Application of nano-silver synthesized by extract of Alhagi sparsifolia shap in preparation of inhibitor for root rot of kidney bean
Nano-silver particles synthesized from camel thorn extract have solved the problem of controlling bean root rot, achieving efficient and environmentally friendly disease control, and are suitable for inhibiting bean root rot pathogens.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- 浙江省植保检疫与农药管理总站
- Filing Date
- 2024-04-11
- Publication Date
- 2026-07-03
Smart Images
Figure CN118556712B_ABST
Abstract
Description
Technical Field
[0001] This invention belongs to the field of biosynthetic nanomaterials technology, specifically relating to the application of nano-silver synthesized from camel thorn extract in the preparation of inhibitors for root rot fungi of common bean. Background Technology
[0002] Common bean (Phaseolus vulgaris L.), also known as string bean, is an annual herbaceous plant belonging to the genus Phaseolus in the legume family. It is high in protein and rich in nutrients, making it a popular food. Root rot is a common soil-borne disease in common bean production, severely impacting yield. Root rot affects the taproot and underground stem. Symptoms in seedlings include dark brown, sunken lesions, browning of the vascular bundles in the roots and stems, and damping-off. Infection during the flowering and pod-setting stage leads to stunted growth, with varying sizes of blight spots appearing on the lower edges of leaves. Leaves gradually turn yellow from the bottom up but do not fall off. In severe cases, the entire plant wilts and dies. In humid weather, pink or white mold may appear at the base of the stem of diseased plants.
[0003] Common bean root rot is mainly caused by four pathogens: *Macrophomina phaseolina*, *Pythium graminicola*, *Rhizoctonia solani*, and *Sclerotium rolfsii*. The pathogens overwinter primarily as mycelium or spores in diseased plant debris. The following year, under suitable conditions, the spores are spread by wind, rain, or agricultural operations, entering the plant through wounds or stomata on the host surface, causing disease. Current control techniques mainly include agricultural and chemical control. Agricultural control involves selecting superior varieties, crop rotation, eliminating diseased plant debris, proper planting density, and good water and fertilizer management. Chemical control involves disinfecting seeds and soil with fungicides and treating bean plants with pesticides.
[0004] Using biosynthesized nanomaterials to prevent and control pathogens offers advantages such as high safety, good stability, strong bactericidal ability, and environmental friendliness. For example, Chinese patent document CN116921688A discloses the inhibitory effect of nano-silver prepared from peach leaf extract on Haffnia bacteria. Haffnia is a common spoilage bacterium found in vacuum-packed and refrigerated meat products. Nano-silver prepared from peach leaf extract can inhibit Haffnia biofilm formation, inhibit the production of extracellular proteases, and inhibit the production of extracellular polysaccharides, showing high application potential in aquaculture, storage, and transportation. Chinese patent document CN111685129A discloses the antibacterial effect of a complex of phenylethanol glycoside extract of Callicarpa nudiflora and nano-silver, which has a good inhibitory effect on Staphylococcus aureus and four yeasts (Candida albicans, Candida tropicalis, Candida krusei, and Candida parapsilosis).
[0005] Therefore, it is necessary to develop a nanomaterial synthesized using biological methods to prevent and control root rot in common beans. Summary of the Invention
[0006] This invention provides the application of nano-silver synthesized from camel thorn extract in the preparation of inhibitors for bean root rot. The method of synthesizing nano-silver using camel thorn extract is green and environmentally friendly, with simple and efficient steps. The nano-silver product has uniform size, excellent stability and dispersibility, and has good application prospects in the preparation of inhibitors for bean root rot.
[0007] The specific technical solution adopted is as follows:
[0008] The application of silver nanoparticles synthesized from camel thorn extract in the preparation of inhibitors for root rot of common bean is described. Specifically, the method for synthesizing silver nanoparticles using camel thorn extract includes the following steps:
[0009] (1) After washing the camel thorn leaves, drying and crushing them, mixing them with deionized water, heating them in a water bath, and filtering them after standing, camel thorn extract was obtained.
