Biochar-based bacterial fertilizer for improving soil obstacles and preparation method of biochar-based bacterial fertilizer

Abstract

The invention relates to the field of agriculture, and discloses a biochar-based bacterial fertilizer for improving soil obstacles and a preparation method of the biochar-based bacterial fertilizer. The biochar-based bacterial fertilizer comprises the following raw materials: a multifunctional complex microbial inoculant fermentation broth; an organic nutritional carrier; porous biochar; the microbial agent comprises azotobacter chroococcum, bacillus megatherium, bacillus subtilis and rhodopseudomonas palustris. According to the biochar-based bacterial fertilizer, through compounding of the specific multifunctional bacterial agent and scientific combination of the specific multifunctional bacterial agent, the porous biochar and the organic nutrient carrier, synergistic interaction between bacterial strains and between the bacterial agent and the carrier is achieved, and the soil pH and the content of organic matter, alkali-hydrolyzable nitrogen, available phosphorus and available potassium can be remarkably increased at the same time.

Description

Technical Field

[0001] This invention relates to the agricultural field, and more particularly to a biochar-based microbial fertilizer for improving soil obstacles and its preparation method. Background Technology

[0002] Agricultural production has long faced severe challenges due to excessive application of chemical fertilizers, including soil compaction, acidification, decline in organic matter, and nutrient imbalance. Particularly concerning is the fact that large amounts of nitrogen, phosphorus, and potassium in the soil exist in ineffective forms that are difficult for crops to absorb, resulting in extremely low fertilizer utilization and creating a vicious cycle of "worse soil → more fertilizer → worse soil." Therefore, developing new, environmentally friendly fertilizers that can efficiently activate the soil's inherent nutrients and reduce chemical fertilizer input has become an urgent need for sustainable agricultural development.

[0003] To address this challenge, microbial fertilizers utilizing functional microorganisms (such as nitrogen-fixing, phosphorus-solubilizing, and potassium-solubilizing bacteria) have emerged. However, direct application of microbial agents has significant drawbacks: exogenous microorganisms are difficult to colonize in the soil, their activity is easily affected by environmental factors leading to unstable effects, and most products have limited functionality, making it difficult to achieve simultaneous and synergistic enhancement of nutrients.

[0004] To further improve the effectiveness of microbial agents, biochar has been studied as a microbial carrier. Its porous structure can provide a "refuge" for microorganisms, theoretically improving the survival rate and colonization ability of strains (e.g., CN120554177A). However, existing biochar-based microbial fertilizer technologies still have shortcomings: on the one hand, biochar is often only regarded as a physical carrier, lacking targeted design for its synergistic effect with specific functional microbial communities; on the other hand, the strain combinations are usually relatively simple, failing to systematically construct a multifunctional microbial community covering the complete nutrient cycle of "nitrogen fixation, phosphorus solubilization, potassium solubilization, and growth promotion," resulting in limited practical effects.

[0005] In summary, there is an urgent need in this field for an innovative product that not only overcomes the shortcomings of single-agent microbial agents and ordinary biochar carriers, but also achieves synergistic effects through precise matching of materials and microbial communities. However, currently, no publicly available technology effectively addresses the compatibility issue between specific multifunctional compound microbial agents and straw biochar carriers, nor how to maximize the comprehensive effect of simultaneously enhancing key available nutrients such as soil alkaline nitrogen, available phosphorus, and readily available potassium through process optimization. Summary of the Invention

[0006] To address the problems of existing microbial agents in soil colonization difficulties, limited functionality, and unstable effects, as well as the simple functions of existing biochar-based microbial fertilizers and insufficient synergistic effects between carriers and agents, this invention provides a biochar-based microbial fertilizer for improving soil conditions and its preparation method. This biochar-based microbial fertilizer, through the compounding of specific multifunctional microbial agents and their scientific combination with porous biochar and organic nutrient carriers, achieves synergistic effects between strains and between agents and carriers, simultaneously and significantly increasing soil pH, organic matter, available nitrogen, available phosphorus, and available potassium content.

[0007] The specific technical solution of the present invention includes: In a first aspect, the present invention provides a biochar-based microbial fertilizer for improving soil obstacles, comprising the following raw materials: multifunctional compound microbial agent fermentation broth, organic nutrient carrier, and porous biochar.

[0008] The bacteria mentioned include Azotobacter chroococcum, Bacillus megaterium, Bacillus subtilis, and Rhodopseudomonas palustris.

