Fertilizer synergist based on biological enzyme regulation and control and preparation method thereof

Abstract

The invention discloses a fertilizer synergist based on biological enzyme regulation and control and a preparation method thereof, and belongs to the technical field of fertilizer synergists. The preparation method comprises the following steps: mixing beta-cyclodextrin, a 50wt% polyethyleneimine aqueous solution and an immobilized enzyme compound for reaction to obtain a mixture; rice hull ash and a 1mol / L sodium hydroxide solution are mixed and react to obtain rice hull ash sol; butyl titanate and manganese sulfate are mixed to obtain a first solution, and then a 95% ethanol solution is added for mixed reaction to obtain manganese-doped titanium oxide; mixing and reacting the mixture, the rice hull ash sol and manganese-doped titanium oxide to obtain a modified enzyme compound; and mixing the modified enzyme compound, potassium humate and magnesium sulfate to obtain the fertilizer synergist. The prepared fertilizer synergist plays a role in improving the fertilizer utilization rate, improving soil, enhancing crop resistance and the like through a multi-way synergistic effect.

Description

Technical Field

[0001] This invention relates to the field of fertilizer synergists, specifically to a fertilizer synergist based on bio-enzyme regulation and its preparation method. Background Technology

[0002] Against the backdrop of global sustainable agricultural development, improving fertilizer utilization efficiency and reducing resource waste and environmental pollution have become core issues in agricultural technology. With increasing population growth and growing constraints on arable land resources, ensuring both food yield and quality while simultaneously protecting the ecological environment places higher demands on modern agricultural technologies. In traditional agricultural production, while excessive application of chemical fertilizers may have increased crop yields in the short term, in the long run, it not only leads to low fertilizer utilization rates but also causes soil degradation problems such as soil compaction, acidification, and decreased organic matter. Furthermore, it poses ecological risks such as eutrophication of water bodies and increased greenhouse gas emissions through surface runoff and groundwater infiltration, severely hindering the healthy cycle of the agricultural ecosystem.

[0003] Against this backdrop, fertilizer synergists, as a key technology for improving nutrient utilization efficiency, have received widespread attention. Bioenzymes, as a class of biomolecules with highly efficient catalytic activity, play a crucial role in soil nutrient transformation. They can promote the activation and release of insoluble nutrients such as organic nitrogen, phosphorus, and potassium in the soil through catalytic hydrolysis and redox reactions, converting ineffective nutrients into readily absorbable forms for crops, thereby reducing fertilizer usage and improving crop absorption efficiency. However, naturally occurring free bioenzymes face challenges in the soil environment, including susceptibility to inactivation due to factors such as temperature, pH, and heavy metal ions, short half-lives, and difficulty in recycling and reuse, which significantly limit their practical application in agricultural production.

[0004] Patent CN117362101A discloses a fertilizer synergist and its preparation method. Its main components are polyaspartic acid, γ-polyglutamic acid, p-benzoquinone, calcium diphenylureasulfonate, and trace elements, aiming to improve fertilizer utilization efficiency and crop yield through the synergistic effect of these components. However, this technical solution mainly relies on the combination of chemical substances to achieve fertilizer synergy, lacking in-depth research on the regulatory mechanisms of biological enzymes. It fails to effectively address the stability issues of biological enzymes in complex soil environments and does not fully explore the synergistic effects between biological enzymes and other components to further enhance the slow-release performance of fertilizers and crop absorption efficiency.

[0005] Therefore, providing a fertilizer enhancer with good stability and high fertilizer utilization rate is an important problem that urgently needs to be solved in this field. Summary of the Invention

[0006] To address the problems in the prior art, this invention provides a fertilizer synergist based on bio-enzyme regulation and its preparation method. Specifically, the technical solution of this invention includes the following:

[0007] A method for preparing a fertilizer synergist based on bio-enzyme regulation, the method comprising the following steps:

[0008] A mixture was obtained by reacting β-cyclodextrin, 50 wt% aqueous polyethyleneimine solution, and immobilized enzyme complex.

[0009] Rice husk ash was mixed with 1 mol / L sodium hydroxide solution to obtain rice husk ash sol;

[0010] A first solution is obtained by mixing tetrabutyl titanate and manganese sulfate, and then 95% ethanol solution is added to react and manganese-doped titanium oxide is obtained.

[0011] The mixture, rice husk ash sol, and manganese-doped titanium dioxide were reacted to obtain a modified enzyme complex;

[0012] The fertilizer synergist was prepared by mixing the modified enzyme complex, potassium humate, and magnesium sulfate.

[0013] Furthermore, the weight ratio of the β-cyclodextrin, 50wt% aqueous solution of polyethyleneimine, and immobilized enzyme complex is 2-4:10-15:100.

[0014] Furthermore, the reaction conditions for the mixture of β-cyclodextrin, 50wt% polyethyleneimine aqueous solution, and immobilized enzyme complex include a reaction temperature of 40–60°C and a reaction time of 2–4 h.

[0015] Furthermore, the method for preparing the immobilized enzyme complex includes the following steps:

[0016] Biochar powder was obtained by high-temperature treatment and grinding of water hyacinth stems;

[0017] Modified biochar was obtained by mixing biochar powder with 30 wt% hydrogen peroxide and reacting the mixture.

[0018] Modified biochar and sodium alginate were mixed and reacted, and then spray-dried to obtain biochar microspheres;

[0019] The first complex solution is obtained by mixing neutral protease and urease inhibitor;

[0020] The alkaline protease was dispersed in a phosphate buffer solution at pH 8.0 to obtain the second enzyme solution;

[0021] Acid phosphatase, phytase, and cellulase were dispersed in a phosphate buffer solution at pH 6.0 to obtain the third enzyme solution.

[0022] The first enzyme solution, the second enzyme solution, the third enzyme solution, and trehalose are mixed to obtain a mixed enzyme solution;

[0023] Zinc nitrate was dispersed in 35 wt% methanol to obtain solution A, and 2-methylimidazole was dispersed in 35 wt% methanol to obtain solution B;

[0024] The first intermediate was obtained by mixing and reacting biochar microspheres with a mixed enzyme solution.

[0025] The immobilized enzyme complex was prepared by reacting the first intermediate, solution A, and solution B together.

[0026] Furthermore, the conditions for the high-temperature treatment include a treatment temperature of 500–600°C and a treatment time of 2–4 hours.

[0027] Furthermore, the weight ratio of the biochar powder to 30 wt% hydrogen peroxide is 10-20:50.

[0028] Furthermore, the reaction conditions for the mixture of biochar powder and 30 wt% hydrogen peroxide include a reaction temperature of 40–60 °C and a reaction time of 12–14 h.

[0029] Furthermore, the weight ratio of the modified biochar to sodium alginate is 14–16:3–5.

[0030] Furthermore, the conditions for the mixed reaction of the modified biochar and sodium alginate include a reaction temperature of 40–60°C and a reaction time of 30–50 min.

