Compound microorganism enhanced heavy metal passivated organic fertilizer and preparation method thereof

Abstract

The invention provides a compound microorganism enhanced heavy metal passivated organic fertilizer and a preparation method thereof, and belongs to the technical field of fertilizers. 22 to 28 parts of a passivation material; 6-9 parts of a compound microbial agent; 3.8 to 4.5 parts of a nutrition synergist; the adsorption capacity is improved to be larger than or equal to 350 mg / g through modification of a nanometer passivation material, a C8-AHL / c-di-GMP / anthraquinone-1, 5-disulfonate signal combination activates synthesis of a phenazine gene and beta-glucan, the Cd fixation rate reaches 86.3%, the HRP / GOx double-enzyme response gel accurately releases a microbial agent in a polluted environment, a magnetic field is used for assisting granulation, 85.6% of the microorganism survival rate is guaranteed, passivation-fertilizer efficiency-ecology synergistic improvement is achieved, and the adsorption capacity is improved to be larger than or equal to 350 mg / g. The problems of poor environmental adaptability and fertilizer efficiency splitting are solved.

Description

Technical Field

[0001] The invention belongs to the technical field of fertilizers, in particular to a composite microbial enhanced heavy metal passivation organic fertilizer and a preparation method thereof. Background Art

[0002] Currently, heavy metal passivation organic fertilizers are mainly achieved through two technical routes, namely passivation material carrier type and microbial addition type. The passivation material carrier type uses bentonite, biochar and other materials to load passivation components (such as phosphates and iron oxides), and fixes heavy metals through adsorption and precipitation. However, there are problems such as limited passivation capacity (nanomaterials are easy to agglomerate) and insufficient phosphorus activation (passivators competitively fix available phosphorus in the soil), which makes it difficult to coordinate passivation and fertilizer efficiency. The microbial addition type combines phosphate-solubilizing bacteria, heavy metal-resistant bacteria and organic fertilizers, and relies on microorganisms to secrete organic acids or extracellular polymers (EPS) to passivate heavy metals. However, there are three major defects: low survival rate of bacterial agents (high-temperature inactivation during granulation), delayed function activation (no pre-induction of key protein expression) and blind release (non-targeted environmental loss).

[0003] Currently, traditional passivation materials (such as hydroxyapatite) strongly adsorb available phosphorus while immobilizing Cd²⁺, resulting in a 20-40% decrease in soil phosphorus availability. Simple bacterial agent combinations lack metabolic synergy. Conventional coating materials (such as polyvinyl alcohol) have a release rate of >60% in non-contaminated soils, resulting in agent waste and the risk of salinization. In response to these issues, the present invention proposes a composite microbial-enhanced heavy metal passivation organic fertilizer and its preparation method. Through the synergistic modification of nano-passivation materials, precise regulation of bacterial metabolism, intelligent environmentally responsive release, and gradient structure process design, this method simultaneously achieves efficient Cd / Pb passivation, synergistically improves fertilizer efficiency, and protects the soil microbiome. This addresses the problems of poor environmental adaptability and the disconnect between bacterial agent inactivation and passivation fertilizer efficiency encountered in traditional technologies. Summary of the Invention

[0004] The purpose of the present invention is to address the above-mentioned problems in the existing technology and to propose a composite microbial enhanced heavy metal passivation organic fertilizer and a preparation method thereof.

[0005] The purpose of the present invention can be achieved through the following technical solutions: A composite microbial enhanced heavy metal passivation organic fertilizer, comprising, by mass: 65-72 parts of organic carrier; 22-28 parts of passivation material; 6-9 parts of composite microbial agent; 3.8-4.5 parts of nutrient synergist; The organic carrier is a mixture of humic acid and soybean meal, with a humic acid:soybean meal ratio of 3.2-3.5:1, providing highly active humic acid (≥45%), significantly improving heavy metal complexing capacity, while the soybean meal slowly releases nitrogen to reduce nutrient loss; The passivation material includes lanthanum phosphate-doped nano-phosphate rock powder and bentonite, with a mass ratio of 1.8:1 to 2.2:1; Among them, La:Ce=1:1; Among them, the bentonite is pretreated by ammonium thiosulfate intercalation; through synergistic effect, the saturated adsorption capacity of Cd²⁺ is ≥350mg / g; Among them, the particle size of nano phosphate rock powder is ≤80nm, and the specific surface area is ≥120m² / g; Bentonite interlayer spacing ≥ 2.0nm, S2O3²⁻ intercalation amount ≥ 12wt%; The composite microbial agent includes a fluorescent Pseudomonas agent and a composite Bacillus agent, the ratio of live bacteria counts of Pseudomonas to Bacillus is 2.3-2.5:1, and the total live bacteria count is ≥8×10 9 CFU / g; The Pseudomonas fluorescens agent used was a new-generation agent (Pseudomonas fluorescens DSX-2020) from Qingdao Dexin Biotechnology. The Bacillus spores used were a Hyvoda Bio-complex Bacillus spore agent (Hyvofeng), containing Bacillus subtilis HSZ-2101, Bacillus licheniformis, Paenibacillus gelatinus, and Bacillus amyloliquefaciens. Pseudomonas fluorescens DSX-2020 fixes nitrogen and chelates Cd / Pb, while the Hyvofeng complex Bacillus spore agent solubilizes phosphate and secretes β-glucan. Compared to single-bacterial agents, the field deactivation efficiency increased by 25%.

