CN110169958A - A kind of preparation method of the multi-functional mesoporous silicon oxide composite nano materials based on Fluorescent silicon nanoparticle - Google Patents

Abstract

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the steps are as follows: using N-(2-aminoethyl)-3-aminopropyltrimethoxysilane as a silicon source, Co-coating of silicon nanoparticles prepared by one-pot hydrothermal method and photosensitizer 5,10,15,20-tetrakis(N-methyl-4-pyridine)porphine p-toluenesulfonate by oil-water two-phase separation method Mesoporous silica forms MSN@SiNPs@TMPyP; and it is further modified with folic acid to obtain the final product MSN@SiNPs@TMPyP‑FA. The invention innovatively combines SiNPs with two-photon fluorescence, photosensitizer and mesoporous silicon, and successfully realizes the preparation of multifunctional nanocomposite particles with two-photon fluorescence imaging-guided drug chemotherapy/photodynamic synergistic therapy.

Description

A Multifunctional Mesoporous Silica Composite Nanomaterial Based on Fluorescent Silicon Nanoparticles Material preparation method

technical field

The invention belongs to the field of nanomaterial preparation, in particular to a method for preparing a multifunctional mesoporous silicon dioxide composite nanomaterial based on fluorescent silicon nanoparticles.

Background technique

Silicon nanoparticles (SiNPs), as a novel fluorescent nanomaterial, have the advantages of low preparation cost, strong fluorescence, strong resistance to photobleaching, low toxicity, and good biocompatibility. See: Zhong, Y.L.J. Am. Chem. Soc. 2013, 135, 8350-8356. In addition, recent studies have shown that silicon nanoparticles have a two-photon absorption effect, and can perform two-photon fluorescence imaging under near-infrared wavelength laser excitation, thereby enhancing tissue penetration while effectively avoiding biological autofluorescence interference. See: Ye, H.L. Anal Chem. 2016, 88(23), 11631-11638. Therefore, SiNPs can be used as a promising fluorescent probe for biodiagnostic therapy.

At present, the research on SiNPs-based nanocomposites for the integration of diagnosis and treatment has developed rapidly. For example, He loaded anticancer drug doxorubicin (DOX) through SiNPs to achieve cancer cell drug chemotherapy. See: Ji, X. Adv. Mater. 2015, 27, 1029-1034. In addition, Chen synthesized a silicon dot and photosensitizer (Ps) nanocomposite successfully applied in photodynamic therapy (PDT). See: Wang, R.G.J. Mater. Chem. B, 2018, 6, 4592-4601. But these are usually limited to a single treatment modality. It remains a challenge to prepare Sidot-based multifunctional nanocomposites for multimodal synergistic, more effective therapeutic applications.

Mesoporous silica nanoparticles (MSNs) have become a potential multifunctional nanocarrier due to their high specific surface area, tunable pore size, easy surface modification and good biocompatibility. See: Wu, S.H.Chem.Soc.Rev., 2013, 42, 3862-3875. At present, fluorescent mesoporous silica nanocomposites based on up-conversion nanoparticles, carbon nanomaterials, and long-lasting materials have been reported. These materials can not only perform effective fluorescent bioimaging, but also realize multiple modal treatments. , see: Han, R. Chem.-Asian J., 2017, 12, 2197-2201.; Liu, Y, RSC Adv., 2017, 7, 31133-31141.; Wang, T. ACS Appl. Mater. Interfaces 2011,3,2479–2486., but the preparation of multifunctional mesoporous silica composite nanomaterials based on fluorescent silicon nanoparticles has not been reported yet.

Contents of the invention

The purpose of the present invention is to overcome the above-mentioned problems in the prior art and provide a method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles.

The method of the present invention first uses a one-pot microwave method to synthesize silicon nanoparticles (SiNPs) with two-photon fluorescence, and then the obtained SiNPs and photosensitizer (Ps) are jointly coated with mesoporous silicon by an oil-water two-phase layering method to form mesoporous pores. Silica composite nanoparticles, and further modify folic acid to have targeting ability. Utilizing the mesoporous structure can be used to load the anticancer drug doxorubicin (DOX), the intercalated Ps can generate singlet oxygen for photodynamic therapy. Finally, multifunctional nanocomposite particles capable of two-photon fluorescence targeted imaging-guided drug chemotherapy/photodynamic synergistic therapy are formed.

