A nanoassembly and its preparation method and application

Abstract

The present invention relates to the field of biomedicine, and in particular to a nanoassembly, its preparation method, and application. The nanoassembly is composed of three components: an amphiphilic block polymer, an antibiotic, and a sonosensitizer. The antibiotic is a hydrophobic antibiotic, and the sonosensitizer is a porphyrin sonosensitizer. The amphiphilic block polymer has the following structural formula: #imgabs0#, where m is an integer from 40 to 120, and n is an integer from 50 to 100. The nanoassembly exhibits excellent sonodynamic properties, antibacterial properties, pH responsiveness, and magnetic resonance imaging capabilities, and has broad application prospects in the treatment of bacterial and fungal infections, bacterial biofilm infections, and magnetic resonance imaging.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine technology, and in particular to a nanoassembly, a preparation method and an application thereof. Background Art

[0002] Biofilms are membrane-like structures formed by microorganisms secreting extracellular polymeric matrices that adhere to the surface of a medium, encapsulating the microorganisms. When bacteria form biofilms, the shielding effect of the biofilm prevents them from being eliminated by the host immune system, reducing the antimicrobial activity of antibiotics and leading to a significant increase in bacterial resistance. Therefore, developing therapeutic strategies that can effectively disrupt biofilm structures, enhance the antimicrobial activity of antibiotics, and reduce bacterial resistance is of great significance.

[0003] In recent years, the rapid development of nanotechnology has provided new insights into the treatment of bacterial biofilm infections. Numerous nanomaterials have been developed to disrupt biofilm structures, enhance the penetration of antibiotics within the biofilm, and thereby improve their antimicrobial activity. However, existing nanomaterials still suffer from insufficient antimicrobial properties, limiting their application.

[0004] Sonodynamic therapy (SDT) is a novel non-invasive treatment approach based on the synergistic effects of sonosensitizers and ultrasound. It works by specifically generating reactive oxygen species (ROS) at the tumor site under the influence of ultrasound and sonosensitizers, thereby eliminating rapidly dividing tumor cells. Compared with photodynamic therapy (PDT), SDT offers deeper tissue penetration and a higher safety profile, and has been widely used in the field of tumor treatment. Studies have shown that ROS, as a strong oxidant, possess broad bactericidal properties, effectively eliminating bacteria, spores, viruses, and fungi (such as mold). It also has a significant killing effect on protozoa and their oocysts, and can destroy bacterial toxins and hepatitis B surface antigen.

[0005] Therefore, combining sonodynamic therapy (SDT) with nanotechnology to develop a nanomaterial with both excellent sonodynamic properties and antibacterial properties is an urgent problem to be solved. Summary of the Invention

[0006] The invention provides a nano assembly having excellent sonodynamic performance, antibacterial performance, pH responsiveness and nuclear magnetic resonance imaging capability.

[0007] The present invention also provides a method for preparing a nanoassembly, by which the nanoassembly having excellent sonodynamic performance, antibacterial performance, pH responsiveness and nuclear magnetic resonance imaging capability can be prepared.

[0008] The present invention also provides the use of the above-mentioned nanoassembly or the nanoassembly prepared by the preparation method of the above-mentioned nanoassembly in the preparation of products for treating bacterial and fungal infections or in the preparation of products for treating biofilm infections. The inventors' research shows that the nanoassembly is used in the antibacterial activity test of oxygen-resistant Klebsiella pneumoniae, Escherichia coli, Staphylococcus aureus and mold. Under the action of ultrasound, it releases sonosensitizers and antibiotics, kills bacteria through the sonodynamic properties of the sonosensitizer (releasing ROS) and the antibacterial properties of the antibiotics, and shows strong antibacterial activity. After sterilizing oxygen-resistant Klebsiella pneumoniae, the survival rate of oxygen-resistant Klebsiella pneumoniae is 8%, after sterilizing Escherichia coli, the survival rate of Escherichia coli is 9%, and after sterilizing Staphylococcus aureus, the survival rate of Staphylococcus aureus is 9%. After sterilization of the mold, the survival rate of the mold was 11%, indicating that the nanoassembly can effectively reduce bacterial resistance, kill bacteria and fungi, and has excellent antibacterial activity against both bacteria and fungi. Therefore, it can be used to prepare products for treating bacterial and / or fungal infections. The nanoassembly was used in a bacterial biofilm inhibition test. Under the action of ultrasound, it can effectively remove bacterial biofilms, destroy the structure of bacterial biofilms, release sonosensitizers and antibiotics, and kill bacteria through the sonodynamic properties of the sonosensitizer (releasing ROS) and the antibacterial properties of the antibiotics. Therefore, it can be used to prepare products for treating bacterial biofilm infections.

[0009] The present invention also provides the use of the aforementioned nanoassembly or a nanoassembly prepared by the aforementioned method for preparing the nanoassembly in the preparation of a magnetic resonance imaging contrast agent. The inventors' research has shown that, when used in magnetic resonance imaging experiments, the nanoassembly can respond to an acidic microenvironment, significantly improving the magnetic resonance signal-to-noise ratio (SNR) of T1-weighted magnetic resonance imaging, achieving signal enhancement. Therefore, the nanoassembly can be used to prepare a magnetic resonance imaging contrast agent.

[0010] The first aspect of the present invention provides a nanoassembly, wherein the nanoassembly is assembled from the following three components:

[0011] An amphiphilic block polymer, an antibiotic, and a sonosensitizer; wherein the amphiphilic block polymer has a structural formula as shown in Formula I, the antibiotic is a hydrophobic antibiotic, and the sonosensitizer is a porphyrin sonosensitizer;

[0012] Formula I

[0013] In formula I, m is an integer of 40-120, and n is an integer of 50-100.

[0014] The nanoassembly as described above has a particle size of 50 to 200 nm.

[0015] The nanoassembly as described above, wherein the mass ratio of the amphiphilic block polymer, the antibiotic and the sonosensitizer is (10-20): (1-3): (1-5).

[0016] The nanoassembly as described above, wherein the hydrophobic antibiotic comprises at least one of levofloxacin, tetracycline, chloramphenicol, ciprofloxacin, azithromycin, doxycycline, linezolid, and norfloxacin;

[0017] And / or, the porphyrin sonosensitizer includes 5,10,15,20-tetraphenyl-21H,23H-porphyrin manganese (II).

[0018] The second aspect of the present invention provides a method for preparing the nanoassembly, comprising the following steps:

[0019] dissolving the amphiphilic block polymer, the antibiotic and the sonosensitizer in an organic solvent respectively to prepare an amphiphilic block polymer mother solution, an antibiotic mother solution and a sonosensitizer mother solution;

[0020] The amphiphilic block polymer mother solution, the antibiotic mother solution and the sonosensitizer mother solution are mixed with a PBS solution and then subjected to stirring and dialysis treatment to obtain the nanoassembly.

