CN104923777A - High-salt-tolerance metal nanoparticle assembly and preparing method thereof - Google Patents

Abstract

The invention discloses a highly salt-tolerant metal nanoparticle assembly and a preparation method thereof. The method is as follows: take metal nanoparticles a and ssDNA, add them to the mixture a of coating agent, buffer solution and salt, separate and purify after mixed culture, and obtain product a; take metal nanoparticles b and sscDNA, add them to the coating agent, In the mixture b of buffer solution and salt, separate and purify after mixed culture to obtain product b; after mixing product a and product b, dissolve in the mixed solution of coating agent, buffer solution and salt, centrifuge and wash after stirring and culturing to obtain Metal nanoparticle assembly with high salt tolerance. The preparation method of the invention is simple and efficient, has strong controllability, and the obtained product has uniform appearance and good dispersibility. More meaningfully, the product prepared by the present invention has strong salt tolerance, and is expected to be used as a nanocarrier in biomedicine and other fields.

Description

A kind of highly salt-tolerant metal nanoparticle assembly and its preparation method

technical field

The invention belongs to the technical field of nanometer materials, and in particular relates to a metal nanoparticle assembly with high salt tolerance and a preparation method thereof.

Background technique

In recent years, metal nanoparticle assemblies have been widely used in nanodevices, nanosensors, and nanomedicine. Metal nanoassemblies not only have excellent optical, electrical, and catalytic properties of isolated nanoparticles, but also have some new properties due to assembly, such as surface-enhanced Raman characteristics, which greatly broaden the application of metal nanoparticles in the field of nanotechnology. within the scope of application. With the increasing application of metal nanoparticle assemblies, especially in biomedicine, the requirements for their salt tolerance are very high. Metal nanoparticles, including metal nanoparticle assemblies, are colloids and are particularly sensitive to electrolyte solutions such as saline solutions. Adding a small amount of salt can easily aggregate nanoparticles, and the concentration of salt in organisms is very high. Therefore, in order to apply metal nanoparticle assemblies in the field of biomedicine, it is necessary to prepare an assembly with high salt tolerance.

At present, domestic invention patents on metal nanoparticle assemblies with high salt tolerance are still blank. In fact, there have been very few technical inventions on isolated salt-tolerant metal nanoparticles. The Chinese invention patent "A Preparation Method of Highly Stable and Functionalized Gold Nanoparticles" with the application number 201010286955.X discloses a method for the synthesis of gold nanoparticles assisted by DNA. The gold nanoparticles prepared by this method have regular shape, uniform particle size, and salt resistance can be enhanced to 20 times, but this method is only for isolated gold nanoparticles.

Contents of the invention

In order to overcome the shortcomings and deficiencies of the prior art, the primary purpose of the present invention is to provide a method for preparing a metal nanoparticle assembly with high salt tolerance, which is simple and efficient, with uniform product appearance and good dispersion.

Another object of the present invention is to provide a metal nanoparticle assembly obtained by the above-mentioned preparation method, and to conduct a salt tolerance test of the assembly.

The object of the present invention is achieved through the following technical solutions:

A method for preparing a metal nanoparticle assembly with high salt tolerance, comprising the steps of:

(1) Take metal nanoparticles a and ssDNA, add them to the mixture a of coating agent, buffer solution and salt, separate and purify after mixing and culturing, and obtain product a;

(2) Take metal nanoparticles b and sscDNA, add them to the mixture b of coating agent, buffer solution and salt, separate and purify after mixing and culturing, and obtain product b;

(3) After mixing the product a and product b of steps (1) and (2), dissolve them in the mixed solution of coating agent, buffer solution and salt, centrifuge and wash after stirring to obtain highly salt-tolerant metal nanoparticle assembly body.

In the above method, the metal nanoparticle a described in step (1) is one or both of Au, Ag or Cu; the size of the metal nanoparticle is 1-100 nm; the metal nanoparticle a The molar ratio to ssDNA is 1:40~400.

In the above method, the encapsulating agent described in step (1) is sodium dodecyl sulfate (SDS); the mass ratio of the encapsulating agent to the mixed liquid a added with metal nanoparticles a and ssDNA is 0.01%~1% The buffer solution is phosphate buffer solution (PBS), pH=7.3, and the concentration of the buffer solution added in the mixed solution a of metal nanoparticles a and ssDNA is 10 ~ 100 mmol/L; the salt The concentration in the mixture a added with metal nanoparticles a and ssDNA gradually increased to 50-500 mmol/L within 6-24 h.

In the above method, the time for the mixed culture in step (1) is 6-24 h; the separation and purification method is centrifugation or dialysis.