[0031] Furthermore, the weight ratio of the neutral protease, urease inhibitor, alkaline protease, acid phosphatase, phytase, cellulase and trehalose is 6-7:1.5-2.0:2.5-3.0:2.5-3.0:3.6-4.0:4.1-4.5:8-10.

[0032] Furthermore, the weight ratio of zinc nitrate, 2-methylimidazole, biochar microspheres, and mixed enzyme solution is 2.7–2.9:30–50:80–100:2000.

[0033] Furthermore, the conditions for the reaction of the biochar microspheres and the mixed enzyme solution include a reaction temperature of 3-4°C and a reaction time of 4-6 hours.

[0034] Furthermore, the conditions for the mixed reaction of the first intermediate, solution A, and solution B include a reaction temperature of 23–25°C and a reaction time of 30–50 min.

[0035] Furthermore, the weight ratio of the rice husk ash to the sodium hydroxide solution is 10-15:100-200.

[0036] Furthermore, the weight ratio of tetrabutyl titanate to manganese sulfate is 3.4–3.9:0.03–0.05.

[0037] Furthermore, the reaction conditions for mixing the first solution and the 95% ethanol solution include standing at 25°C for 24 hours, followed by heating to 180°C and reacting for 12 hours.

[0038] Furthermore, the weight ratio of the mixture, rice husk ash sol, and manganese-doped titanium oxide is 5–10:50:0.5–2.5.

[0039] Furthermore, the reaction conditions for the mixture, rice husk ash sol, and manganese-doped titanium oxide include a reaction temperature of 23–25°C and a reaction time of 8–12 h.

[0040] Furthermore, the weight ratio of the modified enzyme complex, potassium humate, and magnesium sulfate is 100:3-5:1-2.

[0041] Furthermore, the mixing conditions for the modified enzyme complex, potassium humate, and magnesium sulfate include a mixing temperature of 40–50°C and a mixing time of 20–40 min.

[0042] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0043] The fertilizer synergist prepared in this invention uses biochar microspheres as a carrier to immobilize and load multiple functional enzymes and adds urease inhibitors, which can improve the stability and long-term effect of the enzymes. The synergistic effect of multiple enzymes can effectively activate insoluble phosphorus in the soil, promote organic nitrogen mineralization, inhibit urea nitrogen loss, and accelerate organic matter decomposition, thereby significantly improving nutrient utilization efficiency. The outer layer, composed of β-cyclodextrin and polyethyleneimine, is a smart responsive coating layer that can achieve intelligent slow release and targeted delivery of enzymes and nutrients according to changes in the soil microenvironment, further reducing nutrient loss. In addition, rice husk ash sol improves soil structure and provides beneficial silicon, manganese-doped titanium dioxide decomposes harmful substances in the soil under light and regulates the nitrogen cycle, while potassium humate and magnesium sulfate supplement key nutrients and stimulate crop growth. Through multi-pathway synergistic effects, fertilizer utilization is improved, soil is improved, and crop resistance is enhanced. Detailed Implementation

[0044] The technical solution of the present invention will be clearly and completely described below through embodiments. Obviously, the described embodiments are only a part of the embodiments of the present invention, and not all of them. 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.

[0045] Unless otherwise stated, all raw materials and reagents used in this invention are commercially available or can be prepared by known methods.

[0046] Preparation Example 1:

[0047] The method for preparing the immobilized enzyme complex includes the following steps:

[0048] Water hyacinth stems were placed in a tube furnace and heated to 500℃ at a rate of 10℃ / min under nitrogen protection for 2 hours. After cooling, the stems were ground through a 200-mesh sieve to obtain biochar powder. 10 parts by weight of biochar powder and 50 parts by weight of 30wt% hydrogen peroxide were mixed and stirred at 40℃ at a speed of 200r / min for 12 hours. After the reaction was completed, the mixture was centrifuged at 8000r / min for 10 minutes, the supernatant was discarded, and the precipitate was washed with deionized water until neutral. The precipitate was then vacuum dried at 80℃ for 12 hours to obtain modified biochar.

[0049] 14 parts by weight of modified biochar and 3 parts by weight of sodium alginate were dispersed in 200 parts by weight of deionized water and stirred at 40°C for 30 min to obtain a mixture. The mixture was atomized into small droplets on a high-speed rotating centrifugal atomizing plate and sprayed into a 2 wt% calcium chloride solution through a nozzle. After stirring at 100 r / min for 30 min, the mixture was filtered and freeze-dried to obtain biochar microspheres.

[0050] Six parts by weight of neutral protease were dispersed in 500 parts by weight of phosphate buffer at pH 7.0 to obtain the first enzyme solution. 1.5 parts by weight of urease inhibitor were dispersed in 500 parts by weight of anhydrous ethanol and then added to the first enzyme solution to obtain the first composite solution. 2.5 parts by weight of alkaline protease were dispersed in 1000 parts by weight of phosphate buffer at pH 8.0 to obtain the second enzyme solution. 2.5 parts by weight of acid phosphatase, 3.6 parts by weight of phytase and 4.1 parts by weight of cellulase were dispersed in 1000 parts by weight of phosphate buffer at pH 6.0 to obtain the third enzyme solution. The first, second and third enzyme solutions were mixed, and 8 parts by weight of trehalose were added and stirred for 20 min to obtain the mixed enzyme solution.

[0051] 2.7 parts by weight of zinc nitrate were dispersed in 50 parts by weight of 35 wt% methanol to obtain solution A, and 30 parts by weight of 2-methylimidazole were dispersed in 500 parts by weight of 35 wt% methanol to obtain solution B. 80 parts by weight of biochar microspheres were dispersed in 2000 parts by weight of mixed enzyme solution. After stirring and mixing at 3°C ​​for 4 h, the mixture was transferred to solution A and stirred and dispersed for 10 min. Then, solution B was added, and the mixture was stirred and reacted at 25°C for 30 min. After washing three times with methanol, the mixture was vacuum dried to obtain the immobilized enzyme complex.

[0052] Preparation Example 2:

[0053] The method for preparing the immobilized enzyme complex includes the following steps:

[0054] Water hyacinth stems were placed in a tube furnace and heated to 520℃ at a heating rate of 10℃ / min under nitrogen protection for 2.5 hours. After cooling, the stems were ground through a 200-mesh sieve to obtain biochar powder. 12 parts by weight of biochar powder and 50 parts by weight of 30wt% hydrogen peroxide were mixed and stirred at 45℃ at a speed of 200 r / min for 12.5 hours. After the reaction was completed, the mixture was centrifuged at 8000 r / min for 10 minutes, the supernatant was discarded, the precipitate was washed with deionized water until neutral, and then vacuum dried at 80℃ for 12 hours to obtain modified biochar.