[0009] Existing biochar-based microbial fertilizers often treat biochar merely as a physical carrier, lacking targeted design for synergistic effects with specific functional microbial communities and often featuring relatively simple strain combinations. Firstly, this invention combines a multifunctional compound microbial agent fermentation broth, an organic nutrient carrier, and porous biochar. The organic nutrient carrier provides initial nutrients for the microorganisms, facilitating their smooth transition to the soil environment, while the porous structure of the biochar provides a long-term, stable habitat and sanctuary for the microorganisms, significantly improving the survival rate and colonization success rate of exogenous microbial communities. Synergistic effects are achieved through precise matching of the carrier material and the microbial community, thus solving the compatibility problem between the multifunctional compound microbial agent fermentation broth and the biochar carrier. Secondly, the strains in the multifunctional compound microbial agent fermentation broth of this invention perform functions such as nitrogen fixation, phosphorus solubilization, organic matter decomposition, and photosynthesis promotion, forming complementary microbial communities and constructing a complete "nitrogen fixation-phosphorus solubilization-potassium decomposition" microbial cycle system, which can simultaneously act on multiple nutrients such as nitrogen, phosphorus, and potassium in the soil.

[0010] Furthermore, the reason this invention specifically selects *Azotobacter chrysophagus*, *Bacillus megaterium*, *Bacillus subtilis*, and *Rhodopseudomonas palustris* for combination is based on their unique functional synergy and niche complementarity, rather than a simple functional superposition. Specifically, this combination can construct a complete micro-ecological cycle: *Azotobacter chrysophagus* is responsible for nitrogen fixation, *Bacillus megaterium* specializes in phosphorus solubilization, *Bacillus subtilis* provides the organic acids needed for phosphorus solubilization and releases potassium by decomposing organic matter, while *Rhodopseudomonas palustris*, as a key enabler, creates a favorable microenvironment and secretes growth-promoting substances through its unique photosynthetic metabolism that does not produce oxygen, thus nourishing the entire microbial community. Crucially, these four strains exhibit niche separation in terms of nutritional needs and habitat preferences, resulting in minimal competition among them. Instead, they establish a mutually beneficial symbiotic chain of "nitrogen fixation-phosphorus solubilization-decomposition-growth promotion," ensuring stable colonization and synergistic effects of the microbial community in the soil and biochar carrier, ultimately achieving a simultaneous and significant increase in the content of available nitrogen, available phosphorus, and available potassium in the soil.

[0011] Preferably, the viable bacteria concentration of the multifunctional compound microbial agent fermentation broth is not less than 1×10⁻⁶. 8 CFU / mL.

[0012] This concentration limit is the primary prerequisite for ensuring a sufficient number of functional microorganisms in the final product. The concentration of the fermentation broth, as the "inoculum source," directly determines the initial microbial quantity after inoculation onto the organic carrier.

[0013] Preferably, the ratio of the organic nutrient carrier to the multifunctional compound microbial agent fermentation broth is (90-110) kg: 1 L.

[0014] The core objective of determining the above ratio is to achieve efficient integration and dynamic balance of nutrient supply between the microbial agent and the carrier. If the ratio is too low, insufficient carrier and excessive microbial solution will result in an excessively high initial inoculation density. This will lead to excessive competition among the microbial community for space, oxygen, and nutrients, inhibiting bacterial activity and affecting their normal growth and proliferation. Conversely, if the ratio is too high, i.e., too much carrier and insufficient microbial solution, the microbial solution cannot adhere evenly to the surface of the carrier particles, creating "blank spots" in some areas due to a lack of microbial strains. This results in uneven distribution of the microbial community in the final product, affecting the overall performance. Therefore, this invention controls the ratio within the range of (90-110) kg:1 L, which is the empirically proven optimal ratio range determined based on the specific carrier properties of this invention to achieve high microbial activity, uniform distribution, and good production cost control. This ratio helps to achieve ideal colonization of the microbial strain on the carrier, creating a good initial growth environment and effectively avoiding problems such as internal competition or uneven coverage caused by improper inoculation density.

[0015] Preferably, the total amount of the organic nutrient carrier and the multifunctional compound bacterial agent fermentation broth is in a mass ratio of (0.8-1.2):1 with the porous biochar.

[0016] The aforementioned ratio is crucial for achieving a synergistic effect between "nutrient supply" and "habitat space" for the microorganisms. When the ratio is too low, insufficient organic fertilizer leads to inadequate initial nutrition for the functional microorganisms. Although they can colonize the biochar pores, their proliferation is slow, making it difficult to form a dominant microbial community, resulting in slow fertilizer effect and poor efficacy. Conversely, when the ratio is too high, excessive organic fertilizer can clog the biochar pores, reducing its specific surface area and adsorption capacity, weakening its function as a microbial protective carrier, causing the microbial community to remain in the organic matter, reducing its ability to migrate and colonize the soil, and affecting the improvement effect. Therefore, this invention controls the ratio within the range of (0.8-1.2):1, providing sufficient nutrition for the microbial community in the initial stage, while simultaneously utilizing biochar to construct a protective microenvironment, promoting the gradual diffusion of microorganisms into the soil, thus achieving an organic combination of short-term effects and long-term maintenance.