[0006] The nutrient synergist is chitosan and nano-cerium oxide-modified calcium alginate, with the ratio of chitosan to nano-cerium oxide-modified calcium alginate being 1:1.2-1:1.5. The deacetylation degree is ≥95%. Nano-cerium oxide-modified calcium alginate (CeO2 loading ≥8%) can eliminate free radicals and protect microbial activity. Combined with temperature-responsive pores (0.2-0.5 μm), the fertilizer utilization rate is increased by 30%.

[0007] Preferably, the Pseudomonas fluorescens agent (DSX-2020) secretes metallothionein after being induced by Cd²⁺, and its cysteine ​​content is ≥30mol%; The β-1,3-glucan content in the EPS synthesized by the composite Bacillus agent (Haiwofeng) is ≥65%.

[0008] Preferably, the passivation material is treated with low-temperature plasma hydrogen reduction at a power of 300 W for 5 minutes. After treatment, a LaPO4-CePO4 heterojunction is formed on the surface of the nano-phosphate rock powder, and the bentonite intercalation structure contains a S4O6²⁻ / S2O3²⁻ dual sulfur source, the effective valence state of S is ≥+2.5, and the saturated adsorption capacity for Cd²⁺ is ≥350 mg / g.

[0009] Preferably, the fertilizer further comprises 0.06-0.08 parts of a microbial interaction synergist, wherein the microbial interaction synergist comprises a C8-AHL quorum sensing molecule (purity ≥ 98%), a cyclic diguanylate (c-di-GMP) signal molecule, and anthraquinone-1,5-disulfonate (sulfonation degree ≥ 90%). Among them, C8-AHL:c-di-GMP:anthraquinone-1,5-disulfonate = 1:0.8:0.6.

[0010] Preferably, a thermo-induced phase-changing membrane-forming agent is added during granulation of the nutritional enhancer, wherein the thermo-induced phase-changing membrane-forming agent is poly (N-isopropylacrylamide), which forms temperature-responsive pores (pore size 0.2-0.5 μm) inside the granules.

[0011] A method for preparing a composite microbial-enhanced heavy metal passivation organic fertilizer comprises: S1. Enhanced bacterial function: Pseudomonas fluorescens was cultured in a medium containing 15 mg / L CdCl2 + 0.3 mM glutathione (the bacterial agent has been pre-optimized for heavy metal stress response), and the complex Bacillus sp. was cultured in a medium containing 5 g / L calcium magnesium phytate + 0.5% chitin oligosaccharide to activate its phosphate-solubilizing gene phoD; S2. Biomimetic mineralization loading: Pseudomonas adsorbed on ammonium thiosulfate intercalated bentonite at the silane coupling agent modified interface, and Bacillus loaded on lanthanum phosphate-doped nanophosphate rock powder via Ca²⁺-mediated bridging; S3, Directed Assembly Granulation: The mixed material is extruded with the assistance of a magnetic field (magnetic field strength 0.5T), and a radial gradient pore structure (porosity 35±1%) is formed inside the granules; S4, staged curing: initial drying at 40℃ (moisture content 15%), final drying at 25℃ (moisture content ≤10%); S5. Smart coating: The coating material is horseradish peroxidase (HRP) / glucose oxidase (GOx) dual enzyme response gel, and the coating layer thickness is 80±5μm.

[0012] Preferably, in step S2, the loading process parameters include a silane coupling agent modification concentration of 1.5 wt%, Ca 2+ Bridging pH 7.5 ± 0.2, bacterial load density ≥ 10 10 CFU / g.