[0055] 14.5 parts by weight of modified biochar and 3.5 parts by weight of sodium alginate were dispersed in 200 parts by weight of deionized water and stirred at 45°C for 35 min to obtain a mixture. The mixture was atomized into small droplets on a high-speed rotating centrifugal atomizing plate and sprayed into a 2 wt% calcium chloride solution through a nozzle. After stirring at 100 r / min for 30 min, the mixture was filtered and freeze-dried to obtain biochar microspheres.

[0056] 6.2 parts by weight of neutral protease were dispersed in 500 parts by weight of phosphate buffer at pH 7.0 to obtain the first enzyme solution; 1.6 parts by weight of urease inhibitor were dispersed in 500 parts by weight of anhydrous ethanol, and then added to the first enzyme solution to obtain the first composite solution; 2.6 parts by weight of alkaline protease were dispersed in 1000 parts by weight of phosphate buffer at pH 8.0 to obtain the second enzyme solution; 2.6 parts by weight of acid phosphatase, 3.7 parts by weight of phytase and 4.2 parts by weight of cellulase were dispersed in 1000 parts by weight of phosphate buffer at pH 6.0 to obtain the third enzyme solution; the first, second and third enzyme solutions were mixed, and 8.5 parts by weight of trehalose were added and stirred for 25 min to obtain the mixed enzyme solution;

[0057] 2.75 parts by weight of zinc nitrate were dispersed in 50 parts by weight of 35 wt% methanol to obtain solution A, and 35 parts by weight of 2-methylimidazole were dispersed in 500 parts by weight of 35 wt% methanol to obtain solution B. 85 parts by weight of biochar microspheres were dispersed in 2000 parts by weight of mixed enzyme solution. After stirring and mixing at 3°C ​​for 4.5 h, the mixture was transferred to solution A and stirred and dispersed for 15 min. Then, solution B was added, and the mixture was stirred and reacted at 25°C for 35 min. After washing three times with methanol, the mixture was vacuum dried to obtain the immobilized enzyme complex.

[0058] Preparation Example 3:

[0059] The method for preparing the immobilized enzyme complex includes the following steps:

[0060] Water hyacinth stems were placed in a tube furnace and heated to 550℃ at a heating rate of 10℃ / min under nitrogen protection for 3 hours. After cooling, the stems were ground through a 200-mesh sieve to obtain biochar powder. 15 parts by weight of biochar powder and 50 parts by weight of 30wt% hydrogen peroxide were mixed and stirred at 50℃ at a speed of 200 r / min for 13 hours. After the reaction was completed, the mixture was centrifuged at 8000 r / min for 10 minutes, the supernatant was discarded, the precipitate was washed with deionized water until neutral, and then vacuum dried at 80℃ for 12 hours to obtain modified biochar.

[0061] 15 parts by weight of modified biochar and 4 parts by weight of sodium alginate were dispersed in 200 parts by weight of deionized water and stirred at 50°C for 40 min to obtain a mixture. The mixture was atomized into small droplets on a high-speed rotating centrifugal atomizing plate and sprayed into a 2 wt% calcium chloride solution through a nozzle. After stirring at 100 r / min for 30 min, the mixture was filtered and freeze-dried to obtain biochar microspheres.

[0062] 6.5 parts by weight of neutral protease were dispersed in 500 parts by weight of phosphate buffer at pH 7.0 to obtain the first enzyme solution; 1.7 parts by weight of urease inhibitor were dispersed in 500 parts by weight of anhydrous ethanol, and then added to the first enzyme solution to obtain the first composite solution; 2.7 parts by weight of alkaline protease were dispersed in 1000 parts by weight of phosphate buffer at pH 8.0 to obtain the second enzyme solution; 2.7 parts by weight of acid phosphatase, 3.8 parts by weight of phytase and 4.3 parts by weight of cellulase were dispersed in 1000 parts by weight of phosphate buffer at pH 6.0 to obtain the third enzyme solution; the first, second and third enzyme solutions were mixed, and 9 parts by weight of trehalose were added and stirred for 30 min to obtain the mixed enzyme solution;

[0063] 2.8 parts by weight of zinc nitrate were dispersed in 50 parts by weight of 35 wt% methanol to obtain solution A, and 40 parts by weight of 2-methylimidazole were dispersed in 500 parts by weight of 35 wt% methanol to obtain solution B. 90 parts by weight of biochar microspheres were dispersed in 2000 parts by weight of mixed enzyme solution. After stirring and mixing at 4℃ for 5 h, the mixture was transferred to solution A and stirred and dispersed for 20 min. Then, solution B was added, and the mixture was stirred and reacted at 25℃ for 40 min. After washing three times with methanol, the mixture was vacuum dried to obtain the immobilized enzyme complex.

[0064] Preparation Example 4:

[0065] The method for preparing the immobilized enzyme complex includes the following steps:

[0066] Water hyacinth stems were placed in a tube furnace and heated to 570℃ at a heating rate of 10℃ / min under nitrogen protection for 3.5 hours. After cooling, the stems were ground through a 200-mesh sieve to obtain biochar powder. 17 parts by weight of biochar powder and 50 parts by weight of 30wt% hydrogen peroxide were mixed and stirred at 55℃ at a speed of 200 r / min for 13.5 hours. After the reaction was completed, the mixture was centrifuged at 8000 r / min for 10 minutes, the supernatant was discarded, the precipitate was washed with deionized water until neutral, and then vacuum dried at 80℃ for 12 hours to obtain modified biochar.

[0067] 15.5 parts by weight of modified biochar and 4.5 parts by weight of sodium alginate were dispersed in 200 parts by weight of deionized water and stirred at 55°C for 45 min to obtain a mixture. The mixture was atomized into small droplets on a high-speed rotating centrifugal atomizing plate and sprayed into a 2 wt% calcium chloride solution through a nozzle. After stirring at 100 r / min for 30 min, the mixture was filtered and freeze-dried to obtain biochar microspheres.

[0068] 6.8 parts by weight of neutral protease were dispersed in 500 parts by weight of phosphate buffer at pH 7.0 to obtain the first enzyme solution; 1.8 parts by weight of urease inhibitor were dispersed in 500 parts by weight of anhydrous ethanol, and then added to the first enzyme solution to obtain the first composite solution; 2.8 parts by weight of alkaline protease were dispersed in 1000 parts by weight of phosphate buffer at pH 8.0 to obtain the second enzyme solution; 2.8 parts by weight of acid phosphatase, 3.9 parts by weight of phytase and 4.4 parts by weight of cellulase were dispersed in 1000 parts by weight of phosphate buffer at pH 6.0 to obtain the third enzyme solution; the first, second and third enzyme solutions were mixed, and 9.5 parts by weight of trehalose were added and stirred for 35 min to obtain the mixed enzyme solution;

[0069] 2.85 parts by weight of zinc nitrate were dispersed in 50 parts by weight of 35 wt% methanol to obtain solution A, and 45 parts by weight of 2-methylimidazole were dispersed in 500 parts by weight of 35 wt% methanol to obtain solution B. 95 parts by weight of biochar microspheres were dispersed in 2000 parts by weight of mixed enzyme solution. After stirring and mixing at 3°C ​​for 5.5 h, the mixture was transferred to solution A and stirred and dispersed for 25 min. Then, solution B was added, and the mixture was stirred and reacted at 25°C for 45 min. After washing three times with methanol, the mixture was vacuum dried to obtain the immobilized enzyme complex.