[0017] Preferably, the number of live bacteria in the biochar-based microbial fertilizer is 6.0 × 10⁻⁶. 7 CFU / g or higher.

[0018] Preferably, the organic nutrient carrier comprises fermented and decomposed sheep manure, fermented and decomposed chicken manure, and rice husks in a mass ratio of (1.5-2.5):(0.8-1.2):1.

[0019] The aforementioned raw materials can provide initial nutrition for functional bacteria and facilitate the survival of the inoculant in the soil. This invention preferably controls the mass ratio of fermented and decomposed sheep manure, chicken manure, and rice husks in the organic nutrient carrier within the range of (1.5–2.5):(0.8–1.2):1. This ratio comprehensively considers the functional synergy of each component: sheep manure has a high organic matter content and provides sustained nutrient release, playing a fundamental supporting role; chicken manure is rich in readily available nutrients such as nitrogen and phosphorus, providing rapid nutrition for microbial growth; rice husks mainly play a physical regulating role, and their loose structure effectively improves the porosity of the compost pile, ensuring aeration. If the proportion of rice husks is too low and the proportion of manure is too high, it easily leads to the carrier becoming sticky and compacted, with insufficient porosity, creating an anaerobic environment during fermentation, affecting the penetration of the inoculum and the respiration of the microbial community; if the proportion of rice husks is too high, it will excessively dilute the nutrient content, leading to an imbalance in the carbon-nitrogen ratio and reducing the initial proliferation efficiency of microorganisms. Therefore, this optimal ratio achieves a balance between nutrient supply and physical structure, taking into account both the fertilizer retention and air permeability of the carrier, and creating a suitable microenvironment for the survival and colonization of functional microorganisms.

[0020] Preferably, the porous biochar is porous straw biochar, which is a porous material made from rice straw through high-temperature pyrolysis under limited oxygen conditions.

[0021] Secondly, the present invention provides a method for preparing the biochar-based bacterial fertilizer, which includes the following steps: S1: Liquid fermentation culture was carried out on Azotobacter chrysophyte, Bacillus megaterium, Bacillus subtilis and Rhodopseudomonas palustris to obtain single-strain fermentation broth.

[0022] S2: Mix the four single-strain fermentation broths to obtain a multifunctional compound microbial agent fermentation broth.

[0023] S3: Ferment and compost the raw materials of the organic nutrient carrier.

[0024] S4: Mix the multifunctional compound microbial agent fermentation liquid with an organic nutrient carrier to produce an organic microbial fertilizer rich in functional bacteria.

[0025] S5: Mix organic microbial fertilizer with porous biochar to obtain biochar-based microbial fertilizer.

[0026] Preferably, in S1, the viable cell concentration in each of the single-strain fermentation broths is 1×10⁻⁶. 8 CFU / mL or higher.

[0027] Preferably, in S2, the volume ratio of the four single-strain fermentation broths is (0.8-1.2):(0.8-1.2):(0.8-1.2):(0.8-1.2).

[0028] Thirdly, the present invention provides the application of the above-mentioned biochar-based microbial fertilizer in simultaneously increasing soil pH, organic matter, alkaline nitrogen, available phosphorus and available potassium content.

[0029] Compared with the prior art, the beneficial effects of the present invention are: (1) Functional synergy: This invention constructs a complete “nitrogen fixation-phosphorus solubilization-potassium decomposition” microbial cycle system by combining four complementary strains, which can simultaneously act on multiple nutrients such as nitrogen, phosphorus and potassium in the soil, overcoming the limitations of single-function microbial agents.

[0030] (2) Highly efficient colonization: This invention innovatively adopts a dual carrier system of "organic nutrient carrier + straw biochar". The organic carrier provides initial nutrition for the microbial agent, helping it to smoothly transition to the soil environment; the porous structure of biochar provides a long-term stable habitat and protection for microorganisms, significantly improving the survival rate and colonization success rate of exogenous microbial communities.

[0031] (3) Significant and stable effects: The carbon-based microbial fertilizer of the present invention showed a simultaneous and significant increase in the content of soil alkaline nitrogen, available phosphorus and available potassium in pot experiments. The effect was better than the application of microbial fertilizer or biochar alone, which proved its synergistic effect. The field application effect was stable and reliable.

[0032] (4) Environmentally friendly and resource-efficient: The biochar-based microbial fertilizer of this invention uses agricultural waste straw as the main raw material, realizing the high-value utilization of waste resources, which is in line with the development direction of green agriculture. Detailed Implementation

[0033] The present invention will be further described below with reference to embodiments.

