A composite sonosensitizer and its bacterial-targeting delivery system, and related preparation methods and applications

Abstract

The present application provides a composite sonosensitizer and its bacterial target delivery system BPT-ICG@Bd capable of treating pseudomonas aeruginosa biofilm related infection, wherein the composite sonosensitizer (BPT-ICG) is composed of inorganic sonosensitizer (BPT) and organic sonosensitizer (ICG), Bd is natural gram-negative bacteria predator Bdellovibrio, the composite sonosensitizer (BPT-ICG) is modified to the surface of Bdellovibrio (Bd) by polydopamine to form BPT-ICG@Bd. The high-speed movement characteristics of Bdellovibrio predator gram-negative bacteria are used to destroy the biofilm structure, invade bacteria, and at the same time, the composite sonosensitizer is triggered to produce a large amount of active oxygen by applying ultrasound, so as to quickly remove the biofilm and bacteria, and be used for biofilm related lung infection and bone infection.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of medical technology, in particular to a composite sonosensitizer and its bacterial targeting delivery system, and related preparation method and application. BACKGROUND

[0002] Bacterial infection is a major disease that seriously threatens human health. Antibiotic therapy is currently the most effective means to deal with bacterial infection in clinical practice. However, due to the extensive use of antibiotics, bacteria have become increasingly resistant to antibiotics, which has become a serious global public health problem. On the other hand, in most clinical cases of bacterial infection, biofilms are formed. Biofilms can act as a physical barrier to prevent the entry of antibiotics, and the unique biofilm microenvironment (such as hypoxia and acidity) severely limits the efficacy of antibacterial drugs, which makes the bacteria in the biofilm 10-1000 times more resistant to antibiotics than planktonic bacteria, posing a serious challenge to current antibiotic therapy.

[0003] Clinical treatment of bacterial pneumonia and osteomyelitis often relies on empirical treatment, so antibiotics are used in large quantities, leading to the increasing prevalence of multi-drug resistant pathogens such as Pseudomonas aeruginosa, and these bacterial infections are often accompanied by the presence of biofilms, making it difficult for antibiotics to effectively treat them. Moreover, large doses and long-term intravenous or oral administration of antibiotics can cause varying degrees of damage to the body, such as liver and kidney function impairment, gastrointestinal dysfunction, and immune system damage. Therefore, it is urgent to develop an antibiotic-free, efficient, and safe treatment strategy.

[0004] Bdellovibrio bacteriovorus (Bd) is a unique microorganism that can prey on a variety of pathogenic gram-negative bacteria and is known as a "live antibiotic". Bdellovibrio is usually smaller than bacteria and can pass through bacterial filters, with special flagella at the end, providing high mobility. During the predation phase, Bdellovibrio collides with the identified prey at a speed of up to 160 m / s. After the collision, the bacteria usually rotate at a speed of more than 100 rpm / s. After that, Bdellovibrio penetrates into the cytoplasm through the attachment point and grows and divides in it to kill the prey. This provides a biological basis for Bdellovibrio to kill gram-negative bacteria and invade biofilms. Previous studies have shown that Bdellovibrio cannot invade mammalian cells and has no obvious pathogenic effect, and can be easily isolated from the gastrointestinal tract and feces of mammals. Although Bdellovibrio has the above advantages, it is difficult to achieve satisfactory antibacterial effect when used alone, mainly because of its slow antibacterial rate. Therefore, in order to improve the antibacterial treatment effect, it is necessary to combine live bacteria with other strategies.

[0005] Ultrasound-driven antibacterial sonodynamic therapy (SDT) is a treatment that kills bacteria by generating a large amount of reactive oxygen species (ROS) through ultrasound excitation of a sonosensitizer. Ultrasound has a very high tissue penetration, which can be used to destroy the structure of biofilm and treat related infections. However, the current limitations of sonosensitizers greatly hinder the widespread clinical application of SDT, one of the common limiting factors is that reactive oxygen species are easily quenched, and their action distance is very short, which makes it difficult to play a good antibacterial effect in vivo when used alone. In addition, traditional organic sonosensitizers such as indocyanine green (ICG), doxorubicin, curcumin, etc. often have the problems of low bioavailability, fast metabolism in the body, poor biofilm accumulation, etc. Inorganic sonosensitizers such as titanium dioxide (TiO2) have a relatively low quantum yield of ultrasound (US) triggered ROS generation due to the rapid combination of electrons (e) and holes (h+) (50±30 ns). Therefore, new methods are needed to improve the quantum yield of TiO2.

[0006] The purpose of the present application is to develop a new type of composite sonosensitizer and its bacterial targeting delivery system that can effectively treat Pseudomonas aeruginosa infection without damaging the body's tissues. The composite sonosensitizer bacterial targeting delivery system provided by the present application is composed of platinum-modified titanium dioxide nanoparticles loaded with organic sonosensitizer ICG and Bdellovibrio for targeted delivery of the nanoparticles. The organic-inorganic composite composite sonosensitizer (BPT-ICG) has high sonodynamic properties and excellent antibacterial effect. At the same time, Bdellovibrio acts as a targeted delivery system for the composite sonosensitizer, which delivers the composite sonosensitizer to the surface or inside of the bacteria by destroying the biofilm and the predatory characteristics of the bacteria, and then generates a large amount of reactive oxygen species under the excitation of ultrasound to quickly kill Pseudomonas aeruginosa and eradicate the biofilm. In addition, this live bacterial delivery system does not significantly harm mammalian cells and has little side effects on the body's tissues. Therefore, the composite sonosensitizer and its bacterial targeting delivery system provided by the present application are expected to be used for targeted treatment of clinically difficult-to-treat biofilm-related bacterial infections. SUMMARY