[0070] Preparation Example 5:

[0071] The method for preparing the immobilized enzyme complex includes the following steps:

[0072] Water hyacinth stems were placed in a tube furnace and heated to 600℃ at a rate of 10℃ / min under nitrogen protection for 4 hours. After cooling, the stems were ground through a 200-mesh sieve to obtain biochar powder. 20 parts by weight of biochar powder and 50 parts by weight of 30wt% hydrogen peroxide were mixed and stirred at 60℃ at a speed of 200 r / min for 14 hours. After the reaction was completed, the mixture was centrifuged at 8000 r / min for 10 minutes, the supernatant was discarded, and the precipitate was washed with deionized water until neutral. The precipitate was then vacuum dried at 80℃ for 12 hours to obtain modified biochar.

[0073] 16 parts by weight of modified biochar and 5 parts by weight of sodium alginate were dispersed in 200 parts by weight of deionized water and stirred at 60°C for 50 min to obtain a mixture. The mixture was atomized into small droplets on a high-speed rotating centrifugal atomizing plate and sprayed into a 2 wt% calcium chloride solution through a nozzle. After stirring at 100 r / min for 30 min, the mixture was filtered and freeze-dried to obtain biochar microspheres.

[0074] 7 parts by weight of neutral protease were dispersed in 500 parts by weight of phosphate buffer at pH 7.0 to obtain the first enzyme solution; 2.0 parts by weight of urease inhibitor were dispersed in 500 parts by weight of anhydrous ethanol, and then added to the first enzyme solution to obtain the first composite solution; 3.0 parts by weight of alkaline protease were dispersed in 1000 parts by weight of phosphate buffer at pH 8.0 to obtain the second enzyme solution; 3.0 parts by weight of acid phosphatase, 4.0 parts by weight of phytase and 4.5 parts by weight of cellulase were dispersed in 1000 parts by weight of phosphate buffer at pH 6.0 to obtain the third enzyme solution; the first, second and third enzyme solutions were mixed, and 10 parts by weight of trehalose were added and stirred for 40 min to obtain the mixed enzyme solution;

[0075] 2.9 parts by weight of zinc nitrate were dispersed in 50 parts by weight of 35 wt% methanol to obtain solution A, and 50 parts by weight of 2-methylimidazole were dispersed in 500 parts by weight of 35 wt% methanol to obtain solution B. 100 parts by weight of biochar microspheres were dispersed in 2000 parts by weight of mixed enzyme solution. After stirring and mixing at 4°C for 6 h, the mixture was transferred to solution A and stirred and dispersed for 30 min. Then, solution B was added, and the mixture was stirred and reacted at 25°C for 50 min. After washing three times with methanol, the mixture was vacuum dried to obtain the immobilized enzyme complex.

[0076] Preparation Example 6:

[0077] The method for preparing the immobilized enzyme complex includes the following steps:

[0078] Water hyacinth stems were placed in a tube furnace and heated to 600℃ at a rate of 10℃ / min under nitrogen protection for 4 hours. After cooling, the stems were ground through a 200-mesh sieve to obtain biochar powder. 16 parts by weight of biochar powder and 5 parts by weight of sodium alginate were dispersed in 200 parts by weight of deionized water and stirred at 60℃ for 50 minutes to obtain a mixture. The mixture was atomized into small droplets on a high-speed rotating centrifugal atomizing disc and sprayed into a 2wt% calcium chloride solution through a nozzle. The solution was stirred at 100 r / min for 30 minutes and then filtered. After freeze-drying, biochar microspheres were obtained.

[0079] 7 parts by weight of neutral protease were dispersed in 500 parts by weight of phosphate buffer at pH 7.0 to obtain the first enzyme solution; 2.0 parts by weight of urease inhibitor were dispersed in 500 parts by weight of anhydrous ethanol, and then added to the first enzyme solution to obtain the first composite solution; 3.0 parts by weight of alkaline protease were dispersed in 1000 parts by weight of phosphate buffer at pH 8.0 to obtain the second enzyme solution; 3.0 parts by weight of acid phosphatase, 4.0 parts by weight of phytase and 4.5 parts by weight of cellulase were dispersed in 1000 parts by weight of phosphate buffer at pH 6.0 to obtain the third enzyme solution; the first, second and third enzyme solutions were mixed, and 10 parts by weight of trehalose were added and stirred for 40 min to obtain the mixed enzyme solution;

[0080] 2.9 parts by weight of zinc nitrate were dispersed in 50 parts by weight of 35 wt% methanol to obtain solution A, and 50 parts by weight of 2-methylimidazole were dispersed in 500 parts by weight of 35 wt% methanol to obtain solution B. 100 parts by weight of biochar microspheres were dispersed in 2000 parts by weight of mixed enzyme solution. After stirring and mixing at 4°C for 6 h, the mixture was transferred to solution A and stirred and dispersed for 30 min. Then, solution B was added, and the mixture was stirred and reacted at 25°C for 50 min. After washing three times with methanol, the mixture was vacuum dried to obtain the immobilized enzyme complex.

[0081] Preparation Example 7:

[0082] The method for preparing the immobilized enzyme complex includes the following steps:

[0083] Water hyacinth stems were placed in a tube furnace and heated to 600℃ at a rate of 10℃ / min under nitrogen protection for 4 hours. After cooling, the stems were ground through a 200-mesh sieve to obtain biochar powder. 20 parts by weight of biochar powder and 50 parts by weight of 30wt% hydrogen peroxide were mixed and stirred at 60℃ at a speed of 200 r / min for 14 hours. After the reaction was completed, the mixture was centrifuged at 8000 r / min for 10 minutes, the supernatant was discarded, and the precipitate was washed with deionized water until neutral. The precipitate was then vacuum dried at 80℃ for 12 hours to obtain modified biochar.

