate, the n-hexadecane, and the tween was 0.5:0.5:0.6:2: 0.2. The weight ratio can be adjusted according to the actual requirement, and is not limited herein.

In this embodiment, the dialysis bag is a Mw5000 dialysis bag.

In this example, the dialysis purification time was 40 to 50 hours. The dialysis purification time can be selected according to the actual situation and is not limited herein.

In the second step of the present example, the oil phase and the aqueous phase are sufficiently homogenized at 10000rpm for 5min at room temperature. This operation enables an emulsion to be produced quickly.

In this example, tween is tween 80.

Performance verification and structural characterization:

in order to verify the performance and structure of the pesticide microspheres of the invention, the control pesticide microspheres 1 and the control pesticide microspheres 2 are prepared in a small yield by an experimental operation mode and are prepared according to the following steps:

1. the preparation method of the pesticide microspheres comprises the following steps:

preparation of a Redox-responsive modified carboxymethyl cellulose (CMC-SS-NH) ₂ )：

1g of carboxymethylcellulose (CMC) was weighed out and dissolved in a 35ml PBS solution (pH 6) with good stirring, 287mg of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) (1.5mmol) and 173mg of N-hydroxysuccinimide (NHS) (1.5mmol) were added to the CMC solution at 40 ℃ and stirring was continued for 1h to activate the carboxyl groups on the CMC. Then, 2g of cystamine dihydrochloride (CYS) was dissolved in a 10ml pbs solution (pH 6) and added to the above CMC solution, and the reaction was stirred at 40 ℃ for 24 hours. Finally, the obtained sample is dialyzed in a dialysis bag (Mw5000) for 48h to remove the catalyst and unreacted CYS, and freeze-dried to obtain the purified redox-responsive modified carboxymethyl cellulose (CMC-SS-NH) ₂ )。

The reaction formula of cystamine dihydrochloride modified carboxymethyl cellulose is shown in fig. 1.

Preparation of stimulating multiple stimulus response type pesticide microcapsule (AVM @ CM-SS-PU)

The multiple response type pesticide microcapsule AVM @ CM-SS-PU loaded with Abamectin (AVM) is prepared by an interfacial polymerization method. Gym for doing thingsThe process flow is as follows: to 2.8g of n-hexadecane were added 0.2g of AVM, 0.62g of isophorone diisocyanate (IPDI), and 0.1g of dibutyltin dilaurate (DBTDL) and mixed thoroughly to form an oil phase. Adding 0.5g of CMC-SS-NH into 25mL of deionized water ₂ And 0.1g of Tween80 (Tween80) were dissolved with stirring to form an aqueous phase, and the oil phase and the aqueous phase were mixed and homogenized at room temperature at 10000rpm for 3 min. Transferring the homogenized emulsion into a flask, reacting for 4h under magnetic stirring at 70 ℃ to ensure that IPDI isocyanate groups fully react with CMC-SS-NH2 amino and hydroxyl groups, and preparing the pesticide microcapsule (AVM @ CM-SS-PU) by interfacial polymerization

2. The contrast pesticide microsphere 1 is a blank response type microcapsule CM-SS-PU without AVM, and the preparation method of the contrast pesticide microsphere 1 is different from the preparation method of the AVM @ CM-SS-PU of the invention in that no insecticidal component AVM is added.

3. The contrast pesticide microsphere 2 is prepared by replacing CMC-SS-NH with CMC _{2 of} The difference between the preparation method of the response type pesticide microcapsule AVM @ CM and the preparation method of the AVM @ CM-SS-PU of the invention is that CMC-SS-NH is saved ₂ Preparation of (1) CMC replacing CMC-SS-NH ₂ 。

Encapsulation Performance test

The testing steps are as follows: 1. weighing 20mg of freeze-dried drug-loaded microspheres, placing the drug-loaded microspheres in a centrifuge tube, adding 4mL of ethanol, shaking up, centrifuging the mixture for 5min at 10000rpm, taking 1mL of supernatant, and metering the volume to 25 mL. The absorbance A of the free AVM was measured using an ultraviolet spectrophotometer at a wavelength of 245 nm. The standard curve a ═ 0.03321C-0.006 (R) was used ² 0.999) the mass of free AVM in the supernatant was determined. Then, Encapsulation Efficiency (EE) and drug loading efficiency (LC) of AVM were calculated using formulas (4-1) and (4-2).