[0007] In order to overcome the shortcomings and deficiencies of the prior art, the purpose of the present application is to provide a composite sonosensitizer and a related preparation method for its bacterial targeting delivery system. Among them, the composite sonosensitizer (BPT-ICG) is composed of inorganic sonosensitizer (BPT) and organic sonosensitizer (ICG), and the composite sonosensitizer bacterial targeting delivery system (BPT-ICG@Bd) is the composite sonosensitizer nanoparticle modified by in-situ polymerization of surface dopamine on the surface of Bd (Bd). The composite sonosensitizer on the surface of Bd not only can catalyze H2O2 in the environment to produce oxygen as a nano-enzyme, but also has sonodynamic activity, which can convert oxygen into active oxygen under ultrasonic treatment, and has a certain bactericidal effect. Bd can further utilize its high-speed hunting ability to deliver the composite sonosensitizer on its surface to the surface or inside of bacteria, thereby achieving the purpose of treating Pseudomonas aeruginosa infection. In addition, Bd can effectively penetrate the biofilm structure, and the sonodynamic nanosystem can catalyze H2O2 in the biofilm to produce oxygen, relieve the hypoxic condition of the infection site, and under the action of ultrasonic, utilize the generated oxygen to generate ROS to dissolve the biofilm and kill the bacteria therein. The combination of Bd predation characteristics and sonodynamic therapy can rapidly eliminate Pseudomonas aeruginosa and biofilm in a synergistic manner without harming mammalian cells.

[0008] The term "ICG" in the present application refers to the organic sonosensitizer indocyanine green.

[0009] The term "TiO2" in the present application refers to white hollow mesoporous titanium dioxide.

[0010] The term "Pt-TiO2" in the present application refers to platinum nanoparticle modified white hollow mesoporous titanium dioxide.

[0011] The term "BPT" in the present application refers to platinum nanoparticle modified black hollow mesoporous titanium dioxide.

[0012] The term "BPT-ICG" in the present application refers to a composite sonosensitizer composed of platinum nanoparticle modified black hollow mesoporous titanium dioxide and organic sonosensitizer indocyanine green.

[0013] The term "BPT-ICG@Bd" in the present application refers to a composite sonosensitizer bacterial targeting delivery system with bacterial and biofilm targeting effect formed by modifying the composite sonosensitizer (BPT-ICG) to the surface of Bd (Bd).

[0014] One object of the present application is to provide a composite sonosensitizer composed of inorganic sonosensitizer and organic sonosensitizer, wherein the inorganic sonosensitizer is selected from one or more of titanium dioxide nanoparticles, metal-organic framework nanoparticles, and titanium carbide two-dimensional nanosheets, and the organic sonosensitizer is selected from one or more of indocyanine green, hematoporphyrin monomethyl ether, and dihydrophenanthrene.

[0015] Further, the mass ratio of the platinum nanoparticle modified black hollow mesoporous titanium dioxide to the indocyanine green is (0.2-5):1.

[0016] Further, the titanium dioxide nanoparticle is platinum nanoparticle modified black hollow mesoporous titanium dioxide.

[0017] Another object of the present application is to provide a preparation method of the composite sonosensitizer, which comprises the following steps:

[0018] S1, blending and reacting ethanol, polyvinylpyrrolidone aqueous solution, hydrochloric acid and titanium tetrafluoride aqueous solution, centrifuging after heating, and washing to obtain white hollow mesoporous titanium dioxide;

[0019] S2, dispersing the white hollow mesoporous titanium dioxide powder in water, adding chloroplatinic acid hexahydrate, stirring and reacting, then adding sodium borohydride, and reacting to obtain platinum nanoparticle modified white hollow mesoporous titanium dioxide;

[0020] S3, heating the platinum nanoparticle modified white hollow mesoporous titanium dioxide in an inert gas atmosphere to obtain the platinum nanoparticle modified black hollow mesoporous titanium dioxide;

[0021] S4, dispersing the platinum nanoparticle modified black hollow mesoporous titanium dioxide in water, ultrasonicating, adding the indocyanine green, and reacting to obtain the composite sonosensitizer.

[0022] Further, in step S2, the mass ratio of the white hollow mesoporous titanium dioxide powder to the chloroplatinic acid hexahydrate is (1:0.16-0.3).

[0023] Further, in step S3, the heating temperature is 300-700℃, and the heating time is 0.1-3h.

[0024] Further, in step S4, the reaction time is 8-24h.

[0025] Another object of the present application is to provide a bacterial-targeted delivery system encapsulating the composite sonosensitizer on the surface.

[0026] Another object of the present application is to provide applications of the bacterial-targeted delivery system in antibacterial products and treatment of bacterial infections.

[0027] The present application has the following beneficial effects:

[0028] (1) The composite sonosensitizer BPT-ICG provided by the present application comprises an organic-inorganic composite nanoparticle sonosensitizer, has high-efficiency sonodynamic properties, and has excellent antibacterial effect.

[0029] (2) The composite sonosensitizer bacterial targeting delivery system BPT-ICG@Bd provided by the application can penetrate and destroy a biofilm by using the characteristics of the Bd that can actively and rapidly prey on bacteria, and deliver a new type of sonosensitizer to the surface or inside of the bacteria, so that a large amount of active oxygen is generated under the excitation of ultrasound to rapidly kill the bacteria and eradicate the biofilm.

[0030] (3) The composite sonosensitizer bacterial targeting delivery system BPT-ICG@Bd provided by the application has strong antibacterial and anti-biofilm effects and excellent treatment effects on bacterial infections in vitro and in vivo, and the treatment effects are superior to the treatment effects of commonly used antibiotics in the clinic, and the bacteria killing by the ROS generated in response to ultrasound can prevent the bacteria from developing drug resistance.