In the above method, the metal nanoparticle b in step (2) is the same or different from the metal nanoparticle a in step (1), and the metal nanoparticle b is one or both of Au, Ag or Cu ; The size of the metal nanoparticles b is 1-100 nm; the molar ratio of the metal nanoparticles b to sscDNA is 1:0.2-20.

In the above method, the encapsulating agent described in step (2) is SDS, and the mass ratio of the encapsulating agent to the mixed solution b added with metal nanoparticles b and sscDNA is 0.01%~1%; the buffer solution is PBS , pH=7.3, the concentration of the buffer solution added in the mixed solution b of metal nanoparticles b and sscDNA is 10 ~ 100 mmol/L; the salt added in the mixed solution b of metal nanoparticles b and sscDNA The concentration in it is 50~500 mmol/L.

In the above method, the time for the mixed culture in step (2) is 6-24 h; the separation and purification method is centrifugation or dialysis.

In the above method, the molar ratio of product a and product b described in step (3) is 1:10~400; the encapsulating agent is SDS; the encapsulating agent is mixed with the mixed solution of product a and product b Mass ratio is 0.01%～1%; Described buffer solution is PBS, pH=7.3; The concentration of described buffer solution in the mixed solution that is mixed with product a and product b is 10～100 mmol/L; Described The concentration of salt in the mixed solution mixed with product a and product b is 50~500 mmol/L; the mixed culture time is 6~24 h; the centrifugal speed is 5000~20,000 rpm; the centrifugal time is 5 ~30 min, centrifuge 1~3 times.

The salt-tolerance experiment of the above-mentioned metal nanoparticle assembly includes the stability in normal saline and the stability in different concentrations of sodium chloride (NaCl) solutions.

Compared with the prior art, the present invention has the following advantages and effects:

The method is easy to operate, has strong controllability and high assembly efficiency, and the obtained product has a uniform structure and good dispersion, and the product has strong salt tolerance, and can withstand a salt concentration up to three times the concentration of normal saline. Therefore, the metal nanoassembly obtained by this preparation method has potential advantages in biological applications.

Description of drawings

Fig. 1 is the transmission electron micrograph (TEM) of the gold nanoparticle assembly obtained in embodiment 2;

Fig. 2 is the transmission electron micrograph (TEM) of the gold nanoparticle assembly obtained in Example 2 after being cultivated in physiological saline for 12 hours;

Fig. 3 is the result of the salt tolerance test of the gold nanoparticle assembly obtained in Example 2 in different concentrations of NaCl solutions, i.e. the color change diagram;

FIG. 4 is the results of the salt tolerance experiment of the gold nanoparticle assembly obtained in Example 2 in different concentrations of NaCl solutions, that is, the ultraviolet absorption spectrum.

Detailed ways

The technical invention will be further described below through the examples and accompanying drawings, but the embodiments of the present invention are not limited thereto.

The metal nanoparticles used in the method of the present invention can be purchased from the market, or can be prepared by using the existing technology.

The ssDNA and sscDNA used in the following examples were purchased from Shanghai Sangon Bioengineering Co., Ltd.

Its serial number is as follows:

ssDNA: 5'-HS-(CH ₂ ) ₆ ATC CTG ACA TCG GCA CGA GTA TTT CTA CCA TGT ATC-3'

sscDNA: 5'-HS-( _CH2 ) _6GAT ACA TGG TAG AAA TAC TCGTGC CGA TGT CAG GAT-3'.

Example 1

(1) Add 18 nm gold nanoparticles and ssDNA at a molar ratio of 1:100 to a final concentration of 0.2% (mass fraction) In the mixture a of SDS, 20 mM PBS (pH = 7.3) and 300 mM NaCl, after 20 h of mixed culture, centrifuge at 15000 rpm for 3 times, 15 min each time. Remove the lower layer of sediment;

(2) 4 nm gold nanoparticles and sscDNA were added to the mixture b with a final concentration of 0.2% (mass fraction) SDS, 20 mM PBS (pH= 7.3) and 100 mM NaCl at a molar ratio of 1:10, After 6 hours of mixed culture, dialyze with dialysis bag for 20 hours to purify;

(3) After mixing the products of steps (1) and (2) according to the molar ratio of 1:20, add to the final concentration of 0.2% (mass fraction) SDS, 20 mM PBS (pH = 7.3) and 150 mM NaCl In the mixed solution, cultured with slow stirring for 20 h, and then centrifuged at 15,000 rpm for 3 times, each time for 15 min, to obtain gold nanoparticle assemblies.

