A method for preparing plant disease resistance agent using biomass carbon nanomaterials and its application

Abstract

The invention discloses a method for preparing a plant disease-resistant agent by using a biomass carbon nano material and its application. The method uses a biomass material as a raw material, and sequentially sintering, cooling, ball milling, washing and reducing the biomass material to obtain a nano material-level plant disease-resistant agent. The content of O element on the outer surface of the nano material-level plant disease-resistant agent accounts for 18-22% of the total element content on the surface of the material, and the reduction process is carried out under a methane system. The invention reduces the content of O element on the outer surface of the biomass carbon nano material, so that the material itself can better promote the synthesis of hydrogen peroxide in plant tissues. Compared with other disease-resistant preparations of the same type, the amount of hydrogen peroxide produced by the biomass carbon nano material is increased by 98.09%. Because the content of hydrogen peroxide as an endogenous antibacterial compound in plants is increased, the disease resistance of plants is ultimately improved, and no damage is caused to plants and the environment.

Description

Technical Field

[0001] The present invention relates to the field of agricultural protective agents, and in particular to a method for preparing a plant disease-resistant agent by utilizing biomass carbon nanomaterials and the application of the plant disease-resistant agent in targeting soil pathogenic bacteria pollution. Background Art

[0002] At present, soil-borne diseases are causing serious damage worldwide. According to statistics from the Food and Agriculture Organization of the United Nations, plant diseases cause losses of more than 220 billion US dollars to the global economy each year. According to statistics in 2021, my country loses 14 million tons of grain each year due to pests and diseases. Among the many soil-borne diseases, Fusarium spp. is a typical soil-borne pathogenic fungus that can cause plant wilt and root rot. The fungus can infect more than 100 crops with important economic value, including Musaceae plants such as bananas, Rosaceae plants such as strawberries, Cucurbitaceae plants such as watermelons, Cruciferae plants such as cabbage, Solanaceae plants such as tomatoes, and Leguminosae plants such as alfalfa. Fusarium colonizes in the vascular bundles of the plant roots, causing blockage of the ducts, withering of the aboveground parts, brown necrotic spots on the root epidermis, and later root rot and even death of the entire plant, seriously affecting the growth, development, yield and quality of the plant. In recent years, my country has reported many cases of crop yield reduction caused by Fusarium infection. For example, watermelon fields infected by Fusarium have a 20%-30% reduction in yield, and in severe cases, it can reach 50%-60%, or even no yield; in banana orchards, the infection rate of diseased plants is 10%-40%, and in severe cases, it can reach more than 90%, resulting in the abandonment of banana orchards; alfalfa yields drop by 20%-40% after being infected by Fusarium, and in severe cases, it can drop by more than 60%. For the damage caused by Fusarium in agriculture, it is urgent to find an effective method to control the damage of Fusarium to crops.

[0003] Nanomaterials refer to materials with at least one dimension in the three-dimensional space at the nanoscale. They are a new generation of materials composed of nanoparticles whose sizes are between atoms, molecules and macroscopic systems. Studies have reported that nanomaterials developed and produced by nanotechnology not only have good adsorption and bactericidal capabilities, but also have very good absorption effects, so nanomaterials have gradually been applied to soil-borne disease prevention and control and environmental pollutant treatment. In soil-borne disease prevention and control, exogenous application of copper-based nano-ions can effectively improve the disease resistance of tomatoes to Fusarium, and the yield of tomatoes has increased by 58%; foliar spraying of silicon-based nanomaterials can also effectively improve the resistance of watermelons to Fusarium, and the yield of watermelons has recovered by 81.5%; and exogenous addition of copper-based nanomaterials can effectively improve the disease resistance of soybean roots to Fusarium, thereby ensuring the growth of soybeans. It is calculated that the cost of nanomaterials as plant disease-resistant agents is only 40% of that of traditional pesticides.

[0004] Compared with other engineered nanomaterials, biochar materials are green, environmentally friendly, and low-carbon. Studies have reported that metal or metal oxide nanomaterials such as gold, silver, copper, zinc, magnesium, molybdenum oxide, zinc oxide, titanium dioxide, and copper iodide all have anti-pathogenic activity. For example, salts such as heavy metal silver can react with sulfhydryl groups in proteins, or replace metal ions in enzymes, inactivating most enzymes and killing pathogens. However, in agricultural production, the large-scale use of metal nanomaterials will inevitably cause environmental and ecological risks. Therefore, as an environmentally friendly material, biochar nanomaterials are of great significance in plant disease resistance. Summary of the invention

[0005] In view of the above-mentioned problems, the present invention aims to provide an environmentally friendly and effective method for preparing plant disease resistance agents using biochar nanomaterials and its application in treating soil pathogen pollution.

