Method for preparing silver nanoparticles from water extract of caulis microspheraeae and product thereof

Abstract

The application discloses a method for preparing silver nanoparticles by using a morinda officinalis water extract and a product, and relates to the technical field of preparing nanometer materials by using a biological water extract, and the method comprises the following steps: after morinda officinalis is crushed and dried, the morinda officinalis is added into ultrapure water and heated for leaching, the supernatant is obtained after cooling and centrifugation, and a morinda officinalis stock solution is obtained; the morinda officinalis stock solution is diluted; the pH value of the morinda officinalis diluent is adjusted to 8.0-11.0; the diluent with the adjusted pH value is mixed with a silver nitrate solution according to a material-liquid ratio of 3-1:1; ultrasonic reaction is carried out at 25 DEG C-85 DEG C for 1 min-120 min, and a silver nanoparticle crude solution is obtained; the crude solution is left to stand to precipitate, and the precipitate is washed and dried to obtain silver nanoparticles. The application uses morinda officinalis as a raw material, only uses water and a silver nitrate reagent, no toxic chemical reducing agent and stabilizer is added in the whole process, the reaction condition is mild, and the method is suitable for large-scale production; and the prepared silver nanoparticles have a particle size of 5-10 nm, are spherical, uniformly dispersed and good in stability.

Description

Technical Field

[0001] This invention relates to the field of nanomaterials preparation technology from biological aqueous extracts, and in particular to a method and product for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus*. Background Technology

[0002] Nano-silver (AgNPs) refers to elemental silver particles with a nanoscale size, typically smaller than 100 nm. Nano-silver can inhibit various pathogenic microorganisms and even kill these bacteria. Silver nanoparticles possess advantages such as extremely small particle size, good electrical conductivity, excellent antibacterial properties, and high specific surface area, making them promising for applications in medicine, hygiene, electronics, catalytic materials, and electroplating industries. They are commonly used for sterilization of medical devices, preparation of antibacterial drugs, and the production of high-end silver pastes, conductive coatings, and heat exchange materials. Currently, methods for preparing silver nanoparticles are mainly divided into three categories: physical methods, chemical methods, and biological methods. Physical methods rely on large-scale equipment such as vacuum evaporation and laser evaporation, which suffer from high energy consumption, high preparation costs, and difficulty in mass production. Chemical methods mainly reduce silver ions using chemical reducing agents, requiring the addition of various chemical reagents such as sodium citrate and PVP. This not only involves complex processes but also leaves chemical residues that can cause environmental pollution. Furthermore, the prepared silver nanoparticles suffer from severe aggregation, uneven particle size, and limited biological activity, significantly restricting their application in high-end fields such as biomedicine and food.

[0003] Phytosynthesis in biological methods has become a research hotspot due to its advantages such as readily available raw materials, simple processes, environmental friendliness, and good product dispersibility. Guangxi is rich in traditional Chinese medicinal herbs, many of which contain natural reducing substances such as polyphenols, flavonoids, reducing sugars, and polysaccharides, which can be used as reducing agents and stabilizers to reduce silver ions and prepare silver nanoparticles. While there are reports on the use of various plant extracts for the synthesis of silver nanoparticles, a standardized preparation scheme with clear process parameters and excellent product bioactivity, using *Spatholobus suberectus* aqueous extract as the core raw material, has not yet been established. Furthermore, conventionally prepared silver nanoparticles suffer from uneven particle size and insufficient antioxidant and antibacterial properties. Therefore, developing a stable method for preparing silver nanoparticles using *Spatholobus suberectus* aqueous extract as a raw material, with products exhibiting excellent comprehensive performance, has significant practical application value. Summary of the Invention

[0004] The purpose of this invention is to provide a method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus*. This method is simple, green, pollution-free, and has mild reaction conditions with controllable preparation parameters. It also provides a silver nanoparticle product obtained by this method, which has uniform particle size, good dispersibility, and excellent antioxidant and antibacterial properties.

