CN112889816A

CN112889816A - Insect virus and sex pheromone microcapsule and preparation method thereof

Info

Publication number: CN112889816A
Application number: CN202110018892.8A
Authority: CN
Inventors: 蒲侠; 罗萌; 林俊涛; 周新华; 舒绪刚
Original assignee: Zhongkai University of Agriculture and Engineering
Current assignee: Zhongkai University of Agriculture and Engineering
Priority date: 2021-01-07
Filing date: 2021-01-07
Publication date: 2021-06-04

Abstract

The invention relates to the technical field of biological pesticides, in particular to an insect virus and sex pheromone microcapsule and a preparation method thereof. The preparation method of the microcapsule comprises the following steps: the microcapsule containing the insect virus and the insect sex pheromone are uniformly mixed to obtain a mixture, the mixture is mixed with the chitosan solution, and the mixture is filtered to obtain the insect virus and sex pheromone microcapsule. The insect virus and sex pheromone microcapsule is safe and environment-friendly, can effectively attract insects, improves the insecticidal efficiency, protects insect viruses from being inactivated by sunlight, particularly ultraviolet light irradiation, and prolongs the storage time of a virus insecticide in the environment.

Description

Insect virus and sex pheromone microcapsule and preparation method thereof

Technical Field

The invention relates to the technical field of biological pesticides, in particular to an insect virus and sex pheromone microcapsule and a preparation method thereof.

Background

China is a big country for producing and consuming pesticides, and people no longer pay attention to high toxicity and high efficiency of pesticides. In contrast, the safety of pesticides to humans and their compatibility with the environment are increasingly emphasized. For example, the use of a large amount of insecticide has resulted in the resistance of insect populations, and further more insecticide needs to be used, which results in the reappearance of pesticide and pesticide residues in agricultural products, causing environmental pollution and ecological balance problems.

The insect virus pesticide is prepared by artificially cultivating, collecting, purifying and processing specific insect viruses, adopts a brand-new insect killing concept of 'combating poison with poison', has unique action mechanism and good prevention and treatment effect, and has a remarkable advantage of being very safe to the quality of agricultural products and the agricultural ecological environment.

The nuclear polyhedrosis virus preparation of insect virus is characterized in that: (1) the specificity is strong, the insect virus has specificity to the host, and does not infect people, livestock, natural enemies and the like, so the use is safe. But the general insecticidal spectrum is narrow, and only one kind of pest can be controlled by one kind of virus; (2) the effect is slow, a long pathogenic process is needed after the virus is eaten by pests, and the drug effect can be exerted after 7-10 days generally; (3) strong continuous control effect and long lasting effect. After the virus is released, the virus can survive in the environment for years, and can control pest populations for a long time. For some pests, the effects of one-time poison application, effective and long-term control for many years can be achieved; (4) is easy to be inactivated by the influence of ultraviolet rays, and can be used in the field without strong sunlight, and is preferably used in cloudy days.

The insect sex pheromone receiving system has high specificity and specificity, plays an important role in interspecific reproductive isolation, and can make directional flight, winging, copulation and other behaviors after receiving the sex pheromone. Thousands of chemical structures of insect sex pheromones have been identified, and hundreds of species have been commercially produced.

Insect viruses and sex pheromones are combined into an insecticide through microencapsulation, and no report is found at present.

Disclosure of Invention

In order to solve the technical problems, the invention provides an insect virus and sex pheromone microcapsule and a preparation method thereof.

In order to realize the technical effect of the invention, the following technical scheme is adopted for realizing the technical effect:

a method of making an insect virus and sex pheromone microcapsule, the method of making comprising: uniformly mixing the microcapsule containing the insect virus and the insect sex pheromone to obtain a mixture (the mixture can be obtained by uniformly mixing the microcapsule containing the insect virus and the insect sex pheromone solution, namely the insect sex pheromone is mixed with the microcapsule containing the insect virus in the form of the insect sex pheromone solution, and the mixture is mixed solution), mixing the mixture and the chitosan solution, and filtering to obtain the insect virus and sex pheromone microcapsule. The mixture (or the mixed solution) and the chitosan solution can be uniformly mixed by stirring and then filtered.

The sex pheromone of the insect attracts the heterosexual insects, and the insect virus kills the insects, so that the insecticidal effect is better than that of single virus.

As a further improvement, the insect virus is capable of causing the onset of insect infection by insect sex pheromones; the filtration is centrifugal filtration.

Insects feed in the traps to take in viruses or carry microcapsules containing viruses to move, so that the possibility of infection transmission is increased, and pests are controlled. For example: the male insects attracted by the insect sex pheromone eat the food containing the insect viruses and enter the body through the mouth, so that the infection is caused to occur or the attracted insects are infected by the insect viruses and the sex pheromone microcapsules and then infect other insects through the viruses.

As a further improvement, the insect virus and the insect in the insect sex pheromone are the same insect.