[0084] 7 parts by weight of neutral protease were dispersed in 500 parts by weight of phosphate buffer at pH 7.0 to obtain the first enzyme solution; 2.0 parts by weight of urease inhibitor were dispersed in 500 parts by weight of anhydrous ethanol, and then added to the first enzyme solution to obtain the first composite solution; 3.0 parts by weight of alkaline protease were dispersed in 1000 parts by weight of phosphate buffer at pH 8.0 to obtain the second enzyme solution; 3.0 parts by weight of acid phosphatase, 4.0 parts by weight of phytase and 4.5 parts by weight of cellulase were dispersed in 1000 parts by weight of phosphate buffer at pH 6.0 to obtain the third enzyme solution; the first, second and third enzyme solutions were mixed, and 10 parts by weight of trehalose were added and stirred for 40 min to obtain the mixed enzyme solution;

[0085] 2.9 parts by weight of zinc nitrate were dispersed in 50 parts by weight of 35 wt% methanol to obtain solution A, and 50 parts by weight of 2-methylimidazole were dispersed in 500 parts by weight of 35 wt% methanol to obtain solution B. 100 parts by weight of modified biochar were dispersed in 2000 parts by weight of mixed enzyme solution. After stirring and mixing at 4°C for 6 h, the mixture was transferred to solution A and stirred and dispersed for 30 min. Then, solution B was added, and the mixture was stirred and reacted at 25°C for 50 min. After washing three times with methanol, the mixture was vacuum dried to obtain the immobilized enzyme complex.

[0086] Example 1:

[0087] A method for preparing a fertilizer synergist based on bio-enzyme regulation includes the following steps:

[0088] Two parts by weight of β-cyclodextrin and 10 parts by weight of 50 wt% polyethyleneimine aqueous solution were dispersed in 100 parts by weight of deionized water and stirred at 40°C for 20 min. Then, 100 parts by weight of the immobilized enzyme complex prepared in Example 1 were added and the mixture was shaken at 200 r / min for 2 h. After centrifugation to remove the residual liquid, the solid was collected to obtain the mixture.

[0089] 10 parts by weight of rice husk ash were dispersed in 100 parts by weight of 1 mol / L sodium hydroxide solution. After stirring and reacting at 150℃ for 5 hours, the supernatant was collected by centrifugation. The pH of the supernatant was adjusted to 10 and then concentrated to a silica mass fraction of 20% to obtain rice husk ash sol.

[0090] 3.4 parts by weight of tetrabutyl titanate and 0.03 parts by weight of manganese sulfate were dispersed in 30 parts by weight of anhydrous ethanol and stirred for 20 min to obtain the first solution. 20 parts by weight of 95% ethanol solution were added to the first solution and stirred for 10 min. The mixture was then allowed to stand at 25°C for 24 h and then transferred to a high-temperature and high-pressure reactor at 180°C for 12 h. After the reaction was completed, the mixture was washed and dried to obtain manganese-doped titanium oxide.

[0091] Five parts by weight of the mixture, 50 parts by weight of rice husk ash sol and 0.5 parts by weight of manganese-doped titanium oxide were dispersed in 200 parts by weight of Tris-HCl buffer at pH 8.5. The mixture was stirred at 23°C for 8 hours. After the reaction was completed, the modified enzyme complex was obtained by centrifugation, washing and drying.

[0092] The fertilizer synergist was prepared by mixing 100 parts by weight of modified enzyme complex, 3 parts by weight of potassium humate and 1 part by weight of magnesium sulfate at 40°C for 20 minutes and then drying.

[0093] Example 2:

[0094] A method for preparing a fertilizer synergist based on bio-enzyme regulation includes the following steps:

[0095] 2.5 parts by weight of β-cyclodextrin and 11 parts by weight of 50 wt% polyethyleneimine aqueous solution were dispersed in 100 parts by weight of deionized water and stirred at 45°C for 25 min. Then, 100 parts by weight of the immobilized enzyme complex prepared in Preparation Example 2 were added and the mixture was shaken at 200 r / min for 2.5 h. The residue was removed by centrifugation and the solid was collected to obtain the mixture.

[0096] 11 parts by weight of rice husk ash were dispersed in 120 parts by weight of 1 mol / L sodium hydroxide solution. After stirring and reacting at 155℃ for 5.2 h, the supernatant was collected by centrifugation. The pH of the supernatant was adjusted to 10 and then concentrated to a silica mass fraction of 20% to obtain rice husk ash sol.

[0097] 3.5 parts by weight of tetrabutyl titanate and 0.035 parts by weight of manganese sulfate were dispersed in 30 parts by weight of anhydrous ethanol and stirred for 22 min to obtain the first solution. 20 parts by weight of 95% ethanol solution were added to the first solution and stirred for 15 min. The mixture was then allowed to stand at 25°C for 24 h and then transferred to a high-temperature and high-pressure reactor at 180°C for 12 h. After the reaction was completed, the mixture was washed and dried to obtain manganese-doped titanium oxide.

[0098] Six parts by weight of the mixture, 50 parts by weight of rice husk ash sol and 1.0 part by weight of manganese-doped titanium oxide were dispersed in 200 parts by weight of Tris-HCl buffer at pH 8.5 and stirred at 24°C for 9 h. After the reaction was completed, the modified enzyme complex was obtained by centrifugation, washing and drying.

[0099] The fertilizer synergist was prepared by mixing 100 parts by weight of modified enzyme complex, 3.5 parts by weight of potassium humate and 1.2 parts by weight of magnesium sulfate at 42°C for 25 minutes and then drying.

[0100] Example 3:

[0101] A method for preparing a fertilizer synergist based on bio-enzyme regulation includes the following steps:

[0102] Three parts by weight of β-cyclodextrin and 12 parts by weight of 50 wt% polyethyleneimine aqueous solution were dispersed in 100 parts by weight of deionized water and stirred at 50°C for 30 min. Then, 100 parts by weight of the immobilized enzyme complex prepared in Preparation Example 3 were added. The mixture was shaken at 200 r / min for 3 h. After centrifugation to remove the residual liquid, the solid was collected to obtain the mixture.

[0103] 12 parts by weight of rice husk ash were dispersed in 150 parts by weight of 1 mol / L sodium hydroxide solution. After stirring and reacting at 160℃ for 5.5 h, the supernatant was collected by centrifugation. The pH of the supernatant was adjusted to 10 and then concentrated to a silica mass fraction of 20% to obtain rice husk ash sol.

[0104] 3.6 parts by weight of tetrabutyl titanate and 0.04 parts by weight of manganese sulfate were dispersed in 30 parts by weight of anhydrous ethanol and stirred for 25 min to obtain the first solution. 20 parts by weight of 95% ethanol solution were added to the first solution and stirred for 20 min. The mixture was then allowed to stand at 25°C for 24 h and then transferred to a high-temperature and high-pressure reactor at 180°C for 12 h. After the reaction was completed, the mixture was washed and dried to obtain manganese-doped titanium oxide.

[0105] 7 parts by weight of the mixture, 50 parts by weight of rice husk ash sol and 1.5 parts by weight of manganese-doped titanium oxide were dispersed in 200 parts by weight of Tris-HCl buffer at pH 8.5 and stirred at 25°C for 10 h. After the reaction was completed, the modified enzyme complex was obtained by centrifugation, washing and drying.

