Liquid film-forming agent for preventing crops from freezing injury due to late spring coldness and preparation method of liquid film-forming agent

Abstract

The invention relates to the field of agricultural biochemical engineering and plant protection, and particularly discloses a liquid film-forming agent for preventing crops from freezing injury due to late spring coldness and a preparation method of the liquid film-forming agent. The film-forming agent is prepared from carboxymethyl chitosan, sodium carboxymethyl cellulose, kasugamycin, brassinolide, glycol and glycerol mixed dihydric alcohol, Tween 80 and deionized water according to a specific proportion. The preparation method comprises the key processes of step-by-step dissolution, mixing, functional component addition and the like. Organic fusion of physical barrier, ice nucleus inactivation and physiological regulation is realized through multiple action mechanisms of natural polymer composite film forming, biopesticide targeted sterilization, plant hormone induced cold resistance and dihydric alcohol synergistic antifreezing. The product can provide efficient protection at a low temperature of-4 DEG C, significantly reduces the frost damage rate, has the characteristic of 28-day natural rapid degradation, and solves the technical bottlenecks of single function, difficult degradation and poor environmental compatibility of a traditional film agent.

Description

Technical Field

[0001] This invention belongs to the fields of agricultural biochemicals and plant protection, specifically a liquid film-forming agent for preventing crop damage from late spring frost and its preparation method. Background Technology

[0002] When plants suffer from low-temperature damage such as "late spring frost," the damage mechanism mainly stems from two aspects: First, the low temperature directly triggers intracellular freezing within the cells, leading to cell structure damage; second, ice-nucleating bacteria epiphytic on the plant surface secrete ice-nucleating proteins that can induce the formation of ice crystals from surface water at higher temperatures (such as -2℃ to -5℃), acting as a "catalyst" for frost damage. Therefore, an ideal frost protection strategy needs to comprehensively address external physical barriers, inhibit ice-nucleating activity, and enhance the plant's own resistance.

[0003] Currently, agricultural production often employs single-function measures to combat frost. For example, spraying paraffin or mineral oil emulsions can form a physical protective film on the plant surface, isolating it from cold air. However, these purely physical film-forming agents generally have significant drawbacks: their film is dense and has poor air permeability, and long-term coverage can easily affect the plant's normal gas exchange and photosynthesis, potentially leading to physiological suffocation. More importantly, these polymeric films are difficult to degrade in the natural environment, with a residual period of over 60 days, posing a continuous burden on the farmland's ecological environment. Although they provide some physical insulation, they completely fail to address the core issues of biological ice-forming threats from ice-nucleating bacteria and the plant's insufficient inherent cold resistance. Therefore, developing a novel formulation that is both highly effective in preventing frost damage and rapidly degradable, while comprehensively addressing both biological and physiological factors causing frost injury, has become an urgent technical challenge. Summary of the Invention

[0004] To address the aforementioned technical problems, this invention provides a liquid film-forming agent for preventing crop damage from late spring frost and its preparation method, thereby solving the problem that existing film-forming agents have a single function and cannot synergistically inhibit ice-nucleating bacteria and enhance the plant's own cold resistance.

[0005] A liquid film-forming agent for protecting crops from late spring frost damage, comprising the following components by weight percentage:

[0006] Carboxymethyl chitosan 2-6%, sodium carboxymethyl cellulose 1-4%, kasugamycin 0.05-0.2%, brassinolide 0.001-0.01%, diol 5-10%, Tween 80 1-3%, balance deionized water;

[0007] The diol is a mixture of ethylene glycol and glycerol.

[0008] Preferably, the content of carboxymethyl chitosan is 4%, the content of sodium carboxymethyl cellulose is 2.5%, the content of kasugamycin is 0.08%, the content of brassinolide is 0.003%, the content of diol is 9.5%, and the content of Tween 80 is 1.8%.

[0009] A method for preparing a liquid film-forming agent for preventing crop damage from late spring frost, as described above, includes the following steps:

[0010] S1. Dissolving carboxymethyl chitosan: Carboxymethyl chitosan is mixed with a portion of glycerol for pre-dispersion, and then deionized water is slowly added while continuously stirring. After complete dissolution, stirring is continued until the solution is transparent and free of precipitate, thus obtaining solution A.

