Animal amino acid-based plant root oriented nanocapsule and preparation method thereof

Abstract

The invention discloses a plant root oriented nanocapsule based on animal amino acid and a preparation method thereof, and relates to the technical field of nanocapsules. The preparation method specifically comprises the following steps: S1, reacting and grafting chitosan and cinnamyl aldehyde, and then reacting and grafting with lactide to obtain a modified chitosan-polylactic acid grafted copolymer; s2, after animal amino acid is coated with the modified chitosan-polylactic acid grafted copolymer, an EDC aqueous solution is added for activating treatment, finally, the animal amino acid reacts with the modified chitosan-polylactic acid grafted copolymer for grafting, and the plant root system guiding nanocapsule based on the animal amino acid is obtained. The plant root system guiding nanocapsule based on the animal amino acid has natural affinity with a plant root system, so that the plant root system has specific adsorbability on the plant root system guiding nanocapsule, and therefore, in practical application, the plant root system can be promoted to grow, and nutrient substances can be enriched towards the root system.

Description

Technical Field

[0001] This invention relates to the field of nanocapsule technology, specifically to plant root-directed nanocapsules based on animal amino acids and their preparation methods. Background Technology

[0002] Plant growth and development depend on a comprehensive supply of nutrients, among which nitrogen is a key component of proteins, nucleic acids, and chlorophyll. Traditional nitrogen fertilizers are mainly applied to the soil in inorganic forms (such as urea and ammonium salts), but their utilization rate is generally low. Most of the nitrogen is lost through volatilization, leaching, and denitrification, resulting not only in resource waste but also in serious non-point source pollution and a series of environmental problems such as eutrophication of water bodies. Amino acids, as small-molecule organic nitrogen sources, can be directly and rapidly absorbed and utilized by plant roots, participating in various physiological metabolic processes. Their absorption efficiency is far higher than that of inorganic nitrogen fertilizers, and they can also stimulate plant growth and enhance stress resistance. At the same time, my country's livestock industry produces a large number of dead livestock and poultry every year. To achieve harmless treatment and resource utilization of these animals, our company's patent CN111375618A discloses a harmless amino acid-containing aqueous solution for dead livestock and poultry and its preparation method.

[0003] Based on this, the present invention provides a plant root-directed nanocapsule based on animal amino acids and its preparation method, thereby realizing the resource utilization of animal amino acids, which is of great significance. Summary of the Invention

[0004] The purpose of this invention is to provide a plant root-directed nanocapsule based on animal amino acids and its preparation method, so as to solve the problems mentioned in the background art.

[0005] To solve the above-mentioned technical problems, the present invention provides the following technical solution: The preparation method of plant root-directed nanocapsules based on animal amino acids includes the following steps: S1: Chitosan is grafted with cinnamaldehyde and then grafted with lactide to obtain a modified chitosan-polylactic acid graft copolymer. S2: After the modified chitosan-polylactic acid graft copolymer is coated with animal amino acids, it is activated by adding EDC aqueous solution, and then it is grafted with animal amino acids to obtain plant root-guided nanocapsules based on animal amino acids.

[0006] Further, the specific process of S1 is as follows: (1) Under nitrogen and light-protected conditions, chitosan and cinnamaldehyde are added to 1-allyl-3-chloromethylimidazolium, respectively, and stirred and mixed evenly at 70~80℃ to prepare chitosan solution and cinnamaldehyde solution, which are kept warm for later use; (2) Under nitrogen and light-protected conditions, acetic acid is added to chitosan solution, stirred and mixed evenly, and then cinnamaldehyde solution is slowly added dropwise to it. The addition is completed within 1 hour. After the addition is completed, the reaction is stirred for 6~12 hours. h, to obtain a modified chitosan solution; (3) under nitrogen and light-proof conditions, add lactide to the modified chitosan solution, stir and mix evenly at 110~120℃, then add stannous octoate, keep warm and stir for 12~24h, end the reaction, let it cool naturally to room temperature, add excess anhydrous ethanol, let stand overnight, collect the precipitate, wash with anhydrous ethanol 2~3 times, and finally vacuum dry at 55~65℃ to obtain the modified chitosan-polylactic acid graft copolymer.

[0007] Further, the modified chitosan-polylactic acid graft copolymer comprises the following raw material components in parts by weight: 6-10 parts chitosan, 0.5-1 part cinnamaldehyde, 50-70 parts 1-allyl-3-chloromethylimidazolium, 2-2.5 parts acetic acid, 12-24 parts lactide, and 0.2-0.4 parts stannous octoate.

[0008] Furthermore, all raw material components of the modified chitosan-polylactic acid graft copolymer need to be vacuum dried overnight at 55~65°C.

[0009] The preparation of the modified chitosan-polylactic acid graft copolymer of the present invention requires strict control of the water content of the raw material components. The reasons are: (1) If water affects the Schiff base reaction between chitosan and cinnamaldehyde, the reaction process will be slowed down, the degree of graft modification of the product will be lower, and even cinnamaldehyde will undergo self-oxidation, condensation, etc., which will trigger side reactions; (2) Water will also cause lactide to hydrolyze and open the ring, which will affect the preparation of the modified chitosan-polylactic acid graft copolymer.

