Gold-silver alloy nanoparticles, preparation method and application thereof
The gold-silver alloy nanoparticles prepared by the aqueous phase method have solved the shortcomings of alloy nanoparticles in the treatment of Alzheimer's disease, achieved effective inhibition of Aβ40 misfolding, and provided a simple and environmentally friendly treatment solution.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- 安徽理工大学第一附属医院(淮南市第一人民医院)
- Filing Date
- 2023-12-04
- Publication Date
- 2026-06-26
AI Technical Summary
The application of alloy nanoparticles in the treatment of Alzheimer's disease is limited in the current technology, and research on existing nanoparticles in inhibiting Aβ40 misfolding is also limited.
Gold-silver alloy nanoparticles with diameters of 1–3 nm were synthesized by an aqueous phase method. This method involves reacting water, silver sulfide quantum dots, and polyvinylpyrrolidone powder at 0 °C, followed by a mixed reaction of chloroauric acid, sodium citrate, and NaOH solution. Hydroxylamine hydrochloride and NaOH solution were then added, and the mixture was finally purified by ultrafiltration centrifuge tubes.
The synthesis process is simple and environmentally friendly, and it can effectively inhibit the misfolding of Aβ protein, thus having potential applications in the treatment of Alzheimer's disease.
Smart Images
Figure CN117900503B_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of functional nanomaterials technology, specifically to a gold-silver alloy nanoparticle, its preparation method, and its application. Background Technology
[0002] Alzheimer's disease (AD) is a neurodegenerative disease characterized by extracellular senile plaque deposition, intracellular neurofibrillary tangles, and abnormal loss of neurons and synapses. Currently, the diagnosis of Alzheimer's disease primarily relies on detecting the presence of misfolded amyloid proteins, including misformed protein aggregates, neurofibrillary tangles composed of hyperphosphorylated tau protein, senile plaques composed of Aβ peptides, widespread oxidative damage, and post-mortem neuronal loss. Among these, senile plaque deposition is formed by the misfolding, abnormal aggregation, and fibrosis of Aβ40; therefore, inhibiting Aβ40 misfolding is currently key to research in Alzheimer's disease treatment.
[0003] Existing research indicates that nanoparticles of varying sizes, designed based on their physicochemical properties (e.g., size, shape, surface modification, charge, composition, and concentration), can be used to inhibit the misfolding of Aβ40 protein. For example, Professor Gao Guanbin and colleagues at Wuhan University of Technology reported on the size effect of L-glutathione-stabilized gold nanoparticles and gold nanoparticle clusters in inhibiting Aβ protein misfolding; furthermore, researchers evaluated the inhibitory effect of nanoparticles with different shapes (nanospheres and cubes) on Aβ protein fibrosis. These studies demonstrate the significant potential therapeutic implications of developing functional nanomaterials for Alzheimer's disease. However, current research on the therapeutic effects of alloy nanoparticles on Alzheimer's disease is limited. Therefore, this invention proposes a gold-silver alloy nanoparticle, its preparation method, and its application in Alzheimer's disease. Summary of the Invention
[0004] Based on the above description, the present invention provides a method for preparing gold-silver alloy nanoparticles, the method for preparation and the application thereof, and the obtained gold-silver alloy nanoparticles are applied to the research of Alzheimer's disease.
[0005] The technical solution of the present invention to solve the above-mentioned technical problems is as follows:
[0006] In a first aspect, the present invention provides a method for preparing gold-silver alloy nanoparticles, comprising:
[0007] Step S1: React water, silver sulfide quantum dots and polyvinylpyrrolidone powder at 0°C until fully reacted. After centrifugation, obtain the silver sulfide supernatant.
[0008] Step S2: Mix chloroauric acid, sodium citrate solution and NaOH solution at room temperature until fully reacted to obtain a reaction solution;
[0009] Step S3: Mix the silver sulfide supernatant obtained in step S1 with the reaction solution obtained in step S2, and add polyvinylpyrrolidone. After the reaction, add hydroxylamine hydrochloride solution and NaOH solution in sequence to allow the reaction to proceed fully and obtain gold-silver alloy ions.
[0010] Step S4: Purify the gold-silver alloy ions obtained in step S3 to obtain gold-silver alloy nanoparticles.
[0011] Based on the above technical solution, the present invention can be further improved as follows.
