Preparation method of fluorescence-traced targeted photo-thermal anti-tumor nanoparticles
By modifying BaTiO3/Fe nanoparticles for tumor targeting and fluorescent labeling, BaTiO3/Fe-PEI-FA/ICG nanoparticles were prepared, solving the problems of tumor targeting and imaging visualization in clinical applications, enabling precise diagnosis and treatment of osteosarcoma and preventing lung metastasis.
Patent Information
- Application Number
- CN202511331936.7
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-09-18
- Publication Date
- 2026-01-02
AI Technical Summary
BaTiO3/Fe nanoparticles lack tumor targeting and imaging visibility, limiting their application in clinical anti-tumor therapy.
By encapsulating BaTiO3/Fe nanoparticles with polyethyleneimine for tumor targeting modification and introducing indocyanine green for fluorescent labeling, BaTiO3/Fe-PEI-FA/ICG nanoparticles were prepared, achieving tumor targeting and imaging visualization.
This approach enables precise localization and targeted therapy of nanoparticles in osteosarcoma tissue. Combined with photothermal and ultrasound dual-response therapy, it effectively kills tumor cells, prevents lung metastasis and recurrence, and reduces systemic adverse reactions.
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Abstract
Description
TECHNICAL FIELD
[0001] The application belongs to the technical field of functionalized nanomaterials, and specifically relates to a preparation method of fluorescently traced targeted photothermal anti-tumor nanoparticles. BACKGROUND
[0002] Barium titanate (BaTiO3, BT) is a classic wide-bandgap ferroelectric semiconductor. Under the alternating acoustic vibration of frequency / time, through its inherent micro-pressure-induced polarization, an internal electric field and surface potential can be immediately established, thereby triggering an oxidation-reduction reaction to produce reactive oxygen species (ROS), and high levels of ROS exceeding the threshold value can induce oxidative stress, destroy intracellular homeostasis, and induce regulatory cell death, that is, BT nanoparticles can achieve high sonodynamic anti-tumor therapy by generating ROS, thereby eradicating tumors.
[0003] In view of the fact that ultrasound (US) and magnetic resonance (MRI) are suitable and safe medical imaging techniques, have the advantages of non-invasiveness, no ionizing radiation, high spatial resolution and strong tissue penetration depth, and iron-based nanoparticles have been widely used as MRI imaging contrast agents for various disease models, and based on the previous research of the team, the dual effect of iron-containing barium titanate scaffolds on anti-tumor and bone repair [Adv. Healthcare Mater. 2023, 2302901], BaTiO3 / Fe generates ROS intelligently through sonodynamic therapy (SDT) under the synergistic action of ultrasound (US) and near-infrared light (NIR), and plays an anti-tumor role, in addition, the introduction of iron ions into the crystal structure of BaTiO3 nanoparticles endows BaTiO3 / Fe with paramagnetism, which can monitor the accumulation and distribution of BaTiO3 / Fe in real time, thereby improving the accurate positioning of ultrasound (US) cardiogram and magnetic resonance (MRI).
[0004] However, BaTiO3 / Fe itself does not have tumor targeting and imaging visibility, which seriously limits its application in clinical anti-tumor therapy, therefore, it is expected to realize the development of diagnosis and targeted treatment of osteosarcoma (OS) by functionalizing and modifying the iron-containing barium titanate nanoparticles (nBaTiO3 / Fe). SUMMARY
[0005] The application aims to provide a preparation method of fluorescently traced targeted photothermal anti-tumor nanoparticles, and the prepared nanoparticles have tumor targeting and imaging visibility, and can realize the development of diagnosis and targeted treatment of osteosarcoma.
