CN112336858B

CN112336858B - Bismuth-manganese-based composite particle and preparation method and application thereof

Info

Publication number: CN112336858B
Application number: CN202011071534.5A
Authority: CN
Inventors: 李丽华; 徐兴毅
Original assignee: South China University of Technology SCUT
Current assignee: South China University of Technology SCUT
Priority date: 2020-10-09
Filing date: 2020-10-09
Publication date: 2021-09-21
Anticipated expiration: 2040-10-09
Also published as: CN112336858A

Abstract

The invention discloses bismuth-manganese-based composite particles and a preparation method and application thereof. The bismuth-manganese-based composite particle comprises an inner core and a triple negative breast cancer cell membrane wrapping the inner core; the inner core is formed by gathering a plurality of amino-modified hollow structure bismuth-bismuth trioxide-manganese oxide composite nano particles; indocyanine green is loaded inside and on the surface of the amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nano-particles. The bismuth-manganese-based composite particle has active targeting capability, can improve the tumor microenvironment and provide multiple treatment modes, has high drug loading rate, novel structure and simple preparation method, and can be used for treating larger and more malignant TNBC.

Description

Bismuth-manganese-based composite particle and preparation method and application thereof

Technical Field

The invention relates to the technical field of organic-inorganic composite biological materials, in particular to bismuth-manganese-based composite particles and a preparation method and application thereof.

Background

Breast cancer is a malignant tumor occurring in mammary epithelial tissues, at least 1 of 10 cancer patients worldwide is a breast cancer patient, and 99% of breast cancer patients are female, which seriously threatens the health of women. About 15% to 20% of breast cancer patients have Triple Negative Breast Cancer (TNBC) with high mortality. TNBC refers to a specific type of breast cancer that is clinically negative for all of the Estrogen Receptor (ER), the Progesterone Receptor (PR), and the human epidermal growth factor receptor-2 (HER-2). TNBC has no specific receptor and antibody, has higher invasiveness, recurrence rate and transfer rate, and has poorer curative effect on the TNBC by the conventional treatment means.

The low cure rate of TNBC is not only related to the lack of drug targets, but also related to abnormal micro-environments such as tumor parts lack of oxygen, more reducing Glutathione (GSH) and the like. The nano-particle materials (NPs) which can improve the tumor microenvironment and provide the targeting capability and the multiple treatment methods have wide application prospects in the field. Currently, reported nanoparticle materials for TNBC include CuS @ mSiO with a core-shell structure₂/ICG, MnO of hollow mesoporous structure₂Ce6, etc. CuS @ mSiO of core-shell structure₂The function of the/ICG is single, only photothermal therapy can be provided, the structure is simple, the porosity is low, the loading rate of the organic molecule ICG is only about 8%, in addition, the material also lacks the targeting capability, the enrichment at the tumor part is less, and the final therapeutic effect is influenced. MnO of hollow mesoporous structure₂the/Ce 6 can improve hypoxic environment and provide two treatment modes of photo-thermal and photodynamic, the hollow mesoporous structure is novel, and the porosity is relative to CuS @ mSiO of the core-shell structure₂the/ICG is improved, but the loading rate of an organic molecule Ce6 (chlorin e6) is only about 17%, and the material can be passively targeted only by a method of adding a magnetic field from the outside, so that the targeting efficiency is low, and the treatment effect on larger and malignant cancer cells is poor.

Therefore, the development of a nanoparticle material which has active targeting property, high drug loading rate, multifunctional cooperative therapy and novel structure and can be used for TNBC therapy is urgently needed.

Disclosure of Invention

It is an object of the present invention to provide a bismuth-manganese-based composite particle.

The second object of the present invention is to provide a method for preparing the bismuth-manganese-based composite particle.

The invention also aims to provide application of the bismuth-manganese-based composite particles.

The technical scheme adopted by the invention is as follows:

a bismuth-manganese-based composite particle comprises an inner core and a triple negative breast cancer cell membrane wrapping the inner core; the inner core is formed by gathering a plurality of amino-modified hollow structure bismuth-bismuth trioxide-manganese oxide composite nano particles; indocyanine green is loaded inside and on the surface of the amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nano-particles.

Preferably, the loading amount of the indocyanine green in the bismuth-manganese-based composite particles is 45-55%.

Preferably, the particle size of the bismuth-manganese-based composite particles is 300nm to 400 nm.