In the formula, m _totalAVM Is the total mass of AVM in the system, m _freeAVM Is the mass of unencapsulated free AVM in the supernatant after centrifugation, m _AVM@CM-SS-PU Is the total mass of the drug-loaded microspheres.

2. The average particle diameter, particle distribution index and Zeta potential of the sample were measured using a laser particle size analyzer. Measurements were performed by Dynamic Light Scattering (DLS) technique at a scattering angle of 90 °, the measurements were repeated three times and averaged.

As a result: as shown in figure 2, in the process of preparing the microcapsule, other conditions are controlled to be unchanged, the drug loading rate (LC) of the microcapsule is improved along with the increase of the abamectin added into the oil phase, the highest drug loading rate (LC) can reach 8.02 +/-0.18 percent of AVM @ CM-SS-PU0.6, the encapsulation rate (EE) is reduced along with the increase of the abamectin, but the AVM @ CM-SS-PU0.6 can still reach 76.98 +/-1.72 percent, and the AVM @ CM-SS-PU0.2 with the highest encapsulation rate can reach 88.23 +/-0.69 percent, which shows that the AVM can be efficiently encapsulated by the microencapsulation of interfacial polymerization. FIG. 3 shows the Zeta potentials of various samples, the Zeta potentials of CMC molecular chains are increased to-58.94 + -0.69 mV due to the existence of a large number of carboxyl groups, and CMC-SS-NH is subjected to grafting reaction after a large number of carboxyl groups are consumed ₂ The potential of (2) was raised to-30.10. + -. 0.26mV, which also reflects the successful progress of the grafting reaction (Butunet. 2011). The potential of the blank microcapsule CM-SS-PU is-37.77 +/-0.71 mV, and the potential of the drug-loaded microspheres is reduced from-42.22 +/-1.06 mV of AVM @ CM-SS-PU0.2 to-48.08 +/-0.69 mV of AVM @ CM-SS-PU0.6 with the increase of the added AVM during interfacial polymerization, probably because the lower encapsulation rate of AVM @ CM-SS-PU0.6 causes more negatively charged AVM to be deposited on the surfaces of the microspheres. FIG. 4 shows the particle size distributions of three samples of AVM @ CM-SS-PU, AVM @ CM-SS-PU0.2, AVM @ CM-SS-PU0.4 and AVM @ CM-SS-PU0.6 having particle sizes of 5.40. + -. 0.76. mu.m, 4.79. + -. 0.73. mu.m, 3.90. + -. 0.92. mu.m, and particle size distribution indices of 0.312. + -. 0.025, 0.302. + -. 0.023 and 0.419. + -. 0.034, respectively, the particle size distributions of the three samples slightly broadened but did not significantly change after 45 days of storage, and no significant precipitation and phase separation was observed in the true graph of AVM CM-SS-PU, indicating that AVM @ CM-SS-PU has good storage stability.

Nuclear magnetic hydrogen spectrum analysis

The samples were tested at D using a NMR spectrometer at 400MHz ₂ O or CDCl ₃ Nuclear magnetic hydrogen spectroscopy to analyze CMC-SS-NH ₂ CYS and CMC, the results of which are shown in FIG. 5, in the CMC spectrum, the peaks at 3.05-3.35ppm (H2), 3.45-4.22ppm (H3,4,5,7), 4.34-4.58ppm (H1,6) represent the proton signals of each carbon on the glucose ring, and 4.70ppm is solvent D ₂ Peak of O (Kono, 2013). The CYS spectrum shows obvious peaks at 2.95ppm (a) and 3.34ppm (b), which are respectively attributed to proton signals of two methylene groups in the CYS structure. At CMC-SS-NH ₂ In the spectra, in addition to the proton signal of CMC, a methylene signal belonging to CYS appears at 2.93ppm (a ') and 3.31ppm (b'), and in addition, the proton peak at 2.79ppm (c) belongs to the methylene group of CMC, which is in agreement with the previous report (parketal, 2017). The results show that the CYS successfully modifies the CMC to synthesize the CMC-SS-NH ₂ 。

Molecular weight analysis of modified carboxymethylcellulose

The molecular weight (Mw) and Polymer Dispersion Index (PDI) of the samples were measured using a gel permeation chromatograph. A2 mg/mL sample was dissolved in water and passed through a two-column PLaquagel-OH (7.5 mm. times.300 mm,8 μm) mixed column at 30 ℃ in a 100 μ L volume with water as eluent at a flow rate of 1 mL/min.

