CN106620702A - Preparation method of gold/nitrogen-doped hollow carbon nanosphere core-shell material - Google Patents

Abstract

A method for preparing a gold/nitrogen-doped hollow carbon nanosphere core-shell material belongs to the field of biomedical materials. First, a gold sol is prepared by a sol-gel method, and secondly, SiO ₂ is coated on the surface of gold particles by an improved stober method. Then Au@SiO ₂ @PDA spheres were formed by self-polymerization of dopamine in a weakly alkaline aqueous environment, and finally calcined at high temperature and etched with NaOH to obtain gold/nitrogen-doped hollow carbon nanosphere core-shell materials. The method of the present invention is simple, mild, and environmentally friendly, and will not increase additional toxicity. The particle size of the prepared gold/nitrogen-doped hollow carbon nanosphere core-shell material is about 165-175 nm, and the product is used as a near-infrared photothermal therapeutic agent for treating tumors Not only has good biocompatibility and photothermal stability, but also has the characteristics of a hollow structure inside, which increases the rate of particle exchange and improves the efficiency of photothermal conversion.

Description

A kind of preparation method of gold/nitrogen-doped hollow carbon nanosphere core-shell material

technical field

The invention belongs to the field of biomedical materials, in particular to the photothermal treatment technology of minimally invasive tumors.

Background technique

Cancer (malignant tumor) is an increasing threat to human health, and its incidence is increasing every year, and its mortality rate remains high. Although many treatments and treatments have been developed to treat cancer, the survival rate of cancer patients is still very low. Nowadays, the clinical methods of treating cancer are mainly through surgery, chemotherapy and radiotherapy. However, due to the metastatic and invasive nature of cancer, in the advanced stage of cancer, general surgery can no longer cure cancer, and chemotherapy and radiotherapy are more feasible methods. However, the specificity of chemotherapy and radiotherapy to tumor tissue is poor, which makes the treatment effect poor and has certain toxic and side effects. Therefore, improving the cure efficiency of cancer patients and reducing the toxic and side effects of cancer treatment methods are urgent problems to be solved in the current cancer treatment process.

Photothermal therapy (PTT) is a minimally invasive tumor treatment developed in recent years.

The technology is mainly to kill tumor cells by directly irradiating light of a certain wavelength to the tumor site to increase the temperature of the tumor site. Photothermal therapy can use long-wavelength light. Due to its low energy, the damage to healthy tissues and cells is relatively small, and it also greatly reduces the toxicity of the systemic system. Therefore, photothermal therapy has become a very potential alternative to surgical treatment today. One of the tumor techniques. Therefore, it is very important to develop photothermal therapeutic agents with good biocompatibility, simple synthesis method, high photothermal conversion efficiency and photothermal stability.

The materials of existing near-infrared photothermal therapeutic agents for treating tumors are mainly composed of noble metal nanoparticles, carbon materials, and organic dyes. Noble metal nanoparticles have high photothermal conversion efficiency and strong absorption capacity in the near-infrared region, but they are expensive; carbon materials have a large photothermal conversion area and high photothermal conversion efficiency, although the absorption capacity in the near-infrared region is general, but the cost is low; The compounding of carbon materials not only has low cost, but also has high light-to-heat conversion efficiency and strong near-infrared absorption ability.

The better absorption and scattering intensities of gold nanoparticles make them have great application potential in photothermal therapy. Gold nanoparticles also have the advantages of high biological safety, simple preparation, stable surface properties, and adjustable optical properties. Carbon materials have good biocompatibility and also show certain application potential in photothermal therapy. Therefore, combining carbon materials with gold nanoparticles and exerting their synergistic effects is expected to establish a multifunctional photothermal therapy platform with good effects. .

Contents of the invention

In order to solve the above problems, the purpose of the present invention is to propose a method for preparing a gold/nitrogen-doped hollow carbon nanosphere core-shell material that can be used as a near-infrared photothermal therapy agent for treating tumors.