Preferably, the particle size of the amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nanoparticle is 30 nm-80 nm.

The preparation method of the bismuth-manganese-based composite particle comprises the following steps:

1) adding bismuth salt into a reductive organic solvent, and carrying out reduction precipitation to obtain bismuth nanoparticles;

2) carrying out a reaction between the bismuth nanoparticles and permanganate to obtain bismuth-bismuth trioxide-manganese oxide composite nanoparticles;

3) modifying the bismuth-bismuth trioxide-manganese oxide composite nano particles by using an aminosilane coupling agent to obtain amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nano particles;

4) carrying out operation of loading indocyanine green on the amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nano-particles to obtain bismuth-bismuth trioxide-manganese oxide composite nano-particles loaded with indocyanine green;

5) and coating the bismuth-bismuth trioxide-manganese oxide composite nano particles loaded with indocyanine green by using a triple-negative breast cancer cell membrane to obtain the bismuth-manganese-based composite particles.

Preferably, the preparation method of the bismuth-manganese-based composite particle comprises the following steps:

1) adding bismuth salt into a reductive organic solvent, stirring for reaction, and separating and purifying a product to obtain bismuth nanoparticles;

2) adding bismuth nanoparticles, an emulsifier and permanganate into a solvent, stirring for reaction, and separating and purifying a product to obtain bismuth-bismuth trioxide-manganese oxide composite nanoparticles;

3) adding an aminosilane coupling agent into the bismuth-bismuth trioxide-manganese oxide composite nanoparticle dispersion liquid, stirring for reaction, and separating and purifying a product to obtain amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nanoparticles;

4) adding indocyanine green into the amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nanoparticle dispersion liquid, stirring and mixing, and separating a product to obtain bismuth-bismuth trioxide-manganese oxide composite nanoparticles loaded with indocyanine green;

5) adding the cell membrane of the triple-negative breast cancer into the bismuth-bismuth trioxide-manganese oxide composite nanoparticle dispersion liquid loaded with indocyanine green, stirring and mixing, and separating the product to obtain the bismuth-manganese-based composite particles.

Preferably, the bismuth salt in step 1) is at least one of bismuth nitrate and bismuth chloride.

More preferably, the bismuth salt in the step 1) is Bi (NO)₃)₃·5H₂O。

Preferably, the organic solvent having reducibility in step 1) is dodecanethiol.

Preferably, the permanganate in step 2) is at least one of potassium permanganate and sodium permanganate.

Further preferably, the permanganate in step 2) is potassium permanganate.

Preferably, the mass ratio of the bismuth nanoparticles to the permanganate in the step 2) is 1: (0.5-2).

Preferably, the emulsifier in step 2) is alkylphenol ethoxylate.

Preferably, the volume ratio of the aminosilane coupling agent to the bismuth-bismuth trioxide-manganese oxide composite nanoparticle dispersion liquid in the step 3) is 1: (10-30).

Preferably, the aminosilane coupling agent in step 3) is Aminopropyltriethoxysilane (APTES).

Preferably, the mass ratio of the indocyanine green to the amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nanoparticle in the step 4) is 1: (0.5-2).

Preferably, the mass ratio of the triple-negative breast cancer cell membrane in the step 5) to the bismuth-bismuth trioxide-manganese oxide composite nanoparticle loaded with indocyanine green is 1: (2-10).

The invention has the beneficial effects that: the bismuth-manganese-based composite particle has active targeting capability, can improve the tumor microenvironment and provide multiple treatment modes, has high drug loading rate, novel structure and simple preparation method, and can be used for treating larger and more malignant TNBC.

Specifically, the method comprises the following steps:

1) according to the invention, the bismuth-manganese-based composite particles are wrapped by the TNBC cell membrane, so that the bismuth-manganese-based composite particles can be endowed with active targeting capability, and further, the accurate targeting of TNBC cells lacking target spots can be realized;

2) according to the invention, the bismuth-bismuth trioxide-manganese oxide composite nano particles are treated by the aminosilane coupling agent, so that amino groups can be modified on the surfaces of the bismuth-bismuth trioxide-manganese oxide composite nano particles to enable the bismuth-bismuth trioxide-manganese oxide composite nano particles to have positive charges, and the outermost MnO of the bismuth-bismuth trioxide-manganese oxide composite nano particles can be enabled to be MnO by utilizing the Kerkadar effect_xThe bismuth-bismuth trioxide-manganese oxide composite nano-particles are mutually diffused with the innermost Bi to form hollowing, so that the specific surface area and the porosity of the bismuth-bismuth trioxide-manganese oxide composite nano-particles are greatly increased, and the loading capacity of organic molecule indocyanine green (ICG) is improved;