[0012] Furthermore, step S1 specifically includes:
[0013] 8–12 mL of water, 180–230 μL of silver sulfide quantum dots, and 0.09–1.2 g of polyvinylpyrrolidone powder were reacted thoroughly in a water bath at 0 °C. After centrifugation, the supernatant of silver sulfide was obtained.
[0014] Further, step S2 specifically includes: adding 180-220 μL of chloroauric acid, 17-23 μL of sodium citrate solution and 30-38 μL of NaOH solution to the reaction flask, and reacting at room temperature for 25-40 min;
[0015] The concentration of sodium citrate solution is 36–40 mM, and the concentration of NaOH solution is 0.8–1.2 M.
[0016] Furthermore, step S3 specifically includes:
[0017] Add the supernatant of silver sulfide obtained from centrifugation in step S1 to the reaction flask in step S2, and add 0.03-0.06 g of polyvinylpyrrolidone. React at 38-42°C until fully saturated. Then add 18-22 μL of 0.09-1.2 M hydroxylamine hydrochloride solution and 1 drop of 1 M NaOH solution, and allow the reaction to proceed fully.
[0018] Furthermore, step S4 specifically includes:
[0019] The gold-silver alloy ions obtained in step S3 were purified using an ultrafiltration centrifuge tube to obtain gold-silver alloy nanoparticles; wherein the molecular weight cutoff of the ultrafiltration centrifuge tube was 5K Daltons.
[0020] In a second aspect, the present invention also provides gold-silver alloy nanoparticles, which are prepared by the preparation method described in the first aspect.
[0021] Based on the above technical solution, the present invention can be further improved as follows.
[0022] Furthermore, the gold-silver alloy nanoparticles include a gold core, a silver core, and ligands;
[0023] The gold core and the silver core are gold-silver alloy cores, and the ligand is modified on the surface of the gold-silver alloy core.
[0024] Furthermore, the ligand is a polyvinylpyrrolidone with a molecular weight of 5000.
[0025] Furthermore, the diameter of the gold-silver alloy nanoparticles is 1–3 nm.
[0026] Thirdly, the present invention also provides the use of gold-silver alloy nanoparticles in the detection of Alzheimer's disease, specifically for inhibiting Aβ fibrosis in the pathogenesis of Alzheimer's disease.
[0027] Compared with the prior art, the technical solution of this application has the following beneficial technical effects:
[0028] Compared with existing technologies, the gold-silver alloy nanoparticles and their preparation method provided by this invention use an aqueous phase method to synthesize gold-silver alloy nanoparticles. The synthesis process is simple, green and environmentally friendly, and enriches the means of synthesizing gold-silver alloy nanoparticles. It has potential applications in the study of Alzheimer's disease and can effectively inhibit the size effect of Aβ protein misfolding. Attached Figure Description
[0029] Figure 1 Here is a high-resolution transmission electron microscope image of the gold-silver alloy nanoparticles prepared in Example 1;
[0030] Figure 2 X-ray energy dispersive spectroscopy (EDS) of the gold-silver alloy nanoparticles prepared in Example 1, obtained by transmission electron microscopy.
[0031] Figure 3 A double aberration transmission electron microscope image of the gold-silver alloy nanoparticles prepared in Example 1;
[0032] Figure 4 EDS mapping image of the gold-silver alloy nanoparticles prepared in Example 1;
[0033] Figure 5 X-ray diffraction spectrum of the gold-silver alloy nanoparticles prepared in Example 1;
[0034] Figure 6 The fluorescence spectrum of the gold-silver alloy nanoparticles prepared in Example 1 is shown below.
[0035] Figure 7 The ultraviolet-visible emission spectrum of the gold-silver alloy nanoparticles prepared in Example 1;
[0036] Figure 8 The hydrated particle size results are for the gold-silver alloy nanoparticles prepared in Example 1;
[0037] Figure 9 A schematic diagram of the fibrillation dynamics of 20 μmol / L Aβ40 co-incubated with gold-silver alloy nanoparticles of different concentrations prepared in Example 1 at 37 °C for 140 h.