The present invention comprises the following steps:

1) Mix boiling water with HAuCl ₄ aqueous solution and trisodium citrate aqueous solution, stir and reflux, then cool down, then mix with PVP aqueous solution for modification reaction, centrifuge after the reaction, take the solid phase and disperse it in water to obtain gold sol;

2) After mixing and stirring gold sol, ethanol, deionized water, and ammonia water, add TEOS dropwise to the mixed solution and stir at room temperature, then centrifuge, take the solid matter, wash it with water and alcohol, and dry it to obtain Au@SiO ₂ balls;

3) Dissolve Au@SiO ₂ balls in Tris aqueous solution with a concentration of 0.01-0.02 M and a pH value of 8.0-8.5, add dopamine hydrochloride aqueous solution after ultrasonication, stir at room temperature, centrifuge, take the solid phase for washing, and dry. Obtain Au@SiO ₂ @PDA spherical solid powder;

4) Under the protection of N ₂ , the Au@SiO ₂ @PDA spherical solid powder was calcined in a tube furnace to obtain a carbonized product; then the carbonized product was soaked in a NaOH aqueous solution with a concentration of 1 M, and centrifuged to obtain a solid phase Washing to neutrality and drying to obtain gold/nitrogen-doped hollow carbon nanosphere core-shell materials (Au@N-HCNs).

In the present invention, the gold sol is firstly prepared by the sol-gel method, secondly, SiO ₂ is coated on the surface of gold particles by the improved Stober method, and then Au@SiO ₂ @PDA balls are formed by the self-polymerization of dopamine in a weakly alkaline aqueous environment, and finally The gold/nitrogen doped hollow carbon nanosphere core-shell material is obtained after high-temperature calcination and NaOH etching.

In particular, the present invention uses a Tris aqueous solution with a concentration of 0.01-0.02 M and a pH value of 8.0-8.5. This environment is conducive to the polymerization of dopamine, and dopamine will self-polymerize and adhere to the surface of Au@SiO ₂ spheres uniformly.

The method of the invention is not only simple, mild and environmentally friendly, but also does not increase additional toxicity. The particle size of the gold/nitrogen-doped hollow carbon nanosphere core-shell material prepared by the present invention is about 165-175 nm. As a near-infrared photothermal therapeutic agent for treating tumors, the product of the present invention not only has good biocompatibility and photothermal Stability, and has the characteristics of a hollow structure inside, which increases the rate of particle exchange and improves the efficiency of light-to-heat conversion.

Further, in step 1) of the present invention, the mixing molar ratio of AuCl ₃ in the HAuCl ₄ aqueous solution, trisodium citrate in the trisodium citrate aqueous solution, and PVP in the PVP aqueous solution is 1:6:2. Strictly control the molar ratio of HAuCl ₄ and trisodium citrate to 1:6. After adding the sodium citrate dropwise, the color of the solution will obviously change from yellow to colorless to black to purple, and finally turn into wine red. The particle size of the gold particles after the chloroauric acid is reduced by trisodium citrate is about 15 nm. After reacting for half an hour, the solution was cooled to room temperature, modified with PVP and stirred overnight. The molar ratio of trisodium citrate and PVP was 3:1. After the reaction is completed, centrifuge, and dissolve the obtained PVP-modified gold sol in H ₂ O, which is convenient for coating SiO ₂ in the later stage.

In step 1) of the present invention, the stirring speed is 600-700 r/min, and the gold nanoparticles formed at this speed are relatively uniform.

In step 1) of the present invention, the centrifugal speed is 11800 r/min, which can minimize the loss of gold particles. If the rotation speed is too high, the centrifuge tube will be damaged; if the rotation speed is too low, the gold particles will be lost too much. Centrifuge for 1-2 times. If the centrifugation times are too many, the gold nanoparticles are more likely to aggregate.

In step 2) of the present invention, the mixing volume ratio of TEOS, ammonia water, gold sol, deionized water and ethanol is 1:2:4:12:20, and the particle size of _SiO2 supported by gold nanoparticles is relatively uniform under this ratio .

In the step 2), the stirring speed at room temperature is 600-700 r/min, and the centrifugal speed is 10000 r/min. The Au@SiO ₂ spheres formed in this way are relatively uniform, with a particle size of about 150 nm, which not only yields more, but also samples with small particle sizes are more suitable for photothermal conversion.

