A method for synthesizing large-size colloidal gold microspheres

Abstract

The application relates to the field of material chemical synthesis, in particular to a synthesis method of large-size colloidal gold microspheres, which comprises the following steps: dissolving oleylamine and chloroauric acid in toluene, heating to boiling and continuously reacting for 3 hours, adding ethanol for precipitation after cooling to room temperature, centrifuging to obtain oil-phase colloidal gold nanoparticles OA-AuNPs, and redissolving with toluene to obtain OA-AuNPs toluene solution; mixing and dissolving the OA-AuNPs toluene solution obtained in the step S1 with distearoyl phosphatidyl ethanolamine-polyethylene glycol-n-heptanoic acid, adding sodium dodecyl sulfate SDS aqueous solution, and forming a microemulsion through ultrasonic emulsification. The synthesis method is efficient, easy to prepare and stable in performance; the prepared large-size colloidal gold microspheres are large in loading capacity, excellent in colorimetric characteristics, strong in stability, have rich functional groups on the surface which can be directly coupled with proteins, and overcome the related defects of traditional colloidal gold.

Description

Technical Field

[0001] This invention relates to the field of materials chemical synthesis, specifically to a method for synthesizing large-size colloidal gold microspheres. Background Technology

[0002] Colloidal gold nanoparticles have wide applications in food safety testing and medical examination due to their unique optical properties and biocompatibility. Currently, the preparation of colloidal gold nanoparticles mainly falls into two categories: Water-soluble traditional colloidal gold nanoparticles (SC-AuNPs): prepared by sodium citrate reduction, with a particle size typically of 20-40 nm. Although they exhibit good biocompatibility, as the particle size increases, the particle color tends to turn purple and the stability decreases significantly, affecting the reliability and accuracy of detection results and limiting their application in high-sensitivity detection scenarios. Lipid-soluble colloidal gold nanoparticles (OA-AuNPs): Prepared by reducing chloroauric acid with oleylamine, they have uniform particle size and similar optical properties to SC-AuNPs. However, OA-AuNPs need to be stored in an oil phase environment, while practical detection applications are mostly in aqueous systems. The incompatibility between oil and aqueous phases seriously limits their widespread application.

[0003] Given the shortcomings of existing technologies, there is an urgent need to develop a method for preparing colloidal gold microspheres that combines large size, high stability, good aqueous phase compatibility, and abundant surface functional groups. Summary of the Invention

[0004] The purpose of this invention is to provide a method for synthesizing large-sized colloidal gold microspheres, in order to solve the problems of poor stability, abnormal color, and incompatibility with water phases of lipid-soluble colloidal gold after the particle size is increased.

[0005] To achieve the above objectives, the present invention provides the following technical solution: a method for synthesizing large-size colloidal gold microspheres, comprising: S1. Preparation of oil-phase colloidal gold nanoparticles (OA-AuNPs): Oleylamine and chloroauric acid were dissolved in toluene, heated to boiling and reacted for 3 hours. After cooling to room temperature, ethanol was added to precipitate the mixture. The mixture was centrifuged to obtain oil-phase colloidal gold nanoparticles OA-AuNPs, which were then redissolved in toluene to obtain an OA-AuNPs toluene solution. S2. Preparation of microemulsion: The OA-AuNPs toluene solution obtained in step S1 is mixed and dissolved with distearate phosphatidylethanolamine-polyethylene glycol-n-heptanoic acid (DSPE-PEG-COOH), and then added to sodium dodecyl sulfate SDS aqueous solution. The mixture is then ultrasonically emulsified to form a microemulsion. S3. Solidification into spheres: Remove toluene from the microemulsion in step S2, collect the precipitate by centrifugation of the remaining system, wash and re-dissolve to obtain large-sized colloidal gold microspheres (AuSPs).

[0006] Preferably, in step S2, the microemulsion is prepared by ultrasonic emulsification using a cell disruptor, with an emulsification time of 1-10 min, an emulsification power of 100-300 W, and an ultrasonic working mode of 5 s operation followed by a 10 s interval. The concentration of the SDS aqueous solution in step S2 is 8.7 mM.