[0031] (4) The composite sonosensitizer bacterial targeting delivery system BPT-ICG@Bd provided by the application does not significantly harm mammalian cells and has small side effects on the tissues of the body, and therefore the composite sonosensitizer BPT-ICG and the live bacterial targeting delivery system BPT-ICG@Bd thereof are expected to be used for targeted treatment of biofilm infections that are difficult to treat in the clinic. BRIEF DESCRIPTION OF DRAWINGS

[0032] Figure 1 (a) shows a preparation schematic diagram of the composite sonosensitizer bacterial targeting live bacterial delivery system BPT-ICG@Bd with bacterial targeting effect, which is formed by modifying the platinum nanoparticle-modified black hollow mesoporous titanium dioxide (BPT-ICG) containing the organic sonosensitizer indocyanine green (ICG) to the surface of Bd in Example 1; Figure 1 (b) shows a schematic diagram of the process of destroying the bacterial biofilm by BPT-ICG@Bd, in the attack stage, the Bd collides with the host and enters the bacteria, and the active oxygen ROS is generated by activating the sonosensitizer system by applying ultrasound, the biofilm is destroyed, and the bacteria therein are killed.

[0033] Figure 2 shows the characterization data of the composite sonosensitizer prepared in Example 1, wherein, Figure 2 (a)- Figure 2 (c) are transmission electron microscope images of white hollow mesoporous titanium dioxide TiO2, platinum nanoparticle-modified titanium oxide Pt-TiO2, and platinum nanoparticle-modified black hollow mesoporous titanium dioxide BPT, respectively; Figure 2 (d) is an XRD pattern of TiO2, Pt-TiO2, and BPT; Figure 2 (e) is an electron spin resonance spectrum (ESR) pattern of Pt-TiO2 and BPT; Figure 2 (f) is an N2 adsorption-desorption isotherm of BPT; Figure 2(g) Particle size distribution of Ti02, Pt-Ti02, BPT and BPT-ICG; Figure 2 (h) Zeta potential of Ti02, Pt-Ti02, BPT and BPT-ICG; Figure 2 (i) Drug release profile of BPT-ICG under different conditions (Mean ± S.D., N = 3).

[0034] Figure 3 The sonodynamic properties, mechanism and antibacterial effect of the composite sonosensitizer BPT-ICG in Example 2 are shown, wherein, Figure 3 (a) is a schematic diagram of the mechanism of ROS produced by BPT-ICG under ultrasonic (US) irradiation; Figure 3 (b) is the band diagram of Ti02, BPT; Figure 3 (c) is the comparison of the amount of O2 produced by Pt-Ti02 and BPT with the same Ti concentration catalyzing H202 in an anaerobic environment; Figure 3 (d) is the concentration change of reactive oxygen probe DPBF immediately after ultrasonic mixing of Ti02, Pt-Ti02, BPT, BPT-ICG with Ti concentration of 50 μg / mL Ti02; Figure 3 (e) is the relative absorption intensity of singlet oxygen fluorescent probe SOSG under ultrasonic irradiation immediately after ultrasonic mixing of Ti02, Pt-Ti02, BPT, BPT-ICG with Ti concentration of 50 μg / mL Ti02; Figure 3 (f) is the number of CFU after BPT-ICG of different concentrations was co-incubated with P. aeruginosa for 2 h and ultrasonic treatment for 2 min (N = 3; Means ± S.D.).

[0035] Figure 4 The characterization, antibacterial effect and predation efficiency of the bacterial targeting delivery system (BPT-ICG@Bd) of Bd surface loaded with composite sonosensitizer in Example 3 are shown, wherein, Figure 4 (a) is the colocalization picture of BPT-ICG@Bd by laser confocal microscopy (CLSM), from left to right, the green fluorescence signal of ICG in the nanoparticles, the red dye dyed Bd cell membrane and the fluorescence signal after the combination of the two; Figure 4 (b) is the predation ability of pure Bd and Bd surface modified with BPT-ICG, from left to right, the Bd is dyed green with SYTO 9, the P. aeruginosa is dyed blue with Hoechst 33342 and the predation of Bd on P. aeruginosa; Figure 4(c) From left to right, the pictures are the colony of P. aeruginosa after incubation with PBS, Bd, BPT-ICG, BPT-ICG@Bd for 2 h, and the colony of P. aeruginosa after incubation with BPT-ICG@Bd for 2 h and sonication for 2 min; Figure 4 (d) From left to right, the pictures are the colony of P. aeruginosa after incubation with PBS, Bd, BPT-ICG, BPT-ICG@Bd for 2 h, and the colony of P. aeruginosa after incubation with BPT-ICG@Bd for 2 h and sonication for 2 min;

[0036] Figure 5 Figures 6A-6E show the in vitro anti-biofilm activity of the surface-encapsulated composite sonosensitizer-targeted bacteria delivery system (BPT-ICG@Bd) in Example 4, wherein Figure 5 (a) From top to bottom, the pictures are the invasion of P. aeruginosa biofilm by BPT-ICG@Bd at 30, 60 and 120 min, respectively, as observed by laser confocal microscopy (CLSM), BPT-ICG@Bd is stained red by CY 5-NHS, and P. aeruginosa biofilm is stained green by SYTO 9; Figure 5 (b) The pictures are the laser confocal microscopy (CLSM) images of P. aeruginosa biofilm after different treatments and staining with SYTO 9 / PI; Figure 5 (c) The pictures are the relative quantification of biofilm after different treatments and staining with crystal violet; Figure 5 (d) and Figure 5 (e) The pictures are the colony of P. aeruginosa biofilm after different treatments and the CFU counting, respectively.

[0037] Figure 6 Figures 7A-7C show the in vivo therapeutic effect of BPT-ICG@Bd on P. aeruginosa-induced lung infection in mice in Example 5, wherein Figure 6 (a) The schematic diagram of modeling and treatment of P. aeruginosa-induced pneumonia infection in mice; Figure 6 (b) and Figure 6 (c) The pictures are the colony of P. aeruginosa biofilm after different treatments and the CFU counting, respectively. 7 PFU / mL; BPT-ICG: intratracheal aerosol injection of 40 μL BPT-ICG 50 μg / mL, sonication for 5 min after 2 h; BPT-ICG@Bd: intratracheal aerosol injection of BPT-ICG@Bd (40 μL, 10 7PFU / mL Bd, 50 μg / mL BPT-ICG, 100 μL), 5 min ultrasound treatment 2 h later; CAZ: quantification of the number of bacteria in the lungs and survival curve over 14 days after treatment with cefazidime (30 mg / kg) injected in the tail vein.