As a further improvement, the insect disease-containing food comprisesThe mass ratio of the toxic microcapsule to the chitosan is as follows: (20-40): (0.4-1); the mass ratio of the microcapsule containing the insect virus to the insect sex pheromone is 20 multiplied by 10⁶: 110; the stirring temperature is 25-37 ℃, and the stirring time is 0.5-2 hours; the solvent used for dissolving the insect sex pheromone in the insect sex pheromone solution is an organic solvent, and the organic solvent is one or more of n-hexane, ethanol, cyclohexanone, cyclohexanol, dimethyl phthalein amine, toluene and xylene; the insect sex pheromone is prodenia litura sex pheromone and/or beet armyworm sex pheromone.

As a further improvement, after filtration, the obtained filter residue is pre-frozen at the temperature of minus 90 to minus 60 ℃, and then taken out and frozen and dried at the temperature of minus 80 to minus 40 ℃ to obtain the insect virus and sex pheromone microcapsule; the insect virus and sex pheromone microcapsule is powder. The prodenia litura sex pheromone is a substance consisting of cis-9-trans-11-tetradecenol acetate and cis-9-trans-12-tetradecenol acetate; the beet armyworm sex pheromone is a substance consisting of cis-9, trans-12-tetradecenol acetate, cis-9-tetradecenol and cis-11-hexadecenol acetate.

As a further improvement, the preparation method of the chitosan solution comprises the following steps: adding chitosan into water, adding acetic acid while stirring until chitosan is fully dissolved, adjusting pH to 5.5-6 with sodium hydroxide, and filtering; an anti-ultraviolet agent and an antioxidant are also added into the mixed solution; the ultraviolet resistant agent comprises one or more of salicylate, benzophenone, benzotriazole and ultraviolet light stabilizer; the antioxidants include hindered phenolic antioxidants and phosphites. Antioxidant is antioxidant and can be hindered phenol antioxidant and phosphite ester with mass ratio of 1: 0.5. The addition of anti-ultraviolet agent and antioxidant can prolong the storage time of virus and sex pheromone, and further improve the insecticidal effect.

As a further improvement, the preparation method of the microcapsule containing the insect virus comprises the following steps:

stirring and uniformly mixing insect virus dry powder, a milk fat powder water mixed solution containing lactalbumin and a sodium alginate solution to form a mixture; then atomizing the mixture and spraying the mixture into a calcium chloride solution, and stirring to obtain a mixture C; and filtering the mixture C, taking filter residue, washing with water, and collecting wet microcapsules to obtain the insect virus microcapsules.

the method comprises the following steps: uniformly mixing insect virus dry powder and a milk fat powder water mixed solution containing lactalbumin to form a mixture A;

step two: uniformly mixing the mixture A and a sodium alginate solution to form a mixture B;

step three: atomizing and spraying the mixture B into a calcium chloride solution, and stirring to obtain a mixture C;

step four: filtering the mixture C, taking filter residue, washing with deionized water, and collecting wet microcapsules to obtain insect virus microcapsules;

the method specifically comprises the following steps:

the method comprises the following steps: adding the insect virus dry powder into the milk fat powder water mixed solution containing the lactalbumin, and stirring and mixing uniformly to form a mixture A;

step two: adding the mixture A into a sodium alginate solution, and stirring and mixing uniformly to form a mixture B;

step four: and filtering the mixture C, taking filter residues, washing the filter residues with deionized water, and collecting wet microcapsules to obtain the insect virus microcapsules.

As a further improvement, the mass fraction of the whey protein-containing milk fat powder in the mixture B (or the mixture) is 1-5%; the milk fat powder water mixed solution containing the whey protein is obtained by putting milk fat powder containing the whey protein into water for dispersing and uniformly mixing; and step three, sucking the mixture B (or the mixture), atomizing and spraying the mixture B into a calcium chloride solution, and stirring and solidifying the mixture B while spraying to obtain a mixture C.

As a further improvement, the mass fraction of the sodium alginate in the mixture B (or the mixture) is 1-3%; the milk fat powder water mixed solution containing the whey protein is a skim milk powder solution; the insect virus dry powder is prodenia litura nucleopolyhedrosis virus dry powder and/or beet armyworm nucleopolyhedrosis virus dry powder; in the third step, a peristaltic pump of a spray dryer is adopted to suck the mixture B (or the mixture), the mixture B (or the mixture) is atomized by a nozzle of the spray dryer and sprayed into the calcium chloride solution, the liquid suction speed of the peristaltic pump is 2.5-5mL/min, the inner diameter of the nozzle is 0.5mm, the mass fraction of the calcium chloride solution is 1.1-5.6%, and after the spraying is finished, the mixture B (or the mixture) is stirred for 10-30 min; the stirring in the third step is carried out by adopting a magnetic stirrer; the mass of the insect virus dry powder and the volume ratio of the milk fat powder-water mixed solution containing the lactalbumin are as follows: 0.04 g: (30-200) ml; the volume ratio of the milk fat powder water mixed solution containing the whey protein to the sodium alginate solution is as follows: 1, (2-4); the ratio of the volume of the mixture B (or mixture) sprayed into the calcium chloride solution to the volume of the calcium chloride solution is: (50-200): (800-3000); the mass fraction of the chitosan solution is 0.4-1%.