[0010] (2) Camel thorn extract was mixed with AgNO3 solution and reacted in the dark to obtain a mixture. After centrifugation, washing and vacuum freeze-drying, nano silver was obtained.
[0011] The average particle size of the silver nanoparticles is 2-25 nm.
[0012] Camel thorn (Alhagi graecorum) is a deciduous herb belonging to the genus Alhagi in the subfamily Papilionoideae of the legume family. It is a commonly used traditional Chinese medicine. It is warm in nature, sour and sweet in taste, and has nourishing, thirst-quenching, analgesic, and irritability-relieving effects. It is often used to treat diarrhea, abdominal pain, headaches, and other ailments. Camel thorn also possesses anti-inflammatory and anti-tumor biological activities.
[0013] Camel thorn aqueous extract contains a variety of bioactive components, which can act as a reducing agent and stabilizer in the process of reacting with silver nitrate.
[0014] The aforementioned bean root rot pathogen includes at least one of Macropomina phaseolina, Pythium graminicola, Rhizoctonia solani, and Sclerotium rolfsii.
[0015] Preferably, the ratio of crushed camel thorn leaves to deionized water is 1g:10-20mL. This preferred ratio facilitates the extraction of bioactive components from the camel thorn leaves.
[0016] Preferably, the water bath heating temperature is 60-70℃ and the water bath heating time is 50-70 minutes.
[0017] Preferably, in step (2), the camel thorn extract is mixed with AgNO3 solution and reacted in the dark at a temperature of 60-70℃ and a rotation speed of 500-700 rpm. The reaction is terminated when the color of the mixture turns reddish-brown. If the temperature or rotation speed is set too low, the reaction will be insufficient and the synthesis effect will be poor, which will ultimately affect the particle size, stability and uniformity of the silver nanoparticles, thus affecting their antibacterial effect. If the temperature or rotation speed is set too high, the synthesized product will easily deteriorate, and unnecessary energy consumption will also occur.
[0018] Further preferably, the concentration of the AgNO3 solution is 2-4 mM, and the volume ratio of camel thorn extract to AgNO3 solution is 3-4:7. Excessive content of the effective components of the camel thorn extract in the system will affect the formation of silver nanoparticles, while excessively low content will lead to a decrease in the efficiency of the final product synthesis.
[0019] Preferably, in step (2), the centrifugation conditions are 10000-14000g for 10-20min.
[0020] The nano-silver synthesized using camel thorn extract is structurally stable, nearly spherical, uniform in size, and small in diameter. It exhibits strong inhibitory effects on four root rot pathogens of bean: *M. phaseolina*, *P. graminicola*, *R. solani*, and *S. rolfsii*. This significantly enhances the control efficacy of the nano-silver particles against plant diseases caused by these pathogens, demonstrating a wide range of potential applications.
[0021] This invention also provides the application of nano-silver synthesized from camel thorn extract in the prevention and control of plant diseases caused by bean root rot fungus.
[0022] Specifically, the plant diseases mentioned include bean root rot.
[0023] The present invention also provides an inhibitor of bean root rot fungus, comprising nano-silver synthesized from the camel thorn extract.
[0024] Preferably, the inhibitor of *Pseudolaric acid causal agent* is an aqueous solution of nano-silver synthesized from camel thorn extract, with the concentration of nano-silver preferably being 25-100 μg / mL. Within this preferred range, the nano-silver aqueous solution synthesized from camel thorn extract exhibits excellent inhibitory effects against *Pseudolaric acid causal agent*. Further increasing the inhibitor concentration does not significantly improve its bactericidal effect.
[0025] Compared with the prior art, the beneficial effects of the present invention are as follows:
[0026] (1) The present invention uses camel thorn leaves as raw material to synthesize nano silver particles with small average particle size, uniform size and stable structure. It can effectively inhibit the activity of four root rot fungi of bean, namely M. phaseolina, P. graminicola, R. solani and S. rolfsii, and prevent and control the plant disease bean root rot caused by them. It has a strong bactericidal effect.