[0021] The method for preparing the nanoassembly as described above, wherein the concentration of the amphiphilic block polymer in the amphiphilic block polymer mother solution is 14-18 mg / mL, the concentration of the antibiotic in the antibiotic mother solution is 1.5-3 mg / mL, and the concentration of the sonosensitizer in the sonosensitizer mother solution is 1.5-3 mg / mL;

[0022] The volume ratio of the amphiphilic block polymer mother solution, the antibiotic mother solution and the sonosensitizer mother solution is (1-2):1:1.

[0023] The method for preparing the nanoassembly as described above, wherein the stirring process is performed at a speed of 1000 to 1500 rpm / min, a temperature of 20 to 30°C, and a time of 5 to 10 min;

[0024] And / or, in the dialysis treatment, the molecular weight cut-off of the dialysis bag used is 3000 to 20000 Da, and the dialysis time is 8 to 12 h;

[0025] And / or, the organic solvent comprises dimethyl sulfoxide.

[0026] In the above-mentioned method for preparing the nanoassembly, the amphiphilic block polymer is obtained by a preparation method comprising the following steps:

[0027] 2-(azepan-1-yl)ethyl methacrylate, a macromolecular chain transfer agent PEG-CTA, and an initiator are dissolved in dimethyl sulfoxide to obtain a mixed solution;

[0028] The mixed solution is subjected to polymerization reaction at 50-80° C. for 6-12 hours to obtain the amphiphilic block polymer;

[0029] wherein the initiator comprises azobisisobutyronitrile;

[0030] The mass ratio of the 2-(azepan-1-yl)ethyl methacrylate, the macromolecular chain transfer agent PEG-CTA and the initiator is (50-150):1:(0.1-0.3).

[0031] The third aspect of the present invention provides a use of the nanoassembly or the nanoassembly prepared by the method for preparing the nanoassembly in preparing a product for treating bacterial and fungal infections or in preparing a product for treating bacterial biofilm infections.

[0032] The fourth aspect of the present invention provides a use of the nanoassembly or the nanoassembly prepared by the method for preparing the nanoassembly in preparing a magnetic resonance imaging contrast agent.

[0033] The solution of the present invention has at least the following effects:

[0034] The nanoassembly provided by the present invention has excellent sonodynamic performance and pH responsiveness. It generates reactive oxygen species (ROS) under the action of ultrasound and can respond to an acidic microenvironment to generate more ROS under the action of ultrasound. The nanoassembly has excellent sonodynamic performance and antibacterial properties. Under the action of ultrasound, it can effectively remove bacterial biofilms, destroy bacterial biofilm structures, release sonosensitizers and antibiotics, kill bacteria through the sonodynamic performance of the sonosensitizer (releasing ROS) and the antibacterial properties of the antibiotics, and can be used to prepare products for treating bacterial biofilm infections. Moreover, under the action of ultrasound, the nanoassembly releases sonosensitizers and antibiotics. The sonodynamic properties (releasing ROS) and the antibacterial properties of antibiotics can effectively kill levothroid-resistant Klebsiella pneumoniae, Escherichia coli, Staphylococcus aureus and mold, showing strong antibacterial activity, indicating that the nanoassembly can effectively reduce bacterial resistance and has excellent antibacterial activity against both bacteria and fungi. Therefore, it can be used to prepare products for the treatment of bacterial and fungal infections; the nanoassembly has excellent pH-responsive magnetic resonance imaging capabilities, which can respond to acidic microenvironments and significantly improve the magnetic resonance signal-to-noise ratio (SNR) of T1-weighted magnetic resonance imaging to achieve signal enhancement. Therefore, it can be used to prepare magnetic resonance imaging contrast agents and has broad application prospects in magnetic resonance imaging. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0036] Figure 1 is the H NMR spectrum of the amphiphilic block polymer in Example 1 of the present invention;

[0037] Figure 2 is the ultraviolet absorption spectrum of the nanoassembly in Example 1 of the present invention;

[0038] Figure 3 The particle size distribution diagram and transmission electron microscope (TEM) diagram of the nanoassembly in Example 1 of the present invention are shown in FIG. Figure 3 a is the particle size distribution diagram of the nanoassembly in Example 1, Figure 3 b is the TEM image of the nanoassembly in Example 1;

[0039] Figure 4 The ROS release curves of LEV+MnTCPP@PC7A solutions with different pH values ​​before and after ultrasound treatment are shown in Figure 2. Figure 4 a is the ROS release curve of LEV+MnTCPP@PC7A solution at pH=6.0 before and after ultrasound treatment. Figure 4 b is the ROS release curve of LEV+MnTCPP@PC7A solution at pH=7.4 before and after ultrasound treatment;

[0040] Figure 5 For antibacterial activity test, Figure 5 a is the antibacterial activity of the nanoassembly against levothroid-resistant Klebsiella pneumoniae. Figure 5 b is the antibacterial activity of the nanoassembly against Escherichia coli, Figure 5 c is the antibacterial activity of the nanoassembly against Staphylococcus aureus, Figure 5 d is the antibacterial activity of the nanoassembly against mold;

[0041] Figure 6 This is the crystal violet staining result in the biofilm inhibition test;

[0042] Figure 7 This is the SEM test result diagram of the biofilm inhibition test;

[0043] Figure 8 is a magnetic resonance imaging test; Figure 8a is the result of processing the scanned images of nanoassembly solutions of different pH and concentrations (0 mM, 0.4 mM, 0.2 mM, 0.1 mM, 0.05 mM) using Sante DICOM software. Figure 8 b is the magnetic resonance signal-to-noise ratio (SNR) of the nanoassembly solution at different pH and different concentrations (0 mM, 0.4 mM, 0.2 mM, 0.1 mM, 0.05 mM). DETAILED DESCRIPTION

[0044] In order to make the purpose, technical solutions and advantages of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with embodiments of the present invention. Obviously, the described embodiments are part of embodiments of the present invention, rather than all embodiments. Based on the embodiments in the present invention, all other embodiments obtained by those of ordinary skill in the art without making creative work are within the scope of protection of the present invention. If specific techniques or conditions are not indicated in the embodiments, they are carried out according to the techniques or conditions described in the literature in this area or according to the product specifications. Reagents or instruments used that do not indicate manufacturers are conventional products that can be obtained commercially.

[0045] It should be noted that, in the specification, “and / or” means at least one of the connected objects, and the character “ / ” generally indicates that the previous and subsequent related objects are in an “or” relationship.

[0046] In the present invention, the sonodynamic performance refers to the ability of the sonosensitizer to generate reactive oxygen species (ROS) under the action of ultrasound.

[0047] The first aspect of the present invention provides a nanoassembly, which is assembled from the following three components:

[0048] An amphiphilic block polymer, an antibiotic, and a sonosensitizer; wherein the amphiphilic block polymer has a structural formula as shown in Formula I, the antibiotic is a hydrophobic antibiotic, and the sonosensitizer is a porphyrin sonosensitizer;

[0049] Formula I

[0050] In formula I, m is an integer of 40-120, and n is an integer of 50-100.

