Bio-organic fertilizer containing compound enzyme and functional microbial inoculum and preparation method of bio-organic fertilizer

Abstract

The invention belongs to the technical field of fertilizer preparation, and provides a bio-organic fertilizer containing a composite enzyme and a functional bacterial agent and a preparation method thereof. The method comprises the following steps: firstly, carrying out physical pretreatment and enzymatic hydrolysis on the waste poultry feathers to convert the waste poultry feathers into liquid rich in bioactive peptides, and compounding the liquid with chitosan oligosaccharide to prepare chitosan oligosaccharide biological compound liquid; secondly, performing physical and chemical complexing on natural plant extracts and humic acid to form a humic acid complexing synergist; furthermore, porous biochar and clay mineral are adopted as carriers, the complex functional flora is immobilized through a physical granulation technology, a microbial long-acting slow-release system is constructed, and composite microbial particles are obtained; finally, in different stages of fermentation of the organic materials, the prepared products are integrated into the decomposed base stock, and the biological organic fertilizer prepared by the method can efficiently activate soil nutrients through the synergistic effect of all the components, so that the growth of crops is promoted, and the remarkable yield increasing effect is achieved.

Description

Technical Field

[0001] This invention belongs to the field of fertilizer preparation technology, and relates to bio-organic fertilizer containing compound enzymes and functional microbial agents and its preparation method. Background Technology

[0002] With the profound transformation of modern agriculture towards green and sustainable development, bio-organic fertilizer has become a key production material for replacing or reducing the use of chemical fertilizers, improving soil health, and enhancing the quality of agricultural products. Bio-organic fertilizer generally refers to a type of fertilizer that combines the effects of microbial fertilizer and organic fertilizer, made from raw materials such as livestock and poultry manure, crop straw, and organic waste, through microbial fermentation and composting, and then compounded with functional microbial agents.

[0003] However, the nutrient release process of organic fertilizers prepared by traditional fermentation is relatively passive and slow. A large number of key nutrients (such as phosphorus and potassium) still exist in insoluble or organically bound forms, which cannot be absorbed by crop roots in time, thus limiting the growth rate and robustness of plants. Secondly, functional microorganisms cannot continuously and effectively promote crop growth. Mixing functional microbial agents in a simple physical way results in low survival rate and colonization ability of these beneficial microorganisms after entering the complex soil environment, which greatly reduces their functions of providing nutrients to crops, such as phosphorus solubilization, potassium solubilization, and nitrogen fixation. Summary of the Invention

[0004] To address the shortcomings of existing technologies, the present invention aims to provide a bio-organic fertilizer containing compound enzymes and functional microbial agents, and its preparation method. First, waste feathers are subjected to targeted enzymatic hydrolysis to prepare bioactive peptides, which are then compounded with chitosan oligosaccharides. Second, plant extracts rich in natural surfactants are complexed with humic acid for enhanced efficacy. Third, the functional microbial community is physically immobilized and granulated using a biochar-clay mineral composite carrier. Finally, at a specific stage of organic material fermentation, the product obtained above is mixed with a well-rotted substrate to meet the needs of actual production.

[0005] To achieve this objective, the present invention adopts the following technical solution:

[0006] In a first aspect, the present invention provides a method for preparing a bio-organic fertilizer containing compound enzymes and functional microbial agents, the preparation method comprising:

[0007] S1. Waste poultry feathers are mixed with water and subjected to high-pressure steam heat treatment to obtain feather paste. Feather paste is mixed with deionized water, alkaline keratinase is added and enzymatically hydrolyzed. After enzymatic hydrolysis, the mixture is heated to deactivate the enzyme and the feather peptide stock solution is collected. Feather peptide stock solution is mixed with chitosan oligosaccharide to obtain chitosan oligosaccharide biocomposite solution.

[0008] S2, mix and extract the camellia oil cake powder with deionized water to obtain a crude extract, disperse the mineral potassium humate powder in water and add it to the crude extract, mix thoroughly to obtain humic acid complexing synergist.

[0009] S3, bamboo charcoal powder, sodium bentonite powder and composite functional bacterial powder are dry mixed to obtain mixed powder, the mixed powder is fed into a roller extrusion granulator, and sucrose aqueous solution is sprayed at the same time to obtain wet granules and dry them to obtain composite microbial granules.

[0010] S4. Mix well-rotted chicken manure, mushroom spawn, and crushed rice husks. During the turning process, spray humic acid complexing synergist into the pile to obtain the initial fermented pile. Maintain the core temperature of the pile above 55℃ for a total of 7 days. When the pile temperature naturally drops below 50℃, turn the pile and spray with chitosan oligosaccharide bio-composite liquid. Ferment and crush to obtain well-rotted organic fertilizer base material. Mix the well-rotted organic fertilizer base material with compound microbial particles to obtain bio-organic fertilizer containing compound enzymes and functional bacteria.

[0011] Specifically, it includes:

[0012] S1, waste poultry feathers are mixed with water and subjected to high-pressure steam heat treatment to obtain feather pulp. The feather pulp is mixed with deionized water, the pH is adjusted to 8-9, the temperature is raised to the first temperature, alkaline keratinase is added and enzymatic hydrolysis is performed. After enzymatic hydrolysis, the enzyme is deactivated by heating and solid-liquid separation is performed using a plate and frame filter press. Feather peptide stock solution is collected. Feather peptide stock solution is mixed with chitosan oligosaccharide to obtain chitosan oligosaccharide biocomposite solution.

