Nanomaterials and nanocomposites loaded with biopesticides and their applications

Abstract

The present invention relates to nanomaterials and nanocomposites loaded with biopesticides and their applications. Based on research, the present invention screens out nanomaterials suitable for loading biopesticides and prepares three high-quality nanocomposites. Furthermore, based on the different characteristics of the nanocomposites, the inventors have developed two biological control methods, spraying and root irrigation, which have better colonization effects than the current use of Beauveria bassiana and have very good agricultural application value.

Description

Technical Field

[0001] The invention belongs to the field of pest control, and specifically relates to a preparation method and application of nano materials and nano composites loaded with biological pesticides. Background Art

[0002] Biological pesticides refer to preparations that use living organisms (especially fungi, bacteria, insect viruses, natural enemies, etc.) to kill or inhibit agricultural pests. Common biological pesticides include microorganisms such as nematodes, bacteria and viruses. Biological pesticides play an important role in the pest management system of organic agriculture, but their market share is still quite limited. This is mainly due to various factors that limit the development of biological pesticides, such as short shelf life and susceptibility to environmental influences.

[0003] In the field of pest control, thrips are one of the most important agricultural pests that are difficult to control. They are characterized by their tiny bodies, hidden damage, strong reproductive capacity, and strong drug resistance. Traditional chemical pesticides are difficult to achieve a trace and high-efficiency control effect on them in terms of application methods and effects. Biological pesticides such as Beauveria bassiana have a high lethality to western flower thrips and are parasitic, with little impact on other organisms. As an entomogenous fungus, it colonizes in plants to form a symbiont that can change the behavior and feeding specificity of herbivorous insects, and can indirectly protect plants while improving the plants' ability to resist stress. At present, Beauveria bassiana preparations have been developed for pest control, but there is still room for improvement in their control effects and storage capacity. Summary of the invention

[0004] Therefore, it is necessary to develop better quality Beauveria bassiana vectors for use in green control technology.

[0005] The purpose of the present application is to provide an application of graphene oxide or multi-walled carbon nanotubes in loading biological pesticides, wherein the graphene oxide or multi-walled carbon nanotubes and the biological pesticides form a nanocomposite.

[0006] Furthermore, the volume ratio of the biopesticide to graphene oxide or multi-walled carbon nanotubes is 3-5:1; preferably, the volume ratio of the biopesticide to graphene oxide or multi-walled carbon nanotubes is 4:1.

[0007] Furthermore, the graphene oxide is material No. 1, material No. 2 or material No. 3, and the biological pesticide is fungus; optionally, the method for preparing a nanocomposite composed of graphene oxide and biological pesticide includes: preparing a graphene oxide dispersion; preparing a fungal spore suspension; mixing the fungal spore suspension and graphene oxide in a volume ratio of 4:1, stirring the mixture on a magnetic stirrer at room temperature in the dark, and preparing an aqueous solution of the nanocomposite.

[0008] The purpose of the present application is to provide a nanocomposite, comprising a spherical biopesticide and graphene oxide, wherein the biopesticide is one or more of the following living organisms: fungi, bacteria, insect viruses;

[0009] Optionally, the fungus is Beauveria bassiana; preferably, the Beauveria bassiana is strain GZGY-1-3;

[0010] Optionally, the graphene oxide is material No. 1, material No. 2 or material No. 3.

[0011] Further, the nanocomposite is configured into an aqueous solution, and the method includes: preparing a graphene oxide dispersion; preparing a Beauveria bassiana spore suspension; mixing the Beauveria bassiana spore suspension and the graphene oxide dispersion in a volume ratio of 4:1 and stirring at room temperature in the dark to obtain an aqueous solution of the nanocomposite; preferably, the method includes: preparing a graphene oxide dispersion (concentration 100 μg / ml); preparing a Beauveria bassiana spore suspension (concentration 1×10 8 The suspension of Beauveria bassiana spores and the graphene oxide dispersion were mixed in a volume ratio of 4:1 and stirred on a magnetic stirrer at 1200 rpm in the dark at room temperature for 24 hours to obtain an aqueous nanocomposite.