[0010] Furthermore, the degree of deacetylation of the chitosan is 85-95%, and the molecular weight is ≥100,000.

[0011] The proposed method should select chitosan with a degree of deacetylation of 85-95%, as it has a higher amino content, providing more grafting sites and thus improving the grafting rate. The molecular weight should be controlled above 100,000. If the molecular weight is too low, the modified chitosan-polylactic acid graft copolymer will have a looser entanglement, resulting in faster release of animal amino acids from the subsequent nanocapsules and poor sustained-release properties. If the molecular weight is too high, the entanglement will be too dense, resulting in a longer degradation time for the subsequent nanocapsules and preventing them from acting on the plant roots in a timely manner. Therefore, the preferred molecular weight of chitosan is 100,000 to 200,000.

[0012] This invention first uses acetic acid as a catalyst to induce a Schiff base reaction between the aldehyde group of cinnamaldehyde and the amino group of chitosan, thus completing the initial modification of chitosan. Then, using stannous octoate as a catalyst, lactide is subjected to ring-opening polymerization on the modified chitosan to prepare a modified chitosan-polylactic acid graft copolymer. Since the entire preparation process requires heating, the preparation must be carried out under nitrogen and light-protected conditions to protect cinnamaldehyde as much as possible.

[0013] Further, the specific process of S2 is as follows: (1) Under nitrogen and light-protected conditions, the modified chitosan-polylactic acid graft copolymer is added to dichloromethane, stirred and mixed evenly, and then an animal amino acid aqueous solution is added and stirred and mixed evenly to obtain a mixed primary emulsion; (2) Under nitrogen and light-protected conditions, an emulsifier is added to (1), and stirred and mixed at a speed of 500~600 rpm for 2~5 min, and then the speed is adjusted to 200~400 rpm, and stirred and mixed under ventilation conditions for 6~12 h to obtain a nanocapsule suspension; (3) Under nitrogen and light-protected conditions, an EDC aqueous solution is added to the nanocapsule suspension, stirred and mixed for 10~30 min, the pH is adjusted to neutral, and then an animal amino acid aqueous solution is added, stirred and mixed for 1~3 h, and finally centrifuged at a speed of 8000~10000 rpm for 10~30 min, the precipitate is collected, washed with deionized water 2~3 times, and freeze-dried at -30~-20℃ to obtain plant root-guided nanocapsules based on animal amino acids.

[0014] Furthermore, the plant root-guided nanocapsules based on animal amino acids comprise the following raw material components in parts by weight: 4-6 parts of modified chitosan-polylactic acid graft copolymer, 20-30 parts of dichloromethane, 8-12 parts of animal amino acid aqueous solution, 0.6-1.2 parts of emulsifier, and 3-6 parts of EDC aqueous solution.

[0015] Furthermore, the animal amino acid aqueous solution is prepared according to the preparation method disclosed in CN111375618A, and the amino acid content is 50~80g / L.

[0016] Furthermore, the emulsifier is a nonionic emulsifier, including either Tween 80 or Span 60.

[0017] Furthermore, the concentration of the EDC aqueous solution is 2~3 mM.

[0018] Furthermore, the plant root-directed nanocapsules based on animal amino acids are prepared by the above preparation method.

[0019] This invention uses a modified chitosan-polylactic acid graft copolymer as the nanocapsule wall material to encapsulate animal amino acids, thereby obtaining nanocapsules. Finally, the nanocapsules are activated with an EDC aqueous solution, so that the -COOH at the end of the polylactic acid chain on the surface of the nanocapsules can also react with the animal amino acids, thereby grafting animal amino acids onto the surface of the nanocapsules, resulting in plant root-guided nanocapsules based on animal amino acids.

[0020] Compared with the prior art, the beneficial effects achieved by the present invention are: (1) The plant root-oriented nanocapsules based on animal amino acids prepared in this invention have a natural affinity with plant roots, which makes the plant roots have specific adsorption properties. Therefore, in practical applications, they can promote plant root growth and also achieve the enrichment of nutrients into the roots.

[0021] (2) The plant root-guided nanocapsules based on animal amino acids prepared in this invention have a slow-release effect. As polylactic acid is slowly degraded, the internal animal amino acids are slowly released, which is more conducive to the absorption of plant roots. This avoids the release of amino acids too quickly, which would lead to soil eutrophication and affect plant growth, while also avoiding nutrient loss.