In the step 3), the mixing mass ratio of the Au@SiO ₂ spheres to dopamine hydrochloride is 1:1-3, and the polydopamine wrapped in this ratio is relatively uniform, and the photothermal effect is better. The ultrasonic power was 80 W, and the ultrasonic time was 0.5 h. If the ultrasonic time is too short, the sample will be dispersed unevenly in the Tris aqueous solution, which will affect the coating of polydopamine; if the ultrasonic time is too long, although the sample will be dispersed uniformly, it will consume time and energy. The stirring speed at room temperature is 600-700 r/min; the centrifugal speed is 10000 r/min. Under the reaction conditions, the polydopamine layer adhered uniformly and had a moderate thickness.

In the step 4), during the calcination, the temperature was raised to 800 °C at a heating rate of 5 °C/min and then kept for 2 h. When the present invention is calcined at a temperature of 800°C, the degree of carbonization is the best. If the carbonization temperature is too high, the loss of carbon and nitrogen elements will be serious; if the carbonization temperature is too low, there will be more impurities in the carbon material, and the degree of carbonization will be low, resulting in a lower degree of graphitization. When the heating rate is 5 ℃/min, it can not only ensure polydopamine polycondensation and carbonization to generate carbon materials during pyrolysis, but also reduce the loss of carbon materials during pyrolysis.

In the step 4), the soaking temperature is 75°C and the soaking time is 3 h. The addition of NaOH is to remove SiO ₂ by etching, and a low concentration NaOH solution is sufficient. Increasing the temperature can shorten the etching time, and the temperature of 75 ℃ is more suitable. The reaction time is 3 h to ensure that SiO ₂ is completely etched away.

In the step 4), the centrifugal speed is 10000 r/min. If the centrifugation speed is too small, the sample will be lost too much; if the centrifugation speed is too high, it will consume time and energy.

Description of drawings

Fig. 1 is a TEM image of Au@SiO ₂ spheres prepared in the present invention.

Fig. 2 is a TEM image of Au@SiO ₂ @PDA spheres prepared in the present invention.

Fig. 3 is a TEM image of Au@SiO ₂ @N-CNs prepared in the present invention.

Fig. 4 is a TEM image of the Au@N-HCNs core-shell material prepared in the present invention.

Fig. 5 is an X-ray diffraction pattern of the Au@N-HCNs core-shell material prepared in the present invention.

Fig. 6 is a Raman spectrum of the Au@N-HCNs core-shell material prepared in the present invention.

Fig. 7 is a temperature rise curve of the Au@N-HCNs core-shell material prepared by the present invention under near-infrared irradiation.

Fig. 8 is the cycle rate of the heating curve of the Au@N-HCNs core-shell material prepared by the present invention under near-infrared irradiation.

detailed description

1. Preparation process:

1. Embodiment 1:

(1) Preparation of PVP-modified gold sol: Take 30 mL of water in a 100 mL single-necked round-bottom flask and heat to boiling, add 100 μL of 150 mM HAuCl ₄ aqueous solution and 74 μL of 34 mM trisodium citrate aqueous solution, Reflux for 0.5 h at a stirring speed of r/min. After the solution was cooled to room temperature, 5.86 mL of 1.28 mM PVP aqueous solution was added and stirred overnight.

Put the reacted solution in a centrifuge tube and centrifuge at 11800 r/min for 1 to 2 times, take out the supernatant with a dropper, and disperse the remaining solid-phase gold sol in 8 mL deionized water to obtain gold sol for later use.

(2) Preparation of Au@SiO ₂ spheres: take 2 mL of gold sol and sonicate for 3 to 5 min at a power of 80 W, place 1 mL of ammonia water, 6 mL of deionized water, and 10 mL of ethanol in a 50 mL single-port In a round bottom flask, stir at 700 r/min for 0.5 h at room temperature. Finally, 0.5 mL TEOS was added, stirred at room temperature at 700 r/min for 6 h, then centrifuged at 1000 r/min to obtain the solid phase, washed with water and alcohol three times each, and dried in an oven at 60 °C to obtain Au@ _SiO2 balls.

(3) Preparation of Au@SiO ₂ @PDA balls: Weigh 50 mg of Au@SiO ₂ balls into 100 mL Tris aqueous solution ( p H=8.5, 0.01 M), and sonicate at 80 W for 0.5 h, then add 50 mg of dopamine hydrochloride, stir at room temperature at 700 r/min for 10 h, centrifuge at 1000 r/min to obtain the solid phase, wash with water and alcohol for 3 times, and dry in an oven at 60 °C. Au@SiO ₂ @PDA spherical solid powder was obtained.