[0013] Preferably, in step S5, among the enzyme activities of the dual-enzyme response gel, HRP activity is ≥200 U / g, and GOx activity is ≥150 U / g.

[0014] Compared with the existing technology, the composite microbial enhanced heavy metal passivation organic fertilizer and its preparation method have the following beneficial effects: 1. The present invention provides a composite microbial-enhanced heavy metal passivation organic fertilizer and a preparation method thereof. Through the synergistic modification of LaPO4-CePO4 heterojunction nano-phosphate rock powder and double-sulfur source intercalated bentonite, the passivation material has a saturated adsorption capacity of Cd²⁺ ≥380 mg / g and a Cd dissolution rate as low as 4.2%, significantly improving the heavy metal fixation capacity.

[0015] 2. The present invention provides a composite microbial-enhanced heavy metal passivation organic fertilizer and a preparation method thereof, which utilizes a microbial interaction synergist to activate the Pseudomonas phenazine gene and promote the synthesis of β-1,3-glucan by Bacillus, thereby increasing the Cd fixation rate to 89.2% and the effective phosphorus release by 67.3%, thereby resolving the contradiction between the passivator and the fertilizer in competing for nutrients.

[0016] 3. The present invention provides a composite microbial enhanced heavy metal passivation organic fertilizer and its preparation method, which uses HRP / GOx dual enzyme response gel coating to trigger targeted release in heavy metal polluted environments, so that the utilization rate of the microbial agent reaches 90%, and the rhizosphere colonization amount is increased to 10 7 CFU / g root to avoid non-target loss.

[0017] 4. The present invention provides a composite microbial-enhanced heavy metal passivation organic fertilizer and a preparation method thereof, which ensures a 30-day microbial survival rate of ≥85.6% in the field through magnetic field-assisted gradient granulation and segmented low-temperature curing process.

[0018] In summary, the present invention provides a composite microbial-enhanced heavy metal passivation organic fertilizer and a preparation method thereof. The adsorption capacity is increased to ≥350 mg / g through modification of nano-passivation materials. The C8-AHL / c-di-GMP / anthraquinone-1,5-disulfonate signal combination activates phenazine genes and β-glucan synthesis, so that the Cd fixation rate reaches 86.3%. The HRP / GOx dual-enzyme response gel accurately releases bacterial agents in polluted environments, and magnetic field-assisted granulation ensures an 85.6% microbial survival rate, achieving a synergistic improvement in passivation, fertilizer efficiency, and ecology, and solving the problem of poor environmental adaptability and the disconnection between fertilizer efficiency. DETAILED DESCRIPTION

[0019] The following are specific embodiments of the present invention, which further describe the technical solutions of the present invention, but the present invention is not limited to these embodiments.

[0020] A composite microbial enhanced heavy metal passivation organic fertilizer, comprising, by mass: 65-72 parts of organic carrier; 22-28 parts of passivation material; 6-9 parts of composite microbial agent; 3.8-4.5 parts of nutrient synergist; The organic carrier is a mixture of humic acid and soybean meal, with a humic acid:soybean meal ratio of 3.2-3.5:1, providing highly active humic acid (≥45%), significantly improving heavy metal complexing capacity, while the soybean meal slowly releases nitrogen to reduce nutrient loss; The passivation material comprises lanthanum phosphate-doped nano-phosphate rock powder and bentonite, with a mass ratio of 1.8:1-2.2:1, wherein the bentonite is pre-treated by ammonium thiosulfate intercalation; through synergistic effect, the saturated adsorption capacity of Cd²⁺ is ≥350mg / g; Among them, the particle size of nano phosphate rock powder is ≤80nm, and the specific surface area is ≥120m² / g; Bentonite interlayer spacing ≥ 2.0nm, S2O3²⁻ intercalation amount ≥ 12wt%; The composite microbial agent includes a fluorescent Pseudomonas agent and a composite Bacillus agent, the ratio of live bacteria counts of Pseudomonas to Bacillus is 2.3-2.5:1, and the total live bacteria count is ≥8×10 9 CFU / g; The Pseudomonas fluorescens agent used was a new-generation agent (Pseudomonas fluorescens DSX-2020) from Qingdao Dexin Biotechnology. The Bacillus spores used were a Hyvoda Bio-complex Bacillus spore agent (Hyvofeng), containing Bacillus subtilis HSZ-2101, Bacillus licheniformis, Paenibacillus gelatinus, and Bacillus amyloliquefaciens. Pseudomonas fluorescens DSX-2020 fixes nitrogen and chelates Cd / Pb, while the Hyvofeng complex Bacillus spore agent solubilizes phosphate and secretes β-glucan. Compared to single-bacterial agents, the field deactivation efficiency increased by 25%.

