Size-controllable ultra-small gold nanoparticle GSH-Au-coated DPA and preparation method thereof

Abstract

The invention relates to a size-controllable ultra-small gold nanoparticle GSH-Au-DPA and a preparation method thereof, the preparation method comprises the following steps: S1, mixing a HAuCl4. 3H2O solution, a sodium carbonate solution and a glutathione solution to obtain a GSH-HAuCl4 solution, adding a reducing agent under ice-water bath and stirring conditions, and reacting to obtain a GSH-AuNPs solution; and S2, the GSH-AuNPs solution and the dopamine hydrochloride-buffer solution are mixed, stirring reaction is conducted, and the ultra-small gold nanoparticles GSH-Au-DPA with the controllable size are obtained. The sodium carbonate is adopted to adjust the pH value, the water phase preparation method is adopted to prepare the size-controllable ultra-small AuNPs, the particle size of the AuNPs prepared in the water phase is reduced, DPA is adopted for surface coating, the biocompatibility is improved, and the cytotoxicity is reduced. The obtained product is uniform and stable in particle size and simple and non-toxic in process.

Description

Technical Field

[0001] The invention relates to the technical field of nanomaterials, in particular to size-controllable ultra-small gold nanoparticle GSH-Au@DPA and a preparation method thereof. Background Art

[0002] Gold nanoparticles (AuNPs) refer to nanoscale gold particles, which often exist in the form of colloids. The size of AuNPs dispersed in a medium or solution is between 1 and 100 nm. People have prepared AuNPs of various shapes, mainly rods, spheres, shells, cages, polyhedrons, stars, etc. Due to the small size of AuNPs, the transfer of electrons between the ground state and the excited state causes collective oscillation of electrons, which then interacts with light, which makes AuNPs have excellent optical properties. In addition, AuNPs have the following characteristics: (1) The morphology and size are controllable and have good biocompatibility; (2) The properties are stable and can be retained in the blood for a long time; (3) The surface of AuNPs is easy to be modified; (4) It has a surface plasmon resonance effect. Due to the surface plasmon resonance effect, AuNPs have been proven to be excellent photothermal agents with high photothermal conversion efficiency and good tumor photothermal efficacy. In addition, AuNPs have a strong EPR effect and are easy to accumulate in tumor sites. These excellent characteristics mean that AuNPs have potential application value in tumor hyperthermia, delivery of anticancer drugs, CT angiography, etc.

[0003] Considering biocompatibility and toxicity, ultrasmall water-dispersible AuNPs are an attractive option for clinical applications. The commonly used AuNPs synthesis methods are the Turkevich method and the Brust-Schiffrin method, which can obtain 15-150 nm AuNPs in water and ultrasmall AuNPs in organic phases, respectively. In theory, ultrasmall water-dispersible AuNPs can be easily obtained by using a stabilizer and a reducing agent in an appropriate ratio. However, the particle size of AuNPs prepared in the aqueous phase is mostly greater than 10 nm, and the ultrasmall AuNPs prepared in the organic phase have the defects of using organic solvents to affect biocompatibility and cytotoxicity. Therefore, it is of particular practical significance to develop a simple synthetic method for the stable preparation of ultrasmall (<10 nm) AuNPs with controllable size in an aqueous phase. Summary of the invention

[0004] The purpose of the present invention is to overcome the above technical deficiencies, provide a size-controllable ultrasmall gold nanoparticle GSH-Au@DPA and a preparation method thereof, and solve the technical problem that it is difficult to synthesize stable aqueous phase ultrasmall gold nanoparticles when preparing gold nanoparticles in the prior art.

[0005] In order to achieve the above technical purpose, the technical solution provided by the present invention is: In a first aspect, the present invention provides a method for preparing size-controllable ultrasmall gold nanoparticles GSH-Au@DPA, comprising the following steps: S1, HAuCl4·3H2O solution, sodium carbonate solution and glutathione solution are mixed to obtain GSH-HAuCl4 solution, and a reducing agent is added in an ice-water bath with stirring to react to obtain a GSH-AuNPs solution; S2, mixing the GSH-AuNPs solution with dopamine hydrochloride-buffered solution, stirring and reacting, and obtaining size-controllable ultrasmall gold nanoparticles GSH-Au@DPA.

[0006] In a second aspect, the present invention provides a size-controllable ultrasmall gold nanoparticle GSH-Au@DPA prepared by the above preparation method, which is a nanoparticle with GSH-Au as the core and dopamine coated on the surface, and the average particle size is 4-6nm.