[0007] Preferably, the method for removing toluene in step S3 is rotary evaporation or vacuum drying.

[0008] Preferably, the centrifugation conditions for the system after toluene removal in step S3 are: rotation speed 8000-12000 rpm, centrifugation time 10-30 min, and the purpose of centrifugation is to collect the precipitate.

[0009] Preferably, the precipitate washing step in step S3 is as follows: The precipitate was washed twice with ethanol as a detergent, then washed with ultrapure water until the supernatant was colorless, and finally reconstituted with ultrapure water or buffer solution.

[0010] Preferably, the buffer solution is one or a mixture of ultrapure water, phosphate buffer, or Tris-HCl buffer.

[0011] Preferably, the volume ratio of toluene in step S1 to sodium dodecyl sulfate aqueous solution in step S2 is 1:4-1:10; In step S1, the amount of distearate phosphatidylethanolamine-polyethylene glycol-n-heptanoic acid added is 4 mg.

[0012] Preferably, the large-sized colloidal gold microspheres obtained in step S3 have a diameter of 0.1-0.2 μm (i.e., 100-200 nm), have carboxyl active groups attached to their surface, and have a polydispersity index ≤0.299.

[0013] Compared with the prior art, the beneficial effects of the present invention are: This invention uses OA-AuNPs as the core, combined with DSPE-PEG-COOH and SDS as interface modifiers. Through steps such as dissolution and heating to prepare OA-AuNPs, ultrasonic formation of microemulsion, removal of toluene, and centrifugation and washing, colloidal gold microspheres with a diameter of 100-200 nm are obtained. This synthesis method is efficient and easy to operate. The product not only has a large loading capacity, excellent colorimetric performance and strong stability, but also has a surface rich in active groups such as carboxyl groups, which can be directly coupled with proteins. It effectively overcomes the application limitations of traditional colloidal gold and provides a material support with better performance for related detection fields. Attached Figure Description

[0014] Figure 1 This is a schematic diagram of the molecular structure of DSPE-PEG-COOH, a method for synthesizing large-size colloidal gold microspheres according to the present invention. Figure 2 This is a schematic diagram of the synthesis route of AuSPs, a method for synthesizing large-size colloidal gold microspheres according to the present invention. Figure 3 This is a schematic diagram of the ultraviolet-visible spectra of the OA-AuNPs and AuSPs of the present invention; Figure 4 This is a schematic diagram illustrating the hydration particle size characterization of AuSPs synthesized in Example 1 of the present invention; Figure 5 This is a schematic diagram illustrating the hydration particle size characterization of AuSPs synthesized in Example 2 of the present invention; Figure 6 These are physical schematic diagrams of Examples 1 and 2 of the present invention and a conventional 30nm colloidal gold. Detailed Implementation

[0015] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0016] Please see Figure 1-6 This invention provides a technical solution: a method for synthesizing large-size colloidal gold microspheres, comprising: S1. Preparation of oil-phase colloidal gold nanoparticles (OA-AuNPs): Oleylamine and chloroauric acid were dissolved in toluene, heated to boiling and reacted for 3 hours to generate dark red OA-AuNPs. After cooling to room temperature, ethanol was added to precipitate the nanoparticles. The nanoparticles were obtained by centrifugation and redissolved in toluene to obtain a toluene solution of OA-AuNPs. S2. Preparation of microemulsion: The OA-AuNPs toluene solution obtained in step S1 is mixed and dissolved with 4 mg of distearate phosphatidylethanolamine-polyethylene glycol-n-heptanoic acid (DSPE-PEG-COOH), and then added to an 8.7 mM sodium dodecyl sulfate (SDS) aqueous solution. The mixture is then ultrasonically emulsified using a cell disruptor to form a microemulsion. The microemulsion is prepared by ultrasonic emulsification using a cell disruptor. The emulsification time is 1-10 min, the emulsification power is 100-300 W, and the ultrasonic working mode is 5 s working and 10 s interval. The volume ratio of toluene to sodium dodecyl sulfate aqueous solution is 1:4 to 1:10; S3. Solidification into spheres: Toluene in the microemulsion from step S2 is removed by rotary evaporation or vacuum drying. The remaining system is then centrifuged at 8000-12000 rpm for 10-30 min to collect the precipitate. The precipitate is washed and then reconstituted to obtain large-sized colloidal gold microspheres (AuSPs). The diameter of the large-sized colloidal gold microspheres is 0.1-0.2 μm (i.e., 100-200 nm). The surface is connected with carboxyl active groups, which can be directly coupled with proteins. The polydispersity index of the colloidal gold microspheres is ≤0.299. The precipitate is washed twice with ethanol as the detergent, then washed with ultrapure water until the supernatant is colorless, and finally reconstituted with ultrapure water or buffer solution. The buffer solution is one or more of ultrapure water, phosphate buffer, or Tris-HCl buffer. Example 1