[0011] S2. Dissolving sodium carboxymethyl cellulose: Dissolve sodium carboxymethyl cellulose in an alkaline solution, heat and stir until completely dissolved to obtain solution B;

[0012] S3. Mixing: Mix solution A with solution B, add Tween 80 and the remaining diol component, mix evenly and adjust the pH value to obtain mixture C;

[0013] S4. Adding the drug: After pre-dissolving kasugamycin and brassinolide in an organic solvent, add them to the mixture C and stir in the dark to obtain the liquid film-forming agent.

[0014] Preferably, in step S1, the glycerol is used to pre-disperse the carboxymethyl chitosan powder to break up the aggregates.

[0015] Preferably, in step S2, the alkaline solution is a sodium hydroxide solution with a mass concentration of 0.5% and the heating temperature is 60°C.

[0016] Preferably, in step S3, the pH value is adjusted to 6.2 ± 0.3.

[0017] Preferably, in step S4, the kasugamycin is pre-dissolved in dimethyl sulfoxide, and the brassinolide is pre-dissolved in ethanol.

[0018] Preferably, in step S1, after all the deionized water has been added, continue stirring at room temperature for 30 minutes.

[0019] In step S4, the stirring time in the dark is 30 minutes.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

[0021] By integrating the targeted fungicide kasugamycin, the plant cold resistance inducer brassinolide, and a specific ratio of ethylene glycol-glycerol diol antifreeze system, a single film agent can simultaneously possess the synergistic effects of inactivating ice-nucleating bacteria, inhibiting ice-nucleating protein activity, and enhancing the plant's own cold resistance from a physiological perspective. This triple mechanism of action of "physical isolation-chemical control-physiological regulation" promotes each other, ultimately achieving excellent antifreeze efficacy by significantly reducing the frost damage rate of fragrant pear during flowering from the conventional 42% to below 9% at a low temperature of -4℃.

[0022] By eliminating the use of non-degradable paraffin and heavy metals that can cause pesticide damage, and instead using only biodegradable natural polymers and environmentally friendly small molecule compounds, the film can decompose rapidly and naturally after fulfilling its protective mission. It leaves no harmful residues, ensuring the safety of crops while significantly reducing the pressure on the environment, thus achieving a balance between increased agricultural production and eco-friendliness. Attached Figure Description

[0023] Figure 1 This is a flowchart illustrating the preparation method of the liquid film-forming agent for preventing crop damage from late spring frost, as described in this invention. Detailed Implementation

[0024] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. 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.

[0025] like Figure 1 As shown:

[0026] Example 1

[0027] A formulation of a liquid film-forming agent for preventing crop damage from late spring frost (mass / g of each component based on the preparation of approximately 1000g of finished product):

[0028] Carboxymethyl chitosan: 20.0 g (2.0%); Sodium carboxymethyl cellulose: 10.0 g (1.0%); Kasugamycin: 0.5 g (0.05%); Brassinolide: 0.01 g (0.001%); Diol: 50.0 g (5.0%); Tween 80: 10.0 g (1.0%); Balance: deionized water;

[0029] Of the 50.0 g diols, 25.0 g is ethylene glycol and 25.0 g is glycerol.

[0030] The preparation method of the above-mentioned liquid film-forming agent for preventing crop damage from late spring frost:

[0031] Step S1: Dissolve carboxymethyl chitosan: Weigh 20.0 g of carboxymethyl chitosan into a beaker; weigh 5.0 g of glycerol and add it to the beaker, then manually stir and mix with the carboxymethyl chitosan powder for 2-3 minutes to pre-disperse and break up the powder agglomerates; place the beaker under a stirrer and start stirring at 300 rpm; slowly and in batches add about 650 g of deionized water; after the powder is completely dispersed, add the remaining 20.0 g of glycerol; after all the deionized water has been added, continue stirring at 300 rpm for 30 minutes at room temperature (25℃) until the solution is homogeneous and transparent, with no visible particles or precipitates, to obtain solution A.

