Compound fertilizer containing high-activity ARC microbial inoculant and preparation method and application thereof

Abstract

The present application relates to a kind of high-activity ARC microbial inoculant-containing compound fertilizer and its preparation method and application, the compound fertilizer includes: as the core layer fertilizer particle, the ARC microbial inoculant encapsulated by quaternary crosslinking composite wall material is coated in the outside of fertilizer particle;The quaternary crosslinking composite wall material is obtained after mutual crosslinking of polyvinyl alcohol, soybean peptide, dopamine, ammonium molybdate.This application compound fertilizer overall design not only can provide necessary inorganic available nutrient for plant, again, through quaternary crosslinking composite wall material encapsulation ARC microbial inoculant, the survival rate of microorganism in compound fertilizer is improved, in soil, to realize intelligent control release to respond to soil moisture condition, the compound fertilizer preparation process provided in the present application is simple, easy to industrialization, with good application prospect.

Description

Technical Field

[0001] This application belongs to the field of microbial fertilizer technology, and relates to a compound fertilizer containing highly active ARC microbial agents, its preparation method and application. Background Technology

[0002] For leguminous symbiotic nitrogen-fixing crops such as soybeans and peanuts, excessive application of chemical fertilizers, especially nitrogen fertilizers, can inhibit nitrogen fixation in root nodules, reduce the activity of rhizobia, decrease nodule formation, and weaken their symbiotic nitrogen-fixing capacity. Simply applying chemical fertilizers cannot solve the problem of continuous cropping obstacles caused by repeated planting over many years. The accumulation of pathogens due to continuous cropping, especially aflatoxin contamination from Aspergillus flavus, severely restricts the development of the soybean and peanut industries.

[0003] Microbial fertilizers, as a new type of eco-friendly fertilizer, can effectively solve these problems. They play a vital role in improving and maintaining soil fertility, accelerating nutrient conversion, promoting crop growth, controlling soil-borne diseases, and purifying the environment. Addressing the continuous cropping obstacles in peanuts and soybeans, particularly the aflatoxin contamination caused by Aspergillus flavus and the reduced root nodules after chemical fertilizer application, the Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences has developed an ARC microbial inoculant. This inoculant is composed of a series of aflatoxin-producing biocontrol strains, including Bacillus amyloliquefaciens, Bacillus laterosporus, Bacillus mucilaginosus, and Enterobacter ludwig's bacterium. It simultaneously achieves green control of aflatoxin at the field source and enhances nitrogen fixation capacity in root nodules, exhibiting significant effects in promoting growth, increasing yield, controlling diseases, and ensuring safety.

[0004] The core of microbial fertilizers lies in their content of specific, effective live microorganisms. Any factor affecting their survival rate will impact the effectiveness of microbial fertilizers. However, existing microbial fertilizer technologies face the following pressing technical problems that severely restrict their effectiveness and widespread application: First, it is incompatible with chemical fertilizers: the high-salt environment caused by chemical fertilizers (especially NPK compound fertilizers) leads to severe dehydration and inactivation of microbial cells, and existing external wrapping isolation technology cannot effectively block the penetration of salt ions.

[0005] Second, the survival rate is low: after granulation, storage, transportation and application into the soil, the microbial cells are easily affected by high temperature, mechanical extrusion, acid and alkali stress, high salt osmotic pressure stress, ultraviolet radiation and other factors, resulting in a large number of deaths, short shelf life and poor field persistence.

[0006] Third, it is easy to fall off: Existing coating technology uses adhesives to wrap microorganisms on the surface of fertilizer particles. In a dry state, the microorganisms and fertilizer particles are only bound by physical forces such as van der Waals forces and hydrogen bonds. The binding strength is weak, and it is easy to fall off due to mechanical forces during storage and transportation.

[0007] Fourth, poor controlled release: Traditional coating and adhesion technology cannot achieve intelligent response to soil moisture, resulting in irregular release of bacteria after the inoculant is applied to the soil under alternating wet and dry conditions. The bacteria are easily lost with water, and their growth and reproduction cannot be synchronized with crop growth.

[0008] Fifth, the encapsulation wall materials have limited functionality and poor synergy: Existing single-wall material encapsulation systems (polyvinyl alcohol, polydopamine, natural polysaccharides, etc.) have significant shortcomings. Polyvinyl alcohol has poor salt resistance and a loose structure; polydopamine has weak acid resistance and insufficient biocompatibility; many synthetic polymer encapsulation materials (such as certain epoxy resins and polyurethanes) are toxic to microorganisms, affecting their growth and reproduction. Single modified systems cannot simultaneously achieve multiple functions such as salt barrier, UV resistance, erosion resistance, acid-base buffering, intelligent controlled release, and germination promotion.

[0009] Sixth, defects in fertilizer loading process: Traditional microbial fertilizers mostly use wet spraying and wet adhesion processes to coat microorganisms on the surface of fertilizer particles. Moisture can easily cause spores to germinate prematurely and become inactive, and can also easily cause compound fertilizer particles to deliquesce and clump together, making it impossible to achieve waterless, damage-free, and stable loading.

[0010] Therefore, there is an urgent need to develop a multifunctional synergistic technology for combining microbial agents with fertilizer granules, characterized by high barrier properties, high survival rate, strong binding, intelligent response, and good biocompatibility. Summary of the Invention

[0011] In view of the shortcomings of the prior art, the purpose of this application is to provide a compound fertilizer containing highly active ARC microbial agents, its preparation method and application.

[0012] To achieve this objective, the present application adopts the following technical solution: In a first aspect, this application provides a compound fertilizer containing a highly active ARC microbial agent, the compound fertilizer comprising: fertilizer granules as a core layer, and ARC microbial agent encapsulated by a quaternary cross-linked composite wall material covering the fertilizer granules; the quaternary cross-linked composite wall material is obtained by cross-linking polyvinyl alcohol, soybean peptide, dopamine, and ammonium molybdate.

[0013] This application creatively designs a compound fertilizer that encapsulates ARC microbial agents using a quaternary cross-linked composite wall material composed of polyvinyl alcohol, soybean peptides, dopamine, and ammonium molybdate. The components work synergistically to achieve multifunctional integration. This application utilizes the self-polymerization of dopamine under alkaline conditions to form polydopamine active centers. The catechol groups of the polydopamine undergo coordination cross-linking with molybdenum ions in ammonium molybdate, forming rigid chemical cross-linking nodes. Simultaneously, the long-chain hydroxyl groups of polyvinyl alcohol and the amino / carboxyl groups of soybean peptides intertwine with the polydopamine backbone and coordination network through multiple hydrogen bonds, jointly constructing an organic-inorganic hybrid interpenetrating network structure that combines chemical and physical cross-linking. In this network, coordination cross-linking imparts structural stability to the wall material, while hydrogen bond cross-linking imparts flexibility and film-forming properties. Both work synergistically to achieve efficient encapsulation and controlled release of the ARC microbial agents. The specific cross-linking principles and effects of each component are as follows: In this application, dopamine can self-assemble into polydopamine, which, as the main encapsulation framework, exhibits excellent physicochemical stability, mechanical stability, and high-temperature resistance. This protects the encapsulated microorganisms from mechanical compression and physical impact during production and transportation, and resists high-temperature environments. Simultaneously, polydopamine possesses excellent photothermal conversion effects and the ability to maintain structural stability in high-salt environments. The aromatic ring structure of polydopamine provides hydrophobicity, blocking water and salt. Furthermore, polydopamine has abundant active sites, enabling it to form a dense cross-linked network with soybean peptides and polyvinyl alcohol, and can also complex molybdate ions.

[0014] Ammonium molybdate acts as a crosslinking reinforcing agent, with molybdate ions (MoO4) 2- It can interact with polydopamine to significantly increase the network cross-linking density of composite wall materials, physically fill pores, and strengthen salt barrier; at the same time, molybdenum is an essential component of nitrogenase and nitrate reductase, which can enhance the nitrogen fixation ability of rhizobia in legumes and improve protein synthesis efficiency.

[0015] Polyvinyl alcohol, as a hydrophilic and water-retaining basic physical barrier framework, has high crystallinity and a tightly arranged molecular chain. Under dry conditions, it can form a low-permeability continuous film layer, which can significantly block the direct penetration of water molecules and salt ions. It can form hydrogen bonds with polydopamine and soybean peptides to enhance the network stability of the composite wall material. Moreover, its polyhydroxy groups provide good hydration capacity. After being applied to the soil, it can quickly absorb and transfer water to the microcapsule core when the soil moisture is suitable, breaking the dormancy of spores and promoting the rapid reproduction of ARC microbial agents.

[0016] Soybean peptides, a product of soybean enzymatic hydrolysis, are rich in organic nutrients such as small-molecule amino acids, oligopeptides, and polypeptides. They possess both hydrophilic polar groups (amino and carboxyl groups) and nonpolar groups derived from hydrophobic amino acid residues, exhibiting amphiphilic-hydrophobic properties. This enhances the density and toughness of quaternary cross-linked composite wall materials. Soybean peptides bind a small amount of water molecules through hydrogen bonds, forming a stable hydration layer on the material surface. This prevents internal spores from directly contacting high-salt solutions and enables intelligent controlled release in response to soil moisture, without completely blocking adequate water penetration. Furthermore, amino acids such as L-alanine, L-glycine, and L-glutamic acid can serve as germination agents for dormant Bacillus spores in ARC microbial agents. Additionally, soybean peptides and their degradation products can directly provide high-quality organic nitrogen sources for ARC microbial agents and leguminous crops.

