Microbial-active-substance green adjuvant, preparation method and application thereof

Abstract

The application discloses a kind of microbial- active substance green adjuvant, preparation method and its application, the adjuvant includes the following mass content raw materials: bacillus velezensis bacteria active substance 80% to 85%;Cyanophycin 5% to 10%;The balance is brown alga polyphenol modified halloysite nanotube;Bacillus velezensis bacteria active substance is obtained by fermentation culture of bacillus velezensis;The preservation number of the bacillus velezensis is CGMCC NO.16523.The microbial- active substance green adjuvant provided in the application has the characteristics of environmental protection, safety, good dispersibility, emulsibility, formulation stability and environmental stability, and has a synergistic effect on pesticide formulation products, and is mainly applied to pesticide formulation products and pesticide barrel mixing scenes.

Description

Technical Field

[0001] This application belongs to the field of pesticide technology, specifically relating to a microbial-active substance green adjuvant, its preparation method and its application. Background Technology

[0002] Pesticide adjuvants are a general term for various auxiliary materials used in pesticide formulation and application. Although they belong to the adjuvant category and generally lack direct biological activity, they are indispensable additives in pesticide formulation and application. They can refine pesticide formulations, ensuring that small amounts of pesticide are evenly and efficiently distributed in the field to exert their control effects; they can compensate for pesticide defects and improve operational safety; they can reduce the environmental impact of pesticides, optimize operational performance, and enhance the overall efficacy of existing pesticides. In addition, they can enhance the physical stability of formulations, facilitate the diffusion and penetration of pesticide active ingredients on target crops, enhance efficacy, prolong the duration of action, and improve pesticide utilization.

[0003] my country produces millions of tons of various pesticide formulations annually, requiring approximately half a million tons of pesticide adjuvants. This demonstrates the interconnectedness and mutual promotion between my country's pesticide adjuvant industry and pesticide industry. Furthermore, the technological level of pesticide adjuvants plays a crucial role in improving pesticide formulation quality, reducing costs, and maximizing efficacy.

[0004] However, while chemical adjuvants can enhance pesticide dispersibility and adhesion, they also have significant drawbacks. Chemical adjuvants may contain toxic components, such as alkylphenols, which can directly damage crops and severely pollute soil, air, and water sources. Some recalcitrant adjuvants accumulate in the soil, disrupting the soil's microbial ecological balance and endangering ecosystem security. Emulsified oil-based formulations easily penetrate plant epidermis, increasing the risk of phytotoxicity, such as clogging plant stomata or interfering with normal metabolism. Existing pesticide formulation systems have poor stability and are easily affected by environmental factors, exhibiting phenomena such as stratification, flocculation, and precipitation, which reduce pesticide efficacy and increase the risk of pesticide residues. These problems not only threaten sustainable agricultural development but may also harm human health through the food chain.

[0005] Therefore, there is an urgent need to develop a new type of green pesticide adjuvant that is environmentally friendly, safe, and has good environmental stability to replace traditional chemical adjuvants, thereby effectively solving the above problems and ensuring sustainable agricultural development, the living environment, and human health. Summary of the Invention

[0006] In view of this, this application provides an environmentally friendly, safe, and well-dispersible, emulsifying, formulation-stable, and environmentally stable microbial-active substance green adjuvant, its preparation method, and its application.

[0007] In a first aspect, this application provides a green adjuvant containing microbial active substances, the adjuvant comprising the following raw materials in the indicated mass amounts:

[0008] Bacillus belye contains 80% to 85% active substances; Phycocyanin 5% to 10%; The remainder consists of halloysite nanotubes modified with brown algae polyphenols; The active substance of Bacillus belesi is obtained by fermentation culture of Bacillus belesi. The preservation number of the Bacillus belyssus is CGMCC NO.16523.

[0009] Through the above technical solution, this application provides an environmentally friendly, safe, and well-dispersible, emulsifying, formulation-stable, and environmentally stable microbial-active substance green adjuvant, which can completely replace the application of traditional chemical adjuvants in solid and liquid pesticide formulations. This adjuvant constructs a multi-level synergistic functional system through the scientific compounding of Bacillus belysus active substances (containing live microorganisms and their metabolites), phycocyanin, and halloysite nanotubes modified with brown algal polyphenols, exhibiting balanced hydrophilic and lipophilic properties.

[0010] Bacillus belyssaeus has its own active ingredient as the core active ingredient, combining the advantages of both microorganisms and metabolites. Its biosurfactant components are directionally adsorbed onto the interface through their amphiphilic molecular structure, reducing the interfacial energy of the system. Leveraging its solubilizing and spreading properties, it achieves excellent dispersion, emulsification, and stabilization effects, while also promoting target contact and penetration for enhanced efficacy. Simultaneously, the biosurfactant components optimize the hydrophilic-lipophilic balance of the system, enhancing its affinity and spreading ability for pesticide active ingredients and interfaces of different polarities. When live microorganisms are loaded into halloysite nanotubes, they are in a dormant state, actively releasing and restoring their metabolic activity under suitable conditions, continuously producing functional metabolites, thus achieving long-lasting and intelligent response in pesticide efficacy.

[0011] Phycocyanin, as an auxiliary active component, exhibits amphiphilic characteristics, enabling its directional adsorption at oil-water and solid-liquid interfaces. This reduces interfacial tension and enhances the emulsification, dispersion, and suspension stability of the system. This mechanism provides stability support for both water-based and oil-based pesticide formulations and synergizes with the active substances of Bacillus belyssum at the interfacial level. Phycocyanin reduces interfacial tension and provides a biocompatible interfacial environment, promoting the dispersion and activity retention of Bacillus belyssum's active substances. Its protein interfacial layer co-adsorbs with the surface active components of bacterial metabolites, forming a denser and more stable interfacial film. The complementary hydrophilic and hydrophobic regions optimize the interfacial film structure, jointly enhancing the overall stability, formulation adaptability, and bioactivity expression of the system.

[0012] Halloysite nanotubes modified with brown algae polyphenols serve as the functional carrier and stabilizer in this system. After polyphenol modification, their interfacial affinity and steric hindrance effect are enhanced. Because brown algae polyphenols are rich in polar hydroxyl groups and have hydrophobic aromatic ring structures, they improve the affinity of nanotubes for both hydrophilic and hydrophobic components, enabling efficient loading of phycocyanin and Bacillus belyssum active substances. Their tubular mesoporous structure provides protection and a slow-release space for the microbial active ingredients. Halloysite nanotubes loaded with microbial active substances can form a network interface structure, selectively blocking insect stomata without affecting plant stomata. Simultaneously, the halloysite nanotubes, with their sharp, nanoscale rigid structure, physically pierce the cuticle or intestinal cell membrane of insects, causing dehydration, intestinal collapse, secondary infection, and ultimately death, thereby enhancing the insecticidal effect. These structural and functional integrations enable halloysite nanotubes not only to act as carriers for stability, but also to synergize with microbial active ingredients to enhance the dispersibility, stability, delivery, and efficacy of active substances within pests, thereby strengthening the overall control efficacy of the system.