As shown in FIG. 6, the modified product CMC-SS-NH was analyzed by GPC ₂ The molecular weight of CMC is 32.7X 10 compared to the molecular weight change of CMC ⁴ g/mol, and CMC-SS-NH ₂ The molecular weight of (A) is increased to 38.3X 10 ⁴ g/mol，CMC-SS-NH ₂ The PDI of (a) was also increased from 3.819 to 4.328 for CMC, indicating that CYS modification increased the molecular weight of CMC while also broadening the molecular weight distribution. The grafting yield of CYS was calculated to be 17.13% from the change in molecular weight. In addition, CMC-SS-NH was measured by an elemental analyzer ₂ The content of the S element in the resin was 7.83%, and the grafting ratio of CYS obtained by conversion by calculation was 18.49%, which was in agreement with the results of the grafting ratio measured by GPC.

Infrared spectroscopic analysis

Grinding the dried sample to be detected and potassium bromide into fine powder and pressing into slices, and scanning for 32 times by a Fourier infrared spectrometer to obtain a sample with a density of 450cm ^-1 -4000cm ^-1 Infrared spectrum of the wavelength range.

The chemical structure changes of various substances in the synthesis process of the pesticide microspheres are characterized by Fourier infrared spectroscopy, and the result is shown in FIG. 7. 3000cm in CMC spectrogram ^-1 The above signal is due to the stretching vibration of hydroxyl group and is at 1603cm ^-1 And 1422cm ^-1 The peak of (A) is an asymmetric and symmetric stretching vibration about the carboxyl group, 1328cm ^-1 The peak of (a) then represents the bending vibration of the hydroxyl group, consistent with previous literature reports (chenet al, 2018). CMC-SS-NH compared with CMC ₂ The spectrum is 1644cm ^-1 And 1460cm ^-1 A new absorption peak appeared, of which 1644cm ^-1 Representing a stretching vibration peak of-C ═ O (amide absorption band I) in an amide bond, 1460cm ^-1 The C-N stretching vibration in CYS indicates that CMC successfully reacts with CYS and synthesizes CMC-SS-NH with disulfide bonds and reactive amino groups ₂ . After further synthesizing cellulose microspheres by interfacial polymerization, 1641CM in CM-SS-PU spectrogram ^-1 And 1566cm ^-1 Absorption peaks respectively representing-C ═ O (amide absorption band I) and N-H bending vibration (amide absorption band II) due to isocyanate group and CMC-SS-NH ₂ The amino group and the hydroxyl group form a urea bond and a urethane bond. 2267cm ^-1 Signals of (2) are some unreacted isocyanate groups in CM-SS-PU, 2924CM ^-1 The peak of (a) is caused by C-H oscillation of methylene group in n-hexadecane. After the AVM is added into the system, the spectrum of the AVM @ CM-SS-PU is 1734CM ^-1 And (3) a characteristic absorption peak of C ═ O in the AVM structure appears, which indicates that the stimulus-responsive pesticide microspheres are successfully synthesized.

Analysis of ultraviolet resistance

The AVM ethanol solution is added into deionized water and stirred to prepare AVM water dispersion with the concentration of 500mg/L, and 5 percent of abamectin commercial emulsifiable concentrate is also diluted to the concentration of 500mg/L which is the same as that of AVM @ CM-SS-PU by the deionized water. 50mL of the different sample solutions were added to the tube and placed in the photochemical reactor, each sample being 5cm from the mercury lamp (300W, Emax. 365 nm). After ultraviolet irradiation for a certain time interval, 1mL of the sample solution was taken out and placed in a brown volumetric flask, and the volume was adjusted to 10mL with absolute ethanol, and then the absorbance was measured at a wavelength of 245nm using an ultraviolet-visible spectrophotometer. Equations (2-4) were used to calculate the residual rate of AVM in the sample (RR) after different times of irradiation.

In the formula, C _t Represents the concentration of AVM in the sample after a specific time of irradiation, C ₀ Represents the concentration of AVM in the sample prior to irradiation.