The invention also provides an insect virus and sex pheromone microcapsule which is prepared by the preparation method.

Compared with the prior art, the invention has the following beneficial effects:

1) the insect virus and sex pheromone microcapsule can be applied to control insects such as prodenia litura and beet armyworm, is safe and environment-friendly, effectively attracts insects, improves action timeliness, protects insect viruses from being inactivated by light, particularly ultraviolet irradiation, prolongs the storage time of virus insecticides and improves insecticidal efficiency.

2) The microcapsule is prepared by adopting alginate and chitosan in multiple steps, so that the microcapsule not only has double effects of killing insects and attracting, but also can better protect insect viruses from being inactivated by irradiation of light, particularly ultraviolet light, and the insecticidal efficiency is improved.

Detailed Description

Insect virus and sex pheromone microcapsule adopt sex pheromone and virus cooperation to use, on the one hand through virus killing insect, on the other hand through sex pheromone, lure the insect, and then improved insecticidal efficiency, reduce the insecticidal ageing. Particularly, insect viruses can cause insect infection induced by insect sex pheromone to occur, for example, the insect viruses and the insects in the insect sex pheromone are the same insects, the insects are induced by the sex pheromone, then the insects are induced to move with the viruses so that the insects without infection are infected, and simultaneously the insects carrying the viruses also carry the sex pheromone so as to induce other insects to be infected, so that the insecticidal efficiency is improved, and the insecticidal time is reduced.

Insect viruses can cause insect infection to cause epidemic diseases and/or progeny sterility and other problems. Insects (such as male moths) are attracted by utilizing the insect virus and sex pheromone microcapsules, so that the attracted insects are polluted by the sex pheromone and the virus and then return to the field, the problems of epidemic diseases and/or progeny sterility and the like of the whole population are caused, and the purpose of controlling the population density is achieved.

Alginate is an attractive matrix for microencapsulation of various foods, goods for agricultural and pharmaceutical applications. This material is natural and relatively low cost.

Sodium alginate is derived from brown algae, is a negatively charged polysaccharide, and is a linear polyanionic high polymer formed by beta-D-mannuronic acid (M) and alpha-L-guluronic acid (G) through beta- (1,4) glycosidic bonds. Crosslinking of adjacent polymer chains is induced by electrostatic interaction with divalent cations (e.g., calcium) that fit tightly between the polygulonate regions (G blocks) on adjacent alginate chains, forming an "egg-box" structure.

The protein material has controllable permeability, and the formed gel has better strength and acid-base buffering property, so that the protein is used as the wall material of the microcapsule to resist adverse environment. Whey protein has two advantages as a wall material: easily mixed with negatively charged polysaccharides such as alginate, pectin, etc. (also easily mixed with sodium alginate); when the environmental pH is lower than the isoelectric point, the charge of the whey protein is positive, the whey protein is combined with polysaccharide and the like more compactly, and the permeability is reduced. Therefore, the sodium alginate and the lactalbumin are compounded to make up the defect of the porosity of the sodium alginate, and the water-soluble calcium salt is used for crosslinking with the sodium alginate, so that the wall of the microcapsule is more compact, and the activity of insect viruses of the core material is protected.

The present invention will be further described with reference to the following examples.

It should be noted that the raw materials used in the technical scheme provided by the present invention, except for the specific description, are prepared by conventional means or purchased through commercial channels.

A method of making an insect virus and sex pheromone microcapsule, the method of making comprising: the microcapsule containing the insect virus and the insect sex pheromone are uniformly mixed to obtain a mixture, the mixture is mixed with the chitosan solution, and the mixture is filtered to obtain the insect virus and sex pheromone microcapsule. The microcapsule containing the insect virus and the insect sex pheromone solution are uniformly mixed to obtain a mixed solution, the mixed solution and the chitosan solution are mixed and filtered to obtain the insect virus and sex pheromone microcapsule;

the mixture can be added into chitosan solution, stirred, coated by chitosan (electrostatic binding), and filtered (centrifugal filtering for three times) to obtain microcapsule of insect virus and sex pheromone.

The insect virus of the present invention may be an insect virus capable of causing an insect attracted by an insect sex pheromone to infect and attack, and for example, the insect virus and the insect in the insect sex pheromone may be the same insect.

Preferably, the preparation method of the microcapsule containing the insect virus comprises the following steps:

Preferably, the mass ratio of the insect virus dry powder to the milk fat powder-water mixed solution containing the whey protein is as follows: 0.04 g: (30-200) ml; the volume ratio of the milk fat powder water mixed solution containing the whey protein to the sodium alginate solution is as follows: 1, (2-4); the volume ratio of the mixture B sprayed into the calcium chloride solution to the calcium chloride solution is as follows: (50-200): (800-3000); the mass ratio of the microcapsule containing the insect virus to the chitosan is as follows: (20-40): (0.4-1); the mass fraction of the chitosan solution is 0.4-1%.