[0051] The nanoassembly of the invention is assembled from an amphiphilic block polymer, an antibiotic and a sonosensitizer, and has excellent sonodynamic performance, antibacterial performance, pH responsiveness and nuclear magnetic resonance imaging capability. The inventors' research shows that: (1) the nanoassembly can generate reactive oxygen species (ROS) under the action of ultrasound, and it can respond to the acidic microenvironment and generate more ROS under the action of ultrasound, so it has excellent sonodynamic performance and pH responsiveness; (2) the nanoassembly can effectively remove bacterial biofilms, destroy the bacterial biofilm structure, release sonosensitizers and antibiotics under the action of ultrasound, and kill bacteria through the sonodynamic performance of sonosensitizers (releasing ROS) and the antibacterial properties of antibiotics. Moreover, the nanoassembly can also effectively kill left oxygen-resistant Klebsiella pneumoniae, Escherichia coli, Staphylococcus aureus and mold under the action of ultrasound, showing strong antibacterial activity, indicating that the nanoassembly can effectively reduce bacterial resistance and has excellent antibacterial activity against both bacteria and fungi, so it has excellent sonodynamic performance and antibacterial properties; (3) the nanoassembly can respond to the acidic microenvironment and significantly improve the magnetic resonance signal-to-noise ratio (SNR) of T1-weighted magnetic resonance imaging, achieving signal enhancement, so it has pH responsiveness and nuclear magnetic resonance imaging capabilities.

[0052] In a specific embodiment, the particle size of the nanoassembly is 50 to 200 nm. When the particle size of the nanoassembly is within the above range, the nanoassembly can effectively enter the biofilm and eliminate bacteria.

[0053] In a specific embodiment, in the above-mentioned nanoassembly, the mass ratio of the amphiphilic block polymer, the antibiotic and the sonosensitizer is (10-20): (1-3): (1-5).

[0054] When the mass ratio of the amphiphilic block polymer, the antibiotic and the sonosensitizer in the nanoassembly is within the above range, the amphiphilic block polymer, the antibiotic and the sonosensitizer are better matched, thereby obtaining a nanoassembly with excellent sonodynamic properties, antibacterial properties, pH responsiveness and nuclear magnetic resonance imaging capabilities.

[0055] In a specific embodiment, the hydrophobic antibiotics include at least one of levofloxacin, tetracycline, chloramphenicol, ciprofloxacin, azithromycin, doxycycline, linezolid, and norfloxacin.

[0056] In a specific embodiment, the porphyrin sonosensitizer includes 5,10,15,20-tetraphenyl-21H,23H-porphyrin manganese (II).

[0057] A second aspect of the present invention provides a method for preparing the above-mentioned nanoassembly, comprising the following steps:

[0058] Dissolving the amphiphilic block polymer, antibiotics and sonosensitizer in organic solvents respectively to prepare amphiphilic block polymer mother solution, antibiotic mother solution and sonosensitizer mother solution;

[0059] The amphiphilic block polymer mother solution, the antibiotic mother solution, the sonosensitizer mother solution and the PBS solution are mixed and then stirred and dialyzed to obtain a nanoassembly.

[0060] In a specific embodiment, the concentration of the amphiphilic block polymer in the amphiphilic block polymer mother solution is 14 to 18 mg / mL, for example, the concentration of the amphiphilic block polymer in the amphiphilic block polymer mother solution is 14 mg / mL, 15 mg / mL, 16 mg / mL, 17 mg / mL, 18 mg / mL, etc.; the concentration of the antibiotic in the antibiotic mother solution is 1.5 to 3 mg / mL, for example, the concentration of the antibiotic in the antibiotic mother solution is 1.5 mg / mL, 2 mg / mL, 2.5 mg / mL, 3 mg / mL, etc.; the concentration of the sonosensitizer in the sonosensitizer mother solution is 1.5 to 3 mg / mL, for example, the concentration of the sonosensitizer in the sonosensitizer mother solution is 1.5 mg / mL, 2 mg / mL, 2.5 mg / mL, 3 mg / mL, etc.

[0061] When the concentrations of the amphiphilic block polymer in the amphiphilic block polymer mother solution, the antibiotic in the antibiotic mother solution, and the sonosensitizer in the sonosensitizer mother solution are each within the above ranges, the ultimately prepared nanoassembly can effectively kill bacteria.

[0062] In a specific embodiment, the volume ratio of the amphiphilic block polymer mother solution, the antibiotic mother solution and the sonosensitizer mother solution is (1-2):1:1.

[0063] In a specific embodiment, during the above stirring process, the rotation speed is 1000-1500 rpm / min, the temperature is 20-30° C., and the time is 5-10 min.

[0064] When the parameters of rotation speed, temperature and time during the stirring process are each within the above range, the amphiphilic block polymer, antibiotic solution and sonosensitizer solution are fully assembled, thereby preparing a nanoassembly with excellent sonodynamic properties, antibacterial properties, pH responsiveness and nuclear magnetic resonance imaging capabilities.

[0065] For example, in the above stirring process, the rotation speed may be any one of 1000 rpm / min, 1100 rpm / min, 1200 rpm / min, 1300 rpm / min, 1400 rpm / min, and 1500 rpm / min, or a range consisting of any two thereof;

[0066] The temperature is any one of 20°C, 22°C, 24°C, 26°C, 28°C, and 30°C, or any two of them;

[0067] The time is any one of 5 min, 6 min, 7 min, 8 min, 9 min, and 10 min, or a range consisting of any two of them.

[0068] In a specific embodiment, in the above dialysis treatment, the molecular weight cut-off of the dialysis bag used is 3000 to 20000 Da, and the dialysis time is 8 to 12 h.

[0069] When the pore size of the dialysis bag and the dialysis time used in the dialysis treatment are within the above ranges, the loss of loaded drugs (antibiotics and sonosensitizers) is reduced and the organic solvent is removed to the greatest extent.

[0070] In one embodiment, the organic solvent includes dimethyl sulfoxide.

[0071] In one embodiment, the amphiphilic block polymer is obtained by a preparation method comprising the following steps:

[0072] 2-(azepan-1-yl)ethyl methacrylate, a macromolecular chain transfer agent PEG-CTA, and an initiator are dissolved in dimethyl sulfoxide to obtain a mixed solution;

[0073] The mixed solution is subjected to polymerization reaction at 50-80° C. for 6-12 hours to obtain the amphiphilic block polymer.

[0074] The present invention can prepare an amphiphilic block polymer through the above-mentioned preparation method, and assemble the amphiphilic block polymer with an antibiotic and a sonosensitizer to prepare a nanoassembly with excellent sonodynamic performance, antibacterial performance, pH responsiveness and nuclear magnetic resonance imaging capability.