[0013] S2, mix camellia seed cake powder with deionized water, heat to the second temperature and stir to extract, to obtain crude extract, disperse mineral potassium humate powder in water and add to crude extract, mix thoroughly to obtain humic acid complexing synergist;

[0014] S3, bamboo charcoal powder, sodium bentonite powder and composite functional bacterial powder are dry mixed to obtain mixed powder, the mixed powder is fed into a roller extrusion granulator, and sucrose aqueous solution is sprayed at the same time to obtain wet granules and dry them to obtain composite microbial granules.

[0015] S4. Mix well-rotted chicken manure, mushroom spawn, and crushed rice husks. During the turning process, spray humic acid complexing synergist into the pile to obtain the initial fermented pile. Maintain the core temperature of the pile above 55℃ for a cumulative period of 7 days. When the pile temperature naturally drops below 50℃, turn the pile and spray with chitosan oligosaccharide bio-composite liquid. Ferment and crush to obtain well-rotted organic fertilizer base material. Mix the well-rotted organic fertilizer base material with compound microbial particles to obtain bio-organic fertilizer containing compound enzymes and functional bacteria.

[0016] In the high-pressure steam heat treatment stage, saturated steam hydrothermally cleaves the keratin molecular structure, breaking the disulfide and hydrogen bonds that maintain its structure, reducing the crystallinity and structural density of keratin, and increasing the contact area between protein and enzyme in subsequent enzymatic reactions. In the enzymatic hydrolysis stage, alkaline keratinase, a specific proteolytic enzyme, catalyzes the hydrolysis of exposed peptide chains, degrading them into bioactive peptides, oligopeptides, and free amino acids with narrow molecular weight distributions. Finally, the resulting feather peptide stock solution is compounded with chitosan oligosaccharides. Feather peptides are not only a highly efficient organic nitrogen source but also participate in plant growth regulation as signaling molecules; while chitosan oligosaccharides, as an oligosaccharide-like plant defense inducer, can induce systemic acquired resistance by activating jasmonic acid or salicylic acid signaling pathways in plants.

[0017] Tea saponin extracted from camellia oil cake exhibits a typical amphiphilic molecular structure, possessing a hydrophilic glycosyl moiety and a hydrophobic saponin backbone. This structural characteristic enables it to effectively reduce the surface tension of liquids in aqueous solutions and demonstrates excellent wetting and emulsifying properties. Potassium humate, derived from minerals, is a complex, heterogeneous high-molecular-weight electrolyte. Its molecular network is rich in various oxygen-containing functional groups such as carboxyl and phenolic hydroxyl groups, giving it strong ion exchange, adsorption, and chelation capabilities. When tea saponin and potassium humate coexist in the same system, they mainly form supramolecular associations or stable colloidal suspensions through van der Waals forces, hydrogen bonds, and possible hydrophobic interactions. This association does not form new covalent bonds, but rather a physicochemical synergistic effect: tea saponin lowers the surface energy of the fertilizer solution, promoting its penetration and diffusion on the surface of soil aggregates; simultaneously, it may stabilize the colloidal structure of fulvic acid molecules through steric hindrance or charge shielding effects, enhancing its complexation ability for metal ions, thereby improving the effectiveness of fertilizer components.

[0018] Bamboo charcoal, serving as the carrier framework, possesses a micron- to nano-scale porous network structure formed through high-temperature pyrolysis, providing a large specific surface area and offering attachment and colonization sites for microorganisms. Simultaneously, the microporous structure effectively protects microorganisms from adverse factors in the soil environment, such as ultraviolet radiation and extreme pH fluctuations. Sodium-based bentonite, acting as a structural reinforcing agent and binder, utilizes its layered silicate structure to polymerize the components into particles with a certain mechanical strength during granulation through mechanical pressure and the action of a small amount of water molecules. Furthermore, the hydrophilicity and interlayer cation exchange capacity of bentonite give it excellent water retention properties. The composite microbial particles achieve physical encapsulation and controlled release of functional microorganisms, extending their survival period and the effective duration of their biological functions in complex soil ecosystems. The composite bacterial powder includes Bacillus subtilis, Bacillus megaterium, Bacillus mucilaginosus HB-02 strain, Trichoderma harzianum, and Bacillus lichenformis B4 strain. The *Bacillus subtilis* strain is deposited at the China General Microbiological Culture Collection Center (CGMCC), with accession number CGMCC No. 24947. The *Bacillus megaterium* strain is deposited at the CGMCC, with accession number CGMCC No. 17890. The *Bacillus mucilaginosus* HB-02 strain is deposited at the CGMCC, with accession number CGMCC No. 24636. The *Trichoderma harzianum* strain is deposited at the CGMCC, with accession number CGMCC No. 40142. The *Bacillus lichenfformis* B4 strain is deposited at the CGMCC, with accession number CGMCC No. 6677.

[0019] The addition of humic acid complexing synergists during the initial fermentation stage optimizes the initial conditions of the fermentation substrate using its physicochemical properties: surfactants promote the penetration of water into hydrophobic organic materials (such as rice husks), ensuring the homogeneity of the fermentation system; humic acid, as an readily available carbon source and pH buffer, provides a favorable environment for the rapid initiation of the microbial community. During the fermentation stage, high temperatures (>55℃) effectively inactivate pathogenic microorganisms, insect eggs, and weed seeds, completing the harmless treatment of the materials. The chitosan oligosaccharide bio-complex liquid added during the post-ripening period after cooling provides directly usable nitrogen sources and growth factors for mesophilic functional microorganisms, accelerating the degradation and humification of complex polymers such as cellulose, hemicellulose, and lignin.