[0034] General Implementation Examples In the first aspect, a biochar-based microbial fertilizer for improving soil obstacles comprises the following raw materials: multifunctional compound microbial agent fermentation broth, organic nutrient carrier, and porous biochar.

[0035] The bacteria mentioned include Azotobacter chroococcum, Bacillus megaterium, Bacillus subtilis, and Rhodopseudomonas palustris.

[0036] In some preferred embodiments, the viable bacteria concentration of the multifunctional compound microbial agent fermentation broth is not less than 1×10⁻⁶. 8 CFU / mL.

[0037] In some preferred embodiments, the ratio of the organic nutrient carrier to the multifunctional compound microbial agent fermentation broth is (90-110) kg: 1 L.

[0038] In some preferred embodiments, the total amount of the organic nutrient carrier and the multifunctional compound microbial agent fermentation broth is in a mass ratio of (0.8-1.2):1 with the porous biochar.

[0039] In some preferred embodiments, the number of live bacteria in the biochar-based microbial fertilizer is 6.0 × 10⁻⁶. 7 CFU / g or higher.

[0040] In some preferred embodiments, the organic nutrient carrier comprises fermented and decomposed sheep manure, fermented and decomposed chicken manure, and rice husks in a mass ratio of (1.5-2.5):(0.8-1.2):1.

[0041] In some preferred embodiments, the porous biochar is porous straw biochar.

[0042] Secondly, a method for preparing the biochar-based bacterial fertilizer includes the following steps: S1: Liquid fermentation culture was carried out on Azotobacter chrysophyte, Bacillus megaterium, Bacillus subtilis and Rhodopseudomonas palustris to obtain single-strain fermentation broth.

[0043] In some preferred embodiments, in S1, the viable cell concentration in each of the single-strain fermentation broths is 1×10⁻⁶. 8 CFU / mL or higher.

[0044] S2: Mix the four single-strain fermentation broths to obtain a multifunctional compound microbial agent fermentation broth.

[0045] In some preferred embodiments, in S2, the volume ratio of the four single-strain fermentation broths is (0.8-1.2):(0.8-1.2):(0.8-1.2):(0.8-1.2).

[0046] S3: Ferment and compost the raw materials of the organic nutrient carrier.

[0047] S4: Mix the multifunctional compound microbial agent fermentation liquid with an organic nutrient carrier to produce an organic microbial fertilizer rich in functional bacteria.

[0048] S5: Mix organic microbial fertilizer with porous biochar to obtain biochar-based microbial fertilizer.

[0049] Thirdly, the application of the aforementioned biochar-based microbial fertilizer in simultaneously increasing soil pH, organic matter, alkaline nitrogen, available phosphorus, and readily available potassium content.

[0050] Specific implementation examples.

[0051] Example 1: Preparation of straw-based biochar fertilizer (1) Preparation of fermentation broth of multifunctional compound microbial agent Strain activation: The test tube strains of Azotobacter chroococcum (purchased from China General Microbiological Culture Collection Center, strain number bio-05484), Bacillus megaterium (purchased from China General Microbiological Culture Collection Center, strain number bio-74912), Bacillus subtilis (purchased from China General Microbiological Culture Collection Center, strain number bio-00028), and Rhodopseudomonas palustris (purchased from China General Microbiological Culture Collection Center, strain number bio-72803), which were preserved at low temperature (4°C), were streaked onto their respective LB solid medium and incubated at 35°C for 48 hours to activate the strains.

[0052] Primary seed culture: Pick single colonies from freshly activated plates and inoculate them into Erlenmeyer flasks (250 mL) containing 100 mL of the corresponding liquid culture medium. Place the flasks in a shaker and culture at 35°C and 180 rpm for 24 hours to obtain the primary seed culture.

[0053] Fermentation scale-up culture: The above primary seed culture was inoculated at a rate of 5% into fermenters or larger Erlenmeyer flasks containing 1 L of liquid culture medium, and the culture was continued under the same conditions for 24 hours. During this period, the bacterial concentration was monitored using the plate count method.

[0054] Fermentation broth mixing: When the concentration of the fermentation broth for each strain stabilizes at ≥1×10⁻⁶ 8 When the concentration of CFU / mL reaches a certain level, fermentation is stopped. The fermentation broths of the four strains are mixed under aseptic conditions in equal volume ratios (i.e., each strain accounts for 25% of the total volume) to obtain a compound bacterial agent fermentation broth, which is then stored in a 4°C refrigerator for no more than 7 days.