[0006] The application is realized by the following technical solutions:
[0007] A preparation method of fluorescently traced targeted photothermal anti-tumor nanoparticles, comprising the following steps:
[0008] Step 1, preparation of iron-containing barium titanate nanoparticles BaTiO3 / Fe;
[0009] Step 2, using electrostatic adsorption method, BaTiO3 / Fe is coated with polyethyleneimine to obtain PEI-coated photothermal antitumor nanoparticles, denoted as BaTiO3 / Fe-PEI;
[0010] Step 3, preparation of nanoparticles BaTiO3 / Fe-PEI-FA, the specific process is as follows:
[0011] Step 3.1, 2-6 g of folic acid, 0.4-0.8 g of N-hydroxysuccinimide and 0.5-0.9 g of dichloroethane are dissolved in 150 mL of ultrapure water, magnetically stirred for 2 h, and the pH value is adjusted to 10 using NaOH solution to activate FA, to obtain a mixed solution A;
[0012] Step 3.2, according to 1 g:(3-4 mL), BaTiO3 / Fe-PEI is dispersed in mixed solution A, and the reaction is stirred for 24 h to obtain reaction liquid B;
[0013] Step 3.3, dialysis of reaction liquid B to remove unreacted reagents, centrifugal separation of solid product, washing to obtain tumor targeting nanoparticles, denoted as BaTiO3 / Fe-PEI-FA;
[0014] Step 4, preparation of nanoparticles BaTiO3 / Fe-PEI-FA / ICG, the specific process is as follows:
[0015] Step 4.1, 5-10 mg of indocyanine green is added to 10 mL of phosphate buffer solution with a concentration of 0.01 mol / L and a pH of 7.4 to obtain a phosphate buffer solution containing indocyanine green;
[0016] Step 4.2, according to 1 g:(3-5 mL), BaTiO3 / Fe-PEI-FA is dispersed in the phosphate buffer solution containing indocyanine green, and the reaction is carried out in the dark for 12 h, the solid product is separated by centrifugation, and the free indocyanine green is removed by washing, and then freeze-drying to obtain fluorescent tracing of targeted photothermal antitumor nanoparticles, denoted as BaTiO3 / Fe-PEI-FA / ICG.
[0017] Further, the specific process of step 1 is as follows: KOH solution, Ba(OH)2 solution, nano-TiO2 powder and Fe2O3 powder are mixed according to the molar ratio of KOH:Ba(OH)2:nano-TiO2:Fe2O3 (0.05-0.15):(0.005-0.015):(0.005-0.015):(0.05-0.15), and placed in a polytetrafluoroethylene-lined high-temperature high-pressure reaction kettle, and hydrothermally reacted at 85-95℃ for 4h, and the solid product is separated by centrifugation, washed, and the iron-containing barium titanate nanoparticles BaTiO3 / Fe are obtained.
[0018] Further, the specific process of step 2 is as follows: first, BaTiO3 / Fe is taken in a single-mouth round-bottom flask containing distilled water according to 1g:(3-5mL), and ultrasonic dispersion is performed to obtain solution C, then polyethyleneimine is added to solution C according to the volume ratio of polyethyleneimine to distilled water 1:(2-3) at 80-90℃, while stirring solution C, and the stirring reaction is continued for 3h to obtain reaction liquid D, and the solid product is separated by centrifugation and washed to obtain PEI-coated photothermal antitumor nanoparticles.
[0019] Further, the process of separating the solid product by centrifugation and washing is as follows: the reaction liquid D is centrifuged at 12000rpm for 10min at 4℃, the solid product is collected, and ultrapure water is used for vortex and centrifugal washing 3 times to remove free polyethyleneimine.
[0020] Further, the concentration of the NaOH solution in step 3.1 is 1.5mol / L.
[0021] Further, the process of dialyzing reaction liquid B in step 3.3 is as follows: reaction liquid B is transferred to a dialysis bag and dialyzed in ultrapure water for 24h.
[0022] Further, the washing in steps 3.3 and 4 is vortexed and centrifuged with ultrapure water until the washing liquid is clear.
[0023] Further, the freeze-drying of step 4 is freeze-dried at -60℃ for 12h.