[0106] The fertilizer synergist was prepared by mixing 100 parts by weight of modified enzyme complex, 4 parts by weight of potassium humate and 1.5 parts by weight of magnesium sulfate at 45°C for 30 minutes and then drying.

[0107] Example 4:

[0108] A method for preparing a fertilizer synergist based on bio-enzyme regulation includes the following steps:

[0109] 3.5 parts by weight of β-cyclodextrin and 13 parts by weight of 50 wt% polyethyleneimine aqueous solution were dispersed in 100 parts by weight of deionized water and stirred at 55°C for 35 min. Then, 100 parts by weight of the immobilized enzyme complex prepared in Preparation Example 4 were added and the mixture was shaken at 200 r / min for 3.5 h. The residue was removed by centrifugation and the solid was collected to obtain the mixture.

[0110] 13 parts by weight of rice husk ash were dispersed in 170 parts by weight of 1 mol / L sodium hydroxide solution. After stirring and reacting at 170℃ for 5.7 h, the supernatant was collected by centrifugation. The pH of the supernatant was adjusted to 10 and then concentrated to a silica mass fraction of 20% to obtain rice husk ash sol.

[0111] 3.7 parts by weight of tetrabutyl titanate and 0.045 parts by weight of manganese sulfate were dispersed in 30 parts by weight of anhydrous ethanol and stirred for 27 min to obtain the first solution. 20 parts by weight of 95% ethanol solution were added to the first solution and stirred for 25 min. The mixture was then allowed to stand at 25°C for 24 h and then transferred to a high-temperature and high-pressure reactor at 180°C for 12 h. After the reaction was completed, the mixture was washed and dried to obtain manganese-doped titanium oxide.

[0112] Eight parts by weight of the mixture, 50 parts by weight of rice husk ash sol and 2.0 parts by weight of manganese-doped titanium oxide were dispersed in 200 parts by weight of Tris-HCl buffer at pH 8.5 and stirred at 23°C for 11 h. After the reaction was completed, the modified enzyme complex was obtained by centrifugation, washing and drying.

[0113] The fertilizer synergist was prepared by mixing 100 parts by weight of modified enzyme complex, 4.5 parts by weight of potassium humate and 1.8 parts by weight of magnesium sulfate at 48°C for 35 minutes and then drying.

[0114] Example 5:

[0115] A method for preparing a fertilizer synergist based on bio-enzyme regulation includes the following steps:

[0116] Four parts by weight of β-cyclodextrin and 15 parts by weight of 50 wt% polyethyleneimine aqueous solution were dispersed in 100 parts by weight of deionized water and stirred at 60°C for 40 min. Then, 100 parts by weight of the immobilized enzyme complex prepared in Preparation Example 5 were added. The mixture was shaken at 200 r / min for 4 h. The residue was removed by centrifugation and the solid was collected to obtain the mixture.

[0117] 15 parts by weight of rice husk ash were dispersed in 200 parts by weight of 1 mol / L sodium hydroxide solution. After stirring and reacting at 180℃ for 6 hours, the supernatant was collected by centrifugation. The pH of the supernatant was adjusted to 10 and then concentrated to a silica mass fraction of 20% to obtain rice husk ash sol.

[0118] 3.9 parts by weight of tetrabutyl titanate and 0.05 parts by weight of manganese sulfate were dispersed in 30 parts by weight of anhydrous ethanol and stirred for 30 min to obtain the first solution. 20 parts by weight of 95% ethanol solution were added to the first solution and stirred for 30 min. The mixture was then allowed to stand at 25°C for 24 h and then transferred to a high-temperature and high-pressure reactor at 180°C for 12 h. After the reaction was completed, the mixture was washed and dried to obtain manganese-doped titanium oxide.

[0119] 10 parts by weight of the mixture, 50 parts by weight of rice husk ash sol and 2.5 parts by weight of manganese-doped titanium oxide were dispersed in 200 parts by weight of Tris-HCl buffer at pH 8.5 and stirred at 25°C for 12 h. After the reaction was completed, the modified enzyme complex was obtained by centrifugation, washing and drying.

[0120] The fertilizer synergist was prepared by mixing 100 parts by weight of modified enzyme complex, 5 parts by weight of potassium humate and 2 parts by weight of magnesium sulfate at 50°C for 40 minutes and then drying.

[0121] Comparative Example 1:

[0122] A method for preparing a fertilizer synergist based on bio-enzyme regulation includes the following steps:

[0123] The immobilized enzyme complex prepared in Example 5 was replaced with the immobilized enzyme complex prepared in Example 6, and all other operations were the same as in Example 5.

[0124] Comparative Example 2:

[0125] A method for preparing a fertilizer synergist based on bio-enzyme regulation includes the following steps:

[0126] The immobilized enzyme complex prepared in Example 5 was replaced with the immobilized enzyme complex prepared in Example 7, and all other operations were the same as in Example 5.

[0127] Comparative Example 3:

[0128] A method for preparing a fertilizer synergist based on bio-enzyme regulation includes the following steps:

[0129] 15 parts by weight of rice husk ash were dispersed in 200 parts by weight of 1 mol / L sodium hydroxide solution. After stirring and reacting at 180℃ for 6 hours, the supernatant was collected by centrifugation. The pH of the supernatant was adjusted to 10 and then concentrated to a silica mass fraction of 20% to obtain rice husk ash sol.

[0130] 3.9 parts by weight of tetrabutyl titanate and 0.05 parts by weight of manganese sulfate were dispersed in 30 parts by weight of anhydrous ethanol and stirred for 30 min to obtain the first solution. 20 parts by weight of 95% ethanol solution were added to the first solution and stirred for 30 min. The mixture was then allowed to stand at 25°C for 24 h and then transferred to a high-temperature and high-pressure reactor at 180°C for 12 h. After the reaction was completed, the mixture was washed and dried to obtain manganese-doped titanium oxide.

[0131] 10 parts by weight of the immobilized enzyme complex prepared in Example 5, 50 parts by weight of rice husk ash sol and 2.5 parts by weight of manganese-doped titanium oxide were dispersed in 200 parts by weight of Tris-HCl buffer at pH 8.5. The mixture was stirred at 25°C for 12 h. After the reaction was completed, the modified enzyme complex was obtained by centrifugation, washing and drying.

[0132] The fertilizer synergist was prepared by mixing 100 parts by weight of modified enzyme complex, 5 parts by weight of potassium humate and 2 parts by weight of magnesium sulfate at 50°C for 40 minutes and then drying.

[0133] Comparative Example 4:

[0134] A method for preparing a fertilizer synergist based on bio-enzyme regulation includes the following steps:

[0135] Four parts by weight of β-cyclodextrin and 15 parts by weight of 50 wt% polyethyleneimine aqueous solution were dispersed in 100 parts by weight of deionized water and stirred at 60°C for 40 min. Then, 100 parts by weight of the immobilized enzyme complex prepared in Preparation Example 5 were added. The mixture was shaken at 200 r / min for 4 h. The residue was removed by centrifugation and the solid was collected to obtain the mixture.