[0055] (2) Preparation of organic nutrient carriers Raw material proportioning and pretreatment: Sheep manure, chicken manure, and rice husks were weighed and mixed at a mass ratio of 2:1:1. The rice husks were mainly used to adjust the porosity of the compost pile.

[0056] Fermentation and composting: Add an appropriate amount of water to the mixture to adjust the moisture content to 55% (ideally, it should clump together when squeezed in the hand but crumble easily when dropped). Pile the material into a windrow-like heap approximately 1.2 meters high for aerobic fermentation. In the initial stage of fermentation, the heap temperature rapidly rises to 65°C, maintaining this high temperature for 7 days to kill pathogens and weed seeds. During this period, turn the heap twice a week to provide oxygen, dissipate heat, and ensure uniform fermentation.

[0057] Determination of the end point of composting: When the temperature of the compost pile drops to basically the same as the ambient temperature, and the material is dark brown, loose, and odorless, it is judged to be fully composted. Allow the composted organic fertilizer to air dry naturally, then crush it through a 2mm sieve to obtain a homogeneous organic nutrient carrier. Seal and store it in a cool, dry place.

[0058] (3) Preparation of straw biochar Raw material processing: Cut the dry and clean rice straw into small pieces of 2-3 cm using a shredder.

[0059] Oxygen-limited pyrolysis: The treated straw is placed in a continuous carbonization furnace. Under the condition of creating an oxygen-limited environment by introducing nitrogen, the temperature is increased to 500°C at a rate of 10°C / minute, and calcined at this final temperature for 2 hours to ensure that the straw is fully carbonized.

[0060] Post-processing: After pyrolysis, the furnace body is allowed to cool naturally to room temperature under continuous nitrogen gas circulation. The generated biochar is then removed, pulverized using a pulverizer, and passed through a 20-mesh sieve to obtain a finished straw biochar product with uniform particles and rich pore structure, which is then sealed and stored.

[0061] (4) Compound preparation of biochar-based microbial fertilizer Inoculation with bacterial culture: Accurately weigh 10 kg of the organic nutrient carrier prepared in step (2) and place it in a clean, sterile mixing device. Using a quantitative spraying device, spray 100 mL of the compound bacterial agent fermentation broth prepared in step (1) (live bacteria concentration ≥ 1×10⁻⁶). 8 The bacterial solution (CFU / mL) was evenly sprayed onto the organic nutrient carrier. During this process, a mechanical stirrer was used to continuously stir at low speed to ensure full and uniform contact between the bacterial solution and the carrier.

[0062] Charcoal-bacterial mixing: Add 10 kg of straw biochar prepared in step (3) to the organic fertilizer that has been inoculated with bacterial solution. Continue to stir and mix thoroughly for 30 minutes until the material is uniform in color and there are no visible lumps.

[0063] Product quality testing: Random samples were taken, and viable bacteria were counted using the plate dilution method. After incubation on agar plates at 35°C for 48 hours, the total viable bacteria count in the final product was found to be 4.3 × 10⁻⁶. 7 CFU / g up to 8.5×10 7 The concentration of CFU / g is within the range required for product quality. Thus, the straw-based bio-fertilizer that simultaneously enhances key soil nutrients, as described in this invention, is obtained.

[0064] Example 2: Verification of the synergistic effect of straw-based bio-fertilizer on soil fertility and rice growth This embodiment verifies the significant effect of the straw-based biochar fertilizer prepared in Example 1 on simultaneously improving key soil nutrients and rice growth through an indoor pot experiment. The experimental soil was taken from a typical barren plot after "non-grain conversion" remediation, with each pot containing 5 kg of soil. Four treatment groups were set up: a blank control group (CK), with no added materials; an organic biochar fertilizer group (BF), with 100 g of the organic biochar fertilizer prepared in Example 1 (i.e., without biochar); a biochar group (BC), with 100 g of the straw biochar prepared in Example 1; and a biochar fertilizer group (BMF), with 100 g of the complete biochar fertilizer product prepared in Example 1. Healthy rice seedlings with uniform growth were selected, and 4 seedlings were transplanted into each pot. The rice variety used was "Yongyou 15". Each treatment had 3 replicates. All potted plants were uniformly fertilized with inorganic fertilizer as base fertilizer at a dosage of 150 mg N / kg. -1 30 mg Pkg -1 and 75.5 mg K kg -1 After 60 days of cultivation in pots under room temperature and natural light, rice was harvested, and plant height and biomass were measured. Root length and root surface area were also measured using a root scanner. Soil samples were collected simultaneously to determine key fertility indicators.

[0065] Fertility index analysis of soil samples showed that the carbon-based microbial fertilizer of the present invention has excellent improvement effect.