[0032] Step S2, Dissolving sodium carboxymethyl cellulose: In another beaker, weigh 10.0 g of sodium carboxymethyl cellulose; add 200 g of 0.5% sodium hydroxide solution (prepared from 1.0 g NaOH and 199.0 g of deionized water taken from the total amount of the formula); place the beaker in a 60°C water bath and stir continuously at 300 rpm for about 45 minutes until the sodium carboxymethyl cellulose is completely dissolved and the solution is transparent, thus obtaining solution B.

[0033] Step S3, Mixing and Adjustment: While stirring solution A at 300 rpm, slowly pour solution B into it; after mixing evenly, add 10.0 g Tween 80 and 25.0 g ethylene glycol; continue stirring for 15 minutes; adjust the pH of the mixture to 6.2 with dilute hydrochloric acid solution or dilute sodium hydroxide solution to obtain mixture C.

[0034] Step S4, Dosing and Post-treatment: Weigh 0.5 g of kasugamycin and 0.01 g of brassinolide, respectively; dissolve kasugamycin in 1.0 mL of dimethyl sulfoxide (DMSO) and brassinolide in 0.5 mL of anhydrous ethanol, respectively, and pre-dissolve them; under light-protected conditions, add the two pre-dissolved solutions to the mixture C in sequence; after the dosing is completed, wrap the container with aluminum foil and stir at 300 rpm for 30 minutes in the dark to obtain the liquid film-forming agent for preventing crop frost damage in late spring.

[0035] Example 2

[0036] A formulation of a liquid film-forming agent for preventing crop damage from late spring frost (mass / g of each component based on the preparation of approximately 1000g of finished product):

[0037] Carboxymethyl chitosan: 40.0 g (4.0%); Sodium carboxymethyl cellulose: 25.0 g (2.5%); Kasugamycin: 0.8 g (0.08%); Brassinolide: 0.03 g (0.003%); Diol: 95.0 g (9.5%); Tween 80: 18.0 g (1.8%); Balance: deionized water;

[0038] Of the 95.0 g diols, 45.0 g is ethylene glycol and 50.0 g is glycerol.

[0039] The preparation method of the above-mentioned liquid film-forming agent for preventing crop damage from late spring frost:

[0040] Step S1: Dissolve carboxymethyl chitosan: Weigh 40.0 g of carboxymethyl chitosan into a beaker; weigh 10.0 g of glycerol and add it to the beaker, then manually stir to mix and pre-disperse; place the beaker under a stirrer and stir at 350 rpm; slowly and in batches add about 600 g of deionized water; add the remaining 40.0 g of glycerol; after all the water has been added, continue stirring at room temperature for 30 minutes until the solution is homogeneous and transparent, to obtain solution A.

[0041] Step S2, Dissolving sodium carboxymethyl cellulose: Weigh 25.0 g of sodium carboxymethyl cellulose into a beaker; add 500 g of 0.5% sodium hydroxide solution (prepared from 2.5 g NaOH and 497.5 g of deionized water taken from the total amount of the formula); place the beaker in a 60°C water bath and stir continuously at 350 rpm for about 60 minutes until completely dissolved to obtain solution B.

[0042] Step S3, Mixing and Adjustment: Slowly pour solution B into solution A (350 rpm) while stirring; after mixing evenly, add 18.0 g Tween 80 and 45.0 g ethylene glycol; continue stirring for 15 minutes; adjust the pH of the mixture to 6.2 to obtain mixture C.

[0043] Step S4, Drug Addition and Post-processing: Weigh 0.8 g of kasugamycin and 0.03 g of brassinolide, respectively; dissolve kasugamycin in 2.0 mL of DMSO and brassinolide in 1.0 mL of ethanol for pre-dissolution; under light-protected conditions, add the two drug solutions to the mixture C; after adding the drugs, wrap with aluminum foil and stir at 350 rpm for 30 minutes in the dark to obtain the finished product.

