A BiOCl@yeast ultrasound biohybrid material and its preparation method and application

Abstract

The present invention discloses a BiOCl@yeast ultrasonic biohybrid material and its preparation method and application. The BiOCl@yeast biohybrid material provided by the present invention is an ultrasonic catalytic biohybrid material. The present invention has developed for the first time a localized ultrasonic catalytic gas therapy platform that uses live yeast to actively target tumor areas and in situ fermentation to produce carbon dioxide. The BiOCl@yeast biohybrid material provided by the present invention can actively target tumor sites. On the one hand, it uses yeast targeted enrichment materials to improve ultrasonic catalytic performance. On the other hand, it uses yeast fermentation metabolites to increase the original high carbon dioxide level in the tumor microenvironment, thereby achieving the function of increasing the carbon monoxide concentration in tumor tissue under ultrasound, seriously affecting the respiratory metabolism of cancer cells, and reducing the carbon monoxide concentration in nearby normal tissues to a safe value as much as possible. It not only overcomes the problem of low targeting of nanomaterials in the body, but also makes this ultrasonic hybrid material have a very excellent specific tumor killing effect.

Description

Technical Field

[0001] The present invention relates to the technical field of biohybrid materials, and in particular to a BiOCl@yeast ultrasonic biohybrid material and a preparation method and application thereof. Background Art

[0002] Tumor tissue is the product of abnormal tissue proliferation caused by the loss of normal growth regulation by localized cells under the influence of various carcinogenic factors. Cancer cells' unlimited replication potential increases their energy requirements compared to normal cells, triggering abnormal proliferation and creating a tumor microenvironment characterized by hypoxia, elevated carbon dioxide concentrations, and a weak acidity compared to normal tissue.

[0003] Carbon monoxide (CO) is an odorless and colorless gas. Its affinity for hemoglobin is 200 times greater than that of oxygen. It can competitively replace oxygen (O2) and bind to hemoglobin to form carboxyhemoglobin (COHb), which can easily cause symptoms of carbon monoxide poisoning in humans. This greatly hinders the transport of O2 throughout the body. Therefore, people have long been cautious about its use and avoid it. At the cellular level, CO affects mitochondria by interfering with cytochrome c oxidase, blocking oxidative phosphorylation (OXPHOS), causing respiratory abnormalities, severely reducing energy (adenosine triphosphate, ATP) production, and depleting ATP, promoting cell apoptosis (WEGIEL B, GALLO D, CSIZMADIA E, et al. Carbon Monoxide Expedites Metabolic Exhaustion to Inhibit Tumor Growth [J]. Cancer Research, 2013, 73(23): 7009-21.). However, the effect of CO is highly dependent on its concentration. Low concentrations promote cell growth, while high concentrations can induce cell apoptosis. In addition, given that CO can freely penetrate the tumor interstitium, the CO production site must be firmly controlled at the tumor site in order to achieve the desired effect of increasing the local CO concentration in the tumor and reducing the concentration in the blood ([1] MOTTERLINI R, OTTERBEIN LE. The therapeutic potential of carbon monoxide [J]. Nat Rev Drug Discov, 2010, 9(9): 728-43. [2] ZHOU Y, YU W, CAO J, et al. Harnessing carbon monoxide-releasing platforms for cancer therapy [J]. Biomaterials, 2020, 255: 120193.).

[0004] The current clinical methods for cancer treatment (including surgical resection, radiotherapy, chemotherapy, etc.) have long cycles and large side effects, which not only affect the effect of tumor treatment, but also have a great impact on normal tissues and the normal life of patients. Ultrasound dynamic therapy (SDT) utilizes the cavitation effect of ultrasound (US) and its high penetration and spatiotemporal controllability of tissues. With the help of sonosensitizers that are activated by US and can greatly aggravate the cavitation effect, it can achieve targeted ablation of tumor tissue (SON S, KIM JH, WANGX, et al. Multifunctional sonosensitizers in sonodynamic cancer therapy [J]. ChemSoc Rev, 2020, 49 (11): 3244-61.). However, nanomaterials are limited to the process of targeting tumors and currently still rely heavily on the high permeability and long retention effect (EPR effect) or certain characteristics of the tumor microenvironment. It is difficult to achieve high concentration aggregation at the tumor site, which greatly affects the actual clinical treatment effect. Summary of the Invention

[0005] In view of the above-mentioned deficiencies in the prior art, the present invention provides a BiOCl@yeast ultrasound biohybrid material and a preparation method and application thereof.