[0136] 3.9 parts by weight of tetrabutyl titanate and 0.05 parts by weight of manganese sulfate were dispersed in 30 parts by weight of anhydrous ethanol and stirred for 30 min to obtain the first solution. 20 parts by weight of 95% ethanol solution were added to the first solution and stirred for 30 min. The mixture was then allowed to stand at 25°C for 24 h and then transferred to a high-temperature and high-pressure reactor at 180°C for 12 h. After the reaction was completed, the mixture was washed and dried to obtain manganese-doped titanium oxide.

[0137] 10 parts by weight of the mixture and 2.5 parts by weight of manganese-doped titanium oxide were dispersed in 200 parts by weight of Tris-HCl buffer at pH 8.5 and stirred at 25°C for 12 h. After the reaction was completed, the modified enzyme complex was obtained by centrifugation, washing and drying.

[0138] The fertilizer synergist was prepared by mixing 100 parts by weight of modified enzyme complex, 5 parts by weight of potassium humate and 2 parts by weight of magnesium sulfate at 50°C for 40 minutes and then drying.

[0139] Comparative Example 5:

[0140] A method for preparing a fertilizer synergist based on bio-enzyme regulation includes the following steps:

[0141] Four parts by weight of β-cyclodextrin and 15 parts by weight of 50 wt% polyethyleneimine aqueous solution were dispersed in 100 parts by weight of deionized water and stirred at 60°C for 40 min. Then, 100 parts by weight of the immobilized enzyme complex prepared in Preparation Example 5 were added. The mixture was shaken at 200 r / min for 4 h. The residue was removed by centrifugation and the solid was collected to obtain the mixture.

[0142] 15 parts by weight of rice husk ash were dispersed in 200 parts by weight of 1 mol / L sodium hydroxide solution. After stirring and reacting at 180℃ for 6 hours, the supernatant was collected by centrifugation. The pH of the supernatant was adjusted to 10 and then concentrated to a silica mass fraction of 20% to obtain rice husk ash sol.

[0143] 10 parts by weight of the mixture and 50 parts by weight of rice husk ash sol were dispersed in 200 parts by weight of Tris-HCl buffer solution at pH 8.5 and stirred at 25°C for 12 h. After the reaction was completed, the modified enzyme complex was obtained by centrifugation, washing and drying.

[0144] The fertilizer synergist was prepared by mixing 100 parts by weight of modified enzyme complex, 5 parts by weight of potassium humate and 2 parts by weight of magnesium sulfate at 50°C for 40 minutes and then drying.

[0145] Comparative Example 6:

[0146] A method for preparing a fertilizer synergist based on bio-enzyme regulation includes the following steps:

[0147] Four parts by weight of β-cyclodextrin and 15 parts by weight of 50 wt% polyethyleneimine aqueous solution were dispersed in 100 parts by weight of deionized water and stirred at 60°C for 40 min. Then, 100 parts by weight of the immobilized enzyme complex prepared in Preparation Example 5 were added. The mixture was shaken at 200 r / min for 4 h. The residue was removed by centrifugation and the solid was collected to obtain the mixture.

[0148] 15 parts by weight of rice husk ash were dispersed in 200 parts by weight of 1 mol / L sodium hydroxide solution. After stirring and reacting at 180℃ for 6 hours, the supernatant was collected by centrifugation. The pH of the supernatant was adjusted to 10 and then concentrated to a silica mass fraction of 20% to obtain rice husk ash sol.

[0149] 10 parts by weight of the mixture, 50 parts by weight of rice husk ash sol and 2.5 parts by weight of titanium dioxide were dispersed in 200 parts by weight of Tris-HCl buffer at pH 8.5 and stirred at 25°C for 12 h. After the reaction was completed, the modified enzyme complex was obtained by centrifugation, washing and drying.

[0150] The fertilizer synergist was prepared by mixing 100 parts by weight of modified enzyme complex, 5 parts by weight of potassium humate and 2 parts by weight of magnesium sulfate at 50°C for 40 minutes and then drying.

[0151] Effect evaluation:

[0152] An agricultural science and technology demonstration park in Henan Province was selected as the experimental area. The soil chemical properties were: pH 6.4, organic matter 15.7 g / kg, available phosphorus 27.4 mg / kg, available potassium 134.5 mg / kg, available nitrogen 88.2 mg / kg, and total nitrogen 0.79 g / kg. Corn was selected as the experimental crop, and Stanley brand compound fertilizer 17-17-17 was used. A zoned experiment was conducted, with each zone having an area of ​​48 m². 2Eleven treatment groups were established, consisting of a control group receiving only compound fertilizer without fertilizer synergist, and eleven treatment groups receiving fertilizer synergists prepared in Examples 1-5 and Comparative Examples 1-6. Except for the control group, all other areas received both compound fertilizer and fertilizer synergist simultaneously, with three parallel trials per group. The experimental areas were rectangular with protective rows around the perimeter. Each area received 4.5 kg of fertilizer, with 60% applied as basal fertilizer and 40% as topdressing at the large trumpet stage. Each area received 0.6 kg of fertilizer synergist, mixed with the basal fertilizer and applied to the soil before corn sowing. After the corn matured, relevant indicators were tested, and the results are shown in Table 1.

[0153] Table 1. Corn Index Test

[0154] Plant height (cm) Ear length (cm) Ear diameter (cm) Yield per mu (kg) Example 1 214 20.7 5.2 748.3 Example 2 215 21.1 5.4 756.4 Example 3 216 20.9 5.6 768.9 Example 4 218 21.6 5.7 774.5 Example 5 220 21.9 5.9 782.6 Comparative Example 1 204 17.5 4.4 689.1 Comparative Example 2 203 17.1 4.3 671.4 Comparative Example 3 206 18.4 4.5 705.6 Comparative Example 4 207 18.7 4.5 716.2 Comparative Example 5 209 19.8 4.6 728.9 Comparative Example 6 213 20.5 4.9 739.8 control group 201 15.2 3.9 639.7

[0155] The data in Table 1 show that the fertilizer synergists prepared in Examples 1-5 of this invention can effectively increase the yield of corn. Although the fertilizer synergists prepared in Comparative Examples 1-6 can also increase the yield of corn to a certain extent, their performance is lower than that of Examples 1-5.

[0156] The reason for the performance decline in Comparative Example 1 may be that hydrogen peroxide modification can increase the oxygen-containing functional groups on the surface of biochar, thereby enhancing its hydrophilicity and enzyme adsorption capacity. Unmodified biochar has a hydrophobic surface and insufficient functional groups, resulting in low loading capacity and weak binding of subsequent immobilized enzymes. The enzymes are prone to loss or inactivation, and the decline in enzyme activity may affect the catalytic nutrient release effect of fertilizer synergists.