[0006] In order to achieve the above-mentioned purpose, the technical scheme adopted by the present invention is as follows: a method for preparing a plant disease-resistant agent using biomass carbon nanomaterials, wherein the method uses biomass materials as raw materials, and the biomass materials are sintered, cooled, ball-milled, washed, and reduced in sequence to obtain a nano-grade plant disease-resistant agent; the content of O element on the outer surface of the nano-grade plant disease-resistant agent accounts for 18-22% of the total element content on the surface of the material; the biomass material is corn stalks, the sintering temperature is 350-650°C, the ball-to-material volume ratio in the ball milling process is 3:1, and the reduction process is carried out under a methane system.

[0007] The nanomaterial-grade plant disease resistance agent of the present invention greatly reduces the content of O element on its outer surface, so that when the biomass carbon nanomaterial is applied to plants, it can better activate plant tissues to produce hydrogen peroxide. Compared with other carbon nanomaterials in the prior art, the amount of hydrogen peroxide is increased by 21.3% to 98.09%, ultimately improving the resistance of crops to soil pathogens.

[0008] The method of the present invention specifically comprises the following steps:

[0009] 1) Sinter the corn stalks at 650°C and cool them to room temperature after sintering;

[0010] 2) Put the cooled material into an omnidirectional ball mill and grind for 6 hours;

[0011] 3) The ground material was washed with alcohol for 4 times, and the biomass carbon nanomaterial with a particle size of less than 100 nm was precipitated through a dialysis bag. The alcohol was removed by a rotary evaporator, and the biomass carbon nanomaterial was obtained after standing.

[0012] 4) The biomass carbon nanomaterial obtained in step 3) is evenly distributed in a 300°C ventilation pipe, and methane is introduced into the ventilation pipe at a flow rate of 2 mL / min for 30 minutes; after the reaction is completed, nitrogen is introduced for cooling to obtain a nanomaterial-grade plant disease resistance agent;

[0013] 5) The nanomaterial-grade plant disease-resistant agent obtained in step 4) is placed in a vacuum bottle and sealed for storage.

[0014] The content of O element on the outer surface of the nano material-grade plant disease resistance agent of the present invention accounts for 18% of the total element content on the outer surface.

[0015] In step 4) of the present invention, the sealed storage temperature is ≤ 4°C.

[0016] The application of the plant disease-resistant agent of the present invention in soil pathogenic bacteria contamination, the application method is as follows:

[0017] 1) Mix the plant disease-resistant agent and the aqueous solution in a volume ratio of 1:5-20, then add 1% of the total volume of a biological surfactant, and ultrasonically shake the prepared mixed solution;

[0018] 2) applying the mixed solution obtained in step 1) to the leaves of crops with mild soil pathogen contamination or to the seedling stage of crops with severe soil pathogen contamination;

[0019] 3) Evaluate the effectiveness of plant disease resistance agents based on the pathogen content, yield and quality of crops.

[0020] In step 1) of the application method of the present invention, the volume ratio of the plant disease-resistant agent to the aqueous solution is preferably 1:10; and the biological surfactant is preferably sophorolipid.

[0021] In step 2) of the application method of the present invention, for mild soil pathogen contamination: take a certain amount of the mixed solution and spray it on the surface of crop leaves through a spray bottle; after the entire leaf surface is evenly moistened, let it stand and dry; after no dripping, transplant the seedlings.

[0022] In step 2) of the application method of the present invention, for severe soil pathogen contamination: a certain amount of the mixed solution is injected into a syringe, and then injected into the root ball of the plant seedling through the syringe, so that the root ball is completely infiltrated by the mixed solution; after standing until there is no dripping, the seedlings are transplanted; during the transplanting process, the bulbs are completely buried in the soil to complete the soil covering operation.

[0023] In step 3) of the application method of the present invention, the yield and quality of crops are detected:

[0024] ① When crops enter the growth period, regularly check the growth of crops: record the plant height, morphology and leaf area every 7 days;

[0025] ② After the crops reach full maturity, take the above-ground parts of leafy crops and measure the biomass.