(4) Preparation of Au@N-HCNs: Au@SiO ₂ @PDA spherical solid powder was placed in a tube furnace, and N ₂ was introduced for 10-20 min in advance to exhaust the air in the quartz tube, and then 5 The heating rate of ℃/min was increased to 800 ℃ for 2 h to obtain carbide Au@SiO ₂ @N-CNs. Soak the carbonized material in a 0.1 M NaOH aqueous solution at 75 °C for 3 h in a water bath, centrifuge at 1000 r/min, take the solid phase and wash it with deionized water until neutral, and dry it in an oven at 60 °C to obtain gold/nitrogen Doped hollow carbon nanosphere core-shell material Au@N-HCNs.

2. Embodiment 2:

(1) Preparation of PVP-modified gold sol: Take 30 mL of water in a 100 mL single-necked round-bottom flask and heat to boiling, add 100 μL of 150 mM HAuCl ₄ aqueous solution and 74 μL of 34 mM trisodium citrate aqueous solution, Reflux for 0.5 h at a stirring speed of r/min. After the solution was cooled to room temperature, 5.86 mL of 1.28 mM PVP aqueous solution was added and stirred overnight.

Put the reacted solution in a centrifuge tube and centrifuge at 11800 r/min for 1-2 times, take out the supernatant with a dropper, and disperse the remaining solid-phase gold sol in 8 mL deionized water to obtain the gold sol for later use.

(2) Preparation of Au@SiO ₂ spheres: take 2 mL of gold sol and sonicate for 3-5 min at a power of 80 W, place 1 mL of ammonia water, 6 mL of deionized water, and 10 mL of ethanol in a 50 mL single-port circle. In a bottom flask, stir at 700 r/min for 0.5 h at room temperature. Finally, 0.5 mL TEOS was added, stirred at room temperature at 700 r/min for 6 h, then centrifuged at 1000 r/min to obtain the solid phase, washed with water and alcohol three times each, and dried in an oven at 60 °C to obtain Au@ _SiO2 balls.

(3) Preparation of Au@SiO ₂ @PDA balls: Weigh 50 mg of Au@SiO ₂ balls into 100 mL Tris aqueous solution ( p H=8.5, 0.01 M), and sonicate at 80 W for 0.5 h, then add 100 mg of dopamine hydrochloride, stir at room temperature at 700 r/min for 10 h, then centrifuge at 1000 r/min to obtain the solid phase, wash with water and alcohol for 3 times, and dry in an oven at 60°C to obtain Au@SiO ₂ @PDA spherical solid powder.

(4) Preparation of Au@N-HCNs: Au@SiO ₂ @PDA spherical solid powder was placed in a tube furnace, and N ₂ was introduced for 10-20 min in advance to exhaust the air in the quartz tube, and then 5 The heating rate of ℃/min was increased to 800 ℃ for 2 h to obtain carbide Au@SiO ₂ @N-CNs. Soak the carbonized material in a 0.1M NaOH aqueous solution at 75 °C for 3 h in a water bath, centrifuge at 1000 r/min, take the solid phase and wash it with deionized water until neutral, and dry it in an oven at 60 °C to obtain gold/nitrogen Doped hollow carbon nanosphere core-shell material Au@N-HCNs.

3. Embodiment 3:

(1) Preparation of PVP-modified gold sol: Take 30 mL of water and heat it to boiling in a 100 mL single-necked round-bottom flask, add 100 μL of 150 mM HAuCl ₄ aqueous solution and 74 μL of 34 mM trisodium citrate aqueous solution, Reflux for 0.5 h at a stirring speed of r/min. After the solution was cooled to room temperature, 5.86 mL of 1.28 mM PVP aqueous solution was added and stirred overnight.

Put the reacted solution in a centrifuge tube and centrifuge at 11800 r/min for 1-2 times, take out the supernatant with a dropper, and disperse the remaining solid-phase gold sol in 8 mL deionized water to obtain the gold sol for later use.