Due to the special sixteen-membered macrolide structure of the abamectin, the abamectin is sensitive to ultraviolet light and the ultraviolet light, and the activity of the abamectin is rapidly reduced due to the breakage of a macrocyclic ring. The avermectin has short half-life period in the practical application process, so that the light stability of the avermectin is urgently needed to be improved. The photostability of different AVM formulations was tested using photoreactor, and the results are shown in FIG. 8, where AVM aqueous dispersion and AVM commercially available emulsifiable concentrate rapidly decomposed under UV light, leaving only 18.48 + -0.23% and 26.16 + -0.05% after 370h irradiation, respectively, indicating that they do not have UV resistance. And after the AVM @ CM-SS-PU0.2, the AVM @ CM-SS-PU0.4 and the AVM @ CM-SS-PU0.6 are irradiated for 370 hours, 65.04 +/-0.20%, 57.57 +/-0.08% and 37.35 +/-0.07% still remain, the encapsulation efficiency is the main reason for the difference of the uvioresistant performance of the AVM @ CM-SS-PU, and compared with other two unencapsulated AVM preparations, the three AVM @ CM-SS-PUs have obvious uvioresistant performance and can be stabilized for a long time under ultraviolet light.

The UV degradation data of these AVM formulations was fitted using first order kinetics and the results are shown in FIG. 8 and Table 1, AVM @ CM-SS-PU0.2 possesses the best UV resistance, its half-life under UV (t @) _1/2) 550.11min, while the half-life of the aqueous AVM dispersion is 89.32min, therefore, after encapsulation, the light stability of AVM @ CM-SS-PU0.2 is improved by more than 5 times compared with that of the aqueous AVM dispersion. AVM which is easy to photolyze is encapsulated in the polymer microsphere, and a thicker shell layer can effectively resist ultraviolet light through physical barrier; on the other hand, chromophoric groups such as carboxyl and carbonyl on the carrier can absorb part of ultraviolet light to provide better photostability for AVM (Haoetal., 2020). The AVM @ CM-SS-PU with good light stability and adhesion can be stably stored on leaves after being sprayed, and long-acting effect is provided for crop growthThe protective measures of (1) effectively reduce the use frequency and the use dosage of the pesticide.

TABLE 1 first order kinetics fitting degradation results

Pesticide microcapsule stimulation release performance analysis

The testing steps are as follows: 5mL of the sample solution were placed in dialysis bags (MWCO. RTM. 5000Da) in 50mL brown flasks with different pH 40% ethanol in water as release medium. Placing the conical flask in a shaker at 25 ℃ to shake, taking a 1mL volumetric flask of the release medium at a specific time interval, using 40% ethanol water solution to perform constant volume till 10mL, and adding the corresponding release medium with the same volume back to the conical flask. The absorbance of the diluted solution at 245nm was measured by a UV spectrophotometer by passing through a standard curve A of 0.0281C-0.00182 (R) ² 0.999) and equation (2-5) for calculating cumulative release rate (R) _i )。

In the formula: c. C _i Represents the concentration (mg/L) of AVM in the release medium at a particular time, m _AVM Representing the total mass of AVM added to the dialysis bag.

As a result: figure 9(a) shows the release characteristics of AVM @ CM-SS-PU under temperature stimulation, and from the release curves of three temperatures, the release speed of the drug-loaded microcapsule is accelerated along with the temperature rise, and the release is accumulated for 33.67%, 38.77% and 49.61% after 465h at 15 ℃, 25 ℃ and 35 ℃. The phase transition temperature of the n-hexadecane in the core of the AVM @ CM-SS-PU microcapsule is 20 ℃, when the microcapsule is placed in an environment with the temperature higher than 20 ℃, the n-hexadecane in the microcapsule is converted into a flowable liquid state from a solid state, the dissolution rate of the AVM in the microcapsule is accelerated, and therefore the AVM @ CM-SS-PU can be released in an accelerated manner in an environment with higher temperature. The release of the temperature stimulation shows that the AVM @ CM-SS-PU is expected to regulate and control the release rate according to the environmental temperature in the practical application process so as to adapt to the growth and propagation speeds of plant diseases and insect pests at different temperatures.