[0012] Optionally, the preparation method of the Beauveria bassiana spore suspension comprises: picking up the spore powder of the Beauveria bassiana strain after activation culture, mixing it with a Tween sterilized solution, grinding it evenly with a tissue grinder, fully mixing it with a vortex oscillator, determining the spore content, and finally diluting it with a Tween sterilized solution to prepare a spore suspension of desired concentration; preferably, picking up the spore powder of the strain after activation culture with a disposable sterile inoculation loop, mixing it with a 0.05% Tween-80 sterilized solution, grinding it evenly with a glass tissue grinder, fully mixing it with a vortex oscillator, determining the spore content with a 25-grid blood cell counting plate, counting three times, taking an average value, and finally diluting it with a 0.05% Tween-80 sterilized solution to prepare a spore suspension of desired concentration.

[0013] Preferably, the preparation method of the nanocomposite is as follows: obtain material No. 1, material No. 2 or material No. 3 (material No. 3 is solid and is first crushed in sterilized ultrapure water using an ultrasonic cleaner), prepare a graphene oxide dispersion (concentration 100 μg / ml); prepare a Beauveria bassiana spore suspension (concentration 1×10 8 The suspension of Beauveria bassiana spores and the graphene oxide dispersion were mixed in a volume ratio of 4:1 and stirred on a magnetic stirrer at 1200 rpm in the dark at room temperature for 24 hours to obtain an aqueous nanocomposite.

[0014] The purpose of the present application is to provide the use of a nanocomposite in controlling agricultural pests, wherein the nanocomposite is the nanocomposite described above;

[0015] Optionally, the aqueous solution of the No. 1 complex is applied to the crops by root irrigation to complete the colonization of Beauveria bassiana;

[0016] Optionally, the aqueous solution of compound No. 2 or compound No. 3 is applied to crops by spraying to complete the colonization of Beauveria bassiana.

[0017] The purpose of the present application is to provide a method for preparing a nanocomposite, comprising:

[0018] A graphene oxide dispersion is prepared; a Beauveria bassiana spore suspension is prepared; the Beauveria bassiana spore suspension and the graphene oxide dispersion are mixed in a volume ratio of 4:1, and then stirred and compounded at room temperature to prepare an aqueous solution of a nanocomposite, and the nanocomposite powder is obtained after drying;

[0019] Preferably, a graphene oxide dispersion with a concentration of 100 μg / ml is prepared; 1×10 8 A spore suspension of Beauveria bassiana spores / ml, a Beauveria bassiana spore suspension and a graphene oxide dispersion are mixed in a volume ratio of 4:1, and then stirred and compounded with a magnetic stirrer at room temperature for 24 hours to obtain an aqueous nanocomposite, and then dried to obtain a nanocomposite powder.

[0020] The purpose of this application is to provide a method for spraying and controlling agricultural pests, the method comprising:

[0021] The above-mentioned material No. 2 or material No. 3 is used to prepare an aqueous solution of the nanocomposite; the aqueous solution is sprayed on the leaves of crops to complete the colonization of Beauveria bassiana; the leaves of crops after colonization have contact killing and / or stomach poisoning effects on agricultural pests;

[0022] Optionally, the spraying amount is 6-10 ml of the nanocomposite aqueous solution per crop, preferably, 8 ml of the nanocomposite aqueous solution per crop.

[0023] The purpose of this application is to provide a root irrigation control method for agricultural pests, the method comprising:

[0024] The above-mentioned material No. 1 is used to prepare an aqueous solution of the nanocomposite; the water is irrigated along the roots of the crops for three consecutive days to complete the colonization of Beauveria bassiana;

[0025] Optionally, 15-25 ml of the nanocomposite aqueous solution is used for root irrigation of each crop each time; preferably, 20 ml of the nanocomposite aqueous solution is used for root irrigation of each crop each time.

[0026] Furthermore, the crops include cereal crops, legume crops and / or vegetables;

[0027] More preferably, the crops include one or more of the following: wheat, rice, corn, soybean, broad bean, pea, mung bean, tomato, eggplant, kidney bean, cowpea, pepper;

[0028] Optionally, the pests are corn pests; preferably, the pests include one or more of the following: corn borer, fall armyworm, thrips, small cutworm, yellow cutworm, mole cricket, grub, wireworm, corn sunflower mealybug, corn spinner, grass borer, rice leaf roller, Spodoptera litura, corn armyworm, armyworm, yellow-bellied lantern moth, red-margined lantern moth, brown-footed horned leaf beetle, spotted stink bug, red-barbed blind bug, pasture blind bug, three-spotted leafhopper, large green leafhopper, corn aphid, truncate spider mite, cinnabar spider mite, Asian corn borer, large borer, peach borer, cotton bollworm, white-spotted flower beetle, small blue flower beetle, two-spotted glossy leaf beetle, green rice stink bug.