[0017] The overall design of the compound fertilizer in this application not only provides plants with the necessary inorganic fast-acting nutrients, but also improves the survival rate of ARC microbial agents in the compound fertilizer by encapsulating ARC microbial agents with quaternary cross-linked composite wall materials.

[0018] Furthermore, the preparation method of ARC microbial agent encapsulated in the polyvinyl alcohol-soybean peptide-dopamine-ammonium molybdate quaternary crosslinked composite wall material provided in this application is simple in process, easy to industrialize, and has good application prospects.

[0019] In some embodiments, the ARC microbial agent encapsulated in the quaternary cross-linked composite wall material is adhered to the outside of fertilizer particles by a dry anhydrous thermal activation method.

[0020] This application uses a dry, waterless, additive-free, thermally activated adhesive technology to combine fertilizer granules and ARC microbial agents encapsulated in quaternary cross-linked composite wall materials. No additional water and / or binders are required. It can be seamlessly integrated with existing compound fertilizer granulation production lines, and the raw materials are all industrially mass-produced products, making it easy to scale up industrial production.

[0021] In some embodiments, the compound fertilizer comprises, by weight parts: 99.0-99.9 parts fertilizer granules (e.g., 99 parts, 99.1 parts, 99.2 parts, 99.4 parts, 99.6 parts, 99.8 parts, 99.9 parts, etc.) and 0.1-1.0 parts ARC microbial agent encapsulated in quaternary cross-linked composite wall material (e.g., 0.1 parts, 0.2 parts, 0.4 parts, 0.6 parts, 0.8 parts, 1 part, etc.).

[0022] In some embodiments, the mass ratio of polyvinyl alcohol, soybean peptide, dopamine, and ammonium molybdate is (2-3):(0.5-1.5):(0.3-0.5):(0.088-0.353) (wherein, the specific values ​​of 2-3 can be 2, 2.2, 2.4, 2.6, 2.8, 3, etc.; the specific values ​​of 0.5-1.5 can be 0.5, 0.7, 0.9, 1, 1.3, 1.5, etc.; the specific values ​​of 0.3-0.5 can be 0.3, 0.35, 0.4, 0.45, 0.5, etc.; and the specific values ​​of 0.088-0.353 can be 0.088, 0.1, 0.15, 0.2, 0.25, 0.3, 0.353, etc.).

[0023] In some embodiments, the initial encapsulation concentration of the ARC microbial agent in the quaternary cross-linked composite wall material is 1×10⁻⁶. 8 ~1×10 9 CFU / mL (e.g., 1×10⁻⁶) 8 CFU / mL, 1×10 9 (CFU / mL, etc.).

[0024] In some embodiments, the strains in the ARC microbial agent are a combination of Bacillus laterosporus, Bacillus amyloliquefaciens, Enterobacter ludwig's bacillus, and Bacillus mucilaginosus.

[0025] In some embodiments, the raw materials for preparing the fertilizer granules include a combination of nitrogen raw materials, phosphorus raw materials, potassium raw materials, and clay.

[0026] In some embodiments, the nitrogen raw material includes any one or a combination of at least two of urea, monoammonium phosphate, liquid nitrogen, ammonium nitrate, and ammonium sulfate.

[0027] In some embodiments, the phosphorus raw material includes any one or a combination of at least two of monoammonium phosphate, diammonium phosphate, and phosphoric acid.

[0028] In some embodiments, the potassium raw material includes potassium sulfate and / or potassium chloride.

[0029] In some embodiments, the nitrogen raw material in the fertilizer granules comprises 12%-80% of the total mass of the fertilizer granules (e.g., 12%, 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, etc.), the phosphorus raw material comprises 10%-80% of the total mass of the fertilizer granules (e.g., 10%, 30%, 40%, 50%, 60%, 70%, 80%, etc.), and the potassium raw material comprises 12-80% of the total mass of the fertilizer granules (e.g., 12%, 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, etc.).

[0030] In some embodiments, the ARC microbial agent encapsulated in the quaternary cross-linked composite wall material is obtained by a preparation method comprising the following steps: (1) Mix polyvinyl alcohol and soybean peptides, then mix with dopamine salt solution and ammonium molybdate to obtain a quaternary wall material premix; (2) Mix the ARC microbial agent with the quaternary wall material premix liquid to obtain the ARC microbial agent encapsulated in the quaternary cross-linked composite wall material.

[0031] In some embodiments, the temperature at which polyvinyl alcohol and soybean peptides are mixed in step (1) is 20-30°C (e.g., 20°C, 22°C, 24°C, 26°C, 28°C, 30°C, etc.).

[0032] In some embodiments, after mixing polyvinyl alcohol and soybean peptide in step (1), the solution pH is further adjusted to 8.0-8.5 (e.g., 8.0, 8.1, 8.2, 8.3, 8.4, 8.5, etc.).

[0033] In some embodiments, the temperature at which the dopamine salt solution and ammonium molybdate are mixed in step (1) is 25-35°C (e.g., 25°C, 27°C, 29°C, 31°C, 33°C, 35°C, etc.).

[0034] In some embodiments, before mixing the ARC microbial agent with the quaternary wall material premix in step (2), the step of mixing the ARC microbial agent with a freeze-drying protectant is further included, wherein the freeze-drying protectant is preferably a mixture of soybean peptides and glycerol.

[0035] In some embodiments, the temperature at which the ARC microbial agent and the quaternary wall material premix liquid are mixed in step (2) is 25-35°C (e.g., 25°C, 27°C, 29°C, 31°C, 33°C, 35°C, etc.).

[0036] In some embodiments, the ARC microbial agent in step (2) is further dried after being mixed with the quaternary wall material premix.

[0037] Secondly, this application provides a method for preparing the compound fertilizer as described in the first aspect, the method comprising: The ARC microbial agent powder encapsulated in quaternary cross-linked composite wall material is mixed with fertilizer granules to obtain a compound fertilizer containing highly active ARC microbial agent.

[0038] In some embodiments, the mixing temperature is 80-95°C (e.g., 80°C, 85°C, 90°C, 95°C, etc.), and the time is 3-10 min (e.g., 3 min, 4 min, 6 min, 8 min, 10 min, etc.).

[0039] All other specific point values ​​not listed above within the numerical ranges mentioned above are optional and are all within the scope of protection of this application. For the sake of brevity, they will not be elaborated here.

[0040] Thirdly, this application provides the use of the compound fertilizer as described in the first aspect in the preparation of products for agricultural planting.

[0041] Compared with the prior art, this application has the following beneficial effects: First, drying and salt barrier: In a dry state, the quaternary cross-linked composite wall material forms a highly cross-linked interpenetrating network structure, effectively blocking the rapid intrusion of external liquid water and fertilizer salt ions. The hydrophilic groups of the quaternary cross-linked composite wall material can "bind water," reducing the free water activity. The hydroxyl groups of polyvinyl alcohol and the amino, carboxyl, and peptide bonds of soybean peptides have strong hydrogen bonding effects on water molecules, "fixing" the infiltrated water into bound / structured water, rather than freely diffused water, thereby reducing the water vapor permeability coefficient. Furthermore, polyvinyl alcohol, as the basic hydrophilic and water-retaining physical barrier framework, has high crystallinity and a tightly packed molecular chain, forming a low-permeability continuous film layer under dry conditions, significantly blocking the direct penetration of water molecules and salt ions. The dopamine in the quaternary cross-linked composite wall material can self-assemble into polydopamine, which contains a large number of aromatic ring structures providing hydrophobicity, thus blocking water and salt; molybdate ions (MoO4) 2- It can interact with polydopamine to significantly increase the network cross-linking density of composite wall materials, physically fill pores, and strengthen salt barrier; soybean peptides have hydrophilic-hydrophobic amphiphilic properties, which can enhance the density and toughness of composite wall materials. Soybean peptides bind a small amount of water molecules through hydrogen bonds to form a stable hydration layer on the surface of the material, preventing internal microorganisms from directly contacting high-salt solutions.

[0042] Secondly, high survival rate: Quaternary cross-linked composite wall materials have good physical and chemical stability, mechanical stability, UV resistance, acid and alkali resistance and high temperature resistance. They can protect the encapsulated ARC microbial agents from the adverse mechanical forces such as mechanical extrusion and physical impact during production and transportation, and resist high temperature environments. Quaternary cross-linked composite wall materials have excellent salt barrier properties, which can protect the encapsulated ARC microbial agents from the effects of high salt osmotic pressure, and significantly improve the storage stability and survival rate of ARC microbial agents in fertilizer environments.

[0043] Third, built-in spore germination nutrition: Soybean peptides not only participate in the construction of quaternary cross-linked composite wall materials, but their own rich amino acids and small peptides and their degradation products (amino acids, small peptides) can also directly serve as signaling substances and carbon and nitrogen sources for spore germination in ARC microbial agents.

[0044] Fourth, intelligent controlled release in response to soil moisture: During fertilizer storage, the highly cross-linked structure of the quaternary cross-linked composite wall material plays a protective role. After the fertilizer is applied to the soil, when the soil moisture is suitable, the polyvinyl alcohol and soybean peptides in the quaternary cross-linked composite wall material ensure hydrophilicity, and water can slowly penetrate into the microcapsules, providing water activity for ARC microbial agents and spore germination and growth. When used in conjunction with the sowing technology of seed fertilizer for soybeans, peanuts and other legumes, the germination, rooting and growth of soybeans, peanuts and other legumes can be synchronized with the germination, growth and reproduction of ARC microbial agents.