[0006] The present invention discloses a BiOCl@yeast ultrasound biohybrid material, which utilizes yeast to carry nanomaterials to achieve material enrichment at the tumor site. In the tumor microenvironment and the high carbon dioxide concentration environment achieved after yeast fermentation, the nanomaterials convert carbon dioxide into carbon monoxide under US induction, which has good application potential in tumor treatment.

[0007] A method for preparing a BiOCl@yeast ultrasound biohybrid material comprises the following steps:

[0008] S1: Sodium chloride and bismuth salt are dissolved in a solvent, and then a solvothermal reaction is performed to obtain BiOCl;

[0009] S2: Modify the BiOCl obtained in step S1 with polyethyleneimine to obtain BiOCl-PEI;

[0010] S3: Modifying the BiOCl-PEI obtained in step S2 with bovine serum albumin to obtain BiOCl-PEI-BSA;

[0011] S4: The BiOCl-PEI-BSA obtained in step S3 is mixed with yeast to obtain the BiOCl@yeast ultrasound biohybrid material.

[0012] After modification with polyethyleneimine (PEI) to correct the positive charge, subsequent cell experiments found that the material had no obvious cell killing effect. Modification with BSA increased the material's phagocytosis rate and improved the material's killing performance.

[0013] Preferably, the bismuth salt is at least one of the following: bismuth chloride, bismuth nitrate, and bismuth citrate.

[0014] Preferably, in step S1, the molar ratio of sodium chloride to bismuth salt is 1:1-2.

[0015] Preferably, in step S1, the temperature of the solvent thermal reaction is 160-180° C., and the time is 12-20 h.

[0016] Preferably, in step S1, the solvent is water or a mixture of water and ethylene glycol, the volume ratio of water in the solvent is 50% to 100%, and the volume ratio of ethylene glycol is 0 to 50%.

[0017] Preferably, in step S2, the molecular weight of polyethyleneimine is 1800 to 25000.

[0018] Preferably, in step S2, the mass ratio of polyethyleneimine to BiOCl is 0.5 to 1:1;

[0019] In step S3, the mass ratio of bovine serum albumin to BiOCl-PEI is 0.3-0.5:1.

[0020] Preferably, in step S4, the ratio of BiOCl-PEI-BSA to yeast is 10 6 ~10 7 A yeast.

[0021] The present invention further provides a BiOCl@yeast ultrasonic biohybrid material prepared by the preparation method.

[0022] The present invention also provides the use of the BiOCl@yeast ultrasound biohybrid material in preparing a sonodynamic therapy preparation for treating tumors.

[0023] The present invention has at least the following advantages:

[0024] The BiOCl@yeast biohybrid material provided by this invention is an ultrasonic catalytic biohybrid material. This invention is the first to develop a localized ultrasonic catalytic gas therapy platform that uses live yeast to actively target tumor areas and produce carbon dioxide through in situ fermentation.

[0025] 2. The BiOCl@yeast biohybrid material provided by this invention can actively target tumors. This not only utilizes yeast-targeted enrichment to enhance ultrasonic catalytic performance, but also utilizes yeast fermentation metabolites to elevate the already elevated carbon dioxide levels in the tumor microenvironment. This increases carbon monoxide concentrations in tumor tissue under ultrasound, significantly impacting cancer cell respiratory metabolism and minimizing carbon monoxide concentrations in nearby normal tissue to safe levels. This overcomes the inherent inability of nanomaterials to target tumors in vivo and also enables this ultrasonic hybrid material to exhibit exceptionally specific tumor-killing efficacy.

[0026] 3. The preparation method of the present invention has the advantages of simple process, low price, high repeatability, good stability, etc., and is suitable for large-scale production. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 These are electron micrographs of the BiOCl nanoparticles in Example 9, where (a) is a low-magnification scanning electron microscopy (SEM) image of BiOCl; (b) is a transmission electron microscopy (TEM) image, and the upper left corner is a high-angle dark-field scanning electron microscopy (HADDF-STEM) image.