[0075] The present invention does not particularly limit the specific preparation method of the above-mentioned macromolecular chain transfer agent PEG-CTA, and it can be prepared according to methods well known to those skilled in the art. In some embodiments, the preparation method of the macromolecular chain transfer agent PEG-CTA is as follows:

[0076] Dissolve 2.5 mmol of polyethylene glycol monomethyl ether in 50 mL of anhydrous tetrahydrofuran to obtain a polyethylene glycol monomethyl ether solution;

[0077] 5 mmol of 4-cyano-4-(((ethylthio)thiocarbonyl)thio)pentanoic acid, 5 mmol of dicyclohexylcarbodiimide and 0.5 mmol of 4-dimethylaminopyridine were dissolved in 40 mL of anhydrous tetrahydrofuran to obtain a mixture solution;

[0078] The mixture solution was mixed with the polyethylene glycol monomethyl ether solution and then subjected to esterification reaction at 25° C. under stirring conditions for 72 hours to obtain a reaction product;

[0079] The reaction product is sequentially subjected to first suction filtration, reduced pressure distillation, precipitation, second suction filtration and vacuum drying to obtain a macromolecular chain transfer agent PEG-CTA.

[0080] In one specific embodiment, the initiator includes azobisisobutyronitrile.

[0081] In a specific embodiment, the mass ratio of the 2-(azepan-1-yl)ethyl methacrylate, the macromolecular chain transfer agent PEG-CTA and the initiator is (50-150):1:(0.1-0.3).

[0082] A third aspect of the present invention provides the use of the aforementioned nanoassembly, or a nanoassembly prepared by the aforementioned method for preparing the nanoassembly, in the preparation of a product for treating bacterial and fungal infections, or in the preparation of a product for treating bacterial biofilm infections. The inventors' research has shown that, under the action of ultrasound, the nanoassembly can effectively reduce bacterial drug resistance and exhibit excellent antibacterial activity against both bacteria and fungi. Therefore, it can be used in the preparation of products for treating bacterial and / or fungal infections. Furthermore, under the action of ultrasound, the nanoassembly can effectively remove bacterial biofilms, disrupt their structure, release sonosensitizers and antibiotics, and kill bacteria through the sonosensitizer's sonodynamic properties (release of ROS) and the antibiotic's antibacterial properties. Therefore, it can be used in the preparation of products for treating bacterial biofilm infections.

[0083] A fourth aspect of the present invention provides the use of the aforementioned nanoassembly, or a nanoassembly prepared by the aforementioned method for preparing the nanoassembly, in the preparation of a magnetic resonance imaging contrast agent. The inventors' research has shown that, when used in magnetic resonance imaging experiments, the nanoassembly can respond to an acidic microenvironment, significantly improving the magnetic resonance signal-to-noise ratio (SNR) of T1-weighted magnetic resonance imaging, achieving signal enhancement. Therefore, the nanoassembly can be used to prepare a magnetic resonance imaging contrast agent.

[0084] The present invention is further described below through specific examples.

[0085] Example 1

[0086] This embodiment provides a method for preparing a nanoassembly, comprising the following steps:

[0087] (1) Preparation of macromolecular chain transfer agent PEG-CTA:

[0088] Dissolve polyethylene glycol monomethyl ether (2.5 mmol, 12.5 g) in 50 mL of anhydrous tetrahydrofuran to obtain polyethylene glycol monomethyl ether solution;

[0089] 4-Cyano-4-(((ethylthio)thiocarbonyl)thio)pentanoic acid (5 mmol, 1.38 g), dicyclohexylcarbodiimide (5 mmol, 1.03 g) and 4-dimethylaminopyridine (0.5 mmol, 0.06 g) were dissolved in 40 mL of anhydrous tetrahydrofuran to obtain a mixture solution;

[0090] The mixture solution was mixed with polyethylene glycol monomethyl ether solution and then subjected to esterification reaction at 25°C under stirring conditions for 72 hours to obtain a reaction product;

[0091] The reaction product was sequentially subjected to first suction filtration, reduced pressure distillation, precipitation, second suction filtration, and vacuum drying to obtain a macromolecular chain transfer agent PEG-CTA (abbreviated as PEG-CTA). The chemical structure of PEG-CTA is as follows:

[0092] .

[0093] (2) Preparation of amphiphilic block polymers:

[0094] 552 mg of 2-(azepan-1-yl)ethyl methacrylate, 207 mg of PEG-CTA, and 2 mg of azobisisobutyronitrile (AIBN) were dissolved in 500 μL of dimethyl sulfoxide and subjected to three freeze-thaw cycles to obtain a mixed solution;

[0095] The mixed solution was polymerized at 70°C for 8 h to obtain a polymerized product. 2 mL of tetrahydrofuran was added to the polymerized product to dilute it to obtain a diluted product. The diluted product was dialyzed and freeze-dried to obtain an amphiphilic block polymer (denoted as PC7A). The chemical structure of the amphiphilic block polymer is as follows:

[0096] , where m is 113 and n is 57.

[0097] (3) Preparation of nanoassemblies:

[0098] Amphiphilic block polymer (PC7A), levofloxacin (LEV), and 5,10,15,20-tetraphenyl-21H,23H-porphyrin manganese (II) were dissolved in dimethyl sulfoxide (DMSO) to prepare 16 mg / mL PC7A stock solution, 1.8 mg / mL LEV stock solution, and 2 mg / mL MnTCPP stock solution, respectively;

[0099] After mixing 0.3 mL of PC7A mother solution, 0.2 mL of LEV mother solution, and 0.2 mL of MnTCPP mother solution, a mixture solution was obtained;

[0100] The mixture solution was placed in an assembly bottle containing 7 mL of phosphate buffer solution (PBS solution), and stirred at a speed of 1500 rpm / min and a temperature of 20° C. for 5 minutes to obtain a stirred product;

[0101] The stirred product was dialyzed for 12 h using a dialysis bag with a pore size of 3000 Da to obtain a nanoassembly (denoted as LEV+MnTCPP@PC7A).

[0102] Performance testing:

[0103] 1. Nuclear magnetic hydrogen spectrum test

[0104] The amphiphilic block polymer in Example 1 of the present invention was tested by nuclear magnetic hydrogen spectrum, and the results were as follows: Figure 1 As shown; Figure 1 This is the hydrogen NMR spectrum of the amphiphilic block polymer in Example 1 of the present invention.

[0105] Depend on Figure 1 It can be seen that the amphiphilic block polymer having the structure shown in Formula I was successfully prepared.

[0106] 2. Ultraviolet absorption spectrum test

[0107] The nanoassembly in Example 1 was tested by ultraviolet absorption spectrum, and the results were as follows: Figure 2 As shown; Figure 2 This is the ultraviolet absorption spectrum of the nanoassembly in Example 1.

[0108] Depend on Figure 2 It can be seen that the nanoassembly in Example 1 has two obvious absorption peaks at wavelengths of 330 nm and 470 nm, wherein the absorption peak at 330 nm is the characteristic peak of the absorption spectrum of LEV, and the absorption peak at 470 nm is the characteristic peak of the absorption spectrum of MnTCPP.