3) the bismuth-manganese-based composite particles of the invention are loaded with organic molecules ICG, which can enhance the photo-thermal and photodynamic treatment effects of the material;

4) the multivalent manganese exists on the surface of the bismuth-manganese-based composite particle, so that the material can be endowed with the capability of catalyzing oxygen production and improving oxygen deficiency, the photodynamic therapy can be promoted, and the chemodynamic therapy can be realized;

5) the bismuth-manganese-based composite particle has multiple treatment effects and has obvious treatment effect on larger and more malignant TNBC.

Drawings

FIG. 1 is a schematic diagram illustrating the synthesis of bismuth-manganese-based composite particles according to examples 1 to 3.

FIG. 2 is Bi-Bi₂O₃-MnO_x/NH₂Transmission electron micrograph (D).

FIG. 3 shows Bi-Bi₂O₃-MnO_x/NH₂A tunnel scanning electron microscopy image and a line scanning elemental analysis image.

FIG. 4 shows Bi-Bi₂O₃-MnO_x/NH₂XPS spectra of (a).

FIG. 5 shows CM and Bi-Bi₂O₃-MnO_x/NH₂-ICG and Bi-Bi₂O₃-MnO_x/NH₂-results of western blot testing of ICG @ CM.

FIG. 6 shows an ICG solution, Bi-Bi₂O₃-MnO_x/NH₂Solution of Bi-Bi₂O₃-MnO_x/NH₂-ICG solution and Bi-Bi₂O₃-MnO_x/NH₂-absorption spectrum of ICG @ CM solution.

FIG. 7 shows Bi-Bi₂O₃-MnO_x/NH₂-graph of catalytic oxygen evolution performance test results for ICG @ CM.

FIG. 8 shows Bi-Bi₂O₃-MnO_x/NH₂-graph of photodynamic therapy performance test results for ICG @ CM.

FIG. 9 shows Bi-Bi₂O₃-MnO_x/NH₂-chart of photothermal therapy performance test results for ICG @ CM.

FIG. 10 shows Bi-Bi₂O₃-MnO_x/NH₂-graph of results of chemokinetic therapeutic performance tests of ICG @ CM.

FIG. 11 shows Bi-Bi₂O₃-MnO_x/NH₂-graph of targeting test results for ICG @ CM.

FIG. 12 shows Bi-Bi₂O₃-MnO_x/NH₂-graph of biocompatibility test results for ICG @ CM.

FIG. 13 shows Bi-Bi₂O₃-MnO_x/NH₂-graph of results of a multiple therapy ability test of ICG @ CM.

Figure 14 is a graph of the results of the test of the therapeutic effect of different materials on larger, more malignant TNBC mice.

Detailed Description

The invention will be further explained and illustrated with reference to specific examples.

Example 1:

a bismuth-manganese-based composite particle is prepared by the following steps (the synthetic schematic diagram is shown in figure 1):

1) 0.9701g of Bi (NO) are added under the protection of argon₃)₃·5H₂Adding O into 10mL of dodecanethiol, stirring for 10min, placing in an oil bath, heating until the reaction liquid turns black, keeping the temperature for 1min, washing the reaction liquid with cyclohexane and ethanol, centrifuging at 10000rpm for 10min, and taking the precipitate to obtain bismuth nanoparticles (marked as Bi NPs);

2) adding 50mg of bismuth nanoparticles into 50mL of cyclohexane solution, performing ultrasonic dispersion to obtain bismuth nanoparticle dispersion liquid, adding 10mL of emulsifier legal-CO-520 (alkylphenol ethoxylate) into 50mL of cyclohexane solution, uniformly mixing to obtain emulsifier solution, adding the bismuth nanoparticle dispersion liquid into the emulsifier solution, uniformly mixing, and dropwise adding 5mL of KMnO with the concentration of 5mg/mL₄The solution is dripped for 15min, stirred for 10h after dripping, then the reaction solution is washed by ethanol, centrifuged, and the precipitate is taken to obtain the bismuth-bismuth trioxide-manganese oxide composite nano-particles (marked as Bi-Bi)₂O₃-MnO_x)；