In order to achieve the purpose of quickly releasing the pesticide microcapsule in the body of pests, the microcapsule with disulfide bonds as a capsule wall material is designed, so that the microcapsule can quickly release active ingredients under the stimulation of a reducing condition. FIG. 9(b) shows the release behavior of AVM @ CM-SS-PU under stimulation by glutathione, wherein AVM @ CM-SS-PU is rapidly released in a reducing environment containing glutathione, and the release of AVM can be accelerated by increasing the concentration of glutathione, and 21h stimulation of AVM @ CM-SS-PU in a 0mg/mL, 1mg/mL, 2mg/mL glutathione environment respectively cumulatively releases 16.63%, 29.20%, 52.97%, 1mg/mL and 2mg/mL glutathione respectively increases the early release rate of AVM @ CM-SS-PU by 2-fold and 3-fold, respectively, and cumulatively releases 79.67%, 98.10% after 465 h. Therefore, after the AVM @ CM-SS-PU microcapsule with reduction response is applied, the AVM can be slowly released from the microcapsule to avoid the active ingredients from being decomposed by light, and after the AVM microcapsule is ingested and eaten by pests, the AVM can be quickly released in a reduction environment provided by rich glutathione in the pests, so as to achieve the effect of quickly controlling the pests (Liangetal, 2020).

FIG. 9(c) shows the release behavior of AVM @ CM-SS-PU at different pH values, and the drug-loaded microcapsules release faster at pH3 and pH9 than at pH 7. The rapid release under acidic conditions is attributed to the fact that the protonation of the amino groups increases the electrostatic repulsion between the molecular chains, resulting in microcapsule swelling that accelerates the release of AVM. Cleavage of urea bond under alkaline conditions and OH ^- Nucleophilic attack of the disulfide bond by ions leads to cleavage of the disulfide bond, which is responsible for the rapid release of the microcapsules in alkaline environments (Liangelet al, 2020; Dopieralskietal, 2017). Acidic conditions are the optimal microenvironment for the growth and reproduction of many plant pathogens, and release in an acidic environment facilitates the targeted release of AVM @ CM-SS-PU at the site of plant pathology (camaraetal, 2019). The midgut of phytophagous lepidoptera pests has unique alkaline stripsThese conditions favor the rapid release of AVM @ CM-SS-PU in the midgut to achieve pest killing.

When an organism is attacked by a plant pathogenic fungus, the pathogenic fungus releases cellulase and pectinase for degrading the cell wall, and the cellulase can hydrolyze cellulose into small glucose monomers (gaoetal, 2021). FIG. 9(d) shows the release behavior of AVM @ CM-SS-PU under cellulase stimulation, and it can be seen that the release rate and cumulative release rate of the microcapsules under cellulase stimulation are higher than those without enzyme stimulation, and therefore this cellulase-responsive release pesticide system is expected to be useful for delivering pesticides when plants are attacked by pathogens.

Urease is widely present in microorganisms and tissues of animals and plants, and plant roots secrete urease and degrade urea into a nitrogen source fertilizer, so urease-triggered pesticide systems can be used to treat plant root diseases (polacoet al, 1989; wenet al, 2020). Because of the existence of CMC-SS-NH on the wall of the AVM @ CM-SS-PU ₂ The amino group of (A) and the isocyanate group of IPDI (B) synthesized a urea linkage (-NH-CO-NH-), thus the release behavior of AVM @ CM-SS-PU under urease stimulation was investigated. FIG. 9(e) shows the release profiles of 18.35%, 38.18%, and 55.52% after 45h release of AVM @ CM-SS-PU in 0mg/mL, 1mg/mL, and 2mg/mL urease environments, respectively, indicating that urease can break the urea bond in AVM @ CM-SS-PU, and facilitate release of AVM by changing the structure of the microcapsules. Therefore, the AVM @ CM-SS-PU can be used for targeted treatment of crop root diseases by using urease as a stimulus factor.

FIG. 9AVM @ CM-SS-PU response to release under stimulation by (a) temperature, (b) glutathione, (c) pH, (d) cellulase, (e) urease