[0045] Fifth, good biocompatibility: All components of the quaternary cross-linked composite wall material are biodegradable or can directly provide nutrients for microorganisms and crops, making it biocompatible and environmentally friendly; polydopamine, polyvinyl alcohol and soybean peptides are all biodegradable, soybean peptides can directly provide nutrients for microorganisms and crops, and the molybdenum element in ammonium molybdate can promote nitrogen fixation in the root nodules of leguminous crops.

[0046] Furthermore, this application uses a dry, anhydrous, additive-free, and thermally activated adhesive technology to combine fertilizer granules and ARC microbial agents encapsulated in quaternary cross-linked composite wall materials. No additional water or binder is required, which can be seamlessly integrated with existing compound fertilizer granulation production lines. Moreover, the raw materials are all industrially mass-produced products, making it easy to scale up industrial production.

[0047] The overall design of the compound fertilizer in this application not only provides plants with the necessary inorganic fast-acting nutrients, but also improves the survival rate of microorganisms in the compound fertilizer by encapsulating ARC microbial agents with quaternary cross-linked composite wall materials. The preparation method of ARC microbial agents encapsulated with quaternary cross-linked composite wall materials provided in this application is easy to industrialize and has good application prospects. Attached Figure Description

[0048] Figure 1 The figure shows the results of the static release test under constant moisture gradient for Preparation Example 2-1 and Comparative Preparation Example 2-4 in this application. Detailed Implementation

[0049] To further illustrate the technical means and effects adopted in this application, the following describes the technical solution of this application in conjunction with preferred embodiments, but this application is not limited to the scope of the embodiments.

[0050] In the embodiments of this application, the ARC microbial inoculant used was provided by the Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, and was prepared by the preparation methods of Examples 1-3 and Example 5 disclosed in patent CN118185797A.

[0051] The soybean peptides used in this application were obtained from Xinyangfeng Agricultural Technology Co., Ltd.; polyvinyl alcohol was purchased from Wuhan Jiyesheng Chemical Co., Ltd.; ammonium molybdate was purchased from Tianjin Yongda Chemical Reagent Co., Ltd., and ammonium molybdate tetrahydrate ((NH4)6Mo7O) was also used. 24 ·4H2O).

[0052] The viable cell count detection methods in the following test examples are as follows: Viable cell counts were performed after pre-swelling ultrasonic disruption of microcapsules: 1.0 g of the microcapsule powder to be tested was accurately weighed and added to an Erlenmeyer flask containing 100 mL of sterile physiological saline (containing 0.9% NaCl and 0.1% Tween 80). The flask was allowed to stand at 4℃ for 30 min to pre-swell, and the flask was shaken every 10 min. The suspension was then transferred to a sterile beaker and placed in an ice-water bath at 0-4℃. A probe-type ultrasonic disruptor was used, with a power of 100 W (amplitude 20%), pulse mode, and ultrasonic waves for 2 s / 5 s intervals, for a total effective ultrasonic working time of 180 s. The ultrasonic probe was immersed 1.5-2 cm into the liquid surface without touching the beaker wall. After lysis, the mixture was immediately and thoroughly vortexed. The mixture was then serially diluted 10-fold with sterile physiological saline and spread onto nutrient agar (NA) plates for viable cell counts. Viable cell counts were performed according to GB / T 20287-2006 Agricultural Microbial Agents.

[0053] Preparation Example 1-1 This preparation example provides a quaternary cross-linked composite wall material, and the preparation method is as follows: (1) Weigh 5 g of polyvinyl alcohol (type 1788, degree of polymerization 1750, degree of alcoholysis 98%) and dissolve it in 100 mL of pure water. Stir at 90°C for 40 min until completely clear, and cool to 30°C for later use. Weigh 2 g of soybean peptides and dissolve them in 100 mL of Tris-HCl (pH 8.0) solution. Centrifuge (10000×g, 10 min, 4℃) to remove insoluble residues and then filter with a 0.22 μm aqueous filter membrane for sterilization. (2) Mix the cooled polyvinyl alcohol solution and soybean peptide solution at a volume ratio of 1:1 (total 200 mL), adjust the pH of the mixture to 8.0, add 0.8 g of dopamine hydrochloride, stir to dissolve, protect from light, add 1 mL of 0.2 M ammonium molybdate sterile solution (sterilized by 0.22 μm aqueous filter membrane), stir to mix well, and obtain the quaternary cross-linked composite wall material mixture.

[0054] Comparative Preparation Example 1-1 This comparative preparation example provides a wall material. The preparation method differs from that of Preparation Example 1-1 only in that polyvinyl alcohol, soybean peptides, and ammonium molybdate are not added, while all other conditions remain unchanged.

[0055] Comparative preparation examples 1-2 This comparative preparation example provides a wall material. The preparation method differs from that of Preparation Example 1-1 only in that soybean peptides, ammonium molybdate, and dopamine are not added, while all other conditions remain unchanged.

[0056] Comparative preparation examples 1-3 This comparative preparation example provides a ternary composite wall material. The only difference between this preparation method and that of Preparation Example 1-1 is that polyvinyl alcohol is not added, and the reduced mass of polyvinyl alcohol is proportionally allocated to soybean peptides, ammonium molybdate, and dopamine, while all other conditions remain unchanged.

[0057] Comparative preparation examples 1-4 The comparative preparation example provides a ternary composite wall material. The only difference between the preparation method and the preparation example 1-1 is that soybean peptides are not added, and the reduced mass of soybean peptides is proportionally allocated to polyvinyl alcohol, ammonium molybdate and dopamine, while other conditions remain unchanged.

[0058] Comparative preparation examples 1-5 The comparative preparation example provides a ternary composite wall material. The only difference between the preparation method and the preparation example 1-1 is that ammonium molybdate is not added, and the reduced mass of ammonium molybdate is proportionally distributed to polyvinyl alcohol, soybean peptides and dopamine. All other conditions remain unchanged.

[0059] Comparative preparation examples 1-6 The comparative preparation example provides a ternary composite wall material. The only difference between the preparation method and the preparation example 1-1 is that dopamine is not added, and the mass reduction of dopamine is proportionally distributed to polyvinyl alcohol, soybean peptide and ammonium molybdate. All other conditions remain unchanged.

[0060] Preparation Example 2-1 This preparation example provides an ARC microbial agent encapsulated in a quaternary cross-linked composite wall material, and the preparation method is as follows: (1) The ARC microbial inoculant fermentation broth composition was centrifuged (8000×g, 10 min, 4℃) to collect the bacterial cells. The cells were washed three times with 10 mM Tris-HCl buffer (pH 7.2), and then resuspended in sterile 10 mM Tris-HCl buffer (pH 8.0). The bacterial cell concentration was adjusted to 1×10⁻⁶ cells using a hemocytometer. 10 CFU / mL, store temporarily at 4℃ for later use; Before use, the bacterial suspension and the freeze-drying protectant (prepared by dissolving soybean peptides in 10 mM Tris-HCl at pH 8.0-8.5 to form a 30% (w / v) solution, mixing with glycerol at a mass ratio of 7:3, centrifuging at 10000×g at 4℃ for 10 min to remove insoluble residue, and filtering with a 0.22 μm hydrophilic PVDF membrane for sterilization) were mixed at a ratio of 1:1 (v / v) and incubated at 4℃ for 30 min to obtain the ARC microbial agent bacterial suspension; The ARC microbial agent suspension was mixed with the quaternary cross-linked composite wall material premix obtained in Preparation Example 1-1 at a ratio of 1:10 (v / v), and reacted for 2 h at 30°C, pH 8.0, in the dark, and with stirring at 150 r / min to obtain a dark brown reaction product. (2) The dark brown reaction product was pre-cooled to 4°C and transported to the atomizer by a peristaltic pump. It was then freeze-dried using a spray freeze-drying device with high-purity nitrogen as the atomizing medium and protective gas. The entire process was carried out in a closed and light-proof manner. The atomization parameters were set as follows: atomization pressure 0.25 MPa, feed rate 20 mL / min, sprayed into a liquid nitrogen tank at a temperature of -196°C for instantaneous freezing granulation, and spray height 15 cm. (3) After the spraying is finished, collect the frozen particles and transfer them to the freeze-drying chamber. Set the cold trap temperature to -80℃ and the drying procedure is as follows: pre-freeze at -45℃ for 2 h; sublimation drying for 15 h, control the shelf temperature to rise linearly from -30℃ to -10℃ at a rate of 0.5℃ / min, and control the vacuum degree at 30 Pa; desorption drying for 5 h, raise the shelf temperature to 25℃ at a rate of 1℃ / min, and maintain the vacuum degree at 15 Pa. After drying, the bacterial powder was collected under nitrogen protection in a relative humidity of less than 10%, yielding the ARC microbial agent encapsulated in the quaternary cross-linked composite wall material, with an effective viable count of 2.46 × 10⁻⁶. 10 CFU / g (of which the effective viable count of Bacillus amyloliquefaciens is 1.21 × 10⁻⁶) 10 The effective viable count of Bacillus laterosporus was 0.84 × 10⁻⁶ CFU / g. 10 The effective viable count of Bacillus subtilis (CFU / g) was 0.28 × 10⁻⁶. 10 The effective viable count of CFU / g and Ludwig's Enterobacter lumbricoides was 0.13 × 10⁻⁶. 10 (CFU / g), powder moisture content 2.4%.