[0007] Compared with the prior art, the present invention has the following beneficial effects: The invention provides a simple aqueous phase preparation method for size-controllable ultrasmall gold nanoparticles GSH-Au@DPA. The method adopts sodium carbonate to adjust the pH value and adopts an aqueous phase preparation method to prepare size-controllable ultrasmall AuNPs, thereby avoiding the defects of low biocompatibility and high cytotoxicity of ultrasmall AuNPs prepared by using organic solvents, reducing the particle size of AuNPs prepared in the aqueous phase, and making the average particle size reach 3-5nm. The GSH-Au@DPA prepared by the method has a very small particle size, with an average particle size of 4-6nm, and is surface-coated with DPA, thereby further improving biocompatibility, reducing cytotoxicity, and being beneficial to subsequent biological applications. In addition, the particle size of the obtained product is uniform and stable, and the process is simple and non-toxic. BRIEF DESCRIPTION OF THE DRAWINGS

[0008] Figure 1 is a particle size distribution diagram of GSH-AuNPs and GSH-Au@DPA prepared in Example 1 of the present invention; Figure 2 is a particle size distribution diagram of GSH-AuNPs prepared with different concentrations of GSH in Examples 1 to 3 of the present invention; Figure 3 is the infrared spectra of GSH-AuNPs and GSH-Au@DPA prepared in Example 1 of the present invention; Figure 4 is the UV spectra of GSH-AuNPs and GSH-Au@DPA prepared in Example 1 of the present invention; Figure 5is a TEM image of GSH-AuNPs prepared in Example 1 of the present invention; Figure 6 is a TEM image of GSH-Au@DPA prepared in Example 1 of the present invention; Figure 7 This is a graph showing the particle size-time variation of GSH-Au@DPA prepared in Example 1 of the present invention. DETAILED DESCRIPTION

[0009] In order to make the purpose, technical solution and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the present invention.

[0010] In a first aspect, the present invention provides a method for preparing size-controllable ultrasmall gold nanoparticles GSH-Au@DPA, comprising the following steps: S1, HAuCl4·3H2O solution, sodium carbonate solution and glutathione solution are mixed to obtain GSH-HAuCl4 solution, and a reducing agent is added in an ice-water bath with stirring to react to obtain a GSH-AuNPs solution; S2, mixing the GSH-AuNPs solution with dopamine hydrochloride-buffered solution, stirring and reacting, and obtaining size-controllable ultrasmall gold nanoparticles GSH-Au@DPA.

[0011] Preferably, in step S1, in the GSH-HAuCl4 solution, the mass concentration of HAuCl4·3H2O is 0.01%, the concentration of sodium carbonate is 0.5-1.5 mmol / L, and the concentration of glutathione is 1.5-3.5 mg / L.

[0012] Preferably, in step S1, the reducing agent is NaBH4 solution.

[0013] More preferably, the concentration of NaBH4 solution is 0.4-0.6 mg / mL, and the volume ratio of GSH-HAuCl4 solution to NaBH4 solution is 50:(1-2). In the present invention, the concentration of NaBH4 solution in the reaction system is controlled by the concentration and the added volume, which is conducive to obtaining ultra-small gold nanoparticles with controllable size.

[0014] Preferably, in step S1, the temperature of the ice water bath is 0-4°C; and the stirring speed in the stirring condition is 1300-1500 rpm.

[0015] Preferably, in step S1, HAuCl4·3H2O solution, sodium carbonate solution and glutathione solution are mixed and stirred for 15 to 20 minutes to obtain GSH-HAuCl4 solution; after adding a reducing agent, react for 15 to 20 minutes to obtain GSH-AuNPs solution.

[0016] Preferably, in step S2, the dopamine hydrochloride buffer is obtained by mixing a dopamine hydrochloride solution, a Tris-HCl buffer and water; in the dopamine hydrochloride buffer, the ratio of dopamine hydrochloride to Tris-HCl is (1-3) mg: (0.5-1.5) mmol.

[0017] Further preferably, every 100 mL of dopamine hydrochloride-buffer includes 2 to 3 mL of dopamine hydrochloride solution and 1.0 to 1.5 mL of Tris-HCl buffer; wherein the concentration of the dopamine hydrochloride solution is 1 mg / mL; the concentration of the Tris-HCl buffer is 1 mol / L, and the pH is 8.5.

[0018] Preferably, in step S2, the volume ratio of the GSH-AuNPs solution to the dopamine hydrochloride-Tris solution is 1:(1.0-1.2).

[0019] Preferably, in step S2, the reaction is stirred at room temperature for 3 to 4 h.

[0020] In a second aspect, the present invention provides a size-controllable ultra-small gold nanoparticle GSH-Au@DPA prepared by the above preparation method, which is a nanoparticle with GSH-Au as the core and dopamine coated on the surface, and has an average particle size of 4 to 6 nm.