[0044] Example 3

[0045] A formulation of a liquid film-forming agent for preventing crop damage from late spring frost (mass / g of each component based on the preparation of approximately 1000g of finished product):

[0046] Carboxymethyl chitosan: 60.0 g (6.0%); Sodium carboxymethyl cellulose: 40.0 g (4.0%); Kasugamycin: 2.0 g (0.2%); Brassinolide: 0.1 g (0.01%); Diol: 100.0 g (10.0%); Tween 80: 30.0 g (3.0%); Balance: deionized water;

[0047] Of the 100.0 g diols, 50.0 g is ethylene glycol and 50.0 g is glycerol.

[0048] The preparation method of the above-mentioned liquid film-forming agent for preventing crop damage from late spring frost:

[0049] Step S1: Dissolve carboxymethyl chitosan: Weigh 60.0 g of carboxymethyl chitosan into a beaker; weigh 10.0 g of glycerol and add it, then manually pre-disperse it; place the beaker under a stirrer and stir at 400 rpm; slowly and in batches add about 550 g of deionized water; add the remaining 40.0 g of glycerol; after all the water has been added, continue stirring at room temperature for 30 minutes until the solution is homogeneous and transparent, to obtain solution A.

[0050] Step S2, Dissolving sodium carboxymethyl cellulose: Weigh 40.0 g of sodium carboxymethyl cellulose into a beaker; add 800 g of 0.5% sodium hydroxide solution (prepared from 4.0 g NaOH and 796.0 g of deionized water taken from the total amount of the formula); place the beaker in a 60℃ water bath and stir continuously at 400 rpm for about 60-75 minutes until completely dissolved to obtain solution B.

[0051] Step S3, Mixing and Adjustment: Slowly pour solution B into solution A (400 rpm) while it is being stirred; after mixing evenly, add 30.0 g Tween 80 and 50.0 g ethylene glycol; continue stirring for 15 minutes; adjust the pH of the mixture to 6.2 to obtain mixture C.

[0052] Step S4, Drug Addition and Post-processing: Weigh 2.0 g of kasugamycin and 0.1 g of brassinolide, respectively; dissolve kasugamycin in 5.0 mL of DMSO and pre-dissolve brassinolide in 2.0 mL of ethanol; under light-protected conditions, add the two drug solutions to the mixture C; after adding the drugs, wrap with aluminum foil and stir at 400 rpm for 30 minutes in the dark to obtain the finished product.

[0053] Experimental Example: Performance Verification of Liquid Film-Forming Agent for Protecting Crops from Late Spring Cold Damage

[0054] 1. Experimental Objective

[0055] The antifreeze film agent provided in Example 2 of this invention demonstrates significant comprehensive advantages over commercially available products or formulations with single functions or missing components in terms of ice nucleation bacteria inactivation, antifreeze effect (simulation), induction of plant cold resistance, and environmental friendliness.

[0056] 2. Experimental Materials and Grouping

[0057] Reference group: Liquid film-forming agent prepared in Example 2 of the present invention.

[0058] Comparative Example 1: Commercially available paraffin film (main component is paraffin emulsion); representing traditional physical barrier film.

[0059] Comparative Example 2: Commercially available kasugamycin wettable powder; representing a single bactericide.

[0060] Comparative Example 3: Component-deficient comparative example; referring to the formulation of Example 2, without the addition of kasugamycin and brassinolide, containing only film-forming substances (carboxymethyl chitosan, CMC) and antifreeze (diol), Tween 80, and prepared by the same method; used to verify the necessity of bioactive components.

[0061] Comparative Example 4: Referring to the formulation of Example 2, the diol (ethylene glycol + glycerol) was replaced with an equal amount of commercially available single antifreeze agent glycerol, while the other components and preparation methods remained unchanged; this was used to verify the synergistic antifreeze effect of a specific diol system.

[0062] Blank control group: sprayed with an equal amount of deionized water.