[0038] Figure 7 Figures showing the study of the therapeutic effect of BPT-ICG@Bd on P. aeruginosa-induced rat osteomyelitis in Example 6, wherein, Figure 7 (a) is a schematic diagram of the modeling and treatment of P. aeruginosa-induced rat osteomyelitis; Figure 7 (b) and Figure 7 (c) are, respectively, pictures of colonies in the bone marrow and the number of CFU after treatment of infected rats with different samples (Con: no treatment; US: injection of PBS in situ, 5 min ultrasound treatment 2 h later; CAZ: cefazidime (CAZ) 30 mg / kg injected in the tail vein; BPT-ICG@Bd: BPT-ICG@Bd (10 7 PFU / mL Bd, 50 μg / mL BPT-ICG, 100 μL), 5 min ultrasound treatment 2 h later; CAZ: quantification of the number of bacteria in the lungs and survival curve over 14 days after treatment with cefazidime (30 mg / kg) injected in the tail vein. Figure 7 (d) is a graph showing the weight change of rats in each group during the treatment period. DETAILED DESCRIPTION

[0039] In order to more clearly illustrate the technical solutions of the present application, the following examples are listed. The raw materials and processing methods appearing in the examples are all common raw materials on the market and technical means well known to those skilled in the art, unless otherwise stated. The present application will be further specifically and in detail described below in combination with specific examples, but the embodiments of the present application are not limited thereto.

[0040] Reagents: Indocyanine green (ICG) was purchased from Shanghai Yuan Ye Biological Co., Ltd.; titanium tetrafluoride (TiF4) was purchased from Sigma-Aldrich Co.; concentrated hydrochloric acid (HCl) and anhydrous ethanol were purchased from Guangzhou Chemical Reagent Factory; chloroplatinic acid hexahydrate (H2PtCl6·6H2O) and sodium borohydride (NaBH4, 98%) were purchased from Shanghai Aladdin Reagent Co., Ltd.; polyvinylpyrrolidone (PVP-K30, M w = 40,000) were purchased from Solvay; and the test water was deionized water.

[0041] Strains: Pseudomonas aeruginosa strain numbered ATCC 27853, from American Type Culture Collection (ATCC, USA); Bdellovibrio bacteriovorus HD 100 strain numbered DSM 50701, from Deutsche Sammlung von Mikroorganismen und Zellkulturen (DSMZ, Germany).

[0042] Experimental animals: BALB / c mice (6-8 weeks, female, body weight 18-20 grams, SPF level) were used as the animal experimental objects of lung infection in this study, purchased from Zhuhai Boshitong Biotechnology Co., Ltd., Animal Production License No.: SCXK (Yue) 2020-0051, Animal Qualification Certificate No.: No. 44822700015055; SD rats (male, body weight 200-250 grams, SPF level) were used as the animal experimental objects of osteomyelitis in this study, purchased from Zhuhai Boshitong Biotechnology Co., Ltd., Animal Production License No.: SCXK (Yue) 2020-0051, Animal Qualification Certificate No.: No. 44822700016227.

[0043] Example 1: Preparation of a composite sonosensitizer (BPT-ICG)

[0044] S1, 55.2 mL of anhydrous ethanol, 8 mL of polyvinylpyrrolidone (PVP) aqueous solution (9.75 mg / mL, M w = 40,000), 500 μL of HCl (50 mM) and 5 mL of TiF4(40 mM) aqueous solution were mixed and stirred at room temperature for 1 h. Then it was poured into a 100 mL stainless steel reactor, heated at 180°C for 3 h. Centrifugal collection, washed with pure water and anhydrous ethanol for 3 times, to obtain white hollow mesoporous titanium dioxide (TiO2).

[0045] S2, the dried TiO2powder (50 mg) was dispersed in 50 mL of deionized water, and ultrasonicated in water bath for 15 min to make it uniformly dispersed. 1 mL of H2PtCl6·6H2O (10 mM) was added to the previous TiO2dispersion, and stirred at room temperature for 1 h. After adding 2 mL of 4 mg / mL NaBH4under vigorous stirring, continue to stir for 3 h. Centrifugal collection, washed with pure water for 3 times, to obtain platinum nanoparticle modified white hollow mesoporous titanium dioxide (Pt-TiO2).

[0046] S3, the synthesized Pt-TiO2 was transferred into a tube furnace, and was heated to 500℃ at 5℃ / min in the atmosphere of hydrogen:nitrogen=1:9, and was kept for 1 h. When the reaction was completed and the temperature was recovered to room temperature, BPT was obtained, and was taken out and sealed in vacuum for preservation.

[0047] S4, 10 mg of BPT nanoparticles was dispersed in 10 mL of deionized water, and was uniformly dispersed by water bath ultrasonic for 15 min. 10 mg of indocyanine green (ICG) was added into the BPT dispersion, and was stirred at room temperature for 12 h. After centrifugation, pure water was used for washing until the supernatant was colorless to remove free ICG, and the precipitate, i.e. BPT-ICG, was collected, and the supernatant of each washing was collected.

[0048] The loading amount was determined by the characteristic absorbance of the ICG aqueous solution at 780 nm wavelength, and the loading efficiency (LE) could be obtained by the following formula: LE (%)=(m 总 -m 上清 ) / (m BPT +m 总 -m 上清 )×100%; the phase analysis of TiO2, Pt-TiO2 and BPT samples was performed by using an X-ray powder diffractometer of SmartLab produced by Rigaku Company of Japan; the electron spin resonance spectrum (ESR) data were obtained by using an electron spin resonance spectrometer of A300-10 / 12 produced by Bruker Company of Germany; the N2 adsorption-desorption isotherm data of BPT were obtained by using an analyzer of ASAP2460 produced by Micromeritics Company of the United States; and the particle size and Zeta potential data were obtained by using a Zetasizer Pro (Malvern) instrument.