[0026] In step 3) of the application method of the present invention, the pathogen content of crops is detected:

[0027] ① When crops enter the growth period, the content of hydrogen peroxide in the leaves is measured by the diaminobenzidine method;

[0028] ② After the crops are fully mature, take a longitudinal section of the crop stems to test the growth of pathogens.

[0029] Plant disease-resistant preparations can enter the human body through the food chain when sprayed on plants, causing more serious consequences. Germany once found toxic pesticides in popular herbal teas on the market. These pesticides can inhibit human cholinesterase and accumulate in the nervous system, causing nausea, dizziness, and even unconsciousness. Exposure to high concentrations can also cause respiratory paralysis and death. Compared with such high-risk chemical preparations, this antibacterial preparation does not have strong biological toxicity in terms of its composition.

[0030] The advantages of the present invention are as follows: the present invention provides an environmentally friendly biomass carbon nanomaterial plant disease-resistant preparation; by reducing the content of O element on the outer surface of the biomass carbon nanomaterial, the material itself can better promote the synthesis of hydrogen peroxide in plant tissues; compared with other disease-resistant preparations of the same type, the amount of hydrogen peroxide produced by plant tissues is increased by 98.09%; because the content of hydrogen peroxide as an endogenous antibacterial compound in plants is increased, the disease resistance of plants is ultimately improved; and at the same time, no damage is caused to plants and the environment.

[0031] The plant disease-resistant preparation of the present invention has the characteristics of high efficiency and rapid plant disease-resistant response; it has a positive effect on the prevention and control of soil-borne diseases in my country's agricultural production, and can also provide inspiration and ideas for improving plant disease resistance with nanomaterials. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] Figure 1 This is a graph showing the results of the biomass charcoal material fired at 650° C. in Example 2 of the present invention in controlling tobacco phytophthora;

[0033] Figure 2 This is a graph showing the results of the biomass charcoal material fired at 350° C. in Example 2 of the present invention in controlling tobacco phytophthora;

[0034] Figure 3 A blank control diagram of tobacco leaves processed with biomass charcoal material not used in Example 2 of the present invention. Implementation

[0035] The present invention is further described in detail below in conjunction with the accompanying drawings and specific implementation methods.

[0036] Example 1: Preparation method;

[0037] Take 500 g of corn stalks harvested that year and burn them in a muffle furnace at 350 °C and 650 °C to make biochar materials; take 20 g of the biochar materials burned at 350 °C and 650 °C respectively, wash them with alcohol for 4 times, and then use an omnidirectional ball mill with a ball-to-material ratio of (3:1) for 6 hours;

[0038] After grinding, the material was filtered through a dialysis membrane and then transferred to a rotary evaporator to obtain the biomass carbon nano-antibacterial material (350°C and 650°C);

[0039] The biomass carbon nano-antibacterial material (350℃ and 650℃) was placed in the ventilation tube, heated to 300℃, and then 4 mL of methane gas was introduced per minute. After 30 minutes, the heating was stopped and nitrogen was introduced. The sample was taken out after about 20 minutes and set aside. The XPS test showed that:

[0040] The content of O element on the outer surface of the sample of this embodiment accounts for 18.06% of the total element content on the outer surface (650 ℃);

[0041] The content of O element on the outer surface of the sample in this example accounts for 25.90% of the total element content on the outer surface (350°C)

[0042] The detailed test results are shown in the following table:

[0043]

[0044] Storage: Seal the prepared biomass carbon nano-antibacterial material in a sterile vacuum bottle.

[0045] Example 2: Application of tobacco phytophthora control;

[0046] 0.1 g of biochar nano-antibacterial material (350℃ and 650℃) was dissolved in 1000 mL of aqueous solution, 1% of sophorolipid was added, and after ultrasonic treatment for 30 min, 100 ppm of biochar nano-antibacterial liquid was prepared for standby use.

[0047] Take 5 mL of 100 ppm biochar nanoantibacterial liquid fired at 350℃ and 650℃, expose it to the roots of tobacco, and then transplant it into the soil infected by tobacco phytophthora.

[0048] After 7 days, the amount of hydrogen peroxide in the leaves measured by the diaminobenzidine method increased by 98.09% (biomass charcoal nano-antibacterial liquid fired at 650 °C) and 47.88% (biomass charcoal nano-antibacterial liquid fired at 350 °C). Both were confirmed to be immune. The results of 350 °C and 650 °C for tobacco leaves are as follows Figure 2 and 1 As shown, Figure 3 This is a blank control diagram of tobacco leaves processed without using biomass carbon materials; in comparison, the amount of hydrogen peroxide in plant leaves of the present invention is significantly increased.