[0024] The present application has the following beneficial technical effects:
[0025] This invention first encapsulates nBaTiO3 / Fe nanoparticles with polyethyleneimine (PEI) to obtain PEI-encapsulated photothermal antitumor nanoparticles BaTiO3 / Fe-PEI. Then, through the conjugation reaction between the amino groups of PEI and the carboxyl groups of folic acid (FA), the PEI-encapsulated BaTiO3 / Fe nanoparticles are chemically grafted to achieve tumor-targeting modification, resulting in photothermal targeted antitumor nanoparticles BaTiO3 / Fe-PEI-FA. Folic acid not only possesses the advantages of stable properties, non-immunogenicity, and low cost, but also... The nanoparticles exhibit a very high affinity for tumor cell surface receptors. Their short chains and small size facilitate phagocytosis and internalization by the cells, specifically targeting the tumor cell receptors bound to folic acid. This endows the BaTiO3 / Fe-PEI-FA nanoparticles with tumor targeting specificity. Then, for imaging visualization and fluorescence tracing to locate their position in osteosarcoma tissue, indocyanine green (ICG) was introduced for fluorescent labeling, resulting in fluorescently traced targeted photothermal antitumor nanoparticles BaTiO3 / Fe-PEI-FA / ICG. These nanoparticles were then visualized using a fluorescence display in osteosarcoma tissue. The localization and distribution of BaTiO3 / Fe-PEI-FA / ICG within tissues, under dual-mode ultrasound and magnetic resonance imaging, enables precise localization diagnosis and targeted therapy for osteosarcoma. Specifically: First, its paramagnetic and ultrasound properties allow for imaging under both MRI and ultrasound to diagnose the localization and distribution of osteosarcoma. Second, its dual photothermal and ultrasound response, combining low-intensity ultrasound (SDT) sonodynamic therapy (SDT) with mild near-infrared light (PDT), induces BaTiO3 / Fe-PEI-FA / ICG to generate a large amount of reactive oxygen species (ROS) to kill osteosarcoma, thus achieving precise targeted therapy. Third, for osteosarcoma with lung metastases, the combined action of PDT and SDT effectively inhibits OS growth, preventing recurrence and metastasis. Fourth, based on the biocompatibility of BaTiO3 / Fe-PEI-FA / ICG, the combined PDT and SDT targeted therapy for OS reduces systemic adverse reactions and avoids the problem of poor NIR penetration, providing an effective strategy for the precise diagnosis and targeted therapy of OS in deep tissues and metastatic lungs. In summary, this invention, through the size advantage and customized properties of functionalized nanoparticles BaTiO3 / Fe-PEI-FA / ICG, enables highly efficient targeted therapy and metastasis prevention of osteosarcoma, demonstrating outstanding advantages in intelligent diagnostic imaging and nanotherapy. Attached Figure Description
[0026] Figure 1 This is a flowchart of the preparation of fluorescently traced targeted photothermal antitumor nanoparticles according to the present invention;
[0027] Figure 2Molecular structural formula of the nanoparticles BaTiO3 / Fe, BaTiO3 / Fe-PEI, BaTiO3 / Fe-PEI-FA and BaTiO3 / Fe-PEI-FA / ICG prepared in Example 1 of the present application;
[0028] Fig. 3(a) is a transmission electron microscope image and element distribution diagram of the nanoparticles BaTiO3-PEI-FA prepared in Comparative Example 1 of the present application;
[0029] Fig. 3(b) is a transmission electron microscope image and element distribution diagram of the nanoparticles BaTiO3 / Fe-PEI-FA prepared in Example 1 of the present application;
[0030] Figure 4 Fig. 4 is a Fourier transform infrared spectrogram of BaTiO3-PEI-FA prepared in Comparative Example 1 of the present application, and the nanoparticles BaTiO3 / Fe-PEI and BaTiO3 / Fe-PEI-FA prepared in Example 1 of the present application;
[0031] Figure 5 Fig. 5 is the effect of the nanoparticles BaTiO3 prepared in Comparative Example 1 of the present application and the nanoparticles BaTiO3 / Fe prepared in Example 1 of the present application on human osteosarcoma cells under different treatment conditions;
[0032] Figure 6 Fig. 6 is a fluorescence image of the uptake of BaTiO3 / Fe-PEI-FA / ICG prepared in Example 1 of the present application by Saos-2 cells and mouse osteogenic precursor cells. DETAILED DESCRIPTION
[0033] The present application will be further described in detail below in combination with specific examples, which are an explanation rather than a limitation of the present application.