[0049] Figure 2 The characterization data of the composite photosensitizer prepared in Example 1 are shown, wherein, Figure 2 (a)- Figure 2 (c) are the transmission electron microscope images of white hollow mesoporous titanium dioxide TiO2, platinum nanoparticle modified titanium oxide Pt-TiO2 and platinum nanoparticle modified black hollow mesoporous titanium dioxide BPT, respectively; Figure 2 (d) is the XRD pattern of TiO2, Pt-TiO2 and BPT; Figure 2 (e) is the electron spin resonance spectrum (ESR) pattern of Pt-TiO2 and BPT; Figure 2 (f) is the N2 adsorption-desorption isotherm of BPT; Figure 2 (g) is the particle size distribution of TiO2, Pt-TiO2, BPT and BPT-ICG; Figure 2 (h) is the Zeta potential of TiO2, Pt-TiO2, BPT and BPT-ICG;Figure 2 (i) shows the drug release curves of BPT-ICG under different conditions (Mean±SD, N=3).

[0050] Morphological characterization revealed that the obtained TiO2, Pt-TiO2, and BPT nanoparticles were hollow spherical with an average diameter of approximately 100 nm. Figure 2 (a)- Figure 2 (c)). From Figure 2 (a)- Figure 2 (c) It can also be seen that Pt nanoparticles are deposited on the surface of Pt-TiO2 and BPT nanoparticles, and maintain good dispersion. High-temperature hydrogenation reduction enables the Pt nanoparticles to migrate and distribute uniformly on the TiO2 surface. XRD was used to characterize the changes in the main crystal structure of Pt-TiO2 nanoparticles before and after hydrogenation reduction. Figure 2 (d) As shown in the figure, a distinct Pt peak appears when Pt nanoparticles are deposited on TiO2. The reduced titanium dioxide still corresponds to the anatase type (JCPDS 21-1272), indicating that no phase transition occurred during hydrogenation reduction. This is confirmed by the ESR spectrum ( Figure 2 (e) This demonstrates the successful preparation of oxygen vacancies in BPT nanoparticles. Furthermore, N2 adsorption / desorption was used to detect the change in BET surface area, and a typical type IV isotherm confirmed the hollow structure of BPT. Figure 2 (f) , and it can be seen that the reduction process did not significantly damage the nanoparticle structure. The hollow structure of the prepared TiO2 makes it a potential drug carrier. Its drug loading properties were evaluated using indocyanine green (ICG), approved by the US Food and Drug Administration (FDA). Based on the characteristic absorbance of the ICG aqueous solution at 780 nm, the ICG loading efficiency in BPT was calculated to be 45%. The change in zeta potential confirmed the success of the stepwise synthesis, with the zeta potential of BPT-ICG being -22.3 mV ( Figure 2 (g)), the hydrated particle size is approximately 166.1 nm. Figure 2 (h)). The release behavior of indocyanine green was investigated in phosphate-buffered saline (PBS) at different pH values. Figure 3 (i) Both a decrease in pH and ultrasound can increase the release of indocyanine green. This may be because, on the one hand, a lower pH reduces the electrostatic adsorption of indocyanine green and BPT nanoparticles, and on the other hand, ultrasound cavitation promotes the release of indocyanine green from the nanoparticle cavities. Both of these results will lead to an accelerated release of indocyanine green.

[0051] Example 2: Study on the sonodynamic properties and antibacterial activity of BPT-ICG against Pseudomonas aeruginosa

[0052] The sonodynamic properties and antibacterial effect of BPT-ICG prepared in Example 1 on P. aeruginosa were studied in this example.

[0053] The antibacterial effect of BPT-ICG on P. aeruginosa was detected by dilution plate method. P. aeruginosa was dispersed in PBS after 12 h of culture. Bacteria (10 8 CFU / mL, 400 μL) were mixed with BPT-ICG nanoparticles (100 μL) with different Ti concentrations (0, 25, 50, 100 μg / mL final concentration) respectively. Ultrasonic instrument was used for ultrasonic treatment for 2 min, and the ultrasonic parameters were as follows: frequency 1.0 MHz, 50% duty cycle, power 1.5 W cm -2 . Among them, Con was the control group without adding BPT-ICG nanoparticles and without ultrasonic treatment, and US was the ultrasonic group with Ti final concentration of 0 μg / ml.

[0054] The band data of TiO2 and BPT were tested by X-ray photoelectron spectrometer (ESCALAB 250Xi, Thermo, USA).

[0055] Dissolved oxygen meter (JPSJ-605F, Lei-ci, Shanghai, China) was used to detect the content of O2 generated by H2O2 decomposition by Pt-TiO2 and BPT. At room temperature, 1 mL of Pt-TiO2 and BPT with Ti concentration of 50 μg / mL was added to 200 μM of H2O2 solution. The dissolved oxygen data was recorded every 10 s.

[0056] DPBF is a ROS detection probe. 50 μL of DPBF DMSO solution (1 mg / mL) was added to TiO2, Pt-TiO2, BPT and BPT-ICG solutions with Ti concentration of 50 μg / mL. After mixing evenly, the mixture was irradiated by ultrasonic treatment at fixed time intervals (ultrasonic parameters: 1.5 W cm -2 , 1.0 MHz, 50% duty cycle). The concentration change of DPBF was recorded by ultraviolet spectrophotometer (YOKE T2602, Shanghai, China).

[0057] SOSG is a fluorescent molecular probe widely used to detect 1 O2. 1 μL of SOSG methanol solution (5 mM) was mixed with TiO2, Pt-TiO2, BPT and BPT-ICG solutions with Ti concentration of 50 μg / mL. The mixture was irradiated by ultrasonic treatment at fixed time intervals (ultrasonic parameters: 1.5 W cm -21.0 MHz, 50% duty cycle). The fluorescence intensity of SOSG was measured by a fluorescence spectrophotometer.