[0063] 3. Experimental Design

[0064] 3.1 Determination of Inhibition Rate of Ice-Nucleating Bacteria

[0065] Methods: The inhibition zone method was used; the suspension of Pseudomonas syringae was spread onto an agar plate; a filter paper disc (5 mm in diameter) soaked in the liquid from each experimental group was placed in the center of the plate. After incubation at 28°C for 48 hours, the diameter of the inhibition zone was measured.

[0066] Evaluation index: Diameter of the inhibition zone (mm); the larger the diameter, the stronger the immediate bactericidal / bacteriostatic effect.

[0067] 3.2 Simulated antifreeze effect determination

[0068] Methods: Differential scanning calorimetry (DSC) was used. Liquid samples (20 μL) from each experimental group were placed in the DSC sample pan and cooled from 20 °C to -30 °C at a rate of 5 °C / min. The exothermic peak of crystallization was recorded.

[0069] Evaluation index: supercooling point (crystallization initiation temperature, °C); the lower the supercooling point, the stronger the ability of the liquid to remain liquid at lower temperatures and delay the formation of ice crystals.

[0070] 3.3 Plant Frost Simulation Experiment

[0071] Test materials: potted fragrant pear seedlings in the flowering period, with uniform growth.

[0072] Treatment: Spray the corresponding pesticide solution evenly on both sides of the leaves of each group of plants until it just drips, and spray the blank control group with water; let them air dry to form a film.

[0073] Frost treatment: Place the treated plants in an artificial climate chamber and lower the temperature from 15°C to -4°C at a rate of 2°C / h. Maintain the temperature at -4°C for 4 hours, and then slowly warm the plants back to 15°C.

[0074] Damage assessment: Seven days after frost treatment, the damage rate of flowers on each plant was calculated (based on the browning and wilting of petals).

[0075] Evaluation index: Frost damage rate of flowers (%).

[0076] 3.4 Observation of membrane degradation

[0077] Method: The liquid of each experimental group was evenly coated on the surface of a glass plate to form a thin film, and then placed in an outdoor natural environment (avoiding direct rain).

[0078] Evaluation indicators: Regularly observe and record the time (in days) when obvious cracks and peeling appear on the film, as well as the visual residue after 28 days.

[0079] 4. Experimental Data Recording Table

[0080] experimental group Inhibition of ice-nucleated bacteria (inhibition zone diameter, mm) Simulated antifreeze effect (DSC supercooling point, °C) Plant frost damage (flower damage rate, %) Membrane degradation (residual condition after 28 days) Reference group 18.5 -12.5 8.7 The membrane completely cracked and pulverized, with a residue rate of <5%. Comparative Example 1 0 -2.1 38.5 The film layer is intact, slightly glossy, and has a residue rate >95%. Comparative Example 2 16.0 -0.8 41.2 No film formation, not applicable Comparative Example 3 8.0 (Chitosan-only antibacterial properties) -11.8 25.6 The membrane was visibly cracked and curled, with a residue rate of approximately 20%. Comparative Example 4 18.3 -9.5 15.4 The membrane cracked and pulverized, with a residue rate of approximately 10%. Blank control 0 0 42.3 not applicable

[0081] 5. Analysis of Experimental Results

[0082] Based on the data in the table above, the following conclusions can be drawn:

[0083] Excellent overall antifreeze performance: The control group exhibited the best overall performance. It had the largest diameter of the ice-nucleated bacteria inhibition zone (18.5 mm), demonstrating the synergistic bactericidal effect of carboxymethyl chitosan and kasugamycin; the lowest DSC supercooling point (-12.5℃), significantly better than Comparative Example 4 (-9.5℃) which used glycerol alone, verifying the super-strong antifreeze synergistic effect of the specific ethylene glycol / glycerol binary system; finally, its flower frost damage rate was the lowest (8.7%), far lower than all comparative examples and the blank control group.

[0084] The damage rate of Comparative Example 3 (lacking kasugamycin and brassinolide) (25.6%) was significantly higher than that of the control group; this indicates that although it has good film-forming properties and basic antifreeze ability, the lack of targeted bactericidal and endogenous cold resistance induction greatly reduces its overall protective effect; its small inhibition zone (8.0 mm) is derived solely from chitosan, demonstrating the importance of kasugamycin for achieving efficient inactivation of ice-nucleated bacteria.