[0034] The flow of preparing the fluorescently traced targeted photothermal anti-tumor nanoparticles BaTiO3 / Fe-PEI-FA / ICG in Example 1-Example 3 of the present application is as follows: Figure 1As shown, first, nanoparticles BaTiO3 / Fe were prepared by hydrothermal synthesis, then BaTiO3 / Fe was coated with PEI by electrostatic adsorption under magnetic stirring to obtain nanoparticles BaTiO3 / Fe-PEI, then the conjugate reaction between the carboxyl group of folic acid (FA) and the amine group of polyethyleneimine (PEI) was performed to obtain targeted photothermal antitumor nanoparticles BaTiO3 / Fe-PEI-FA, then indocyanine green (ICG) was used to fluorescently label BaTiO3 / Fe-PEI-FA to obtain nanoparticles BaTiO3 / Fe-PEI-FA / ICG, which has fluorescence visualization and tracing functions, wherein the molecular structural formula of nanoparticles BaTiO3 / Fe, BaTiO3 / Fe-PEI, BaTiO3 / Fe-PEI-FA and BaTiO3 / Fe-PEI-FA / ICG are as shown in Figure 2
[0035] The purpose of using ultrapure water vortex and centrifugal washing in Examples 1-3 and Comparative Example 1 is to first mix pure water with nanoparticles uniformly by vortexing to achieve sufficient contact, and then centrifugally separate the solid product to facilitate cleaning of the nanoparticles and thus remove residual reaction solution.
[0036] Example 1
[0037] Step 1, preparation of iron-containing barium titanate nanoparticles BaTiO3 / Fe, the specific process is as follows: KOH solution, Ba(OH)2 solution, nano-TiO2 powder and Fe2O3 powder were mixed according to a molar ratio of KOH:Ba(OH)2:nano-TiO2:Fe2O3 of 0.1:0.01:0.01:0.1, and placed in a polytetrafluoroethylene-lined high-temperature high-pressure reaction kettle, and hydrothermal reaction was carried out at 90°C for 4h. The solid product was centrifugally separated and washed to obtain iron-containing barium titanate nanoparticles BaTiO3 / Fe;
[0038] Step 2, BaTiO3 / Fe was coated with polyethyleneimine by electrostatic adsorption to obtain PEI-coated photothermal antitumor nanoparticles, the specific process is as follows: first, BaTiO3 / Fe was added to a single-neck round-bottom flask containing distilled water according to a ratio of 1g:4mL, and ultrasonic dispersion was performed to obtain solution C, then polyethyleneimine was added to solution C according to a volume ratio of polyethyleneimine to distilled water of 1:3 while stirring solution C at 85°C, and the stirring reaction was continued for 3h to obtain reaction liquid D, reaction liquid D was centrifuged at 12000rpm for 10min at 4°C, and the solid product was collected and washed with ultrapure water by vortexing and centrifugation for 3 times to remove free polyethyleneimine, thereby obtaining PEI-coated photothermal antitumor nanoparticles, denoted as BaTiO3 / Fe-PEI;
[0039] Step 3, preparation of nanoparticles BaTiO3 / Fe-PEI-FA, the specific process is as follows:
[0040] Step 3.1, 4g of folic acid, 0.6g of N-hydroxysuccinimide and 0.7g of dichloroethane were dissolved in 150mL of ultrapure water and magnetically stirred for 2h, and a 1.5mol / L NaOH solution was used to adjust the pH value to 10 to activate the FA, and a mixed solution A was obtained;
[0041] Step 3.2, according to the ratio of 1g:3mL, BaTiO3 / Fe-PEI was dispersed in the mixed solution A, and the reaction was stirred for 24h to obtain a reaction liquid B;
[0042] Step 3.3, the reaction liquid B was transferred to a dialysis bag and dialyzed in ultrapure water for 24h to remove unreacted reagents, and the solid product was separated by centrifugation, and washed with ultrapure water by vortex and centrifugation until the washing liquid was clear, to obtain tumor-targeting nanoparticles, denoted as BaTiO3 / Fe-PEI-FA;
[0043] Step 4, preparation of nanoparticles BaTiO3 / Fe-PEI-FA / ICG, the specific process is as follows:
[0044] Step 4.1, 5mg of indocyanine green was added to 10mL of phosphate buffer solution with a concentration of 0.01mol / L and a pH of 7.4 to obtain a phosphate buffer solution containing indocyanine green;
[0045] Step 4.2, according to the ratio of 1g:3mL, BaTiO3 / Fe-PEI-FA was dispersed in the phosphate buffer solution containing indocyanine green, and the reaction was carried out in the dark for 12h, and the solid product was separated by centrifugation, and washed with ultrapure water by vortex and centrifugation until the washing liquid was clear, to remove free indocyanine green, and freeze-dried at-60℃ for 12h to obtain fluorescently labeled targeted photothermal antitumor nanoparticles, denoted as BaTiO3 / Fe-PEI-FA / ICG.