[0058] Figure 3 The sonodynamic properties, mechanism and antibacterial effect of the composite sonosensitizer BPT-ICG in Example 2 are shown, wherein, Figure 3 (a) is a schematic diagram of the mechanism of ROS generated by BPT-ICG under ultrasonic (US) irradiation; Figure 3 (b) is the energy band diagram of TiO2, BPT; Figure 3 (c) is the amount of O2 generated by Pt-TiO2 and BPT with the same Ti concentration catalyzing H2O2 in an anaerobic environment; Figure 3 (d) is the concentration change of reactive oxygen probe DPBF immediately after ultrasonic mixing of TiO2, Pt-TiO2, BPT, BPT-ICG with a Ti concentration of 50 μg / mL TiO2; Figure 3 (e) is the relative absorption intensity of singlet oxygen fluorescence probe SOSG under ultrasonic irradiation immediately after ultrasonic mixing of TiO2, Pt-TiO2, BPT, BPT-ICG with a Ti concentration of 50 μg / mL TiO2; Figure 3 (f) is the number of CFU after co-incubation of BPT-ICG with different concentrations and P. aeruginosa for 2 h and ultrasonic treatment for 2 min (N=3; Means±S.D.).

[0059] The BPT nanoparticles trigger the separation of e - and h + pairs under ultrasonic irradiation, and the generated e - reacts with O2 to generate ·O2 - , which is consumed to produce toxic 1 O2; at the same time, the generated h + reacts with H2O to generate ·OH( Figure 3 (a)). The energy band gap of TiO2 is 3.0-3.2 eV( Figure 3 (b)), which is not enough to achieve effective separation of e- and h+ pairs, resulting in limited ROS production. Compared with TiO2 (3.11 eV), the band gap of BPT is narrower, about 2.24 eV, which is more conducive to the separation of e - and h + pairs under US irradiation( Figure 3 (b)). In summary, compared with TiO2, BPT has stronger ability to promote the separation of e - and h + pairs, thereby generating higher levels of 1 O2 and ·OH.

[0060] BPT as a sonosensitizer for sonodynamic therapy (SDT) shows great prospects. Oxygen is essential for BPT to produce ROS because it is the raw material for the production of ROS. BPT has nanocatalytic activity and can catalyze the production of oxygen from H2O2 in the environment. Under the same Ti concentration conditions, the oxygen production efficiency of BPT is about 1.5 times that of Pt-TiO2, mainly because the specific surface area of platinum nanoparticles increases after reduction, and there are more active sites for catalysis. In order to verify whether BPT has stronger SDT activity than TiO2 and Pt-TiO2, 1,3-diphenyl isobenzofuran (DPBF) probe was used to detect the SDT effect because DPBF can react with the generated ROS, causing its ultraviolet characteristic absorption at a wavelength of about 420 nm to decrease. TiO2, Pt-TiO2 and BPT were treated under the same conditions. As expected, BPT can produce the most ROS Figure 3 (c)). This conclusion shows that the deposition of Pt nanoparticles and the preparation of oxygen vacancies are more conducive to the separation of electrons and holes to produce more ROS. Due to the introduction of the organic sonosensitizer indocyanine green, BPT-ICG can produce more 1 O2( Figure 3 (d)). In addition, the production of 1 O2 under different conditions was characterized by the singlet oxygen fluorescence probe SOSG. With the increase of ultrasonic time, the fluorescence signal intensity of SOSG also increases, indicating that TiO2, Pt-TiO2, BPT and BPT-ICG can all produce 1 O2, the conclusion is consistent with the DPBF experiment, and BPT-ICG produces 1 O2 with the highest efficiency Figure 3 (e)). After verifying the ROS production ability of BPT-ICG under ultrasonic irradiation, we further evaluated the sonodynamic antibacterial performance of the material. Pseudomonas aeruginosa (P. aeruginosa) was selected as a model of gram-negative bacterial strain. According to the residual colony forming units (CFU) of P. aeruginosa after treatment, BPT-ICG nanostructure shows significant killing of P. aeruginosa under ultrasonic conditions, and the sonodynamic bactericidal efficiency gradually increases with the increase of the concentration of nano-composite Figure 4 (f).

[0061] Example 3: Preparation of a composite sonosensitizer bacteria-targeted delivery system

[0062] Bdellovibrio (1 x 10 8BPT-ICG (50 pg / mL, 100 pL) and BPT-ICG@Bd (10 PFU / mL Bd, 50 pg / mL BPT-ICG, 100 pL). The ultrasonic parameters were as follows: frequency 1.0 MHz, 50% duty cycle, power 1.5 W cm2. The ultrasonic instrument was used for 2 min. The mixture was centrifuged at 6500 rpm / min to collect BPT-ICG@Bd, which was washed twice with PBS. The collected BPT-ICG@Bd was resuspended in PBS and stored at 4 °C for further use.

[0063] To observe whether BPT-ICG was successfully loaded on Bd, the Bd membrane in BPT-ICG@Bd was dyed red with FM 4-64, and BPT-ICG emitted green fluorescence because it carried ICG. The co-localization of Bd and BPT-ICG was observed by laser confocal microscopy (CLSM).

[0064] Bd, BPT-ICG@Bd were dyed green with SYTO 9, and P. aeruginosa was dyed blue with Hoechst 33342. Subsequently, Bd, BPT-ICG@Bd and P. aeruginosa were co-incubated for 15 min, and the predation of Bd and BPT-ICG@Bd on P. aeruginosa was observed by laser confocal microscopy (CLSM).

[0065] P. aeruginosa was dispersed in PBS after 12 h of culture. Bacteria (10 8 CFU / mL, 400 pL) were mixed with PBS (100 pL), Bd (10 7 PFU / mL, 100 pL), BPT-ICG (50 pg / mL, 100 pL), BPT-ICG@Bd (10 7 PFU / mL Bd, 50 pg / mL BPT-ICG, 100 pL), respectively. The ultrasonic instrument was used for 2 min. The ultrasonic parameters were as follows: frequency 1.0 MHz, 50% duty cycle, power 1.5 W cm2. -2 The antibacterial effect of BPT-ICG@Bd on P. aeruginosa was detected by the dilution plate method.