[0066] Table 1: Effects of each treatment on soil fertility indicators

[0067] As shown in Table 1, compared with the blank control group (CK), the application of organic microbial fertilizer (BF) or biochar (BC) alone improved soil nutrients to varying degrees. However, the biochar-based microbial fertilizer group (BMF) showed the best effect in all indicators, especially the simultaneous improvement in available nitrogen, available phosphorus, and available potassium. Specifically, the soil organic matter, cation exchange capacity, available nitrogen, available phosphorus, and available potassium content in the BMF treatment were significantly higher than those in the other groups.

[0068] Analysis of rice growth indicators showed that the carbon-based microbial fertilizer of the present invention has a significant promoting effect on rice growth.

[0069] Table 2: Effects of each treatment on rice growth indicators

[0070] As shown in Table 2, compared with the blank control group (CK), the application of organic microbial fertilizer (BF) or biochar (BC) alone improved rice growth to varying degrees, but the biochar-based microbial fertilizer group (BMF) had the best effect. The rice plant height, aboveground dry weight, root length and root surface area of ​​the biochar-based microbial fertilizer (BMF) treatment increased by 32.1%, 28.5%, 37.9% and 36.6% respectively compared with the blank control group.

[0071] The above experimental results fully demonstrate that the straw-based microbial fertilizer prepared by this invention is not a simple superposition of its components (functional microbial agents and biochar), but rather produces a significant synergistic effect. This product can effectively and simultaneously increase the content of multiple key available nutrients such as nitrogen, phosphorus, and potassium in the soil, while improving soil pH and organic matter levels, showing great application potential in cultivating healthy and fertile soil.

[0072] Example 3 (1) Preparation of fermentation broth of multifunctional compound microbial agent Strain activation: The test tube strains of Azotobacter chroococcum (purchased from China General Microbiological Culture Collection Center, strain number bio-05484), Bacillus megaterium (purchased from China General Microbiological Culture Collection Center, strain number bio-74912), Bacillus subtilis (purchased from China General Microbiological Culture Collection Center, strain number bio-00028), and Rhodopseudomonas palustris (purchased from China General Microbiological Culture Collection Center, strain number bio-72803), which were preserved at low temperature (4°C), were streaked onto their respective LB solid medium and incubated at 35°C for 48 hours to activate the strains.

[0073] Primary seed culture: Pick single colonies from freshly activated plates and inoculate them into Erlenmeyer flasks (250 mL) containing 100 mL of the corresponding liquid culture medium. Place the flasks in a shaker and culture at 35°C and 180 rpm for 24 hours to obtain the primary seed culture.

[0074] Fermentation scale-up culture: The above primary seed culture was inoculated at a rate of 5% into fermenters or larger Erlenmeyer flasks containing 1 L of liquid culture medium, and the culture was continued under the same conditions for 24 hours. During this period, the bacterial concentration was monitored using the plate count method.

[0075] Fermentation broth mixing: When the concentration of the fermentation broth for each strain stabilizes at ≥1×10⁻⁶ 8 When the concentration of CFU / mL reaches a certain level, fermentation is stopped. The fermentation broths of the four strains are aseptically mixed in an equal volume ratio of *Azotobacter chrysophagus*: *Bacillus megaterium*: *Bacillus subtilis*: *Rhodopseudomonas palustris* = 1:0.8:0.8:0.8 to obtain a compound bacterial fermentation broth. This broth is stored at 4°C for no more than 7 days.

[0076] (2) Preparation of organic nutrient carriers Raw material proportioning and pretreatment: Sheep manure, chicken manure, and rice husks were weighed and mixed at a mass ratio of 2.5:1:1. The rice husks were mainly used to adjust the porosity of the compost pile.

[0077] Fermentation and composting: Add an appropriate amount of water to the mixture to adjust the moisture content to 55% (ideally, it should clump together when squeezed in the hand but crumble easily when dropped). Pile the material into a windrow-like heap approximately 1.2 meters high for aerobic fermentation. In the initial stage of fermentation, the heap temperature rapidly rises to 65°C, maintaining this high temperature for 7 days to kill pathogens and weed seeds. During this period, turn the heap twice a week to provide oxygen, dissipate heat, and ensure uniform fermentation.

[0078] Determination of the end point of composting: When the temperature of the compost pile drops to basically the same as the ambient temperature, and the material is dark brown, loose, and odorless, it is judged to be fully composted. Allow the composted organic fertilizer to air dry naturally, then crush it through a 2mm sieve to obtain a homogeneous organic nutrient carrier. Seal and store it in a cool, dry place.

[0079] (3) Preparation of straw biochar Raw material processing: Cut the dry and clean rice straw into small pieces of 2-3 cm using a shredder.