[0029] Technical advantages of the present invention:

[0030] 1 This application provides a high-quality nanomaterial that can load biological pesticides (such as Beauveria bassiana). The electron microscopy experimental results show that graphene oxide has obvious advantages in loading biological pesticides;

[0031] 2 The present application prepares a high-quality nanocomposite, namely, graphene oxide loaded with biopesticide, which has good economic and biological benefits when the spore suspension of Beauveria bassiana and the graphene oxide dispersion are mixed in a volume ratio of 4:1;

[0032] 3 The present application provides a spraying biological control method, which is based on the ultraviolet protection function of graphene oxide No. 2 and graphene oxide No. 3 on the spores of Beauveria bassiana, and has a better leaf colonization effect than the conventional method of using Beauveria bassiana;

[0033] 4 The present application provides a root irrigation biological control method, which is based on the insensitivity of the No. 1 compound to temperature and has a better root colonization effect than the conventional method of using Beauveria bassiana. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 This is a scanning electron microscope image of material No. 1 and its loaded Beauveria bassiana nanocomposite. The left side is a scanning electron microscope image of material No. 1, and the right side is a scanning electron microscope image of material No. 1 loaded Beauveria bassiana nanocomposite.

[0035] Figure 2 This is a scanning electron micrograph of material No. 2 and its loaded Beauveria bassiana nanocomposite. The left side is a scanning electron micrograph of material No. 2, and the right side is a scanning electron micrograph of material No. 2 loaded Beauveria bassiana nanocomposite.

[0036] Figure 3 This is a scanning electron microscope image of material No. 3 and its loaded Beauveria bassiana nanocomposite. The left side is a scanning electron microscope image of material No. 3, and the right side is a scanning electron microscope image of material No. 3 loaded Beauveria bassiana nanocomposite.

[0037] Figure 4 This is a scanning electron micrograph of material No. 4 and its loaded Beauveria bassiana nanocomposite. The left side is a scanning electron micrograph of material No. 4, and the right side is a scanning electron micrograph of material No. 4 loaded Beauveria bassiana nanocomposite.

[0038] Figure 5 This is a scanning electron micrograph of material No. 5 and its loaded Beauveria bassiana nanocomposite. The left side is a scanning electron micrograph of material No. 5, and the right side is a scanning electron micrograph of material No. 5 loaded Beauveria bassiana nanocomposite.

[0039] Figure 6 The scanning electron micrographs of silica and its loaded Beauveria bassiana nanocomposite are shown, with the left side showing the scanning electron micrograph of silica and the right side showing the scanning electron micrograph of the silica loaded Beauveria bassiana nanocomposite.

[0040] Figure 7 It is the infrared spectrum of graphene oxide No. 1, Beauveria bassiana, and nanocomposite;

[0041] Figure 8 This is the infrared spectrum of graphene oxide No. 2, Beauveria bassiana, and nanocomposite

[0042] Fig. 9 It is the infrared spectrum of graphene oxide No. 3, Beauveria bassiana, and nanocomposite;

[0043] Fig.10 This is a diagram showing the effects of nanomaterials prepared in different proportions on the germination of Beauveria bassiana spores;

[0044] Fig.11 is a line graph of the germination rate of the nanocomposite at different temperatures;

[0045] Fig.12 is a line graph of the spore inactivation rate of the nanocomposite after UV irradiation;

[0046] Fig.13 It is a line graph of the survival rate of western flower thrips after the nanocomposite treatment. DETAILED DESCRIPTION

[0047] The present invention is further described below in conjunction with specific embodiments, which are only used to explain the present invention and cannot be understood as limiting the present invention. It can be understood by those skilled in the art that various changes, modifications, substitutions and variations can be made to these embodiments without departing from the principles and purpose of the present invention, and the scope of the present invention is defined by the claims and their equivalents.