[0021] The main mechanism of action of the present invention: (1) The present invention adopts an aqueous phase preparation method to prepare ultra-small AuNPs with controllable size, thereby avoiding the defects of using organic solvents to prepare ultra-small AuNPs that affect biocompatibility and cytotoxicity, and reducing the particle size of AuNPs prepared in the aqueous phase, so that the average particle size reaches 4 to 6 nm; and the GSH-Au@DPA prepared by the method of the present invention is surface-coated with DPA, which increases the stability of gold nanoparticles, improves biocompatibility, and reduces cytotoxicity, which is beneficial to subsequent biological applications, such as tumor imaging and tumor photothermal therapy, etc.; and the obtained product has a high yield, uniform and stable particle size, and a simple and non-toxic process; (2) Sodium carbonate is used in the present invention to adjust the pH value of the reaction system. It has a weak alkalinity and a certain buffering effect. It can fine-tune the pH value of the system, which is conducive to the synthesis of gold nanoparticles with small particle size; (3) In the present invention, the size of AuNPs is adjusted by controlling the concentration of the stabilizer GSH and the ratio of gold.

[0022] The present invention is described in detail below through specific embodiments.

[0023] Example 1 The specific steps for preparing GSH-Au@DPA in this embodiment are as follows: (1) 0.5 mL of 1 wt% HAuCl4·3H2O solution, 0.5 mL of 0.1 mol / L Na2CO3 solution and 0.5 mL of 0.25 mg / mL GSH solution were uniformly mixed and added to a 50 mL volumetric flask with double distilled water to obtain a GSH-HAuCl4 solution, in which the mass concentration of HAuCl4·3H2O was 0.01%, the concentration of sodium carbonate was 1 mmol / L, and the concentration of glutathione was 2.5 mg / L. The solution was transferred to a 100 mL round-bottom flask, and after being placed in an ice-water bath and vigorously stirred (at a speed of about 1400 rpm) for 15 min, a reducing agent 1.5 mL of 0.5 mg / mL NaBH4 aqueous solution was added, and the solution quickly turned watermelon red. The reaction was continued for 15 min to obtain naked gold nanoparticles GSH-AuNPs.

[0024] (2) After 1 mL of 1M Tris-HCl solution (pH 8.5) and 2 mL of 1 mg / mL dopamine hydrochloride solution were uniformly mixed, the volume was fixed with double distilled water in a 100 mL volumetric flask to obtain dopamine hydrochloride-Tris solution. Then 100 mL of GSH-AuNPs solution and 100 mL of dopamine hydrochloride-Tris solution were mixed and the reaction was continued under stirring at room temperature for 3 h to obtain a stable GSH-Au@DPA sample 1 with dopamine coated on the surface.

[0025] Example 2 The specific steps for preparing GSH-Au@DPA in this embodiment are as follows: (1) 0.5 mL of 1 wt% HAuCl4·3H2O solution, 0.5 mL of 0.1 mol / L Na2CO3 solution and 0.5 mL of 0.15 mg / mL GSH solution were uniformly mixed and added to a 50 mL volumetric flask with double distilled water to obtain a GSH-HAuCl4 solution, in which the mass concentration of HAuCl4·3H2O was 0.01%, the concentration of sodium carbonate was 1 mmol / L, and the concentration of glutathione was 1.5 mg / L. The solution was transferred to a 100 mL round-bottom flask, and after being placed in an ice-water bath and vigorously stirred for 15 min, a reducing agent 1.5 mL of 0.5 mg / mL NaBH4 aqueous solution was added, and the solution quickly turned into watermelon red. The reaction was continued for 15 min to obtain naked gold nanoparticles GSH-AuNPs.

[0026] (2) After 1 mL of 1M Tris-HCl solution (pH 8.5) and 2 mL of 1 mg / mL dopamine hydrochloride solution were uniformly mixed, the volume was fixed with double distilled water in a 100 mL volumetric flask to obtain dopamine hydrochloride-Tris solution. Then 100 mL of GSH-AuNPs solution and 100 mL of dopamine hydrochloride-Tris solution were mixed and the reaction was continued under stirring at room temperature for 3 h to obtain a stable GSH-Au@DPA sample 2 with dopamine coated on the surface.