[0027] (2) The process of synthesizing nano-silver from camel thorn extract is simple, low-cost, consumes relatively little energy and does not produce toxic byproducts. It does not require the addition of reducing agents and catalysts, is green and environmentally friendly, and the synthesized nano-silver can be used as an effective ingredient in the inhibitor of bean root rot fungus.
[0028] (3) The nano-silver particles synthesized from camel thorn extract are used as inhibitors of bean root rot fungus. They have a good prevention and control effect. They will not only not cause potential harm to the human body, but also will not pollute the environment. The application method is simple and easy to implement, and it has a promising prospect for promotion in the field of agricultural production. Attached Figure Description
[0029] Figure 1 The image shows the UV-Vis spectrum of the AgNPs synthesized in Example 1.
[0030] Figure 2 The image shows the Fourier transform infrared spectrum of the AgNPs synthesized in Example 1.
[0031] Figure 3 The image shows the X-ray diffraction pattern of the AgNPs synthesized in Example 1.
[0032] Figure 4This is a transmission electron microscope image of the AgNPs synthesized in Example 1.
[0033] Figure 5 This is a scanning electron microscope image of the AgNPs synthesized in Example 1.
[0034] Figure 6 The image shows the EDS plot of the AgNPs synthesized in Example 1 and the corresponding analysis results.
[0035] Figure 7 The graph shows the inhibitory effects of different concentrations of AgNPs solutions on the growth of four pathogens in Example 1. In the graph, A is a statistical graph of mycelial inhibition rate, and B is a graph of colony growth under different treatments. Detailed Implementation
[0036] The present invention will be further illustrated below with reference to the embodiments and accompanying drawings. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention.
[0037] Example 1
[0038] (1) Wash and dry the freshly picked camel thorn leaves, crush them in a mortar and pestle, weigh 5g and mix them with 100mL of deionized water, place them in a water bath and heat at 60℃ for 50 minutes. During this process, stir intermittently with a glass rod, let stand, and then filter through cotton cloth and filter paper to ensure that solid impurities are removed, thus obtaining camel thorn extract.
[0039] (2) Take 30 mL of camel thorn extract obtained in step (1) and mix it with 70 mL of AgNO3 solution with a concentration of 2 mM. Place it in a shaker at a temperature of 60℃ and a speed of 500 rpm to react in the dark. Stop the reaction when the color of the mixture turns reddish brown. After taking it out, centrifuge the mixture at a speed of 10000 g for 10 minutes, remove the supernatant, take the precipitate, wash it three times with ultrapure water, and freeze dry it under vacuum to obtain powdered nano silver (denoted as AgNPs).
[0040] Example 2
[0041] (1) Wash and dry the freshly picked camel thorn leaves, crush them in a mortar and pestle, weigh 5g and mix them with 70mL of deionized water, place them in a water bath and heat at 65℃ for 60 minutes. During this process, stir intermittently with a glass rod, let stand, and then filter through cotton cloth and filter paper to ensure that the solid impurities are removed, thus obtaining camel thorn extract.
[0042] (2) Take 35 mL of camel thorn extract obtained in step (1) and mix it with 70 mL of AgNO3 solution with a concentration of 3 mM. Place it in a shaker at a temperature of 65℃ and a speed of 600 rpm to react in the dark. Stop the reaction when the color of the mixture turns reddish brown. After taking it out, centrifuge the mixture at a speed of 12000 g for 15 minutes, remove the supernatant, take the precipitate, wash it three times with ultrapure water, and freeze dry it under vacuum to obtain powdered nano silver (denoted as AgNPs).
[0043] Example 3
[0044] (1) Wash and dry the freshly picked camel thorn leaves, crush them with a mortar and pestle, weigh 5g and mix them with 50mL of deionized water, place them in a water bath and heat at 70℃ for 70 minutes. During this process, stir intermittently with a glass rod, let stand, and then filter through cotton cloth and filter paper to ensure that the solid impurities are removed, thus obtaining camel thorn extract.