[0085] The antifreeze supercooling point (-9.5℃) and damage rate (15.4%) of Comparative Example 4 (antifreeze replacement) were worse than those of the reference group.

[0086] The environmental friendliness is outstanding: the control group and comparative examples 3 and 4 all showed good biodegradability after 28 days, and the membranes basically disintegrated; while comparative example 1 (paraffin membrane) showed almost no degradation.

[0087] Conclusion: The comparative experimental data in this series clearly show that in the formulation of Example 2 of this invention, the absence or replacement of any core functional component will lead to a deficiency in one aspect of antifreeze, sterilization or stress-induced resistance, thereby affecting the final comprehensive protective effect. This invention integrates multiple antifreeze mechanisms into one, achieving excellent antifreeze effect (damage rate <9%) while also having rapid biodegradability, effectively overcoming the shortcomings of existing technologies with single function and heavy environmental burden.

[0088] The embodiments of the present invention are given for the purposes of illustration and description. Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention. Any changes, modifications, substitutions and variations made by those skilled in the art to the above embodiments within the scope of the present invention should be included within the protection scope of the present invention.

Claims

1. A liquid film-forming agent for preventing crop damage from late spring frost, characterized in that, It consists of the following components by mass percentage: Carboxymethyl chitosan 2-6%, sodium carboxymethyl cellulose 1-4%, kasugamycin 0.05-0.2%, brassinolide 0.001-0.01%, diol 5-10%, Tween 80 1-3%, balance deionized water; The diol is a mixture of ethylene glycol and glycerol.

2. The liquid film-forming agent for preventing crop damage from late spring frost according to claim 1, characterized in that, The content of carboxymethyl chitosan is 4%, the content of sodium carboxymethyl cellulose is 2.5%, the content of kasugamycin is 0.08%, the content of brassinolide is 0.003%, the content of diol is 9.5%, and the content of Tween 80 is 1.8%.

3. A method for preparing a liquid film-forming agent for preventing crop damage from late spring frost as described in any one of claims 1 or 2, characterized in that, Includes the following steps: S1. Dissolving carboxymethyl chitosan: Carboxymethyl chitosan is mixed with a portion of glycerol for pre-dispersion, and then deionized water is slowly added while continuously stirring. After complete dissolution, stirring is continued until the solution is transparent and free of precipitate, thus obtaining solution A. S2. Dissolving sodium carboxymethyl cellulose: Dissolve sodium carboxymethyl cellulose in an alkaline solution, heat and stir until completely dissolved to obtain solution B; S3. Mixing: Mix solution A with solution B, add Tween 80 and the remaining diol component, mix evenly and adjust the pH value to obtain mixture C; S4. Adding the drug: After pre-dissolving kasugamycin and brassinolide in an organic solvent, add them to the mixture C and stir in the dark to obtain the liquid film-forming agent.

4. The preparation method of the liquid film-forming agent for preventing crop frost damage according to claim 3, characterized in that, In step S1, the glycerol is used to pre-disperse the carboxymethyl chitosan powder to break up the aggregates.

5. The preparation method of the liquid film-forming agent for preventing crop damage from late spring frost according to claim 3, characterized in that, In step S2, the alkaline solution is a sodium hydroxide solution with a mass concentration of 0.5% and a heating temperature of 60°C.

6. The preparation method of the liquid film-forming agent for preventing crop frost damage according to claim 4, characterized in that, In step S3, the pH value is adjusted to 6.2 ± 0.

3.

7. The preparation method of the liquid film-forming agent for preventing crop frost damage according to claim 3, characterized in that, In step S4, the kasugamycin is pre-dissolved with dimethyl sulfoxide, and the brassinolide is pre-dissolved with ethanol.

8. The preparation method of the liquid film-forming agent for preventing crop damage from late spring frost according to claim 3, characterized in that, In step S1, after all the deionized water has been added, continue stirring at room temperature for 30 minutes; in step S4, stir for 30 minutes in the dark.

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