[0013] In terms of their interactions, the three components form a synergistic network centered on the active substances of Bacillus belyssus: phycocyanin, as an auxiliary active component, provides a suitable hydrophilic-hydrophobic equilibrium interface environment for microorganisms and their metabolites through its amphiphilic structure; the brown algal polyphenol modification layer acts as a smart bridge, enhancing the affinity and loading efficiency of halloysite nanotubes for phycocyanin and active substances through its amphiphilic properties and multiple molecular forces, while optimizing the dispersion stability of the system in polar and non-polar media through the synergistic arrangement of hydrophilic and hydrophobic regions; halloysite nanotubes, as a carrier and protective unit, achieve a rational configuration of hydrophilic and oleophilic microregions through their inner and outer surface structures, ensuring the stable and long-lasting effects of phycocyanin and active substances under different environments, while also expanding the physical control function of the system. The synergistic coupling of the active substances, organic, and inorganic components, through the balance regulation of hydrophilic-hydrophobic structures, the synergy of interfacial energy states, and functional complementarity, constructs a dynamic coupling system with dual interfacial adaptability, achieving efficient adaptation and long-term stability to multiple interfaces in polar and non-polar media. This system can enhance wetting and adhesion on crop surfaces through hydrophilic interaction, and promote penetration and retention across the waxy layer through hydrophobic interaction, thereby improving the delivery efficiency and duration of action of pesticide formulations at complex interfaces, enhancing the integrated pest management effect, and providing an innovative solution for modern agricultural plant protection technology.

[0014] In some embodiments, a microbial-active substance green adjuvant comprises raw materials in the following mass amounts: The active substance of Bacillus belysin contains 80% to 85%; for example, the mass content of the active substance of Bacillus belysin is 80%, 81%, 82%, 83%, 84%, 85%, or any combination thereof; Phycocyanin 5% to 10%; for example, the mass content of phycocyanin is 5%, 6%, 7%, 8%, 9%, 10% or any combination thereof; The remainder consists of halloysite nanotubes modified with brown algae polyphenols.

[0015] In some embodiments, a microbial-active substance green adjuvant comprises raw materials in the following mass amounts: The active substances in Bacillus belye are 81% to 84%; Phycocyanin 6% to 9%; The remainder consists of halloysite nanotubes modified with brown algae polyphenols.

[0016] In some embodiments, the additive comprises raw materials in the following mass amounts: Bacillus belysinus contains 83% active substances; Phycocyanin 8%; The remainder consists of halloysite nanotubes modified with brown algae polyphenols. Based on the above technical solution, this application can further improve the environmental stability of microbial-active green adjuvants.

[0017] In some embodiments, the preparation method of the halloysite nanotubes modified by brown algae polyphenols is as follows: Halloysite nanotubes are placed in a brown algae polyphenol solution, stirred at a mass ratio of 1:10 to 1:20, and dried to obtain the halloysite nanotubes modified by brown algae polyphenols. The brown algae polyphenol solution comprises an aqueous solution of brown algae polyphenols and ethanol at a mass ratio of 1:50 to 1:100. This application employs the above technical solution to uniformly load and stably attach brown algae polyphenols onto the surface of halloysite nanotubes, improving their surface physicochemical properties and enhancing the loading and controlled-release performance of phycocyanin and Bacillus belyssum active substances. Simultaneously, the introduction of brown algae polyphenols enhances the hydrophilicity and dispersibility of the nanotubes, making them easier to disperse in aqueous systems and improving the emulsification efficiency and stability of the adjuvants. Furthermore, brown algae polyphenols possess antioxidant and UV absorption capabilities, synergistically protecting the active ingredients. This solution addresses the problem of low loading rates of the original halloysite nanotubes, improving the functionality, environmental adaptability, and biocompatibility of the adjuvant system, laying the foundation for its widespread application in green pesticide formulations.

[0018] In some embodiments, the halloysite nanotubes have an average particle size of 10 nm to 50 nm. By controlling the average particle size of the halloysite nanotubes within the aforementioned suitable range, this application can effectively improve the adsorption and loading capacity of phycocyanin and Bacillus belysin active substances, while optimizing their dispersibility and stability in aqueous systems.

[0019] In some embodiments, the halloysite nanotubes are modified halloysite nanotubes treated with microbial polysaccharides. By employing the above-described technical solution, the microbial polysaccharides encapsulate the halloysite nanotubes through hydrogen bonds, forming a stable "network-filler" structure. This improves their surface properties and enhances their compatibility and binding force with brown algal polyphenols and active ingredients, thereby increasing the loading efficiency and controlled-release performance of the adjuvant on phycocyanin and Bacillus belyssum active substances. Simultaneously, this modification significantly optimizes the interfacial properties of the halloysite nanotubes in the solid-liquid two-phase system, giving them excellent emulsifying and dispersion stability in the liquid phase and achieving uniform and dense micro-dispersion in the solid phase. This collectively ensures their efficient and homogeneous participation in the formulation process, thereby significantly improving the overall stability, functionality, and comprehensive performance of the final product.

[0020] In some embodiments, the microbial polysaccharide includes xanthan gum.

[0021] In some embodiments, the preparation method of xanthan gum-treated modified halloysite nanotubes includes the following steps: mixing halloysite nanotubes (average particle size of 10 nm to 50 nm) and ethanol at a mass ratio of 1:(15-25), stirring, then adding xanthan gum, the ratio of xanthan gum to halloysite nanotubes being 1:(4-6), stirring, filtering, and washing (washing three times with anhydrous ethanol), drying, to obtain xanthan gum-treated halloysite nanotubes.

[0022] In some embodiments, the method for preparing the Bacillus belye fermentation active substance includes the following steps: S1. Inoculate Bacillus belye into LB medium for the first culture to obtain seed fermentation broth; S2. The seed fermentation broth described in step S1 is inoculated into a fermentation medium for a second culture to obtain a sample of Bacillus belye fermentation broth. S3. Centrifuge the *Bacillus berberis* fermentation broth sample obtained in step S2, remove 50% of the supernatant, and obtain the *Bacillus berberis* active substance. The preparation process provided in this application is simple to operate, has good repeatability, is conducive to large-scale production, and provides a high-quality functional raw material basis for the green application of subsequent additives.

[0023] In some embodiments, the fermentation medium in step S2 comprises the following raw materials in the indicated mass percentages: 2% to 3% corn steep liquor powder, 3.5% to 4.5% sucrose, 1% to 1.5% dextrin, 0.1% to 0.15% dipotassium hydrogen phosphate, 0.2% to 0.25% magnesium sulfate heptahydrate, and purified water to bring the total to 100%. The LB culture medium in step S1 comprises the following raw materials in the following mass percentages: 1% peptone, 0.5% yeast extract, 1% sodium chloride, and purified water to 100%.