[0020] As a preferred technical solution of the present invention, in S1, the mass ratio of the discarded poultry feathers to water is (8-12):(18-22), for example, it can be (8, 8.4, 8.8, 9.2, 9.6, 10, 10.4, 10.8, 11.2, 11.6 or 12):(18, 18.4, 18.8, 19.2, 19.6, 20, 20.4, 20.8, 21.2, 21.6 or 22), but it is not limited to the listed values, and other unlisted values ​​within this range are also applicable.

[0021] In some optional embodiments, the high-pressure steam treatment temperature is 120-125°C, for example, 120°C, 120.5°C, 121°C, 121.5°C, 122°C, 122.5°C, 123°C, 123.5°C, 124°C, 124.5°C, or 125°C, and the pressure is 0.1-0.12 MPa, for example, 0.1 MPa, 0.102 MPa, 0.104 MPa, 0.106 MPa, 0.108 MPa, or 0.12 MPa. The pressure is 0.11MPa, 0.112MPa, 0.114MPa, 0.116MPa, 0.118MPa, or 0.12MPa, and the time is 25-35 minutes, for example, 25 minutes, 26 minutes, 27 minutes, 28 minutes, 29 minutes, 30 minutes, 31 minutes, 32 minutes, 33 minutes, 34 minutes, or 35 minutes, but it is not limited to the listed values. Other unlisted values ​​within this range are also applicable.

[0022] In some alternative examples, the mass ratio of the feather sizing, deionized water, and alkaline keratinase is (30-35):(60-70):(0.18-0.22), for example, (30, 30.5, 31, 31.5, 32, 32.5, 33, 33.5, 34, 34.5, or 35):(60, 61, 62, 63, 64, 65, 66, 67, 68, 69, or 70):(0.18, 0.184, 0.188, 0.192, 0.196, 0.2, 0.204, 0.208, 0.212, 0.216, or 0.22), but is not limited to the listed values; other unlisted values ​​within this range are also applicable.

[0023] In some alternative instances, the first temperature is 53-57°C, for example, it can be 53°C, 53.4°C, 53.8°C, 54.2°C, 54.6°C, 55°C, 55.4°C, 55.8°C, 56.2°C, 56.6°C or 57°C, but is not limited to the listed values, other unlisted values ​​within this range are also applicable.

[0024] In some optional examples, the alkaline keratinase has an enzyme activity ≥200,000 U / g.

[0025] In some optional instances, the enzymatic hydrolysis time is 7-9 hours, for example, 7 hours, 7.2 hours, 7.4 hours, 7.6 hours, 7.8 hours, 8 hours, 8.2 hours, 8.4 hours, 8.6 hours, 8.8 hours, or 9 hours, but is not limited to the listed values; other unlisted values ​​within this range are also applicable.

[0026] In some alternative instances, the volume-to-mass ratio of the feather peptide stock solution to chitosan oligosaccharide is 1L:(18-22)g, for example, it can be 1L:(18, 18.4, 18.8, 19.2, 19.6, 20, 20.4, 20.8, 21.2, 21.6 or 22)g, but is not limited to the listed values, other unlisted values ​​within this range are also applicable.

[0027] In some optional instances, the degree of polymerization of the chitosan oligosaccharide is 2-10, for example, it can be 2, 3, 4, 5, 6, 7, 8, 9 or 10, but is not limited to the listed values, other unlisted values ​​within this range are also applicable.

[0028] As a preferred technical solution of the present invention, in S2, the mass ratio of the camellia seed cake powder to deionized water is (8-12):(90-110), for example, it can be (8, 8.4, 8.8, 9.2, 9.6, 10, 10.4, 10.8, 11.2, 11.6 or 12):(90, 92, 94, 96, 98, 100, 102, 104, 106, 108 or 110), but it is not limited to the listed values, and other unlisted values ​​within this range are also applicable.

[0029] In some alternative instances, the second temperature is 75-85°C, for example, it can be 75°C, 76°C, 77°C, 78°C, 79°C, 80°C, 81°C, 82°C, 83°C, 84°C or 85°C, but is not limited to the listed values, other unlisted values ​​within this range are also applicable.

[0030] In some optional instances, the stirring extraction speed is 40-60 rpm, for example, 40 rpm, 42 rpm, 44 rpm, 46 rpm, 48 rpm, 50 rpm, 52 rpm, 54 rpm, 56 rpm, 58 rpm or 60 rpm, but is not limited to the listed values, and other unlisted values ​​within this range are also applicable.

[0031] In some optional instances, the stirring extraction time is 1.5-2.5 h, for example, it can be 1.5 h, 1.6 h, 1.7 h, 1.8 h, 1.9 h, 2.0 h, 2.1 h, 2.2 h, 2.3 h, 2.4 h or 2.5 h, but is not limited to the listed values, other unlisted values ​​within this range are also applicable.

[0032] In some alternative instances, the mass ratio of the mineral-derived potassium humate powder to water is (8-12):(90-110), for example, it can be (8, 8.4, 8.8, 9.2, 9.6, 10, 10.4, 10.8, 11.2, 11.6 or 12):(90, 92, 94, 96, 98, 100, 102, 104, 106, 108 or 110), but is not limited to the listed values, other unlisted values ​​within this range are also applicable.