[0048] Example 1 Nanomaterial Screening

[0049] 1.1 Materials and methods

[0050] 1.1.1 Test materials:

[0051] Nanocarrier: Purchased graphene oxide materials from MACKLIN (Shanghai MACKLIN Biochemical Technology Co., Ltd.) from Fangguo (Beijing) Technology Co., Ltd. in 2023:

[0052] Material No. 1 (graphene oxide No. 1) is: graphene oxide (dispersion); average radial size 20-30 μm; thickness: 1 nm;

[0053] Material No. 2 (graphene oxide No. 2) is: graphene oxide (dispersion); average radial size 20-30 μm; thickness: 1 nm;

[0054] Material No. 3 (graphene oxide No. 3) is: graphene oxide (solid); purity 99%; sheet diameter 0.5-5 μm; thickness 0.8-1.2 nm;

[0055] The above materials were stored in a dark place at 4°C, and the properties were as follows: solid / dispersion (solid was crushed in sterilized ultrapure water by ultrasonic cleaning), the ratio was 100 μg / ml (dispersed in ddH2O);

[0056] No. 1 multi-walled carbon nanotube (No. 4 material), No. 2 multi-walled carbon nanotube (No. 5 material), silicon dioxide;

[0057] Biopesticide: Beauveria bassiana, commercial Beauveria bassiana or laboratory-produced strains such as GZGY-1-3 strain can be selected; Nanocomposite: Material No. 1 loaded Beauveria bassiana nanocomposite (No. 1 complex), Material No. 2 loaded Beauveria bassiana nanocomposite (No. 2 complex), Material No. 3 loaded Beauveria bassiana nanocomposite (No. 3 complex), Material No. 4 loaded Beauveria bassiana nanocomposite, Material No. 5 loaded Beauveria bassiana nanocomposite, Silica loaded Beauveria bassiana nanocomposite;

[0058] The preparation method of the nanocomposite is as follows: obtain material No. 1, material No. 2 or material No. 3 (the solid is crushed in sterilized ultrapure water by ultrasonic cleaning), prepare graphene oxide dispersion (concentration 100 μg / ml); prepare Beauveria bassiana spore suspension (concentration 1×10 8 The suspension of Beauveria bassiana spores and the graphene oxide dispersion were mixed in a volume ratio of 4:1 and stirred on a magnetic stirrer at 1200 rpm in the dark at room temperature for 24 hours to obtain an aqueous nanocomposite.

[0059] 1.1.2 Machine model: Japan Hitachi cold field emission scanning electron microscope Regulus8100

[0060] 1.1.3 The dried nanocomposite powder was treated with gold spraying. The scanning electron microscope (SEM) was a Hitachi microscope of Japan with an operating voltage of 5KV. The sample surface was bombarded with a finely focused electron beam. The surface morphology of the sample was observed and analyzed by the secondary electrons and backscattered electrons generated by the interaction between the electrons and the sample. After observing the structure of the biopesticide (Beauveria bassiana) and the structure of the nanocarrier (graphene oxide, multi-walled carbon nanotubes, silicon dioxide) under the scanning electron microscope, the more suitable materials were preliminarily screened by the compatibility of the biopesticide and the nanocarrier.

[0061] 1.1.4 Result Analysis

[0062] Experiments were conducted on 6 candidate nanomaterials, and the results are shown in Figure 1-6 , combined with the image observation and analysis, it was found that graphene oxide and multi-walled carbon nanotubes can load a certain amount of Beauveria bassiana, while silicon dioxide without functional group modification cannot adhere to or load Beauveria bassiana, and is in a non-inclusion-related separation state under a scanning electron microscope (see Figure 6 Of the two materials that can be loaded, single-layer graphene oxide has the characteristics of long sheet diameter and thin thickness. After being treated with a magnetic stirrer, it can be layered to adhere to, coat, and load Beauveria bassiana in large quantities (see Figure 1-3 ). Multi-walled carbon nanotubes can be used to adhere and load a small amount of Beauveria bassiana due to their small diameter and small gaps between different nanotubes (see Figure 4 and Figure 5 ). The results of scanning electron microscopy preliminarily show that the three graphene oxide materials can load, wrap and adhere to Beauveria bassiana in large quantities, and are ideal carriers for biological pesticides (Beauveria bassiana).