Technical scheme of the present invention:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, comprising the steps of:

1) Add sodium citrate as a reducing agent to ultrapure water, pass through nitrogen and stir for 10 minutes, then add silicon source N-(2-aminoethyl)-3-aminopropyltrimethoxysilane (DAMO); keep stirring After 15 minutes, move it to a polytetrafluoroethylene reactor, react at 200°C for 1-5 hours, then transfer the obtained yellow solution to a dialysis bag for 24 hours, and change the water every 8 hours to obtain an aqueous solution of SiNPs;

The mass ratio of the ultrapure water, sodium citrate and silicon source N-(2-aminoethyl)-3-aminopropyltrimethoxysilane is 1:0.025-0.075:0.25.

2) Take cetrimonium chloride solution (CTAC), Si NPs aqueous solution obtained in step 1), photosensitizer 5,10,15,20-tetrakis(N-methyl-4-pyridine)porphine p-toluenesulfonate (TMPyP) and triethanolamine (TEA) were dissolved in ultrapure water, and stirred continuously at 60°C for 1 hour at a stirring speed of 150rmp, then the octadecene solution dissolved in ethyl silicate (TEOS) was slowly added, and continued After reacting for 12 hours, remove the upper layer of oil, and collect the product by centrifugation after washing; then the product is dispersed in an ethanol solution containing ammonium nitrate, refluxed at 66° C. for 6 hours, and obtained by centrifugal washing to obtain mesoporous silica composite nanoparticles ( MSN@SiNPs@TMPyP);

Wherein the concentration of the cetrimonium chloride solution is 25wt%; the volume ratio of the ethanol, ultrapure water, CTAC solution, Si NPs aqueous solution, octadecene and TEOS is 1:0.38:0.24:0.02-0.06:0.16: 0.04; the mass ratio of TEOS, TEA, TMPyP and ammonium nitrate is 1:0.048:0.0005-0.006:0.016.

3) Dissolve folic acid (FA), 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC), N-hydroxysuccinimide (NHS) in dimethyl In sulfoxide, keep stirring for 30min, then add 3-aminopropyltriethoxysilane (APTES), stir overnight to obtain FA-APTES; take FA-APTES, MSN@SiNPs@TMPyP obtained in step 2) and add to methanol solution, stirred at room temperature for 24 h, and centrifuged to obtain folic acid-modified multifunctional mesoporous silica composite nanoparticles (MSN@SiNPs@TMPyP-FA).

The mass ratio of APTES, FA, EDC, NHS and MSN@SiNPs@TMPyP is 1:0.17-0.51:0.084-0.25:0.13-0.38:0.21; the volume ratio of methanol, APTES, DMSO and FA-APTES: 1 :0.05:0.4:0.02.

The specific preparation steps of the multifunctional mesoporous silica composite nanomaterial are as follows:

1) Add 0.2-0.6g of sodium citrate as a reducing agent to 8mL of ultrapure water, blow in nitrogen and stir for 10min, then add 2mL of silicon source N-(2-aminoethyl)-3-aminopropyltrimethoxysilane (DAMO); continue to stir for 15 minutes, then transfer to a polytetrafluoroethylene reactor, react at 200 ° C for 1-5 hours, and then transfer the obtained yellow solution to a dialysis bag, the dialysis time is 24 hours, and change the water every 8 hours to obtain SiNPs aqueous solution;

2) Take 12mL cetrimonium chloride solution (CTAC), 1-3mL SiNPs aqueous solution obtained in step 1), 1-10mg photosensitizer 5,10,15,20-tetrakis(N-methyl-4-pyridine) porphyrin Phenyl p-toluenesulfonate (TMPyP) and 90mg triethanolamine (TEA) were dissolved in 19mL ultrapure water, stirred continuously at 60°C for 1h at a stirring speed of 150rmp, and then dissolved in 2mL ethyl silicate (TEOS) Add 10 mL of octadecene mixed solution slowly, continue to react for 12 hours, remove the upper layer of oil, and collect the product by centrifugation after washing; then disperse the product in ethanol solution (50 mL) containing 30 mg of ammonium nitrate, and reflux at 66 ° C for 6 Hours, the mesoporous silica composite nanoparticles (MSN@SiNPs@TMPyP) were obtained by centrifugal washing;