3) Dispersing 50mg of bismuth-bismuth trioxide-manganese oxide composite nanoparticles into 25mL of absolute ethanol, adding 1mL of aminopropyltriethoxysilane, stirring at 120rpm at 50 ℃ in the absence of light for 48h, centrifuging, collecting precipitate, and obtaining amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nanoparticles (marked as Bi-Bi)₂O₃-MnO_x/NH₂)；

4) Dispersing 10mg of amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nanoparticles into 10mL of ultrapure water, adding 6mg of indocyanine green, stirring at 4 ℃ in the dark for 4 hours, centrifuging, and taking the precipitate to obtain bismuth-bismuth trioxide-manganese oxide composite nanoparticles (marked as Bi-Bi) loaded with indocyanine green₂O₃-MnO_x/NH₂-ICG)；

5) Extracting a TNBC cell membrane by using a cell membrane extraction kit to obtain a TNBC cell membrane dispersion solution, dispersing 1mg of bismuth-bismuth trioxide-manganese oxide composite nano particles loaded with indocyanine green into 1mL of sterilized Phosphate Buffered Saline (PBS), adding 200 mu L of the TNBC cell membrane dispersion solution (containing 0.2mg of the TNBC cell membrane), ultrasonically dispersing for 5min at 4 ℃, stirring for 4h in a dark place, centrifuging, taking a precipitate, and obtaining the bismuth-manganese-based composite particles (marked as Bi-Bi)₂O₃-MnO_x/NH₂-ICG@CM)。

And (3) performance testing:

1)Bi-Bi₂O₃-MnO_x/NH₂the transmission electron micrograph (see FIG. 2), the tunneling scanning electron micrograph (left) and the line scanning elemental analysis micrograph (right) are shown in FIG. 3, and the XPS spectrum (specifically: Bi-Bi is used)₂O₃-MnO_x/NH₂The sample was dropped onto a silicon wafer and analyzed for valence states of Bi and Mn by Thermo Fisher XPS) as shown in FIG. 4 (the left graph is a 4f orbital XPS spectrum of Bi, and the right graph is a 2p orbital XPS spectrum of Mn), CM, Bi-Bi₂O₃-MnO_x/NH₂-ICG and Bi-Bi₂O₃-MnO_x/NH₂The results of the Western blot (immunoblot assay) of-ICG @ CM are shown in FIG. 5.

As can be seen from fig. 2: Bi-Bi of hollow structure₂O₃-MnO_x/NH₂The aggregates are aggregated together, the formed aggregates have a plurality of holes and are in a honeycomb shape, and the particle size of the aggregates is about 366 nm.

As can be seen from fig. 3: the existence and distribution of Bi, Mn and O elements are confirmed.

As can be seen from fig. 4: form Bi element with 3 valence as main part and Mn element with 2 valence and 4 valence as main part with 3 valence, which shows Bi-Bi₂O₃-MnO_x/NH₂The diversity of the valence states of the medium Mn element.

As can be seen from fig. 5: Bi-Bi₂O₃-MnO_x/NH₂-ICG @ CM with CM coating Bi-Bi₂O₃-MnO_x/NH₂Structure of-ICG。

2) Preparing ICG solution and Bi-Bi₂O₃-MnO_x/NH₂Solution of Bi-Bi₂O₃-MnO_x/NH₂-ICG solution and Bi-Bi₂O₃-MnO_x/NH₂The absorbance spectra (690nm to 810nm) of the 4 solutions were measured using a multifunctional microplate reader at 100. mu.L each of the ICG @ CM solutions, and the test results are shown in FIG. 6.

As can be seen from fig. 6: ICG successful load, Bi-Bi₂O₃-MnO_x/NH₂-ICG and Bi-Bi₂O₃-MnO_x/NH₂-ICG @ CM was successfully prepared. The following can be calculated through absorption spectrum: Bi-Bi₂O₃-MnO_x/NH₂The loading of ICG in ICG @ CM was up to 50.6%.