[0028] Figure 2 This is the energy spectrum of Bi, Cl, and O elements in the BiOCl nanoparticles in Example 9.

[0029] Figure 3 This is the X-ray diffraction pattern of the BiOCl nanoparticles in Example 9.

[0030] Figure 4 1 are electron microscope images of the BiOCl nanoparticles in Examples 1 to 8, wherein (ah) are electron microscope images of the BiOCl nanoparticles in Examples 1 to 8, respectively.

[0031] Figure 5 These are electron micrographs of the ultrasonic biohybrid materials in Example 9, where (a) is the SEM image of yeast and (b) is the SEM image of BiOCl@yeast.

[0032] Figure 6 This is a diagram of the hydrodynamic radius of the ultrasonic biohybrid material in Example 9.

[0033] Figure 7 This is the electrokinetic potential diagram (Zeta potential diagram) of the ultrasonic biohybrid material and its components in Example 9.

[0034] Figure 8These are performance graphs showing the CO generation performance of the BiOCl nanoparticles prepared in Example 9 in Example 2 under ultrasonic conditions, where (a) shows the conversion of CO2 to CO by BiOCl nanoparticles within 5 minutes, (b) shows the conversion of CO2 to CO by the modified BiOCl-PEI-BSA nanoparticles within 5 minutes, and (c) shows the CO concentrations generated by BiOCl and BiOCl-PEI-BSA over ultrasonic time.

[0035] Figure 9 This is a cytotoxicity graph of BiOCl nanoparticles co-cultured with 4T1 cells for 24 hours in Example 9.

[0036] Figure 10 Graph showing the cell killing ability of BiOCl nanoparticles modified with PEI and BSA in Example 9 when co-cultured with 4T1 cells for 24 hours with or without ultrasound. DETAILED DESCRIPTION

[0037] The method for preparing the BiOCl@Yeast ultrasonic biohybrid material provided by the present invention comprises the following steps:

[0038] S1: Sodium chloride and bismuth salt are fully dissolved in water or a mixed solvent of water and ethylene glycol, and the solution is transferred to a solvothermal reactor for solvothermal reaction. After cooling to room temperature, the solution is centrifuged to obtain a BiOCl solution.

[0039] S2: Modifying the BiOCl nanosheets with polyethyleneimine (PEI) to obtain a BiOCl-PEI solution;

[0040] S3: Modifying the BiOCl-PEI material with bovine serum albumin (BSA) to obtain a BiOCl-PEI-BSA solution;

[0041] S4: The modified BiOCl-PEI-BSA material is added to the yeast-containing solution to obtain BiOCl@yeast biohybrid particles.

[0042] The present invention is described in detail below by way of examples. It should be understood that the following examples are only used to further illustrate the present invention and are not to be construed as limiting the scope of protection of the present invention. Some non-essential improvements and adjustments made by those skilled in the art based on the above content of the present invention all fall within the scope of protection of the present invention. The specific process parameters and the like in the following examples are only examples within a suitable range, and those skilled in the art can make selections within a suitable range through the description herein, and are not to be limited to the specific numerical values ​​exemplified below.

[0043] Example 1

[0044] A method for preparing BiOCl@Yeast biohybrid particles comprises:

[0045] Dissolve 2 mmol of bismuth nitrate pentahydrate and 1 mmol of sodium chloride in 30 ml of pure water and stir at 400 rpm for 30 minutes at room temperature. Transfer the mixed solution to a solvothermal reactor and set the reaction temperature at 160°C for 16 hours. After cooling to room temperature, the product was centrifuged at 12,000 rpm for 10 minutes and washed three times with ethanol and water to remove excess impurities, yielding a BiOCl solution, i.e., BiOCl nanoparticles.

[0046] A polyethyleneimine (PEI) solution with a relative molecular mass of approximately 25,000 and a final concentration of 1 mg / ml was added to a BiOCl solution with a final concentration of 1 mg / ml. The solution was stirred at room temperature for 3 h. The resulting solution was centrifuged at 12,000 rpm and thoroughly washed with water to obtain a BiOCl-PEI solution.

[0047] Then, a bovine serum albumin (BSA) solution with a final concentration of 0.5 mg / ml was added to the BiOCl-PEI solution with a final concentration of 1 mg / ml, and the solution was stirred at room temperature for 8 h. The resulting solution was centrifuged at 12000 rpm and washed thoroughly with water to obtain a BiOCl-PEI-BSA solution.