[0045] (2) Take 40 mL of camel thorn extract obtained in step (1) and mix it with 70 mL of AgNO3 solution with a concentration of 4 mM. Place it in a shaker at a temperature of 70℃ and a speed of 700 rpm to react in the dark. Stop the reaction when the color of the mixture turns reddish brown. After taking it out, centrifuge the mixture at a speed of 14000 g for 20 minutes, remove the supernatant, take the precipitate, wash it three times with ultrapure water, and freeze dry it under vacuum to obtain powdered nano silver (denoted as AgNPs).
[0046] Sample Analysis
[0047] (I) Performance Characterization
[0048] Observation of the reaction process revealed that after adding camel thorn extract to a transparent AgNO3 solution and placing it on a shaker in the dark, the color changed from the initial orange-yellow to reddish-brown.
[0049] The structural characteristics and physicochemical properties of the AgNPs prepared in Example 1 were further evaluated using characterization techniques such as ultraviolet-visible absorption spectroscopy (UV-VIS), Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), transmission electron microscopy (TEM), energy dispersive spectroscopy (EDS), and X-ray diffraction (XRD).
[0050] Figure 1 The UV-Vis spectroscopy results show that the AgNPs have the strongest absorption at 432 nm, indicating that the synthesized AgNPs are relatively stable. Figure 2 The Fourier transform infrared spectrum indicates that the AgNPs have been successfully synthesized.
[0051] Figure 3The X-ray diffraction pattern of AgNPs shows characteristic peaks (101), (111), (200), (220), and (311) at positions of 2θ = 28.81°, 31.98°, 37.85°, 46.03°, and 77.06°, respectively, proving that stable AgNPs were synthesized.
[0052] Figure 4 The image shows a transmission electron microscope image of AgNPs, which reveals that AgNPs are nearly spherical particles. Figure 5 The image shows a scanning electron microscope (SEM) image of the AgNPs, which reveals that the AgNPs are relatively uniform in size, with a particle size range of 4.02-21.90 nm.
[0053] Figure 6 The EDS elemental distribution diagram of AgNPs shows that the AgNPs are mainly composed of Ag, Si, and O elements. The Si element may have originated from the plant extract of camel thorn leaves.
[0054] (II) Effect of AgNPs concentration on antibacterial effect
[0055] To evaluate the inhibitory activity of the AgNPs prepared in Example 1 against the root rot fungi of bean (M. phaseolina, P. graminicola, R. solani, and S. rolfsii), the AgNPs prepared in Example 1 were prepared with water to form solutions with concentrations of 250, 500, 750, and 1000 μg / mL, respectively. The antibacterial performance of different AgNP concentrations was evaluated by measuring the mycelial growth inhibition rate.
[0056] 1 mL of AgNPs solutions with concentrations of 250, 500, 750, and 1000 μg / mL were added dropwise to 9 mL of PDA solution, mixed thoroughly, and poured into petri dishes. After cooling and air-drying, PDA petri dishes containing AgNPs solutions of 25, 50, 75, and 100 μg / mL were obtained. A slice of *Pseudostella apiacea* root rot pathogen was then inoculated into the center of each plate. Each treatment was repeated three times. The plates were placed in an incubator at 25°C and incubated for 5 days. Compared with the control group, the colony diameter on the plates with added AgNPs solution decreased with increasing concentration of nano-silver, reaching the highest inhibition rate at an AgNPs solution concentration of 100 μg / mL. These results indicate that within the concentration range of 25-100 μg / mL, higher AgNPs solution concentrations result in better antibacterial effects. However, further increases in concentration beyond 100 μg / mL do not significantly increase the antibacterial rate or enhance the antibacterial effect.
[0057] The results are as follows Figure 7As shown in A, the higher the concentration of AgNPs solution, the better its antibacterial effect. AgNPs synthesized using camel thorn can significantly inhibit the growth of the root rot fungi of bean, including M. phaseolina, P. graminicola, R. solani, and S. rolfsii.