[0021] The nutrient synergist is chitosan and nano-cerium oxide-modified calcium alginate, with the ratio of chitosan to nano-cerium oxide-modified calcium alginate being 1:1.2-1:1.5. The deacetylation degree is ≥95%. Nano-cerium oxide-modified calcium alginate (CeO2 loading ≥8%) can eliminate free radicals and protect microbial activity. Combined with temperature-responsive pores (0.2-0.5 μm), the fertilizer utilization rate is increased by 30%.

[0022] The preparation method of nano-ceria modified calcium alginate is as follows: Dissolve 10 g of Ce(NO3)3·6H2O in 200 mL of deionized water and stir until clear. Dropwise add ammonia water until pH = 9.0 ± 0.2 to form Ce(OH)3 precipitate. Carry out hydrothermal reaction at 80 °C for 6 h, centrifuge and wash until neutral to obtain CeO2 nano sol (particle size 10–20 nm). Dissolve 5 g of sodium alginate in 100 mL of deionized water, add 0.8 g of PNIPAM, and stir to dissolve at 60 °C. Add CeO2 sol (containing 0.8 g of CeO2), and disperse by ultrasonic wave for 30 min (power 300 W) to make the CeO2 loading reach 8%. Drop the mixed solution into 5% CaCl2 solution (containing 0.1% Tween-80), and stir magnetically (200 rpm). The initial temperature is 40 °C (> LCST of PNIPAM ≈ 32 °C) to make PNIPAM shrink hydrophobically and form a dense network. Suddenly cool down to 10 °C (< LCST), PNIPAM swells hydrophilically to generate temperature-responsive pores with a size of 0.2–0.5 μm. Cure the gel beads for 30 min, wash and dry to obtain a porous CeO2-calcium alginate carrier. Immerse the dried carrier in 2% chitosan acetate solution (degree of deacetylation ≥ 95%) and oscillate at 25 °C for 2 h. Take it out and freeze-dry to obtain the final modified material.

[0023] The fluorescent pseudomonad agent (DSX-2020) secretes metallothionein after being induced by Cd 2+ and its cysteine content ≥ 30 mol%; The β-1,3-glucan in the EPS synthesized by the composite bacillus agent (Haiwofeng) accounts for ≥ 65%.

[0024] The passivation material is treated by low-temperature plasma hydrogen reduction with a power of 300 W and a time of 5 min. After treatment, a LaPO4-CePO4 heterojunction is formed on the surface of nano apatite powder, and the intercalation structure of bentonite contains a double sulfur source of S4O6²⁻ / S2O3²⁻, the effective valence state of S ≥ +2.5, and the saturated adsorption capacity for Cd²⁺ ≥ 350 mg / g.

[0025] The fertilizer also includes 0.06 - 0.08 parts of a bacterial community interaction synergist, and the bacterial community interaction synergist includes a C8-AHL quorum sensing molecule (purity ≥ 98%), a cyclic diguanylate (c-di-GMP) signaling molecule, and anthraquinone-1,5-disulfonate (sulfonation degree ≥ 90%); Among them, C8-AHL:c-di-GMP:anthraquinone-1,5-disulfonate = 1:0.8:0.6.

[0026] When granulating the nutrient synergist, a thermally induced phase change film-forming agent is added, and the thermally induced phase change film-forming agent is poly N-isopropylacrylamide, forming temperature-responsive pores (pore size 0.2 - 0.5 μm) inside the particles.

[0027] A method for preparing a composite microbial-enhanced heavy metal passivation organic fertilizer comprises: S1. Enhanced bacterial function: Pseudomonas fluorescens was cultured in a medium containing 15 mg / L CdCl2 + 0.3 mM glutathione (the bacterial agent has been pre-optimized for heavy metal stress response), and the complex Bacillus sp. was cultured in a medium containing 5 g / L calcium magnesium phytate + 0.5% chitin oligosaccharide to activate its phosphate-solubilizing gene phoD; S2. Biomimetic mineralization loading: Pseudomonas adsorbed on ammonium thiosulfate intercalated bentonite at the silane coupling agent modified interface, and Bacillus loaded on lanthanum phosphate-doped nanophosphate rock powder via Ca²⁺-mediated bridging; S3, Directed Assembly Granulation: The mixed material is extruded with the assistance of a magnetic field (magnetic field strength 0.5T), and a radial gradient pore structure (porosity 35±1%) is formed inside the granules; S4, staged curing: initial drying at 40℃ (moisture content 15%), final drying at 25℃ (moisture content ≤10%); S5. Smart coating: The coating material is horseradish peroxidase (HRP) / glucose oxidase (GOx) dual enzyme response gel, and the coating layer thickness is 80±5μm.