[0022] (3) In this invention, cinnamaldehyde is grafted onto chitosan, which can improve the antibacterial properties of chitosan and thus enhance the plant's disease resistance. Detailed Implementation

[0023] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0024] It should be noted that the following quantities are by weight, and there are no special restrictions on the suppliers of all raw materials involved in this invention. Exemplary examples include: In the following examples, chitosan had the following properties: degree of deacetylation 80%, average molecular weight 50,000; degree of deacetylation 85%, average molecular weight 100,000; degree of deacetylation 90%, average molecular weight 150,000; degree of deacetylation 95%, average molecular weight 200,000; cinnamaldehyde, L-lactide, 1-allyl-3-chloromethylimidazolium, ferrous acetate, ammonium carbonate, and palladium on carbon were all purchased from Merck Reagents Ltd.; other raw materials were commercially available; each part by weight was 100g.

[0025] Preliminary preparations: 1. Preparation of animal amino acid aqueous solution: (1) Weigh 1 part ferrous acetate, 2.3 parts urea, and 1.65 parts ammonium carbonate and add them to 1.8 parts deionized water. Stir at 120 rpm for 35 min at 87 °C, then heat to 110 °C and stir for 25 min. Stop the reaction and let it cool naturally to room temperature. After filtration, washing, and drying, the iron-ammonia complex is obtained. (2) 100 parts of pork were added to a sealed reactor and reacted at 145°C and 0.6 MPa for 40 min. Then 7 parts of concentrated sulfuric acid and 9 parts of deionized water were added, and the temperature was adjusted to 110°C and the pressure to 0.2 MPa. The mixture was stirred at 20 rpm for 1.5 h. Then 0.08 parts of hydrogen peroxide, 0.09 parts of formic acid, 0.045 parts of formaldehyde, and 0.005 parts of palladium on carbon were added. The temperature was adjusted to 67°C and the pressure to 0.03 MPa. The mixture was stirred at 80 rpm for 0.3 h. Finally, 0.05 parts of iron-ammonia complex were added, and the material was transferred to an ultrasonic reactor and ultrasonically treated at 53°C and 20 kHz for 3 s. The mixture was allowed to stand overnight, filtered, and concentrated to obtain a 50 g / L animal amino acid aqueous solution.

[0026] 2. Chitosan, cinnamaldehyde, 1-allyl-3-chloromethylimidazolium, acetic acid, lactide, and stannous octoate were all vacuum dried overnight at 60°C; they were then removed before the experiment.

[0027] Example 1: Preparation method of plant root-directed nanocapsules based on animal amino acids: S1: Preparation of modified chitosan-polylactic acid graft copolymer: (1) Under nitrogen and light-protected conditions, 8 parts of chitosan (degree of deacetylation of 90%, average molecular weight of 150,000) were added to 50 parts of 1-allyl-3-chloromethylimidazolium, and 0.75 parts of cinnamaldehyde were added to 5 parts of 1-allyl-3-chloromethylimidazolium. The mixture was stirred and mixed evenly at 75°C to prepare chitosan solution and cinnamaldehyde solution, and kept warm for later use; (2) Under nitrogen and light-protected conditions, 2.25 parts of acetic acid were added to the chitosan solution and stirred and mixed evenly. Then, cinnamaldehyde was slowly added dropwise to the solution. Cinnamaldehyde solution was added dropwise within 1 hour. After the addition was completed, the reaction was stirred for 9 hours to obtain modified chitosan solution. (3) Under nitrogen and light-proof conditions, 18 parts of L-lactide were added to the modified chitosan solution and stirred and mixed evenly at 115°C. Then 0.3 parts of stannous octoate were added and stirred for 18 hours to stop the reaction. After the reaction was stopped, the solution was allowed to cool naturally to room temperature. Excess anhydrous ethanol was added and the solution was allowed to stand overnight. The precipitate was collected, washed three times with anhydrous ethanol, and finally dried under vacuum at 60°C for 3 hours to obtain modified chitosan-polylactic acid graft copolymer. S2: Preparation of plant root-oriented nanocapsules based on animal amino acids: (1) Under nitrogen and light-proof conditions, 5 parts of modified chitosan-polylactic acid graft copolymer were added to 25 parts of dichloromethane and stirred until homogeneous. Then, 9 parts of animal amino acid aqueous solution were added and stirred until homogeneous to obtain mixed primary emulsion. (2) Under nitrogen and light-proof conditions, 0.9 parts of Tween 80 were added to (1) and stirred at 550 rpm for 3 min. Then, the speed was adjusted to 300 rpm and stirred for 9 h under ventilation conditions to obtain nanocapsule suspension. (3) Under nitrogen and light-proof conditions, 4.5 parts of 2.5 mM EDC aqueous solution were added to nanocapsule suspension and stirred for 20 min. The pH was adjusted to neutral and then 1 part of animal amino acid aqueous solution was added and stirred for 2 h. Finally, the mixture was centrifuged at 9000 rpm for 20 min, the precipitate was collected, washed 3 times with deionized water, and freeze-dried at -25℃ for 3 h to obtain plant root-oriented nanocapsules based on animal amino acids.