[0058] (III) Antibacterial effect of AgNPs
[0059] To further demonstrate the inhibitory effect of the AgNPs prepared in Example 1 on the growth of *M. phaseolina*, *P. graminicola*, *R. solani*, and *S. rolfsii*, PDA culture dishes containing 100 μg / mL AgNPs solution were prepared according to the above experimental method. The experiment was repeated three times. Figure 7 As shown in B, the experimental results show that after treating *M. phaseolina*, *R. solani*, *S. rolfsii*, and *P. graminicola* with a 100 μg / mL AgNPs solution, the growth inhibition rates of the colonies reached 73.33%, 77.78%, 92.95%, and 93.33%, respectively.
[0060] The embodiments described above provide a detailed explanation of the technical solutions of the present invention. It should be understood that the above descriptions are merely specific embodiments of the present invention and are not intended to limit the present invention. Any modifications, additions, or similar substitutions made within the scope of the principles of the present invention should be included within the protection scope of the present invention.
Claims
1. The application of nano-silver synthesized from camel thorn extract in the preparation of inhibitors for root rot fungi of common bean, characterized in that, The method for synthesizing nano-silver using camel thorn extract includes the following steps: (1) After washing and drying the camel thorn leaves, crush them, mix them with deionized water, heat them in a water bath, let them stand and filter to obtain camel thorn extract; the ratio of crushed camel thorn leaves to deionized water is 1 g: 10-20 mL; the water bath heating temperature is 60-70 ℃ and the water bath heating time is 50-70 min. (2) The camel thorn extract was mixed with AgNO3 solution and reacted in the dark at a temperature of 60-70 ℃ and a rotation speed of 500-700 rpm. The reaction was terminated when the color of the mixture turned reddish-brown. The resulting mixture was centrifuged, washed, and freeze-dried under vacuum to obtain nano-silver. The concentration of AgNO3 solution was 2-4 mM, and the volume ratio of camel thorn extract to AgNO3 solution was 3-4:
7. The average particle size of the silver nanoparticles is 2-25 nm; The aforementioned bean root rot pathogen is *Cyclocarya beanica*. Macrophomina phaseolina Phytophthora Pythium graminicola Rhizoctonia solani Rhizoctonia solani Neat and orderly small sclerotium Sclerotium rolfsii At least one of them.
2. The application of the nano-silver synthesized from camel thorn extract according to claim 1 in the preparation of an inhibitor for root rot of common bean, characterized in that, In step (2), the centrifugation conditions are 10000-14000 g for 10-20 min.
3. The application of nano-silver synthesized from camel thorn extract in the prevention and control of plant diseases caused by *Pseudomonas aeruginosa* root rot, characterized in that... The method for synthesizing nano-silver using camel thorn extract includes the following steps: (1) After washing and drying the camel thorn leaves, crush them, mix them with deionized water, heat them in a water bath, let them stand and filter to obtain camel thorn extract; the ratio of crushed camel thorn leaves to deionized water is 1 g: 10-20 mL; the water bath heating temperature is 60-70 ℃ and the water bath heating time is 50-70 min. (2) The camel thorn extract was mixed with AgNO3 solution and reacted in the dark at a temperature of 60-70 ℃ and a rotation speed of 500-700 rpm. The reaction was terminated when the color of the mixture turned reddish-brown. The resulting mixture was centrifuged, washed, and freeze-dried under vacuum to obtain nano-silver. The concentration of AgNO3 solution was 2-4 mM, and the volume ratio of camel thorn extract to AgNO3 solution was 3-4:
7. The average particle size of the silver nanoparticles is 2-25 nm; The aforementioned bean root rot pathogen is *Cyclocarya beanica*. Macrophomina phaseolina Phytophthora Pythium graminicola Rhizoctonia solani Rhizoctonia solani Neat and orderly small sclerotium Sclerotium rolfsii At least one of them.
4. The application of nano-silver synthesized from camel thorn extract according to claim 3 in the prevention and control of plant diseases caused by *Pseudomonas aeruginosa* root rot, characterized in that... The plant diseases mentioned include bean root rot.