[0028] In step S2, the loading process parameters include silane coupling agent modification concentration of 1.5wt%, Ca²⁺ bridging pH 7.5±0.2, and bacterial loading density ≥10¹ 0 CFU / g.

[0029] In step S5 , the enzyme activity of the dual-enzyme response gel is ≥200 U / g, and the GOx activity is ≥150 U / g. Specific embodiment 1

[0031] Nano-phosphate rock powder was impregnated in 0.1M La(NO3)3 / Ce(NO3)3 solution (La:Ce=1:1), hydrolyzed at 60℃ for 2h to form LaPO4-CePO4 heterojunction, and centrifuged for drying; bentonite was intercalated with 0.5M (NH4)2S2O3 solution at 60℃ for 4h and washed to neutrality; lanthanum phosphate-doped nano-phosphate rock powder was composited with ammonium thiosulfate intercalated bentonite and then treated with H2 plasma (300W, 5min, H2 flow rate 20sccm).

[0032] Comparative Example 1: The difference from the first embodiment is that 100 nm hydroxyapatite nanopowder (specific surface area 110 m 2 / g) and calcium bentonite with an interlayer spacing of 1.2 nm were dry-mixed in a 1:1 mass ratio and ball-milled for 30 min (300 rpm).

[0033] Comparative Example 2: 100nm hydroxyapatite nanopowder and 1.2nm interlayer spacing calcium bentonite were mixed in a 1:1 mass ratio and treated with H2 plasma (300W, 5min, H2 flow rate 30sccm) without rare earth doping and sulfur intercalation pretreatment.

[0034] Detection method XPS: Thermo ESCALAB Xi+, S 2p spectrum peak fitting was used to determine the proportions of -S²⁻ (161.5 eV), -S²⁻ (162.8 eV), and S4O6²⁻ (166.2 eV), and the average S valence state was calculated.

[0035] Adsorption performance: Take 0.1g of material and add 50mL of Cd 2+ The solution (initial concentration 800 mg / L, pH = 6.0 ± 0.1) was shaken at a constant temperature of 25°C for 24 h, filtered through a 0.22 μm filter membrane, and the equilibrium concentration was determined by ICP-OES. The saturated adsorption capacity was fitted with the Langmuir model.

[0036] Stability test: Saturated adsorption of Cd 2+ The material was loaded into a leaching column (Φ2.5×30 cm), and simulated acid rain (mixed acid with pH 4.0 (H2SO4:HNO3=3:1) + 0.01M Na2SO4) was passed through it at a flow rate of 10 mL / min. The leaching was continued for 168 h (7 days). The leachate was collected every 24 h, and the cumulative Cd dissolution rate (dissolved Cd amount / total adsorbed Cd amount×100%) was determined.

[0037] Table 1 Verification of passivation material modification effect Performance indicators Comparative Example 1 Comparative Example 2 Specific embodiment 1 <![CDATA[Cd 2+ Adsorption capacity]]> 120±5mg / g 145±8mg / g 398±12mg / g S average valence +2.0 +2.0 +2.58 Cd dissolution rate 32.4±1.8% 28.5±2.1% 4.2±0.3% Table 1 shows that the modified passivation material (Specific Example 1) has the following characteristics: 2+ The material significantly outperformed both Comparative Example 1 (unmodified material) and Comparative Example 2 (plasma treatment alone) in terms of adsorption capacity (398 mg / g), sulfur valence (+2.58), and Cd dissolution rate (4.2%). Adsorption capacity increased by 89.5%, while dissolution rate decreased by 76.8%, demonstrating that the synergistic modification of lanthanum phosphate-doped nanophosphate rock powder with dual-sulfur bentonite significantly enhances the material's adsorption capacity and stability. Specific embodiment 2

[0039] For Pseudomonas enhancement, LB medium containing CdCl2 (15 mg / L) + glutathione (0.3 mM) was used and cultured at 30°C with shaking for 48 h; for Bacillus enhancement, medium containing calcium magnesium phytate (5 g / L) + chitin oligosaccharide (0.5%) was used and cultured at 30°C with shaking for 48 h.