[0033] In some alternative instances, the mass ratio of the camellia oil cake powder to the mineral-derived potassium humate powder is (8-12):(8-12), for example, it can be (8, 8.4, 8.8, 9.2, 9.6, 10, 10.4, 10.8, 11.2, 11.6 or 12):(8, 8.4, 8.8, 9.2, 9.6, 10, 10.4, 10.8, 11.2, 11.6 or 12), but is not limited to the listed values, other unlisted values ​​within this range are also applicable.

[0034] As a preferred embodiment of the present invention, in S3, the mass ratio of bamboo charcoal powder, sodium bentonite powder, composite functional bacterial powder, and sucrose aqueous solution is (55-65):(25-35):(8-12):(13-17), for example, it can be (55, 56, 57, 58, 59, 60, 61, 62, 63, 64 or 65):(25, 26, 27, 28, 29, 30, 31). (32, 33, 34 or 35): (8, 8.4, 8.8, 9.2, 9.6, 10, 10.4, 10.8, 11.2, 11.6 or 12): (13, 13.4, 13.8, 14.2, 14.6, 15, 15.4, 15.8, 16.2, 16.6 or 17), but not limited to the listed values; other unlisted values ​​within this range also apply.

[0035] In some optional examples, the compound microbial powder comprises Bacillus subtilis, Bacillus megaterium, Bacillus mucilaginosus, Trichoderma harzianum, and Bacillus licheniformis B4 strains in a mass ratio of 6:5:5:3:1.

[0036] In some alternative instances, the sucrose aqueous solution has a mass fraction of 4-6 wt.%, for example, 4 wt.%, 4.2 wt.%, 4.4 wt.%, 4.6 wt.%, 4.8 wt.%, 5 wt.%, 5.2 wt.%, 5.4 wt.%, 5.6 wt.%, 5.8 wt.%, or 6 wt.%, but is not limited to the listed values; other unlisted values ​​within this range are also applicable.

[0037] As a preferred technical solution of the present invention, in S4, the mass ratio of the decomposed chicken manure, mushroom spawn, crushed rice husk, humic acid complexing synergist, and chitosan oligosaccharide biocomposite liquid is (40-50):(25-35):(13-17):(2.5-3.5):(1.8-2.2), for example, it can be (40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50):(25, 26, 27, 28, 29, 30, 31, 32, 33, 34 or 35):(1 3, 13.4, 13.8, 14.2, 14.6, 15, 15.4, 15.8, 16.2, 16.6 or 17): (2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4 or 3.5): (1.8, 1.84, 1.88, 1.92, 1.96, 2.0, 2.04, 2.08, 2.12, 2.16 or 2.2), but not limited to the listed values; other unlisted values ​​within this range also apply.

[0038] In some optional examples, the mass ratio of the decomposed organic fertilizer substrate to the composite microbial particles is 95:5.

[0039] In some optional instances, the fermentation involves turning the pile every 7 days, with a fermentation cycle of 40-45 days, such as 40 days, 40.5 days, 41 days, 41.5 days, 42 days, 42.5 days, 43 days, 43.5 days, 44 days, 44.5 days, or 45 days, but is not limited to the listed values; other unlisted values ​​within this range are also applicable.

[0040] Secondly, the present invention provides a bio-organic fertilizer containing compound enzymes and functional microbial agents prepared by the preparation method described in the first aspect.

[0041] Compared with the prior art, the beneficial effects of the present invention are as follows: (1) The present invention improves the permeability of fertilizer in the soil by preparing humic acid complexing synergist and utilizing the physicochemical properties of natural surfactants. At the same time, the introduction of functional microbial communities continuously transforms the insoluble mineral elements such as phosphorus and potassium in the soil into forms that can be absorbed by plants, thereby improving the comprehensive utilization efficiency of nutrients and effectively solving the problems of slow nutrient release and low efficiency of traditional organic fertilizers; (2) The functional microbial communities are physically immobilized by using biochar-bentonite composite particles, ensuring that beneficial microorganisms can survive stably, release slowly and successfully colonize the rhizosphere of crops after being applied to the soil, thereby sustainably exerting their biological functions such as phosphorus and potassium solubilization and disease inhibition; (3) Biostimulants rich in small molecule peptides and amino acids are prepared by targeted enzymatic hydrolysis of waste feathers and compounded with chitosan oligosaccharides. These substances can be directly absorbed by plants, acting as signal molecules to regulate the balance of endogenous hormones in plants, strongly promoting root development and inducing plants to produce systemic stress resistance. Attached Figure Description

[0042] Figure 1 This is a physical image of the bio-organic fertilizer containing compound enzymes and functional microbial agents provided in Example 1 of the present invention. Detailed Implementation

[0043] The technical solution of the present invention will be described in detail below with reference to specific embodiments. The embodiments described herein are specific implementations of the present invention and are used to illustrate the concept of the present invention; these descriptions are explanatory and exemplary and should not be construed as limiting the implementation of the present invention or the scope of protection of the present invention. In addition to the embodiments described herein, those skilled in the art can also adopt other obvious technical solutions based on the content disclosed in the claims and the specification of this application. These technical solutions include those that make any obvious substitutions and modifications to the embodiments described herein.

[0044] The chemical reagents used in the embodiments and comparative examples of this invention are all commercially available products and have not undergone any further purification treatment.