[0066] Figure 4 Characterization, antibacterial effect and predation efficiency of the Bd surface-loaded composite sonosensitizer bacterial targeting delivery system (BPT-ICG@Bd) in Example 3 are shown, wherein, Figure 4(a) Laser confocal microscopy (CLSM) co-localization images of BPT-ICG@Bd from left to right are green fluorescence signal of ICG in nanoparticles, red dye stained Bd cell membrane and fluorescence signal after the combination of the two; Figure 4 (b) Predation ability of pure Bd and Bd surface modified with BPT-ICG from left to right are Bd stained with SYTO 9 in green, P. aeruginosa stained with Hoechst 33342 in blue and predation of Bd on P. aeruginosa; Figure 4 (c) From left to right are phosphate buffer solution (PBS), Bd, BPT-ICG, BPT-ICG@Bd and P. aeruginosa co-incubated for 2 h, and colony pictures after ultrasonic treatment for 2 min; Figure 4 (d) From left to right are phosphate buffer solution (PBS), Bd, BPT-ICG, BPT-ICG@Bd and P. aeruginosa co-incubated for 2 h, and the number of bacterial colonies (CFU) after ultrasonic treatment for 2 min.

[0067] Figure 4 (a) shows Bd labeled with membrane dye FM 4-64, and the successful preparation of BPT-ICG@Bd is verified by laser confocal co-localization using the spontaneous fluorescence of BPT-ICG. In order to study whether the surface modification of nanoparticles on Bd will affect its predation on prey, Bd is stained with SYTO 9 in green, P. aeruginosa is stained with Hoechst 33342 in blue, and laser confocal microscopy is used to observe the predation difference of Bd and BPT-ICG@Bd groups on P. aeruginosa. The results show that the surface modified nanoparticles do not significantly affect the predation ability of Bd on P. aeruginosa Figure 4 (b).

[0068] Due to the natural targeting predation ability of Bd on P. aeruginosa, the interaction between the surface modified nanoparticles and bacteria is greatly enhanced, and the action distance of ROS generated under ultrasound is reduced. Therefore, BPT-ICG@Bd can produce the strongest sonodynamic antibacterial effect under ultrasound Figure 4 (c- Figure 5 (d).

[0069] Example 4: Study on antibacterial biofilm of BPT-ICG@Bd

[0070] This example studies the antibacterial biofilm of the composite sonosensitizer bacteria-targeted delivery system BPT-ICG@Bd prepared in Example 3.

[0071] P. aeruginosa biofilm was stained green with SYTO 9, and then the supernatant was removed and washed with PBS three times. Cy5-labeled BPT-ICG@Bd (10 7 PFU / mL Bd, 50 μg / mL BPT-ICG, 1 mL) was incubated at 37 °C for different times. Next, the supernatant was removed and the biofilm was rinsed with PBS three times to remove excess BPT-ICG@Bd. The location of BPT-ICG@Bd in the biofilm was observed by laser confocal microscopy (CLSM).

[0072] P. aeruginosa (10 8 CFU / mL) was cultured in Mueller-Hinton Broth (MHB) medium for 24 h to form a biofilm. PBS (1 mL), Bd (10 7 PFU / mL Bd, 1 mL), BPT-ICG (50 μg / mL, 1 mL), BPT-ICG@Bd (10 7 PFU / mL Bd, BPT-ICG 50 μg / mL BPT-ICG, 1 mL) were co-incubated for 2 h, and then after 2 min of ultrasound, the remaining biofilm was stained with SYTO 9 / PI and observed by laser confocal microscopy (CLSM). For crystal violet staining, after the plate was rinsed with sterile PBS (pH 7.4) and dried for 10 min, it was stained with 0.01% (w / v) crystal violet for 30 min, and then anhydrous ethanol was added to dissolve the crystal violet, and the absorbance at 570 nm was measured by a microplate reader to determine the biofilm biomass. For biofilm bacterial number determination, PBS (500 μL) was added to the biofilm of different treatment groups, and the biofilm was dispersed by vortexing, and the number of biofilm bacteria was determined by the dilution plate method.

[0073] Figure 5 In vitro anti-biofilm activity of the surface-packaged composite sonosensitizer delivery system (BPT-ICG@Bd) of Example 4 is shown, wherein, Figure 5 (a) is the invasion of P. aeruginosa biofilm by BPT-ICG@Bd observed by laser confocal microscopy (CLSM) at different incubation times, from top to bottom, the time of BPT-ICG@Bd invasion of P. aeruginosa biofilm is 30, 60 and 120 min, respectively, BPT-ICG@Bd is stained red by CY 5-NHS, and P. aeruginosa biofilm is stained green by SYTO 9; Figure 5 (b) is the laser confocal microscopy (CLSM) image of P. aeruginosa biofilm stained with SYTO 9 / PI after different treatments; Figure 5(c) the relative quantification of biofilm stained with crystal violet after different treatments; Figure 5 (d) and Figure 5 (e) the pictures of bacterial colonies and CFU number in P. aeruginosa biofilm after different treatments, respectively.

[0074] Bacterial biofilm is a membrane-like structure formed by a hydrated matrix of biological macromolecules (such as polysaccharides and proteins) secreted by bacteria, which encapsulates various bacteria, and significantly reduces the sensitivity of bacteria to antibiotics and immune attacks of the immune system, so it is extremely difficult to eradicate biofilm in vivo. To investigate the anti-biofilm activity of BPT-ICG@Bd, the ability of BPT-ICG@Bd to penetrate the biofilm was first studied. Figure 5 (a)) By laser confocal microscopy, it was found that after 30 min of BPT-ICG@Bd treatment, red fluorescence was dispersed on the surface of the biofilm, and after 2 h, it diffused in the biofilm, indicating that BPT-ICG@Bd still retained the motility of Bd and could move in the bacterial exopolysaccharide matrix of the biofilm. BPT-ICG@Bd can completely penetrate the biofilm of about 40 μm thick within 2 hours Figure 5 (a)).