[0063] 1.2 Fourier transform infrared absorption spectroscopy (FTIR)

[0064] 1.2.1 Machine model: American Thermo Scientific Nicolet iS20

[0065] 1.2.2 Fourier transform infrared absorption spectroscopy (FT-IR) was analyzed using the iS20 infrared spectrometer of Thermo Company of the United States. The types of functional groups contained on the surface of the material were determined and analyzed by the position, intensity and shape of the characteristic peaks of graphene oxide, Beauveria bassiana and graphene oxide-Beauveria bassiana nanocomposite in the infrared spectrum. The sample to be tested was mixed with potassium bromide (KBr) in a certain proportion and ground evenly in an agate mortar with a grinding particle size of less than 2μm, placed in a die mold, and pressed into a transparent film with a hydraulic press for measurement. The measurement temperature was room temperature and the scanning wavelength range was 4000-500cm -1 , resolution 2cm -1 .

[0066] 1.2.3 Results and Analysis

[0067] like Figure 7 As shown, the infrared spectrum comparison of the sample before and after the composite of graphene oxide No. 1 and Beauveria bassiana. By comparing the infrared spectra before and after the composite, it can be clearly found that the characteristic peaks of the composite sample (nanocomplex, blue solid line) and the spectrum of the biological pesticide (Beauveria bassiana, red solid line) are relatively close, indicating that the composite sample already contains the components of Beauveria bassiana, so the graphene oxide sample No. 1 is successfully composited with the biological pesticide (Beauveria bassiana).

[0068] like Figure 8 As shown, the infrared spectrum comparison of the sample before and after the composite of graphene oxide No. 2 and Beauveria bassiana. By comparing the infrared spectra before and after the composite, it can be clearly found that the characteristic peaks of the composite sample (nanocomplex, blue solid line) and the spectrum of the biological pesticide (Beauveria bassiana, red solid line) are relatively close, indicating that the composite sample already contains the components of Beauveria bassiana, so the graphene oxide sample No. 2 is successfully composited with the biological pesticide (Beauveria bassiana).

[0069] like Fig. 9 As shown, the infrared spectrum comparison of the sample before and after the composite of graphene oxide No. 3 and Beauveria bassiana. By comparing the infrared spectra before and after the composite, it can be clearly found that the characteristic peaks of the spectrum of the composite sample (nanocomplex, blue solid line) and the spectrum of the biological pesticide (Beauveria bassiana, red solid line) are relatively close, indicating that the composite sample already contains the components of Beauveria bassiana, so the graphene oxide sample No. 3 is successfully composited with the biological pesticide (Beauveria bassiana).

[0070] Example 2 Nanomaterial Preparation and Performance Measurement

[0071] 2.1 Screening of optimal concentration ratio of nanocomplexes

[0072] 2.1.1 Configuration 1×10 8A spore suspension with a concentration of spores / ml (the solution is 0.05% Tween solution) was prepared with an ultrasonic cleaner. A nanomaterial dispersion with a concentration of 100 μg / ml (the solution is ddH2O) was prepared. The mixture was mixed at a volume ratio of 1:1, 2:1, 4:1, and 6:1, respectively. After stirring with a magnetic stirrer at room temperature for 24 hours, 500 μl of the composite solution was pipetted with a pipette tip into a standard culture medium for Beauveria bassiana (2% corn flour, 1% wheat bran, 0.5% peptone, 0.3% potassium dihydrogen phosphate, 0.1% magnesium sulfate heptahydrate, 0.1% ammonium nitrate, 2% agar, and 0.5% cicada shells). The growth and status were observed every 6 hours.

[0073] 2.1.2 Results Analysis

[0074] according to Fig.10 As shown, graphene oxide has a protective and sustained-release effect on Beauveria bassiana, taking into account the economic benefits (the input ratio of Beauveria bassiana needs to be greater than that of nanomaterials) and the biological benefits (when the volume ratio of Beauveria bassiana spore suspension and graphene oxide dispersion is greater than 4:1, the nanomaterial inhibits the growth of Beauveria bassiana), so the optimal ratio concentration is when the input ratio of Beauveria bassiana spore suspension to graphene oxide dispersion is 4:1.