(2) Preparation of Au@SiO ₂ spheres: take 2 mL of gold sol and sonicate for 3 to 5 min at a power of 80 W, place 1 mL of ammonia water, 6 mL of deionized water, and 10 mL of ethanol in a 50 mL single-port In a round bottom flask, stir at 600 r/min for 0.5 h at room temperature. Finally, 0.5 mL TEOS was added, stirred at room temperature at 700 r/min for 6 h, centrifuged at 1000 r/min to obtain the solid phase, washed with water and alcohol three times each, and dried in an oven at 60 °C to obtain Au@ _SiO2 balls.

(3) Preparation of Au@SiO ₂ @PDA balls: Weigh 50 mg of Au@SiO ₂ balls into 100 mL Tris aqueous solution ( p H=8.5, 0.01 M), and sonicate at 80 W for 0.5 h, then add 150 mg dopamine hydrochloride, stir at 600 r/min at room temperature for 10 h, then centrifuge at 1000 r/min to obtain the solid phase, wash with water and alcohol for 3 times, and dry in an oven at 60 °C , to get Au@SiO ₂ @PDA spherical solid powder.

(4) Preparation of Au@N-HCNs: Au@SiO ₂ @PDA spherical solid powder was placed in a tube furnace, and N ₂ was introduced for 10-20 min in advance to exhaust the air in the quartz tube, and then 5 The heating rate of ℃/min was increased to 800 ℃ for 2 h to obtain carbide Au@SiO ₂ @N-CNs. Soak the carbonized material in a 0.1 M NaOH aqueous solution at 75 °C for 3 h in a water bath, centrifuge at 1000 r/min, take the solid phase and wash it with deionized water until neutral, and dry it in an oven at 60 °C to obtain gold/nitrogen Doped hollow carbon nanosphere core-shell material Au@N-HCNs.

2. Product characteristics:

The TEM images of Au@SiO ₂ spheres prepared in step 2 of the above examples are shown in Figure 1. It is obvious from Figure 1 that the particle size of gold particles is about 15 nm, while the particle size of Au@SiO ₂ is around 15 nm. 145-155nm.

The TEM images of the Au@SiO ₂ @PDA spheres prepared in step 3 of the above examples are shown in Figure 2. It can be seen from Figure 2 that PDA is uniformly coated on the surface of the Au@SiO ₂ spheres, Au@SiO ₂ The particle size of @PDA balls is about 165-175 nm.

The TEM images of the Au@SiO ₂ @N-CNs spheres prepared in step 4 of the above examples are shown in Figure 3. From Figure 3, it can be clearly seen that the particle size of the carbonized material has shrunk, which indicates that the PDA is carbonized The particle size of Au@SiO ₂ @N-CNs balls is 160-170 nm.

The TEM images of the Au@SiO ₂ @N-HCNs spheres prepared in step 4 of the above examples are shown in Figure 4. From Figure 4, it can be clearly seen that SiO ₂ has been completely removed, and the thickness of the carbon layer is 10-15 nm .

The X-ray diffraction pattern of the Au@N-HCNs core-shell material prepared in step 4 of each of the above examples is shown in FIG. 5 . According to the results of X-ray diffraction, the diffraction peaks of (111), (200), (220), and (311) crystal planes are basically consistent with the standard diffraction peaks of Au, and the diffraction peaks of (002) and (110) crystal planes are consistent with those of C. The shape-setting diffraction peaks are basically consistent. The Au@N-HCNs core-shell material prepared by the present invention is basically consistent with the standard card, which basically proves the composition and structure of the obtained material.

The Raman spectrum of the Au@N-HCNs core-shell material prepared in step 4) of each of the above examples is shown in Figure 6. It can be seen from the figure that the ratio of the _D peak to the G peak is ID / I _G = 0.916, indicating that the Au@N-HCNs core-shell material has a better degree of graphitization.