[0040] A basic compound bacterial agent + bacterial interaction synergist was used, C8-AHL:c-di-GMP:anthraquinone-1,5-disulfonate = 1:0.8:0.6, and the synergist addition amount was 0.07wt%.

[0041] Comparative Example 3: Pseudomonas was fortified using LB medium containing CdCl2 (15 mg / L) and glutathione (0.3 mM) with shaking at 30°C for 48 hours. Bacillus was fortified using medium containing calcium magnesium phytate (5 g / L) and chitin oligosaccharide (0.5%) with shaking at 30°C for 48 hours. A basic inoculum without pre-induction (same strain ratio as in Specific Example 2) was used without a synergist.

[0042] Phenazine gene expression was detected using StepOnePlus RT-PCR; β-1,3-glucan was analyzed using RenishawinVia and glucan standards, and Raman spectroscopy (1085 cm -1 The total amount of EPS was determined by phenol-sulfuric acid method; the Cd content was determined by Agilent 7900 ICP-MS. 2+ (50mg / L) residual amount of culture medium (determined after EDTA washing) was used to calculate the fixation rate; the water-soluble phosphorus content in the insoluble phosphate culture medium (Ca3(PO4)210g / L) was determined using a UV-2600 spectrophotometer and the molybdenum antimony colorimetric method.

[0043] Table 2 Comparison of key indicators Detection indicators Comparative Example 3 Specific embodiment 2 p-value phzM gene expression 1.00±0.08 3.25±0.22 <0.001 The proportion of β-1,3-glucan in EPS 52.3±2.1% 62.5±2.3% <0.01 Cd²⁺ fixation rate 62.4±3.1% 86.3±2.7% <0.001 Available phosphorus release (mg / L) 34.6±1.5 42.7±1.8 <0.001 Microbial survival rate (30 days) 41.2±3.5% 83.1±3.2% <0.001 As can be seen from the above table, in the specific embodiment 2, C8-AHL activates phenazine genes to promote electron transfer and drives metallothionein to efficiently chelate Cd 2+ ; Anthraquinone-1,5-disulfonate (sulfonation degree 92%) is used as an electron mediator. Compared with comparative example three, anthraquinone-1,5-disulfonate accelerates electron transfer, enhances the anabolism of Bacillus EPS, and increases the proportion of β-1,3-glucan; c-di-GMP signal molecules coordinate the division of labor between the two bacteria. Pseudomonas is specialized in fixing Cd, and Bacillus is focused on phosphorus solubilization, which increases the release of effective phosphorus by 67.3% and avoids metabolic competition. In summary, Table 2 compares the effects of bacterial interaction enhancers. The specific example two of adding synergists has an effect on the expression of phzM gene, the proportion of β-1,3-glucan in EPS, and Cd 2+ The fixation rate and effective phosphorus release were significantly higher than those of the comparative example 3 without synergist, proving that the C8-AHL / c-di-GMP / anthraquinone signaling molecule combination simultaneously improved the efficiency of heavy metal fixation and phosphorus activation by activating phenazine genes and optimizing the division of labor in the bacterial community. Specific embodiment three

[0045] The fluorescent Pseudomonas agent was cultured in a medium containing 15 mg / L CdCl2 + 0.3 mM glutathione (the agent had been pre-optimized for heavy metal stress response), and the composite Bacillus agent was cultured in a medium containing 5 g / L calcium magnesium phytate + 0.5% chitin oligosaccharide to activate its phosphate-solubilizing gene phoD; Pseudomonas and ammonium thiosulfate intercalated bentonite were adsorbed on the interface modified by silane coupling agent, and Bacillus was activated by Ca 2+ The mediated bridging was loaded onto lanthanum phosphate-doped nanophosphate rock powder. The mixture was subjected to magnetic field-assisted extrusion (magnetic field strength 0.5 T), resulting in a radially gradient porosity structure within the particles (porosity 35 ± 1%). Initial drying was performed at 40°C (moisture content 15%) and final drying was performed at 25°C (moisture content ≤ 10%). The coating was a horseradish peroxidase (HRP) / glucose oxidase (GOx) dual-enzyme responsive gel with a coating thickness of 80 ± 5 μm. The HRP activity was ≥ 200 U / g, and the GOx activity was ≥ 150 U / g. Contaminated soil (pH 5.5, 30°C) was used. A microbial interaction enhancer (C8-AHL:c-di-GMP:anthraquinone-1,5-disulfonate = 1:0.8:0.6) was added. Comparative Example 4: The fluorescent Pseudomonas agent was cultured in a medium containing 15 mg / L CdCl2 + 0.3 mM glutathione (the agent had been pre-optimized for heavy metal stress response), and the composite Bacillus agent was cultured in a medium containing 5 g / L calcium magnesium phytate + 0.5% chitin oligosaccharide to activate its phosphate-solubilizing gene phoD; Pseudomonas and ammonium thiosulfate intercalated bentonite were adsorbed on the interface modified by silane coupling agent, and Bacillus was adsorbed by Ca 2+ The mediated bridging was loaded onto lanthanum phosphate-doped nanophosphate rock powder. The mixture was subjected to magnetic field-assisted extrusion (magnetic field intensity 0.5 T), forming a radially gradient pore structure (porosity 35±1%) within the particles. Initial drying was performed at 40°C (moisture content 15%) and final drying was performed at 25°C (moisture content ≤10%). The coating was a horseradish peroxidase (HRP) / glucose oxidase (GOx) dual-enzyme responsive gel with a thickness of 80±5μm. The HRP activity was ≥200U / g and the GOx activity was ≥150U / g. Clean soil (pH 7.0, 20°C) was used.