3) For Bi-Bi₂O₃-MnO_x/NH₂-ICG @ CM, the specific test steps are as follows:

a) determination of catalytic oxygen generation performance: the content change of oxygen in the reaction solution is monitored in real time by a portable dissolved oxygen instrument, and the Bi-Bi with different concentrations (0 mug/mL, 12.5 mug/mL and 25 mug/mL) of 4mL is adopted₂O₃-MnO_x/NH ₂2 mu L of hydrogen peroxide solution with the mass fraction of 30% is respectively added into the-ICG @ CM solution, the change of the time and the oxygen content in the reaction liquid is dynamically detected in real time, and the test result is shown in figure 7.

As can be seen from fig. 7: Bi-Bi₂O₃-MnO_x/NH₂the-ICG @ CM has strong catalytic oxygen production capacity, and shows that the material can improve the hypoxic environment of tumors.

b) Determination of photodynamic therapeutic properties: in 3 parts of 200. mu.L each of Bi-Bi₂O₃-MnO_x/NH₂Adding 2 μ L of DPBF solution (concentration 10mM, solvent is dimethyl sulfoxide; 1, 3-diphenyl isobenzofuran (DPBF) can be oxidized by singlet oxygen, DPBF is degraded, the color is gradually changed from yellow green, the wavelength is 420nm, the absorption spectrum is changed), and the No. 1 part is not processed, and the No. 2 part is processed by the solution with wavelength 808nm and power density of 0.8W/cm²The 3 rd portion was irradiated with 2mM hydrogen peroxide at a wavelength of 808nm and a power density of 0.8W/cm²The laser is used for irradiating, the change of the monitoring time and the absorbance of the light with the wavelength of 420nm is dynamically detected in real time by a microplate reader, and the test result is shown in figure 8.

As can be seen from fig. 8: Bi-Bi₂O₃-MnO_x/NH₂Under the condition of laser irradiation, DPBF is obviously degraded, and under the condition of hydrogen peroxide and laser irradiation, the degradation is more obvious, which shows that the material has the capacity of photodynamic therapy, and simultaneously, the effect of photodynamic therapy can be obviously improved by catalyzing oxygen production.

c) Determination of photothermal therapeutic properties: 400 μ L of Bi-Bi at different concentrations (125 μ g/mL, 250 μ g/mL, and 500 μ g/mL)₂O₃-MnO_x/NH₂Adding the-ICG @ CM solution into independent single wells of a 48-well cell culture plate respectively, and using the wavelength of 808nm and the power density of 0.8W/CM²The temperature of the solution was monitored in real time by a thermometer as a function of the irradiation time, and the results of the test are shown in FIG. 9.

As can be seen from fig. 9: Bi-Bi₂O₃-MnO_x/NH₂The temperature of the ICG @ CM is increased under the condition of laser irradiation, and the higher the concentration is, the more the temperature is increased, and the material has the capacity of photothermal therapy.

d) Determination of the chemokinetic therapeutic properties: 6 groups of 500. mu.L Bi-Bi were prepared₂O₃-MnO_x/NH₂-ICG @ CM solution (500. mu.g/mL), GSH of different masses was added to the solution in order of 0mM, 0.5mM, 1mM, 2mM, 4mM and 8mM, and NaHCO was added to the solution in 25mM concentration₃Per 5% CO₂Buffering the solution until the total volume of the solution is 1mL, shaking at 37 deg.C for 15min, centrifuging, collecting the supernatant, respectively labeled A, B, C, D, E and F, and adding 800 μ L of the supernatant to the solution containing H₂O₂(100. mu.L, 80mM) of MB (100. mu.L, 100. mu.g/mL; Methylene Blue (MB) can be oxidized by hydroxyl radicals, MB is degraded, the blue solution gradually fades at 665nm, the absorption spectrum changes), incubating at 37 ℃ for 15min, and taking 1mL of MB solution (10. mu.g/mL) as a controlThe absorbance at 665nm of each solution was measured by a microplate reader, and the measurement results are shown in FIG. 10 (in the figure, the degradation picture of methylene blue).

As can be seen from fig. 10: Bi-Bi₂O₃-MnO_x/NH₂-ICG @ CM at GSH and H₂O₂Under the existing condition, Fenton-like reaction can occur to generate hydroxyl free radicals, and the effect is more obvious when the concentration of GSH is 2mM, which shows that the material has the capacity of chemodynamic treatment.