[0061] Comparative Preparation Example 2-1 This comparative preparation example provides a freeze-dried ARC microbial inoculant powder, prepared by the following method: (1) The ARC microbial inoculant fermentation broth composition was centrifuged (8000×g, 10 min, 4℃) to collect the bacterial cells. The cells were washed three times with 10 mM Tris-HCl buffer (pH 7.2), and the bacterial cells were resuspended in sterile 10 mM Tris-HCl buffer (pH 8.0). The bacterial cell concentration was adjusted to 1×10⁻⁶ cells using a hemocytometer. 10 CFU / mL, store at 4℃ for later use; Before use, the bacterial suspension and the freeze-drying protectant (prepared by dissolving soybean peptides in 10 mM Tris-HCl at pH 8.0-8.5 to form a 30% (w / v) solution, mixing with glycerol at a mass ratio of 7:3, centrifuging at 10000×g at 4℃ for 10 min to remove insoluble residue, and filtering with a 0.22 μm hydrophilic PVDF membrane for sterilization) were mixed at a ratio of 1:1 (v / v) and incubated at 4℃ for 30 min to obtain the ARC microbial agent bacterial suspension; (3) The ARC microbial agent suspension was pre-cooled to 4°C and transported to the atomizer by a peristaltic pump. It was then freeze-dried using a spray freeze-drying device with high-purity nitrogen as the atomizing medium and protective gas. The entire process was carried out in a closed and light-proof manner. The atomization parameters were set as follows: atomization pressure 0.25 MPa, feed rate 20 mL / min, sprayed into a liquid nitrogen tank at a temperature of -196°C for instantaneous freezing granulation, and spray height 15 cm. (4) After the spraying is finished, collect the frozen particles and transfer them to the freeze-drying chamber. Set the cold trap temperature to -80℃ and the drying procedure is as follows: pre-freeze at -45℃ for 2 h; sublimation drying for 15 h, control the shelf temperature to rise linearly from -30℃ to -10℃ at a rate of 0.5℃ / min, and control the vacuum degree at 30 Pa; desorption drying for 5 h, raise the shelf temperature to 25℃ at a rate of 1℃ / min, and maintain the vacuum degree at 15 Pa. After drying, the bacterial powder was collected under nitrogen protection at a relative humidity of less than 10%, yielding the ARC microbial agent freeze-dried bacterial powder with an effective viable count of 2.61 × 10⁻⁶. 10 CFU / g (of which the effective viable count of Bacillus amyloliquefaciens is 1.29 × 10⁻⁶) 10 The effective viable count of Bacillus laterosporus was 0.90 × 10⁻⁶ CFU / g. 10 The effective viable count of Bacillus subtilis (CFU / g) was 0.29 × 10⁻⁶. 10 The effective viable count of CFU / g and Ludwig's Enterobacter lumbricoides was 0.13 × 10⁻⁶. 10 (CFU / g), powder moisture content 2.3%.

[0062] Comparative Preparation Examples 2-2 to 2-7 Comparative Preparation Examples 2-2 to 2-7 each provide an ARC microbial agent encapsulated by a wall material. The only difference between the preparation method and Preparation Example 2-1 is that the quaternary cross-linked composite wall material obtained in Preparation Example 1-1 is replaced by the wall material obtained in Comparative Preparation Examples 2-1 to 2-6 in equal mass, while all other conditions remain unchanged.

[0063] Test Example 1 This test example determined the encapsulation rate of the quaternary cross-linked composite wall material in Preparation Example 2-1 for the ARC microbial agent and examined the release behavior of the encapsulated microbial agent (Preparation Example 2-1) in a simulated environment to evaluate the encapsulation and sustained-release effects of the quaternary cross-linked composite wall material.

[0064] I. Encapsulation Rate Determination (1) Initial total viable count Accurately weigh 1.0 g of the microcapsule bacterial powder sample to be tested (Preparation Example 2-1), add it to an Erlenmeyer flask containing 100 mL of sterile physiological saline (containing 0.9% NaCl, 0.1% Tween 80, pH 7.2), and perform viable count after pre-swelling ultrasonic disruption of the microcapsules. The viable count is recorded as the initial total viable count. Set up 3 parallel groups, and the relative deviation of the viable count of the parallel samples should be ≤10%.

[0065] (2) Count of free viable bacteria in the embedded sample Accurately weigh 1.0 g of the microcapsule bacterial powder sample to be tested (Preparation Example 2-1), and add it to an Erlenmeyer flask containing 100 mL of sterile physiological saline (containing 0.9% NaCl, 0.1% Tween 80, pH 7.2). Place the Erlenmeyer flask in a constant temperature shaker and gently shake at 25℃ and 100 r / min for 2 min to gently wash away the bacteria adsorbed on the surface of the microcapsules. Let it stand for 10 min, place the pipette tip at 1 / 2 depth of the liquid surface, and accurately aspirate 10 mL of the supernatant under sterile conditions. Dilute the supernatant 10-fold with sterile physiological saline and spread it on nutrient agar (NA) plates for viable count. The viable count is recorded as the number of free viable bacteria in the embedded sample. Set up 3 parallel groups, and the relative deviation of the viable count of the parallel samples should be ≤10%.

[0066] (3) Calculation of embedding rate Encapsulation efficiency (%) = [(Initial total viable count - Number of free viable bacteria in the encapsulated sample) / Initial total viable count] × 100% The test results are shown in Table 1.

[0067] II. Release Rate Measurement (1) Initial total viable count Accurately weigh 1.0 g of the microcapsule bacterial powder sample to be tested (Preparation Example 2-1), add it to an Erlenmeyer flask containing 100 mL of sterile physiological saline (containing 0.9% NaCl, 0.1% Tween 80, pH 7.2), and use pre-swollen ultrasonic to break up the microcapsules. Then, use sterile physiological saline to perform a 10-fold serial dilution and spread it on nutrient agar (NA) plates for viable cell count. The viable cell count is recorded as the initial total viable cell count. Set up 3 parallel groups, and the relative deviation of the viable cell count of the parallel samples should be ≤10%.

[0068] (2) Count of free viable bacteria released instantaneously Accurately weigh 1.0 g of the lyophilized microcapsule powder sample (Preparation Example 2-1) and place it in a pre-sterilized 250 mL Erlenmeyer flask with a stopper. Add 200 mL of sterile physiological saline (containing 0.9% NaCl, 0.1% Tween 80, pH 7.2) pre-temperatured to 25°C to the Erlenmeyer flask, gently shake to completely disperse the sample, and seal the mouth of the Erlenmeyer flask to prevent contamination by other microorganisms. Place the sealed Erlenmeyer flask in a constant temperature incubator set at 25°C, keeping it static and protected from light throughout the process. Take samples at 0 h, 3 h, 6 h, 12 h, 24 h, 3 d, 7 d, and 14 d. At the corresponding time points, remove the sample flask from the constant temperature incubator and let it stand at room temperature for 10 min. Under sterile conditions, place the pipette tip at 1 / 2 depth of the liquid surface and accurately aspirate 10 mL of the upper suspension. At the same time, immediately add 10 mL of sterile physiological saline pre-temperatured to 25°C to ensure that the total volume of the system remains constant. Each extract was serially diluted 10-fold with sterile physiological saline and spread onto nutrient agar (NA) plates for viable cell counting. The viable cell count was the number of free viable cells released at that time point. Three parallel groups were set up, and the relative deviation of the viable cell count of the parallel samples should be ≤10%.

[0069] A blank control group (preparation example 2-1) was set up, and the operation was completely consistent with that of the microcapsule group (preparation example 2-1). The initial viable bacteria count at time 0 of the blank control group was used as the benchmark, and the ARC microbial survival rate K at each sampling time point was calculated. n :K n = (Number of free viable bacteria measured at the nth time point of the blank group + total number of free viable bacteria carried away by the count at time points 0 to n-1) / initial total number of viable bacteria in the blank group at time 0.

[0070] (3) Calculation of cumulative release rate Calculate the cumulative release rate at each time point using the formula: Cumulative release rate = (V0 × C) / (V0 × C) n +V s ×(C1 / K1+C2 / K2+…+C n-1 / K n-1 )) / (C t ×V t ) × 100% Where: V0 is the total volume of the extraction system (200 mL), V s For a single sample volume (10 mL), C n K represents the concentration of released free viable bacteria (CFU / mL) measured at the nth time point. n K represents the survival rate K of the free spores in the blank control group measured at the nth time point. n C t V represents the initial total viable bacteria concentration (CFU / mL) of the microcapsule lyophilized powder. tThe total volume of the initial total viable bacteria count system for the microcapsule lyophilized powder is 100 mL.

[0071] The test results are shown in Table 1. Test results showed that the quaternary cross-linked composite wall material achieved a 93.2% encapsulation rate of ARC bacterial agent. Release performance results indicated that the quaternary cross-linked composite wall material continuously and slowly released ARC bacterial agent over 14 days. Initially, the release was slow, reaching 10.9% at 12 hours, followed by a rapid increase in the release rate: 27.7% at 1 day, 59.0% at 2 days, and 72.8% at 3 days. The release rate then slowed down and plateaued over time, reaching 83.5% at 7 days and 86.2% at 14 days, demonstrating significant slow-release characteristics. This indicates that the quaternary cross-linked composite wall material has excellent encapsulation and slow-release effects on ARC bacterial agent.

[0072] Test Example 2 This test example conducts high-temperature accelerated stability tests on the ARC microbial agents prepared in Example 2-1 and Comparative Preparation Examples 2-1 to 2-7.

[0073] Referring to the temperature control conditions of GB / T 19136-2021, the heat storage time was extended to 30 days for accelerated testing of the heat storage stability of the ARC microbial agents in Preparation Example 2-1 and Comparative Preparation Examples 2-1 to 2-7: 10 g of bacterial powder from each of the eight treatment groups was weighed, with three replicates per group. The samples were sealed and numbered in brown glass bottles and stored in a constant temperature oven at 54℃±2℃ for 30 days. Viable cell counts were measured on days 0, 7, 14, 21, and 30 (each time point and replicate was stored in an independently sealed brown glass bottle; the sample was not returned to the oven after sampling). At each time point, the samples were cooled to room temperature for 30 minutes, and then the microcapsules were disrupted using pre-swollen ultrasonication before viable cell counting. The weighed samples were then placed in a constant temperature oven for continuous aging, ensuring complete protection from light and moisture. The survival rate was calculated based on the viable cell count at day 0, and was expressed as the percentage of the tested viable cell count to the viable cell count at day 0.