[0157] The reason for the performance decline in Comparative Example 2 may be that sodium alginate encapsulates modified biochar to form microspheres, providing a physical protective barrier for the enzyme and preventing environmental factors from damaging the enzyme structure. Without the microsphere carrier, the enzyme is directly exposed to the soil and is easily degraded or inactivated by microorganisms, resulting in the failure of catalytic function; nutrient release is out of control, and crops cannot continuously obtain nutrients.

[0158] The performance degradation of Comparative Example 3 may be due to the lack of β-cyclodextrin and polyethyleneimine, which reduced enzyme immobilization efficiency, loosened enzyme-carrier binding, and easy detachment and loss in the soil; decreased enzyme stability and shortened catalytic activity maintenance time.

[0159] The reason for the performance decline in Comparative Example 4 may be that the rice husk ash sol contains nano-silica, which forms a porous network structure, encapsulates the immobilized enzyme complex, and provides slow-release protection. Without sol coating, the enzyme complex directly contacts the soil and is easily inhibited by heavy metals or organic matter, resulting in a decrease in catalytic efficiency; the nutrient release rate is too fast and cannot match the needs of the crop growth cycle.

[0160] The reason for the performance decline in Comparative Example 5 may be that manganese-doped titanium dioxide has photocatalytic activity, which can promote the oxidative decomposition of organic matter in the soil, release trace elements, reduce harmful substances, and improve the availability of nutrients such as phosphorus and potassium. Without the use of titanium dioxide, this catalytic effect is lacking, resulting in reduced nutrient conversion efficiency and thus reduced yield and ear diameter.

[0161] The performance degradation of Comparative Example 6 may be due to the fact that manganese doping expands the photoresponse range of titanium dioxide, enhances its photocatalytic activity, and promotes the decomposition of organic matter in the soil into absorbable nutrients; ordinary titanium dioxide has low catalytic efficiency and cannot effectively activate inert nutrients; and the soil nutrient conversion rate is insufficient, limiting the crops' access to nutrients.

[0162] Furthermore, the fertilizer synergist prepared by this invention exhibits excellent performance, which may be related to the multi-enzyme synergistic scheme employed in this invention. This may be due to the activation of insoluble phosphorus in the soil by acid phosphatase and phytase; the direct or indirect inhibition of urease activity by neutral protease and urease inhibitor by synergistic effect, reducing nitrogen loss and improving nitrogen utilization; the degradation of pathogen cell wall proteins by alkaline protease, which may stimulate crop systemic resistance and reduce disease occurrence; and the decomposition of cellulose into glucose by cellulase, which may shorten the composting cycle and accelerate the humification of organic matter. Through the synergistic effect of these multiple enzymes, the goal of increasing crop yield is achieved.

[0163] The embodiments described above provide a detailed explanation of the technical solutions and beneficial effects of the present invention. It should be understood that the above descriptions are merely specific embodiments of the present invention and are not intended to limit the present invention. Various changes and modifications can be made to the present invention without departing from its spirit and scope, and all such changes and modifications fall within the scope of the present invention as claimed.

Claims

1. A method for preparing a fertilizer synergist based on bio-enzyme regulation, characterized in that, The preparation method includes the following steps: A mixture was obtained by reacting β-cyclodextrin, 50 wt% aqueous solution of polyethyleneimine, and immobilized enzyme complex. Rice husk ash was mixed with 1 mol / L sodium hydroxide solution to obtain rice husk ash sol; A first solution is obtained by mixing tetrabutyl titanate and manganese sulfate, and then 95% ethanol solution is added to react and manganese-doped titanium oxide is obtained. The mixture, rice husk ash sol, and manganese-doped titanium dioxide were reacted to obtain a modified enzyme complex; The fertilizer synergist was prepared by mixing the modified enzyme complex, potassium humate, and magnesium sulfate.

2. The preparation method of a fertilizer synergist based on bio-enzyme regulation as described in claim 1, characterized in that, The weight ratio of the β-cyclodextrin, 50wt% aqueous polyethyleneimine solution, and immobilized enzyme complex is 2–4:10–15:

100.

3. The preparation method of a fertilizer synergist based on bio-enzyme regulation as described in claim 1, characterized in that, The method for preparing the immobilized enzyme complex includes the following steps: Biochar powder was obtained by high-temperature treatment and grinding of water hyacinth stems; Modified biochar was obtained by mixing biochar powder with 30 wt% hydrogen peroxide. Modified biochar and sodium alginate were mixed and reacted, and then spray-dried to obtain biochar microspheres; The first complex solution is obtained by mixing neutral protease and urease inhibitor; The alkaline protease was dispersed in a phosphate buffer solution at pH 8.0 to obtain the second enzyme solution; Acid phosphatase, phytase, and cellulase were dispersed in a phosphate buffer solution at pH 6.0 to obtain the third enzyme solution. The first enzyme solution, the second enzyme solution, the third enzyme solution, and trehalose are mixed to obtain a mixed enzyme solution; Zinc nitrate was dispersed in 35 wt% methanol to obtain solution A, and 2-methylimidazolium was dispersed in 35 wt% methanol to obtain solution B; The first intermediate was obtained by mixing and reacting biochar microspheres with a mixed enzyme solution. The immobilized enzyme complex was prepared by reacting the first intermediate, solution A, and solution B together.

4. The preparation method of a fertilizer synergist based on bio-enzyme regulation as described in claim 3, characterized in that, The weight ratio of the biochar powder to 30wt% hydrogen peroxide is 10-20:

50.

5. The preparation method of a fertilizer synergist based on bio-enzyme regulation as described in claim 3, characterized in that, The weight ratio of the modified biochar to sodium alginate is 14-16:3-5.

6. The preparation method of a fertilizer synergist based on bio-enzyme regulation as described in claim 3, characterized in that, The weight ratio of the neutral protease, urease inhibitor, alkaline protease, acid phosphatase, phytase, cellulase and trehalose is 6-7:1.5-2.0:2.5-3.0:2.5-3.0:3.6-4.0:4.1-4.5:8-10.

7. The preparation method of a fertilizer synergist based on bio-enzyme regulation as described in claim 1, characterized in that, The weight ratio of tetrabutyl titanate to manganese sulfate is 3.4–3.9:0.03–0.

05.

8. The preparation method of a fertilizer synergist based on bio-enzyme regulation as described in claim 1, characterized in that, The weight ratio of the mixture, rice husk ash sol, and manganese-doped titanium oxide is 5–10:50:0.5–2.

5.

9. The preparation method of a fertilizer synergist based on bio-enzyme regulation as described in claim 1, characterized in that, The weight ratio of the modified enzyme complex, potassium humate, and magnesium sulfate is 100:3-5:1-2.

10. A fertilizer synergist based on bio-enzyme regulation, characterized in that, It is prepared by the preparation method of the fertilizer synergist based on bio-enzyme regulation as described in any one of claims 1 to 9.