[0038] Figure 10 Circular dichroism spectrum of 20 μmol / L Aβ40 co-incubated with 20 mg / L gold-silver alloy nanoparticles prepared in Example 1 at 37 °C;
[0039] Figure 11 Transmission electron microscope image of 20 μmol / L Aβ40 incubated at 37°C for 48 h;
[0040] Figure 12 Transmission electron microscope image of 20 μmol / L Aβ40 and 20 mg / L gold-silver alloy nanoparticles prepared in Example 1 after co-incubation at 37 °C for 48 h. Detailed Implementation
[0041] To facilitate understanding of this application, a more complete description will be provided below with reference to the accompanying drawings, which illustrate embodiments of the present application. However, the present application can be implemented in many different forms and is not limited to the embodiments described herein. Rather, these embodiments are provided so that the disclosure of this application will be thorough and complete.
[0042] The terms “comprising,” “including,” and any variations thereof in the following embodiments and comparative examples are intended to cover non-exclusive inclusion, such as a process, method, or product that includes a series of steps or units but is not limited to the steps or units listed, but may optionally include steps or units not listed, or may optionally include other steps or units inherent to such processes, methods, or products.
[0043] Furthermore, the reaction apparatus and chemical reagents involved in the following examples and comparative examples are all commercially available, as are the detection instruments and detection reagents involved.
[0044] The embodiments of the present invention will be described in further detail below with reference to the accompanying drawings and examples. The following examples are used to illustrate the present invention, but should not be used to limit the scope of the present invention.
[0045] Example 1
[0046] This embodiment provides a method for preparing gold-silver alloy nanoparticles, including the following steps:
[0047] Step S1: React 10 mL of water, 200 μL of silver sulfide (AgS) quantum dots and 0.1024 g of polyvinylpyrrolidone (PVP) powder in a 0 °C water bath for 30 minutes, centrifuge at 6000 rpm for 30 minutes, and collect the supernatant.
[0048] Step S2: Add 200 μL of chloroauric acid (HAuCl4), 20 μL of 38.8 mM sodium citrate solution, and 35 μL of 1 M NaOH solution to the reaction flask and react at room temperature for 30 min.
[0049] Step S3: Add the AgS solution centrifuged in step S1 to the reaction flask in step S2, add 0.05g PVP, react at 40℃, then add 20μL of 0.1M hydroxylamine hydrochloride and one drop of 1M NaOH, and react overnight.
[0050] Step S4: Purify the gold-silver alloy nanoparticles AuAg NPs using an ultrafiltration centrifuge tube; wherein the molecular weight cutoff of the ultrafiltration centrifuge tube is 5K Dalton, the ultrafiltration tube is used for filtration, and the product is freeze-dried for later use.
[0051] The product was identified as gold-silver nanoalloy particles. The product was analyzed using TEM, mapping, XRD, EDS, DLS, FT-IR, UV-Vis, and XPS. Specific results are as follows: Figures 1 to 8 As shown.
[0052] Figure 1 The high-resolution transmission electron microscope (TEM) image of the AuAg NPs prepared for this embodiment shows that the size of the AuAg NPs is between 1 and 3 nm.
[0053] Figure 2 The X-ray energy dispersive spectroscopy (EDS) image of the AuAg NPs prepared for this embodiment shows that the AuAg NPs contain Au and Ag elements.
[0054] Figure 3 The image shows a double-spherical aberration transmission electron microscope image of the AuAg NPs prepared in this embodiment. As shown in the figure, the contrast between Au and Ag is different under the double-spherical aberration electron microscope, with one end being the Au nucleus and the other end being the Ag nucleus, which fully proves the successful preparation of AuAg NPs.
[0055] Figure 4 The EDS mapping diagram of the AuAg NPs prepared for this embodiment is shown in the figure, which shows the distribution of Au and Ag elements.
[0056] Figure 5 X-ray diffraction (XRD) pattern of AuAg NPs prepared for this embodiment.
[0057] Figure 6 The fluorescence spectrum of the AuAg NPs prepared in this example.
[0058] Figure 7 The UV-Vis emission spectrum of the AuAg NPs prepared for this embodiment.
[0059] Figure 8 The hydrated particle size results of AuAg NPs prepared for this example.
[0060] Figures 1 to 8 As shown, this effectively demonstrates the acquisition of gold-silver alloy nanoparticles obtained in this embodiment.
[0061] Example 2
[0062] This embodiment provides a method for preparing gold-silver alloy nanoparticles, including the following steps:
[0063] Step S1: React 8 mL of water, 180 μL of silver sulfide (AgS) quantum dots and 0.09 g of polyvinylpyrrolidone (PVP) powder in a 0 °C water bath for 30 minutes, centrifuge at 6000 rpm for 30 minutes, and collect the supernatant.