[0024] Secondly, this application provides a method for preparing a microbial-active substance green adjuvant, the method comprising the following steps: mixing and stirring phycocyanin, Bacillus belyeis active substance, and halloysite nanotubes modified with brown algal polyphenols to obtain the environmentally friendly microbial-active substance green adjuvant. By adopting the above technical solution, this application achieves efficient coupling of the various functional components of the adjuvant, with a simple process, convenient operation, and suitability for large-scale production.

[0025] Thirdly, this application provides a method for preparing a microbial-active substance green adjuvant, which can be used in pesticide formulations. This application employs the above-mentioned technical solution to use a green and environmentally friendly microbial-active substance green adjuvant with excellent emulsifying properties and stability in pesticide formulations, replacing traditional chemical adjuvants and improving the environmental friendliness and stability of pesticide formulations. This adjuvant has good emulsifying and dispersing abilities, enhancing the adhesion and penetration of pesticide active ingredients on the target surface, improving efficacy, and reducing the risk of pesticide use and residues. Moreover, due to its natural origin, it possesses good biodegradability, is not easily accumulated, reduces harm to non-target organisms and the ecological environment, meets the requirements of green agriculture and sustainable development, and has broad market application prospects.

[0026] The beneficial effects of this application are: The adjuvant described in this application can be effectively applied in the pesticide tank mixing process. In practical operation, when mixing pesticides, this adjuvant can solve the flocculation problem that may occur during mixing, ensuring the quality and stability of the mixed pesticide solution. Furthermore, this adjuvant can increase the specific gravity of the pesticide solution, giving it better physical properties such as dispersibility and atomization. Simultaneously, after adding this adjuvant, the pesticide solution settles faster, not only improving application efficiency but also reducing the possibility of pesticide drift in aerial spraying and misting operations, enhancing application accuracy and safety, and providing strong support for the scientific and efficient use of pesticides. Detailed Implementation

[0027] To make the objectives, technical solutions, and advantages of this application clearer, the following detailed description is provided in conjunction with embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the scope of this application.

[0028] The solution of this application will be described below with reference to the following specific embodiments. Unless otherwise specified, the raw materials used in the following embodiments are all from commercially available products, and the devices or equipment used are all purchased from conventional market sales channels.

[0029] The active substance of Bacillus belye in this application is obtained by fermentation culture of Bacillus belye, and the source information of the strain has been disclosed in the previous invention content. The phycocyanin used in this application is E18 grade phycocyanin from Shaanxi Chenming Biotechnology Co., Ltd.

[0030] The brown algae polyphenols used in this application are selected from Shandong Jiejing Group Co., Ltd.

[0031] The halloysite nanotubes used in this application are 10nm-50nm halloysite nanotubes from Angxing New Carbon Materials Changzhou Co., Ltd.

[0032] The Bacillus velezensis strain in this application has the accession number CGMCC NO.16523, is classified as Bacillus velezensis, was deposited on September 21, 2018, and is deposited at the China General Microbiological Culture Collection Center (CGMCC, address: No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, telephone: 010-64807355).

[0033] Unless otherwise specified, all other materials used in the formulation processing are commercially available. The percentages in the examples refer to the mass content of each component. The actual amount of active ingredient added in the formulation processing is the amount calculated based on the purity of the commercially available active ingredient. The following examples are used to illustrate the present invention but are not intended to limit the scope of the invention.

[0034] Preparation Examples 1 to 5 Preparation Example 1 This preparation example provides a Bacillus belye fermentation active substance, and the preparation method includes the following steps: (1) Bacillus belye was inoculated into seed fermentation medium and fermented at 37°C and 220 r / min for 8 h. (2) The fermented seed broth was inoculated into the fermentation medium. The fermentation medium formula was 3% corn steep liquor powder, 4.5% sucrose, 1% dextrin, 0.1% potassium dihydrogen phosphate, 0.25% magnesium sulfate heptahydrate, and purified water was added to 100%. The fermentation was carried out at 37℃ and 220r / min for 72h to obtain the Bacillus berberis fermentation broth sample. (3) The fermentation broth of Bacillus belye was centrifuged at 8000 rpm for 10 min, and 50% of the supernatant was removed according to the mass ratio to obtain the active substance of Bacillus belye.

[0035] Preparation Example 2 The difference between this preparation example and Preparation Example 1 is that the preparation method of this preparation example includes the following steps: (1) Bacillus belye was inoculated into seed fermentation medium and fermented at 37°C and 220 r / min for 8 h. (2) The fermented seed broth was inoculated into the fermentation medium. The fermentation medium formula was 2% corn steep liquor powder, 3.5% sucrose, 1.5% dextrin, 0.15% potassium dihydrogen phosphate, 0.2% magnesium sulfate heptahydrate, and purified water was added to 100%. The fermentation was carried out at 37℃ and 220r / min for 60h to obtain the Bacillus berberis fermentation broth sample. (3) The fermentation broth of Bacillus belye was centrifuged at 8000 rpm for 10 min, and 50% of the supernatant was removed according to the mass ratio to obtain the active substance of Bacillus belye.

[0036] Preparation Example 3 This preparation example provides halloysite nanotubes modified with brown algae polyphenols. The preparation method of halloysite nanotubes modified with brown algae polyphenols is as follows: Halloysite nanotubes are placed in brown algae polyphenol solution at a mass ratio of 1:10, stirred, and dried to obtain halloysite nanotubes modified with brown algae polyphenols. The brown algae polyphenol solution consists of brown algae polyphenols and 95% ethanol aqueous solution in a mass ratio of 1:50.

[0037] The average particle size of halloysite nanotubes is 10 nm.

[0038] Preparation Example 4 This preparation example provides halloysite nanotubes modified with brown algae polyphenols. The preparation method of halloysite nanotubes modified with brown algae polyphenols is as follows: Halloysite nanotubes are placed in brown algae polyphenol solution at a mass ratio of 1:20, stirred, and dried to obtain halloysite nanotubes modified with brown algae polyphenols. The brown algae polyphenol solution consists of brown algae polyphenols and 95% ethanol aqueous solution in a mass ratio of 1:100.

[0039] The average particle size of halloysite nanotubes is 50 nm.

[0040] Preparation Example 5 This preparation example provides a KR-41B halloysite nanotube modified with brown algae polyphenols. The preparation method includes the following steps: A1. Halloysite nanotubes (average particle size of 10nm to 50nm) with ethanol at a mass ratio of 1:20 were mixed and stirred. Then, KR-41B titanate coupling agent (the mass ratio of KR-41B titanate coupling agent to halloysite nanotubes was 1:5) was added dropwise. The mixture was stirred, filtered, and washed (three times with anhydrous ethanol). After drying, KR-41B modified halloysite nanotubes were obtained. A2. Mix the brown algae polyphenol solution (the mass ratio of brown algae polyphenol to ethanol aqueous solution is 1:100) with the KR-41B modified halloysite nanotubes prepared in step A1 at a mass ratio of 1:20, stir, let stand, filter to collect the solid material, dry, and obtain brown algae polyphenol modified KR-41B halloysite nanotubes.