[0075] 2.2 Determination of thermal stability of nanocomposites

[0076] 2.2.1 Use an ultrasonic cleaner to prepare a nanomaterial dispersion with a concentration of 100 μg / ml and 1×10 8 The spore suspension of spores / ml was mixed at a ratio of 4:1 and stirred with a magnetic stirrer at room temperature for 24 hours, and then put into the spore germination solution prepared in advance and sterilized at high temperature (taking 1L as an example, 4% glucose and 1% fermentation solution were added and mixed evenly, and then put into a 50mL conical flask sterilized at high temperature and high pressure, sealed with a sealing film, and the temperature gradient was set to 25°C, 30°C, 35°C, and 40°C, respectively. The mixture was shaken and cultured continuously on a shaker at 180r / min for 18 hours. An appropriate amount of the mixed solution after shaking and culture was placed on a hemocytometer, and the spore germination was observed under a microscope. The germination was considered to be occurred when the length of the germ tube exceeded half of the spore diameter. At least 500 spores were randomly selected each time and counted three times.

[0077] 2.2.2 Result analysis:

[0078] Table 1. Analysis of the significant differences between nanocomplexes and spore suspensions on germination rates at different temperatures

[0079]

[0080] Depend on Fig.11It can be seen that the nanocomposite made of graphene oxide No. 1 has a protective effect on the spores of Beauveria bassiana at high temperatures. The activity of the No. 1 complex is not sensitive to temperature, and the curve in the figure is relatively stable. It can be seen from Table 1 that when the temperature is greater than 25°C, the single factor analysis shows that the protective effect of graphene oxide No. 1 on the spores of Beauveria bassiana is significantly different from that of the spore suspension (Tukey's HSD, P<0.05). Graphene oxide No. 1 is an ideal raw material for root irrigation preparations.

[0081] 2.3 Photostability of nanocomposites

[0082] 2.3.1 Test instrument: Qilin Bell three-purpose ultraviolet analyzer ZF-1

[0083] 2.3.2 Use an ultrasonic cleaner to prepare a nanomaterial dispersion with a concentration of 100 μg / ml and prepare 1×10 8 The spore suspension of spores / ml, the spore suspension of Beauveria bassiana and the graphene oxide dispersion were mixed in a ratio of 4:1 and stirred with a magnetic stirrer at room temperature for 24 hours, then placed in a centrifuge tube, immersed in 4°C water, placed on an ultraviolet analyzer, irradiated with 254nm ultraviolet light, and sampled at 15min, 30min, 1h, 2h, 4h, and 6h, respectively, and added to the spore germination solution, sealed with a sealing film, and continuously shaken and cultured on a shaker at 26°C and 180r / min for 18h. Take an appropriate amount of the mixed solution after shaking and culture on a hemocytometer, observe the spore germination under a microscope, and consider it to be germinated when the length of the germ tube exceeds half of the spore diameter. At least 500 spores are randomly selected each time and counted three times.

[0084] 2.3.3 Results Analysis

[0085] Table 2. Analysis of the significant differences in the spore inactivation rate of the nanocomplex and spore suspension over time under ultraviolet irradiation

[0086]

[0087] Depend on Fig.12 As shown in Table 2, the three graphene oxide materials all have a certain UV protection effect on Beauveria bassiana spores, among which the protective effect of graphene oxide No. 2 is the most obvious. Combined with the single factor difference significance analysis, except that the No. 1 compound has no obvious protective effect compared with the spore suspension after UV irradiation for 15 minutes, the No. 2 compound and the No. 3 compound are significantly better than the untreated suspension during the whole light irradiation process (Tukey's HSD, P < 0.05). Therefore, the UV protection function of the three graphene oxides on spores is ranked from strong to weak: No. 2 graphene oxide > No. 3 graphene oxide > No. 1 graphene oxide. Graphene oxide No. 2 and No. 3 graphene oxide can be used as ideal raw materials for spraying nanopesticides.