[0049] Example 3: Comparative Example - Anti-disease performance of biomass carbon nanomaterials prepared by different methods;

[0050] Take 500 g of corn stalks harvested that year and burn them in a muffle furnace at 650 °C to make biochar materials; take 20 g of the biochar materials burned at 650 °C, wash them with alcohol 4 times, and then use an omnidirectional ball mill with a ball-to-material ratio of (3:1) for 6 hours;

[0051] After grinding, the material was filtered through a dialysis membrane and then transferred to a rotary evaporator to obtain a biomass carbon nano-antibacterial material. XPS testing showed that the content of O element on the outer surface of the biomass carbon nano-antibacterial material accounted for 35.52% of the total element content on the outer surface. The detailed test results are shown in the following table:

[0052]

[0053] 0.1 g of the above-mentioned biochar nanomaterial was dissolved in 1000 mL of aqueous solution, and 1% of sophorolipids was added to prepare an antibacterial material solution; after ultrasonication for 30 min, a 100 ppm biochar nanoantibacterial solution was prepared for standby use;

[0054] Take 5 mL of 100 ppm of the nano-antibacterial liquid of the biochar nanomaterials that has not been treated with methane, expose it to the roots of tobacco, and then transplant it into the soil infected with tobacco phytophthora;

[0055] After 7 days, the amount of hydrogen peroxide in the leaves measured by the diaminobenzidine method increased by 10.33%; compared with the increase in the amount of hydrogen peroxide in the leaves of the biomass carbon nanomaterial prepared in Example 1 of the present invention, the amount of hydrogen peroxide was greatly reduced. Therefore, the biomass carbon nanomaterial obtained after the adjustment method has a significantly reduced disease resistance to plants.

[0056] Example 4: Comparative Example - Preparation ratio of antibacterial agent;

[0057] 0.05, 0.1 and 0.2 g of biochar nano-antibacterial material (650 ° C) were dissolved in 1000 mL of aqueous solution, 1% of sophorolipid was added, and after ultrasonic treatment for 30 min, 50, 100 and 200 ppm of biochar nano-antibacterial liquid were prepared for standby use;

[0058] Take 5 mL of 50, 100 and 200 ppm biochar nano antibacterial liquid and expose it to the roots of tobacco. The experiment found that the 200 ppm antibacterial liquid is thicker and forms particles after entering the root system, while the 50 and 100 ppm nano antibacterial liquid can infiltrate the root system normally; then transplant it into the soil infected with tobacco blight;

[0059] After 7 days, the amount of hydrogen peroxide in the leaves of tobacco plants treated with 50, 100 and 200 ppm biochar nano-antibacterial liquid increased by 40.2%, 98.09% and 21.3% respectively as measured by the diaminobenzidine method. The experiment showed that 100 ppm biochar nano-antibacterial liquid had the best immune effect.

[0060] Example 5: Comparative Example - Comparison of Similar Anti-disease Preparations;

[0061] Comparison of antibacterial effects

[0062] 0.1 g of biochar nano-antibacterial material (650 ° C) was dissolved in 1000 mL of aqueous solution, 1% of sophorolipid was added, and after ultrasonic treatment for 30 min, 100 ppm of biochar nano-antibacterial liquid was prepared for standby use;

[0063] Take 0.1 g of the same dose of CuSO4 reagent, dissolve it in 1000 mL of aqueous solution, and ultrasonicate it for 30 min to obtain a 100 ppm copper sulfate solution;

[0064] Take 5 mL of 100 ppm biochar nanoantibacterial solution and CuSO4 solution, expose the roots of tobacco, and then transplant them into the soil infected with tobacco phytophthora;

[0065] After 7 days, tobacco treated with biochar nano-antibacterial liquid and CuSO4 solution both grew well, but the Cu content in the soil after CuSO4 treatment increased by 35.6% compared with the untreated soil, reaching 20.2 ppm; compared with the environmental risk indicator threshold of 3-300 ppb, the use of CuSO4 reagent has posed a greater risk to environmental health.

[0066] It should be noted that the above are merely preferred embodiments of the present invention and are not intended to limit the protection scope of the present invention. Any combination or equivalent transformation made on the basis of the above embodiments shall fall within the protection scope of the present invention.