[0005] To achieve the above objectives, the technical solution adopted by the present invention is as follows: Its preparation method includes the following steps: (1) After the dried chicken blood vine is crushed and sieved, it is added to ultrapure water and heated for extraction. After cooling and centrifugation, the supernatant is taken to obtain the original chicken blood vine solution. Then the original chicken blood vine solution is diluted to obtain the diluted chicken blood vine solution. (2) Adjust the pH of the chicken blood vine dilution obtained in step (1) to 8.0 to 11.0 using an alkaline solution. Mix the adjusted chicken blood vine dilution with silver nitrate solution evenly, control the material-liquid ratio to be 3:1 to 1:1, and sonicate at 25℃ to 85℃ for 1 min to 120 min to obtain crude silver nanoparticle solution. (3) The crude liquid of silver nanoparticles obtained in step (2) is allowed to stand to precipitate, the precipitate is separated, washed and dried to obtain silver nanoparticles.

[0006] In the above technical solution, a more specific solution can be: in step (1), the chicken blood vine is crushed and passed through a 60-mesh sieve; 200mL of ultrapure water is added to every 10g of crushed chicken blood vine for extraction, and the extraction conditions are heating to a slight boiling state and extraction time is 30 min.

[0007] In some possible implementations, in step (1), the centrifugation speed is 5000 r / min and the centrifugation time is 15 min; the original chicken blood vine solution is diluted 7 times with ultrapure water to obtain a diluted chicken blood vine solution with a pH value of 7.0.

[0008] In some possible implementations, the alkalinity regulator is a 10 mol / L aqueous solution of sodium hydroxide.

[0009] In some possible implementations, in step (2), the pH value of the diluted chicken blood vine solution obtained in step (1) is adjusted to 8.0 using sodium hydroxide aqueous solution. The diluted chicken blood vine solution with adjusted pH value is mixed evenly with silver nitrate solution, and the material-liquid ratio is controlled to be 1:1. The mixture is ultrasonically reacted at 25°C for 10 min to obtain crude silver nanoparticle solution.

[0010] In some possible implementations, in step (3), the concentration of the silver nitrate solution is 0.01 mol / L.

[0011] In some possible implementations, in step (3), the precipitate is washed with ultrapure water, and the washing is performed 3 times; the drying temperature is 60°C, and the precipitate is dried to constant weight.

[0012] The present invention also provides silver nanoparticles of *Spatholobus suberectus* prepared by the method, wherein the silver nanoparticles have a particle size of 5-10 nm, a spherical morphology, and a face-centered cubic crystal structure.

[0013] By adopting the above technical solution, the present invention has the following advantages compared with the prior art: 1. The preparation method of the present invention uses the characteristic Chinese herbal medicine of Guangxi, Chicken Blood Vine, as raw material. It only uses water and silver nitrate as conventional reagents. No toxic chemical reducing agents or stabilizers are added throughout the process. The reaction conditions are mild and there is no pollution from waste. Chicken Blood Vine raw material is widely available and inexpensive, making it suitable for large-scale production.

[0014] 2. The silver nanoparticles prepared by this method have a particle size concentrated in the range of 5-10 nm, are spherical in shape, uniformly dispersed, and have a negatively charged surface, exhibiting strong stability. They can scavenge DPPH free radicals at a rate of over 80%, demonstrating outstanding antioxidant capacity. They also exhibit significant antibacterial and bactericidal effects against Escherichia coli and Staphylococcus aureus, with a minimum inhibitory concentration as low as 312.5 μg / mL and a minimum bactericidal concentration of 625 μg / mL. Their antibacterial performance is superior to that of silver nanoparticles prepared by traditional chemical methods.

[0015] 3. The four core parameters of reaction time, pH value, feed-liquid ratio and reaction temperature were optimized through single-factor experimental system, and the optimal process system was determined, resulting in stable product yield and good repeatability. Attached Figure Description

[0016] Figure 1 This is the UV-Vis absorption spectrum of the reaction between the aqueous extract of *Spatholobus suberectus* and silver nitrate at different times according to the present invention.