[0046] Example 2
[0047] Step 1, preparation of iron-containing barium titanate nanoparticles BaTiO3 / Fe, the specific process is as follows: KOH solution, Ba(OH)2 solution, nano-TiO2 powder and Fe2O3 powder were mixed according to the molar ratio of KOH, Ba(OH)2, nano-TiO2 and Fe2O3 0.05:0.005:0.005:0.05, and placed in a polytetrafluoroethylene-lined high-temperature and high-pressure reaction kettle, and hydrothermal reaction was carried out at 85℃ for 4h, and the solid product was separated by centrifugation, and washed to obtain iron-containing barium titanate nanoparticles BaTiO3 / Fe;
[0048] Step 2, BaTiO3 / Fe was coated with polyethyleneimine by electrostatic adsorption to obtain PEI-coated photothermal anti-tumor nanoparticles. Specifically, 1 g of BaTiO3 / Fe was added to a single-mouth round-bottom flask containing distilled water, and ultrasonic dispersion was performed to obtain solution C. Then, 1 g of polyethyleneimine was added to solution C at a volume ratio of 1:2 under stirring at 80°C for 3 h to obtain reaction solution D. The reaction solution D was centrifuged at 12,000 rpm for 10 min at 4°C to collect the solid product. The collected product was washed with ultrapure water by vortexing and centrifugation for 3 times to remove free polyethyleneimine, and PEI-coated photothermal anti-tumor nanoparticles were obtained, which were denoted as BaTiO3 / Fe-PEI;
[0049] Step 3, nanoparticles BaTiO3 / Fe-PEI-FA were prepared. Specifically,
[0050] Step 3.1, 2 g of folic acid, 0.4 g of N-hydroxysuccinimide, and 0.5 g of dichloroethane were dissolved in 150 mL of ultrapure water and magnetically stirred for 2 h. The pH value was adjusted to 10 using a 1.5 mol / L NaOH solution to activate the FA to obtain a mixed solution A.
[0051] Step 3.2, 1 g of BaTiO3 / Fe-PEI was dispersed in the mixed solution A at a ratio of 1 g:4 mL, and the reaction was stirred for 24 h to obtain reaction solution B.
[0052] Step 3.3, the reaction solution B was transferred to a dialysis bag and dialyzed in ultrapure water for 24 h to remove unreacted reagents. The solid product was separated by centrifugation, and washed with ultrapure water by vortexing and centrifugation until the washing liquid was clear to obtain tumor-targeting nanoparticles, which were denoted as BaTiO3 / Fe-PEI-FA.
[0053] Step 4, nanoparticles BaTiO3 / Fe-PEI-FA / ICG were prepared. Specifically,
[0054] Step 4.1, 7.5 mg of indocyanine green was added to 10 mL of phosphate buffer solution with a concentration of 0.01 mol / L and a pH of 7.4 to obtain a phosphate buffer solution containing indocyanine green.
[0055] Step 4.2, 1 g of BaTiO3 / Fe-PEI-FA was dispersed in the phosphate buffer solution containing indocyanine green at a ratio of 1 g:4 mL, and the reaction was carried out in the dark for 12 h. The solid product was separated by centrifugation, and washed with ultrapure water by vortexing and centrifugation until the washing liquid was clear to remove free indocyanine green. The mixture was freeze-dried at -60°C for 12 h to obtain fluorescently labeled targeted photothermal anti-tumor nanoparticles, which were denoted as BaTiO3 / Fe-PEI-FA / ICG.