[0041] Example 1 This embodiment provides a green adjuvant containing microbial active substances. The adjuvant comprises the following raw materials in the indicated mass amounts: Bacillus belysinii active substances: 85%; Phycocyanin 5%; 10% halloysite nanotubes modified with brown algae polyphenols; The active substance of Bacillus belye in this embodiment was prepared in Preparation Example 1.

[0042] The halloysite nanotubes modified with brown algae polyphenols in this embodiment were prepared in Preparation Example 3.

[0043] The preparation method of the microbial-active substance green adjuvant in this embodiment includes the following steps: mixing and stirring phycocyanin, Bacillus belye active substance, and halloysite nanotubes modified with brown algae polyphenols evenly to obtain an environmentally friendly microbial-active substance green adjuvant.

[0044] Example 2 The difference between this embodiment and Embodiment 1 is that the microbial-active green adjuvant in this embodiment includes the following raw materials in the following mass percentages: Bacillus belyi contains 80% active substances; Phycocyanin 10%; 10% Halloysite nanotubes modified with brown algae polyphenols.

[0045] Example 3 The difference between this embodiment and Embodiment 1 is that the microbial-active green adjuvant in this embodiment includes the following raw materials in the following mass percentages: Bacillus belyi contains 83% active substances; Phycocyanin 8%; Halloysite nanotubes modified with brown algae polyphenols 9%.

[0046] Example 4 The difference between this embodiment and Example 3 is that the active substance of Bacillus belye in this embodiment was prepared by Example 2.

[0047] Example 5 The difference between this embodiment and Example 3 is that the halloysite nanotubes modified with brown algae polyphenols in this embodiment were prepared by Example 4.

[0048] Example 6 The difference between this embodiment and Example 3 is that the halloysite nanotubes modified with brown algae polyphenols in this embodiment were prepared by Example 5.

[0049] Comparative Example 1 The difference between this comparative example and Example 3 is that, in preparing the microbial-active substance green adjuvant, the Bacillus beryl fermentation active substance of Preparation Example 1 is used instead of the microbial-active substance green adjuvant prepared in Example 3. That is, the adjuvant of this comparative example contains only the Bacillus beryl fermentation active substance of Preparation Example 1.

[0050] Comparative Example 2 The difference between this comparative example and Example 3 is that in the preparation of the microbial-active substance green adjuvant in this comparative example, phycocyanin is used instead of the microbial-active substance green adjuvant prepared in Example 3. That is, the adjuvant in this comparative example contains only phycocyanin.

[0051] Comparative Example 3 The difference between this comparative example and Example 3 is that in the preparation of the microbial-active substance green adjuvant, brown algae polyphenol-modified halloysite nanotubes are used instead of the microbial-active substance green adjuvant prepared in Example 3. That is, the adjuvant in this comparative example contains only brown algae polyphenol-modified halloysite nanotubes.

[0052] Comparative Example 4 The difference between this comparative example and Example 3 is that this comparative example does not add the microbial-active substance green adjuvant obtained by modifying halloysite nanotubes with brown algae polyphenols when preparing the microbial-active substance green adjuvant. That is, the adjuvant in this comparative example only contains Bacillus belye bacterial active substances and phycocyanin.

[0053] Comparative Example 5 The difference between this comparative example and Example 3 is that this comparative example does not add phycocyanin to the preparation of the microbial-active green adjuvant. That is, the adjuvant in this comparative example only contains Bacillus belyssus active material and halloysite nanotubes modified with brown algal polyphenols.

[0054] Comparative Example 6 The difference between this comparative example and Example 3 is that this comparative example does not add the microbial-active substance green adjuvant prepared from Bacillus belysinian active substances during the preparation of the microbial-active substance green adjuvant. That is, the adjuvant in this comparative example only contains phycocyanin and halloysite nanotubes modified with brown algal polyphenols.

[0055] Comparative Example 7 The difference between this comparative example and Example 3 is that this comparative example uses unmodified halloysite nanotubes instead of the brown algae polyphenol-modified halloysite nanotubes in Example 3 to prepare the microbial-active substance green adjuvant.

[0056] Comparative Example 8 The difference between this comparative example and Example 3 is that, in preparing the microbial-active substance green adjuvant, this comparative example uses Preparation Example 1 + Ethyl Acetate Polyene Fish Oil (3:0.3) (patent adjuvant application number CN202411571494.9) instead of the microbial-active substance green adjuvant prepared in Example 3. That is, in this comparative example, only the Bacillus belyesii active substance from Preparation Example 1 and ethyl ester polyene fish oil are contained, and the mass ratio of Bacillus belyesii active substance to ethyl ester polyene fish oil is 3:0.3.

[0057] Formulation stability validation test 1: For pesticide formulations such as emulsifiable concentrates, water-in-oil emulsions, and microemulsions, the stability of formulations using different adjuvants is validated by emulsion stability, low-temperature precipitation rate test method, heat storage precipitation rate test method, and water-emulsification dispersion state.

[0058] The mineral oil emulsion products with different additives were formulated according to the formula in Table 1, and the results of the stability test are shown in Table 2.

[0059] Emulsion stability: Referring to GB / T 1603-2001 "Test Method for Stability of Pesticide Emulsions", add 99.5 mL of standard hard water to a 250 mL beaker. Using a pipette, add 0.5 mL of pesticide aqueous solution (i.e., the adjuvant in the above examples and comparative examples) slowly to the hard water while continuously stirring, preparing a 100 mL emulsion. After adding the emulsion, continue stirring at 2-3 r / s for 30 s. Immediately transfer the emulsion to a clean, dry 100 mL graduated cylinder and place it in a constant temperature water bath at 30±2℃ for 1 hour. Remove the cylinder and observe the emulsion separation. If no floating oil, settling oil, or precipitate forms in the graduated cylinder...

[0060] Low-temperature precipitation rate test method: Referring to GB / T 19137-2003 "Determination of Low-Temperature Stability of Pesticides", transfer 100 mL of pesticide aqueous solution (i.e., the adjuvant in the above examples and comparative examples) into a centrifuge tube, cool it to 0±2℃ in a refrigerator, and keep the centrifuge tube and its contents at 0±2℃ for 1 hour, stirring once every 15 minutes for 15 seconds each time, checking and recording whether any solid or oily substances precipitate. Return the centrifuge tube to the refrigerator and continue to place it at 0±2℃ for 7 days. After 7 days, remove the centrifuge tube, let it stand at room temperature for 3 hours, and centrifuge for 15 minutes (the relative centrifugal force at the top of the tube is 500-600 g, where g is the acceleration due to gravity). Record the volume of the precipitate at the bottom of the tube (accurate to 0.05 mL). Perform 3 parallel tests for each sample and calculate the precipitation rate. The calculation formula is as follows:

[0061] Test method for heat storage release rate: Referring to GB / T 19136-2003 "Determination of Heat Storage Stability of Pesticides", approximately 30 mL of pesticide aqueous solution (i.e., the adjuvants in the above examples and comparative examples) is injected into a clean ampoule using a syringe. The ampoule is placed in an ice-salt bath for cooling, sealed with a high-temperature flame, and weighed after cooling to room temperature. The sealed ampoule is then placed in a metal container, and the metal container is placed in a constant temperature chamber at 54±2℃ for 14 days. After removal, the outside of the ampoule is wiped clean and weighed. For samples whose mass has not changed, three parallel sets are performed for each sample. The release rate is calculated using the following formula:

[0062] Water-emulsification and dispersion level: The water-emulsification and dispersion state is determined according to the following methods: Level 1: Automatically dispersed in a cloud-like, uniform manner, with no visible oil droplets, appearing as a milky white emulsion with a blue fluorescence; Secondary: Cloudy dispersion, no visible oil droplets, thick milky white emulsion; Level 3: Cloudy dispersion, with virtually no visible oil droplets, appearing as a milky white emulsion; Level 4: Partially self-dispersed, with coarse emulsion particles, appearing as a white emulsion; Level 5: Does not disperse automatically, forms oil droplets when in water, emulsifies when shaken, and quickly separates when left to stand.

[0063] In the table below, letter A refers to the type of conventional adjuvant, letter B represents the microbial-active substance green adjuvant provided in the embodiments of this application, letter C refers to the adjuvant prepared by Comparative Example 1, letter D refers to the adjuvant prepared by Comparative Example 2, letter E refers to the adjuvant prepared by Comparative Example 3, letter F refers to the adjuvant prepared by Comparative Example 4, letter G refers to the adjuvant prepared by Comparative Example 5, letter H refers to the adjuvant prepared by Comparative Example 6, letter I refers to the adjuvant prepared by Comparative Example 7, and letter J refers to the adjuvant prepared by Comparative Example 8.

[0064] Table 1 Processing Table for Mineral Oil Emulsifiable Concentrates with Different Additives

[0065] Table 2. Stability Test Results of Mineral Oil Emulsifiable Oils Processed with Different Additives

[0066] The experimental results in Tables 1 and 2 show that: (1) Advantages compared to single and compound treatments: Mineral oil emulsifiable concentrates using the safe and environmentally friendly microbial-active substance green adjuvants provided in this application (Application Examples 1-6) show significantly better performance than single microbial active substances (Comparative Application Example 2), single phycocyanin (Comparative Application Example 3), halloysite nanotubes modified with single brown algal polyphenols (Comparative Application Example 4), and the compound system of microbial active substances and phycocyanin (Comparative Application Example 5) in terms of emulsion stability, low-temperature precipitation rate, heat storage precipitation rate, and water emulsification and dispersibility. The following systems were compared: a system of halloysite nanotubes modified with bacterial active substances and brown algae polyphenols (Comparative Application Example 6); a system of halloysite nanotubes modified with phycocyanin and brown algae polyphenols (Comparative Application Example 7); and a system of bacterial active substances + phycocyanin + unmodified halloysite nanotubes (Comparative Application Example 8). These results demonstrate that the microbial-active green adjuvant composed of bacterial active substances + phycocyanin + halloysite nanotubes modified with brown algae polyphenols can significantly improve the emulsion stability, low-temperature precipitation rate, heat storage precipitation rate, and water emulsification and dispersibility of mineral oil products.

[0067] (2) Superiority compared to conventional high-quality additives: Mineral oil emulsion products using the safe and environmentally friendly microbial-active green additives provided in this application (Application Example 1-Application Example 6) are significantly superior to conventional high-quality additives in four aspects: emulsion stability, low temperature precipitation rate, heat storage precipitation rate, and water emulsification and dispersion (Comparative Application Example 1).

[0068] (3) Superiority of patented additives: Mineral oil emulsion products using the safe and environmentally friendly microbial-active green additives provided in this application (Application Example 1-Application Example 6) are significantly superior to the patented additives with application number CN202411571494.9 (Comparative Application Example 6) in terms of emulsion stability, low temperature precipitation rate, heat storage precipitation rate and water emulsification dispersion.

[0069] (4) Formulation optimization: In application examples 1-6, each application example was at the same level in terms of emulsion stability, low-temperature precipitation rate, and water-emulsification and dispersion. In terms of heat storage precipitation rate, application example 6 performed the best, followed by application examples 3-5, while application examples 1 and 2 were relatively poor. This indicates that 83% of the active bacterial substance + 8% of phycocyanin is a better formulation. At the same time, treating halloysite nanotubes with KR-41B titanate coupling agent first, and then modifying the halloysite nanotubes with brown algae polyphenols, can further improve the heat storage stability of mineral oil emulsion products.

[0070] (5) Specifically, regarding emulsion stability, the application example product did not exhibit any floating emulsion or sedimentation, while the comparative application examples showed varying degrees of floating emulsion and sedimentation. Regarding low-temperature precipitation rate, the application example's value was significantly lower than the comparative application example, indicating better low-temperature stability. In terms of heat storage precipitation rate, the application example showed a significant advantage with a lower precipitation rate. Regarding water-based emulsion dispersibility, the application example product could disperse rapidly and uniformly in water, while the comparative application example showed poor dispersion. These data fully demonstrate the superiority of the safe and environmentally friendly microbial-active substance green adjuvant of this application. Further analysis shows that the adjuvant performs excellently in multiple indicators, laying the foundation for formulation stability and enabling it to maintain good condition under various environments. The modified halloysite nanotubes enhanced the compatibility of the adjuvant with mineral oil, thereby improving heat storage stability.

[0071] Based on the above test results, the optimal formulation (Example 6) was selected, and other products (such as emulsifiable concentrates, microemulsions, and water emulsions) were formulated according to Table 3. The results of the stability test are shown in Table 4.

[0072] Table 3 Processing Table for Different Additives

[0073] Table 4. Stability Test Table for Products Processed with Different Additives

[0074] As can be seen from the test results in Tables 3 and 4, the different products using the safe and environmentally friendly microbial-active substance green adjuvants provided in this application (Application Examples 7 to 12) are significantly superior to high-quality adjuvants in the conventional market in terms of four indicators: emulsion stability, low-temperature precipitation rate, heat storage precipitation rate, and water emulsification and dispersibility (Comparative Application Examples 6 to 11).

[0075] Specifically, regarding emulsion stability, products using the additives described in this application exhibited no floating emulsion and no sedimentation, while high-quality additives in the conventional market generally showed a small amount of floating emulsion and even a small amount of sedimentation. Regarding low-temperature precipitation rate, the precipitation rate of products using the additives described in this application was significantly lower than that of products using conventional additives, demonstrating superior low-temperature stability. Regarding heat storage precipitation rate, the additives described in this application also performed excellently, with the precipitation rate of the corresponding products being far lower than that of conventional additives. In terms of emulsification and dispersibility in water, products using the additives described in this application also exhibited excellent performance, all meeting the first-class standard, while products using conventional additives showed significant differences.

[0076] Formulation stability verification test 2: For pesticide formulations such as suspension concentrates and dispersible oil suspension concentrates, the stability verification test of formulation products using different adjuvants is carried out by pesticide suspension rate, pesticide pourability, low temperature precipitation rate test method and heat storage precipitation rate test method.