[0017] Figure 2 This is the UV-Vis absorption spectrum of the reaction between the aqueous extract of *Spatholobus suberectus* at different pH values ​​and silver nitrate according to the present invention.

[0018] Figure 3 This is the UV-Vis absorption spectrum of the reaction between the aqueous extract of *Spatholobus suberectus* and silver nitrate at different temperatures according to the present invention.

[0019] Figure 4 This is the UV-Vis absorption spectrum of the reaction between the aqueous extract of *Spatholobus suberectus* and silver nitrate under different material-to-liquid ratios according to the present invention.

[0020] Figure 5 This is a zeta potential diagram of the silver nanoparticles from the chicken blood vine of this invention.

[0021] Figure 6 This is a morphology image of the silver nanoparticles of *Spatholobus suberectus* prepared in Example 1 of this invention under a spherical aberration electron microscope.

[0022] Figure 7 This is the XRD diffraction pattern of the silver nanoparticles of *Chicken Blood Vine* according to the present invention.

[0023] Figure 8 This is a diagram illustrating the scavenging effect of the silver nanoparticles from *Spatholobus suberectus* on DPPH free radicals. Detailed Implementation

[0024] The present invention will be further described in detail below with reference to embodiments: Example 1

[0025] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: (1) Weigh 10.0g of chicken blood vine, crush it and pass it through a 60-mesh sieve. Add 200mL of ultrapure water to the crushed chicken blood vine for extraction. The extraction conditions are heating to a slight boiling state and extraction time of 30 min. After cooling, centrifuge at 5000r / min for 15 min and take the supernatant to obtain the original chicken blood vine solution. Dilute the original chicken blood vine solution with ultrapure water 7 times to obtain a diluted chicken blood vine solution with a pH of 7.0. (2) The pH value of the chicken blood vine dilution prepared in step (1) was adjusted by using 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution was adjusted to 8.0. The chicken blood vine dilution with adjusted pH value was mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 1:1. The mixture was ultrasonically reacted at 25℃ for 10 min to obtain crude silver nanoparticle solution. (3) The crude liquid of silver nanoparticles obtained in step (2) was allowed to stand to precipitate, the precipitate was separated and washed with ultrapure water three times, and then dried at 60°C until constant weight was obtained to obtain silver nanoparticles. Example 2

[0026] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 8.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 1:1. The mixture is ultrasonically reacted at 25℃ for 1 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1. Example 3

[0027] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 8.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 1:1. The mixture is ultrasonically reacted at 25℃ for 30 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1. Example 4

[0028] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 8.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 1:1. The mixture is ultrasonically reacted at 25℃ for 60 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1. Example 5

[0029] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 8.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 1:1. The mixture is ultrasonically reacted at 25℃ for 90 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1. Example 6

[0030] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 8.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 1:1. The mixture is ultrasonically reacted at 25℃ for 120 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1. Example 7

[0031] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 9.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 1:1. The mixture is ultrasonically reacted at 25℃ for 10 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1. Example 8

[0032] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 10.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 1:1. The mixture is ultrasonically reacted at 25℃ for 10 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1. Example 9

[0033] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 11.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 1:1. The mixture is ultrasonically reacted at 25℃ for 10 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1. Example 10

[0034] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 8.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 1:1. The mixture is ultrasonically reacted at 40℃ for 10 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1. Example 11

[0035] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 8.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 1:1. The mixture is ultrasonically reacted at 55℃ for 10 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1. Example 12

[0036] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 8.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 1:1. The mixture is ultrasonically reacted at 70℃ for 10 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1. Example 13

[0037] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 8.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 1:1. The mixture is ultrasonically reacted at 85℃ for 10 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1. Example 14

[0038] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 8.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 3:1. The mixture is ultrasonically reacted at 25℃ for 10 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1. Example 15

[0039] A method for preparing silver nanoparticles from an aqueous extract of *Spatholobus suberectus* includes the following steps: Step (1) is the same as step (1) in Example 1; Step (2) The pH value of the chicken blood vine dilution obtained in step (1) is adjusted with 10 mol / L NaOH solution. The pH value of the chicken blood vine dilution is adjusted to 8.0. The chicken blood vine dilution with adjusted pH value is mixed with 0.01 mol / L silver nitrate solution at a material-liquid ratio of 2:1. The mixture is ultrasonically reacted at 25℃ for 10 min to obtain crude silver nanoparticle solution. Step (3) is the same as step (3) in Example 1.

