A method for preparing hollow mesoporous nanoparticles composited with protein and silicon dioxide

Abstract

The present invention relates to a method for preparing hollow mesoporous nanoparticles composited with protein and silicon dioxide, comprising the steps of stirring ethanol, ammonia water, bovine serum albumin solution and aminopropyltriethoxysilane at a temperature of 30-50°C for 5-7h, then adding tetraethyl orthosilicate, stirring and reacting at a temperature of 30-50°C for 5-7h, centrifuging, washing and drying after the reaction is completed, and obtaining the prepared hollow mesoporous nanoparticles. The preparation method is green and friendly, easy to operate, and has a high yield. The prepared particles have a small size (<50 nm), a clear hollow morphology, and the protein and silicon dioxide can be effectively composited.

Description

Technical Field

[0001] The invention belongs to the technical field of nanomaterials, and specifically relates to a method for preparing small-sized (<50 nm) hollow mesoporous nanoparticles composited with protein and silicon dioxide. Background Art

[0002] As we all know, nanoparticles of proteins and silica composites of different sizes and morphologies have attracted much attention in the biomedical field for many years due to their good biocompatibility and biodegradability, as well as their efficient adsorption of various types of drug molecules. In particular, silica nanoparticles with hollow mesoporous morphology are more significant in drug delivery and clinical applications due to their high specific surface area, large cavity volume and mesoporous structure.

[0003] In addition, many in vitro and in vivo bioactivity experiments have found that the smaller the size of the hollow mesoporous particles (<50 nm), the more conducive it is to maintaining the circulation and biodistribution of drug molecules in the blood after loading drug molecules, and the more conducive it is to capturing drug molecules in the tumor microenvironment ( small 2009, 5 , 1408–1413). Especially in clinical applications, the smaller the size of the nanoparticles, the more beneficial they are to kidney metabolism and the more practical their application value ( Nat. Nanotechnol. 2007, 25 , 1165−1170).

[0004] However, in the preparation process of this type of material, it was found that without adding any surfactants or cross-linking agents, it is generally difficult to use protein molecules as effective templates (structure-directing agents) due to the instability of their secondary or tertiary structures, and it is also difficult to further effectively synthesize silica materials with uniform particle size and stable hollow morphology. In short, it is always a challenge to effectively synthesize small-sized (<50 nm) hollow mesoporous nanoparticles composited with proteins and silica in a green and friendly manner! Summary of the invention

[0005] The purpose of the present invention is to overcome the technical problems and defects of the prior art and to provide a method for preparing a novel hollow mesoporous nanoparticle composited with protein and silica. The preparation method is green and friendly, easy to operate, and has a high yield. The prepared particles have a small size (<50 nm) and a clear hollow morphology, and the protein and silica can be effectively composited.

[0006] To achieve the above object, the present invention adopts the following technical solution:

[0007] A method for preparing hollow mesoporous nanoparticles composited with protein and silicon dioxide comprises the following steps:

[0008] Stir ethanol, ammonia water, bovine serum albumin solution and aminopropyltriethoxysilane at 30-50°C for 5-7h, then add tetraethyl orthosilicate, continue stirring and reacting at 30-50°C for 5-7h. After the reaction is completed, centrifuge, wash and dry to obtain the product.

[0009] In the above-mentioned preparation method, in the reaction system, the final concentration of the ammonia water is 0.40-0.60 mol / L.

[0010] Specifically, in the reaction system, the final concentration of bovine serum albumin (BSA) is 0.008-0.01 mmol / L.

[0011] Furthermore, in the reaction system, the final concentration of aminopropyltriethoxysilane (APTES) is 0.01-0.02 mol / L.

[0012] Specifically, in the reaction system, the final concentration of tetraethyl orthosilicate (TEOS) is 0.01-0.02 mol / L.

[0013] The above-mentioned preparation method, drying specifically comprises: placing at 50-70°C for 6-10 hours.

[0014] The invention also provides hollow mesoporous nanoparticles composited with protein and silicon dioxide prepared by the preparation method.