Insecticidal Activity assay

As an insecticide, the biological activity of the AVM preparation is one of the most important indicators, and therefore the biological activity of the agent is evaluated by testing the insecticidal effect of the preparation on plutella xylostella using the leaf dipping method. As can be seen from fig. 10, the insecticidal activity of all three formulations increased with increasing concentration of the active ingredient. In the lower concentration range of 0.625-2.5mg/L, the diamondback moth mortality rate of AVM commercial emulsifiable concentrate and AVM @ CM-SS-PU is higherHigher than the original AVM drug and from Table 2, the semilethal concentrations (LC) of the commercial AVM emulsifiable concentrate and AVM @ CM-SS-PU are known ₅₀ ) 3.44mg/L and 3.54mg/L, respectively, are less than 6.01mg/L of the bulk AVM, probably because the commercial emulsifiable concentrate of AVM and AVM @ CM-SS-PU possess smaller contact angles and better liquid retention rates, and therefore more active ingredient is deposited on the fed leaf for killing diamondback moth. On the other hand, the encapsulated AVM @ CM-SS-PU may be better attached to the bodies of diamondback moths (Suetal, 2020), which increases the contact toxicity of AVM @ CM-SS-PU against diamondback moths, and thus the AVM @ CM-SS-PU has higher biological activity.

TABLE 2 insecticidal Activity results for AVM technical, AVM commercial emulsifiable concentrate and AVM @ CM-SS-PU

Research shows that the growth speed of eggs, larvae and pupae of the diamondback moth increases along with the temperature rise in the temperature range of 17.5 ℃ to 32.5 ℃. Therefore, when the temperature is higher than 18 ℃, insect pests caused by diamondback moth are easier to erupt due to the shortening of the development period. Therefore, the phase-change material n-hexadecane is used as a core to be encapsulated in the polymer microsphere, and the pesticide microsphere is endowed with a temperature response property.

It is known from the release profile of AVM @ CM-SS-PU microspheres that an increase in temperature promotes release of AVM within the microspheres. The biological activity of the temperature response type pesticide microspheres on diamond back moths at different temperatures is further explored, and the result is shown in fig. 11. After the leaves soaked with different AVM preparations are fed with the plutella xylostella for 24 hours (figure 11(a)), compared with two microcapsule preparations (AVM @ CM and AVM @ CM-SS-PU), the AVM technical product and the AVM commercial emulsifiable solution have higher insecticidal effects at three temperatures, because active ingredients in the AVM technical product and the AVM commercial emulsifiable solution can directly contact with the digestive system of the plutella xylostella, the plutella xylostella can be poisoned more quickly, and meanwhile, the temperature does not influence the biological activity of the plutella xylostella. The bioactivity of AVM @ CM and AVM @ CM-SS-PU increases with increasing temperature, because higher temperatures facilitate the release of the active ingredient in the microspheres. The insecticidal effect of the AVM @ CM-SS-PU at three temperatures is better than that of the AVM @ CM, and the effect of AVM technical and AVM commercial emulsifiable concentrate is achieved at 35 ℃, because glutathione rich in bodies of diamondback moths provides a reducing environment, the reducing environment destroys disulfide bonds on an AVM @ CM-SS-PU carrier, the carrier is disintegrated and the effective components are quickly released, the quick insecticidal effect is achieved, and even at a lower temperature, the biological activity of the AVM @ CM-SS-PU is higher than that of the AVM @ CM.

After 48h (fig. 11(b)), the insecticidal effect was higher than 24h for all four AVM formulations. The biological activities of AVM @ CM and AVM @ CM-SS-PU can be improved correspondingly with the temperature, and the biological activities of AVM @ CM and AVM @ CM-SS-PU can reach or even exceed those of AVM technical and AVM commercial emulsifiable concentrate at 35 ℃, probably because the microsphere preparation deposits more effective components on the fed leaves and the higher temperature stimulates the AVM in the microspheres to rapidly collect and release. In addition, the biological activity of the AVM @ CM-SS-PU at 15 ℃ and 25 ℃ is higher than that of the AVM @ CM, and the insecticidal effect of the AVM technical material and the AVM @ EC is achieved at 25 ℃, which shows that the breakage of disulfide bonds in the AVM @ CM-SS-PU in diamondback moth plays a key role, and the insecticidal activity of the microcapsule preparation is improved together with temperature response release. FIG. 11(c) shows a schematic diagram of AVM @ CM-SS-PU insecticidal at different temperatures.

The relative arrangement of the components and steps, the numerical expressions, and numerical values set forth in these embodiments do not limit the scope of the present application unless specifically stated otherwise. Meanwhile, it should be understood that the sizes of the respective portions shown in the drawings are not drawn in an actual proportional relationship for the convenience of description. Techniques, methods, and apparatus known to one of ordinary skill in the relevant art may not be discussed in detail but are intended to be part of the specification where appropriate. In all examples shown and discussed herein, any particular value should be construed as merely illustrative, and not limiting. Thus, other examples of the exemplary embodiments may have different values. It should be noted that: like reference numbers and letters refer to like items in the following figures, and thus, once an item is defined in one figure, further discussion thereof is not required in subsequent figures.