[0040] 1. Effect of reaction time on the formation of silver nanoparticles Take 20 ml of the diluted chicken blood vine solution prepared in step (1) of Example 1, add 20 ml of 0.01 mol / L silver nitrate solution, and sonicate at 25°C. Then, take the reaction solution with sonication time of 1 min, 10 min, 30 min, 60 min, 90 min, and 120 min respectively for ultraviolet spectral measurement. Figure 1 As shown, the diluted *Spatholobus suberectus* solution reacts with silver nitrate. Immediately after the addition of silver nitrate solution (1 minute), a characteristic absorption peak appears at 410 nm, indicating that the active ingredients in the *Spatholobus suberectus* extract can reduce silver nitrate to silver nanoparticles. Furthermore, the solution color changes from orange-yellow to brownish-black immediately upon addition of silver nitrate, indicating the formation of silver nanoparticles. With increasing reaction time, the absorbance at 410 nm gradually increases, indicating that longer ultrasonic time results in more silver nanoparticles. The absorbance change is greatest from the start to 10 minutes, indicating the fastest reaction rate. At 1 minute, the UV absorption peak is relatively broad, possibly due to insufficient reaction time. Between 10 and 30 minutes, 30 and 60 minutes, and 60 and 90 minutes, the reaction efficiency between the extract and silver nitrate decreases, slowing the formation of silver nanoparticles. The reaction rate is fastest at 10 minutes, and a larger number of silver nanoparticles are formed.

[0041] 2. Effect of pH on the formation of silver nanoparticles Take five 3 mL portions of the *Spatholobus suberectus* diluted solution prepared in step (1) of Example 1. Adjust the pH of the diluted solution to 8.0, 9.0, 10.0, and 11.0 respectively using 10 mol / L NaOH solution, and add 3 mL of 0.01 mol / L AgNO3 solution to each. Sonicate at room temperature for ten minutes. Use a UV spectrophotometer to detect whether nano-silver is formed during the synthesis process, and measure the intensity of the absorption peak at 410 nm. Select the most suitable pH by observing the magnitude of the absorbance value and the changes in the two absorbance curves.

[0042] like Figure 2 As shown, at pH 7.0, the absorbance value near 410 nm of the diluted *Spatholobus suberectus* solution is low, indicating a slow rate of silver nanoparticle formation. With increasing pH, the reaction rate accelerates, and the number of silver nanoparticles formed also increases. The largest change in absorbance value occurs between pH 7.0 and pH 8.0, indicating that at pH 8.0, the reaction rate between the reducing substances in *Spatholobus suberectus* and silver nitrate is significantly accelerated, and alkaline conditions are more conducive to the formation of silver nanoparticles. The absorbance value changes are relatively small between pH 8.0 and pH 9.0, and the curves for pH 9.0 and pH 10.0 almost overlap, indicating that at these two pH values, the reaction rates of the diluted *Spatholobus suberectus* solution and the silver nitrate solution are similar, the absorbance values ​​are not significantly different, and the number of silver nanoparticles formed is also basically the same. From pH 10.0 to pH 11.0, the reaction rate between the *Spatholobus suberectus* extract and silver nitrate changes little, and the number of silver nanoparticles formed increases slightly. Considering all factors, pH 8.0 represents the most suitable pH, as it results in a faster reaction rate and a higher number of silver nanoparticles formed.