Example 1

A method for preparing insect virus and sex pheromone microcapsules comprises the following steps:

the method comprises the following steps: adding 0.04g of insect virus dry powder into 30mL of milk fat powder water mixed solution containing lactalbumin, and stirring and mixing uniformly to form a mixture A;

step two: adding the mixture A into 63.5mL of sodium alginate solution, and stirring and mixing uniformly to form a mixture B;

wherein, in the mixture B, the mass fraction of the milk fat powder containing the whey protein is 4 percent, and the mass fraction of the sodium alginate is 2.5 percent;

step three: sucking the mixture B, atomizing and spraying the mixture B into the calcium chloride solution while stirring to obtain a mixture C; the method specifically comprises the following steps: sucking the mixture B by a peristaltic pump of a spray dryer at a speed of 2.5-5mL/min, atomizing the feed liquid by a nozzle with an inner diameter of 0.5mm, placing 800-3000 mL of a calcium chloride solution with a mass fraction of 2% below the nozzle, matching the calcium chloride solution with a magnetic stirrer, and continuously stirring for 15min after spraying to obtain a mixture C;

step four: filtering the mixture C, taking filter residue, washing with deionized water, and collecting wet microcapsules;

step five: adding 0.4g of chitosan into 98mL of water to obtain a mixed liquid, adding 1mL of acetic acid into the mixed liquid because the chitosan is soluble in an acid solution and is easy to agglomerate under neutral and alkaline conditions, fully dissolving, adjusting the pH value to 5.5-6 by using a NaOH solution after the dissolution is finished, and filtering to obtain a chitosan solution with the mass fraction of 0.4%.

Step six: and (3) taking 20g of wet microcapsules prepared in the fourth step, 0.33 mu g of ultraviolet resistant agent, 0.55 mu g of antioxidant and insect sex pheromone solution (the content of the insect sex pheromone is 110 mu g), dispersing and mixing uniformly to obtain mixed solution D, dropwise adding the mixed solution D into the chitosan solution, stirring for 40min at 37 ℃, carrying out chitosan coating on the microcapsules, and filtering (taking filter residues) to collect wet insect virus and sex pheromone microcapsules (sodium alginate/milk fat powder/chitosan microcapsules).

Step seven: the wet insect virus and sex pheromone microcapsules are put into an ultra-low temperature refrigerator to be pre-frozen at the temperature of (-90) - (-70) DEG C, and then the wet insect virus and sex pheromone microcapsules are taken out and put into a freeze drier to be dried for 12 hours to obtain the insect virus and sex pheromone microcapsule powder.

Example 2

step two: adding the mixture A into 65mL of sodium alginate solution, and stirring and mixing uniformly to form a mixture B;

wherein in the mixture B, the mass fraction of the whey protein-containing milk fat powder is 4%, and the mass fraction of the sodium alginate is 1%;

step five: adding 0.4g of chitosan into 98mL of water to obtain a mixed liquid, adding 1mL of acetic acid into the mixed liquid for fully dissolving because the chitosan can be dissolved in an acidic solution and is easy to agglomerate under neutral and alkaline conditions, adjusting the pH of the solution to 5.5-6 by using 5mol/L of NaOH after the dissolution is finished, filtering and obtaining a chitosan solution.

Step six: and (3) taking 20g of wet microcapsules prepared in the fourth step, 0.33 mu g of ultraviolet resistant agent, 0.55 mu g of antioxidant and insect sex pheromone solution (the content of the insect sex pheromone is 110 mu g), dispersing and mixing uniformly to obtain mixed liquor D, dropwise adding the mixed liquor D into the chitosan solution, stirring for 40min at 37 ℃, coating the chitosan of the microcapsules, and filtering and collecting the wet insect virus and sex pheromone microcapsules (sodium alginate/milk fat powder/chitosan microcapsules).

Example 3

step two: adding the mixture A into 64.5mL of sodium alginate solution, and stirring and mixing uniformly to form a mixture B;

wherein, in the mixture B, the mass fraction of the milk fat powder containing the whey protein is 4 percent, and the mass fraction of the sodium alginate is 1.5 percent;

step five: adding 0.4g of chitosan into 98mL of water to obtain a mixed liquid, adding 1mL of acetic acid into the mixed liquid for fully dissolving because the chitosan is soluble in an acidic solution and is easy to agglomerate under neutral and alkaline conditions, adjusting the pH of the solution to 5.5-6 by using 5mol/L of NaOH after the dissolution is finished, filtering and obtaining a chitosan solution.

Example 4

step two: adding the mixture A into 64mL of sodium alginate solution, and stirring and mixing uniformly to form a mixture B;

wherein in the mixture B, the mass fraction of the whey protein-containing milk fat powder is 4%, and the mass fraction of the sodium alginate is 2%;

Example 5

step two: adding the mixture A into 63mL of sodium alginate solution, and stirring and mixing uniformly to form a mixture B;

wherein, in the mixture B, the mass fraction of the milk fat powder containing whey protein is 4 percent, and the mass fraction of the sodium alginate is 3 percent;

step five: adding 0.4g of chitosan into 98mL of water to obtain a mixed liquid, adding 1mL of acetic acid into the mixed liquid for fully dissolving because the chitosan is soluble in an acidic solution and is easy to agglomerate under neutral and alkaline conditions, adjusting the pH of the solution to 5.5-6 by using a 5mol/L NaOH solution after the dissolution is finished, filtering and obtaining a chitosan solution.