[0017] Synthesis of AuSPs with a particle size of 126.0 nm Preparation of lipid-soluble colloidal gold nanoparticles (OA-AuNPs): 18 mL of toluene and 7 mL of oleylamine were mixed and heated to boiling. Under rapid stirring, 2.895 mL of 2% chloroauric acid was added. After reacting for 3 h, 25 mL of ethanol was added to precipitate OA-AuNPs, followed by centrifugation at 2500 g for 10 min. Finally, the synthesized OA-AuNPs were completely dissolved in 5 mL of toluene. S2. Preparation of microemulsion: Take 100 μL of the above OA-AuNPs toluene solution, add 4 mg of DSPE-PEG-COOH and dissolve it completely. Disperse the mixture in 0.9 mL of 8.7 mM SDS aqueous solution and sonicate it for 5 min (5 s working time, 10 s interval) using an ultrasonic emulsifier with a working power of 300 W. S3. Solidification into spheres: Place the above microemulsion in a rotary evaporator and evaporate until no toluene is detected. Centrifuge the concentrate at 10,000 rpm for 15 min. Wash the precipitate twice with ethanol and then wash it with ultrapure water until the supernatant is colorless. Finally, redissolve the precipitate with 1 mL of ultrapure water. Example 2

[0018] Synthesis of AuSPs with a particle size of 182.5 nm Preparation of lipid-soluble colloidal gold nanoparticles (OA-AuNPs): 18 mL of toluene and 7 mL of oleylamine were mixed and heated to boiling. Under rapid stirring, 2.895 mL of 2% chloroauric acid was added. After reacting for 3 h, 25 mL of ethanol was added to precipitate OA-AuNPs, followed by centrifugation at 2500 g for 10 min. Finally, the synthesized OA-AuNPs were completely dissolved in 5 mL of toluene. S2. Preparation of microemulsion: Take 200 μL of the above OA-AuNPs toluene solution, add 4 mg of DSPE-PEG-COOH and dissolve it completely. Disperse the mixture in 0.8 mL of 8.7 mM SDS aqueous solution and sonicate it for 5 min (5 s working time, 10 s interval) using an ultrasonic emulsifier with a working power of 300 W. S3. Solidification into spheres: Place the above microemulsion in a rotary evaporator and evaporate until no toluene is detected. Centrifuge the concentrate at 10,000 rpm for 15 min. Wash the precipitate twice with ethanol and then wash it with ultrapure water until the supernatant is colorless. Finally, redissolve the precipitate with 1 mL of ultrapure water.

[0019] Comparative Example Synthesis of traditional colloidal gold Synthesis of conventional 30nm colloidal gold: 1 mL of 1% chloroauric acid was added to 99 mL of ultrapure water and heated to boiling. Under high-speed stirring, 1.45 mL of 1% sodium citrate was added. After reacting for 15 min, the mixture was cooled to synthesize conventional colloidal gold (SC-AuNPs).

[0020] The OA-AuNPs and AuSPs prepared in Example 1 were subjected to UV-Vis absorption spectroscopy tests, and the results are as follows: Figure 3 As shown, both have a UV absorption peak of 530 nm, indicating that the AuSPs prepared in this invention retain the excellent optical properties of OA-AuNPs.