[0045] Example 1

[0046] This embodiment provides a bio-organic fertilizer containing compound enzymes and functional microbial agents, and its preparation method. The preparation method specifically includes the following steps:

[0047] S1. 10 kg of waste poultry feathers are mixed with 21 kg of water and subjected to high-pressure steam heat treatment at a temperature of 123°C, a pressure of 0.11 MPa, and a time of 30 min to obtain feather slurry. 32 kg of feather slurry is mixed with 68 kg of deionized water, the pH is adjusted to 8.8, the temperature is raised to 56°C, 210 g of alkaline keratinase is added and enzymatic hydrolysis is performed. The alkaline keratinase has an enzyme activity ≥200,000 U / g, and the enzymatic hydrolysis time is 8.5 h. After enzymatic hydrolysis, the enzyme is deactivated by heating and solid-liquid separation is performed using a plate and frame filter press to collect the feather peptide stock solution. 5 L of the feather peptide stock solution is mixed with 105 g of chitosan oligosaccharide. The degree of polymerization of the chitosan oligosaccharide is 2-10 to obtain a chitosan oligosaccharide biocomposite solution.

[0048] S2, 9 kg of camellia seed cake powder is mixed with 105 kg of deionized water, heated to 82°C and stirred for extraction. The stirring speed is 55 rpm and the stirring time is 2.2 h to obtain crude extract. 10 kg of mineral potassium humate powder is dispersed in 100 kg of water and added to the crude extract. After thorough mixing, humic acid complexing synergist is obtained.

[0049] S3, 6.2 kg of bamboo charcoal powder, 3.2 kg of sodium bentonite powder and 9 kg of composite functional bacterial powder are dry-mixed to obtain a mixed powder. The composite bacterial powder includes Bacillus subtilis, Bacillus megaterium, Bacillus mucilaginosus HB-02 strain, Trichoderma harzianum and Bacillus licheniformis B4 strain, in a mass ratio of 6:5:5:3:1. The mixed powder is fed into a roller extrusion granulator, and 16 kg of 5.5 wt.% sucrose aqueous solution is sprayed on simultaneously to obtain wet granules, which are then dried to obtain composite microbial granules.

[0050] S4. Mix 48kg of decomposed chicken manure, 32kg of mushroom spawn, and 16kg of crushed rice husks. During the turning process, spray 3.2kg of humic acid complexing synergist into the pile to obtain the initial fermented pile. Maintain the core temperature of the pile above 55℃ for a cumulative period of 7 days. When the pile temperature naturally drops below 50℃, turn the pile and spray 2.1kg of chitosan oligosaccharide bio-composite liquid. Ferment and crush to obtain decomposed organic fertilizer base material. The fermentation is carried out by turning the pile every 7 days, and the fermentation cycle is 43 days. Mix the decomposed organic fertilizer base material with compound microbial particles to obtain bio-organic fertilizer containing compound enzymes and functional bacteria.

[0051] Figure 1 This is a photograph of the bio-organic fertilizer containing compound enzymes and functional microbial agents provided in this embodiment.

[0052] Example 2

[0053] This embodiment provides a bio-organic fertilizer containing compound enzymes and functional microbial agents, and its preparation method. The preparation method specifically includes the following steps:

[0054] S1, 8 kg of waste poultry feathers are mixed with 18 kg of water and subjected to high-pressure steam heat treatment at a temperature of 120°C, a pressure of 0.1 MPa, and a time of 25 min to obtain feather slurry. 30 kg of feather slurry is mixed with 60 kg of deionized water, the pH is adjusted to 8, the temperature is raised to 53°C, 180 g of alkaline keratinase is added and enzymatic hydrolysis is performed. The alkaline keratinase has an enzyme activity ≥200,000 U / g, and the enzymatic hydrolysis time is 7 h. After enzymatic hydrolysis, the enzyme is deactivated by heating and solid-liquid separation is performed using a plate and frame filter press to collect the feather peptide stock solution. 5 L of the feather peptide stock solution is mixed with 90 g of chitosan oligosaccharide. The degree of polymerization of the chitosan oligosaccharide is 2-10 to obtain a chitosan oligosaccharide biocomposite solution.

[0055] S2, mix 8 kg of camellia seed cake powder with 90 kg of deionized water, heat to 75°C and stir to extract. The stirring speed is 40 rpm and the stirring time is 1.5 h to obtain crude extract. Disperse 8 kg of mineral potassium humate powder in 90 kg of water and add it to the crude extract. After thorough mixing, obtain humic acid complexing synergist.

[0056] S3. 5.5 kg of bamboo charcoal powder, 2.5 kg of sodium bentonite powder, and 8 kg of composite functional bacterial powder are dry-mixed to obtain a mixed powder. The composite bacterial powder includes Bacillus subtilis, Bacillus megaterium, Bacillus mucilaginosus HB-02 strain, Trichoderma harzianum, and Bacillus lichenfformis B4 strain in a mass ratio of 6:5:5:3:1. The mixed powder is fed into a roller extrusion granulator, and 13 kg of 4 wt.% sucrose aqueous solution is sprayed on simultaneously to obtain wet granules, which are then dried to obtain composite microbial granules.