[0015] The present invention also provides a method for preparing a preferred hollow mesoporous nanoparticle composite of protein and silicon dioxide, which is as follows:

[0016] In ethanol, after controlling the reaction temperature to be constant at 30-50°C, ammonia water, bovine serum albumin (BSA) solution and aminopropyltriethoxysilane (APTES, analytical grade) are added in sequence under stirring conditions. After stirring and reacting at 30-50°C for 5-7 hours, tetraethyl orthosilicate (TEOS, analytical grade) is added and stirring and reacting is continued at 30-50°C for 5-7 hours. After the reaction is completed, centrifuge, wash with water, and dry at 50-70°C for 6-10 hours to obtain pure, small-sized (<50 nm), hollow mesoporous nanoparticles with effective composite of protein and silica.

[0017] The basic principle and beneficial effects of the method of the present invention are as follows:

[0018] The method of the present invention adopts a sol-gel / emulsion method. Bovine serum albumin and aminopropyltriethoxysilane may first form a relatively soft spherical aggregate as a soft template in an alcohol / water mixed solution, which can play a good hollow template role when the silica shell is subsequently coated. Among them, the BSA molecules are mainly distributed inside the spherical aggregate, and the APTES molecules are distributed on the surface of the aggregate. The amino group of APTES and the carboxyl group of the protein molecule can be connected to each other through electrostatic or hydrogen bonding, and the siloxane group of the APTES molecule is exposed on the outermost layer of the soft template. When the subsequent TEOS molecules are added, under the catalytic action of a weak alkaline reagent (ammonia water), TEOS and APTES molecules undergo a slow co-hydrolysis / condensation process, which will continuously promote the nucleation and growth of silica on the outer surface of the aggregate. Since there are still a large number of free BSA molecules in the solution system at this time, under their synergistic action, a mesoporous structure may be formed when the silica hollow shell is formed. Therefore, the hollow mesoporous nanoparticles of protein and silicon dioxide composite prepared by the method of the present invention are simpler, greener and more efficient. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 is a transmission electron microscope (TEM) image of hollow mesoporous nanoparticles composited with protein and silica obtained according to Example 1 of the present invention;

[0020] Figure 2 This is an infrared spectrum image of hollow mesoporous nanoparticles composited with protein and silicon dioxide obtained according to Example 1 of the present invention;

[0021] Figure 3 The circular dichroism spectrum image of the protein and silicon dioxide composite nanoparticles obtained according to Example 1 of the present invention;

[0022] Figure 4 The nitrogen adsorption-desorption curve of the hollow mesoporous nanoparticles composited with protein and silicon dioxide obtained according to Example 1 of the present invention;

[0023] Figure 5 This is the mesopore distribution curve of the hollow mesoporous nanoparticles composited with protein and silica obtained according to Example 1 of the present invention. DETAILED DESCRIPTION

[0024] The following are basic conditions used in the embodiments of the present invention, but the scope of implementation of the present invention is not limited to these conditions or these embodiments.

[0025] The raw materials used in the following examples are all common commercially available products that can be directly purchased, and some parameters are listed below:

[0026] Ambient temperature 25 ℃, 1 atmosphere pressure;

[0027] Ethanol (super pure), density 0.79 g / mL, molecular weight 46.00 g / mol;

[0028] Water, density 1.00 g / mL, molecular weight 18.00 g / mol;

[0029] Ammonia water, density 0.90~0.91g / mL, mass fraction 25%, molar concentration 13.30 mol / L, molecular weight 17.00 g / mol;

[0030] Bovine serum albumin (BSA), molecular weight 6,6446 Da (g / mol);

[0031] Aminopropyltriethoxysilane (APTES, analytical grade), density 0.946 g / mL, molecular weight 221.37 g / mol;

[0032] Tetraethyl orthosilicate (TEOS, analytical grade), density 0.931 g / mL, molecular weight 208.33 g / mol.