[0077] Different additive products (such as suspending agents and dispersible oil suspensions) were formulated according to the formula in Table 5, and the emulsification performance test results are shown in Table 6. Pesticide suspension rate: Referring to GB / T 14825-2023 "Determination of Pesticide Suspension Rate", add 100 mL of standard hard water to a 250 mL graduated cylinder. Weigh an appropriate amount of sample into a 100 mL beaker and transfer the entire sample to the 250 mL graduated cylinder using standard hard water. Dilute to the 250 mL mark with standard hard water, stopper the cylinder, and invert the cylinder 30 times within 1 minute, using the center of the cylinder as the axis (one inversion is approximately 2 seconds). Open the stopper, place the graduated cylinder vertically on a flat surface, and let it stand for 30 minutes, avoiding vibration and direct sunlight. Use a pipette to remove 9 / 10 (i.e., 225 mL) of the suspension within 10-15 seconds. Do not shake or stir the sediment in the graduated cylinder, and ensure that the tip of the pipette is always a few millimeters below the liquid surface. The mass of the effective components in the 25 mL suspension and precipitate remaining at the bottom of the graduated cylinder was determined according to the method for determining the effective components in the product standard. The formula for calculating the suspension rate of the effective components in the sample is as follows.

[0078]

[0079]

[0080] In the formula: m1 — The numerical value of the effective component in the graduated cylinder, in grams (g). m0 — The numerical value of the mass of the sample in the graduated cylinder, in grams (g). w0—The mass content of the effective component in the sample, expressed as a percentage (%); w1—The suspension rate of the effective component in the sample, expressed as a percentage (%); m2 — The mass of the active ingredient remaining in the 25mL suspension at the bottom of the graduated cylinder, expressed in grams (g). —Conversion factor.

[0081] Pesticide dumping property: Referring to GB-T 31737-2015 "Test Method for Pesticide Dumping Property", two indicators need to be measured: (1) Determination of residue after pouring: Weigh the graduated cylinder with stopper (accurate to 0.1g); add the sample to 80% of the total volume of the graduated cylinder, stopper it, and weigh it again (accurate to 0.1g). After standing at room temperature (the specific temperature can be determined according to actual needs) for 24 hours, first rotate the graduated cylinder 135° from an upright position, pour it for 60s, and then invert it for 60s. Weigh the graduated cylinder with stopper again (accurate to 0.1g). (2) Determination of residue after washing: Add 20℃ distilled water to 80% of the total volume of the graduated cylinder, stopper it, and invert the graduated cylinder 10 times (when inverting the graduated cylinder, ensure that each time the cylinder is rotated 180° from an upright position and returned to its original position, the time should be about 2 seconds, and the operation should be completed smoothly and evenly). Then pour out the water using the same method as when pouring, and then cover it and weigh it (accurate to 0.1g). The calculation formula is as follows.

[0082]

[0083]

[0084] In the formula: w1 — Residue after dumping, expressed as a percentage; w2 — Residue after washing, expressed as a percentage; m1 — The sum of the sample mass and the mass of the stoppered graduated cylinder, in grams (g).

[0085] m2 — The sum of the mass of the residue after pouring and the mass of the stoppered graduated cylinder, in grams (g). m3 — The sum of the mass of the residue after washing and the mass of the stoppered graduated cylinder, in grams (g). m0 — the mass of the stoppered graduated cylinder, in grams (g). Low-temperature stability of pesticides: 80 mL of the sample was placed in a 100 mL beaker and cooled to (0 ± 2) °C in a refrigerator for 1 hour. The sample was stirred for 15 seconds every 15 minutes, and the appearance was observed for any changes. The beaker was then returned to the refrigerator and kept at (0 ± 2) °C for 7 days. After 7 days, the beaker was removed and allowed to return to room temperature. The stratification under low-temperature treatment was recorded.

[0086] Table 5 Processing Table for Different Additives

[0087] Table 6. Stability Test Table for Products Processed with Different Additives

[0088] As can be seen from the test results in Tables 5 and 6, the different products using the microbial-active green adjuvants provided in this application (Application Examples 13-16) are significantly superior to high-quality adjuvants in the conventional market (Comparative Application Examples 13-16) in five aspects: suspension rate, pesticide dumping properties (residue content after dumping and residue content after washing), low-temperature stratification, and heat storage release rate.

[0089] Specifically, in terms of suspension rate, products using the microbial-active substance green adjuvant of this application all showed higher values, indicating that it can better suspend pesticide particles in liquid and improve pesticide application efficiency. Regarding pesticide pourability, both the residue content after pouring and the residue content after washing were significantly lower in products using the adjuvant of this application than in conventional adjuvants, demonstrating superior pourability and less residue. Low-temperature stratification is an important indicator of the stability of adjuvants at low temperatures. The test results show that products using the adjuvant of this application did not exhibit stratification at low temperatures, while conventional adjuvants generally showed stratification problems. Finally, in terms of heat storage release rate, products using the adjuvant of this application also showed a lower release rate, indicating that it maintains good stability even at high temperatures.

[0090] Formulation stability verification test 3: For pesticide formulations such as wettable powders and water-dispersible granules, the pesticide suspension rate, wetting time and heat storage stability test methods are used to verify the stability of formulation products using different adjuvants.

[0091] Different additive products (such as wettable powder and water-dispersible granules) were formulated according to the formula in Table 7, and the results of the stability test are shown in Table 8. Pesticide suspension rate: The suspension rate was determined in accordance with GB / T 14825-2023, "Method for Determination of Pesticide Suspension Rate".

[0092] Wetting Time: Referring to GB / T 5451-2001 "Determination of Wetting Properties of Pesticide Wettable Powders", take 100mL±1mL of standard hard water and pour it into a 250mL beaker. Place this beaker in a constant temperature water bath at 25℃±1℃, ensuring the liquid level is flush with the water bath's surface. When the hard water reaches 25℃±1℃, weigh 5g±0.1g of the sample (the sample should be a representative, uniform powder, and clumps or agglomerates are not allowed), place it on a watch glass, and pour the entire sample evenly onto the liquid surface of the beaker from a position flush with the rim, without excessively disturbing the liquid surface. Immediately start timing with a stopwatch while adding the sample until the sample is completely wetted (the fine powder film remaining on the liquid surface is negligible). Record the wetting time (accurate to the second). Repeat this process 5 times and take the average value as the wetting time of the sample.

[0093] Pesticide thermal storage stability: Place 20g of sample into a beaker without applying any pressure, allowing it to spread into a smooth, uniform layer of equal thickness. Place a disc on top of the sample, and put the beaker in an oven at (54±2)℃ for 14 days. Remove the beaker and disc, and place them in a desiccator to cool the sample to room temperature. Within 24 hours, complete the tests for the specified items such as active ingredient content, pesticide suspension rate, and wetting time. Perform three parallel tests for each sample, and take the average value of the final results. Simultaneously calculate the active ingredient degradation rate, suspension rate reduction rate, and wetting time growth rate using the following formulas.