The temperature rise curve of the Au@N-HCNs core-shell material prepared in step 4) of each of the above examples under near-infrared irradiation is shown in Fig. 7 . The incident wavelength is 808 nm, and the laser intensity is 2 W/cm ² . Since the human body temperature is around 37°C, cancer cells can be blocked from the DNA synthesis of cancer cells and cause cancer cell apoptosis if kept at a high temperature of 45-50°C for 40 minutes. It can be seen from the figure that the temperature rise curves of materials with different concentrations after being illuminated, the suspensions containing 50 μg/mL, 100 μg/mL, 150 μg/mL, and 200 μg/mL of Au@N-HCNs core-shell materials were subjected to After 10 min of 808 nm laser irradiation, the temperature of the suspension increased by 9.5 ℃, 15.2 ℃, 15.9 ℃, and 17.2 ℃, respectively. The results indicate that Au@N-HCNs core-shell materials are more effective near-infrared photothermal therapeutic agents for treating tumors.

The cycle rate of the heating curve of the Au@N-HCNs core-shell material prepared in step 4) of each embodiment under near-infrared irradiation is shown in Figure 8 . It can be seen from the figure that the temperature of the 150 μg/mL suspension was increased by 15.9 ℃, 15.2 ℃, and 14.5 ℃ for three times, respectively. The core-shell material of HCNs will settle, which will reduce the particle collision velocity. But in general, Au@N-HCNs core-shell materials are relatively stable near-infrared photothermal therapeutic agents for treating tumors.

Claims (10)

1. a kind of preparation method of the hollow Nano carbon balls core-shell material of gold/N doping, it is characterised in that comprise the following steps：

1）By the water and HAuCl of boiling₄The aqueous solution, trisodium citrate aqueous solution mixing, after being stirred at reflux cool down, then again with PVP Aqueous solution mixing carries out modification reaction, and reaction is centrifuged after terminating, and takes solid phase dispersion in water, obtains final product aurosol；

2）By aurosol, ethanol, deionized water, ammoniacal liquor mix after, in mixed solution be added dropwise TEOS be stirred at room temperature after from The heart, takes solid matter with water and alcohol washing post-drying, obtains Au@SiO₂Ball；

3）By Au@SiO₂Ball be dissolved in concentration for 0.01～0.02 M,pH values are in 8.0～8.5 Tris aqueous solution, to add after ultrasound Enter the Dopamine hydrochloride aqueous solution, Jing centrifugations after being stirred at room temperature take solid phase washing, drying, obtain Au SiO₂@PDA ball pressed powders；

4）In N₂Under protection, by Au@SiO₂@PDA ball pressed powders are placed in tube furnace and calcine, and obtain carbide；Again by carbide It is placed in the NaOH aqueous solution that concentration is 1 M and soaks, Jing centrifugations obtains solid phase and wash to neutrality, drying, obtain golden/N doping Hollow Nano carbon balls core-shell material.

2. preparation method according to claim 1, it is characterised in that the step 1）In, HAuCl₄AuCl in the aqueous solution₃、 The mixing mol ratio of PVP is 1: 6: 2 in trisodium citrate and the PVP aqueous solution in trisodium citrate aqueous solution.

3. preparation method according to claim 2, it is characterised in that the step 1）In, the mixing speed is 600 ～700 r/min, the centrifugal rotational speed is 11800 r/min, is centrifuged 1～2 time.

4. preparation method according to claim 1, it is characterised in that the step 2）In, the TEOS, ammoniacal liquor, Jin Rong The mixed volume ratio of glue, deionized water and ethanol is 1: 2: 4: 12: 20.

5. preparation method according to claim 4, it is characterised in that the step 2）In, the speed that is stirred at room temperature is 600～700 r/min, the centrifugal rotational speed is 10000 r/min.

6. preparation method according to claim 1, it is characterised in that the step 3）In, the Au@SiO₂Ball is more with hydrochloric acid The mixing quality ratio of bar amine is 1: 1～3, and ultrasonic power is 80 W, and ultrasonic time is 0.5 h.

7. preparation method according to claim 6, it is characterised in that the step 3）In, the speed that is stirred at room temperature is 600～700 r/min；The centrifugal rotational speed is 10000 r/min.

8. preparation method according to claim 1, it is characterised in that the step 4）In, with 5 DEG C/min during the calcining Heating rate be warming up to 800 DEG C after keep 2 h.

9. the preparation method according to claim 1 or 8, it is characterised in that the step 4）In, the soaking temperature is 75 DEG C, the time is 3 h.

10. preparation method according to claim 1, it is characterised in that the step 4）In, the centrifugal rotational speed is 10000 r/min。