[0074] The results are shown in Table 2, which summarizes the survival rate (%) of ARC microbial agents at 5 time points under accelerated conditions of 54℃±2℃ for 8 treatment groups. The results showed that the unencapsulated ARC microbial agent (Comparative Preparation 2-1) was rapidly deactivated at high temperatures, with the lowest survival rate at 30 days, only 9.2%, a decrease of 90.8%. The ARC microbial agents of Comparative Preparations 2-2 to 2-3, which were encapsulated with a single wall material, had relatively poor high-temperature resistance, with a survival rate of less than 50% at 30 days. The thermal stability of the ARC microbial agent encapsulated with pure polyvinyl alcohol was weaker than that treated with pure dopamine. The heat resistance of the ternary groups lacking any one component of the wall material (comparative preparation examples 2-4 to 2-7) was significantly lower than that of the complete quaternary group (preparation example 2-1). The heat resistance of the ternary groups (comparative preparation examples 2-4 to 2-7) was better than that of comparative preparation examples 2-2 and 2-3, which were encapsulated with a single wall material. The ARC microbial agent of comparative preparation example 2-1, which was encapsulated with a quaternary synergistic system, had the best heat resistance, and still retained 85.4% survival rate after 30 days of high-temperature acceleration, which was much higher than all other treatments.

[0075] Test Example 3 This test example examines the acid and alkali resistance of the ARC microbial agents prepared in Example 2-1 and the comparative preparations 2-1 to 2-7.

[0076] 1.0 g of ARC microbial inoculant powder from each of the 8 groups was accurately weighed and placed in a sterile Erlenmeyer flask. After pre-swelling and ultrasonic disruption, the powder was serially diluted 10 times with sterile physiological saline and then plated for viable count. The viable count was recorded as the initial viable count before stress (0 h).

[0077] Each of the eight groups of ARC microbial inoculant powders was accurately weighed to 1.0 g and added to a sterile Erlenmeyer flask containing 100 mL of sterile 0.1 mol / L acetate-sodium acetate buffer (pH 4.0). After being sealed and allowed to stand at 25°C for 24 h, the mixture was stirred, pre-swollen and ultrasonically broken up, and then serially diluted 10 times with sterile physiological saline. Finally, viable bacteria were counted on agar plates, and the viable bacteria count was recorded as the acid stress viable bacteria count.

[0078] Each of the eight groups of ARC microbial inoculant powders was accurately weighed to 1.0 g and added to a sterile Erlenmeyer flask containing 100 mL of sterile 0.1 mol / L CAPS buffer (pH 10.0). After being sealed and allowed to stand at 25°C for 24 h, the powders were mixed and then subjected to pre-swelling ultrasonic disruption. The mixtures were then serially diluted 10-fold with sterile physiological saline and plated for viable count. The viable count was recorded as the alkali stress viable count.

[0079] Survival rate was calculated based on the initial viable bacterial count before stress (0 h), using the following formula: Acid / alkali stress survival rate (%) = (Number of viable bacteria under acid / alkali stress / Initial number of viable bacteria before stress) × 100% The test results are shown in Table 3. The results showed that all encapsulation treatments (Preparation Example 2-1, Comparative Preparation Examples 2-1 to 2-7) had higher survival rates under acid and alkali conditions than the unencapsulated treatments. The unencapsulated ARC microbial agent had no protection, and the microbial cells directly exposed to acid and alkali ion stress had the lowest survival rate.

[0080] ARC microbial agents encapsulated with a single wall material exhibit relatively poor acid and alkali resistance. ARC microbial agents encapsulated with pure polyvinyl alcohol show better acid resistance than those treated with pure dopamine. At pH 4.0, the pure polydopamine encapsulation layer structure is relatively stable, but its porous structure makes it difficult to prevent hydrogen ion penetration. In contrast, pure polyvinyl alcohol can form a dense film-forming matrix, effectively blocking hydrogen ion penetration. ARC microbial agents encapsulated with pure polyvinyl alcohol show weaker alkali resistance than those treated with pure dopamine. At pH 10.0, pure polyvinyl alcohol, as the sole wall material, is prone to molecular chain breakage and disintegration after prolonged immersion, resulting in a significant decrease in the survival rate of the encapsulated ARC microbial agents.

[0081] The acid and alkali resistance of wall materials with ternary components is better than that of wall materials with single components. Comparative preparations 2-4 (wall material: soybean peptide, ammonium molybdate, and dopamine), 2-5 (wall material: polyvinyl alcohol, ammonium molybdate, and dopamine), and 2-6 (wall material: polyvinyl alcohol, soybean peptide, and dopamine) all showed higher survival rates at pH 4.0 and 10.0 than Comparative preparation 2-2 (wall material: dopamine). Similarly, Comparative preparations 2-5 (wall material: polyvinyl alcohol, ammonium molybdate, and dopamine), 2-6 (wall material: polyvinyl alcohol, soybean peptide, and dopamine), and 2-7 (wall material: polyvinyl alcohol, soybean peptide, and ammonium molybdate) all showed higher survival rates at pH 4.0 and 10.0 than Comparative preparation 2-3 (wall material: polyvinyl alcohol). The acid and alkali resistance of ternary wall materials lacking any one component was significantly lower than that of the complete quaternary wall material preparation 2-1.

[0082] Test Example 4 This test example examines the salt tolerance of the ARC microbial agents prepared in Example 2-1 and the comparative preparations 2-1 to 2-7.

[0083] 1.0 g of ARC microbial inoculant powder from each of the 8 groups was accurately weighed and placed in a sterile Erlenmeyer flask. After pre-swelling and ultrasonic disruption, the powder was serially diluted 10 times with sterile physiological saline and then plated for viable count. The viable count was recorded as the initial viable count before stress (0 h).

[0084] Each of the eight groups of ARC microbial inoculant powders was accurately weighed to 1.0 g and added to a sterile Erlenmeyer flask containing 100 mL of 15% (w / v) sodium chloride sterile solution. After mixing, the mixture was sealed and allowed to stand at 25°C for 24 h. After pre-swelling and ultrasonic disruption, the mixture was serially diluted with sterile physiological saline and then plated for viable count. The viable count was recorded as the salt stress viable count.

[0085] Survival rate was calculated based on the initial viable bacterial count before stress (0 h): Salt stress survival rate (%) = (Number of viable bacteria under salt stress / Initial number of viable bacteria before stress) × 100% The test results are shown in Table 4. The results showed that under high salt stress of 15% NaCl, the encapsulation wall material significantly improved the survival rate of the ARC microbial agent. In contrast, the ARC microbial agent prepared in Example 2-1 without encapsulation protection suffered significant inactivation and exhibited the lowest salt tolerance when directly exposed to a high osmotic pressure environment. The survival rate of the ARC microbial agent encapsulated with a single wall material was improved, and the survival rate of the ARC microbial agent with polyvinyl alcohol as the wall material was higher than that of the ARC microbial agent with dopamine as the wall material, indicating that the polyvinyl alcohol group had better barrier salt tolerance than the dopamine group.

[0086] The salt resistance of any ternary encapsulation system is better than that of pure dopamine or pure polyvinyl alcohol single wall material, indicating that there is a synergistic salt resistance effect of multiple components; the survival rate of preparation example 2-1 in the high salt environment of 15% NaCl is the highest, indicating that the four components work synergistically and the salt resistance protection effect is the best.

[0087] Test Example 5 This test example examines the UV radiation resistance of the ARC microbial agents prepared in Example 2-1 and Comparative Preparation Examples 2-1 to 2-7.

[0088] 0.1 g of each of the ARC microbial inoculant powders from the 8 groups was accurately weighed and added to 10 mL of sterile physiological saline (containing 0.9% NaCl and 0.1% Tween 80). The powders were then subjected to pre-swelling ultrasonic disruption, serially diluted 10-fold, and plated for viable cell counting. The viable cell count was recorded as the initial viable cell count before irradiation (0 min).

[0089] Eight groups of ARC microbial inoculant powders were accurately weighed at 0.1 g each and spread evenly on the open bottom of sterile petri dishes, forming a thin layer ≤1 mm thick. Stacking was avoided to prevent obstruction of UV light. Three parallel samples were prepared for each group. A 40 W, 254 nm UV lamp was preheated for 10 min to stabilize the irradiation intensity. All samples were simultaneously placed 1 m below the lamp to ensure consistent irradiation intensity. Irradiation was precisely timed for 30 min, and the entire process was performed in a dark room to avoid interference from ambient light.

[0090] Immediately after irradiation, pre-swollen ultrasonic disruption was performed (with 10 mL of sterile physiological saline (containing 0.9% NaCl and 0.1% Tween 80)). After a 10-fold serial dilution, viable bacteria were counted on agar plates. The viable bacteria count was recorded as the viable bacteria count after UV irradiation. The survival rate was calculated based on the initial viable bacteria count before stress. Survival rate (%) = (Number of viable bacteria after UV irradiation / Initial number of viable bacteria before irradiation) × 100% The test results are shown in Table 5. The results showed that the four components of the quaternary preparation example 2-1 provided synergistic protection and exhibited the best UV resistance. Under 254 nm UV stress (40 W, 30 min, irradiation distance 1 m), the ARC microbial agent of the control example 2-1 without encapsulation protection was significantly inactivated upon direct exposure to UV light. The encapsulation wall material can improve the survival rate of the ARC microbial agent under UV irradiation.