Example 2

(1) 18 nm gold nanoparticles and ssDNA were added to the mixture a with a final concentration of 0.2% (mass fraction) SDS, 20 mM PBS (pH= 7.3) and 300 mM NaCl at a molar ratio of 1:100, After 20 h of mixed culture, centrifuge at 15000 rpm for 3 times, 15 min each time. Remove the lower layer of sediment;

(2) Add 4 nm gold nanoparticles and sscDNA at a molar ratio of 1:10 to the mixture b with a final concentration of 0.2% SDS, 20 mM PBS (pH= 7.3) and 100 mM NaCl, and incubate for 6 h Afterwards, it was dialyzed with a dialysis bag for 20 h to purify;

(3) After mixing the products of steps (1) and (2) according to the molar ratio of 1:40, they were added to the mixed solution with a final concentration of 0.2% SDS, 20 mM PBS (pH= 7.3) and 150 mM NaCl, Slowly agitate and cultivate for 20 h, and then centrifuge at 15,000 rpm for 3 times, each time for 15 min, to obtain gold nanoparticle assemblies.

The obtained gold nanoparticle assembly was characterized by TEM, and the results are shown in Fig. 1 . It can be seen from Figure 1 that 4 nm gold nanoparticles are successfully connected to the surface of 18 nm gold nanoparticles, forming a "core-satellite" assembly. The average number of satellites is 9, and the dispersion of the assembly is good. , with a relatively uniform size.

Example 3

(1) 18 nm gold nanoparticles and ssDNA were added to the mixture a with a final concentration of 0.2% (mass fraction) SDS, 20 mM PBS (pH= 7.3) and 300 mM NaCl at a molar ratio of 1:100, After 20 h of mixed culture, centrifuge at 15000 rpm for 3 times, 15 min each time. Remove the lower layer of sediment;

(2) Add 4 nm gold nanoparticles and sscDNA into the mixture b of 0.2% SDS, 20 mM PBS (pH= 7.3) and 100 mM NaCl according to the molar ratio of 1:10, mix and incubate for 6 h, then use dialysis The bag was dialyzed for 20 h to purify;

(3) After mixing the products of steps (1) and (2) according to the molar ratio of 1:80, they were added to the mixture with a final concentration of 0.2% SDS, 20 mM PBS (pH= 7.3) and 150 mM NaCl, Slowly agitate and cultivate for 20 h, and then centrifuge at 15,000 rpm for 3 times, each time for 15 min, to obtain gold nanoparticle assemblies.

Example 4

(1) 18 nm gold nanoparticles and ssDNA were added to the mixture a with a final concentration of 0.2% (mass fraction) SDS, 20 mM PBS (pH= 7.3) and 300 mM NaCl at a molar ratio of 1:100, After 20 h of mixed culture, centrifuge at 15000 rpm for 3 times, 15 min each time. Remove the lower layer of sediment;

(2) Add 4 nm gold nanoparticles and sscDNA into the mixture b of 0.2% SDS, 20 mM PBS (pH= 7.3) and 100 mM NaCl according to the molar ratio of 1:10, mix and incubate for 6 h, then use dialysis The bag was dialyzed for 20 h to purify;

(3) After mixing the products of steps (1) and (2) according to the molar ratio of 1:160, they were added to a mixed solution with a final concentration of 0.2% SDS, 20 mM PBS (pH= 7.3) and 150 mM NaCl, Slowly agitate and cultivate for 20 h, and then centrifuge at 15,000 rpm for 3 times, each time for 15 min, to obtain gold nanoparticle assemblies.

Salt tolerance test

(1) After the gold nanoparticle assembly of Example 2 was cultured in physiological saline for 12 h, it was characterized by a transmission electron microscope (TEM). Its TEM image is shown in Figure 2.

It can be seen from Figure 2 that the gold nanoparticle assembly prepared by the method of the present invention still maintains a complete shape after being cultured in physiological saline for up to 12 h, indicating that it has good stability in a salt concentration of 0.9%.

(2) Take the gold nanoparticle assembly of Example 2 and carry out the salt tolerance test in NaCl solutions of 0.9%, 1.8%, 3.0%, 3.3% and 4.5% (mass fraction) respectively, observe the initial color change of the solution and Perform UV characterization. The results are shown in Figure 3 and Figure 4.

The stability of gold nanoparticle assemblies is easily affected by salt concentration. The normal homogeneously dispersed aqueous solution of the assembly is generally ruby red, and may agglomerate when it is in a high-concentration salt solution. This is reflected in the appearance as a color change of the solution (usually to purple) and in the UV spectrum as a red shift of the maximum absorption peak. Therefore, its stability can be verified by solution color and ultraviolet absorption spectrum. The color change in Figure 3 shows: in 0.9% NaCl solution (equivalent to the concentration of physiological saline), the gold nanoparticle assembly obtained in Example 2 did not agglomerate, and the color was the initial ruby red, which is consistent with the salt tolerance The results of experiment (1) are consistent with each other; when the salt concentration is increased by about two times (3.0 %), there is still no agglomeration phenomenon, and the solution is still ruby red, indicating that the assembly still has good stability at this time. It can be seen from Figure 4 that when the salt concentration is as high as 3.0%, the maximum absorption peak of the gold nanoparticle assembly still has no obvious red shift, which is also consistent with the color change results in Figure 3. This shows that the product has high salt tolerance and is expected to be applied in a physiological environment.