[0109] 3. Particle size and morphology test

[0110] The particle size and morphology of the nanoassembly in Example 1 of the present invention were tested respectively. The results are as follows: Figure 3 As shown in ab; Figure 3 The particle size distribution diagram and transmission electron microscope (TEM) diagram of the nanoassembly in Example 1 of the present invention are shown in FIG. Figure 3 a is the particle size distribution diagram of the nanoassembly in Example 1, Figure 3 b is the TEM image of the nanoassembly in Example 1.

[0111] Depend on Figure 3 From a, we can see that the particle size of the nanoassembly in Example 1 is 164 nm.

[0112] Depend on Figure 3 As can be seen from b, the nanoassemblies in Example 1 are round particles.

[0113] 4. pH responsiveness and sonodynamic performance test

[0114] The acoustic dynamic performance test of the nanoassembly in Example 1 of the present invention was carried out, and the specific method is as follows:

[0115] The nanoassembly (LEV+MnTCPP@PC7A) in Example 1 was prepared into a nanoassembly (LEV+MnTCPP@PC7A) solution with a pH of 6.0 using a phosphate buffer solution (PBS solution). The LEV+MnTCPP@PC7A solution with a pH of 6.0 was used as the experimental group A, and the phosphate buffer solution with a pH of 6.0 was used as the control group A.

[0116] The nanoassembly (LEV+MnTCPP@PC7A) in Example 1 was prepared into a LEV+MnTCPP@PC7A solution with a pH of 7.4 using PBS solution. The LEV+MnTCPP@PC7A solution with a pH of 7.4 was used as experimental group B, and the phosphate buffer solution with a pH of 7.4 was used as control group B.

[0117] Under the action of 1.5W ultrasound, the experimental group A, control group A, experimental group B and control group B were ultrasonically treated for 2 minutes, and the release of reactive oxygen species (ROS) in each group was measured using 2,7-dichlorodihydrofluorescein diacetate (DCFH-DA) as a fluorescent probe. During the ultrasonic treatment, the excitation wavelength of 488 nm was used to record the changes in the fluorescence intensity of each group. The results are shown in Figure 2. Figure 4 As shown in a and b; Figure 4 The ROS release curves of LEV+MnTCPP@PC7A solutions with different pH values ​​before and after ultrasound treatment are shown in Figure 2. Figure 4 a is the ROS release curve of LEV+MnTCPP@PC7A solution at pH=6.0 before and after ultrasound treatment. Figure 4 b is the ROS release curve of LEV+MnTCPP@PC7A solution with pH = 7.4 before and after ultrasound action.

[0118] Depend on Figure 4As shown by ab, under 1.5W ultrasound, LEV+MnTCPP@PC7A solutions of different pH values ​​released varying amounts of reactive oxygen species (ROS). Compared with the LEV+MnTCPP@PC7A solution at pH 7.4, the LEV+MnTCPP@PC7A solution at pH 6.0 produced more ROS with prolonged ultrasound treatment. These results demonstrate that the nanoassemblies provided by the present invention can respond to acidic microenvironments and generate more ROS under ultrasound, demonstrating excellent sonodynamic performance and pH responsiveness.

[0119] 5. Antibacterial activity test

[0120] The antibacterial performance of the nanoassembly in Example 1 of the present invention was tested, and the specific method is as follows:

[0121] (1) Bacterial culture:

[0122] Levooxygen-resistant Klebsiella pneumoniae, Escherichia coli, and Staphylococcus aureus were inoculated into fresh tryptone soy broth (TSB) medium and incubated at 37°C for 15 h to obtain bacterial suspensions. 50 μL of the bacterial suspension was then added to 5 mL of fresh TSB medium and incubated at 37°C until the bacteria reached the logarithmic growth phase. The bacteria were collected by centrifugation and washed twice with sterile phosphate buffered saline (PBS). Each wash was followed by centrifugation at 10,000 rpm for 5 min at 4°C. After the two washes, the bacterial concentration was adjusted to 1.5 × 10 6 CFU / mL, and obtained the oxygen-resistant Klebsiella pneumoniae suspension, Escherichia coli suspension and Staphylococcus aureus suspension respectively.

[0123] (2) Fungal (mold) culture:

[0124] The fungus was inoculated into yeast extract peptone dextrose medium (YPD), placed in a shaking incubator at 30°C and 200 rpm / min, and cultured for 15 h. The fungal cells were collected by centrifugation and diluted to 2 × 10 5 CFU / mL to obtain fungal suspension.

[0125] (3) Antibacterial performance test of nanoassemblies:

[0126] 1. Antibacterial activity of the nanoassembly against levothiocyanate-resistant Klebsiella pneumoniae: ① Experimental group without ultrasonic treatment (-US): 100 μL of the assembly was added to a 96-well plate, and then 50 μL of levothiocyanate-resistant Klebsiella pneumoniae suspension was added to each well, and the plates were incubated at 37°C for 20 min to obtain an incubated levothiocyanate-resistant Klebsiella pneumoniae suspension; the incubated levothiocyanate-resistant Klebsiella pneumoniae suspension was further incubated at 37°C for 30 min, and then the bacterial suspension was diluted 100-fold with sterile PBS to obtain a diluted levothiocyanate-resistant Klebsiella pneumoniae suspension; 20 μL of the diluted levothiocyanate-resistant Klebsiella pneumoniae suspension was placed on a tryptone soy agar plate and incubated at 37°C for 15 h to obtain a nanoassembly-treated levothiocyanate-resistant Klebsiella pneumoniae suspension (LEV+MnTCPP@PC7A);

[0127] Control group without ultrasonic treatment (-US): The steps were basically the same as those of the experimental group without ultrasonic treatment (-US), except that 100 μL of nanoassemblies was replaced with 100 μL of phosphate buffered saline (PBS), thus finally obtaining a PBS-treated suspension of levothroid-resistant Klebsiella pneumoniae (PBS);

[0128] ② Experimental group subjected to ultrasonic treatment (+US): 100 μL of the nanoassembly was added to a 96-well plate, and then 50 μL of the levodopa-resistant Klebsiella pneumoniae suspension was added to each well. The plates were incubated at 37°C for 20 min to obtain the incubated levodopa-resistant Klebsiella pneumoniae suspension. The incubated levodopa-resistant Klebsiella pneumoniae suspension was ultrasonically treated with 1.5 W ultrasound for 2 min, and then incubated at 37°C for 30 min. The bacterial suspension was then diluted 100-fold with sterile PBS to obtain the diluted levodopa-resistant Klebsiella pneumoniae suspension. 20 μL of the diluted levodopa-resistant Klebsiella pneumoniae suspension was placed on a tryptone soy agar plate and incubated at 37°C for 15 h to obtain the nanoassembly-treated levodopa-resistant Klebsiella pneumoniae suspension (LEV+MnTCPP@PC7A).