[0048] Take 3ml of 1mg / ml BiOCl-PEI-BSA solution and add it to 10ml containing 10 7 The product was centrifuged at 3000 rpm for 4 minutes and washed thoroughly with pure water. Ultrasonic dispersion was avoided. The resulting BiOCl@yeast ultrasonic biohybrid material was stored in pure water.

[0049] Example 2

[0050] Preparation of BiOCl@yeast biohybrid particles, comprising:

[0051] Dissolve 1 mmol of bismuth nitrate pentahydrate and 1 mmol of sodium chloride in 30 ml of pure water and stir at 400 rpm for 30 minutes at room temperature. Transfer the mixed solution to a solvothermal reactor and set the reaction temperature at 160°C for 16 hours. After cooling to room temperature, the product is centrifuged at 12,000 rpm for 10 minutes and washed three times with ethanol and water to remove excess impurities, yielding a BiOCl solution, i.e., BiOCl nanoparticles.

[0052] A polyethyleneimine (PEI) solution with a relative molecular mass of approximately 25,000 and a final concentration of 1 mg / ml was added to a BiOCl solution with a final concentration of 1 mg / ml. The solution was stirred at room temperature for 3 h. The resulting solution was centrifuged at 12,000 rpm and thoroughly washed with water to obtain a BiOCl-PEI solution.

[0053] Then, a bovine serum albumin (BSA) solution with a final concentration of 0.5 mg / ml was added to the BiOCl-PEI solution with a final concentration of 1 mg / ml, and the solution was stirred at room temperature for 8 h. The resulting solution was centrifuged at 12000 rpm and washed thoroughly with water to obtain a BiOCl-PEI-BSA solution.

[0054] Take 3ml of 1mg / ml BiOCl-PEI-BSA solution and add it to 10ml containing 10 7 The product was centrifuged at 3000 rpm for 4 minutes and washed thoroughly with pure water. Ultrasonic dispersion was avoided. The resulting BiOCl@yeast ultrasonic biohybrid material was stored in pure water.

[0055] Example 3

[0056] Preparation of BiOCl@yeast biohybrid particles, comprising:

[0057] Dissolve 1 mmol of bismuth nitrate pentahydrate and 1 mmol of sodium chloride in 15 ml of pure water and 15 ml of ethylene glycol. Stir at 400 rpm for 30 minutes at room temperature. Transfer the mixed solution to a solvothermal reactor and set the reaction temperature at 160°C for 16 hours. After cooling to room temperature, the product is centrifuged at 12,000 rpm for 10 minutes and washed three times with ethanol and water to remove excess impurities, yielding a BiOCl solution, i.e., BiOCl nanoparticles.

[0058] A polyethyleneimine (PEI) solution with a relative molecular mass of approximately 25,000 and a final concentration of 1 mg / ml was added to a BiOCl solution with a final concentration of 1 mg / ml. The solution was stirred at room temperature for 3 h. The resulting solution was centrifuged at 12,000 rpm and thoroughly washed with water to obtain a BiOCl-PEI solution.

[0059] Then, a bovine serum albumin (BSA) solution with a final concentration of 0.5 mg / ml was added to the BiOCl-PEI solution with a final concentration of 1 mg / ml, and the solution was stirred at room temperature for 8 h. The resulting solution was centrifuged at 12000 rpm and washed thoroughly with water to obtain a BiOCl-PEI-BSA solution.

[0060] Take 3ml of 1mg / ml BiOCl-PEI-BSA solution and add it to 10ml containing 10 7 The product was centrifuged at 3000 rpm for 4 minutes and washed thoroughly with pure water. Ultrasonic dispersion was avoided. The resulting BiOCl@yeast ultrasonic biohybrid material was stored in pure water.

[0061] Example 4

[0062] Preparation of BiOCl@yeast biohybrid particles, comprising:

[0063] Dissolve 1 mmol of bismuth nitrate pentahydrate and 1 mmol of sodium chloride in 20 ml of pure water and 10 ml of ethylene glycol. Stir at 400 rpm for 30 minutes at room temperature. Transfer the mixed solution to a solvothermal reactor and set the reaction temperature at 160°C for 16 hours. After cooling to room temperature, the product is centrifuged at 12,000 rpm for 10 minutes and washed three times with ethanol and water to remove excess impurities, yielding a BiOCl solution, i.e., BiOCl nanoparticles.