[0080] Oxygen-limited pyrolysis: The treated straw is placed in a continuous carbonization furnace. Under the condition of creating an oxygen-limited environment by introducing nitrogen, the temperature is programmed to rise to 550°C at a heating rate of 12°C / min, and calcined at this final temperature for 1.5 hours to ensure that the straw is fully carbonized.

[0081] Post-processing: After pyrolysis, the furnace body is allowed to cool naturally to room temperature under continuous nitrogen gas circulation. The generated biochar is then removed, pulverized using a pulverizer, and passed through a 20-mesh sieve to obtain a finished straw biochar product with uniform particles and rich pore structure, which is then sealed and stored.

[0082] (4) Compound preparation of biochar-based microbial fertilizer Inoculation with bacterial culture: Accurately weigh 10 kg of the organic nutrient carrier prepared in step (2) and place it in a clean, sterile mixing device. Using a quantitative spraying device, spray 100 mL of the compound bacterial agent fermentation broth prepared in step (1) (live bacteria concentration ≥ 1×10⁻⁶). 8 The bacterial solution (CFU / mL) was evenly sprayed onto the organic nutrient carrier. During this process, a mechanical stirrer was used to continuously stir at low speed to ensure full and uniform contact between the bacterial solution and the carrier.

[0083] Charcoal-bacterial mixing: Add 10 kg of straw biochar prepared in step (3) to the organic fertilizer that has been inoculated with bacterial solution. Continue to stir and mix thoroughly for 30 minutes until the material is uniform in color and there are no visible lumps.

[0084] Example 4 (1) Preparation of fermentation broth of multifunctional compound microbial agent Strain activation: The test tube strains of Azotobacter chroococcum (purchased from China General Microbiological Culture Collection Center, strain number bio-05484), Bacillus megaterium (purchased from China General Microbiological Culture Collection Center, strain number bio-74912), Bacillus subtilis (purchased from China General Microbiological Culture Collection Center, strain number bio-00028), and Rhodopseudomonas palustris (purchased from China General Microbiological Culture Collection Center, strain number bio-72803), which were preserved at low temperature (4°C), were streaked onto their respective LB solid medium and incubated at 35°C for 48 hours to activate the strains.

[0085] Primary seed culture: Pick single colonies from freshly activated plates and inoculate them into Erlenmeyer flasks (250 mL) containing 100 mL of the corresponding liquid culture medium. Place the flasks in a shaker and culture at 35°C and 180 rpm for 24 hours to obtain the primary seed culture.

[0086] Fermentation scale-up culture: The above primary seed culture was inoculated at a rate of 5% into fermenters or larger Erlenmeyer flasks containing 1 L of liquid culture medium, and the culture was continued under the same conditions for 24 hours. During this period, the bacterial concentration was monitored using the plate count method.

[0087] Fermentation broth mixing: When the concentration of the fermentation broth for each strain stabilizes at ≥1×10⁻⁶ 8 When the concentration of CFU / mL reaches a certain level, fermentation is stopped. The fermentation broths of the four strains are aseptically mixed in an equal volume ratio of *Azotobacter chrysophagus*: *Bacillus megaterium*: *Bacillus subtilis*: *Rhodopseudomonas palustris* = 1:1.2:1.2:1.2 to obtain a compound bacterial fermentation broth. This broth is stored at 4°C for no more than 7 days.

[0088] (2) Preparation of organic nutrient carriers Raw material proportioning and pretreatment: Sheep manure, chicken manure, and rice husks were weighed and mixed at a mass ratio of 1.5:1:1. The rice husks were mainly used to adjust the porosity of the compost pile.

[0089] Fermentation and composting: Add an appropriate amount of water to the mixture to adjust the moisture content to 55% (ideally, it should clump together when squeezed in the hand but crumble easily when dropped). Pile the material into a windrow-like heap approximately 1.2 meters high for aerobic fermentation. In the initial stage of fermentation, the heap temperature rapidly rises to 65°C, maintaining this high temperature for 7 days to kill pathogens and weed seeds. During this period, turn the heap twice a week to provide oxygen, dissipate heat, and ensure uniform fermentation.

[0090] Determination of the end point of composting: When the temperature of the compost pile drops to basically the same as the ambient temperature, and the material is dark brown, loose, and odorless, it is judged to be fully composted. Allow the composted organic fertilizer to air dry naturally, then crush it through a 2mm sieve to obtain a homogeneous organic nutrient carrier. Seal and store it in a cool, dry place.

[0091] (3) Preparation of straw biochar Raw material processing: Cut the dry and clean rice straw into small pieces of 2-3 cm using a shredder.

