CN109604632B - Method for preparing gold nanoparticles by using polymyxin E as template - Google Patents
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Abstract
The invention relates to a method for preparing gold nanoparticles by taking polymyxin E as a template. The method comprises the following specific steps: preparing 0.1-3mg/mL polymyxin E solution, 0.01-10mg/mL chloroauric acid solution and 0.1-10mg/mL ascorbic acid solution; uniformly mixing the polymyxin E solution and the chloroauric acid solution according to the molar ratio of the polymyxin E solution to the chloroauric acid solution of 1:0.1-10, putting the mixed solution into a metal bath, adding a phosphate buffer solution with the pH value of 7.4, adding distilled water, carrying out incubation, then adding an equal amount of ascorbic acid solution in portions, wherein the interval time of each time is 1-10min, and changing the solution from light yellow to blue-violet to obtain the gold nanoparticles. The prepared nano gold particles have photo-thermal conversion performance, uniform particle size and good stability.
Description
Technical Field
The invention belongs to the technical field of metal materials, and relates to a method for preparing gold nanoparticles by using polymyxin E as a template.
Background
The metal nano-particles have wide application prospect in various fields of molecular devices, chemical/biological sensors, catalysis, photoelectronic materials and the like. The preparation of metal nanoparticles is mainly performed by a sol-gel method, namely, a reducing agent metal salt is utilized and then the nanoparticles are obtained by sintering, and the nanoparticles obtained by the method have larger particle size and wide particle size distribution and are difficult to solve the problem of agglomeration.
In recent years, photothermal therapy of cancer has become a hot point of research by utilizing the unique optical properties of noble metal nanoparticles. The noble metal nanoparticles have plasma resonance capability, absorb light energy and convert the light energy into heat energy, so that the temperature is locally raised, and cancer cells are killed. The photothermal conversion ability of the noble metal nanoparticles depends on their shape and size. However, it is still difficult to prepare noble metal nanoparticles having high photothermal conversion performance and high biocompatibility. The nano gold particles are an inert metal thermotherapy agent with high biocompatibility and good photothermal conversion performance at present. At this time, the gold nanoparticles have special morphology including spherical shell, rod, dendritic, etc. Therefore, the photo-thermal therapeutic agent of the nano gold particles with special shapes has good development and application prospects.
At present, the template method is a very important method for preparing nanoparticles. The template method is characterized in that no matter the reaction occurs in a liquid phase or a gas phase, the nano gold particles can be reduced and generated in an effective area of the template. In addition, the template can regulate and control the particle size and the morphology of the nano gold particles and play a role in stabilizing the nano particles. Nowadays, most of the templates used are artificially synthesized organic polymers. Most of the polymers are toxic and not easy to be applied in biological systems. And the biological macromolecular templates such as lysozyme, trypsin, bovine serum albumin and the like with good biocompatibility are difficult to form nano particles with the light-heat conversion performance. Therefore, there is a need to further develop a new template for preparing gold nanoparticles having high light-heat conversion properties.
Disclosure of Invention
Aiming at the defects of the prior art, the invention provides a method for preparing nano gold particles by taking polymyxin E as a template. Simple preparation conditions and simple operation. The raw materials are green and environment-friendly, and the prepared nano gold particles have good photo-thermal effect.
The invention is realized by the following steps:
a method for preparing gold nanoparticles by using polymyxin E as a template comprises the following steps:
s1, preparing polymyxin E solution:
preparing 0.1-3mg/mL polymyxin E solution according to the mass ratio of 1:0.1-3 of distilled water to polymyxin;
s2, preparing a chloroauric acid solution:
preparing a chloroauric acid solution of 0.01-10mg/mL according to the mass ratio of 1:0.01-10 of distilled water to the chloroauric acid;
s3, preparing an ascorbic acid solution:
preparing 0.1-10mg/mL ascorbic acid solution according to the mass ratio of 1:0.1-10 of distilled water to ascorbic acid;
s4, preparing nano gold particles:
uniformly mixing the polymyxin E solution and the chloroauric acid solution according to the molar ratio of the polymyxin E solution to the chloroauric acid solution of 1:0.1-10, putting the mixed solution into a metal bath, adding a phosphate buffer solution with the pH value of 7.4, adding distilled water, carrying out incubation, then adding an equal amount of ascorbic acid solution in portions, wherein the interval time of each time is 1-10min, and changing the solution from light yellow to blue-violet to obtain the gold nanoparticles.
Preferably, in step S4, the phosphate buffered saline solution is used in an amount of 0 to 1000. mu.L, and the distilled water is used in an amount of 0 to 1000. mu.L.