[0064] A polyethyleneimine (PEI) solution with a relative molecular mass of approximately 25,000 and a final concentration of 1 mg / ml was added to a BiOCl solution with a final concentration of 1 mg / ml. The solution was stirred at room temperature for 3 h. The resulting solution was centrifuged at 12,000 rpm and thoroughly washed with water to obtain a BiOCl-PEI solution.

[0065] Then, a bovine serum albumin (BSA) solution with a final concentration of 0.5 mg / ml was added to the BiOCl-PEI solution with a final concentration of 1 mg / ml, and the solution was stirred at room temperature for 8 h. The resulting solution was centrifuged at 12000 rpm and washed thoroughly with water to obtain a BiOCl-PEI-BSA solution.

[0066] Take 3ml of 1mg / ml BiOCl-PEI-BSA solution and add it to 10ml containing 10 7 The product was centrifuged at 3000 rpm for 4 minutes and washed thoroughly with pure water. Ultrasonic dispersion was avoided. The resulting BiOCl@yeast ultrasonic biohybrid material was stored in pure water.

[0067] Example 5

[0068] Preparation of BiOCl@yeast biohybrid particles, comprising:

[0069] Dissolve 1 mmol of bismuth nitrate pentahydrate and 1 mmol of sodium chloride in 10 ml of pure water and 20 ml of ethylene glycol. Stir at 400 rpm for 30 minutes at room temperature. Transfer the mixed solution to a solvothermal reactor and set the reaction temperature at 160°C for 16 hours. After cooling to room temperature, the product is centrifuged at 12,000 rpm for 10 minutes and washed three times with ethanol and water to remove excess impurities, yielding a BiOCl solution, i.e., BiOCl nanoparticles.

[0070] A polyethyleneimine (PEI) solution with a relative molecular mass of approximately 25,000 and a final concentration of 1 mg / ml was added to a BiOCl solution with a final concentration of 1 mg / ml. The solution was stirred at room temperature for 3 h. The resulting solution was centrifuged at 12,000 rpm and thoroughly washed with water to obtain a BiOCl-PEI solution.

[0071] Then, a bovine serum albumin (BSA) solution with a final concentration of 0.5 mg / ml was added to the BiOCl-PEI solution with a final concentration of 1 mg / ml, and the solution was stirred at room temperature for 8 h. The resulting solution was centrifuged at 12000 rpm and washed thoroughly with water to obtain a BiOCl-PEI-BSA solution.

[0072] Take 3ml of 1mg / ml BiOCl-PEI-BSA solution and add it to 10ml containing 10 7 The product was centrifuged at 3000 rpm for 4 minutes and washed thoroughly with pure water. Ultrasonic dispersion was avoided. The resulting BiOCl@yeast ultrasonic biohybrid material was stored in pure water.

[0073] Example 6

[0074] Preparation of BiOCl@yeast biohybrid particles, comprising:

[0075] Dissolve 1 mmol of bismuth nitrate pentahydrate and 1 mmol of sodium chloride in 15 ml of pure water and 15 ml of ethylene glycol. Stir at 400 rpm for 30 minutes at room temperature. Transfer the mixed solution to a solvothermal reactor and set the reaction temperature at 160°C for 20 hours. After cooling to room temperature, the product is centrifuged at 12,000 rpm for 10 minutes and washed three times with ethanol and water to remove excess impurities, yielding a BiOCl solution, i.e., BiOCl nanoparticles.

[0076] A polyethyleneimine (PEI) solution with a relative molecular mass of approximately 25,000 and a final concentration of 1 mg / ml was added to a BiOCl solution with a final concentration of 1 mg / ml. The solution was stirred at room temperature for 3 h. The resulting solution was centrifuged at 12,000 rpm and thoroughly washed with water to obtain a BiOCl-PEI solution.

[0077] Then, a bovine serum albumin (BSA) solution with a final concentration of 0.5 mg / ml was added to the BiOCl-PEI solution with a final concentration of 1 mg / ml, and the solution was stirred at room temperature for 8 h. The resulting solution was centrifuged at 12000 rpm and washed thoroughly with water to obtain a BiOCl-PEI-BSA solution.