[0094]

[0095]

[0096]

[0097] Table 7 Processing Table for Different Additives

[0098] Table 8. Stability Test Table for Products Processed with Different Additives

[0099] As shown in Tables 7 and 8, the different products using the microbial-active substance green adjuvant provided in this application (Application Examples 17-21) are significantly superior to high-quality adjuvants in the conventional market in terms of the content of active ingredients before heat storage, the content of active ingredients after heat storage, and the degradation rate of active ingredients after heat storage (Comparative Application Examples 17-21). They are also significantly superior to high-quality adjuvants in the conventional market in terms of the suspension rate before heat storage, the suspension rate after heat storage, and the reduction rate of suspension rate after heat storage (Comparative Application Examples 17-21). Furthermore, they are significantly superior to high-quality adjuvants in the conventional market in terms of the wetting time before heat storage, the wetting time after heat storage, and the growth rate of the wetting time after heat storage (Comparative Application Examples 17-21).

[0100] Further analysis reveals that this significant advantage is reflected in several key performance dimensions. Regarding the content of active ingredients, products using the microbial-active substance green adjuvant of this application show minimal changes in active ingredient content and low degradation rates before and after heat storage, indicating that it better maintains the stability of the active ingredients, extending the product's shelf life and efficacy. In terms of suspension rate, the suspension rate remains high and decreases at a low rate before and after heat storage, demonstrating that the product maintains uniformity and stability during use. Regarding wetting time, the wetting time shows minimal change and a low rate of increase before and after heat storage, meaning the product can quickly wet the target object, improving efficiency and effectiveness. In summary, the microbial-active substance green adjuvant provided in this application exhibits significant performance superiority.

[0101] The following application tests were conducted on the mineral oil emulsifiable concentrate products from Application Examples 3 and 6, and Comparative Application Examples 1-8, which used different additives: Application Experiment 1: Effect of the experiment on the control of red spider mites on citrus trees; This application experiment was conducted in a citrus orchard in Rong'an County, Liuzhou City, Guangxi Province, where citrus red spider mites were severely infested. The experimental design was as follows to verify its control effect on citrus red spider mite disease.

[0102] (1) Application of pesticides: Pesticides were applied during periods of severe spider mite infestation in the citrus orchard, ensuring the leaves were fully moistened. The cultivation period, growth period, and planting density were consistent for all treatments. Weeding, irrigation, and fertilization were carried out according to conventional management practices. No other pesticides were used during the experiment. The dosage of mineral oil products was 150g / mu.

[0103] (2) The experimental steps are as follows: First, the experimental site was divided into uniformly sized application areas, with a no-application treatment area set up between each area.

[0104] Next, the mineral oil emulsifiable concentrate products of different adjuvants application examples 3, 6, and comparative application examples 1-6 were diluted by a dilution factor of 300 times, and the diluted mineral oil products were sprayed onto the experimental field (until the leaves were fully wetted). Each treatment was repeated 3 times.

[0105] (3) Control experiment (blank control area): Set up a control experiment, use water instead of mineral oil products, and spray the experimental field (until the leaves are fully wet).

[0106] (4) Survey time and methods: Three plants were surveyed at each diluted mineral oil product site. One branch from each plant was selected in the east, south, west, north, or center directions and tagged. The number of adult and nymph spider mites on each shoot was counted on five leaves upwards from the tag (note the number on both sides of the leaves). One survey was conducted before application, and three more surveys were conducted at 7, 14, and 21 days after application. The pest reduction rate and control effect were calculated using the following formula. The control effect of application experiment 1 is shown in Table 9.

[0107]

[0108]

[0109] Table 9 – Prevention and Control Effects of Application Experiment 1

[0110] From the experimental results in Table 9, the following conclusions can be drawn: (1) The mineral oil emulsifiable concentrate products using the safe and environmentally friendly microbial-active substance green adjuvant provided in this application (Application Example 3 and Application Example 6) showed significantly better insect population reduction rates and control effects than single bacterial active substance (Comparative Application Example 2), single phycocyanin (Comparative Application Example 3), halloysite nanotubes modified with single brown algal polyphenols (Comparative Application Example 4), the bacterial active substance and phycocyanin compound system (Comparative Application Example 5), and the bacterial active substance and brown algal polyphenol modified halloysite nanotube compound system (Comparative Application Example 6). Example 6), a system of halloysite nanotubes modified with phycocyanin and brown algal polyphenols (comparative application example 7), a system of bacterial active substances + phycocyanin + unmodified halloysite nanotubes (comparative application example 8), and mineral oil alone and microbial-active substance green adjuvant alone (Example 6), demonstrate that the microbial-active substance green adjuvant composed of bacterial active substances + phycocyanin + brown algal polyphenols modified halloysite nanotubes can significantly improve the control effect of mineral oil products on citrus red spider mites and prolong the duration of effectiveness, while the microbial-active substance green adjuvant itself has no activity.

[0111] (2) The mineral oil emulsifiable concentrate products using the safe and environmentally friendly microbial-active green adjuvant provided in this application (Application Example 3 and Application Example 6) showed significantly better insect population reduction rate and control effect than conventional high-quality adjuvants on the market after 7, 14 and 21 days of pesticide application (Comparative Application Example 1).

[0112] (3) The mineral oil emulsion products using the safe and environmentally friendly microbial-active substance green adjuvant provided in this application (Application Example 3 and Application Example 6) showed significantly better insect population reduction rate and control effect than the patent adjuvant with application number CN202411571494.9 (Comparative Application Example 6) after 7, 14 and 21 days of pesticide application.

[0113] (4) The mineral oil emulsifiable concentrate products used in Application Examples 3 and 6 showed better control effects 21 days after application than those 7 and 14 days after application. This indicates that the effective period of the mineral oil emulsifiable concentrate products using safe and environmentally friendly microbial-active green adjuvants was extended to 21 days, significantly improving the effective period of mineral oil products.

[0114] (5) Application Example 6 showed that the insect population reduction rate and control effect were slightly better than those of Application Example 3 after 7, 14 and 21 days of pesticide application. This indicates that treating halloysite nanotubes with KR-41B titanate coupling agent and then modifying them with brown algae polyphenols further improved the control effect of mineral oil emulsifiable concentrate on citrus red spider mites.

[0115] Application Experiment 2: Control Effect of Citrus Canker: Using a severely affected citrus canker orchard in Xixiangtang District, Nanning City, Guangxi Province as the experimental site, the following experimental design was conducted to verify its control effect on citrus canker. The test results of this application experiment are shown in Table 10.

[0116] (1) Application treatment: The mineral oil emulsifiable concentrate products of Application Example 3, Application Example 6, and Comparative Application Example 1-Comparative Application Example 6 with different adjuvants were diluted by a dilution ratio of 300 times, and the diluted mineral oil products were sprayed on the experimental field (until the leaves were fully moistened). Each treatment was repeated 3 times. The first application was carried out at the early stage of citrus canker disease in the citrus plantation, and the leaves were fully moistened. The application was carried out every 10 days, for a total of 3 applications. The cultivation period, growth period and density of each treatment were the same. Weeding, watering and fertilization were carried out in accordance with conventional management. No other agents were used during the experiment.