Preferably, the ascorbic acid solution is added in an amount of 5 to 50. mu.L per time in step S4.
Preferably, the incubation in the metal bath is carried out in particular by: the rotation speed is 200-1000 rpm, and the temperature is 10-40 ℃ for incubation for 0-120 min.
Compared with the prior art, the invention has the following beneficial effects:
1. the preparation method is simple, the reaction condition is mild, and the operation is simple and convenient.
2. The nano gold particles prepared by the method have the advantages of photo-thermal conversion performance, uniform particle size and good stability.
3. The template polymyxin E of the invention has high biocompatibility, and the product can be used in the biomedical field.
Drawings
Fig. 1 is a transmission diagram of the nanogold particles prepared in example 1.
FIG. 2 is a full wavelength scan of the gold nanoparticles prepared in example 1.
Fig. 3 is a graph showing a temperature increase of the gold nanoparticles prepared in example 1.
Fig. 4 is a temperature distribution diagram of the gold nanoparticles prepared in example 1 during 5 light irradiation cycles.
Fig. 5 is a transmission diagram of the nanogold particles prepared in example 2.
Fig. 6 is a graph showing temperature increase of the gold nanoparticles prepared in example 2 compared with distilled water.
Fig. 7 is a transmission diagram of the nanogold particles prepared in example 3.
Fig. 8 is a graph showing temperature increase of the gold nanoparticles prepared in example 3 compared with distilled water.
Detailed Description
Exemplary embodiments, features and performance aspects of the present invention will be described in detail below with reference to the accompanying drawings. In the drawings, like reference numbers can indicate functionally identical or similar elements. While the various aspects of the embodiments are presented in drawings, the drawings are not necessarily drawn to scale unless specifically indicated.
Firstly, preparing polymyxin E solution, reducing a part of gold particles by utilizing the reducibility of polymyxin E to be used as seed crystals, and then adding ascorbic acid to reduce to obtain the nano gold particles with the photo-thermal effect.
A method for preparing gold nanoparticles by using polymyxin E as a template comprises the following steps:
s1, preparing polymyxin E solution:
preparing 0.1-3mg/mL polymyxin E solution according to the mass ratio of 1:0.1-3 of distilled water to polymyxin;
s2, preparing a chloroauric acid solution:
preparing a chloroauric acid solution of 0.01-10mg/mL according to the mass ratio of 1:0.01-10 of distilled water to the chloroauric acid;
s3, preparing an ascorbic acid solution:
preparing 0.1-10mg/mL ascorbic acid solution according to the mass ratio of 1:0.1-10 of distilled water to ascorbic acid;
s4, preparing nano gold particles:
adding a polymyxin E solution into a 2mL centrifuge tube, uniformly mixing the polymyxin E solution and a chloroauric acid solution according to the molar ratio of the polymyxin E solution to the chloroauric acid solution of 1:0.1-10, putting the mixed solution into a metal bath, adding 0-1000 muL of phosphate buffer solution with the pH value of 7.4, adding 0-1000 muL of distilled water at the rotation speed of 200-1000 rpm, incubating at the temperature of 10-40 ℃ for 0-120min, then adding an equivalent amount of ascorbic acid solution in portions, wherein the interval time is 1-10min each time, and changing the solution from light yellow to blue-purple to obtain the gold nanoparticles.
Preferably, the ascorbic acid solution is added in an amount of 5 to 50. mu.L per time in step S4.
And (4) taking a small amount of the solution obtained in the step S4, carrying out ultrafiltration centrifugation, adding excessive sodium borohydride into the centrifuged solution, and indicating that the solution is colorless and transparent, thereby indicating that all the nano gold particles are generated.
Example 1
S1, preparing polymyxin E solution:
dissolving polymyxin E in distilled water to prepare a polymyxin E solution with the concentration of 3 mg/mL;
s2, preparing a chloroauric acid solution:
preparing a chloroauric acid solution with the concentration of 0.4mg/mL according to the mass ratio of 1:0.4 of distilled water to the chloroauric acid;
s3, preparing an ascorbic acid solution:
preparing an ascorbic acid solution with the concentration of 1mg/mL according to the mass ratio of 1:1 of distilled water to ascorbic acid;
s4, preparing nano gold particles:
adding 400 mu L of polymyxin E solution into a 2mL centrifuge tube, uniformly mixing the polymyxin E solution and chloroauric acid solution according to the molar ratio of the polymyxin E solution to the chloroauric acid solution of 1:1, putting the mixed solution into a metal bath, adding 400 mu L of phosphate buffer solution with the pH value of 7.4 and 400 mu L of distilled water, incubating at the rotation speed of 600rpm and the temperature of 20 ℃ for 15min, then adding equal amount of ascorbic acid solution in portions, wherein the interval time of each time is 5min, the adding amount of each time is 20 mu L, and the solution is changed from light yellow to blue-purple to obtain the gold nanoparticles.