[0028] Example 2: Preparation method of plant root-directed nanocapsules based on animal amino acids: Example 2 is based on Example 1, with the following adjustments: chitosan with a degree of deacetylation of 85% and an average molecular weight of 100,000 is selected, while other processes remain unchanged. Specifically: S1: Preparation of modified chitosan-polylactic acid graft copolymer: (1) Under nitrogen and light-protected conditions, 8 parts of chitosan (degree of deacetylation of 85%, average molecular weight of 100,000) were added to 50 parts of 1-allyl-3-chloromethylimidazolium, and 0.75 parts of cinnamaldehyde were added to 5 parts of 1-allyl-3-chloromethylimidazolium. The mixture was stirred and mixed evenly at 75°C to prepare chitosan solution and cinnamaldehyde solution, and kept warm for later use; (2) Under nitrogen and light-protected conditions, 2.25 parts of acetic acid were added to the chitosan solution and stirred and mixed evenly. Then, cinnamaldehyde was slowly added dropwise to the solution. Cinnamaldehyde solution was added dropwise within 1 hour. After the addition was completed, the reaction was stirred for 9 hours to obtain modified chitosan solution. (3) Under nitrogen and light-proof conditions, 18 parts of L-lactide were added to the modified chitosan solution and stirred and mixed evenly at 115°C. Then 0.3 parts of stannous octoate were added and stirred for 18 hours to stop the reaction. After the reaction was stopped, the solution was allowed to cool naturally to room temperature. Excess anhydrous ethanol was added and the solution was allowed to stand overnight. The precipitate was collected, washed three times with anhydrous ethanol, and finally dried under vacuum at 60°C for 3 hours to obtain modified chitosan-polylactic acid graft copolymer. S2: Preparation of plant root-oriented nanocapsules based on animal amino acids: (1) Under nitrogen and light-proof conditions, 5 parts of modified chitosan-polylactic acid graft copolymer were added to 25 parts of dichloromethane and stirred until homogeneous. Then, 9 parts of animal amino acid aqueous solution were added and stirred until homogeneous to obtain mixed primary emulsion. (2) Under nitrogen and light-proof conditions, 0.9 parts of Tween 80 were added to (1) and stirred at 550 rpm for 3 min. Then, the speed was adjusted to 300 rpm and stirred for 9 h under ventilation conditions to obtain nanocapsule suspension. (3) Under nitrogen and light-proof conditions, 4.5 parts of 2.5 mM EDC aqueous solution were added to nanocapsule suspension and stirred for 20 min. The pH was adjusted to neutral and then 1 part of animal amino acid aqueous solution was added and stirred for 2 h. Finally, the mixture was centrifuged at 9000 rpm for 20 min, the precipitate was collected, washed 3 times with deionized water, and freeze-dried at -25℃ for 3 h to obtain plant root-oriented nanocapsules based on animal amino acids.

[0029] Example 3: Preparation method of plant root-directed nanocapsules based on animal amino acids: Example 3 is based on Example 1, with the following adjustments: chitosan with a degree of deacetylation of 95% and an average molecular weight of 200,000 is selected, while other processes remain unchanged. Specifically: S1: Preparation of modified chitosan-polylactic acid graft copolymer: (1) Under nitrogen and light-protected conditions, 8 parts of chitosan (degree of deacetylation of 95%, average molecular weight of 200,000) were added to 50 parts of 1-allyl-3-chloromethylimidazolium, and 0.75 parts of cinnamaldehyde were added to 5 parts of 1-allyl-3-chloromethylimidazolium. The mixture was stirred and mixed evenly at 75°C to prepare chitosan solution and cinnamaldehyde solution, and kept warm for later use; (2) Under nitrogen and light-protected conditions, 2.25 parts of acetic acid were added to the chitosan solution and stirred and mixed evenly. Then, cinnamaldehyde was slowly added dropwise to the solution. Cinnamaldehyde solution was added dropwise within 1 hour. After the addition was completed, the reaction was stirred for 9 hours to obtain modified chitosan solution. (3) Under nitrogen and light-proof conditions, 18 parts of L-lactide were added to the modified chitosan solution and stirred and mixed evenly at 115°C. Then 0.3 parts of stannous octoate were added and stirred for 18 hours to stop the reaction. After the reaction was stopped, the solution was allowed to cool naturally to room temperature. Excess anhydrous ethanol was added and the solution was allowed to stand overnight. The precipitate was collected, washed three times with anhydrous ethanol, and finally dried under vacuum at 60°C for 3 hours to obtain modified chitosan-polylactic acid graft copolymer. S2: Preparation of plant root-oriented nanocapsules based on animal amino acids: (1) Under nitrogen and light-proof conditions, 5 parts of modified chitosan-polylactic acid graft copolymer were added to 25 parts of dichloromethane and stirred until homogeneous. Then, 9 parts of animal amino acid aqueous solution were added and stirred until homogeneous to obtain mixed primary emulsion. (2) Under nitrogen and light-proof conditions, 0.9 parts of Tween 80 were added to (1) and stirred at 550 rpm for 3 min. Then, the speed was adjusted to 300 rpm and stirred for 9 h under ventilation conditions to obtain nanocapsule suspension. (3) Under nitrogen and light-proof conditions, 4.5 parts of 2.5 mM EDC aqueous solution were added to nanocapsule suspension and stirred for 20 min. The pH was adjusted to neutral and then 1 part of animal amino acid aqueous solution was added and stirred for 2 h. Finally, the mixture was centrifuged at 9000 rpm for 20 min, the precipitate was collected, washed 3 times with deionized water, and freeze-dried at -25℃ for 3 h to obtain plant root-oriented nanocapsules based on animal amino acids.