[0091] The survival rate of the ARC microbial agent with polyvinyl alcohol as the wall material was only 8.5% higher than that of the control preparation 2-1. This is because polyvinyl alcohol is transparent as an encapsulation layer, has almost no UV absorption, and only provides weak physical shielding.

[0092] The survival rate of the comparative preparation example 2-2, with dopamine as the wall material, reached 43.7%, which was significantly higher than that of the comparative preparation example 2-3, indicating that dopamine is a key component for UV protection in the quaternary system.

[0093] Compared with preparation example 2-7 (wall material is polyvinyl alcohol, soybean peptide and ammonium molybdate), which is dopamine-free, its survival rate (18.1%) is only higher than that of preparation example 2-3, further confirming that the UV resistance of the dopamine-free system is greatly weakened. This is because polydopamine has an aromatic ring / conjugated structure and has strong UV absorption near 254 nm.

[0094] The survival rates of comparative preparations 2-4 to 2-6 were all higher than those of comparative preparation 2-2, indicating that soybean peptides and ammonium molybdate can enhance the cross-linking density of the system and improve the UV protection performance of the dopamine system.

[0095] Test Example 6 This test example conducts a soil moisture response test on the ARC microbial agents prepared in Example 2-1 and the comparative preparation example 2-4.

[0096] Borosilicate glass beads were used to simulate soil matrix, and static release tests under constant soil moisture gradient and alternating wet and dry release tests were conducted to study the microbial release behavior of ARC microbial agent.

[0097] Glass bead pretreatment: Select 40-100 mesh solid borosilicate glass microspheres, soak the glass beads in 10% dilute hydrochloric acid for 24 hours to remove surface metal ions and impurities, rinse repeatedly with deionized water until the supernatant is neutral, then calcine in a muffle furnace at 400℃ for 4 hours to completely remove trace organic matter, cool and then perform moist heat sterilization at 121℃ for 1 hour, dry at 105℃ until anhydrous, and cool to room temperature for later use.

[0098] Constant moisture gradient static release test: Under aseptic conditions, 100 g of pretreated borosilicate glass beads were weighed into a 250 mL sterile blue-capped bottle. Sterile deionized water was added according to the weight of the oven-dry beads to adjust the substrate moisture content to 8%, 16%, and 24% (w / w), corresponding to low moisture (drought-prone), moderate moisture (medium moisture), and high moisture (near field capacity) conditions, respectively, with corresponding volumetric moisture contents of approximately 12%, 24%, and 36%. After the moisture was adjusted, the bottle was sealed at 25°C for 24 h to allow the moisture to be fully redistributed within the borosilicate glass bead system and the liquid bridge structure to stabilize. The ARC microbial agents of Preparation Example 2-1 and Comparative Preparation Example 2-4 were added to the borosilicate glass beads at a dosage of 1% (w / w), respectively. After thorough mixing, the bottle cap was tightened and sealed. The bottle was then incubated at 25°C in the dark, with the entire process sealed to prevent uncontrolled moisture loss.

[0099] The experiment employed an independent parallel endpoint sacrifice sampling strategy, with independent parallel culture systems set up at each culture time point. Each sample was only tested once at the preset culture endpoint, and the samples were discarded after sampling without further culture or monitoring. No rinsing was performed throughout the experiment to eliminate interference from the instantaneous release of bacteria. Each treatment was set up with 3 parallel replicates, and endpoint sampling and testing were carried out at 0 days, 0.5 days (12 hours), 1 day, 3 days, and 7 days after culture.

[0100] Wet-dry release experiment: A four-stage wet-dry culture program was adopted (the order of change of the absolute dry substrate moisture content was 8%→24%→8%→24%, w / w) to simulate the fluctuation of soil moisture in farmland due to alternating drought and flooding. Under aseptic conditions, 100g of pretreated borosilicate glass beads were weighed into a 250mL sterile blue-capped bottle. Sterile deionized water was added according to the weight of the absolute dry beads to adjust the substrate moisture content to 8%. The bottle cap was tightened and the bottle was sealed at 25 ℃ for 24 h to ensure uniform initial substrate water potential. The ARC microbial agents of Preparation Example 2-1 and Comparative Preparation Example 2-4 were added to the borosilicate glass beads at a dosage of 1% (w / w). After thorough mixing, the bottle cap was tightened and sealed as the starting point of Stage I (low moisture).

[0101] The process for switching between cultivation and soil moisture conditions is as follows: Phase I (low moisture, 8% w / w): Incubate at 25℃ in the dark for 3 days, and immediately after the incubation period, take samples for final analysis. Phase II (high moisture, 24% w / w): After Phase I, sterile deionized water was sprayed onto the surface of the bead bed in small amounts multiple times, while turning and mixing. The moisture content was checked in real time according to the change in the total mass of the blue cap bottle until it reached 24% (w / w). The cap was tightened and the bottle was incubated at a constant temperature of 25°C in the dark for 3 days. At the end of the incubation, the endpoint sample was taken and tested immediately. Phase III (Low Moisture, 8% w / w): After Phase II, remove the caps from the blue-capped bottles and cover the bottle openings with a 0.22 μm hydrophobic ePTFE microporous breathable membrane, ensuring full coverage. Secure the membrane with rubber bands and place the treated blue-capped bottles in a 25℃ constant temperature incubator for controlled evaporation dehydration (simulating slow transpiration water loss at ambient field temperature, lasting 24 hours). Gently rotate the bead bed during this period. When monitoring moisture content, quickly remove the breathable membrane and rubber bands and tighten the caps. Weigh the total weight of the blue-capped bottles and the internal matrix system. After weighing, quickly open the caps, re-cover and reseal the bottles, and continue dehydration. The entire operation should be completed within 90 seconds to minimize uncontrolled water evaporation during the weighing stage. Allow the oven-dry matrix to dehydrate gently and evenly to the target moisture content of 8%. Once the target is reached, remove the breathable membrane and rubber bands, tighten the caps to completely seal, and incubate at 25℃ in the dark for 3 days. Immediately after incubation, take samples for endpoint testing. Phase IV (high moisture, 24% w / w): Same as Phase II procedure, with endpoint sampling and testing performed immediately after 3 days of culture.

[0102] Each stage of sampling adopted an independent parallel endpoint sacrifice sampling strategy. The blue-capped bottles were kept sealed throughout the process except for moisture adjustment operations to avoid uncontrolled moisture loss.

[0103] Sampling and viable cell count: Before sampling, gently invert and mix thoroughly. Aseptically weigh 1 g of sample and add it to a sterile centrifuge tube containing 10 mL of 0.1% Tween 80 sterile physiological saline. Vortex at medium-low speed for 3 sets × 10 s, with a 20 s resting time between sets. Then, slowly invert the centrifuge tube 30 times to mix thoroughly and let it stand for 2 min to allow the free bacteria adsorbed on the surface of the borosilicate glass beads to fully desorb and disperse into the liquid phase. Centrifuge the suspension at 25℃ and 1000 × g for 5 min to allow the intact microcapsules and borosilicate glass beads to settle. The free microbial cells released into the liquid phase are mainly retained in the supernatant. After standing for 2 min, aseptically aspirate 8 mL of the upper layer of the supernatant (avoiding the precipitation interface) and transfer it to a sterile tube for instantaneous free viable cell count. Take the initial matrix and ARC microbial agent mixture sample, use pre-swelling ultrasonic to break up the microcapsules, and then perform the initial total viable cell count (converted uniformly based on sample mass). The cumulative release rate is the percentage of the instantaneous free viable bacteria count to the initial total number of viable bacteria.

[0104] The results of the alternating dry and wet release test are shown in Table 6. The results of the static release experiment under constant moisture gradient are shown in [the table]. Figure 1 Compared with the ternary system of preparation examples 2-4, the cumulative release curves of ARC live bacteria encapsulated in the ARC microbial agent highly overlapped under low moisture (8% w / w), medium moisture (16% w / w), and high moisture (24% w / w) conditions. The system exhibited significant early burst release characteristics, with a cumulative release rate of 40%-50% in 0-1 days, a continuous rapid release phase in 1-3 days, and a cumulative release rate rising to 60%-80%. The release rate slowed down in 3-7 days, and finally the cumulative release rate stabilized at 80%-90%. The release curve showed a steep rise in the early stage and a slow and gradual rise in the later stage. Changes in environmental soil moisture could not regulate the release behavior of ARC microorganisms, and there was no water (moisture) response characteristic. Preparation Example 2-1: In a low-moisture (8% w / w) environment, the cumulative release curve of the ARC live bacteria encapsulated in the quaternary system was extremely flat, with minimal increase in the cumulative release rate, indicating that the capsule wall of the quaternary system inhibited the release of the encapsulated ARC microorganisms under drought conditions. In a high-moisture (24% w / w) environment, the cumulative release curve of the ARC live bacteria exhibited a typical S-shaped kinetic curve, with slow initial release effectively inhibiting burst release; the release rate gradually increased with prolonged immersion time; after reaching swelling equilibrium, the release slowed down and entered a plateau phase. Under medium-moisture conditions (16% w / w), the release level was between that of high and low moisture, indicating that the release rate could be continuously controlled by the moisture content (soil moisture).