[0043] 3. Effect of temperature on the formation of silver nanoparticles Take 5 portions of 3 mL of the *Spatholobus suberectus* diluted solution obtained in step (1) of Example 1, adjust the pH of the diluted solution to 8.0 with 10 mol / L NaOH solution, and add 3 mL of 0.01 mol / L AgNO3 solution to each portion. Sonicate the solutions at 25℃, 40℃, 55℃, 70℃, and 85℃ for ten minutes each. Use a UV spectrophotometer to detect whether nano-silver is formed during the synthesis process, and measure the intensity of the absorption peak at 410 nm. Select the optimal temperature by analyzing the magnitude of the absorbance value and the changes in the two absorbance curves.

[0044] like Figure 3As shown, when the pH of the *Spatholobus suberectus* extract was adjusted to the optimal value of 8.0, the absorbance gradually increased with increasing temperature, and the number of silver nanoparticles generated also increased slightly, but the yield was not significantly different from that generated at 25℃. At these five different temperatures, the absorbance of the five curves did not change much, indicating that the reaction rate and the number of silver nanoparticles generated were not significantly different at different temperatures, suggesting that temperature has little effect on the reaction rate of the solution and the formation of silver nanoparticles. Therefore, the reaction at room temperature (25℃) is sufficient.

[0045] 4. Effect of feed-to-liquid ratio on the formation of silver nanoparticles Take 5 portions of 3 mL of the diluted chicken blood vine solution prepared in step (1) of Example 1, adjust the pH of the diluted chicken blood vine solution to 8.0 with 10 mol / L NaOH solution, and add 3 mL of 0.01 mol / L AgNO3 solution respectively. Adjust the ratio of chicken blood vine solution to silver nitrate to 3:1, 2:1, 1:1, 1:2, and 1:3 respectively. Sonicate at room temperature for 10 minutes, use a UV spectrophotometer to detect whether there is the formation of nano-silver during the synthesis process, and measure the intensity of the absorption peak at 410 nm. Select the most suitable material-liquid ratio by the magnitude of the absorbance value and the change of the two absorbance value curves.

[0046] like Figure 4 As shown, when the material-to-liquid ratio is 3:1, the absorbance gradually increases as the ratio increases from 3:1 to 2:1 and then to 1:1, indicating that there are more reducing substances in the *Spatholobus suberectus* solution and fewer silver nanoparticles are generated. Increasing the amount of silver nitrate also increases the number of silver nanoparticles generated. The peak width of the UV absorption peak is related to the particle size distribution; larger particles result in a larger maximum absorption wavelength. When the material-to-liquid ratio reaches 1:2, the absorbance decreases, possibly indicating aggregation. When the material-to-liquid ratio is 1:3, the UV absorption peak width increases, possibly due to uneven distribution of silver nanoparticles and a significant increase in particle size. In conclusion, a material-to-liquid ratio of 1:1 is the optimal ratio.

[0047] 5. Characterization of AgNPS The crystal characteristics, morphology, particle size and surface properties of nanosilver were studied using X-ray diffraction (XRD) and spherical aberration electron microscopy (TEM).

[0048] 5.1 Zeta potential of silver nanoparticles prepared by different methods The average particle size and PDI of silver nanoparticles were measured using a Malvern laser particle size analyzer. A trace amount of silver nanoparticles prepared in Example 1 was dissolved in 100 mL of ultrapure water and ultrasonically dispersed for 10 min using a 150 W ultrasonic cleaner. The sample was then placed in a cuvette, and the following parameters were set: test time: 6 min, dispersion medium: water, temperature: 25 °C, refractive index: 0.230, absorptivity: 0.05, test temperature: 25 °C, delay time: 50 μs. Each sample was measured seven times, and the average was taken. The particle size and PDI displayed on the screen were recorded. A Zeta potential analyzer was used. The page for measuring particle size was switched to the page for measuring Zeta potential, and measurements were performed according to the above parameters. Each sample was measured three times, and the average was taken.