3) Mix 40-120mg folic acid (FA), 20-60mg 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC), 30-90mg N-hydroxysuccinimide (NHS) was dissolved in 2mL dimethyl sulfoxide, stirred continuously for 30min, then added 250μL 3-aminopropyltriethoxysilane (APTES), stirred overnight to obtain FA-APTES; take 100μL FA-APTES, step 2) The obtained 50 mg of MSN@SiNPs@TMPyP was added to 5 mL of methanol solution, stirred and reacted at room temperature for 24 h, and multifunctional mesoporous silica composite nanoparticles modified with folic acid (MSN@SiNPs@TMPyP-FA) were obtained by centrifugal washing.

Advantages and beneficial effects of the present invention:

1) By co-packing mesoporous silicon with silicon nanoparticles and photosensitizers, a multifunctional mesoporous silica composite nanoparticle based on silicon nanoparticles was synthesized for the first time, which can realize the dual mode of drug-loaded chemotherapy/photodynamic therapy treatment, enhance the treatment effect.

2) Due to the introduction of silicon nanoparticles and folic acid modification, the material can be used for two-photon targeted imaging under near-infrared wavelength laser irradiation, which avoids autofluorescence and improves imaging accuracy.

Description of drawings

Figure 1 is the TEM images of (a) SiNPs and (b) MSN@SiNPs@TMPyP-FA.

Figure 2 shows the release efficiency of MSN@SiNPs@TMPyP-FA loaded Dox at different pH.

Figure 3 is a comparison of two-photon targeted imaging of different cells by MSN@SiNPs@TMPyP-FA.

Figure 4 shows the therapeutic effect of MSN@SiNPs@TMPyP-FA on MCF-7 cells.

Detailed ways

Example 1:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, in which the two-photon fluorescent silicon nanoparticles synthesized by the one-pot hydrothermal method and the photosensitizer are co-incorporated by the oil-two-phase layering method Porous silicon forms a nanocomposite, and further modifies folic acid to make it have a targeting ability, and finally forms a multifunctional nanocomposite particle that can perform drug chemotherapy/photodynamic synergistic therapy guided by two-photon fluorescence targeting imaging.

Including the following steps:

1) Add 0.4g of sodium citrate as a reducing agent to 8mL of ultrapure water, and then add 2mL of silicon source N-(2-aminoethyl)-3-aminopropyltrimethoxysilane (DAMO ); after continuing to stir for 15 minutes, move it to a polytetrafluoroethylene reactor, react at 200 ° C for 3 hours, and then transfer the obtained yellow solution to a dialysis bag for 24 hours, and change the water every 8 hours to obtain an aqueous solution of SiNPs;

2) Take 12 mL of cetrimonium chloride solution (CTAC), 1.5 mL of Si NPs aqueous solution obtained in step 1), 5 mg of photosensitizer 5,10,15,20-tetrakis(N-methyl-4-pyridine) porphine Tosylate (TMPyP) and 90 mg triethanolamine (TEA) were dissolved in 19 mL of ultrapure water, stirred continuously at 60 ° C for 1 h at a stirring speed of 150 rpm, and then 2 mL of ethyl silicate (TEOS) was dissolved in ten Slowly add 10 mL of octane mixed solution, continue to react for 12 hours, remove the upper layer of oil, and collect the product by centrifugation after washing; then disperse the product in ethanol solution (50 mL) containing 30 mg of ammonium nitrate, and reflux at 66 ° C for 6 hours. Mesoporous silica composite nanoparticles (MSN@SiNPs@TMPyP) were obtained by centrifugal washing;

3) Dissolve 80mg of folic acid (FA), 40mg of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC), 60mg of N-hydroxysuccinimide (NHS) in 2mL In dimethyl sulfoxide, keep stirring for 30 min, then add 250 μL 3-aminopropyltriethoxysilane (APTES), and stir overnight to obtain FA-APTES; take 100 μL FA-APTES, step 2) to obtain 50 mg MSN@SiNPs The multifunctional mesoporous silica composite nanoparticles modified with folic acid (MSN@SiNPs@TMPyP-FA) were obtained by centrifugation and washing in 5 mL of methanol solution added to @TMPyP under stirring at room temperature for 24 h.