[0064] Step S2: Add 180 μL of chloroauric acid (HAuCl4), 17 μL of 30 mM sodium citrate solution, and 30 μL of 0.8 M NaOH solution to the reaction flask and react at room temperature for 30 min.
[0065] Step S3: Add the AgS solution centrifuged in step S1 to the reaction flask in step S2, add 0.03g PVP, react at 40℃, then add 18μL of 0.1M hydroxylamine hydrochloride and one drop of 1M NaOH, and react overnight.
[0066] Step S4: Purify the gold-silver alloy nanoparticles AuAg NPs using an ultrafiltration centrifuge tube; wherein the molecular weight cutoff of the ultrafiltration centrifuge tube is 5K Dalton, the ultrafiltration tube is used for filtration, and the product is freeze-dried for later use.
[0067] Example 3
[0068] This embodiment provides a method for preparing gold-silver alloy nanoparticles, including the following steps:
[0069] Step S1: React 12 mL of water, 230 μL of silver sulfide (AgS) quantum dots and 1.2 g of polyvinylpyrrolidone (PVP) powder in a 0 °C water bath for 30 minutes, centrifuge at 6000 rpm for 30 minutes, and collect the supernatant.
[0070] Step S2: Add 220 μL of chloroauric acid (HAuCl4), 23 μL of 30 mM sodium citrate solution, and 38 μL of 0.8 M NaOH solution to the reaction flask and react at room temperature for 30 min.
[0071] Step S3: Add the centrifuged AgS solution from step S1 to the reaction flask from step S2, and add 0.06 g PVP. React at 42 °C, then add 22 μL of 0.1 M hydroxylamine hydrochloride and one drop of 1 M NaOH, and react overnight.
[0072] Step S4: Purify the gold-silver alloy nanoparticles AuAg NPs using an ultrafiltration centrifuge tube; wherein the molecular weight cutoff of the ultrafiltration centrifuge tube is 5K Dalton, the ultrafiltration tube is used for filtration, and the product is freeze-dried for later use.
[0073] Example 4
[0074] This example demonstrates the use of the gold-silver alloy nanoparticles obtained in Example 1 in the detection of Alzheimer's disease, specifically in the Aβ40 protein assay.
[0075] First, take 5 mg of Aβ40 out of the -80℃ freezer and transfer it to a sterilized 2 mL EP tube. Add 1 mL of hexafluoroisopropanol solution and sonicate in an ice bath for 2 hours. After the treatment, take out the experimental volume and dry it with nitrogen.
[0076] Then, experiments were conducted using the gold-silver alloy nanoparticles obtained in Example 1 and the treated Aβ40 protein:
[0077] Step 1: Prepare solutions of gold-silver alloy nanoparticles with concentrations of 0 mg / L, 2.5 mg / L, 25 mg / L, 62.5 mg / L, and 125 mg / L, using phosphate-buffered saline (PBS) as the solvent.
[0078] Step 2: Weigh 15.9 mg of ThT (benzothiazole dye) powder and dissolve it in 12.5 mL of PBS. Then take 100 μL of the solution and dilute it to 4 mL to obtain the final ThT solution for later use.
[0079] Step 3: Dissolve the nitrogen-dried Aβ40 protein in PBS to a concentration of 50 μmol / L. Take a 96-well plate and add 50 μL of ThT solution, 100 μL of AuAg NPs solution at different concentrations, and 100 μL of protein solution to each well in the following order, until the final protein concentration in the well is 20 μmol / L. The AuAg NPs solution concentrations are 0 mg / L, 1 mg / L, 10 mg / L, 25 mg / L, and 50 mg / L, with 5 replicates for each concentration. The entire experiment, including sample addition and solution preparation, must be conducted in the dark.
[0080] Step 4: Add the 96-well plate to the microplate reader, set the incubation temperature to 37℃, select an excitation wavelength of 445nm and an emission wavelength of 485nm, and shake the plate and perform a test every 10 minutes.
[0081] Relevant test results, such as Figures 9 to 12 As shown.
[0082] Figure 9 The diagram shows the fibrillation kinetics of 20 μmol / L Aβ40 co-incubated with 1 mg / L, 5 mg / L, 10 mg / L, 25 mg / L and 50 mg / L AuAg NPs prepared in Example 1 at 37 °C for 140 h. It can be seen that, compared with the control group without gold and silver alloy nanoparticles, the addition of gold and silver alloy nanoparticles can inhibit the fiberization process of Aβ40.