In the description of the present application, it is to be understood that the directions or positional relationships indicated by the directional terms such as "front, rear, upper, lower, left, right", "lateral, vertical, horizontal" and "top, bottom", etc., are generally based on the directions or positional relationships shown in the drawings, and are for convenience of description and simplicity of description only, and in the case of not making a reverse description, these directional terms do not indicate and imply that the device or element referred to must have a particular orientation or be constructed and operated in a particular orientation, and therefore, should not be considered as limiting the scope of the present application; the terms "inner and outer" refer to the inner and outer relative to the profile of the respective component itself.

Spatially relative terms, such as "above … …," "above … …," "above … …," "above," and the like, may be used herein for ease of description to describe one device or feature's spatial relationship to another device or feature as illustrated in the figures. It will be understood that the spatially relative terms are intended to encompass different orientations of the device in use or operation in addition to the orientation depicted in the figures. For example, if a device in the figures is turned over, devices described as "above" or "on" other devices or configurations would then be oriented "below" or "under" the other devices or configurations. Thus, the exemplary term "above … …" can include both an orientation of "above … …" and "below … …". The device may be otherwise variously oriented (rotated 90 degrees or at other orientations) and the spatially relative descriptors used herein interpreted accordingly.

It should be noted that the terms "first", "second", and the like are used to define the components, and are only used for convenience of distinguishing the corresponding components, and the terms have no special meanings unless otherwise stated, and therefore, the scope of protection of the present application is not to be construed as being limited.

The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A multiple stimulus response type pesticide microsphere is characterized in that: the insecticidal composition comprises modified carboxymethyl cellulose, an insecticidal component, isophorone diisocyanate and n-hexadecane, wherein the modified carboxymethyl cellulose comprises carboxymethyl cellulose and cystamine dihydrochloride.

2. The multiple stimulus-responsive pesticide microsphere of claim 1, wherein: the insecticidal component is one or more of abamectin, ivermectin, chlorpyrifos, 2, 4-dichlorophenoxyacetic acid, emamectin benzoate, chlorantraniliprole and high-efficiency cyhalothrin.

3. A preparation method of a multiple stimulus response type pesticide microsphere is characterized by comprising the following steps: the preparation of the multi-stimulus-responsive pesticide microsphere of claim 1 or 2, comprising the steps of,

step two, preparing pesticide microspheres, comprising the following steps:

4. The method for preparing the multi-stimulus-response pesticide microsphere according to claim 3, which is characterized by comprising the following steps of: in the first step, the weight ratio of the carboxymethyl cellulose to the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride to the N-hydroxysuccinimide to the cystamine dihydrochloride is 1 (0.1-0.3) to (0.1-0.2) to (1-3).

5. The method for preparing the multi-stimulus-response pesticide microsphere according to claim 3, which is characterized by comprising the following steps of: the weight ratio of the modified carboxymethyl cellulose, the insecticidal component, the isophorone diisocyanate, the n-hexadecane and the Tween is (0.2-0.6): 0.5-0.7): 1-3): 0.1-0.3.

6. The method for preparing the multi-stimulus-response pesticide microsphere according to claim 3, which is characterized by comprising the following steps of: in the first step, a dialysis bag is adopted to carry out dialysis purification on the material.

7. The method for preparing the multi-stimulus response type pesticide microsphere as claimed in claim 6, which is characterized in that: the dialysis bag is a Mw5000 dialysis bag.

8. The method for preparing the multi-stimulus-response pesticide microsphere according to claim 6, which is characterized by comprising the following steps of: the dialysis purification time is 40-50 h.

9. The method for preparing the multi-stimulus-response pesticide microsphere according to claim 3, which is characterized by comprising the following steps of: in the second step, the oil phase and the water phase are fully homogenized at the rotation speed of 5000-15000 rpm for 2-6 min at room temperature in a fully homogenized mode.

10. The method for preparing the multi-stimulus-response pesticide microsphere according to claim 1, which is characterized by comprising the following steps of: the tween is tween 80.