[0030] The following is a control experiment based on Example 1, with comparative examples 1 to 5, as detailed below: Comparative Example 1: Preparation method of plant root-directed nanocapsules based on animal amino acids: Comparative Example 1 is based on Example 1, with the following adjustments: chitosan with a degree of deacetylation of 80% and an average molecular weight of 50,000 was selected, while other processes remained unchanged. Specifically: S1: Preparation of modified chitosan-polylactic acid graft copolymer: (1) Under nitrogen and light-protected conditions, 8 parts of chitosan (degree of deacetylation of 80%, average molecular weight of 50,000) were added to 50 parts of 1-allyl-3-chloromethylimidazolium, and 0.75 parts of cinnamaldehyde were added to 5 parts of 1-allyl-3-chloromethylimidazolium. The mixture was stirred and mixed evenly at 75°C to prepare chitosan solution and cinnamaldehyde solution, and kept warm for later use; (2) Under nitrogen and light-protected conditions, 2.25 parts of acetic acid were added to the chitosan solution and stirred and mixed evenly. Then, cinnamaldehyde was slowly added dropwise. Aldehyde solution was added dropwise within 1 hour. After the addition was completed, the reaction was stirred for 9 hours to obtain modified chitosan solution. (3) Under nitrogen and light-protected conditions, 18 parts of L-lactide were added to the modified chitosan solution and stirred and mixed evenly at 115°C. Then, 0.3 parts of stannous octoate were added and stirred for 18 hours to stop the reaction. After the reaction was stopped, the solution was allowed to cool naturally to room temperature. Excess anhydrous ethanol was added and the solution was allowed to stand overnight. The precipitate was collected, washed three times with anhydrous ethanol, and finally dried under vacuum at 60°C for 3 hours to obtain modified chitosan-polylactic acid graft copolymer. S2: Preparation of plant root-oriented nanocapsules based on animal amino acids: (1) Under nitrogen and light-proof conditions, 5 parts of modified chitosan-polylactic acid graft copolymer were added to 25 parts of dichloromethane and stirred until homogeneous. Then, 9 parts of animal amino acid aqueous solution were added and stirred until homogeneous to obtain mixed primary emulsion. (2) Under nitrogen and light-proof conditions, 0.9 parts of Tween 80 were added to (1) and stirred at 550 rpm for 3 min. Then, the speed was adjusted to 300 rpm and stirred for 9 h under ventilation conditions to obtain nanocapsule suspension. (3) Under nitrogen and light-proof conditions, 4.5 parts of 2.5 mM EDC aqueous solution were added to nanocapsule suspension and stirred for 20 min. The pH was adjusted to neutral and then 1 part of animal amino acid aqueous solution was added and stirred for 2 h. Finally, the mixture was centrifuged at 9000 rpm for 20 min, the precipitate was collected, washed 3 times with deionized water, and freeze-dried at -25℃ for 3 h to obtain plant root-oriented nanocapsules based on animal amino acids.

[0031] Comparative Example 2: Preparation method of plant root-directed nanocapsules based on animal amino acids: Comparative Example 2 is based on Example 1, with the following adjustment: cinnamaldehyde was not used to modify chitosan, while other processes remained unchanged. Specifically: S1: Preparation of chitosan-polylactic acid graft copolymer: (1) Under nitrogen and light-protected conditions, 8 parts of chitosan (deacetylation degree of 90%, average molecular weight of 150,000) were added to 50 parts of 1-allyl-3-chloromethylimidazolium and stirred and mixed evenly at 75°C to prepare a chitosan solution, which was kept warm for later use; (2) Under nitrogen and light-protected conditions, 18 parts of L-lactide were added to the chitosan solution and stirred and mixed evenly at 115°C. Then 0.3 parts of stannous octoate were added and stirred and reacted for 18 hours. The reaction was stopped and allowed to cool naturally to room temperature. Excess anhydrous ethanol was added and allowed to stand overnight. The precipitate was collected, washed 3 times with anhydrous ethanol, and finally dried under vacuum at 60°C for 3 hours to obtain chitosan-polylactic acid graft copolymer; S2: Preparation of plant root-oriented nanocapsules based on animal amino acids: (1) Under nitrogen and light-protected conditions, 5 parts of chitosan-polylactic acid graft copolymer were added to 25 parts of dichloromethane and stirred until homogeneous. Then, 9 parts of animal amino acid aqueous solution were added and stirred until homogeneous to obtain a mixed primary emulsion. (2) Under nitrogen and light-protected conditions, 0.9 parts of Tween 80 were added to (1) and stirred at 550 rpm for 3 min. The stirring speed was then adjusted to 300 rpm and stirred for 9 h under ventilation conditions to obtain a nanocapsule suspension. (3) Under nitrogen and light-protected conditions, 4.5 parts of 2.5 mM EDC aqueous solution were added to the nanocapsule suspension and stirred for 20 min. The pH was adjusted to neutral and then 1 part of animal amino acid aqueous solution was added and stirred for 2 h. Finally, the mixture was centrifuged at 9000 rpm for 20 min, the precipitate was collected, washed 3 times with deionized water, and freeze-dried at -25℃ for 3 h to obtain plant root-oriented nanocapsules based on animal amino acids.