[0105] The results of the alternating dry and wet release test are shown in Table 5. The release pattern of the ternary system encapsulated with ARC microbial agents from Preparation Examples 2-4 is basically consistent with the static release test under constant moisture gradient. Regardless of whether it is in a dry or wet phase, ARC microorganisms are continuously released. When switching from high moisture to dry conditions, the release process cannot be effectively blocked, and the cumulative release rate continues to increase. The switching between dry and wet environments does not change the overall release kinetics, further confirming that the ternary system does not possess the ability to regulate release due to moisture stimulation.

[0106] The quaternary system encapsulating ARC microbial agents in Preparation Example 2-1 exhibited reversible moisture-response characteristics: during the dry period, the quaternary composite capsule wall contracted, strongly restricting the diffusion of ARC microorganisms and resulting in a minimal increase in the cumulative release rate; when the environment shifted to high moisture conditions, the release rate of ARC microorganisms significantly increased, and the cumulative release rate rose markedly; upon switching back to a dry environment, the release almost ceased again. The ARC microbial release kinetics during the moist phase were consistent with the S-shaped release pattern under high moisture conditions in the static release experiment with a constant moisture gradient. The system can achieve "drought-induced slow-release dormancy and moist-oriented release" by relying on alternating soil dryness and wetness.

[0107] Example 1 This embodiment provides a compound fertilizer containing highly active ARC microbial inoculant, and the preparation method is as follows: (1) Preparation of fertilizer granules: 6.0% urea, 2.6% liquid ammonia, 15.0% ammonium sulfate, 2% sulfuric acid, 42.0% monoammonium phosphate, 27.0% potassium sulfate and 5.4% clay were mixed and granulated by spraying to obtain fertilizer granules (12-19-14). After drying, the granules were screened by a screening machine. The particle size of the fertilizer granules was 1.00 mm to 4.75 mm. (2) Adhesion of ARC microbial agent: 1000 g of compound fertilizer (12-19-14) fertilizer granules from the screening process were placed in the sugar coating machine and the heating was turned on to the material temperature of 90°C. 10 g of ARC microbial agent encapsulated in the quaternary cross-linked composite wall material obtained in Preparation Example 2-1 was evenly sprinkled into the rotating drum for mixing. The mixture was rotated and mixed for 3 min, without adding any water or binder. After the microbial powder is firmly embedded in the micro-melting layer on the surface of the fertilizer granules, heating is stopped, and rotation continues until the material cools to 25°C. The material is then discharged and sieved through an 18-mesh screen to remove fine powder, yielding the compound fertilizer (12-19-14) containing highly active ARC microbial agent, with a viable count ≥2×10⁻⁶. 8 CFU / g.

[0108] Comparative Example 1 This comparative example provides a compound fertilizer containing ARC microbial agent. The difference between the preparation method and Example 1 is that step (2) is as follows: 1000 g of compound fertilizer (12-19-14) fertilizer granules from the screening process are placed in a sugar coating machine, and the heating is turned on to the material temperature of 90°C. 10 g of ARC microbial agent obtained from Comparative Preparation Example 2-1 is evenly sprinkled into the rotating drum for mixing. 0.1% starch paste binder is sprayed by a high-pressure spray gun to adhere the ARC microbial agent to the surface of the fertilizer granules. After the ARC microbial agent adheres, continue rotating until the material cools to 25°C, then discharge. Sieve through an 18-mesh sieve to remove fine powder, yielding the compound fertilizer (12-19-14) containing the highly active ARC microbial agent, with a viable count ≥2×10⁻⁶. 8 CFU / g.

[0109] Comparative Examples 2-7 Comparative Examples 2-7 each provide a compound fertilizer containing a highly active ARC microbial agent. The only difference between the preparation method and Example 1 is that the ARC microbial agent encapsulated in the quaternary cross-linked composite wall material obtained in Preparation Example 2-1 is replaced by the encapsulated ARC microbial agent obtained in Comparative Preparation Examples 2-2 to 2-7 by equal mass, while all other conditions remain unchanged.

[0110] Comparative Example 8 This comparative example provides a compound fertilizer containing ARC microbial agent. The difference between the preparation method and Example 1 is that step (2) is: 1000 g of compound fertilizer (12-19-14) fertilizer particles from the screening process are placed in a sugar coating machine, and the heating is turned on to the material temperature of 90°C. 10 g of ARC microbial agent encapsulated in the quaternary cross-linked composite wall material obtained in Preparation Example 2-1 is evenly sprinkled into the rotating drum for mixing. The ARC microbial agent is adhered to the surface of the fertilizer particles by spraying atomized 0.1% starch paste binder through a high-pressure spray gun. After the ARC microbial agent adheres, continue rotating until the material cools to 25°C, then discharge. Sieve through an 18-mesh sieve to remove fine powder, yielding the compound fertilizer (12-19-14) containing the highly active ARC microbial agent, with a viable count ≥2×10⁻⁶. 8 CFU / g.

[0111] Comparative Example 9 This comparative example provides a compound fertilizer containing inactivated ARC microbial agents, and the preparation method is as follows: (1) Preparation of fertilizer granules: 6.0% urea, 2.6% liquid ammonia, 15.0% ammonium sulfate, 2% sulfuric acid, 42.0% monoammonium phosphate, 27.0% potassium sulfate and 5.4% clay were mixed and granulated by spraying to obtain fertilizer granules (12-19-14). After drying, the granules were screened by a screening machine. The particle size of the fertilizer granules was 1.00 mm to 4.75 mm. (2) Place 1000 g of compound fertilizer (12-19-14) granules from the screening process into a sugar coating machine, turn on the heating to the material temperature of 90°C, and after high-pressure steam sterilization of 10 g of ARC microbial agent encapsulated in the quaternary cross-linked composite wall material obtained in Preparation Example 2-1, evenly sprinkle it into the rotating drum for mixing, and rotate and mix for 3 min without adding any water or binder. After the bacterial powder is firmly embedded in the micro-melting layer on the surface of the fertilizer granules, heating is stopped, and the material is rotated until it cools to 25°C. The material is then discharged, and fine powder is removed by sieving through an 18-mesh sieve to obtain the compound fertilizer (12-19-14) containing inactivated ARC microbial agent.

[0112] Test Example 7 This test case examines the shelf-life viable count of compound fertilizers containing ARC microbial agents in Example 1 and Comparative Examples 1-8.

[0113] The compound fertilizers containing ARC microbial inoculant from Examples 1 and Comparative Examples 1-8 were sealed and stored at 25°C. The effective viable count of the compound fertilizers containing ARC microbial inoculant was tested every 6 months. The effective viable count was determined according to the plate count method in 5.1 of "NY / T 2321-2013 Microbial Fertilizer Product Inspection Procedure". Pre-swelling ultrasonic crushing treatment was added. The results are shown in Table 7 (unit: 100 million / g). The results showed that in Example 1, after 18 months of storage at room temperature, the number of viable bacteria in the compound fertilizer containing ARC microbial agent decreased from the initial value of 217 million / g to 209 million / g, with a microbial survival rate of 96.3%. In Comparative Example 1, no ARC microbial agent was encapsulated, resulting in poor microbial stability and a rapid decrease in the number of viable bacteria; after 18 months, the microbial survival rate was 68.0%. In Comparative Examples 2 and 3, the ARC microbial agent was encapsulated with a single wall material, and after 18 months, the microbial survival rates were 84.8% and 87.5%, respectively. In Comparative Examples 4 to 7, the ARC microbial agent was encapsulated with a ternary composite wall material, and after 18 months, the microbial survival rates were 78.9%, 83.4%, 84.2%, and 85.5%, respectively. In Comparative Example 8, the microbial survival rate was 81.9% after 18 months. These results indicate that encapsulation of the ARC microbial agent in this application significantly improves the microbial survival rate of the compound fertilizer containing ARC microbial agent.

[0114] Test Example 8 This test case examines the bacterial loading stability of compound fertilizers containing ARC microbial agents in Example 1, Comparative Example 1, and Comparative Example 8.

[0115] The storage and transportation environment was simulated by shaking and free fall. The binding and shedding of ARC microbial agent with compound fertilizer was monitored by detecting the viable bacteria retention rate before and after the test. 10.0 g of compound fertilizer containing ARC microbial agent from Example 1, Comparative Example 1, and Comparative Example 8 were accurately weighed, added to 100 mL of sterile physiological saline and mixed well. After a 10-fold serial dilution, viable bacteria were counted on plate. The number of viable bacteria was recorded as the initial viable bacteria count.

[0116] Take 100 g of each of the compound fertilizers containing ARC microbial agent from Example 1, Comparative Example 1, and Comparative Example 8, seal them in a sealed bag, place them on a shaker at 150 r / min and 25℃ for 24 h. After shaking, remove the sealed bag and drop it freely from a height of 1 m onto a hard surface. Repeat the free fall 3 times. Pass the mixture through a 100-mesh sieve. Collect and weigh the material that passes through the sieve (shed microbial powder and abraded fertilizer powder). The shedding rate is the percentage of the total mass of the fine powder passing through the sieve to the total mass of the fertilizer before the test. Take 10.0 g of the fertilizer that passes through the sieve after sieving, pre-swell and ultrasonically crush it, dilute it 10 times, and then perform plate coating for viable count. The number of viable bacteria is recorded as the number of viable bacteria after treatment. The viable bacteria retention rate is the percentage of the number of viable bacteria after treatment to the initial number of viable bacteria.