Example 6

step two: adding the mixture A into 66.5mL of sodium alginate solution, and stirring and mixing uniformly to form a mixture B;

wherein in the mixture B, the mass fraction of the whey protein-containing milk fat powder is 1%, and the mass fraction of the sodium alginate is 2.5%;

step five: adding 0.4g of chitosan into 98mL of water to obtain a mixed liquid, adding 1mL of acetic acid into the mixed liquid for fully dissolving because the chitosan is soluble in an acidic solution and is easy to agglomerate under neutral and alkaline conditions, adjusting the pH of the solution to 5.5-6 by using a 5mol/L NaOH solution after the dissolution is finished, and filtering the solution to obtain a chitosan solution.

Example 7

step two: adding the mixture A into 65.5mL of sodium alginate solution, and stirring and mixing uniformly to form a mixture B;

wherein in the mixture B, the mass fraction of the whey protein-containing milk fat powder is 2%, and the mass fraction of the sodium alginate is 2.5%;

Example 8

wherein, in the mixture B, the mass fraction of the milk fat powder containing whey protein is 3 percent, and the mass fraction of the sodium alginate is 2.5 percent;

Example 9

step two: adding the mixture A into 62.5mL of sodium alginate solution, and stirring and mixing uniformly to form a mixture B;

wherein, in the mixture B, the mass fraction of the milk fat powder containing whey protein is 5 percent, and the mass fraction of the sodium alginate is 2.5 percent;

step five: adding 0.4g of chitosan into 98mL of water to obtain a mixed liquid, adding 1mL of acetic acid into the mixed liquid because the chitosan is soluble in an acidic solution and is easy to agglomerate under neutral and alkaline conditions, fully dissolving, adjusting the pH of the solution to 5.5-6 by using 5mol/L of NaOH after the dissolution is finished, filtering, and obtaining a chitosan solution.

Example 10

step two: adding the mixture A into 62mL of sodium alginate solution, and stirring and mixing uniformly to form a mixture B;

wherein in the mixture B, the mass fraction of the whey protein-containing milk fat powder is 5%, and the mass fraction of the sodium alginate is 3%;

step three: sucking the mixture B, atomizing and spraying the mixture B into the calcium chloride solution while stirring to obtain a mixture C; the method specifically comprises the following steps: sucking the mixture B by a peristaltic pump of a spray dryer at a speed of 2.5-5mL/min, atomizing the feed liquid by a nozzle with an inner diameter of 0.5mm, placing 800-3000 mL of a calcium chloride solution with a mass fraction of 1.1% below the nozzle, matching the calcium chloride solution with a magnetic stirrer, and continuously stirring for 10min after spraying to obtain a mixture C;

Step six: and (3) taking 20g of wet microcapsules prepared in the step four, 0.33 mu g of ultraviolet resistant agent, 0.55 mu g of antioxidant and insect sex pheromone solution (the content of the insect sex pheromone is 110 mu g), dispersing and mixing uniformly to obtain mixed liquor D, dropwise adding the mixed liquor D into the chitosan solution, stirring for 30min at 25 ℃, coating the chitosan of the microcapsules, and filtering and collecting the wet insect virus and sex pheromone microcapsules (sodium alginate/milk fat powder/chitosan microcapsules).

Step seven: the wet insect virus and sex pheromone microcapsules are put into an ultra-low temperature refrigerator to be pre-frozen at the temperature of (-80) - (-60) DEG C, and then the wet insect virus and sex pheromone microcapsules are taken out and put into a freeze drier to be dried for 12 hours to obtain the insect virus and sex pheromone microcapsule powder.

Example 11

step three: sucking the mixture B, atomizing and spraying the mixture B into the calcium chloride solution while stirring to obtain a mixture C; the method specifically comprises the following steps: sucking the mixture B by a peristaltic pump of a spray dryer at a speed of 2.5-5mL/min, atomizing the feed liquid by a nozzle with an inner diameter of 0.5mm, placing 800-3000 mL of a calcium chloride solution with a mass fraction of 5.6% below the nozzle, matching the calcium chloride solution with a magnetic stirrer, and continuously stirring for 30min after spraying to obtain a mixture C;

Step six: and (3) taking 20g of wet microcapsules prepared in the step four, 0.33 mu g of ultraviolet resistant agent, 0.55 mu g of antioxidant and insect sex pheromone solution (the content of the insect sex pheromone is 110 mu g), dispersing and mixing uniformly to obtain mixed liquor D, dropwise adding the mixed liquor D into the chitosan solution, stirring for 2 hours at 37 ℃, coating the chitosan of the microcapsules, and filtering and collecting the wet insect virus and sex pheromone microcapsules (sodium alginate/milk fat powder/chitosan microcapsules).