[0083] Figure 10 The circular dichroism spectrum of 20 μmol / L Aβ40 co-incubated with 20 mg / L AuAg NPs prepared in Example 1 at 37 °C shows that, compared with the control group without gold-silver alloy nanoparticles, the addition of gold-silver alloy nanoparticles can maintain the Aβ40 protein structure in the α-helix structure and inhibit the transformation of Aβ40 to the β-helix structure, which further verifies the results of Aβ40 fibrillation dynamics: effectively inhibiting Aβ40 fibrillation.
[0084] Figure 11 Transmission electron microscope image of 20 μmol / L Aβ40 incubated at 37℃ for 48 h; Figure 12 Transmission electron microscope image of 20 μmol / L Aβ40 and 20 mg / L AuAg NPs prepared in Example 1, co-incubated at 37 °C for 48 h. Figure 11 and Figure 12 The comparison shows that AuAg NPs have a good inhibitory effect on the folding of Aβ40 in vitro.
[0085] Therefore, the gold-silver alloy nanoparticles prepared in the embodiments of the present invention can be used to inhibit Aβ40 fibrosis in the pathogenesis of Alzheimer's disease.
[0086] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.
Claims
1. A gold-silver alloy nanoparticle, characterized in that, The gold-silver alloy nanoparticles include a gold core, a silver core, and ligands; The gold core and the silver core are gold-silver alloy cores, and the ligand is modified on the surface of the gold-silver alloy core. The ligand is polyvinylpyrrolidone with a molecular weight of 5000. The method for preparing the gold-silver alloy nanoparticles includes: Step S1: React water, silver sulfide quantum dots and polyvinylpyrrolidone powder at 0°C until fully reacted. After centrifugation, obtain the silver sulfide supernatant. Step S2: Mix chloroauric acid, sodium citrate solution and NaOH solution at room temperature until fully reacted to obtain a reaction solution; Step S3: Mix the silver sulfide supernatant obtained in step S1 with the reaction solution obtained in step S2, and add polyvinylpyrrolidone. React at 38~42℃ until fully reacted. After the reaction, add hydroxylamine hydrochloride solution and NaOH solution in sequence and react fully to obtain gold-silver alloy particles. Step S4: Purify the gold-silver alloy particles obtained in step S3 to obtain gold-silver alloy nanoparticles.
2. The gold-silver alloy nanoparticles according to claim 1, characterized in that, Step S1 specifically includes: 8-12 mL of water, 180-230 μL of silver sulfide quantum dots, and 0.09-1.2 g of polyvinylpyrrolidone powder were reacted in a 0°C water bath until fully reacted. After centrifugation, the supernatant of silver sulfide was obtained.
3. The gold-silver alloy nanoparticles according to claim 2, characterized in that, Step S2 specifically includes: adding 180~220μL of chloroauric acid, 17~23μL of sodium citrate solution and 30~38μL of NaOH solution to the reaction flask, and reacting at room temperature for 25~40min; The concentration of sodium citrate solution is 36~40mM, and the concentration of NaOH solution is 0.8~1.2M.
4. The gold-silver alloy nanoparticles according to claim 3, characterized in that, Step S3 specifically includes: Add the supernatant of silver sulfide obtained from centrifugation in step S1 to the reaction flask in step S2, and add 0.03~0.06g of polyvinylpyrrolidone. React at 38~42℃ until fully developed. Then add 18~22μL of 0.09~1.2M hydroxylamine hydrochloride solution and 1 drop of 1M NaOH solution and allow to react fully.
5. The gold-silver alloy nanoparticles according to claim 4, characterized in that, Step S4 specifically includes: The gold-silver alloy ions obtained in step S3 were purified using an ultrafiltration centrifuge tube to obtain gold-silver alloy nanoparticles; wherein the molecular weight cutoff of the ultrafiltration centrifuge tube was 5K Daltons.
6. The gold-silver alloy nanoparticles according to claim 1, characterized in that, The diameter of the gold-silver alloy nanoparticles is 1~3nm.
7. The use of the gold-silver alloy nanoparticles according to any one of claims 1 to 6 in the detection of Alzheimer's disease, specifically for inhibiting Aβ40 fibrosis in the pathogenesis of Alzheimer's disease.