[0032] Comparative Example 3: Preparation method of plant root-directed nanocapsules based on animal amino acids: Comparative Example 3 is based on Example 1, with the following adjustment: polylactic acid is not grafted onto the modified chitosan, while other processes remain unchanged. Specifically: S1: Preparation of modified chitosan: (1) Under nitrogen and light-protected conditions, 8 parts of chitosan (deacetylation degree of 90%, average molecular weight of 150,000) were added to 50 parts of 1-allyl-3-chloromethylimidazolium, and 0.75 parts of cinnamaldehyde were added to 5 parts of 1-allyl-3-chloromethylimidazolium. The mixture was stirred and mixed evenly at 75°C to prepare chitosan solution and cinnamaldehyde solution. The mixture was kept warm for later use. (2) Under nitrogen and light-protected conditions, 2.25 parts of acetic acid were added to the chitosan solution and stirred and mixed evenly. Then, cinnamaldehyde solution was slowly added dropwise. The addition was completed within 1 hour. After the addition was completed, the reaction was stirred for 9 hours to obtain modified chitosan solution. The reaction was stopped and allowed to cool naturally to room temperature. Excess anhydrous ethanol was added to the solution and allowed to stand overnight. The precipitate was collected and washed 3 times with anhydrous ethanol. Finally, the mixture was vacuum dried at 60°C for 3 hours to obtain modified chitosan. S2: Preparation of plant root-guided nanocapsules based on animal amino acids: (1) Under nitrogen and light-protected conditions, 5 parts of modified chitosan were added to 25 parts of dichloromethane and stirred until homogeneous. Then, 9 parts of animal amino acid aqueous solution were added and stirred until homogeneous to obtain a mixed primary emulsion. (2) Under nitrogen and light-protected conditions, 0.9 parts of Tween 80 were added to (1) and stirred at 550 rpm for 3 min. The stirring speed was then adjusted to 300 rpm and stirred for 9 h under ventilation conditions to obtain a nanocapsule suspension. (3) Under nitrogen and light-protected conditions, 4.5 parts of 2.5 mM EDC aqueous solution were added to the nanocapsule suspension and stirred for 20 min. The pH was adjusted to neutral and then 1 part of animal amino acid aqueous solution was added and stirred for 2 h. Finally, the mixture was centrifuged at 9000 rpm for 20 min, the precipitate was collected, washed 3 times with deionized water, and freeze-dried at -25℃ for 3 h to obtain plant root-guided nanocapsules based on animal amino acids.

[0033] Comparative Example 4: Preparation method of plant root-directed nanocapsules based on animal amino acids: Comparative Example 4 is based on Example 1, with the following adjustment: no EDC aqueous solution was added for activation treatment, while other processes remained unchanged. Specifically: S1: Preparation of modified chitosan-polylactic acid graft copolymer: (1) Under nitrogen and light-protected conditions, 8 parts of chitosan (degree of deacetylation of 90%, average molecular weight of 150,000) were added to 50 parts of 1-allyl-3-chloromethylimidazolium, and 0.75 parts of cinnamaldehyde were added to 5 parts of 1-allyl-3-chloromethylimidazolium. The mixture was stirred and mixed evenly at 75°C to prepare chitosan solution and cinnamaldehyde solution, and kept warm for later use; (2) Under nitrogen and light-protected conditions, 2.25 parts of acetic acid were added to the chitosan solution and stirred and mixed evenly. Then, cinnamaldehyde was slowly added dropwise to the solution. Cinnamaldehyde solution was added dropwise within 1 hour. After the addition was completed, the reaction was stirred for 9 hours to obtain modified chitosan solution. (3) Under nitrogen and light-proof conditions, 18 parts of L-lactide were added to the modified chitosan solution and stirred and mixed evenly at 115°C. Then 0.3 parts of stannous octoate were added and stirred for 18 hours to stop the reaction. After the reaction was stopped, the solution was allowed to cool naturally to room temperature. Excess anhydrous ethanol was added and the solution was allowed to stand overnight. The precipitate was collected, washed three times with anhydrous ethanol, and finally dried under vacuum at 60°C for 3 hours to obtain modified chitosan-polylactic acid graft copolymer. S2: Preparation of plant root-oriented nanocapsules based on animal amino acids: (1) Under nitrogen and light-proof conditions, 5 parts of modified chitosan-polylactic acid graft copolymer were added to 25 parts of dichloromethane and stirred until homogeneous. Then, 9 parts of animal amino acid aqueous solution were added and stirred until homogeneous to obtain mixed primary emulsion. (2) Under nitrogen and light-proof conditions, 0.9 parts of Tween 80 were added to (1) and stirred at 550 rpm for 3 min. The stirring speed was then adjusted to 300 rpm and stirred for 9 h under ventilation conditions to obtain nanocapsule suspension. (3) Under nitrogen and light-proof conditions, 1 part of animal amino acid aqueous solution was added to nanocapsule suspension and stirred for 2 h. Finally, the mixture was centrifuged at 9000 rpm for 20 min, the precipitate was collected, washed 3 times with deionized water, and freeze-dried at -25℃ for 3 h to obtain plant root-oriented nanocapsules based on animal amino acids.