[0088] Example 3 Determination of the biological activity of nanocomplexes

[0089] 3.1 Test materials: graphene oxide No. 1 (material No. 1), graphene oxide No. 2 (material No. 2), graphene oxide No. 3 (material No. 3), Beauveria bassiana;

[0090] 3.2 Nanocomposite preparation: Prepare a nanocarrier dispersion with a concentration of 100 μg / ml using an ultrasonic cleaner; prepare 1×10 8 A spore suspension of Beauveria bassiana spores / ml, a Beauveria bassiana spore suspension and a graphene oxide dispersion were mixed in a ratio of 4:1 and then stirred with a magnetic stirrer at room temperature for 24 hours to prepare three aqueous nanocomposites (composite No. 1, complex No. 2, and complex No. 3);

[0091] The leaves and backs of pepper plants with more than three true leaves were sprayed evenly with the aqueous solution of the three nanocomposites (composite No. 1, complex No. 2, and complex No. 3), spore suspension (spore suspension of Beauveria bassiana), and 0.05% Tween solution. After spraying, a layer of plastic film was gently covered to increase the humidity and ensure the colonization rate. One week after spraying, the colonized leaves (the Tween treatment group was not colonized) were picked, and a layer of filter paper was laid flat on the bottom of the culture dish, and distilled water was sprayed to keep it moist. A fresh and clean leaf with the reverse side facing up (the leaves were of the same size) was placed on the filter paper, and the petiole of the leaf was wrapped with wet absorbent cotton to extend and keep the fresh time of the leaf. The western flower thrips adults were attached to the leaves using a fluke tube, and 20 thrips adults were attached to each dish, with 5 replicates. The lids were sealed with plastic wrap with fine pinholes. After sealing, the leaves were placed in an artificial climate box (temperature 25±1℃, relative humidity 60%±5%, photoperiod 16L:8D) for breeding. Use a small spray bottle to spray an appropriate amount of distilled water around the filter paper to keep the leaves moist every day. Observe for seven consecutive days and record the number of dead insects every day. Check the death of western flower thrips every 24 hours after the treatment for seven consecutive days. Divide it into death (gently stimulate the insect body with the tip of a brush, and the inactive one is recorded as dead) and survival (gently stimulate the insect body with the tip of a brush and it can crawl freely), and record them separately.

[0092] Table 3. Analysis of the significance of the effects of nanocomplex and spore suspension on the survival rate of western flower thrips

[0093] Survival rate (%) Complex No. 1 3.00±6.71a Complex II 0.00±0.00c Complex III 0.00±0.00b Spore suspension 11.00±13.42a

[0094] The bioassay process Fig.13As shown in Table 3, the three nanocomposites all showed faster insecticidal speed than the spore suspension from the time of application. The first compound had the fastest effect in the first two days, and the second and third compounds had better lethal effects on western flower thrips after the second day. The biological test results are shown in Table 3. Except for the first compound, which had no significant difference, the second and third compounds could achieve significant insect control effects (Tukey's HSD, P<0.05), which was superior to the traditional application method of Beauveria bassiana in terms of biological benefits.

Claims

1. Application of graphene oxide in loading biological pesticides, characterized in that: The graphene oxide and the biological pesticide form a nanocomposite, wherein the biological pesticide is Beauveria bassiana; a graphene oxide dispersion with a concentration of 100 μg / ml is prepared; and 1×10 8 The invention discloses a spore suspension of Beauveria bassiana spores / ml, wherein the volume ratio of the spore suspension of the biological pesticide to the graphene oxide dispersion is 3-5:1, and the graphene oxide is material No. 1 or material No.

3. The material No. 1 is a graphene oxide dispersion with an average radial size of 20-30 μm and a thickness of 1 nm; the material No. 3 is a graphene oxide solid with a purity of 99%, a flake diameter of 0.5-5 μm and a thickness of 0.8-1.2 nm.

2. The use according to claim 1, characterized in that: The volume ratio of the biological pesticide to graphene oxide is 4:

1.

3. The use according to claim 1, characterized in that: The preparation method of the nanocomposite composed of graphene oxide and biological pesticide comprises: preparing a graphene oxide dispersion; preparing a fungal spore suspension; mixing the fungal spore suspension and the graphene oxide dispersion in a volume ratio of 4:1, stirring the mixture on a magnetic stirrer at room temperature in the dark, and preparing an aqueous solution of the nanocomposite.

4. The use according to claim 3, characterized in that: The third material, solid graphene oxide, is crushed in sterilized ultrapure water using an ultrasonic cleaner to prepare a graphene oxide dispersion.