[0049] like Figure 5 As shown, the zeta potential of the silver nanoparticles prepared from *Spatholobus suberectus* is -1.41 mV. This indicates that the prepared silver nanoparticles have good stability, and that negatively charged compounds are adsorbed on the surface of the silver nanoparticles. This is because active substances such as flavonoids from *Spatholobus suberectus* are adsorbed onto the surface of the silver nanoparticles in the form of negative charges.

[0050] 5.2 Aberration-corrected electron microscopy (AC-TEM) analysis A certain amount of the silver nanoparticles of *Chicken Blood Vine* prepared in Example 1 were dissolved in ultrapure water, ultrasonically dispersed in an ultrasonic cleaner for 30 minutes, and a portion of the sample was dropped onto a copper grid for testing. The results are as follows: Figure 6 As shown, the silver nanoparticles prepared from chicken blood vine have a complete structure, are spherical or near-spherical in shape, have a smooth surface, and a particle size of about 5-10 nm. They are uniformly dispersed, which is smaller than the nanoparticle size reported in most related literature.

[0051] 5.3 X-ray diffraction analysis A certain amount of the *Spatholobus suberectus* silver nanoparticles prepared in Example 1 were spread evenly at the center of the sample stage and flattened with a glass plate (without cracks appearing), then placed in the diffraction instrument. The parameters were set in the software as follows: Voltage: 40 kV, Current: 40 mA, Time: 0.1, 2Theta, Start: 5°, Stop: 70°, Increment: 0.0195°. The test was then started.

[0052] like Figure 7As shown, the silver nanoparticles synthesized by the chicken blood vine exhibit obvious absorption peaks. The four diffraction peaks of the silver nanoparticles at 2θ = 38.02°, 44.15°, 64.50°, and 77.48° can be attributed to the (111), (200), (220), and (311) planes, respectively, indicating that the silver nanoparticles synthesized by the biological method have a face-centered cubic structure. From the XRD pattern, it can be inferred that the particles prepared by the biological method have small particle size, uniform dispersion, and are polycrystalline. Polycrystalline particles are small, have a large specific surface area, and have more active sites, which will result in better activity in applications such as catalysis and sterilization.

[0053] 6. Antioxidant performance test Weigh 2.5 mg of DPPH and dissolve it in ultrapure water, then dilute to a 100 mL amber volumetric flask. Prepare AgNPs solutions of 50 μg / mL, 100 μg / mL, 150 μg / mL, and 200 μg / mL, and sonicate for ten minutes. Take 4 mL of each AgNPs solution of different concentrations, add 4 mL of DPPH, and react in the dark for half an hour. Measure the absorbance at 517 nm. Measure each sample three times and take the average value. Take another 4 mL of each AgNPs solution of different concentrations, add 4 mL of ethanol, and react in the dark for half an hour. Measure the absorbance at 517 nm. Measure each sample three times and take the average value. Take 4 mL of DPPH and add 4 mL of ultrapure water, and 4 mL of ethanol and add 4 mL of ultrapure water. Sonicate again, react in the dark for half an hour, and measure the absorbance at 517 nm. Use vitamin C as a control and prepare vitamin C solutions with the same concentration gradient. Take 4 mL of vitamin C solution of different concentrations, add 4 mL of DPPH, and react in the dark for half an hour. Measure the absorbance at 517 nm. Measure each sample three times and take the average value. Then take 4 mL of vitamin C solution of different concentrations, add 4 mL of ethanol, and react in the dark for half an hour. Measure the absorbance at 517 nm. Measure each sample three times and take the average value. Finally, calculate the DPPH scavenging rate according to the following formula based on the measured absorbance values. Simultaneously, for the control experiment, prepare vitamin C (VC) solution of the same mass concentration as the AgNPs sample solution, prepare the same concentration, and measure its absorbance at 517 nm as a positive control.

[0054] Free radical scavenging rate = [1 - (A1 - A2) / A0] × 100% In the formula, A0: absorbance of 4 mL DPPH + 4 mL ultrapure water at 517 nm; A1: absorbance of 4 mL DPPH + 4 mL sample at 517 nm; A2: absorbance of 4 mL sample + 4 mL ethanol at 517 nm.