4)Bi-Bi₂O₃-MnO_x/NH₂-targeting test of ICG @ CM: Bi-Bi with a concentration of 50 mug/mL₂O₃-MnO_x/NH₂-ICG solution and Bi₂O₃-MnO_x/NH₂After co-incubation of the ICG @ CM solution with 4T1 cells and L929 cells for 4h, respectively, the residual material was washed away with PBS, and then each group of cells was collected for flow cytometry to obtain the mean fluorescence intensity map (a and b in the figure) and the flow cell histogram (c in the figure), wherein the untreated group was the control group, and the test results are shown in fig. 11.

As can be seen from fig. 11: cell membrane-coated Bi-Bi₂O₃-MnO_x/NH₂The ICG @ CM has good targeting effect on the three-negative breast cancer 4T1 cells, almost has no targeting effect on normal cells L929, and has no Bi-Bi of cell membranes₂O₃-MnO_x/NH₂Lack of targeting ability of ICG for both cells, suggesting Bi-Bi₂O₃-MnO_x/NH₂the-ICG @ CM has good targeting function on TNBC.

5)Bi-Bi₂O₃-MnO_x/NH₂-biocompatibility testing of ICG @ CM: gradient concentration of Bi-Bi₂O₃-MnO_x/NH₂-ICG solution and Bi-Bi₂O₃-MnO_x/NH₂ICG @ CM solutions (concentrations of 500. mu.g/mL, 250. mu.g/mL, 125. mu.g/mL, 62.5. mu.g/mL, and 0. mu.g/mL, respectively) were co-cultured with 4T1 cells and L929 cells (both purchased from Shanghai cell Bank of Chinese academy), respectively, for 24h, and the activity of the cells was assayed by the CCK-8 method, as shown in FIG. 12.

As can be seen from fig. 12: even at a concentration of 500. mu.g/mL of Bi-Bi₂O₃-MnO_x/NH₂The activity of L929 cells, namely non-cancer normal cells in the-ICG @ CM solution is more than 80 percent, and meanwhile, the activity of Bi-Bi is₂O₃-MnO_x/NH₂The activity of L929 in the-ICG @ CM solution is slightly larger than that of Bi-Bi₂O₃-MnO_x/NH₂The activity of L929 in ICG solution proves that the coating of cell membrane is favorable for improving the safety of materials in normal cells, and the Bi-Bi prepared by the method is illustrated₂O₃-MnO_x/NH₂ICG @ CM has good biocompatibility, but for 4T1 cells an abnormal microenvironment (high GSH, high H) due to cancer cells₂O₂) Targeting Bi-Bi₂O₃-MnO_x/NH₂ICG @ CM is toxic, indicating that the material can be used to kill cancer cells.

6)Bi-Bi₂O₃-MnO_x/NH₂-multiplex capacity test for ICG @ CM: Bi-Bi with a concentration of 50 mug/mL₂O₃-MnO_x/NH₂-ICG solution and Bi-Bi₂O₃-MnO_x/NH₂-ICG @ CM solution and ICG solution with concentration of 25. mu.g/mL were co-cultured with 4T1 cells for 4h, and after washing away residual material with PBS, with or without 808nm wavelength and 0.8W/CM power density²The laser of (1) was irradiated for 10min, and after the irradiation, the cells were cultured for 4 hours, and the activity of the cells was measured by the CCK-8 method, wherein the group without any material was designated as a control group, and the test results are shown in FIG. 13.

As can be seen from fig. 13: Bi-Bi₂O₃-MnO_x/NH₂The ICG @ CM has obvious killing effect on TNBC cells 4T1 under the condition of light or no light, and shows that Bi-Bi is used for killing the TNBC cells₂O₃-MnO_x/NH₂The multiple treatment triggered by-ICG @ CM has excellent killing capacity on TNBC cells.