Figure 1 (a) TEM images of SiNPs and (b) MSN@SiNPs@TMPyP-FA, which show that the average particle size of silicon nanoparticles is about 3.7nm, and the characteristic lattice of silicon nanoparticles is built in. The average particle size of the mesoporous composite nanoparticles is 88nm.

Figure 2 shows the release efficiency of DOX loaded with MSN@SiNPs@TMPyP-FA at different pH. The figure shows: under acidic (pH=5.5) conditions, the release rate of DOX reaches 66%, while under alkaline (pH=7.4) conditions, the release rate is 32%

Figure 3 is a comparison of two-photon targeted imaging of different cells by MSN@SiNPs@TMPyP-FA. The figure shows that under the same culture conditions, the fluorescence intensity of MCF-7 (folate receptor positive) cells is higher than that of A549 (folate receptor negative).

Figure 4 shows the therapeutic effect of MSN@SiNPs@TMPyP-FA on MCF-7 cells. The figure shows: the survival rate of the MCF-7 drug-loaded + laser irradiation group was the lowest, down to 30%

Example 2:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the quality of sodium citrate in step 1) is 0.2g, in The two-photon fluorescent silicon nanoparticles SiNPs were prepared by reacting at 200°C for 3 h.

Example 3:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the quality of sodium citrate in step 1) is 0.6g, in The two-photon fluorescent silicon nanoparticles SiNPs were prepared by reacting at 200°C for 3 hours.

Example 4:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the quality of sodium citrate in step 1) is 0.4g, in The two-photon fluorescent silicon nanoparticles SiNPs were prepared by reacting at 200°C for 1 h.

Example 5:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the quality of sodium citrate in step 1) is 0.4g, in The two-photon fluorescent silicon nanoparticles SiNPs were prepared by reacting at 200°C for 5 h.

Embodiment 6:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the volume of SiNPs in step 2) is 1mL, and the obtained Multifunctional hybrid MSN@SiNPs@TMPyP for chemotherapy/photodynamic therapy guided by two-photon fluorescence imaging.

Embodiment 7:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the volume of SiNPs in step 2) is 3mL, and the obtained Multifunctional hybrid MSN@SiNPs@TMPyP for chemotherapy/photodynamic therapy guided by two-photon fluorescence imaging.

Embodiment 8:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the quality of TMPyP in step 2) is 1mg, and the obtained Multifunctional hybrid MSN@SiNPs@TMPyP for chemotherapy/photodynamic therapy guided by two-photon fluorescence imaging.

Embodiment 9:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the quality of TMPyP in step 2) is 10mg, and the obtained Multifunctional hybrid MSN@SiNPs@TMPyP for chemotherapy/photodynamic therapy guided by two-photon fluorescence imaging.

Example 10:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the quality of folic acid in step 3) is 40mg, and the obtained Multifunctional complex MSN@SiNPs@TMPyP-FA targeting two-photon fluorescence imaging-guided chemotherapy/photodynamic therapy.

Example 11:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the quality of folic acid in step 3) is 120mg, and the obtained Multifunctional complex MSN@SiNPs@TMPyP-FA targeting two-photon fluorescence imaging-guided chemotherapy/photodynamic therapy.

Example 12:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the quality of EDC in step 3) is 20mg, and the obtained Multifunctional complex MSN@SiNPs@TMPyP-FA targeting two-photon fluorescence imaging-guided chemotherapy/photodynamic therapy.