[0092] Oxygen-limited pyrolysis: The treated straw is placed in a continuous carbonization furnace. Under the condition of creating an oxygen-limited environment by introducing nitrogen, the temperature is programmed to rise to 450°C at a heating rate of 8°C / min, and calcined at this final temperature for 3 hours to ensure that the straw is fully carbonized.

[0093] Post-processing: After pyrolysis, the furnace body is allowed to cool naturally to room temperature under continuous nitrogen gas circulation. The generated biochar is then removed, pulverized using a pulverizer, and passed through a 20-mesh sieve to obtain a finished straw biochar product with uniform particles and rich pore structure, which is then sealed and stored.

[0094] (4) Compound preparation of biochar-based microbial fertilizer Inoculation with bacterial culture: Accurately weigh 10 kg of the organic nutrient carrier prepared in step (2) and place it in a clean, sterile mixing device. Using a quantitative spraying device, spray 100 mL of the compound bacterial agent fermentation broth prepared in step (1) (live bacteria concentration ≥ 1×10⁻⁶). 8 The bacterial solution (CFU / mL) was evenly sprayed onto the organic nutrient carrier. During this process, a mechanical stirrer was used to continuously stir at low speed to ensure full and uniform contact between the bacterial solution and the carrier.

[0095] Charcoal-bacterial mixing: Add 10 kg of straw biochar prepared in step (3) to the organic fertilizer that has been inoculated with bacterial solution. Continue to stir and mix thoroughly for 30 minutes until the material is uniform in color and there are no visible lumps.

[0096] Unless otherwise specified, the raw materials and equipment used in this invention are all commonly used in the field; unless otherwise specified, the methods used in this invention are all conventional methods in the field.

[0097] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any way. Any simple modifications, alterations, and equivalent transformations made to the above embodiments based on the technical essence of the present invention shall still fall within the protection scope of the present invention.

Claims

1. A biochar-based microbial fertilizer for improving soil obstacles, characterized in that: Including the following raw materials: Multifunctional compound microbial agent fermentation broth; Organic nutrient carrier; Porous biochar; The bacteria mentioned include Azotobacter chrysophyte, Bacillus megaterium, Bacillus subtilis, and Rhodopseudomonas palustris.

2. The biochar-based bacterial fertilizer according to claim 1, characterized in that: The viable bacteria concentration of the multifunctional compound microbial agent fermentation broth is not less than 1×10⁻⁶. 8 CFU / mL.

3. The biochar-based bacterial fertilizer according to claim 1, characterized in that: The ratio of the organic nutrient carrier to the multifunctional compound microbial agent fermentation broth is (90-110) kg: 1 L; The total amount of the organic nutrient carrier and the multifunctional compound microbial agent fermentation broth, and the mass ratio of the porous biochar, are (0.8-1.2):

1.

4. The biochar-based bacterial fertilizer according to claim 1, 2, or 3, characterized in that: The number of live bacteria in the biochar-based microbial fertilizer is 6.0 × 10⁻⁶. 7 CFU / g or higher.

5. The biochar-based bacterial fertilizer according to claim 1 or 3, characterized in that: The organic nutrient carrier comprises fermented and decomposed sheep manure, fermented and decomposed chicken manure, and rice husks in a mass ratio of (1.5-2.5):(0.8-1.2):

1.

6. The biochar-based bacterial fertilizer according to claim 1 or 3, characterized in that: The porous biochar is porous straw biochar.

7. A method for preparing biochar-based bacterial fertilizer according to any one of claims 1-6, characterized in that: include: S1: Liquid fermentation culture was carried out on Azotobacter chrysophagus, Bacillus megaterium, Bacillus subtilis and Rhodopseudomonas palustris to obtain single-strain fermentation broth; S2: Mix the four single-strain fermentation broths to obtain a multifunctional compound microbial agent fermentation broth; S3: Ferment and compost the raw materials of the organic nutrient carrier; S4: Mix the multifunctional compound microbial agent fermentation liquid with an organic nutrient carrier to make organic microbial fertilizer; S5: Mix organic microbial fertilizer with porous biochar to obtain biochar-based microbial fertilizer.

8. The preparation method according to claim 7, characterized in that: In S1, the viable cell concentration in each of the single-strain fermentation broths is 1×10⁻⁶. 8 CFU / mL or higher.

9. The preparation method according to claim 7, characterized in that: In S2, the volume ratio of the four single-strain fermentation broths is (0.8-1.2):(0.8-1.2):(0.8-1.2):(0.8-1.2).

10. The application of the biochar-based microbial fertilizer according to any one of claims 1-6 or the biochar-based microbial fertilizer obtained by the preparation method according to any one of claims 7-9 in simultaneously increasing soil pH, organic matter, alkaline nitrogen, available phosphorus and available potassium content.

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