7) Construction of mouse model of TNBC and treatment thereof:

4T1 cells were injected subcutaneously into the axilla of 30 female BALB/c mice (purchased at the Experimental center for medical animals in Guangdong province) for 6-8 weeksMiddle (1X 10 per mouse)⁷Individual cells) was modeled when tumor volume reached 300mm, which was larger and more malignant³～500mm³Meanwhile, tumor-bearing mice are randomly divided into 5 groups, different treatments are respectively carried out, and the mice in each group are subjected to the following operations:

1) control group: tail vein injection of 100 μ L PBS;

2) ICG + Laser group: tail vein injection of 100 μ L ICG solution (0.5mg/mL) at 808nm power density of 0.8W/cm after 4h²Irradiating for 10min by the laser;

3)Bi-Bi₂O₃-MnO_x/NH₂-ICG + Laser group: tail vein injection of 100 mu L Bi-Bi₂O₃-MnO_x/NH₂ICG solution (1mg/mL), at a wavelength of 808nm after 4h, at a power density of 0.8W/cm²Irradiating for 10min by the laser;

4)Bi-Bi₂O₃-MnO_x/NH₂-ICG @ CM group: tail vein injection of 100 mu L Bi-Bi₂O₃-MnO_x/NH₂-ICG @ CM solution (1 mg/mL);

5)Bi-Bi₂O₃-MnO_x/NH₂-ICG @ CM + Laser group: tail vein injection of 100 mu L Bi-Bi₂O₃-MnO_x/NH₂-ICG @ CM solution (1mg/mL) at 808nm after 4h with a power density of 0.8W/CM²Irradiating for 10min by the laser;

tumor sizes were measured every two days and the relative sizes of the tumors were calculated, and the results are shown in fig. 14.

As can be seen from fig. 14: Bi-Bi₂O₃-MnO_x/NH₂-ICG @ CM group and Bi-Bi₂O₃-MnO_x/NH₂the-ICG @ CM + Laser group can obviously inhibit or even reduce the growth of larger and more malignant TNBC, which indicates that Bi-Bi₂O₃-MnO_x/NH₂ICG @ CM has superior therapeutic potential for larger, more malignant TNBC.

Example 2:

2) adding 50mg of bismuth nanoparticles into 50mL of cyclohexane solution, performing ultrasonic dispersion to obtain bismuth nanoparticle dispersion liquid, adding 10mL of emulsifier legal-CO-520 (alkylphenol ethoxylate) into 50mL of cyclohexane solution, uniformly mixing to obtain emulsifier solution, adding the bismuth nanoparticle dispersion liquid into the emulsifier solution, uniformly mixing, and dropwise adding 5mL of KMnO with the concentration of 10mg/mL₄The dripping time of the solution is 15min, the solution is stirred for 8h after the dripping is finished, the reaction solution is washed by ethanol, the solution is centrifuged, and the precipitate is taken to obtain the bismuth-bismuth trioxide-manganese oxide composite nano-particles (marked as Bi-Bi)₂O₃-MnO_x)；

4) Dispersing 10mg of amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nanoparticles into 10mL of ultrapure water, adding 5mg of indocyanine green, stirring at 4 ℃ in the dark for 4h, centrifuging, and collecting precipitate to obtain bismuth-bismuth trioxide-manganese oxide composite nanoparticles (marked as Bi-Bi) loaded with indocyanine green₂O₃-MnO_x/NH₂-ICG)；

5) Extracting TNBC cell membrane with cell membrane extraction kit to obtain TNBC cell membrane dispersion, dispersing 1mg of indocyanine green-loaded bismuth-bismuth trioxide-manganese oxide composite nanoparticles in 1mL of sterilized Phosphate Buffered Saline (PBS), and adding 200 μ L of TNBC cellUltrasonically dispersing the membrane dispersion solution (containing TNBC cell membrane 0.5mg) at 4 deg.C for 5min, stirring in dark for 4 hr, centrifuging, and collecting precipitate to obtain bismuth-manganese based composite particles (marked as Bi-Bi)₂O₃-MnO_x/NH₂-ICG@CM)。

Upon testing, the Bi-Bi prepared in this example₂O₃-MnO_x/NH₂Properties of-ICG @ CM Bi-Bi prepared in example 1₂O₃-MnO_x/NH₂-ICG @ CM is very close.