[0021] The colloidal gold microspheres prepared in Example 1 were characterized for hydration particle size, and the results are as follows: Figure 4 As shown, the hydrated particle size of AuSPs is 126.0 nm, and the polydispersity index is 0.160.

[0022] The colloidal gold microspheres prepared in Example 2 were characterized for hydration particle size, and the results are as follows: Figure 5 As shown, the hydrated particle size of AuSPs is 182.5 nm, and the polydispersity index is 0.299. Therefore, the colloidal gold microspheres prepared in this invention have relatively uniform particle size and good dispersibility.

[0023] The AuNPs prepared in Examples 1 and 2 were compared with the conventional 30nm colloidal gold of the comparative example in terms of physical color. The results are as follows: Figure 6 As shown, the colors of Examples 1, 2 and the comparative examples remained basically unchanged, but the AuSPs of the present invention were darker, indicating that the colloidal gold microspheres of the present invention had superior colorimetric performance and a stronger colorimetric signal.

[0024] It should be noted that, in this document, relational terms such as "first" and "second" are used only to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such process, method, article, or apparatus.

[0025] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A method for synthesizing large-size colloidal gold microspheres, characterized in that: include: S1. Preparation of oil-phase colloidal gold nanoparticles: Dissolve oleylamine and chloroauric acid in toluene, heat to boiling and continue the reaction for 3 hours, cool to room temperature, add ethanol to precipitate, centrifuge to obtain oil-phase colloidal gold nanoparticles, and redissolve with toluene to obtain a toluene solution of oil-phase colloidal gold nanoparticles. S2. Preparation of microemulsion: The toluene solution of oil-phase colloidal gold nanoparticles obtained in step S1 is mixed and dissolved with distearate phosphatidylethanolamine-polyethylene glycol-n-heptanoic acid, and then added to an aqueous solution of sodium dodecyl sulfate. The mixture is then ultrasonically emulsified to form a microemulsion. S3. Solidification into spheres: Remove toluene from the microemulsion in step S2, collect the precipitate by centrifugation of the remaining system, wash and re-dissolve to obtain large-sized colloidal gold microspheres.

2. The method for synthesizing large-size colloidal gold microspheres according to claim 1, characterized in that: In step S2, the microemulsion is prepared by ultrasonic emulsification using a cell disruptor. The emulsification time is 1-10 min, the emulsification power is 100-300 W, and the ultrasonic working mode is 5 s operation followed by 10 s interval. The concentration of sodium dodecyl sulfate aqueous solution in step S2 is 8.7 mM.

3. The method for synthesizing large-size colloidal gold microspheres according to claim 2, characterized in that: The toluene removal method in step S3 is rotary evaporation or vacuum drying.

4. The method for synthesizing large-size colloidal gold microspheres according to claim 3, characterized in that: In step S3, the centrifugation conditions for the system after toluene removal are: rotation speed 8000-12000 rpm, centrifugation time 10-30 min, and the purpose of centrifugation is to collect the precipitate.

5. The method for synthesizing large-size colloidal gold microspheres according to claim 4, characterized in that: The precipitate washing step in step S3 is as follows: The precipitate was washed twice with ethanol as a detergent, then washed with ultrapure water until the supernatant was colorless, and finally reconstituted with ultrapure water or buffer solution.

6. The method for synthesizing large-size colloidal gold microspheres according to claim 5, characterized in that: The buffer solution is one or more of ultrapure water, phosphate buffer, or Tris-HCl buffer.

7. The method for synthesizing large-size colloidal gold microspheres according to claim 6, characterized in that: The volume ratio of toluene in step S1 to sodium dodecyl sulfate aqueous solution in step S2 is 1:4-1:10; In step S2, the amount of distearate phosphatidylethanolamine-polyethylene glycol-n-heptanoic acid added is 4 mg.

8. The method for synthesizing large-size colloidal gold microspheres according to claim 7, characterized in that: The large-sized colloidal gold microspheres prepared in step S3 have a diameter of 0.1-0.2 μm, are attached to the surface with carboxyl active groups, and have a polydispersity index ≤0.299.

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