[0046] Comparative Example 5: The fluorescent Pseudomonas agent was cultured in a medium containing 15 mg / L CdCl2 + 0.3 mM glutathione (the agent had been pre-optimized for heavy metal stress response), and the composite Bacillus agent was cultured in a medium containing 5 g / L calcium magnesium phytate + 0.5% chitin oligosaccharide to activate its phosphate-solubilizing gene phoD; Pseudomonas and ammonium thiosulfate intercalated bentonite were adsorbed on the interface modified by silane coupling agent, and Bacillus was adsorbed by Ca 2+The mediated bridging loading was performed on lanthanum phosphate-doped nanophosphate rock powder. The mixture was subjected to magnetic field-assisted extrusion (magnetic field intensity 0.5 T), resulting in a radially gradient pore structure (porosity 35±1%) within the particles. Initial drying was performed at 40°C (moisture content 15%) and final drying was performed at 25°C (moisture content ≤10%). Conventional polyvinyl alcohol coating was applied to a thickness of 80±5μm, resulting in no enzyme response. Contaminated soil (pH 5.5, 30°C) was used.

[0047] Detection indicators Release kinetics: The cumulative release rate of the signal molecule (C8-AHL) was detected by HPLC-MS. Samples were taken every 24 hours, and the cumulative release rate over 3 days was calculated.

[0048] Microbial activity: Flow cytometry was used to detect the number of viable bacteria in fertilizer granules, with the number of viable bacteria at the time of preparation as the benchmark (≥8×10 9 CFU / g) and calculate the survival rate after 30 days.

[0049] Field matching: The colonization of in situ fluorescently labeled bacteria in the crop rhizosphere was measured using corn seedlings, with samples taken 30 days after planting to determine the rhizosphere colonization.

[0050] Table 3 Comparison of coating targeted release index Comparative Example 5 Specific embodiment three Comparative Example 4 3-day signal molecule release rate 68.0% 95.0% 12.0% 30-day microbial survival rate 40.0% 85.6% 85.0% Rhizosphere colonization (CFU / g root) <![CDATA[10 5 ]]> <![CDATA[10 7 ]]> <![CDATA[10 4 ]]> In Cd-contaminated soil (pH 5.5, 30°C), the specific example 3 of the dual-enzyme response gel coating showed significant differences in signal molecule release rate (95%), microbial survival rate (85.6%), and rhizosphere colonization rate (10 7 CFU / g root) were much better than the comparative example 5 (68%, 40%, 10 5 ) and the comparative example 4 of non-polluted environment (12%, 85%, 10 4 ), indicating that the HRP / GOx dual-enzyme responsive gel can accurately match the polluted environment and avoid non-targeted loss of bacterial agents.

[0051] The specific embodiments described herein are merely illustrative of the spirit of the present invention. Persons skilled in the art may make various modifications, additions, or substitutions to the described specific embodiments without departing from the spirit of the present invention or exceeding the scope of the appended claims.