[0057] S4. Mix 40kg of decomposed chicken manure, 25kg of mushroom spawn, and 13kg of crushed rice husks. During the turning process, spray 2.5kg of humic acid complexing synergist into the pile to obtain the initial fermented pile. Maintain the core temperature of the pile above 55℃ for a cumulative period of 7 days. When the pile temperature naturally drops below 50℃, turn the pile and spray 1.8kg of chitosan oligosaccharide bio-composite liquid. Ferment and crush to obtain decomposed organic fertilizer base material. The fermentation is carried out by turning the pile every 7 days, and the fermentation cycle is 40 days. Mix the decomposed organic fertilizer base material with compound microbial particles to obtain bio-organic fertilizer containing compound enzymes and functional bacteria.

[0058] Example 3

[0059] This embodiment provides a bio-organic fertilizer containing compound enzymes and functional microbial agents, and its preparation method. The preparation method specifically includes the following steps:

[0060] S1, 12 kg of waste poultry feathers are mixed with 20 kg of water and subjected to high-pressure steam heat treatment at a temperature of 125°C, a pressure of 0.12 MPa, and a time of 35 min to obtain feather slurry. 35 kg of feather slurry is mixed with 65 kg of deionized water, the pH is adjusted to 8.5, the temperature is raised to 55°C, 200 g of alkaline keratinase is added and enzymatic hydrolysis is performed. The alkaline keratinase has an enzyme activity ≥200,000 U / g, and the enzymatic hydrolysis time is 8 h. After enzymatic hydrolysis, the enzyme is deactivated by heating and solid-liquid separation is performed using a plate and frame filter press to collect the feather peptide stock solution. 5 L of the feather peptide stock solution is mixed with 100 g of chitosan oligosaccharide. The degree of polymerization of the chitosan oligosaccharide is 2-10 to obtain a chitosan oligosaccharide biocomposite solution.

[0061] S2, mix 10kg of camellia seed cake powder with 100kg of deionized water, heat to 80℃ and stir to extract, the stirring speed is 50rpm and the stirring time is 2.0h to obtain crude extract, disperse 12kg of mineral potassium humate powder in 110kg of water and add to crude extract, mix thoroughly to obtain humic acid complexing synergist;

[0062] S3, 6.0 kg of bamboo charcoal powder, 3.0 kg of sodium bentonite powder and 10 kg of compound functional bacterial powder are dry-mixed to obtain a mixed powder. The compound bacterial powder includes Bacillus subtilis, Bacillus megaterium, Bacillus mucilaginosus HB-02 strain, Trichoderma harzianum and Bacillus licheniformis B4 strain, in a mass ratio of 6:5:5:3:1. The mixed powder is fed into a roller extrusion granulator, and 15 kg of 5 wt.% sucrose aqueous solution is sprayed on simultaneously to obtain wet granules, which are then dried to obtain compound microbial granules.

[0063] S4. Mix 45kg of decomposed chicken manure, 30kg of mushroom spawn, and 15kg of crushed rice husks. During the turning process, spray 3.0kg of humic acid complexing synergist into the pile to obtain the initial fermented pile. Maintain the core temperature of the pile above 55℃ for a cumulative period of 7 days. When the pile temperature naturally drops below 50℃, turn the pile and spray 2.0kg of chitosan oligosaccharide bio-composite liquid. Ferment and crush to obtain decomposed organic fertilizer base material. The fermentation is carried out by turning the pile every 7 days, and the fermentation cycle is 42 days. Mix the decomposed organic fertilizer base material with compound microbial particles to obtain bio-organic fertilizer containing compound enzymes and functional bacteria.

[0064] Example 4

[0065] This embodiment provides a bio-organic fertilizer containing compound enzymes and functional microbial agents, and its preparation method. The preparation method specifically includes the following steps:

[0066] S1, 9 kg of waste poultry feathers are mixed with 22 kg of water and subjected to high-pressure steam heat treatment at a temperature of 122°C, a pressure of 0.115 MPa, and a time of 28 min to obtain feather slurry. 33 kg of feather slurry is mixed with 70 kg of deionized water, the pH is adjusted to 9, the temperature is raised to 57°C, 220 g of alkaline keratinase is added and enzymatic hydrolysis is performed. The alkaline keratinase has an enzyme activity ≥200,000 U / g, and the enzymatic hydrolysis time is 9 h. After enzymatic hydrolysis, the enzyme is deactivated by heating and solid-liquid separation is performed using a plate and frame filter press to collect the feather peptide stock solution. 5 L of the feather peptide stock solution is mixed with 110 g of chitosan oligosaccharide. The degree of polymerization of the chitosan oligosaccharide is 2-10 to obtain a chitosan oligosaccharide biocomposite solution.

[0067] S2, 12kg of camellia seed cake powder is mixed with 110kg of deionized water, heated to 85℃ and stirred for extraction. The stirring speed is 60rpm and the stirring time is 2.5h to obtain crude extract. 9kg of mineral potassium humate powder is dispersed in 105kg of water and added to the crude extract. After thorough mixing, humic acid complexing synergist is obtained.

[0068] S3, 6.5 kg of bamboo charcoal powder, 3.5 kg of sodium bentonite powder and 12 kg of composite functional bacterial powder are dry-mixed to obtain a mixed powder. The composite bacterial powder includes Bacillus subtilis, Bacillus megaterium, Bacillus mucilaginosus HB-02 strain, Trichoderma harzianum and Bacillus licheniformis B4 strain, in a mass ratio of 6:5:5:3:1. The mixed powder is fed into a roller extrusion granulator, and 17 kg of 6 wt.% sucrose aqueous solution is sprayed on simultaneously to obtain wet granules, which are then dried to obtain composite microbial granules.