Step seven: the wet insect virus and sex pheromone microcapsules are put into an ultra-low temperature refrigerator to be pre-frozen at the temperature of (-80) - (-60) DEG C, and then the wet insect virus and sex pheromone microcapsules are taken out and put into a freeze drier to be dried for 24 hours to obtain the insect virus and sex pheromone microcapsule powder.

Example 12

The difference from example 1 is that the UV resistant agent and the antioxidant are not added, and the preparation method is the same.

Antioxidant is antioxidant and can be hindered phenol antioxidant and phosphite ester with mass ratio of 1: 0.5. The antioxidants selected in examples 1-12 were 2, 8-di-tert-butyl-4-methylphenol and dipentaerythritol diphosphite.

The anti-uv agent may include one or more of salicylates, benzophenones, benzotriazoles, uv light stabilizers. The UV blocker selected in examples 1-12 was octyl 2-hydroxybenzoate.

In examples 1 to 12, 0.33. mu.g of the ultraviolet inhibitor and 0.55. mu.g of the antioxidant were added in step six, and they were added in the form of 1mL of an ethanol solution (containing 0.33. mu.g of the ultraviolet inhibitor and 0.55. mu.g of the antioxidant). The ethanol solution is prepared by dissolving an ultraviolet resistant agent and an antioxidant in ethanol. For example, in the sixth step of example 1, "take 20g of the wet microcapsule prepared in the fourth step, 0.33. mu.g of the ultraviolet resistant agent, 0.55. mu.g of the antioxidant and the insect sex pheromone solution (the content of the insect sex pheromone is 110. mu.g)", there may be "take 20g of the wet microcapsule prepared in the fourth step, 1mL of the ethanol solution (the ethanol solution contains 0.33. mu.g of the ultraviolet resistant agent, 0.55. mu.g of the antioxidant) and the insect sex pheromone solution (the content of the insect sex pheromone is 110. mu.g).

In examples 1 to 12, the freeze-drying treatment can be carried out at a temperature of-80 to-40 ℃ in the freeze-dryer.

In examples 1 to 12, in the third step, the mixture is stirred while spraying until the mixture is uniformly stirred, that is, the sprayed mixture B and the calcium chloride solution are uniformly stirred and mixed to obtain a mixture C; alternatively, a magnetic stirrer may be placed in the calcium chloride solution, and the mixture B may be sprayed in during the stirring process.

In examples 1 to 12, the ultra-low temperature refrigerator may be a medical refrigerator and freezer, and a hail brand ultra-low temperature refrigerator.

In examples 1 to 12, the powdered milk containing whey protein was an aqueous solution of powdered skim milk. In the sixth step, the filtration is centrifugal filtration, and the centrifugal rate can be 500-.

In examples 1 to 12, the air flow rate of the spray dryer was set to 10 to 40L/min.

In example 1, the dosage of each substance (i.e. the mass of the insect virus dry powder, the volume of the milk fat powder water mixed solution and the sodium alginate solution) can be doubled in the first step and the second step, and the third step can be that 100mL of the mixture B is sucked and atomized and sprayed into the calcium chloride solution, and the result is the same.

In examples 1 to 12, the insect sex pheromone was a sex pheromone of Spodoptera litura, and 2 sex pheromone compounds of cis-9-trans-11-tetradecenol acetate and cis-9-trans-12-tetradecenol acetate were mixed at a ratio of 100: 10.

In examples 1 to 12, the insects were spodoptera litura, and the spodoptera litura nuclear polyhedrosis virus was used as the insect virus dry powder.

In examples 1 to 12, the insect sex pheromone solution may be prepared by dissolving insect sex pheromone in an organic solvent to prepare an insect sex pheromone solution with a certain concentration, wherein the organic solvent is ethanol, and the mass ratio of the insect sex pheromone to the organic solvent is 1: 5.

of course, the organic solvent used in the insect sex pheromone solution of the invention can also be one or more of N-hexane, ethanol, cyclohexanone, cyclohexanol, N-dimethyl phthalein amine, toluene and xylene, as long as the insect sex pheromone can be dissolved. The mass ratio of the insect sex pheromone to the organic solvent can also be 1: 0.1, as long as it can dissolve insect sex pheromones.

In each example, the mass fraction of whey protein-containing powdered milk fat in mixture B was calculated by calculating the mass of powdered milk fat added to the whey protein-containing powdered milk fat-water mixture in step one. And similarly, the mass fraction of the sodium alginate is the mass of the sodium alginate calculated by the mass of the sodium alginate added into the sodium alginate solution in the step two.

In the third step of each embodiment, the spraying and stirring are carried out simultaneously, the curing is carried out, and the stirring time can be 10-30 minutes after the spraying is finished.

In examples 1 to 12, the powdered skim milk solution (or the aqueous mixed solution of powdered skim milk) may be a mixed solution of powdered cream and water containing whey protein.

Corresponding comparative examples were also made in each example, i.e., step five and step six were removed from each example, step seven "wet insect virus and sex pheromone microcapsule" was changed to "wet microcapsule", and the remaining steps were the same as step seven in the corresponding examples, to obtain sodium alginate/milk fat powder insect virus microcapsule powder. Example n (n is an integer from 1 to 13) the corresponding comparative example n is identified as comparative example n1, e.g., example 1 corresponds to comparative example 11.