[0078] Take 3ml of 1mg / ml BiOCl-PEI-BSA solution and add it to 10ml containing 10 7 The product was centrifuged at 3000 rpm for 4 minutes and washed thoroughly with pure water. Ultrasonic dispersion was avoided. The resulting BiOCl@yeast ultrasonic biohybrid material was stored in pure water.

[0079] Example 7

[0080] Preparation of BiOCl@yeast biohybrid particles, comprising:

[0081] Dissolve 1 mmol of bismuth nitrate pentahydrate and 1 mmol of sodium chloride in 15 ml of pure water and 15 ml of ethylene glycol. Stir at 400 rpm for 30 minutes at room temperature. Transfer the mixed solution to a solvothermal reactor and set the reaction temperature at 160°C for 12 hours. After cooling to room temperature, the product is centrifuged at 12,000 rpm for 10 minutes and washed three times with ethanol and water to remove excess impurities, yielding a BiOCl solution, i.e., BiOCl nanoparticles.

[0082] A polyethyleneimine (PEI) solution with a relative molecular mass of approximately 1800 and a final concentration of 0.5 mg / ml was added to a BiOCl solution with a final concentration of 1 mg / ml. The solution was stirred at room temperature for 12 h. The resulting solution was centrifuged at 12,000 rpm and thoroughly washed with water to obtain a BiOCl-PEI solution.

[0083] Then, a bovine serum albumin (BSA) solution with a final concentration of 0.5 mg / ml was added to the BiOCl-PEI solution with a final concentration of 1 mg / ml, and the solution was stirred at room temperature for 8 h. The resulting solution was centrifuged at 12000 rpm and washed thoroughly with water to obtain a BiOCl-PEI-BSA solution.

[0084] Take 3ml of 1mg / ml BiOCl-PEI-BSA solution and add it to 10ml containing 10 7 The product was centrifuged at 3000 rpm for 4 minutes and washed thoroughly with pure water. Ultrasonic dispersion was avoided. The resulting BiOCl@yeast ultrasonic biohybrid material was stored in pure water.

[0085] Example 8

[0086] Preparation of BiOCl@yeast biohybrid particles, comprising:

[0087] Dissolve 1 mmol of bismuth nitrate pentahydrate and 1 mmol of sodium chloride in 15 ml of pure water and 15 ml of ethylene glycol. Stir at 400 rpm for 30 minutes at room temperature. Transfer the mixed solution to a solvothermal reactor and set the reaction temperature at 180°C for 16 hours. After cooling to room temperature, the product is centrifuged at 12,000 rpm for 10 minutes and washed three times with ethanol and water to remove excess impurities, yielding a BiOCl solution, i.e., BiOCl nanoparticles.

[0088] A polyethyleneimine (PEI) solution with a relative molecular mass of approximately 1800 and a final concentration of 0.5 mg / ml was added to a BiOCl solution with a final concentration of 1 mg / ml. The solution was stirred at room temperature for 12 h. The resulting solution was centrifuged at 12,000 rpm and thoroughly washed with water to obtain a BiOCl-PEI solution.

[0089] Then, a bovine serum albumin (BSA) solution with a final concentration of 0.3 mg / ml was added to the BiOCl-PEI solution with a final concentration of 1 mg / ml, and the solution was stirred at room temperature for 8 h. The obtained solution was centrifuged at 12000 rpm and washed thoroughly with water to obtain a BiOCl-PEI-BSA solution.

[0090] Take 3ml of 1mg / ml BiOCl-PEI-BSA solution and add it to 10ml containing 10 7 The product was centrifuged at 3000 rpm for 4 minutes and washed thoroughly with pure water. Ultrasonic dispersion was avoided. The resulting BiOCl@yeast ultrasonic biohybrid material was stored in pure water.

[0091] Example 9

[0092] Preparation of BiOCl@yeast biohybrid particles, comprising:

[0093] Dissolve 1 mmol of bismuth nitrate pentahydrate and 1 mmol of sodium chloride in 15 ml of pure water and 15 ml of ethylene glycol. Stir at 400 rpm for 30 minutes at room temperature. Transfer the mixed solution to a solvothermal reactor and set the reaction temperature at 160°C for 16 hours. After cooling to room temperature, the product is centrifuged at 12,000 rpm for 10 minutes and washed three times with ethanol and water to remove excess impurities, yielding a BiOCl solution, i.e., BiOCl nanoparticles.