[0055] The scavenging rates of DPPH free radicals by *Spatholobus suberectus* nano-silver and vitamin C are shown in Table 1. Table 1. Free radical scavenging rates of nano silver and vitamin C

[0056] like Figure 8 As shown, the silver nanoparticles from *Spatholobus suberectus* exhibit a scavenging effect on DPPH free radicals. Furthermore, the scavenging rate of DPPH free radicals increases with increasing concentration of silver nanoparticles from *Spatholobus suberectus*. When the concentration of silver nanoparticles from *Spatholobus suberectus* reaches 100 μg / mL, the free radical scavenging rate reaches 81.7%, indicating that the silver nanoparticles prepared from *Spatholobus suberectus* possess good free radical scavenging ability.

[0057] 7. Antibacterial performance test Escherichia coli and Staphylococcus aureus were separately inoculated into LB solid medium and streaked, then incubated at 37°C for 18 h. Two sterilized Erlenmeyer flasks were then filled with LB liquid medium. A loopful of a single colony was picked up and transferred to a test tube, which was then placed in a water bath and incubated at 37°C and 150 rpm for 18 h to obtain a bacterial suspension of 1×10⁸ CFU / mL. On a clean bench, 1 mL of the Escherichia coli suspension was placed in a tube, and 100 μL of the suspension was added to 900 μL of LB liquid medium to obtain an Escherichia coli suspension of 1×10⁷ CFU / mL. This suspension was then diluted sequentially until a final concentration of 1×10⁵ CFU / mL was obtained. Similarly, Staphylococcus aureus was diluted sequentially to a 1×10⁵ CFU / mL suspension using the same method.

[0058] The minimum inhibitory concentration (MIC) of silver nanoparticles was determined using a serial dilution method. Appropriate amounts of the *Spatholobus suberectus* silver nanoparticles prepared in Example 1 and those prepared by conventional chemical methods were weighed and dissolved in sterile water to prepare suspensions of 5000 μg / mL. These suspensions were then diluted with sterile water to create ten concentration gradients: 2500 μg / mL, 1250 μg / mL, 625 μg / mL, 312.5 μg / mL, and 156.25 μg / mL. 50 μL of each concentration was added dropwise to a 96-well plate (from highest to lowest concentration, from top to bottom), followed by 50 μL of bacterial suspension. Each sample was repeated three times. Additionally, 100 μL of LB broth and 100 μL of bacterial suspension were used as negative and positive controls, respectively. The 96-well plates were incubated at 37°C for 24 h. Observe the turbidity of the well plate. If it becomes turbid, it indicates that it cannot inhibit bacteria. The lowest concentration corresponding to the well that does not produce turbidity is the minimum inhibitory concentration.

[0059] After obtaining the minimum inhibitory concentration (MIC), 10 μL of culture was taken from the clear wells of the two 96-well plates in the above experiment, and then 90 μL of LB liquid medium was added. Each sample was repeated three times. Additionally, 100 μL of LB liquid medium and 100 μL of bacterial suspension were taken as negative and positive controls, respectively. The 96-well plates were incubated at 37°C for 24 h. The turbidity of the plates was observed; if the plates became turbid, it indicated that no bacteria were inhibited. The lowest concentration corresponding to the wells that did not produce turbidity was the minimum bactericidal concentration.

[0060] The results are shown in Table 2: Table 2 Antibacterial activity of aqueous extract and nano silver