Claims

1. A composite microbial enhanced heavy metal passivation organic fertilizer, characterized in that: Included by mass: 65-72 parts of organic carrier; 22-28 parts of passivation material; 6-9 parts of composite microbial agent; 3.8-4.5 parts of nutrient synergist; The organic carrier is a mixture of humic acid and soybean meal, with a ratio of humic acid to soybean meal of 3.2-3.5:1; The passivation material includes LaPO4-CePO4 heterojunction nano-phosphate rock powder and bentonite formed by impregnation with La(NO3)3 and Ce(NO3)3 solution (La:Ce=1:1) and then H2 plasma treatment, with a mass ratio of 1.8:1-2.2:

1. Among them, La:Ce=1:1; Among them, bentonite was pretreated by ammonium thiosulfate intercalation; Among them, the particle size of nano phosphate rock powder is ≤80nm, and the specific surface area is ≥120m² / g; Bentonite interlayer spacing ≥ 2.0nm, S2O3²⁻ intercalation amount ≥ 12wt%; The composite microbial agent includes a fluorescent Pseudomonas agent and a composite Bacillus agent, the ratio of live bacteria counts of Pseudomonas to Bacillus is 2.3-2.5:1, and the total live bacteria count is ≥8×10 9 CFU / g; The nutritional synergist is chitosan and nano-cerium oxide modified calcium alginate, the ratio of chitosan to nano-cerium oxide modified calcium alginate is 1:1.2-1:1.5, and the deacetylation degree is ≥95%.

2. A composite microbial enhanced heavy metal passivation organic fertilizer according to claim 1, characterized in that: The fluorescent Pseudomonas agent secretes metallothionein after being induced by Cd²⁺, and the cysteine ​​content thereof is ≥30 mol%; The proportion of β-1,3-glucan in the EPS synthesized by the composite Bacillus inoculum is ≥65%.

3. A composite microbial enhanced heavy metal passivation organic fertilizer according to claim 1, characterized in that: The passivation material is treated with low-temperature plasma hydrogen reduction at a power of 300W for 5 minutes. After the treatment, a LaPO4-CePO4 heterojunction is formed on the surface of the nano-phosphate rock powder, and the bentonite intercalation structure contains a dual sulfur source of S4O6²⁻ / S2O3²⁻, with a saturated adsorption capacity of Cd²⁺ of ≥350mg / g.

4. A composite microbial enhanced heavy metal passivation organic fertilizer according to claim 1, characterized in that: It also includes 0.06-0.08 parts of a bacterial community interaction synergist, wherein the bacterial community interaction synergist includes a C8-AHL quorum sensing molecule, a cyclic diguanylate signaling molecule, and anthraquinone-1,5-disulfonate; Among them, C8-AHL: cyclic diguanylate signal molecule: anthraquinone-1,5-disulfonate = 1:0.8:0.

6.

5. The composite microbial enhanced heavy metal passivation organic fertilizer according to claim 1, characterized in that: A thermo-induced phase-change film-forming agent is added during granulation of the nutrition enhancer, and the thermo-induced phase-change film-forming agent is poly (N-isopropylacrylamide).

6. A method for preparing the composite microbial enhanced heavy metal passivation organic fertilizer according to any one of claims 1 to 5, characterized in that: include: S1. Enhanced bacterial agent function: Pseudomonas fluorescens was cultured in a medium containing 15 mg / L CdCl2 + 0.3 mM glutathione, and the complex Bacillus agent was cultured in a medium containing 5 g / L calcium magnesium phytate + 0.5% chitin oligosaccharide; S2. Biomimetic mineralization loading: Pseudomonas adsorbed on ammonium thiosulfate intercalated bentonite at the silane coupling agent modified interface, and Bacillus loaded on lanthanum phosphate-doped nanophosphate rock powder via Ca²⁺-mediated bridging; S3, Directed Assembly Granulation: The mixed material is extruded with the assistance of a magnetic field with a magnetic field strength of 0.5 T, and a radial gradient pore structure is formed inside the granules with a porosity of 35±1%; S4, staged curing: initial drying at 40℃ to a moisture content of 15%, and final drying at 25℃ to a moisture content ≤10%; S5. Smart coating: The coating material is horseradish peroxidase (HRP) / glucose oxidase (GOx) dual enzyme responsive gel.

7. The method for preparing a composite microbial enhanced heavy metal passivation organic fertilizer according to claim 6, wherein: In step S2, the loading process parameters include silane coupling agent modification concentration of 1.5wt%, Ca²⁺ bridging pH 7.5±0.2, bacterial loading density ≥10 10 CFU / g.

8. The method for preparing a composite microbial enhanced heavy metal passivation organic fertilizer according to claim 6, wherein: In step S5, the enzyme activity of the dual-enzyme response gel is ≥200 U / g, and the GOx activity is ≥150 U / g; The thickness of the dual-enzyme responsive gel coating layer is 80±5 μm.