[0034] Comparative Example 5: Preparation method of plant root-directed nanocapsules based on animal amino acids: Comparative Example 5 is based on Example 1, with the following adjustment: animal amino acids are not grafted onto the outside of the nanocapsules, while other processes remain unchanged. Specifically: S1: Preparation of modified chitosan-polylactic acid graft copolymer: (1) Under nitrogen and light-protected conditions, 8 parts of chitosan (degree of deacetylation of 90%, average molecular weight of 150,000) were added to 50 parts of 1-allyl-3-chloromethylimidazolium, and 0.75 parts of cinnamaldehyde were added to 5 parts of 1-allyl-3-chloromethylimidazolium. The mixture was stirred and mixed evenly at 75°C to prepare chitosan solution and cinnamaldehyde solution, and kept warm for later use; (2) Under nitrogen and light-protected conditions, 2.25 parts of acetic acid were added to the chitosan solution and stirred and mixed evenly. Then, cinnamaldehyde was slowly added dropwise to the solution. Cinnamaldehyde solution was added dropwise within 1 hour. After the addition was completed, the reaction was stirred for 9 hours to obtain modified chitosan solution. (3) Under nitrogen and light-proof conditions, 18 parts of L-lactide were added to the modified chitosan solution and stirred and mixed evenly at 115°C. Then 0.3 parts of stannous octoate were added and stirred for 18 hours to stop the reaction. After the reaction was stopped, the solution was allowed to cool naturally to room temperature. Excess anhydrous ethanol was added and the solution was allowed to stand overnight. The precipitate was collected, washed three times with anhydrous ethanol, and finally dried under vacuum at 60°C for 3 hours to obtain modified chitosan-polylactic acid graft copolymer. S2: Preparation of plant root-oriented nanocapsules based on animal amino acids: (1) Under nitrogen and light-protected conditions, 5 parts of modified chitosan-polylactic acid graft copolymer were added to 25 parts of dichloromethane and stirred until homogeneous. Then, 9 parts of animal amino acid aqueous solution were added and stirred until homogeneous to obtain a mixed primary emulsion. (2) Under nitrogen and light-protected conditions, 0.9 parts of Tween 80 were added to (1) and stirred at 550 rpm for 3 min. The stirring speed was then adjusted to 300 rpm and stirred for 9 h under ventilation conditions to obtain a nanocapsule suspension. Finally, the suspension was centrifuged at 9000 rpm for 20 min, the precipitate was collected, washed 3 times with deionized water, and freeze-dried at -25℃ for 3 h to obtain plant root-oriented nanocapsules based on animal amino acids.

[0035] Performance testing: The following performance tests were conducted on the plant root-guided nanocapsules based on animal amino acids prepared in Examples 1-3 and Comparative Examples 1-5, as detailed below: 1. Guiding performance test: (1) Prepare Hogland nutrient solution containing 1wt% agarose, use hydroponics to cultivate several tomato seeds, cultivate at 25℃ for 7 days to obtain tomato seedlings; (2) Transfer the tomato seedlings to clean soil obtained from the same place, and add 2g of plant root guiding nanocapsules 2cm away from the roots. After cultivating at 25℃ for 7 days, measure the amino acid residue on both sides of the roots and the fresh weight of the roots. During this period, keep the soil moisture at 65%.

[0036] 2. Antibacterial performance test: (1) Hogland nutrient solution containing 1 wt% agarose was prepared, and several tomato seeds were cultured using hydroponics. After 7 days of cultivation at 25℃, tomato seedlings were obtained; (2) The bacterial count (tomato bacterial wilt fungus) in 10 mL was 10 7 ~10 8 The soil was inoculated with a bacterial suspension of CFU / mL at a ratio of 1 kg to obtain pathogenic soil; (3) Tomato seedlings were planted in pathogenic soil and 2 g of plant root-guided nanocapsules were added to the roots. After culturing at 25℃ for 7 days, the residual amino acid content on both sides of the roots and the fresh weight of the roots were measured. During this period, the soil moisture was kept at 65%.