The morphology of the prepared gold nanoparticles is characterized by using a transmission electron microscope, as shown in figure 1, the gold nanoparticles have different morphologies, no obvious spherical morphology exists, and the particle size is about 15-30 nm. The ultraviolet spectrophotometer is used to scan the nano gold particles at full wavelength, as shown in FIG. 2, the nano gold particles have very wide absorption band at 700-800 nm.
In order to study the photothermal effect of the gold nanoparticles, the following experiment was set up:
1) 1mL of the nano gold particle solution prepared in example 1 was taken and added into a 5mL cuvette; using 808nm infrared light emitter at 4W/cm2Under the condition (1), the nano gold particle solution in the cuvette is irradiated, the solution temperature is recorded every 30s, a dot line graph is drawn, and the photo-thermal conversion performance of the nano gold particles is analyzed. As can be seen from fig. 3, the nanogold particle solution rises from room temperature to 43.5 ℃ in 750s, which means that the nanogold particles can efficiently convert the optical energy of the NIR laser into thermal energy.
2) Using 808nm infrared light emitter at 4W/cm2Under the condition (1), the nano gold particle solution in the cuvette is irradiated for about 10min, the irradiation is stopped when the temperature rises to about 45 ℃, the nano gold particle solution is naturally cooled to the room temperature at the room temperature, the 808nm infrared light emitter is used again to irradiate the sample, the temperature is raised again, and the circulation stability performance is observed for five times in a circulating manner. The samples still have high photothermal conversion performance after 5 photothermal cycling tests as shown in fig. 4. In conclusion, the gold nanoparticles have high photothermal conversion performance and photothermal stability, so that the gold nanoparticles can be used as a PTT reagent in the photothermal treatment of cancers.
Example 2
S1, preparing polymyxin E solution:
dissolving polymyxin E in distilled water to prepare a polymyxin E solution with the concentration of 3 mg/mL;
s2, preparing a chloroauric acid solution:
preparing a chloroauric acid solution with the concentration of 0.4mg/mL according to the mass ratio of 1:0.4 of distilled water to the chloroauric acid;
s3, preparing an ascorbic acid solution:
preparing an ascorbic acid solution with the concentration of 1mg/mL according to the mass ratio of 1:1 of distilled water to ascorbic acid;
s4, preparing nano gold particles:
adding 400 mu L of polymyxin E solution into a 2mL centrifuge tube, uniformly mixing the polymyxin E solution and the chloroauric acid solution according to the molar ratio of 1:1, putting the mixed solution into a metal bath, incubating at 25 ℃ for 60min at the rotating speed of 500rpm, then adding equivalent ascorbic acid solution in portions, wherein the interval time of each time is 5min, the adding amount of each time is 20 mu L, and the solution is changed from light yellow to bluish purple to obtain the gold nanoparticles.
The morphology of the prepared gold nanoparticles was characterized by using a transmission electron microscope, as shown in FIG. 5, the gold nanoparticles had irregular protrusions with a particle size of about 100-200 nm.
In order to study the photothermal effect of the gold nanoparticles, the following experiment was set up:
taking 1mL of the nano gold particle solution prepared in the example 2, and adding the nano gold particle solution into a 5mL cuvette; irradiating the nano gold particle solution in the cuvette by using an 808nm infrared light emitter under the condition of 4W/cm2, recording the solution temperature every 30s, carrying out the same detection by using distilled water as a contrast, drawing a point line graph, and analyzing the photothermal conversion performance of the nano gold particles. As can be seen from fig. 6, the nanogold particle solution rose from 22.4 ℃ to 45.6 ℃ within 750 seconds, as a control group, the distilled water solution showed no significant response to the radiation, and the temperature rose from 22.5 ℃ to 27 ℃ within 750 seconds only. This indicates that the rough surface structure of the gold nanoparticles results in a very large local electric field enhancement, so that the gold nanoparticles can more effectively convert the optical energy of the NIR laser into thermal energy.