[0027] Example 3 The specific steps for preparing GSH-Au@DPA in this embodiment are as follows: (1) 0.5 mL 1 wt% HAuCl4·3H2O solution, 0.5 mL 0.1 mol / L Na2CO3 solution and 0.5 mL 0.35 mg / mL GSH solution were uniformly mixed and added to a 50 mL volumetric flask with double distilled water to obtain a GSH-HAuCl4 solution, in which the mass concentration of HAuCl4·3H2O was 0.01%, the concentration of sodium carbonate was 1 mmol / L, and the concentration of glutathione was 3.5 mg / L. The solution was transferred to a 100 mL round-bottom flask, and after being placed in an ice-water bath and vigorously stirred for 15 min, a reducing agent 1.5 mL 0.5 mg / mL NaBH4 aqueous solution was added, and the solution quickly turned watermelon red. The reaction was continued for 15 min to obtain naked gold nanoparticles GSH-AuNPs.

[0028] (2) After 1 mL of 1M Tris-HCl solution (pH 8.5) and 2 mL of 1 mg / mL dopamine hydrochloride solution were uniformly mixed, the volume was fixed with double distilled water in a 100 mL volumetric flask to obtain dopamine hydrochloride-Tris solution. Then 100 mL of GSH-AuNPs solution and 100 mL of dopamine hydrochloride-Tris solution were mixed and the reaction was continued under stirring at room temperature for 3 h to obtain a stable GSH-Au@DPA sample 3 with dopamine coated on the surface.

[0029] The particle sizes of GSH-AuNPs and GSH-Au@DPA in Examples 1 to 3 above were tested, wherein the average particle size of GSH-AuNPs was in the range of 3 to 5 nm, and the average particle size of GSH-Au@DPA was in the range of 4 to 6 nm.

[0030] Specifically, the GSH-AuNPs and GSH-Au@DPA prepared in Example 1 were selected for particle size distribution statistics. The results are as follows: Figure 1 As shown, the particle size distribution is narrow, all below 10 nm.

[0031] The particle size distribution of GSH-AuNPs prepared with different concentrations of GSH in Examples 1 to 3 is as follows: Figure 2 As shown, as the GSH concentration increases, the particle size of the obtained GSH-AuNPs tends to decrease, but high or low GSH concentrations will widen the particle size distribution range of GSH-AuNPs. Therefore, in the present invention, it is preferred to use 0.25 mg / mL GSH solution to prepare the GSH-HAuCl4 solution.

[0032] The GSH-AuNPs and GSH-Au@DPA prepared in Example 1 were characterized by infrared spectroscopy. Figure 3 shown.

[0033] The UV spectra of GSH-AuNPs and GSH-Au@DPA prepared in Example 1 of the present invention are shown in FIG. Figure 4 As shown, the maximum ultraviolet absorption peak red-shifted after dopamine was coated.

[0034] The morphology of GSH-AuNPs and GSH-Au@DPA prepared in Example 1 of the present invention was characterized. Figure 5 and Figure 6 shown.

[0035] At the same time, the particle size-time variation of GSH-Au@DPA was also tested. The results are as follows: Figure 7 As shown, it can be seen that GSH-Au@DPA can always maintain a relatively stable particle size range, and will not produce large particle size fluctuations over time, which proves that the use of a simple size-controllable ultrasmall gold nanoparticle GSH-Au@DPA aqueous phase preparation method of the present invention can obtain GSH-Au@DPA with uniform and stable particle size and good storage stability.

[0036] Comparative Example 1 The only difference from Example 1 is that in step (1), the sodium carbonate solution is replaced with a sodium hydroxide solution of the same concentration, and the other steps and conditions are the same as those in Example 1.

[0037] The results showed that after replacing the sodium carbonate solution with sodium hydroxide solution, the gold nanoparticles formed had a larger particle size, ranging from 100nm to 300nm. This is because the alkalinity of sodium carbonate is weak, and the pH value adjustment of the entire reaction system is relatively gentle and will not cause strong pH fluctuations, which has a good effect on stabilizing the particle size.

[0038] Comparative Example 2 The only difference from Example 1 is that in step (1), the vigorous stirring in the ice water bath is replaced with a room temperature water bath, and the other steps and conditions are the same as those in Example 1.

[0039] The results showed that the synthesized nanoparticles were large and uncontrollable in size (100nm, 500nm), and had an uneven size distribution (100-5000nm).

[0040] Comparative Example 3 The only difference from Example 1 is that in step (2), the reaction time is extended to 5 h, and the other steps and conditions are the same as in Example 1.

[0041] The results showed that the size of the synthesized nanoparticles was not uniform. There were fewer nanoparticles with a size of about 10 nm, and most of the particles increased to 100-500 nm. There were large differences in size and a wide particle size distribution.