Example 13:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the quality of EDC in step 3) is 60mg, and the obtained Multifunctional complex MSN@SiNPs@TMPyP-FA targeting two-photon fluorescence imaging-guided chemotherapy/photodynamic therapy.

Example 14:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the quality of NHS in step 3) is 30mg, and the obtained Multifunctional complex MSN@SiNPs@TMPyP-FA targeting two-photon fluorescence imaging-guided chemotherapy/photodynamic therapy.

Example 15:

A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles, the preparation steps are basically the same as in Example 1, the difference is that the quality of NHS in step 3) is 90mg, and the obtained Multifunctional complex MSN@SiNPs@TMPyP-FA targeting two-photon fluorescence imaging-guided chemotherapy/photodynamic therapy.

Claims (4)

1. A method for preparing a multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles. Two-photon fluorescent silicon nanoparticles synthesized by a one-pot hydrothermal method and a photosensitizer are separated by an oil-water two-phase layering method. Mesoporous silicon is co-encapsulated to form a mesoporous silica nanocomposite, and folic acid is further modified to have targeting ability, and finally a multifunctional nanocomposite capable of two-photon fluorescence targeted imaging-guided drug chemotherapy/photodynamic synergistic therapy is formed particle; Including the following steps: 1) Add sodium citrate as a reducing agent to ultrapure water, pass through nitrogen and stir for 10 minutes, then add silicon source N-(2-aminoethyl)-3-aminopropyltrimethoxysilane (DAMO); keep stirring After 15 minutes, move it to a polytetrafluoroethylene reactor, react at 200°C for 1-5 hours, then transfer the obtained yellow solution to a dialysis bag for 24 hours, and change the water every 8 hours to obtain an aqueous solution of SiNPs; 2) Take cetrimonium chloride solution (CTAC), SiNPs aqueous solution obtained in step 1), photosensitizer 5,10,15,20-tetrakis(N-methyl-4-pyridine)porphine p-toluenesulfonate ( TMPyP) and triethanolamine (TEA) were dissolved in ultrapure water, and kept stirring at 60°C for 1h at a stirring speed of 150rmp, then slowly added octadecene solution dissolved in ethyl silicate (TEOS) to continue the reaction After 12 hours, remove the upper layer of oil, the product was washed and collected by centrifugation; then the product was dispersed in an ethanol solution containing ammonium nitrate, refluxed at 66 ° C for 6 hours, and the mesoporous silica composite nanoparticles (MSN @SiNPs @TMPyP); 3) Dissolve folic acid (FA), 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC), N-hydroxysuccinimide (NHS) in dimethyl sulfoxide (DMSO), continued to stir for 30min, then added 3-aminopropyltriethoxysilane (APTES), and stirred overnight to obtain FA-APTES; take FA-APTES, step 2) to obtain MSN@SiNPs@TMPyP Added into methanol solution, stirred at room temperature for 24 hours, and centrifuged to obtain multifunctional mesoporous silica composite nanoparticles modified with folic acid (MSN@SiNPs@TMPyP-FA). 2. the preparation method of the multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles according to claim 1, is characterized in that, ultrapure water, sodium citrate and silicon source N described in step 1) The mass ratio of -(2-aminoethyl)-3-aminopropyltrimethoxysilane is 1:0.025-0.075:0.25. 3. the preparation method of the multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles according to claim 1, is characterized in that, the concentration of cetrimonium chloride solution described in step 2) is 25wt%; The volume ratio of the ethanol, ultrapure water, CTAC solution, SiNPs aqueous solution, octadecene and TEOS is 1:0.38:0.24:0.02-0.06:0.16:0.04; the mass ratio of TEOS, TEA, TMPyP and ammonium nitrate is 1 :0.048:0.0005-0.006:0.016. 4. The preparation method of the multifunctional mesoporous silica composite nanomaterial based on fluorescent silicon nanoparticles according to claim 1, characterized in that, in step 3), APTES, FA, EDC, NHS and MSN@SiNPs The mass ratio of @TMPyP is 1:0.17-0.51:0.084-0.25:0.13-0.38:0.21; the volume ratio of methanol, APTES, DMSO and FA-APTES: 1:0.05:0.4:0.02.