[0037] The test results are shown in Table 1 below: Table 1

[0038] Results Analysis: As shown in Table 1 above, the plant root-guided nanocapsules based on animal amino acids prepared in this invention have a natural affinity with plant roots and a sustained-release effect, which significantly promotes plant root growth and also has significant disease resistance.

[0039] Finally, it should be noted that the above descriptions are merely preferred embodiments of the present invention and are not intended to limit the present invention. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of the technical features. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for the preparation of animal amino acid based plant root guiding nanocapsules, characterized by: Includes the following steps: S1: Chitosan is grafted with cinnamaldehyde and then grafted with lactide to obtain a modified chitosan-polylactic acid graft copolymer. S2: After the modified chitosan-polylactic acid graft copolymer is coated with animal amino acids, it is activated by adding EDC aqueous solution, and then it is grafted with animal amino acids to obtain plant root-guided nanocapsules based on animal amino acids.

2. The method for preparing plant root-directed nanocapsules based on animal amino acids according to claim 1, characterized in that: The specific process of S1 is as follows: (1) Under nitrogen and light-protected conditions, chitosan and cinnamaldehyde were added to 1-allyl-3-chloromethylimidazolium and stirred and mixed evenly at 70~80℃ to prepare chitosan solution and cinnamaldehyde solution, which were kept warm for later use. (2) Under nitrogen and light-protected conditions, add acetic acid to the chitosan solution, stir and mix evenly, then slowly add cinnamaldehyde solution dropwise over 1 hour. After the addition is complete, continue stirring and reacting for 6 to 12 hours to obtain the modified chitosan solution. (3) Under nitrogen and light-protected conditions, lactide was added to the modified chitosan solution and stirred and mixed evenly at 110~120℃. Stannous octoate was then added to the solution and the mixture was stirred and kept warm for 12~24h. The reaction was then stopped and allowed to cool naturally to room temperature. Excess anhydrous ethanol was added to the solution and allowed to stand overnight. The precipitate was collected, washed, and dried under vacuum to obtain the modified chitosan-polylactic acid graft copolymer. The modified chitosan-polylactic acid graft copolymer comprises the following raw material components in parts by weight: 6-10 parts chitosan, 0.5-1 part cinnamaldehyde, 50-70 parts 1-allyl-3-chloromethylimidazolium, 2-2.5 parts acetic acid, 12-24 parts lactide, and 0.2-0.4 parts stannous octoate.

3. The method for preparing plant root-directed nanocapsules based on animal amino acids according to claim 2, characterized in that: All raw material components of the modified chitosan-polylactic acid graft copolymer need to be vacuum dried overnight at 55~65℃.

4. The method for preparing plant root-directed nanocapsules based on animal amino acids according to claim 2, characterized in that: The degree of deacetylation of the chitosan is 85-95%, and the molecular weight is ≥100,000.

5. The method for preparing plant root-directed nanocapsules based on animal amino acids according to claim 1, characterized in that: The specific process of S2 is as follows: (1) Under nitrogen and light-protected conditions, the modified chitosan-polylactic acid graft copolymer was added to dichloromethane and stirred until homogeneous. Then, an aqueous solution of animal amino acids was added and stirred until homogeneous to obtain a mixed primary emulsion. (2) Under nitrogen and light-proof conditions, add emulsifier to (1), stir and mix at a speed of 500-600 rpm for 2-5 min, then adjust the speed to 200-400 rpm, stir and mix under ventilation conditions for 6-12 h to obtain nanocapsule suspension; (3) Under nitrogen and light-protected conditions, add EDC aqueous solution to the nanocapsule suspension, stir and mix for 10-30 min, adjust the pH to neutral, then add animal amino acid aqueous solution II, stir and mix for 1-3 h, and finally centrifuge at 8000-10000 rpm for 10-30 min, collect the precipitate, wash, freeze dry, and obtain plant root-guided nanocapsules based on animal amino acids. The plant root-guided nanocapsules based on animal amino acids comprise the following raw material components in parts by weight: 4-6 parts of modified chitosan-polylactic acid graft copolymer, 20-30 parts of dichloromethane, 8-12 parts of animal amino acid aqueous solution, 0.6-1.2 parts of emulsifier, and 3-6 parts of EDC aqueous solution.

6. The method for preparing plant root-directed nanocapsules based on animal amino acids according to claim 5, characterized in that: The animal amino acid aqueous solution was prepared according to the preparation method disclosed in CN111375618A, and the amino acid content was 50~80g / L.

7. The method for preparing plant root-directed nanocapsules based on animal amino acids according to claim 5, characterized in that: The emulsifier is a nonionic emulsifier.

8. The method for preparing plant root-directed nanocapsules based on animal amino acids according to claim 5, characterized in that: The concentration of the EDC aqueous solution is 2~3 mM.

9. Plant root-directed nanocapsules based on animal amino acids prepared by the preparation method according to any one of claims 1 to 8.

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