[0056] Example 3
[0057] Step 1, preparation of iron-containing barium titanate nanoparticles BaTiO3 / Fe, the specific process is: according to the molar ratio of KOH, Ba(OH)2, nano-TiO2 and Fe2O3 0.15:0.015:0.015:0.15, take KOH solution, Ba(OH)2 solution, nano-TiO2 powder and Fe2O3 powder, put into polytetrafluoroethylene lined high temperature and high pressure reactor, hydrothermal reaction at 95℃ for 4h, centrifugal separation of solid product, washing, get iron-containing barium titanate nanoparticles BaTiO3 / Fe;
[0058] Step 2, using electrostatic adsorption method, polyethyleneimine coated BaTiO3 / Fe, get PEI coated photothermal antitumor nanoparticles, the specific process is: first according to 1g:5mL, take BaTiO3 / Fe into the single-mouth round-bottom flask containing distilled water, ultrasonic dispersion, get solution C, then according to the volume ratio of polyethyleneimine and distilled water 1:2.5, at 90℃, stirring solution C, adding polyethyleneimine to it, continue stirring for 3h, get reaction liquid D, at 4℃, centrifugal at 12000rpm speed for 10min, collect the solid product, use ultrapure water to spin and centrifugal wash 3 times to remove free polyethyleneimine, get PEI coated photothermal antitumor nanoparticles, recorded as BaTiO3 / Fe-PEI;
[0059] Step 3, preparation of nanoparticles BaTiO3 / Fe-PEI-FA, the specific process is:
[0060] Step 3.1, 6g folic acid, 0.8g N-hydroxysuccinimide and 0.9g dichloroethane were dissolved in 150mL ultrapure water, magnetic stirring for 2h, using 1.5mol / L NaOH solution to adjust the pH value to 10, to activate FA, get mixed solution A;
[0061] Step 3.2, according to 1g:3.5mL, BaTiO3 / Fe-PEI was dispersed in mixed solution A, stirring reaction for 24h, get reaction liquid B;
[0062] Step 3.3, transfer reaction liquid B to dialysis bag, dialysis in ultrapure water for 24h, remove unreacted reagents, centrifugal separation of solid product, using ultrapure water to spin and centrifugal wash until the washing liquid is clear, get tumor targeting nanoparticles, recorded as BaTiO3 / Fe-PEI-FA;
[0063] Step 4, preparation of nanoparticles BaTiO3 / Fe-PEI-FA / ICG, the specific process is:
[0064] Step 4.1, 10 mg of indocyanine green was added into 10 mL of phosphate buffer solution with a concentration of 0.01 mol / L and a pH of 7.4 to obtain an indocyanine green-containing phosphate buffer solution;
[0065] Step 4.2, BaTiO3 / Fe-PEI-FA was dispersed in the indocyanine green-containing phosphate buffer solution according to 1 g:5 mL, and reacted in the dark for 12 h. The solid product was separated by centrifugation, and washed with ultrapure water by vortexing and centrifugation until the washing liquid was clear, so as to remove free indocyanine green. The fluorescent tracer targeted photothermal antitumor nanoparticles, denoted as BaTiO3 / Fe-PEI-FA / ICG, were obtained by freeze-drying at-60℃ for 12 h.
[0066] Comparative Example 1
[0067] Step 1, barium titanate nanoparticles BaTiO3 were prepared as follows: KOH solution, Ba(OH)2 solution and nano-TiO2 powder were mixed according to a molar ratio of KOH:Ba(OH)2:nano-TiO2 of 0.1:0.01:0.01, and placed in a polytetrafluoroethylene-lined high-temperature high-pressure reaction kettle. Hydrothermal reaction was carried out at 180℃ for 4 h. The solid product was separated by centrifugation, and washed to obtain barium titanate nanoparticles BaTiO3;
[0068] Step 2, BaTiO3 was coated with polyethyleneimine by electrostatic adsorption to obtain PEI-coated nanoparticles as follows: BaTiO3 was added to a single-mouth round-bottom flask containing distilled water according to 1 g:4 mL, and ultrasonically dispersed to obtain solution E. Polyethyleneimine was added to solution E while stirring at 85℃ according to a volume ratio of polyethyleneimine to distilled water of 1:3, and the stirring was continued for 3 h to obtain reaction liquid F. Reaction liquid F was centrifuged at 12000 rpm for 10 min at 4℃ to collect the solid product, which was washed with ultrapure water by vortexing and centrifugation for 3 times to remove free polyethyleneimine, and PEI-coated nanoparticles, denoted as BaTiO3-PEI, were obtained.