[0033] Embodiment 1:

[0034] A method for preparing hollow mesoporous nanoparticles composited with protein and silicon dioxide is as follows:

[0035] In 50 mL of ethanol, after the reaction temperature is constant at 40°C, 2 mL of 25% ammonia water, 1 mL of 0.5 mM bovine serum albumin (BSA) solution and 0.2 mL of APTES analytical grade are added in sequence under stirring conditions. After stirring and reacting at 40°C for 6 hours (within this time range, the synthesis results are similar), 0.2 mL of TEOS analytical grade is added and the reaction is continued at a constant temperature of 40°C for 6 hours. In the reaction system, the final concentration of the ammonia water is 0.50 mol / L; the final concentration of the bovine serum albumin (BSA) is 9.36×10 -3 mmol / L; the final concentration of aminopropyltriethoxysilane (APTES) is 0.016 mol / L; the final concentration of tetraethyl orthosilicate (TEOS) is about 0.017 mol / L. After the reaction is completed, the product is first centrifuged and washed twice with water, and then placed at 60°C for 6 hours for drying, and finally pure, small-sized (<50nm), protein-silica composite hollow mesoporous nanoparticles are obtained, with a yield of about 92%.

[0036] The TEM photo of the particle product prepared in Example 1 is as follows: Figure 1 As shown, from Figure 1 It can be seen that the particle size is relatively uniform and has a clear hollow morphology.

[0037] The infrared spectrum of the particle product prepared in Example 1 is as follows: Figure 2 As shown, from Figure 2 It can be seen that the particles have a clear -Si-O-Si- skeleton vibration peak (1080 cm -1 ) and the characteristic peak band of protein (1663 cm -1 and 1547 cm -1 ).

[0038] The circular dichroism spectrum of the particle product prepared in Example 1 is as follows: Figure 3 As shown, from Figure 3 It can also be clearly seen that the protein in the particles has clear secondary structure characteristic peaks (a-helix 208 nm; β-fold 227 nm). The infrared spectrum data and circular dichroism spectrum data can clearly show that bovine serum albumin is effectively complexed with silica particles.

[0039] The nitrogen adsorption-desorption curve of the particle product prepared in Example 1 is as follows: Figure 4 As shown, from Figure 4 It can be seen that the particle is at 0.4 P / P o A very obvious N2 adsorption-desorption hysteresis loop is shown near the surface, indicating that it has a clear hollow mesoporous structure with a pore volume of 0.58 cm 3 / g, BET specific surface area is 143.40 m 2 / g.

[0040] The mesopore distribution curve of the particle product prepared in Example 1 is as follows: Figure 5 As shown, Figure 5 The data in the figure show that the particles have a relatively narrow mesopore distribution, with an average pore size of about 3.6 nm.

[0041] In summary, the particles prepared by the method of the present invention have a small size (<50 nm), a clear hollow morphology, and the protein and silica can be effectively compounded; and the preparation method is green and friendly, easy to operate, and has a high yield, and is of great value for promotion and application.

Claims

1. A method for preparing hollow mesoporous nanoparticles composited with protein and silicon dioxide, characterized in that: Stir ethanol, ammonia water, bovine serum albumin solution and aminopropyltriethoxysilane at 30-50°C for 5-7h, then add tetraethyl orthosilicate, continue stirring for 5-7h at 30-50°C, after the reaction, centrifuge, wash and dry to obtain; In the reaction system, the final concentration of the ammonia water is 0.40-0.60 mol / L; The final concentration of the bovine serum albumin is 0.008-0.01 mmol / L; The final concentration of the aminopropyltriethoxysilane is 0.01-0.02 mol / L; The final concentration of tetraethyl orthosilicate is 0.01-0.02 mol / L.

2. The method for preparing hollow mesoporous nanoparticles of protein and silicon dioxide composite according to claim 1, characterized in that: The specific drying process is as follows: placing at 50-70°C for 6-10 hours.

3. Hollow mesoporous nanoparticles of protein and silicon dioxide composite prepared by the preparation method according to any one of claims 1 to 2.

Images