The above-mentioned embodiment is a preferred embodiment of the present invention, but the embodiment of the present invention is not limited by the above-mentioned embodiment, and any other changes, modifications, substitutions, combinations, Simplifications should be equivalent replacement methods, and all are included in the protection scope of the present invention.

Claims (9)

1. A preparation method of a highly salt-tolerant metal nanoparticle assembly, characterized in that, comprising the steps: (1) Take metal nanoparticles a and ssDNA, add them to the mixture a of coating agent, buffer solution and salt, separate and purify after mixing and culturing, and obtain product a; (2) Take metal nanoparticles b and sscDNA, add them to the mixture b of coating agent, buffer solution and salt, separate and purify after mixing and culturing, and obtain product b; (3) After mixing the product a and product b of steps (1) and (2), dissolve them in the mixed solution of coating agent, buffer solution and salt, centrifuge and wash after stirring to obtain highly salt-tolerant metal nanoparticle assembly body. 2. The method for preparing a metal nanoparticle assembly with high salt tolerance according to claim 1, wherein the metal nanoparticle a in step (1) is one of Au, Ag or Cu or Two kinds; the size of the metal nanoparticles is 1-100 nm; the molar ratio of the metal nanoparticles a to ssDNA is 1:40-400. 3. The preparation method of a metal nanoparticle assembly with high salt tolerance according to claim 1, characterized in that: the encapsulating agent in step (1) is sodium dodecyl sulfate (SDS); the The mass ratio of the encapsulating agent to the mixture a of metal nanoparticles a and ssDNA is 0.01%~1%; the buffer solution is phosphate buffer solution (PBS), pH=7.3, and the buffer solution is added to the metal The concentration in the mixture a of nanoparticles a and ssDNA is 10-100 mmol/L; the concentration of the salt in the mixture a added with metal nanoparticles a and ssDNA increases to the final concentration within 6-24 h. 50~500mmol/L. 4. The method for preparing a highly salt-tolerant metal nanoparticle assembly according to claim 1, characterized in that: the time for the mixed culture in step (1) is 6 to 24 h; the separation and purification The method is centrifugation or dialysis. 5. The preparation method of a metal nanoparticle assembly with high salt tolerance according to claim 1, characterized in that: the metal nanoparticles b described in step (2) and the metal nanoparticles in step (1) a is the same or different, the metal nanoparticles b are one or both of Au, Ag or Cu; the size of the metal nanoparticles b is 1-100 nm; the metal nanoparticles b and sscDNA The molar ratio is 1:0.2~20. 6. The preparation method of a metal nanoparticle assembly with high salt tolerance according to claim 1, characterized in that: the encapsulating agent in step (2) is SDS, and the encapsulating agent and metal nanoparticles are added The mass ratio of the mixture b of b and sscDNA is 0.01%~1%; the buffer solution is PBS, pH=7.3, and the concentration of the buffer solution in the mixture b of metal nanoparticles b and sscDNA is added 10-100 mmol/L; the concentration of the salt in the mixture b added with metal nanoparticles b and sscDNA is 50-500 mmol/L. 7. The method for preparing a highly salt-tolerant metal nanoparticle assembly according to claim 1, characterized in that: the time for the mixed culture in step (2) is 6 to 24 h; the separation and purification The method is centrifugation or dialysis. 8. The method for preparing a metal nanoparticle assembly with high salt tolerance according to claim 1, characterized in that: the molar ratio of product a and product b in step (3) is 1:10~400; Described wrapping agent is SDS; The mass ratio of described wrapping agent and the mixed solution that is mixed with product a and product b is 0.01%～1%; Described buffer solution is PBS, pH=7.3; Described buffer solution is in The concentration in the mixed solution mixed with product a and product b is 10-100 mmol/L; the concentration of the salt in the mixed solution mixed with product a and product b is 50-500 mmol/L; the The mixing culture time is 6-24 h; the centrifugation speed is 5000-20,000 rpm; the centrifugation time is 5-30 min, centrifuged 1-3 times. 9. According to the preparation method described in any one of claims 1 to 8, the metal nanoparticle assembly with high salt tolerance is prepared.