[0069] S4. Mix 50kg of well-rotted chicken manure, 35kg of mushroom spawn, and 17kg of crushed rice husks. During the turning process, spray 3.5kg of humic acid complexing synergist into the pile to obtain the initial fermented pile. Maintain the core temperature of the pile above 55% for a cumulative period of 7 days. When the pile temperature naturally drops below 50%, turn the pile and spray 2.2kg of chitosan oligosaccharide bio-composite liquid. Ferment and crush to obtain well-rotted organic fertilizer base material. The fermentation is carried out by turning the pile every 7 days, and the fermentation cycle is 45 days. Mix the well-rotted organic fertilizer base material with compound microbial particles to obtain bio-organic fertilizer containing compound enzymes and functional bacteria.

[0070] Comparative Example 1

[0071] Use only basic garden soil, without applying any external fertilizers, and only water regularly.

[0072] Comparative Example 2

[0073] Apply conventional nitrogen, phosphorus, and potassium compound fertilizer (N-P2O5-K2O=15-15-15) to simulate conventional agricultural production.

[0074] Comparative Example 3

[0075] The same basic materials as in Example 1 were used, but without the addition of humic acid complexing synergist, chitosan oligosaccharide biocomposite liquid, and composite microbial particles.

[0076] Comparative Example 4

[0077] This comparative example provides a bio-organic fertilizer containing compound enzymes and functional microbial agents and its preparation method. The difference between this example and Example 1 is that the mass of the chitosan oligosaccharide bio-compound liquid in S4 is 0, while other process parameters and operating conditions are exactly the same as in Example 1.

[0078] Comparative Example 5

[0079] This comparative example provides a bio-organic fertilizer containing compound enzymes and functional microbial agents and its preparation method. The difference between this example and Example 1 is that the mass of the humic acid complexing synergist in S4 is 0, while the other process parameters and operating conditions are exactly the same as in Example 1.

[0080] Comparative Example 6

[0081] This comparative example provides a bio-organic fertilizer containing compound enzymes and functional microbial agents and its preparation method. The difference between this example and Example 1 is that the mass of the compound microbial particles in S4 is 0, while the other process parameters and operating conditions are exactly the same as in Example 1.

[0082] Test crop: Tomato, Provence variety. A greenhouse pot experiment was conducted, with 10 groups corresponding to Examples 1-4 and Comparative Examples 1-6, each with 10 replicates, for a total of 100 pots. Plastic pots with a top diameter of 30cm and a height of 25cm were used, each filled with 5.0kg of sieved garden soil. Basal fertilizer application: Comparative Example 1 received no fertilizer; Comparative Example 2 received 10g of compound fertilizer per pot, mixed with the potting soil; Comparative Example 3 received 250g of ordinary organic fertilizer per pot, mixed with the potting soil; and in Examples 1-4, each pot received 250g of bio-organic fertilizer containing compound enzymes and functional microbial agents, mixed with the potting soil. Tomato seedlings with uniform growth (4 leaves and 1 heart stage) were transplanted. Comparative Example 2 received 5.0g of compound fertilizer at the initial flowering and peak fruiting stages, while other groups received no topdressing to assess the long-term effectiveness of the basal fertilizer. All treatment groups maintained completely consistent watering, pruning, trellising, and physical pest and disease control measures. The experimental layout employed a completely randomized block design, with experimental pots randomly arranged on seedbeds within the greenhouse and their positions periodically rotated to eliminate environmental errors such as edge effects and uneven light exposure.

[0083] Plant growth indicators: plant height, measured vertically from the soil surface to the plant's growing point using a steel tape measure; stem diameter, measured with vernier calipers below the first branch of the main stem.

[0084] Yield and quality indicators (fruit maturity period): Number of fruits per plant, recording the total number of tomatoes per plant; Single fruit weight, randomly selecting 5 fruits of uniform maturity and weighing them, calculating the average; Yield per plant, calculating the total weight of tomatoes harvested from each plant. The test results are shown in Table 1.

[0085] Table 1. Test results of plant growth indicators, yield and quality indicators (fruit ripening period) of Examples 1-4 and Comparative Examples 1-6

[0086]

[0087] As shown in Table 1, compared to Example 1, the plant height, stem diameter, number of fruits per plant, weight per fruit, and yield per plant all decreased in Comparative Examples 1-6. In Comparative Example 1, the primary nutrients in the soil were rapidly depleted, leading to a comprehensive nutrient deficiency in the plants, directly causing stunted growth and weak stems. Simultaneously, the lack of key reproductive elements such as phosphorus and potassium failed to support flowering, fruit setting, and fruit development, resulting in extremely low yield indicators. Comparative Example 2 only provided readily available macronutrients, lacking organic matter and micronutrients, causing unbalanced nutrient absorption in the plants. This may have promoted vegetative growth faster than structural growth, resulting in relatively weak stems. The lack of organic matter and beneficial microbial communities for soil improvement reduced the overall efficiency of nutrient utilization. Although Comparative Example 3 provided comprehensive basic nutrients, its nutrient release relied on the slow decomposition by native soil microorganisms, resulting in low mineralization rates and low efficiency in the generation of bioactive substances. Comparative Example 4 lacked chitosan oligosaccharide bio-complex liquid, preventing plants from obtaining exogenous small molecule peptides and chitosan oligosaccharides. These two substances directly promote root development, thus weakening root absorption capacity and reducing the final number of fruits and yield. Comparative Example 5 lacked humic acid complexing synergist, resulting in a lack of synergistic effects between humic acid and natural surfactants in the fertilizer, reducing nutrient mobility and availability in the soil, especially easily fixed phosphorus and micronutrients, thereby affecting yield. Comparative Example 6 lacked complex microbial particles, meaning it lacked the core of functional microbial communities, leading to a deficiency in the continuous biological activation process of insoluble phosphorus and potassium minerals in the soil, reducing the long-term nutrient supply capacity of the fertilizer. Meanwhile, the yield per plant is the actual total harvest weight of all mature tomatoes per plant. The deviation from "number of fruits per plant × weight of single fruit" stems from sampling errors when randomly selecting 5 fruits to calculate the weight of a single fruit, as well as weight differences caused by inconsistent maturity of some fruits.