Example 3
S1, preparing polymyxin E solution:
dissolving polymyxin E in distilled water to prepare a polymyxin E solution with the concentration of 3 mg/mL;
s2, preparing a chloroauric acid solution:
preparing a chloroauric acid solution with the concentration of 0.4mg/mL according to the mass ratio of 1:0.4 of distilled water to the chloroauric acid;
s3, preparing an ascorbic acid solution:
preparing an ascorbic acid solution with the concentration of 1mg/mL according to the mass ratio of 1:1 of distilled water to ascorbic acid;
s4, preparing nano gold particles:
adding 400 mu L of polymyxin E solution into a 2mL centrifuge tube, uniformly mixing the polymyxin E solution and the chloroauric acid solution according to the molar ratio of the polymyxin E solution to the chloroauric acid solution of 1:1, putting the mixed solution into a metal bath, adding 800 mu L of phosphate buffer solution with the pH value of 7.4, incubating at 25 ℃ for 60min at the rotation speed of 200rpm, then adding equivalent ascorbic acid solution in portions, wherein the interval time of each time is 5min, the adding amount of each time is 20 mu L, and changing the solution from light yellow to blue-violet to obtain the gold nanoparticles.
The morphology of the prepared gold nanoparticles was characterized by using a transmission electron microscope, and as shown in fig. 7, the edges of the gold nanoparticles had significant irregular protrusions, and the particle size was about 40-100 nm.
In order to study the photothermal effect of the gold nanoparticles, the following experiment was set up:
1mL of the nano gold particle solution prepared in example 3 was taken and added into a 5mL cuvette; using 808nm infrared light emitter at 4W/cm2The same test was performed by irradiating the nanogold particle solution in the cuvette, recording the temperature of the solution every 30 seconds, comparing with distilled water, and plotting a dot-line graph to analyze the photothermal conversion performance of the nanogold particles, and as can be seen from fig. 8, the nanogold particle solution increased from 23.5 ℃ to 42.8 ℃ within 750 seconds, as a control group, the distilled water solution showed no significant response to the radiation, and the temperature increased from 22.5 ℃ to 27 ℃ within 750 seconds. This indicates that the protruding elliptical tips on the surface of the nanogold particles result in a larger local electric field enhancement, so that the nanogold particles can efficiently convert the optical energy of the NIR laser into thermal energy.
In conclusion, the invention has the following advantages:
the preparation method is simple, firstly, polymyxin E solution is prepared, part of gold particles are reduced by utilizing the reducibility of polymyxin E to serve as seed crystals, ascorbic acid is added for reduction, the nano gold particles with the photo-thermal effect are obtained, the reaction conditions are mild, the main reaction process is carried out by adopting metal bath, and the operation is simple and convenient.
The nano gold particles prepared by the method have the photo-thermal conversion performance, and the nano gold particles prepared each time are uniform in particle size and good in stability.
The template polymyxin E of the invention has high biocompatibility, and the product can be used in the biomedical field.
Finally, it should be noted that: the above-mentioned embodiments are only used for illustrating the technical solution of the present invention, and not for limiting the same; although the present invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some or all of the technical features may be equivalently replaced; and the modifications or the substitutions do not make the essence of the corresponding technical solutions depart from the scope of the technical solutions of the embodiments of the present invention.
Claims (2)
1. A method for preparing gold nanoparticles by using polymyxin E as a template is characterized by comprising the following steps: which comprises the following steps:
s1, preparing polymyxin E solution:
preparing 0.1-3mg/mL polymyxin E solution according to the mass ratio of 1:0.1-3 of distilled water to polymyxin;
s2, preparing a chloroauric acid solution:
preparing a chloroauric acid solution of 0.01-10mg/mL according to the mass ratio of 1:0.01-10 of distilled water to the chloroauric acid;
s3, preparing an ascorbic acid solution:
preparing 0.1-10mg/mL ascorbic acid solution according to the mass ratio of 1:0.1-10 of distilled water to ascorbic acid;
s4, preparing nano gold particles:
uniformly mixing a polymyxin E solution and a chloroauric acid solution according to the molar ratio of the polymyxin E solution to the chloroauric acid solution of 1:0.1-10, putting the mixed solution into a metal bath, adding a phosphate buffer solution with the pH =7.4, wherein the amount of the phosphate buffer solution is 400-1000 mu L, adding distilled water, the amount of the distilled water is 0-1000 mu L, carrying out incubation, then adding an ascorbic acid solution in an equivalent manner in portions, wherein the amount of the ascorbic acid solution is 5-50 mu L each time, and the interval time is 1-10min each time, so that the solution is changed from light yellow to blue-purple, and the nano gold particles are obtained, wherein irregular bulges are arranged on the edges of the nano gold particles.
2. The method for preparing gold nanoparticles using polymyxin E as a template according to claim 1, wherein: the method for carrying out the incubation in the metal bath comprises the following specific steps: the rotation speed is 200-1000 rpm, and the temperature is 10-40 ℃ for incubation for 15-120 min.
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