[0129] The control group with ultrasonic treatment (+US) was basically the same as the experimental group with ultrasonic treatment (+US), except that 100 μL of the nanoassembly was replaced with 100 μL of phosphate buffered saline (PBS), thus finally obtaining a PBS-treated suspension of levothroid-resistant Klebsiella pneumoniae (PBS);

[0130] Bacterial viability tests were performed on the nanoassembly-treated levodopa Klebsiella pneumoniae suspension (LEV+MnTCPP@PC7A) in the experimental group without ultrasonic treatment (-US), the PBS-treated levodopa Klebsiella pneumoniae suspension (PBS) in the control group without ultrasonic treatment (-US), the nanoassembly-treated levodopa Klebsiella pneumoniae suspension (LEV+MnTCPP@PC7A) in the experimental group with ultrasonic treatment (+US), and the PBS-treated levodopa Klebsiella pneumoniae suspension (PBS) in the control group with ultrasonic treatment (+US). The results are shown in Figure 2. Figure 5 As shown in a.

[0131] 2. Antibacterial activity of the nanoassembly against Escherichia coli: ① Experimental group without ultrasonic treatment (-US): The procedure was basically the same as the experimental group without ultrasonic treatment (-US) in the antibacterial activity of the nanoassembly against levothiocyanate-resistant Klebsiella pneumoniae, except that the levothiocyanate-resistant Klebsiella pneumoniae suspension was replaced with an Escherichia coli suspension, resulting in a nanoassembly-treated Escherichia coli suspension (LEV+MnTCPP@PC7A);

[0132] Control group without ultrasonic treatment (-US): The steps were basically the same as those of the experimental group without ultrasonic treatment (-US), except that 100 μL of nanoassemblies was replaced by 100 μL of phosphate buffer solution (PBS), thus finally obtaining a PBS-treated E. coli suspension (PBS);

[0133] ② Experimental group with ultrasonic treatment (+US): The procedure was basically the same as that of the experimental group with ultrasonic treatment (+US) in the antibacterial activity of the nanoassembly against levothiocyanate-resistant Klebsiella pneumoniae, except that the levothiocyanate-resistant Klebsiella pneumoniae suspension was replaced with an Escherichia coli suspension, thus finally obtaining a nanoassembly-treated Escherichia coli suspension (LEV+MnTCPP@PC7A);

[0134] Ultrasonication-treated (+US) control group: The steps were basically the same as those for the ultrasonication-treated (+US) experimental group, except that 100 μL of nanoassemblies was replaced with 100 μL of phosphate buffered saline (PBS), thus obtaining a PBS-treated E. coli suspension (PBS);

[0135] Bacterial activity tests were performed on the Escherichia coli suspension (LEV+MnTCPP@PC7A) treated with nanoassemblies in the experimental group without ultrasonic treatment (-US), the Escherichia coli suspension (PBS) treated with PBS in the control group without ultrasonic treatment (-US), the Escherichia coli suspension (LEV+MnTCPP@PC7A) treated with nanoassemblies in the experimental group with ultrasonic treatment (+US), and the Escherichia coli suspension (PBS) treated with PBS in the control group with ultrasonic treatment (+US). The results are shown in Figure 2. Figure 5 As shown in b.

[0136] 3. Antibacterial activity of the nanoassembly against Staphylococcus aureus: ① Experimental group without ultrasonic treatment (-US): The procedure was basically the same as the experimental group without ultrasonic treatment (-US) in the antibacterial activity of the nanoassembly against levothiocyanate-resistant Klebsiella pneumoniae, except that the levothiocyanate-resistant Klebsiella pneumoniae suspension was replaced with a Staphylococcus aureus suspension, thus obtaining a nanoassembly-treated Staphylococcus aureus suspension (LEV+MnTCPP@PC7A);

[0137] Control group without ultrasonic treatment (-US): The steps were basically the same as those of the experimental group without ultrasonic treatment (-US), except that 100 μL of nanoassemblies was replaced by 100 μL of phosphate buffer solution (PBS), thus finally obtaining a PBS-treated Staphylococcus aureus suspension (PBS);

[0138] ② Experimental group with ultrasonic treatment (+US): The procedure was basically the same as that of the experimental group with ultrasonic treatment (+US) in the antibacterial activity of the nanoassembly against levothiocyanate-resistant Klebsiella pneumoniae, except that the levothiocyanate-resistant Klebsiella pneumoniae suspension was replaced with a Staphylococcus aureus suspension, thus finally obtaining a nanoassembly-treated Staphylococcus aureus suspension (LEV+MnTCPP@PC7A);

[0139] Ultrasonication-treated (+US) control group: The steps were basically the same as those of the ultrasonication-treated (+US) experimental group, except that 100 μL of nanoassemblies was replaced with 100 μL of phosphate buffered saline (PBS), thus finally obtaining a PBS-treated Staphylococcus aureus suspension (PBS);

[0140] Bacterial viability tests were performed on the nanoassembly-treated Staphylococcus aureus suspension (LEV+MnTCPP@PC7A) in the experimental group without ultrasonic treatment (-US), the PBS-treated Staphylococcus aureus suspension (PBS) in the control group without ultrasonic treatment (-US), the nanoassembly-treated Staphylococcus aureus suspension (LEV+MnTCPP@PC7A) in the experimental group with ultrasonic treatment (+US), and the PBS-treated Staphylococcus aureus suspension (PBS) in the control group with ultrasonic treatment (+US). The results are shown in Figure 2. Figure 5 As shown in c.

[0141] 4. Antibacterial activity of the nanoassembly against mold: ① Experimental group without ultrasonic treatment (-US): The steps were basically the same as those of the experimental group without ultrasonic treatment (-US) in the antibacterial activity of the nanoassembly against levothiocyanate-resistant Klebsiella pneumoniae, except that the levothiocyanate-resistant Klebsiella pneumoniae suspension was replaced with a mold suspension and the tryptone soy agar plate was replaced with a yeast extract peptone dextrose agar plate, thus finally obtaining a nanoassembly-treated mold suspension (LEV+MnTCPP@PC7A);

[0142] Control group without ultrasonic treatment (-US): The steps were basically the same as those of the experimental group without ultrasonic treatment (-US), except that 100 μL of nanoassemblies was replaced by 100 μL of phosphate buffer solution (PBS), thus finally obtaining a PBS-treated fungal suspension (PBS);

[0143] ② Experimental group with ultrasonic treatment (+US): The steps of the experimental group with ultrasonic treatment (+US) in the antibacterial activity of the nanoassembly against levodopa-resistant Klebsiella pneumoniae were basically the same, except that the levodopa-resistant Klebsiella pneumoniae suspension was replaced with a mold suspension and the tryptone soy agar plate was replaced with a yeast extract peptone dextrose agar plate, thus finally obtaining a nanoassembly-treated mold suspension (LEV+MnTCPP@PC7A);

[0144] The control group with ultrasonic treatment (+US) was similar to the experimental group with ultrasonic treatment (+US), except that 100 μL of nanoassemblies was replaced with 100 μL of phosphate buffered saline (PBS), thus obtaining a PBS-treated fungal suspension (PBS).