Meanwhile, each example is provided with another group of corresponding comparative examples, namely, in each comparative example, no insect virus dry powder is added, (step one is omitted in each example, and a milk fat powder water mixed solution containing whey protein is directly added into a sodium alginate solution in step two, and is stirred and mixed uniformly to form a mixture B), and the other steps and the material consumption are the same as those of the corresponding examples. Example n (n is an integer from 1 to 13) the corresponding comparative example n is designated comparative example n2, e.g., example 1 corresponds to comparative example 12. Clean feed was used as control.

And (3) testing the attraction and ultraviolet resistance:

the samples were: pure virus dry powder (0.04 g), insect virus and sex pheromone microcapsules prepared according to examples 1 to 12 and the product of the comparative example.

1) Using a writing brush to uniformly brush 36 spodoptera litura feed surfaces with the same size and shape (namely, each sample is coated with 3 spodoptera litura feed surfaces) and then inoculating 20 heads of 3-instar spodoptera litura larvae (test insects) to each feed surface, wherein the temperature of an incubator is kept at 25 ℃, and the illumination is 12L: 12D, the illumination time is 24 hours. The observation is carried out for 7 days, and the cumulative death rate of the larvae is calculated.

2) The pure virus dry powder samples and each of the samples prepared in examples 1-12 were placed in an attracting device. The luring device with the sample is placed in a place where prodenia litura often appears and causes harm, such as a vegetable garden and the like, one luring device (the device is a cuboid, a plurality of baffles are arranged inside the luring device, the prodenia litura can move, the prodenia litura is lured through sex pheromone, and the prodenia litura flies into the luring device from the lower part of the device) is placed every 100 meters, and after 12 days, the condition of the prodenia litura in the luring device is checked.

The above experiments were repeated 3 times and the average was taken. The specific results are shown in table 1 below:

TABLE 1

From the above table, it can be seen that: the method provided by the embodiment of the invention can be used for killing insects and inducing, and after the chitosan coating of the microcapsule is carried out, the insecticidal effect is better (the viruses are probably not easy to inactivate), and unexpected technical effects are generated.

In example 2, because the mass fraction of sodium alginate is only 1%, the formed microcapsules are few, the crosslinking degree with calcium chloride is low, the microcapsules have the phenomenon of porosity, and viruses cannot be well protected under ultraviolet irradiation, so the death rate of the test insects is 15%; the lethality of examples 1 and 9 to the test insects reaches 75%, which shows that the viruses in the virus microcapsules of the insect virus insecticide still have most activity, the sodium alginate/milk fat powder is used for wrapping the insect viruses, the viruses can be effectively protected from being killed by ultraviolet rays, and further the mortality of the test insects is high.

The used virus dry powder can also be beet armyworm polyhedrosis virus dry powder and other virus dry powders.

In the experiment, if the prodenia litura polyhedrosis virus dry powder in the examples 1-12 is replaced by the beet armyworm polyhedrosis virus dry powder, the prodenia litura sex pheromone is replaced by the beet armyworm sex pheromone (for example, the sex pheromone consisting of cis-9, trans-12-tetradecenol acetate, cis-9-tetradecenol and cis-11-hexadecenol acetate is mixed at a ratio of 7:3:1, and the addition amount is 1.0mg), the same preparation method, the rest material amounts and the test method are adopted for the attraction and ultraviolet resistance performance test (the test larva is changed into beet armyworm larva, and the feed is changed into beet armyworm feed), the microcapsules prepared by each example are found to be capable of attracting the beet armyworm (the amount of the beet armyworm attracted by each example is 1-7 more than the amount attracted by corresponding example 1-12), each example had 1-3 less deaths compared to the corresponding examples 1-12, and the reduction in the number of deaths was probably due to the slow action time of the Spodoptera exigua polyhedrosis virus within 7 days, but still had a good insecticidal attraction effect.

In addition, in the experiment, if the steps one to four are not carried out, the steps five, six and seven are directly carried out, and no wet microcapsule is added in the step six, the microcapsule can not be produced at all.

The sodium alginate solution, the calcium chloride solution and the NaOH solution can be aqueous solutions.

The technical solutions provided by the embodiments of the present invention are described in detail above, and the principles and embodiments of the present invention are explained herein by using specific examples, and the descriptions of the embodiments are only used to help understanding the principles of the embodiments of the present invention; meanwhile, for a person skilled in the art, according to the embodiments of the present invention, there may be variations in the specific implementation manners and application ranges, and in summary, the content of the present description should not be construed as a limitation to the present invention.

Claims

1. A method for preparing insect virus and sex pheromone microcapsules, which is characterized by comprising the following steps: the microcapsule containing the insect virus and the insect sex pheromone are uniformly mixed to obtain a mixture, the mixture is mixed with the chitosan solution, and the mixture is filtered to obtain the insect virus and sex pheromone microcapsule.