Example 3:

1) under the protection of argon, 0.315g of BiCl₃Adding the bismuth nanoparticles into 10mL of dodecanethiol, stirring for 10min, placing the mixture in an oil bath, heating until the reaction solution turns black, preserving the heat for 1min, washing the reaction solution with cyclohexane and ethanol, centrifuging the reaction solution at 10000rpm for 10min, and taking precipitates to obtain bismuth nanoparticles (marked as Bi NPs);

2) adding 50mg of bismuth nanoparticles into 50mL of cyclohexane solution, performing ultrasonic dispersion to obtain bismuth nanoparticle dispersion liquid, adding 10mL of emulsifier legal-CO-520 (alkylphenol ethoxylate) into 50mL of cyclohexane solution, uniformly mixing to obtain emulsifier solution, adding the bismuth nanoparticle dispersion liquid into the emulsifier solution, uniformly mixing, and dropwise adding 5mL of KMnO with the concentration of 6mg/mL₄The solution is dripped for 20min, stirred for 10h after dripping, then the reaction solution is washed by ethanol, centrifuged, and the precipitate is taken to obtain the bismuth-bismuth trioxide-manganese oxide composite nano-particles (marked as Bi-Bi)₂O₃-MnO_x)；

3) Dispersing 50mg of bismuth-bismuth trioxide-manganese oxide composite nanoparticles into 25mL of absolute ethanol, adding 1mL of aminopropyltriethoxysilane, stirring at 120rpm at 50 ℃ in the absence of light for 48h, centrifuging, collecting precipitate, and obtaining amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nanoparticles (marked as Bi)₂O₃-MnO_x/NH₂)；

4) 10mg of amino-modified hollow structuresDispersing the bismuth-bismuth trioxide-manganese oxide composite nanoparticles in 10mL of ultrapure water, adding 10mg of indocyanine green, stirring at 4 ℃ in the dark for 4 hours, centrifuging, and taking the precipitate to obtain the bismuth-bismuth trioxide-manganese oxide composite nanoparticles (marked as Bi-Bi) loaded with the indocyanine green₂O₃-MnO_x/NH₂-ICG)；

5) Extracting a TNBC cell membrane by using a cell membrane extraction kit to obtain a TNBC cell membrane dispersion solution, dispersing 1mg of bismuth-bismuth trioxide-manganese oxide composite nano particles loaded with indocyanine green into 1mL of sterilized Phosphate Buffered Saline (PBS), adding 200 mu L of the TNBC cell membrane dispersion solution (containing 0.4mg of the TNBC cell membrane), ultrasonically dispersing for 5min at 4 ℃, stirring for 4h in a dark place, centrifuging, taking a precipitate, and obtaining the bismuth-manganese-based composite particles (marked as Bi-Bi)₂O₃-MnO_x/NH₂-ICG@CM)。

The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.

Claims

1. The bismuth-manganese-based composite particle is characterized by comprising an inner core and a triple negative breast cancer cell membrane wrapping the inner core; the core is a hollow structure bismuth-bismuth trioxide-manganese oxide composite nanoparticle Bi-Bi with a plurality of amino groups for modification₂O₃-MnO_x/NH₂Formed by agglomeration of MnO_xThe valence of the medium Mn is 2 valence, 3 valence and 4 valence; indocyanine green is loaded inside and on the surface of the amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nano-particles.

2. The bismuth-manganese-based composite particles of claim 1, wherein: the loading amount of indocyanine green in the bismuth-manganese-based composite particles is 45-55%.

3. The bismuth-manganese-based composite particles of claim 1 or 2, wherein: the particle size of the bismuth-manganese-based composite particles is 300 nm-400 nm; the particle size of the amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nano-particles is 30-80 nm.

4. A method for preparing bismuth-manganese-based composite particles according to any one of claims 1 to 3, comprising the steps of:

5. The method of preparing bismuth-manganese based composite particles according to claim 4, comprising the steps of:

6. The method of producing bismuth-manganese-based composite particles according to claim 4 or 5, characterized in that: the bismuth salt in the step 1) is at least one of bismuth nitrate and bismuth chloride.

7. The method of producing bismuth-manganese-based composite particles according to claim 4 or 5, characterized in that: and in the step 2), the permanganate is at least one of potassium permanganate and sodium permanganate.

8. The method of producing bismuth-manganese-based composite particles according to claim 4 or 5, characterized in that: the mass ratio of the bismuth nanoparticles to the permanganate in the step 2) is 1: (0.5-2).

9. The method of producing bismuth-manganese-based composite particles according to claim 4 or 5, characterized in that: the mass ratio of the indocyanine green to the amino-modified hollow-structure bismuth-bismuth trioxide-manganese oxide composite nano-particles in the step 4) is 1: (0.5-2).

10. Use of the bismuth-manganese-based composite particles according to any one of claims 1 to 3 in the preparation of a medicament for treating triple negative breast cancer and a diagnostic agent for triple negative breast cancer.