[0145] Bacterial activity tests were performed on the fungal suspension (LEV+MnTCPP@PC7A) treated with nanoassemblies in the experimental group without ultrasonic treatment (-US), the fungal suspension (PBS) treated with PBS in the control group without ultrasonic treatment (-US), the fungal suspension (LEV+MnTCPP@PC7A) treated with nanoassemblies in the experimental group with ultrasonic treatment (+US), and the fungal suspension (PBS) treated with PBS in the control group with ultrasonic treatment (+US). The results are shown in Figure 2. Figure 5 As shown in d.

[0146] Figure 5 For antibacterial activity test, Figure 5 a is the antibacterial activity of the nanoassembly against levothroid-resistant Klebsiella pneumoniae. Figure 5 b is the antibacterial activity of the nanoassembly against Escherichia coli, Figure 5 c is the antibacterial activity of the nanoassembly against Staphylococcus aureus, Figure 5 d is the antibacterial activity of the nanoassembly against mold.

[0147] Depend on Figure 5 It can be seen from a that when no ultrasonic treatment is performed, the survival rate of the oxygen-resistant Klebsiella pneumoniae after the nanoassembly is sterilized is 77%; while when ultrasonic treatment is performed, the survival rate of the oxygen-resistant Klebsiella pneumoniae after the nanoassembly is sterilized is 8%.

[0148] Depend on Figure 5 It can be seen from b that when no ultrasonic treatment is performed, the survival rate of Escherichia coli is 82% after the nanoassembly is used to sterilize Escherichia coli; while when ultrasonic treatment is performed, the survival rate of Escherichia coli is 9% after the nanoassembly is used to sterilize Escherichia coli.

[0149] Depend on Figure 5 It can be seen from c that when no ultrasonic treatment is performed, the survival rate of Staphylococcus aureus is 84% ​​after the nanoassembly is sterilized against Staphylococcus aureus; while when ultrasonic treatment is performed, the survival rate of Staphylococcus aureus is 9% after the nanoassembly is sterilized against Staphylococcus aureus.

[0150] Depend on Figure 5 It can be seen from d that when no ultrasonic treatment is performed, the survival rate of the mold is 85% after the nanoassembly is used to sterilize the mold; while when ultrasonic treatment is performed, the survival rate of the mold is 11%.

[0151] The above results show that the nanoassembly provided by the present invention can effectively reduce bacterial resistance, kill bacteria and fungi under the action of ultrasound, and has excellent antibacterial activity against both bacteria and fungi, indicating that the nanoassembly provided by the present invention has excellent antibacterial properties.

[0152] 5. Biofilm inhibition test

[0153] The nanoassembly in Example 1 of the present invention was subjected to a biofilm inhibition test as follows:

[0154] (1) Biofilm culture: 500 μL of tryptone soy broth (TSB) medium and 100 μL of Klebsiella pneumoniae suspension (10 8 CFU / mL), and incubated in a 37°C incubator for 24 h. Subsequently, the old TSB medium was replaced with fresh TSB medium, and incubated for another 24 h to obtain the cultured biofilm.

[0155] (2) The effect of nanoassemblies on the removal of biofilm: ① Experimental group without ultrasonic treatment (-US): The cultured biofilm was mixed with 100 μL of nanoassemblies (nanoassemblies prepared according to the preparation method of Example 1), and then incubated at 37°C for 1 h, followed by further incubation at 37°C for 30 min, washed twice with PBS, added with 0.5 mL of crystal violet staining solution and incubated for 20 min, washed twice with PBS, and finally added with 500 μL of pure ethanol to obtain a biofilm suspension treated with nanoassemblies (LEV+MnTCPP@PC7A);

[0156] Control group without ultrasonic treatment (-US): The steps were basically the same as those of the experimental group without ultrasonic treatment (-US), except that 100 μL of nanoassemblies was replaced by 100 μL of phosphate buffer solution (PBS), thus finally obtaining a PBS-treated biofilm suspension (PBS);

[0157] ② Experimental group subjected to ultrasonic treatment (+US): The cultured biofilm was mixed with 100 μL of the nanoassembly (prepared according to the preparation method of Example 1) and then incubated at 37°C for 1 hour to obtain the incubated biofilm; the incubated biofilm was ultrasonically treated with 1.5 W ultrasound for 2 minutes, incubated at 37°C for another 30 minutes, washed twice with PBS, incubated with 0.5 mL of crystal violet staining solution for 20 minutes, washed twice with PBS, and finally added with 500 μL of pure ethanol to obtain a nanoassembly-treated biofilm suspension (LEV+MnTCPP@PC7A);

[0158] Control group with ultrasonic treatment (+US): The steps were basically the same as those of the experimental group with ultrasonic treatment (+US), except that 100 μL of nanoassemblies was replaced by 100 μL of phosphate buffer solution (PBS), thus finally obtaining a PBS-treated biofilm suspension (PBS).

[0159] The biofilm suspensions treated with nanoassemblies (LEV+MnTCPP@PC7A) in the experimental group without ultrasonic treatment (-US), the biofilm suspensions treated with PBS (PBS) in the control group without ultrasonic treatment (-US), the biofilm suspensions treated with nanoassemblies (LEV+MnTCPP@PC7A) in the experimental group with ultrasonic treatment (+US), and the biofilm suspensions treated with PBS in the control group with ultrasonic treatment (+US) were photographed. The results are shown in Figure 2. Figure 6 As shown; Figure 6 This is the crystal violet staining result in the biofilm inhibition test.

[0160] Depend on Figure 6 It can be seen that the nanoassembly provided in the embodiment of the present invention can effectively remove bacterial biofilms and destroy the bacterial biofilm structure under the action of ultrasound. This is because the nanoassembly releases MnTCPP and LEV under the action of ultrasound, killing bacteria through the sonodynamic properties of MnTCPP (releasing ROS) and the antibacterial properties of LEV, thereby effectively removing bacterial biofilms and destroying the bacterial biofilm structure.

[0161] Scanning electron microscopy (SEM) was used to observe the morphology of the biofilm suspension (LEV+MnTCPP@PC7A) treated with nanoassemblies in the experimental group without ultrasonic treatment (-US), the biofilm suspension (PBS) treated with PBS in the control group without ultrasonic treatment (-US), the biofilm suspension (LEV+MnTCPP@PC7A) treated with nanoassemblies in the experimental group with ultrasonic treatment (+US), and the biofilm suspension (PBS) treated with PBS in the control group with ultrasonic treatment (+US). The specific methods are as follows:

[0162] Each group was washed three times with phosphate buffered saline (PBS) to remove excess culture medium and metabolites. After washing, each group was placed in a 4°C environment and fixed overnight with 1 mL of paraformaldehyde solution (concentration of 2.5 wt%) to ensure the structural stability of the biofilm in each group. After fixation, in order to further facilitate SEM observation, each group of biofilms was serially dehydrated with gradient ethanol solutions. This process included ethanol solutions of different mass fractions (30%, 50%, 70%, 80%, 90%, 95% and 100%). Each mass fraction of ethanol solution was used to dehydrate each group of biofilms. Finally, the dehydrated biofilms of each group were placed in a vacuum state for drying overnight. The dried biofilms of each group were tested and observed using a scanning electron microscope (SEM). The results are shown in Figure 2. Figure 7 As shown; Figure 7 This is the SEM test result of the biofilm inhibition test.