Claims

1. A method for preparing a plant disease-resistant agent using biomass carbon nanomaterials, characterized in that: The method uses biomass materials as raw materials, and sequentially sintering, cooling, ball milling, washing, and reducing the biomass materials to obtain a nano-material-grade plant disease-resistant agent; the molar content of O element on the outer surface of the nano-material-grade plant disease-resistant agent accounts for 18-22% of the total molar content of elements on the outer surface; The biomass material is corn stalks, the sintering temperature is 350-650° C., the ball-to-material volume ratio in the ball milling process is 3:1, and the reduction process is carried out under a methane system.

2. The method for preparing a plant disease resistance agent according to claim 1, characterized in that: The steps include: 1) Sinter the corn stalks at 650°C and cool them to room temperature after sintering; 2) Put the cooled material into an omnidirectional ball mill and grind for 6 hours; 3) The ground material was washed with alcohol for 4 times, and the biomass carbon nanomaterial with a particle size of less than 100 nm was precipitated through a dialysis bag. The alcohol was removed by a rotary evaporator, and the biomass carbon nanomaterial was obtained after standing. 4) The biomass carbon nanomaterial obtained in step 3) is evenly distributed in a 300°C ventilation pipe, and methane is introduced into the ventilation pipe at a flow rate of 2 mL / min for 30 minutes; after the reaction is completed, nitrogen is introduced for cooling to obtain a nanomaterial-grade plant disease resistance agent; 5) The nanomaterial-grade plant disease-resistant agent obtained in step 4) is placed in a vacuum bottle and sealed for storage.

3. The method for preparing a plant disease resistance agent according to claim 1 or 2, characterized in that: The molar content of O element on the outer surface of the nanomaterial-grade plant disease-resistant agent accounts for 18% of the molar content of the total elements on the outer surface.

4. The method for preparing a plant disease resistance agent according to claim 2, characterized in that: In the step 5), the sealed storage temperature is ≤ 4°C.

5. Use of a plant disease-resistant agent prepared by the method according to claim 1 or 2 in treating soil pathogenic bacteria contamination, characterized in that: The application method is as follows: 1) Mix the plant disease-resistant agent and the aqueous solution in a volume ratio of 1:5-20, then add 1% of the total volume of a biological surfactant, and ultrasonically shake the prepared mixed solution; 2) applying the mixed solution obtained in step 1) to the leaves of crops with mild soil pathogen contamination or to the seedling stage of crops with severe soil pathogen contamination; 3) Evaluate the effectiveness of plant disease resistance agents based on the pathogen content, yield and quality of crops.

6. Use of the plant disease resistance agent according to claim 5 in treating soil pathogenic bacteria contamination, characterized in that: In the step 1), the volume ratio of the plant disease-resistant agent to the aqueous solution is 1:10; and the biological surfactant is sophorolipid.

7. Use of the plant disease resistance agent according to claim 5 in treating soil pathogenic bacteria contamination, characterized in that: In the step 2), for mild soil pathogen contamination: take the mixed solution and spray it on the surface of crop leaves through a spray bottle; after the entire leaf surface is evenly moistened, let it stand and dry; after no dripping liquid, transplant the seedlings.

8. Use of the plant disease resistance agent according to claim 5 in treating soil pathogenic bacteria contamination, characterized in that: In the step 2), for severe soil pathogen contamination: inject the mixed solution into a syringe, and inject it into the root ball of the plant seedling through the syringe, so that the root ball is completely infiltrated by the mixed solution; after standing until there is no dripping, transplant the seedlings; during the transplanting process, the bulbs are completely buried in the soil to complete the soil covering operation.

9. Use of the plant disease resistance agent according to claim 5 in treating soil pathogenic bacteria contamination, characterized in that: In step 3), the yield and quality of crops are tested: ① When crops enter the growth period, regularly check the growth of crops: record the plant height, morphology and leaf area every 7 days; ② After the crops reach full maturity, take the above-ground parts of leafy crops and measure their biomass.

10. Use of the plant disease resistance agent according to claim 5 in treating soil pathogenic bacteria contamination, characterized in that: In step 3), the pathogen content of crops is detected: ① When crops enter the growth period, the content of hydrogen peroxide in the leaves is measured by the diaminobenzidine method; ② After the crops are fully mature, take a longitudinal section of the crop stems to test the growth of pathogens.

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