[0117] The test results are shown in Table 8. The results showed that under simulated transportation vibration and loading / unloading drop conditions, the fine powder shedding rate of Comparative Example 1 reached 3.8%, and the viable bacteria retention rate was 77.5%. The fine powder shedding rate of Comparative Example 8 reached 3.6%, and the viable bacteria retention rate was 79.4%. This indicates that the bacterial powder with ordinary physical adhesion is very easy to detach from the fertilizer carrier surface under continuous mechanical shear force and instantaneous impact, forming a large amount of free fine powder. In contrast, the fine powder shedding rate of Example 2 was only 2.1%, and the viable bacteria retention rate was 95.6%. This indicates that Example 2 detached less fine powder, and the detached material was mostly inorganic fertilizer abrasive. The thermal activation method changed the binding mode between the bacterial powder and the carrier, from physical adsorption to physicochemical bonding (such as local melting and coordination bond bonding), which significantly improved the binding capacity, thus showing excellent adhesion strength under the simulated fertilizer storage and transportation environment.

[0118] Test Example 9 This test case involves a field planting experiment of the compound fertilizers from Example 1 and Comparative Example 9.

[0119] Test location: Nenjiang City, Heilongjiang Province; Trial period: May 2025 - October 2025; Experimental crop: soybean, Heihe 43; Experimental treatment: The compound fertilizers of Example 1 and Comparative Example 9 were used as sample groups. The experimental area of ​​each group was 5 mu, and the amount of compound fertilizer used in each group was 18.7 kg / mu. It was applied as base fertilizer on May 17. Except for the fertilizer, the management of the two groups was exactly the same.

[0120] Field growth survey: A total of 3 growth surveys were conducted. The survey results from June 13th showed that the seedlings in Example 1 group exhibited better overall growth, a higher seedling survival rate, and slightly higher plant height than the 9 control groups. The stems were also thicker. The average plant height in Example 1 group was 10.2 cm, while the average in the 9 control groups was 10.0 cm. Regarding seedling survival rate, the seedlings in Example 1 group were stronger and more uniform at emergence, with a higher emergence rate of 92.4%, compared to 88.7% in the 9 control groups. In terms of root system, root nodules formed earlier in Example 1 group, and the average root hair length in Example 1 group was 0.94 cm longer than that in the 9 control groups.

[0121] The survey results on June 28 showed that the seedlings in Example 1 group were growing better overall than those in the control group 9, and the plant height was also significantly higher.

[0122] The survey results on August 3 showed that the average plant height of soybeans in Example 1 group was 101 cm, while the average plant height of comparative group 9 was 85 cm. The average number of pods per plant in Example 1 group was 38, including 3 one-pods, 15 two-pods, 16 three-pods, and 4 four-pods. The average number of pods per plant in comparative group 9 was 28, including 3 one-pods, 5 two-pods, 15 three-pods, and 5 four-pods.

[0123] Field yield measurement: The actual yield measurement was carried out on October 2nd. The yield of Example 1 group was 254.48 kg per mu, and the yield of Comparative Example 9 group was 245.20 kg per mu.

[0124] Test Case 10 This test case involves a field planting experiment of the compound fertilizers from Example 1 and Comparative Example 9.

[0125] Test location: Lühe Township, Zhengyang County, Zhumadian City, Henan Province; Trial period: June 2025 - September 2025; Experimental crop: Peanut, Wanhua No. 7; Experimental treatment: The compound fertilizers of Example 1 and Comparative Example 9 were used as the experimental group and the control group, respectively. The area of ​​each group was 2 mu. The amount of compound fertilizer used in both the experimental group and the control group was 60 kg / mu. It was applied as base fertilizer on June 13. Except for the fertilizer, all other management was the same between the two groups.

[0126] After planting, conduct a field growth survey: On June 25, the peanut seedlings in Example 1 emerged more uniformly and grew more vigorously.

[0127] On July 5th, the flowers in Example 1 group showed stronger growth, darker green leaves, and earlier flowering.

[0128] On July 15, after a period of continuous rainy weather, the peanuts in Example 1 group still showed vigorous growth, with well-developed root systems, abundant capillary roots, and rich rhizobia, and entered the pegging stage ahead of schedule.

[0129] On July 28, the flowers in Example 1 recovered quickly, grew vigorously, had few seedling deaths, developed root systems, abundant root nodules, and thick stems.

[0130] On August 14, the peanuts in Example 1 showed stronger growth, darker green leaves, earlier fruiting, and a larger yield.

[0131] On August 30, the peanuts in Example 1 yielded a larger quantity, had a better appearance, and did not turn yellow or age prematurely.

[0132] On September 4, the peanuts in Example 1 showed vigorous growth, did not suffer from nutrient deficiency in the later stages, and produced a larger yield.

[0133] Field yield measurement: On September 30, the actual yield of peanuts was measured. The peanut yield of Example 1 group was 680 jin / mu, and the peanut yield of Example 1 was 610 jin / mu, an increase of 70 jin / mu. Calculated at 3.8 yuan / jin, the income increased by 266 yuan / mu.

[0134] The applicant declares that this application illustrates the technical solution of this application through the above embodiments, but this application is not limited to the above embodiments, that is, it does not mean that this application must rely on the above embodiments to be implemented. Those skilled in the art should understand that any improvements to this application, equivalent substitutions of the raw materials of this application's products, addition of auxiliary components, selection of specific methods, etc., all fall within the protection scope and disclosure scope of this application.

[0135] The preferred embodiments of this application have been described in detail above. However, this application is not limited to the specific details of the above embodiments. Within the scope of the technical concept of this application, various simple modifications can be made to the technical solution of this application, and these simple modifications all fall within the protection scope of this application.

[0136] It should also be noted that the various specific technical features described in the above embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, this application will not describe the various possible combinations separately.

Claims

1. A compound fertilizer containing highly active ARC microbial inoculant, characterized in that, The compound fertilizer includes: fertilizer granules as the core layer, and ARC microbial inoculant encapsulated by a quaternary cross-linked composite wall material covering the fertilizer granules; The quaternary cross-linked composite wall material is obtained by cross-linking polyvinyl alcohol, soybean peptides, dopamine, and ammonium molybdate.

2. The compound fertilizer according to claim 1, characterized in that, The ARC microbial agent encapsulated in the quaternary cross-linked composite wall material is adhered to the outside of fertilizer particles by a dry, anhydrous thermal activation method.

3. The compound fertilizer according to claim 1 or 2, characterized in that, The compound fertilizer comprises, by weight, 99.0-99.9 parts fertilizer granules and 0.1-1.0 parts ARC microbial inoculant encapsulated in quaternary cross-linked composite wall material; And / or, the mass ratio of the polyvinyl alcohol, soybean peptide, dopamine, and ammonium molybdate is (2-3):(0.5-1.5):(0.3-0.5):(0.088-0.353). And / or, the initial encapsulation concentration of the ARC microbial agent in the quaternary cross-linked composite wall material is 1×10⁻⁶. 8 ~1×10 9 CFU / mL.

4. The compound fertilizer according to any one of claims 1-3, characterized in that, The strains in the ARC microbial agent are a combination of Bacillus laterosporus, Bacillus amyloliquefaciens, Enterobacter ludwig's bacillus, and Bacillus mucilaginosus. And / or, the raw materials for preparing the fertilizer granules include a combination of nitrogen raw materials, phosphorus raw materials, potassium raw materials, and clay.

5. The compound fertilizer according to any one of claims 1-4, characterized in that, The nitrogen raw materials include any one or a combination of at least two of the following: urea, monoammonium phosphate, liquid nitrogen, ammonium nitrate, and ammonium sulfate; And / or, the phosphorus raw material includes any one or a combination of at least two of monoammonium phosphate, diammonium phosphate, and phosphoric acid; And / or, the potassium raw material includes potassium sulfate and / or potassium chloride.

6. The compound fertilizer according to any one of claims 1-5, characterized in that, In the fertilizer granules, the mass percentage of nitrogen raw material is 12%-80% of the total mass of fertilizer granules, the mass percentage of phosphorus raw material is 10%-80% of the total mass of fertilizer granules, and the mass percentage of potassium raw material is 12%-80% of the total mass of fertilizer granules.

7. The compound fertilizer according to any one of claims 1-6, characterized in that, The ARC microbial agent encapsulated in the quaternary cross-linked composite wall material is obtained by a preparation method including the following steps: (1) Mix polyvinyl alcohol and soybean peptides, then mix with dopamine salt solution and ammonium molybdate to obtain a quaternary wall material premix; (2) Mix the ARC microbial agent with the quaternary wall material premix liquid to obtain the ARC microbial agent encapsulated in the quaternary cross-linked composite wall material.

8. The compound fertilizer according to claim 7, characterized in that, The mixing temperature of polyvinyl alcohol and soybean peptides in step (1) is 20-30℃; And / or, after mixing the polyvinyl alcohol and soybean peptides in step (1), the step further includes adjusting the pH of the solution to 8.0-8.5; And / or, the temperature at which the salt solution of dopamine and ammonium molybdate are mixed in step (1) is 25-35°C; And / or, before mixing the ARC microbial agent with the quaternary wall material premix in step (2), the step of mixing the ARC microbial agent with a freeze-drying protectant is also included, wherein the freeze-drying protectant is preferably a mixture of soybean peptides and glycerol; And / or, the temperature at which the ARC microbial agent in step (2) is mixed with the quaternary wall material premix is ​​25-35℃; And / or, after the ARC microbial agent in step (2) is mixed with the quaternary wall material premix, a vacuum freeze-drying step is also included.

9. The method for preparing the compound fertilizer according to any one of claims 1-8, characterized in that, The preparation method includes: The ARC microbial agent powder encapsulated in quaternary cross-linked composite wall material is mixed with fertilizer granules at 80-95℃ for 3-10 minutes to obtain a compound fertilizer containing highly active ARC microbial agent.

10. The use of the compound fertilizer according to any one of claims 1-8 in the preparation of products for agricultural planting.

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