[0061] As shown in Table 2, the ten concentrations of *Spatholobus suberectus* aqueous extract used in the experiment had no inhibitory effect on either *Escherichia coli* or *Staphylococcus aureus*. The *Spatholobus suberectus* silver nanoparticles prepared in Example 1 exhibited the same minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against both *Escherichia coli* and *Staphylococcus aureus*. The MIC was 312.5 μg / mL, and the MBC was 625 μg / mL. In contrast, the chemical method showed a MIC and MBC of 625 μg / mL against *Escherichia coli* and 1250 μg / mL against *Staphylococcus aureus*. The minimum bactericidal concentration (MIC) of silver nanoparticles prepared from *Spatholobus suberectus* against both bacteria was lower than that of silver nanoparticles prepared by chemical methods. The MICs of silver nanoparticles prepared by both methods against *Escherichia coli* were the same. However, for *Staphylococcus aureus*, the MIC of silver nanoparticles prepared from *Spatholobus suberectus* was lower than that of silver nanoparticles prepared by chemical methods. These results demonstrate that the antibacterial properties of silver nanoparticles prepared from *Spatholobus suberectus* are superior to those prepared by chemical methods. This is consistent with the characterization results: silver nanoparticles prepared from *Spatholobus suberectus* are smaller, have a larger specific surface area, are more uniformly dispersed, and have more active sites than chemically prepared nanoparticles, thus exhibiting better bactericidal activity.

Claims

1. A method for preparing silver nanoparticles from a water extract of *Spatholobus suberectus*, characterized in that, Includes the following steps: (1) After the dried chicken blood vine is crushed and sieved, it is added to ultrapure water and heated for extraction. After cooling and centrifugation, the supernatant is taken to obtain the original chicken blood vine solution. Then the original chicken blood vine solution is diluted to obtain the diluted chicken blood vine solution. (2) Adjust the pH of the chicken blood vine dilution obtained in step (1) to 8.0 to 11.0 using an alkaline solution. Mix the adjusted chicken blood vine dilution with silver nitrate solution evenly, control the material-liquid ratio to be 3:1 to 1:1, and sonicate at 25℃ to 85℃ for 1 min to 120 min to obtain crude silver nanoparticle solution. (3) The crude liquid of silver nanoparticles obtained in step (2) is allowed to stand to precipitate, the precipitate is separated, washed and dried to obtain silver nanoparticles of chicken blood vine.

2. The method for preparing silver nanoparticles from chicken blood vine aqueous extract according to claim 1, characterized in that: In step (1), the chicken blood vine is crushed and passed through a 60-mesh sieve; 200mL of ultrapure water is added to every 10g of crushed chicken blood vine for extraction, and the extraction conditions are heating to a slight boiling state and extraction time is 30min.

3. The method for preparing silver nanoparticles from chicken blood vine aqueous extract according to claim 1, characterized in that: In step (1), the centrifugation speed is 5000 r / min and the centrifugation time is 15 min; the original chicken blood vine solution is diluted 7 times with ultrapure water to obtain a diluted chicken blood vine solution with a pH value of 7.

0.

4. The method for preparing silver nanoparticles from chicken blood vine aqueous extract according to claim 1, characterized in that... In step (2), the alkaline regulator is a 10 mol / L sodium hydroxide aqueous solution.

5. The method for preparing silver nanoparticles from the aqueous extract of *Spatholobus suberectus* according to claim 4, characterized in that: In step (2), the pH value of the diluted chicken blood vine solution obtained in step (1) is adjusted to 8.0 using sodium hydroxide aqueous solution. The diluted chicken blood vine solution with adjusted pH value is mixed evenly with silver nitrate solution, and the material-liquid ratio is controlled to be 1:

1. The mixture is ultrasonically reacted at 25℃ for 10 min to obtain crude silver nanoparticle solution.

6. The method for preparing silver nanoparticles from the aqueous extract of *Spatholobus suberectus* according to claim 5, characterized in that: In step (2), the concentration of the silver nitrate solution is 0.01 mol / L.

7. The method for preparing silver nanoparticles from the aqueous extract of *Spatholobus suberectus* according to claim 1, characterized in that: In step (3), ultrapure water is used for precipitate washing, and the washing is performed 3 times; the drying temperature is 60℃, and the product is dried to constant weight.

8. A type of silver nanoparticles prepared according to any one of claims 1-7, characterized in that: The silver nanoparticles have a particle size of 5-10 nm, a spherical morphology, and a face-centered cubic crystal structure.

Images