[0094] A polyethyleneimine (PEI) solution with a relative molecular mass of approximately 1800 and a final concentration of 0.5 mg / ml was added to a BiOCl solution with a final concentration of 1 mg / ml. The solution was stirred at room temperature for 12 h. The resulting solution was centrifuged at 12,000 rpm and thoroughly washed with water to obtain a BiOCl-PEI solution.

[0095] Then, a bovine serum albumin (BSA) solution with a final concentration of 0.3 mg / ml was added to the BiOCl-PEI solution with a final concentration of 1 mg / ml, and the solution was stirred at room temperature for 8 h. The obtained solution was centrifuged at 12000 rpm and washed thoroughly with water to obtain a BiOCl-PEI-BSA solution.

[0096] Take 2ml of 1mg / ml BiOCl-PEI-BSA solution and add it to 5ml containing 10 6 The product was centrifuged at 3000 rpm for 4 minutes and washed thoroughly with pure water. Ultrasonic dispersion was avoided. The resulting BiOCl@yeast ultrasonic biohybrid material was stored in pure water.

[0097] Example 10

[0098] The products prepared in the above examples were tested, and the results were as follows:

[0099] Figure 1 (a) and (b) are electron microscope images of BiOCl nanoparticles in Example 9, wherein (a) is a low-magnification SEM image of BiOCl, (b) is a TEM image of BiOCl, and the upper left corner of (b) is a HADDF-STEM image. Figure 1 It can be seen from the figure that BiOCl nanoparticles have a nanosheet morphology. From HADDF-STEM, it can be seen that the interatomic distance between BiOCl surfaces is , corresponding to the standard (001) crystal plane of BiOCl, and behaves as a direct bandgap semiconductor.

[0100] The BiOCl nanoparticles prepared in Example 9 were subjected to energy spectrum analysis, and the results were as follows: Figure 2 As shown. Figure 2Energy spectrum analysis shows that BiOCl nanoparticles are composed of Bi, Cl and O elements.

[0101] The BiOCl nanoparticles prepared in Example 9 were subjected to X-ray diffraction analysis, and the results were as follows: Figure 3 As shown. Figure 3 The X-ray diffraction pattern of the material can confirm that it is BiOCl.

[0102] Figure 4 Figures (a, b, and c) show electron micrographs of BiOCl nanoparticles, with Figures (a, h, and e) representing the BiOCl nanoparticles used in Examples 1-8, respectively. The materials shown in Figures (a) and (b) are oversized. The ratio of HO and EG, reaction time, and temperature affect the material morphology. As shown in Figures (c, d, and e), the material in Figure (c) exhibits a larger effective surface area, a relatively uniform size of approximately 200 nm, and a stable size.

[0103] Figure 5 The following are electron micrographs of the BiOCl@yeast biohybrid material prepared in Example 9 of the present invention, wherein (a) is a SEM image of yeast and (b) is a SEM image of BiOCl@yeast. Figure 5 It can be seen from the figure that the yeast surface was originally smooth. After BiOCl was adsorbed on the yeast to form BiOCl@yeast, the surface BiOCl was adsorbed on the yeast surface in clusters and easily fell off under ultrasound.

[0104] The hydrodynamic radius of the BiOCl@yeast biohybrid material prepared in Example 9 was tested. Figure 6 As shown. Figure 6 It can also be seen in Figure 3 that BiOCl is adsorbed on the yeast surface.

[0105] The BiOCl@yeast biohybrid material prepared in Example 9 and its components were tested for zeta potential. Figure 7 shown. Figure 7 Zeta potential characterization revealed changes in the surface potential of the nanomaterials before and after modification, further demonstrating that BiOCl and yeast were assembled into BiOCl@yeast ultrasonic biohybrid materials by surface electrostatic adsorption.

[0106] Example 11

[0107] Ultrasonic catalytic performance test of the material prepared in Example 9: monitoring the ability of the material to convert carbon dioxide (CO2) into carbon monoxide (CO) under ultrasonic conditions.