[0069] Step 3, nanoparticles BaTiO3-PEI-FA were prepared as follows:
[0070] Step 3.1, 4 g of folic acid, 0.6 g of N-hydroxysuccinimide and 0.7 g of dichloroethane were dissolved in 150 mL of ultrapure water and magnetically stirred for 2 h. The pH value was adjusted to 10 using a 1.5 mol / L NaOH solution to activate the FA, and a mixed solution G was obtained.
[0071] Step 3.2, BaTiO3-PEI was dispersed in the mixed solution G according to 1 g:3 mL, and stirred for 24 h to obtain reaction liquid H;
[0072] Step 3.3, transfer the reaction solution H to a dialysis bag, dialyze in ultrapure water for 24 h, centrifugalize the solid product, wash with ultrapure water by vortex and centrifugalization until the washing solution is clear, to obtain the tumor-targeting nanoparticles, denoted as BaTiO3-PEI-FA.
[0073] Fig. 3(a) and Fig. 3(b) respectively show the transmission electron microscope images and element distribution maps (Mapping) of the nanoparticles BaTiO3-PEI-FA prepared in Comparative Example 1 and the nanoparticles BaTiO3 / Fe-PEI-FA prepared in Example 1, from which it can be seen that the elements Ba, Ti, O, N, C and Fe are uniformly distributed and correspond to the surface morphological characteristics of the scanning electron microscope (SEM).
[0074] Figure 4 Fig. 4 shows the Fourier transform infrared spectra (FTIR) of BaTiO3-PEI-FA prepared in Comparative Example 1, the nanoparticles BaTiO3 / Fe-PEI prepared in Example 1 and BaTiO3 / Fe-PEI-FA, wherein the three spectra from bottom to top in the figure correspond to BaTiO3 / Fe-PEI, BaTiO3 / Fe-PEI-FA and BaTiO3-PEI-FA, respectively, it can be seen that the -NH2 on the BaTiO3 / Fe-PEI spectrum is a characteristic chemical bond on PEI, the COO- and -OH appearing on the BaTiO3-PEI-FA spectrum are both characteristic chemical bonds of FA, indicating the successful introduction of FA; the BaTiO3 / Fe-PEI-FA spectrum shows all the characteristic peaks of PEI and FA, indicating that the doping of iron does not have a significant effect on the organic modification process, FA is successfully coupled to BaTiO3 / Fe-PEI, and the targeting function of the material is realized.
[0075] Referring to Figure 5 Fig. 5 shows the effects of the nanoparticles BaTiO3 under ultrasonic and the nanoparticles BaTiO3 / Fe prepared in Example 1 under different treatment conditions on human osteosarcoma cells (Saos-2), it can be seen that the nanoparticles BaTiO3 / Fe prepared in Example 1 can completely inhibit the growth of tumor cells under the conditions of photothermal and US dual response, indicating that the nanoparticles BaTiO3 / Fe have good anti-tumor function.
[0076] From Figure 6It can be seen that the uptake of the nanoparticles BaTi03 / Fe-PEI-FA / ICG by human osteosarcoma cells (Saos-2) is more obvious than that by mouse osteogenic precursor cells (MC3T3-E1), indicating that the tumor cell targeting of nBaTi03 / Fe-PEI-FA is achieved by coupling the carboxyl of FA with the amine group of PEI-wrapped BaTi03 / Fe.
Claims
1. A method for preparing fluorescently traced targeted photothermal antitumor nanoparticles, characterized in that, The method comprises the following steps: Step 1, preparing iron-containing barium titanate nanoparticles BaTiO3 / Fe; Step 2, wrapping the BaTiO3 / Fe with polyethyleneimine by electrostatic adsorption to obtain PEI-wrapped photothermal anti-tumor nanoparticles, denoted as BaTiO3 / Fe-PEI; Step 3, preparing nanoparticles BaTiO3 / Fe-PEI-FA, and the specific process is as follows: Step 3.1, dissolving 2-6 g of folic acid, 0.4-0.8 g of N-hydroxysuccinimide and 0.5-0.9 g of dichloroethane in 150 mL of ultrapure water, magnetically stirring for 2 h, adjusting the pH value to 10 by using a NaOH solution to activate the FA, and obtaining a mixed solution A; Step 3.2, dispersing the BaTiO3 / Fe-PEI in the mixed solution A according to 1 g:(3-4 mL), stirring for 24 h to obtain a reaction liquid B; Step 3.3, dialyzing the reaction liquid B to remove unreacted reagents, centrifuging to separate the solid product, washing, and obtaining tumor-targeting nanoparticles, denoted as BaTiO3 / Fe-PEI-FA; Step 4, preparing nanoparticles BaTiO3 / Fe-PEI-FA / ICG, and the specific process is as follows: Step 4.1, adding 5-10 mg of indocyanine green to 10 mL of phosphate buffer solution with a pH of 7.4 to obtain a phosphate buffer solution containing indocyanine green; Step 4.2, dispersing the BaTiO3 / Fe-PEI-FA in the phosphate buffer solution containing indocyanine green according to 1 g:(3-5 mL), avoiding light for 12 h, centrifuging to separate the solid product, washing to remove free indocyanine green, and freeze-drying to obtain fluorescence-traced targeted photothermal anti-tumor nanoparticles, denoted as BaTiO3 / Fe-PEI-FA / ICG.