5. A nanocomposite comprising a biopesticide and graphene oxide, characterized in that: The biological pesticide is Beauveria bassiana, and a graphene oxide dispersion with a concentration of 100 μg / ml is prepared; 1×10 8 The invention discloses a spore suspension of Beauveria bassiana spores / ml, wherein the volume ratio of the spore suspension of the biological pesticide to the graphene oxide dispersion is 3-5:1, and the graphene oxide is material No. 1 or material No.

3. The material No. 1 is a graphene oxide dispersion with an average radial size of 20-30 μm and a thickness of 1 nm; the material No. 3 is a graphene oxide solid with a purity of 99%, a flake diameter of 0.5-5 μm and a thickness of 0.8-1.2 nm.

6. The nanocomposite according to claim 5, characterized in that: The Beauveria bassiana is strain GZGY-1-3.

7. The nanocomposite according to claim 5, characterized in that: The nanocomposite is configured into an aqueous solution, and the method includes: configuring a graphene oxide dispersion; configuring a Beauveria bassiana spore suspension; mixing the Beauveria bassiana spore suspension and graphene oxide in a volume ratio of 4:1, and then stirring and compounding them at room temperature to obtain an aqueous solution of the nanocomposite.

8. Application of nanocomposites in preventing and controlling agricultural pests, characterized in that: The nanocomposite is the nanocomposite according to any one of claims 5-7.

9. The use according to claim 8, characterized in that: The aqueous solution of the compound made of material No. 1 is applied to crops by root irrigation to complete the colonization of Beauveria bassiana, or the aqueous solution of the compound made of material No. 3 is applied to crops by spraying to complete the colonization of Beauveria bassiana.

10. A method for preparing a nanocomposite according to any one of claims 5 to 7, characterized in that: include: Configuration 1×10 8 A spore suspension of Beauveria bassiana spores / ml; a graphene oxide dispersion with a concentration of 100 μg / ml, the Beauveria bassiana spore suspension and the graphene oxide dispersion are mixed in a volume ratio of 4:1, and then stirred and compounded at room temperature to obtain an aqueous solution of a nanocomposite, and the nanocomposite powder is obtained after drying.

11. A method for spraying and controlling agricultural pests, characterized in that: The method comprises: using the aqueous solution prepared by the nanocomposite of claim 5; spraying it on the leaves of crops to complete the colonization of Beauveria bassiana; the colonized leaves of crops have contact killing and / or stomach poisoning effects on agricultural pests.

12. The control method according to claim 11, characterized in that: The spraying amount is 6-10 ml of the nanocomposite aqueous solution per crop.

13. The control method according to claim 12, characterized in that: 8ml of nanocomplex solution per crop.

14. A root irrigation control method for agricultural pests, characterized in that: The method comprises: using the aqueous solution prepared by the nanocomposite of claim 5; irrigating along the roots of crops for three consecutive days to complete the colonization of Beauveria bassiana.

15. The control method according to claim 14, characterized in that: Each crop is irrigated with 15-25 ml of the nanocomposite aqueous solution at a time.

16. The control method according to claim 14, characterized in that: Each crop was irrigated with 20 ml of nanocomposite solution at the root each time.

17. The control method according to claim 11 or 14, characterized in that: The crops include cereal crops, legume crops and / or vegetables.

18. The control method according to claim 17, characterized in that: The crops include one or more of the following: wheat, rice, corn, soybean, broad bean, pea, mung bean, tomato, eggplant, kidney bean, cowpea, and pepper.

19. The control method according to claim 11 or 14, characterized in that: The pests are corn pests.

20. The control method according to claim 19, characterized in that: The pests include one or more of the following: corn borer, fall armyworm, thrips, small cutworm, yellow cutworm, mole cricket, grub, wireworm, corn sunflower mealybug, corn spinner, grass borer, rice leaf roller, Spodoptera litura, corn armyworm, armyworm, yellow-bellied lantern moth, red-margined lantern moth, brown-footed horned leaf beetle, spotted stink bug, red-barbed blind bug, pasture blind bug, three-spotted leafhopper, large green leafhopper, corn aphid, truncate spider mite, cinnabar spider mite, Asian corn borer, large borer, peach borer, cotton bollworm, white-star flower beetle, small blue flower beetle, two-spotted leaf beetle, and green rice stink bug.

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