2. The method of claim 1, wherein the microcapsule containing insect virus and the sex pheromone is mixed with the insect sex pheromone solution to obtain a mixture, the mixture is mixed with the chitosan solution, and the mixture is filtered to obtain the insect virus and sex pheromone microcapsule; the insect virus is capable of causing the onset of insect infection by insect sex pheromones; uniformly mixing the mixed solution and the chitosan solution by stirring, and filtering; the filtration is centrifugal filtration.

3. The method of claim 2, wherein the insect virus and sex pheromone are the same insect species.

4. The process for preparing insect virus and sex pheromone microcapsules according to claim 2,

the microcapsule containing insect virus and chitosanThe quantity ratio is: (20-40): (0.4-1); the mass ratio of the microcapsule containing the insect virus to the insect sex pheromone is 20 multiplied by 10⁶: 110; the stirring temperature is 25-37 ℃, and the stirring time is 0.5-2 hours; the solvent used for dissolving the insect sex pheromone in the insect sex pheromone solution is an organic solvent, and the organic solvent is one or more of N-hexane, ethanol, cyclohexanone, cyclohexanol, N-dimethyl phthalein amine, toluene and xylene; the mass ratio of the insect sex pheromone to the organic solvent is 1: (0.1-5); the insect sex pheromone is prodenia litura sex pheromone and/or beet armyworm sex pheromone.

5. The method for preparing insect virus and sex pheromone microcapsules according to claim 4, wherein the filter residue is pre-frozen at (-90) — (-60) ° C after filtration, and then the filter residue is taken out and is subjected to freeze drying treatment at (-80) — (-40) ° C to obtain the insect virus and sex pheromone microcapsules; the insect virus and sex pheromone microcapsule is powdery; the prodenia litura sex pheromone is a substance consisting of cis-9-trans-11-tetradecenol acetate and cis-9-trans-12-tetradecenol acetate; the beet armyworm sex pheromone is a substance consisting of cis-9, trans-12-tetradecenol acetate, cis-9-tetradecenol and cis-11-hexadecenol acetate.

6. The method of claim 1, wherein the chitosan solution is prepared by: adding chitosan into water, adding acetic acid while stirring until chitosan is fully dissolved, adjusting pH to 5.5-6 with sodium hydroxide, and filtering; an anti-ultraviolet agent and an antioxidant are also added into the mixed solution; the ultraviolet resistant agent comprises one or more of salicylate, benzophenone, benzotriazole and ultraviolet light stabilizer; the antioxidants include hindered phenolic antioxidants and phosphites.

7. The method of claim 1, wherein the method of preparing the insect virus and sex pheromone microcapsule comprises the steps of: stirring and uniformly mixing insect virus dry powder, a milk fat powder water mixed solution containing lactalbumin and a sodium alginate solution to form a mixture; then atomizing the mixture and spraying the mixture into a calcium chloride solution, and stirring to obtain a mixture C; and filtering the mixture C, taking filter residue, washing with water, and collecting wet microcapsules to obtain the insect virus microcapsules.

8. The process for preparing insect virus and sex pheromone microcapsules according to claim 7,

the mixture B contains whey protein-containing milk fat powder with the mass fraction of 1-5%; the milk fat powder water mixed solution containing the whey protein is obtained by putting milk fat powder containing the whey protein into water for dispersing and uniformly mixing; absorbing the mixture B, atomizing and spraying the mixture B into a calcium chloride solution, stirring and solidifying while spraying to obtain a mixture C; the mass of the insect virus dry powder and the volume ratio of the milk fat powder-water mixed solution containing the lactalbumin are as follows: 0.04 g: (30-200) ml; the volume ratio of the milk fat powder water mixed solution containing the whey protein to the sodium alginate solution is as follows: 1, (2-4); the volume ratio of the mixture B sprayed into the calcium chloride solution to the calcium chloride solution is as follows: (50-200): (800-3000); the mass fraction of the chitosan solution is 0.4-1%.

9. The method for preparing insect virus and sex pheromone microcapsules according to claim 7, wherein the mass fraction of sodium alginate in the mixture B is 1-3%; the milk fat powder water mixed solution containing the whey protein is a skim milk powder solution; the insect virus dry powder is prodenia litura nucleopolyhedrosis virus dry powder and/or beet armyworm nucleopolyhedrosis virus dry powder; in the third step, a peristaltic pump of a spray dryer is adopted to suck the mixture B, the mixture B is atomized by a nozzle of the spray dryer and sprayed into the calcium chloride solution, the liquid suction speed of the peristaltic pump is 2.5-5mL/min, the inner diameter of the nozzle is 0.5mm, the mass fraction of the calcium chloride solution is 1.1-5.6%, and after the spraying is finished, the mixture B is stirred for 10-30 min; the stirring in the third step is carried out by adopting a magnetic stirrer; the volume ratio of the milk fat powder water mixed solution containing the whey protein to the sodium alginate solution is as follows: 30, (62-66.5); the mass ratio of the insect virus dry powder to the calcium chloride solution is as follows: 0.04 g: (800-; the mass fraction of the chitosan solution is 0.4-1%.

10. An insect virus and sex pheromone microcapsule, which is prepared by the preparation method of any one of claims 1 to 9.