2. The method for preparing fluorescently traced targeted photothermal antitumor nanoparticles according to claim 1, characterized in that, The specific process of step 1 is as follows: taking KOH solution, Ba(OH)2 solution, nano-TiO2 powder and Fe2O3 powder according to the molar ratio of KOH:Ba(OH)2:nano-TiO2:Fe2O3(0.05-0.15):(0.005-0.015):(0.005-0.015):(0.05-0.15), mixing them, placing them in a polytetrafluoroethylene-lined high-temperature and high-pressure reaction kettle, and hydrothermally reacting at 85-95 ℃ for 4 h, centrifuging to separate the solid product, and washing to obtain iron-containing barium titanate nanoparticles BaTiO3 / Fe.
3. The method of claim 1, wherein the fluorescently traced, targeted photothermal antitumor nanoparticle is prepared by the steps of: (a) synthesizing a fluorescently traced, targeted photothermal antitumor nanoparticle; and (b) purifying the fluorescently traced, targeted photothermal antitumor nanoparticle. The specific process of step 2 is as follows: first, taking BaTiO3 / Fe according to 1 g:(3-5 mL) and adding it into a single-necked round-bottom flask containing distilled water, ultrasonic dispersing to obtain solution C, then adding polyethyleneimine into solution C according to the volume ratio of polyethyleneimine to distilled water 1:(2-3) while stirring solution C at 80-90 ℃, continuously stirring for 3 h to obtain reaction liquid D, centrifuging to separate the solid product and washing to obtain PEI-wrapped photothermal anti-tumor nanoparticles.
4. The method for preparing fluorescently traced targeted photothermal antitumor nanoparticles according to claim 3, characterized in that, The centrifugation and washing of the solid product is as follows: centrifugation of reaction liquid D at 12000 rpm for 10 min at 4℃, collection of the solid product, and vortexing and centrifugation washing with ultrapure water for 3 times to remove free polyethyleneimine.
5. The method of claim 1, wherein the fluorescently traced, targeted photothermal antitumor nanoparticle is prepared by the steps of: (a) synthesizing a fluorescently traced, targeted photothermal antitumor nanoparticle; and (b) purifying the fluorescently traced, targeted photothermal antitumor nanoparticle. The concentration of the NaOH solution in step 3.1 is 1.5 mol / L.
6. The method of claim 1, wherein the fluorescently traced, targeted photothermal antitumor nanoparticle is prepared by the steps of: (a) synthesizing a fluorescently traced, targeted photothermal antitumor nanoparticle; and (b) purifying the fluorescently traced, targeted photothermal antitumor nanoparticle. The dialysis of reaction liquid B in step 3.3 is as follows: transferring reaction liquid B to a dialysis bag and dialysis in ultrapure water for 24 h.
7. The method of claim 1, wherein the fluorescently traced, targeted photothermal antitumor nanoparticle is prepared by the steps of: (a) synthesizing a fluorescently traced, targeted photothermal antitumor nanoparticle; and (b) purifying the fluorescently traced, targeted photothermal antitumor nanoparticle. The washing in step 3.3 and step 4 is vortexing and centrifugation washing with ultrapure water until the washing liquid is clear.
8. The method for preparing fluorescently traced targeted photothermal antitumor nanoparticles according to claim 1, characterized in that, The freeze-drying in step 4 is freeze-drying at -60℃ for 12 h.