[0088] The above description is only a specific embodiment of the present invention, but the protection scope of the present invention is not limited thereto. Those skilled in the art should understand that any changes or substitutions that can be easily conceived by those skilled in the art within the technical scope disclosed in the present invention fall within the protection and disclosure scope of the present invention.

Claims

1. A method for preparing bio-organic fertilizer containing compound enzymes and functional microbial agents, characterized in that, The preparation method includes: S1. Waste poultry feathers are mixed with water and subjected to high-pressure steam heat treatment to obtain feather paste. Feather paste is mixed with deionized water, alkaline keratinase is added and enzymatically hydrolyzed. After enzymatic hydrolysis, the mixture is heated to deactivate the enzyme and the feather peptide stock solution is collected. Feather peptide stock solution is mixed with chitosan oligosaccharide to obtain chitosan oligosaccharide biocomposite solution. S2, mix and extract the camellia oil cake powder with deionized water to obtain a crude extract, disperse the mineral potassium humate powder in water and add it to the crude extract, mix thoroughly to obtain humic acid complexing synergist. S3, bamboo charcoal powder, sodium bentonite powder and composite functional bacterial powder are dry mixed to obtain mixed powder, the mixed powder is fed into a roller extrusion granulator, and sucrose aqueous solution is sprayed at the same time to obtain wet granules and dry them to obtain composite microbial granules. S4. Mix well-rotted chicken manure, mushroom spawn, and crushed rice husks. During the turning process, spray humic acid complexing synergist into the pile to obtain the initial fermented pile. Maintain the core temperature of the pile above 55℃ for a total of 7 days. When the pile temperature naturally drops below 50℃, turn the pile and spray with chitosan oligosaccharide bio-composite liquid. Ferment and crush to obtain well-rotted organic fertilizer base material. Mix the well-rotted organic fertilizer base material with compound microbial particles to obtain bio-organic fertilizer containing compound enzymes and functional bacteria.

2. The method for preparing bio-organic fertilizer containing compound enzymes and functional microbial agents according to claim 1, characterized in that, In S1: The mass ratio of the discarded poultry feathers to water is (8-12):(18-22); The mass ratio of the feather paste, deionized water and alkaline keratinase is (30-35):(60-70):(0.18-0.22).

3. The method for preparing bio-organic fertilizer containing compound enzymes and functional microbial agents according to claim 1, characterized in that, In S1: The alkaline keratinase has an enzyme activity ≥200,000 U / g.

4. The method for preparing bio-organic fertilizer containing compound enzymes and functional microbial agents according to claim 1, characterized in that, In S1: The volume-to-mass ratio of the feather peptide stock solution to chitosan oligosaccharide is 1L:(18-22)g; The degree of polymerization of the chitosan oligosaccharide is 2-10.

5. The method for preparing bio-organic fertilizer containing compound enzymes and functional microbial agents according to claim 1, characterized in that, In S2: The mass ratio of the camellia seed cake powder to deionized water is (8-12):(90-110).

6. The method for preparing bio-organic fertilizer containing compound enzymes and functional microbial agents according to claim 1, characterized in that, In S2: The mass ratio of the mineral-derived potassium humate powder to water is (8-12):(90-110); The mass ratio of the camellia oil cake powder to the mineral-derived potassium humate powder is (8-12):(8-12).

7. The method for preparing bio-organic fertilizer containing compound enzymes and functional microbial agents according to claim 1, characterized in that, In S3: The mass ratio of bamboo charcoal powder, sodium bentonite powder, compound functional bacterial powder and sucrose aqueous solution is (55-65):(25-35):(8-12):(13-17).

8. The method for preparing bio-organic fertilizer containing compound enzymes and functional microbial agents according to claim 1, characterized in that, In S3: The compound bacterial powder includes Bacillus subtilis, Bacillus megaterium, Bacilhus mucilaginosus HB-02 strain, Trichoderma harzianum and Bacillus licheniformis B4 strain, in a mass ratio of 6:5:5:3:

1.

9. The method for preparing bio-organic fertilizer containing compound enzymes and functional microbial agents according to claim 1, characterized in that, In S4: The mass ratio of the decomposed chicken manure, mushroom lees, crushed rice husks, humic acid complexing synergist, and chitosan oligosaccharide biocomposite liquid is (40-50):(25-35):(13-17):(2.5-3.5):(1.8-2.2). The mass ratio of the decomposed organic fertilizer base material to the composite microbial particles is 95:

5.

10. The preparation method according to any one of claims 1-9 yields a bio-organic fertilizer containing compound enzymes and functional microbial agents.

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