[0042] The present invention discloses a simple aqueous phase preparation method for ultra-small gold nanoparticles (GSH-Au@DPA) with controllable size, the steps of which include: mixing chloroauric acid (HAuCl4·3H2O) solution, sodium carbonate (NaCO3) solution and glutathione (GSH), then adding a reducing agent under ice-water bath and vigorous stirring conditions to obtain naked gold nanoparticles (GSH-AuNPs), and finally coating dopamine on the surface of GSH-AuNPs to obtain stable GSH-Au@DPA. The present invention adopts an aqueous phase preparation method to prepare gold nanoparticles, avoiding the defects of using organic solvents in traditional preparation methods to affect biocompatibility and cytotoxicity; solving the problem of excessively large particle size of particles prepared in aqueous phase in traditional preparation methods; the GSH-Au@DPA prepared by the method improves biocompatibility, reduces cytotoxicity, and is conducive to subsequent biological applications; and the obtained product has a high yield, uniform and stable particle size, and a simple and non-toxic process.

[0043] The specific implementation of the present invention described above does not constitute a limitation on the protection scope of the present invention. Any other corresponding changes and modifications made based on the technical concept of the present invention should be included in the protection scope of the claims of the present invention.

Claims

1. A method for preparing size-controllable ultra-small gold nanoparticles GSH-Au@DPA, characterized in that: The following steps are involved: S1, HAuCl4·3H2O solution, sodium carbonate solution and glutathione solution are mixed to obtain GSH-HAuCl4 solution, and a reducing agent is added in an ice-water bath with stirring to react to obtain a GSH-AuNPs solution; S2, mixing the GSH-AuNPs solution with dopamine hydrochloride-buffered solution, stirring and reacting, and obtaining size-controllable ultrasmall gold nanoparticles GSH-Au@DPA.

2. The method for preparing the size-controllable ultrasmall gold nanoparticles GSH-Au@DPA according to claim 1, characterized in that: In step S1, in the GSH-HAuCl4 solution, the mass concentration of HAuCl4·3H2O is 0.01%, the concentration of sodium carbonate is 1-3 mmol / L, and the concentration of glutathione is 3-7 mg / L.

3. The method for preparing the size-controllable ultrasmall gold nanoparticles GSH-Au@DPA according to claim 1, characterized in that: In step S1, the reducing agent is NaBH4 solution.

4. The method for preparing the size-controllable ultrasmall gold nanoparticles GSH-Au@DPA according to claim 3, characterized in that: The concentration of NaBH4 solution is 0.4-0.6 mg / mL, and the volume ratio of GSH-HAuCl4 solution to NaBH4 solution is 50:(1-2).

5. The method for preparing the size-controllable ultrasmall gold nanoparticles GSH-Au@DPA according to claim 1, characterized in that: In step S1, the stirring speed in the stirring condition is 1300-1500 rpm; the HAuCl4·3H2O solution, the sodium carbonate solution and the glutathione solution are mixed and stirred for 15-20 minutes to obtain a GSH-HAuCl4 solution; After adding the reducing agent, the reaction was continued for 15 to 20 minutes to obtain a GSH-AuNPs solution.

6. The method for preparing the size-controllable ultrasmall gold nanoparticles GSH-Au@DPA according to claim 1, characterized in that: In step S2, the dopamine hydrochloride buffer is obtained by mixing a dopamine hydrochloride solution, a Tris-HCl buffer and water; in the dopamine hydrochloride buffer, the ratio of dopamine hydrochloride to Tris-HCl is (1-3) mg: (0.5-1.5) mmol.

7. The method for preparing size-controllable ultrasmall gold nanoparticles GSH-Au@DPA according to claim 6, characterized in that: Each 100 mL of dopamine hydrochloride-buffered solution includes 1 to 3 mL of dopamine hydrochloride solution and 0.5 to 1.5 mL of Tris-HCl buffer; wherein the concentration of the dopamine hydrochloride solution is 1 mg / mL, the concentration of the Tris-HCl buffer is 1 mol / L, and the pH value is 8.

5.

8. The method for preparing the size-controllable ultrasmall gold nanoparticles GSH-Au@DPA according to claim 1, characterized in that: In step S2, the volume ratio of the GSH-AuNPs solution to the dopamine hydrochloride-Tris solution is 1:(0.8-1.2); the reaction is stirred at room temperature for 3-4 hours. 9 . The size-controllable ultrasmall gold nanoparticles GSH-Au@DPA prepared by the preparation method according to any one of claims 1 to 8 .

10. The size-controllable ultrasmall gold nanoparticle GSH-Au@DPA according to claim 9, characterized in that: It is a nanoparticle with GSH-Au as the core and dopamine coated on the surface, with an average particle size of 4~6nm.

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