[0117] (2) Control experiment (blank control area): Set up a control experiment, use water instead of mineral oil products, and spray the experimental field (until the leaves are fully wet).

[0118] (3) Investigation time and method: The control effect was investigated before application and 7 days after each application. Three parallel treatments were conducted in each plot. Two trees were investigated in each plot. For each tree, samples were taken from five points: east, west, south, north, and center. Ten fruits were investigated at each point. The total number of fruits investigated and the number of fruits with diseases at each level were recorded. The disease index and control effect were calculated. The disease grading method and calculation formula are as follows. The experimental results are shown in Table 5 below (the disease index before application is a general statistical analysis of the entire experimental area): Level 0: No disease; Grade 1: Each fruit has 1 to 5 diseased spots; Grade 3: Each fruit has 6 to 10 diseased spots; Grade 5: Each fruit has 11 to 15 diseased spots; Grade 7: Each fruit has 16 to 20 diseased spots; Grade 9: Each fruit has more than 21 disease spots.

[0119]

[0120]

[0121] Table 10 – Control efficacy of various treatment agents against citrus canker

[0122] From the experimental results in Table 10, the following conclusions can be drawn: (1) The mineral oil emulsifiable concentrate products using the safe and environmentally friendly microbial-active substance green adjuvant provided in this application (Application Example 3 and Application Example 6) showed significantly better control efficacy against citrus canker 7 days after three applications compared to single bacterial active substance (Comparative Application Example 2), single phycocyanin (Comparative Application Example 3), halloysite nanotubes modified with single brown algae polyphenols (Comparative Application Example 4), a compound system of bacterial active substance and phycocyanin (Comparative Application Example 5), and a compound system of bacterial active substance and halloysite nanotubes modified with brown algae polyphenols (Comparative Application Example 6). 6) The halloysite nanotube compound system modified with phycocyanin and brown algae polyphenols (comparative application example 7), the system of bacterial active substance + phycocyanin + unmodified halloysite nanotubes (comparative application example 8), and mineral oil alone and microbial-active substance green adjuvant alone (example 6) demonstrate that the microbial-active substance green adjuvant composed of bacterial active substance + phycocyanin + brown algae polyphenol modified halloysite nanotube system can significantly improve the effect of mineral oil products on the prevention and control of citrus canker, while the microbial-active substance green adjuvant itself has no activity.

[0123] (2) The mineral oil emulsifiable concentrate products using the safe and environmentally friendly microbial-active green adjuvant provided in this application (Application Example 3 and Application Example 6) showed significantly better control efficacy against citrus canker than conventional high-quality adjuvants on the market 7 days after three applications (Comparative Application Example 1).

[0124] (3) The mineral oil emulsion products using the safe and environmentally friendly microbial-active green adjuvant provided in this application (Application Example 3 and Application Example 6) showed significantly better control efficacy against citrus canker than the patent adjuvant with application number CN202411571494.9 (Comparative Application Example 6) 7 days after three applications.

[0125] (4) The control effect of application example 6 after three sprays 7 days later was slightly better than that of application example 3. This indicates that the halloysite nanotubes were first treated with KR-41B titanate coupling agent and then modified with brown algae polyphenols, which further improved the control effect of mineral oil emulsion products on citrus canker.

[0126] The above description is merely a preferred embodiment of this application and is not intended to limit this application. Any modifications, equivalent substitutions, and improvements made within the principles of this application should be included within the protection scope of this application.

Claims

1. A microbial-active substance green adjuvant, characterized in that, The additives comprise the following raw materials in the indicated mass percentages: Bacillus belye contains 80% to 85% active substances; Phycocyanin 5% to 10%; The remainder consists of halloysite nanotubes modified with brown algae polyphenols; The active substance of Bacillus belesis is obtained by fermentation culture of Bacillus belesis. The preservation number of the Bacillus belyssus is CGMCC NO.16523.

2. The microbial-active substance green adjuvant according to claim 1, characterized in that, The additives comprise the following raw materials in the indicated mass percentages: Bacillus belysinus contains 83% active substances; Phycocyanin 8%; The remainder consists of halloysite nanotubes modified with brown algae polyphenols.

3. The microbial-active substance green adjuvant according to claim 1, characterized in that, The preparation method of the brown algae polyphenol-modified halloysite nanotubes is as follows: Halloysite nanotubes were placed in a brown algae polyphenol solution, stirred at a mass ratio of 1:10 to 1:20, and dried to obtain the brown algae polyphenol modified halloysite nanotubes. The brown algae polyphenol solution comprises an aqueous solution of brown algae polyphenols and ethanol in a mass ratio of 1:50 to 1:

100.

4. The microbial-active substance green adjuvant according to claim 3, characterized in that, The halloysite nanotubes have an average particle size of 10 nm to 50 nm.

5. The microbial-active substance green adjuvant according to claim 3, characterized in that, The halloysite nanotubes are modified halloysite nanotubes treated with microbial polysaccharides.

6. The microbial-active substance green adjuvant according to claim 5, characterized in that, The microbial polysaccharides include xanthan gum.

7. The microbial-active substance green adjuvant according to claim 1, characterized in that, The method for preparing the active substance of Bacillus belyssus includes the following steps: S1. Inoculate Bacillus belye into LB medium for the first culture to obtain seed fermentation broth; S2. The seed fermentation broth described in step S1 is inoculated into a fermentation medium for a second culture to obtain a sample of Bacillus belye fermentation broth. S3. Centrifuge the Bacillus vesiculosus fermentation broth sample obtained in step S2, remove the supernatant, and obtain the active substance of Bacillus vesiculosus.

8. The microbial-active substance green adjuvant according to claim 7, characterized in that, The LB culture medium in step S1 comprises the following raw materials in the indicated mass amounts: 1% peptone, 0.5% yeast powder, 1% sodium chloride, and purified water to bring the total to 100%; The fermentation medium in step S2 comprises the following raw materials in the indicated mass percentages: The ingredients include 2% to 3% corn steep liquor powder, 3.5% to 4.5% sucrose, 1% to 1.5% dextrin, 0.1% to 0.15% dipotassium hydrogen phosphate, 0.2% to 0.25% magnesium sulfate heptahydrate, and purified water to bring the total to 100%.

9. A method for preparing a microbial-active green adjuvant according to any one of claims 1 to 8, characterized in that, The preparation method includes the following steps: mixing and stirring Bacillus belye active substance, phycocyanin, and halloysite nanotubes modified with brown algae polyphenols until uniform, thereby obtaining the microbial-active substance green adjuvant.

10. The application of a microbial-active green adjuvant according to any one of claims 1 to 8 or an adjuvant prepared by the method according to claim 9 in a pesticide formulation product.

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