[0108] 4.2 μmol / L hemoglobin (HB) was bubbled in N2 for 15-30 minutes, and 10 mg of sulfite was added. 20 mL of the above solution was added with 20 μL of Gox solution and 200 μL of glucose solution, and then 25-30 mL of HB to obtain a reaction substrate solution, wherein the solvent was PBS buffer solution.

[0109] 100 μL of the material was added to the above reaction substrate solution, and the absorption curve of the solution was measured every 2 minutes using a UV-visible spectrophotometer under ultrasound.

[0110] from Figure 8 It can be seen that after the mixed solution is subjected to ultrasound, the ultraviolet characteristic peaks of oxygen and hemoglobin with a maximum absorption wavelength of 555nm decrease, and the ultraviolet characteristic peaks of carboxyhemoglobin with a maximum absorption wavelength of 538nm and 569nm increase. At the same time, the amount of carbon monoxide produced can be calculated based on the change amplitude of the characteristic peaks and the amount of hemoglobin in the substrate solution, as shown in Figure (c).

[0111] Example 12

[0112] This example illustrates the application of the material provided by the present invention in killing tumor cells by the killing effect at the cellular level. The material used is the material prepared in Example 9. The cells selected are 4T1 mouse breast cancer cells, and the data obtained are shown in the figure below. Figure 9 、 Figure 10 shown.

[0113] Figure 9 and Figure 10 The potential cytotoxicity and cell killing effects of BiOCl nanoparticles before and after modification were studied by standard methylthiazolyl tetrazolium (MTT) assay. After co-culture of BiOCl nanoparticles with 4T1 for 4 h, the cells were exposed to 1W / cm 2 After the ultrasound treatment, the co-culture was continued for 24 hours, and the proportion of living cells was marked by MTT staining. Figure 9 As can be seen from the figure, as the concentration of BiOCl increases, the activity of 4T1 cells is basically unaffected. Figure 10 As can be seen in the figure, after modification with polyethyleneimine (PEI), the 100μg / ml BiOCl material has almost no cell killing effect under ultrasound, while the killing performance of the material is improved after modification with BSA, indicating that the material can generate sufficient CO under ultrasound and has good tumor treatment performance.

Claims

1. A method for preparing BiOCl@yeast ultrasound biohybrid material, characterized in that: The following steps are involved: S1: Sodium chloride and bismuth salt are dissolved in a solvent, and then a solvothermal reaction is performed to obtain BiOCl; S2: Modify the BiOCl obtained in step S1 with polyethyleneimine to obtain BiOCl-PEI; S3: Modifying the BiOCl-PEI obtained in step S2 with bovine serum albumin to obtain BiOCl-PEI-BSA; S4: The BiOCl-PEI-BSA obtained in step S3 is mixed with yeast to obtain the BiOCl@yeast ultrasound biohybrid material.

2. The preparation method according to claim 1, characterized in that The bismuth salt is at least one of the following: bismuth chloride, bismuth nitrate, and bismuth citrate.

3. The preparation method according to claim 1, characterized in that In step S1, the molar ratio of sodium chloride to bismuth salt is 1:1-2.

4. The preparation method according to claim 1, characterized in that In step S1, the temperature of the solvent thermal reaction is 160-180° C., and the time is 12-20 hours.

5. The preparation method according to claim 1, characterized in that In step S1, the solvent is water or a mixture of water and ethylene glycol, the volume ratio of water in the solvent is 50% to 100%, and the volume ratio of ethylene glycol is 0 to 50%.

6. The preparation method according to claim 1, characterized in that In step S2, the molecular weight of polyethyleneimine is 1800 to 25000.

7. The preparation method according to claim 1, characterized in that In step S2, the mass ratio of polyethyleneimine to BiOCl is 0.5 to 1:1; In step S3, the mass ratio of bovine serum albumin to BiOCl-PEI is 0.3-0.5:

1.

8. The preparation method according to claim 1, characterized in that In step S4, the ratio of BiOCl-PEI-BSA to yeast is 10 6 ~10 7 A yeast.

9. The BiOCl@yeast ultrasound biohybrid material prepared by the preparation method according to any one of claims 1 to 8.

10. Use of the BiOCl@yeast ultrasound biohybrid material according to claim 9 in the preparation of a sonodynamic therapy preparation for treating tumors.

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