[0163] Figure 7 The results further illustrate that the nanoassembly provided by the embodiment of the present invention can effectively remove bacterial biofilms and destroy the bacterial biofilm structure under the action of ultrasound.

[0164] 6. Magnetic resonance imaging test

[0165] The nanoassemblies (prepared according to the preparation method of the nanoassemblies in Example 1) were respectively prepared into nanoassembly solutions with different concentrations (0 mM, 0.4 mM, 0.2 mM, 0.1 mM, 0.05 mM) at pH = 6.0 using PBS solution, where the concentration refers to the concentration of MnTCPP in the nanoassembly solution;

[0166] The nanoassemblies (prepared according to the preparation method of the nanoassemblies in Example 1) were respectively prepared into nanoassembly solutions with different concentrations (0 mM, 0.4 mM, 0.2 mM, 0.1 mM, 0.05 mM) at pH = 7.4 using PBS solution, where the concentration refers to the concentration of MnTCPP in the nanoassembly solution;

[0167] T1-weighted sequence scanning experiments were performed on a Philips 3.0T magnetic resonance imaging system for nanoassembly solutions with different concentrations (0 mM, 0.4 mM, 0.2 mM, 0.1 mM, 0.05 mM) at pH = 6.0 and nanoassembly solutions with different concentrations (0 mM, 0.4 mM, 0.2 mM, 0.1 mM, 0.05 mM) at pH = 7.4. The scanned images were obtained and then processed using Sante DICOM software ( Figure 8a), the magnetic resonance signal-to-noise ratio (SNR) of the nanoassembly solution with different concentrations (0 mM, 0.4 mM, 0.2 mM, 0.1 mM, 0.05 mM) at pH = 6.0 or pH = 7.4 was calculated and recorded ( Figure 8 b).

[0168] Figure 8 is a magnetic resonance imaging test; Figure 8 a is the result of processing the scanned images of nanoassembly solutions of different pH and concentrations (0 mM, 0.4 mM, 0.2 mM, 0.1 mM, 0.05 mM) using Sante DICOM software. Figure 8 b is the magnetic resonance signal-to-noise ratio (SNR) of the nanoassembly solution at different pH and different concentrations (0 mM, 0.4 mM, 0.2 mM, 0.1 mM, 0.05 mM).

[0169] Depend on Figure 8 From a and b, we can see that with the increase of MnTCPP concentration in the nanoassembly solution, SNR shows a certain trend of change. Specifically, when the MnTCPP concentration in the nanoassembly solution is low, the SNR value is relatively low, indicating that the signal strength is weak; while with the increase of MnTCPP concentration in the nanoassembly solution, the SNR value gradually increases, indicating that the signal strength is enhanced. And by Figure 8 As can be seen from a and b, when the MnTCPP concentration in the nanoassembly solution is the same, the SNR values ​​of the nanoassembly solutions at different pH values ​​are different, with the SNR value of the nanoassembly solution at pH = 6.0 being higher than that of the nanoassembly solution at pH = 7.4. These results demonstrate that the nanoassembly provided by the embodiments of the present invention can respond to an acidic microenvironment, significantly improve the SNR of T1-weighted magnetic resonance imaging, achieve signal enhancement, possess nuclear magnetic resonance imaging capabilities, and can be used as a T1-enhanced magnetic resonance imaging contrast agent.

[0170] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the above embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the above embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A nanoassembly, characterized in that: The nanoassembly is assembled from the following three components: An amphiphilic block polymer, a hydrophobic antibiotic, and a porphyrin sonosensitizer; wherein the structural formula of the amphiphilic block polymer is as shown in Formula I, the hydrophobic antibiotic is levofloxacin, and the porphyrin sonosensitizer is 5,10,15,20-tetraphenyl-21H,23H-porphyrin manganese (II); in the nanoassembly, the mass ratio of the amphiphilic block polymer, the antibiotic, and the sonosensitizer is (10-20): (1-3): (1-5); Formula I In formula I, m is an integer of 40-120, and n is an integer of 50-100.

2. The nanoassembly according to claim 1, characterized in that The particle size of the nanoassembly is 50-200 nm.

3. A method for preparing the nanoassembly according to any one of claims 1 to 2, characterized in that: The following steps are involved: dissolving the amphiphilic block polymer, the antibiotic and the sonosensitizer in an organic solvent respectively to prepare an amphiphilic block polymer mother solution, an antibiotic mother solution and a sonosensitizer mother solution; The amphiphilic block polymer mother solution, the antibiotic mother solution, the sonosensitizer mother solution and PBS solution are mixed and then stirred and dialyzed to obtain the nanoassembly.

4. The method for preparing a nanoassembly according to claim 3, wherein: The concentration of the amphiphilic block polymer in the amphiphilic block polymer mother solution is 14-18 mg / mL, the concentration of the antibiotic in the antibiotic mother solution is 1.5-3 mg / mL, and the concentration of the sonosensitizer in the sonosensitizer mother solution is 1.5-3 mg / mL; The volume ratio of the amphiphilic block polymer mother solution, the antibiotic mother solution and the sonosensitizer mother solution is (1-2):1:

1.

5. The method for preparing a nanoassembly according to claim 3, wherein: During the stirring process, the rotation speed is 1000-1500 rpm / min, the temperature is 20-30°C, and the time is 5-10 min; And / or, in the dialysis treatment, the molecular weight cut-off of the dialysis bag used is 3000 to 20000 Da, and the dialysis time is 8 to 12 h; And / or, the organic solvent comprises dimethyl sulfoxide.

6. The method for preparing a nanoassembly according to claim 3, wherein: The amphiphilic block polymer is obtained by a preparation method comprising the following steps: 2-(azepan-1-yl)ethyl methacrylate, a macromolecular chain transfer agent PEG-CTA, and an initiator are dissolved in dimethyl sulfoxide to obtain a mixed solution; The mixed solution is subjected to polymerization reaction at 50-80° C. for 6-12 hours to obtain the amphiphilic block polymer; Wherein the initiator is azobisisobutyronitrile; The mass ratio of the 2-(azepan-1-yl)ethyl methacrylate, the macromolecular chain transfer agent PEG-CTA and the initiator is (50-150):1:(0.1-0.3).

7. Use of the nanoassembly according to any one of claims 1 to 2 or the nanoassembly prepared by the method for preparing the nanoassembly according to any one of claims 3 to 6 in preparing a product for treating bacterial and fungal infections or in preparing a product for treating bacterial biofilm infections, characterized in that: The bacteria is any one of levooxygen-resistant Klebsiella pneumoniae, Escherichia coli, and Staphylococcus aureus, and the fungus is a mold.

8. Use of the nanoassembly according to any one of claims 1 to 2 or the nanoassembly prepared by the method for preparing a nanoassembly according to any one of claims 3 to 6 in preparing a magnetic resonance imaging contrast agent.

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