[0075] Next, the clearance effect of BPT-ICG@Bd on P. aeruginosa biofilm was explored. After different treatments, the biofilm was stained with a live / dead staining kit and observed by laser confocal microscopy for its integrity Figure 5 (b)) Due to the physical destruction of ultrasound and the predatory ability of Bd, the ultrasound (US) group and the Bd group can destroy the biofilm structure to a certain extent, but the red fluorescence is weak, indicating that it is difficult to kill the bacteria in the biofilm, while BPT-ICG@Bd can significantly disintegrate the bacterial biofilm by combining Bd, physical destruction of ultrasound and ROS produced, and show a strong red fluorescence signal, indicating that BPT-ICG@Bd can not only destroy the biofilm, but also target and kill the bacteria inside the biofilm Figure 5 (b)) By quantitatively analyzing the biofilm stained with crystal violet Figure 5 (c)), the US group, the Bd group and the BPT-ICG@Bd group can remove 46.13%, 33.24% and 30.85% of the biofilm, respectively, while under the action of ultrasound, the BPT-ICG@Bd+US group can remove most of the biofilm (about 88.71%). Figure 6 (d)- Figure 6(e)) Compared with the control group, the BPT-ICG@Bd+US group can reduce the number of bacteria in the biofilm by about 4.62 log, which is 2.56 times the effect of the free nanoparticle BPT-ICG group, indicating that BPT-ICG@Bd combined with US stimulation has excellent effect on removing bacteria in the biofilm.

[0076] Example 5: In vivo antibacterial effect of BPT-ICG@Bd against lung infection

[0077] In this example, the in vivo antibacterial effect of the composite sonosensitizer bacteria-targeted delivery system BPT-ICG@Bd prepared in Example 3 against lung infection was studied.

[0078] The mice were randomly divided into 6 treatment groups: PBS, US, Bd, BPT-ICG, BPT-ICG@Bd, and CAZ groups, with 6 mice in each group. After the mice were anesthetized, 5x10 6 CFU of P. aeruginosa was inoculated intratracheally, and 24 h after infection, PBS (40 μL), Bd (40 μL, 107 PFU / mL), BPT-ICG (40 μL, 75 μg / mL), and BPT-ICG@Bd (40 μL, 10 7 PFU / mL Bd, 75 μg / mL BPT-ICG) were given intratracheally. The CAZ group was given 60 mg / kg of ceftazidime intravenously. The US group, the BPT-ICG group, or the BPT-ICG@Bd group was subjected to ultrasound for 5 min (ultrasound parameters: 1.5 W cm -2 , 1.0 MHz, 50% duty cycle) 2 h after administration. After treatment, all mice were euthanized, and the lungs of the mice were collected for bacterial load quantification.

[0079] Figure 6 The in vivo therapeutic effect of BPT-ICG@Bd on P. aeruginosa-induced lung infection in mice in Example 5 is shown, wherein Figure 6 (a) is a schematic diagram of modeling and treatment of P. aeruginosa-induced pneumonia-infected mice; Figure 6 (b) and Figure 6 (c) are respectively the bacterial counting method for determining the infected mice after different treatment groups (Con: no treatment; US: intratracheal aerosol injection of PBS (40 μL), and ultrasound treatment for 5 min 2 h later; Bd: intratracheal aerosol injection of Bd 40 μL 10 7 PFU / mL; BPT-ICG: intratracheal aerosol injection of 40 μL of 50 μg / mL BPT-ICG, and ultrasound treatment for 5 min 2 h later; BPT-ICG@Bd: intratracheal aerosol injection of 40 μL of BPT-ICG@Bd (10 7PFU / mL Bd, 50μg / mL BPT-ICG), 2h followed by 5min of sonication; CAZ: quantitative analysis of lung bacterial count and 14-day survival curve after tail vein injection of ceftazidime 30mg / kg.

[0080] To investigate the in vivo antibacterial activity of BPT-ICG@Bd, we also evaluated its therapeutic effect on Pseudomonas aeruginosa-induced lung infection. A Pseudomonas aeruginosa-induced pneumonia model was first established using BALB / c mice. Figure 6 (a)). Each mouse was intratracheally inoculated with 4 × 10 6 CFU of Pseudomonas aeruginosa (40 μL, 10) 8 CFU / mL), fed for 24 hours, to establish a pneumonia infection model ( Figure 6 (a)). Subsequently, PBS (US group), Bd, BPT-ICG, or BPT-ICG@Bd were administered via intratracheal spray. The US group or BPT-ICG@Bd+US group underwent sonication (1.5W cm) 2 hours after administration. -2 (1.0 MHz, 50% duty cycle, 5 min). Immediately after treatment, lung tissue was collected and homogenized. The homogenized solution was then serially diluted, and 50 μL of the homogenate was homogenized onto CN agar plates. Figure 6 As shown in (b), the bacterial count in the BPT-ICG@Bd group was reduced by nearly three orders of magnitude compared to the negative control (Con group), and significantly lower than that in the BPT-ICG group. Furthermore, we also monitored the survival rate of mice after infection. Figure 7 (c) More importantly, mice treated with BPT-ICG@Bd had a 100% survival rate during the 14-day study period, in stark contrast to all untreated mice that died within three days, and mice in other treatment groups with survival rates below 80%. Our conclusion is that the prolonged pulmonary retention and targeting properties of BPT-ICG@Bd significantly improved survival compared to the control group. Subsequently, we compared the efficacy of BPT-ICG@Bd with conventional intravenous (IV) administration of a clinically relevant dose (1.2 mg per mouse) of ceftazidime (CAZ). Although no mice died from either treatment, mice treated with BPT-ICG@Bd had a lower bacterial load in their lungs than those treated with CAZ. Figure 7 (b)) The surface exhibits better therapeutic efficacy. Moreover, due to its antibacterial mechanism, this sonosensitive agent delivery system holds promise for treating bacterial infections resistant to the antibiotic CAZ, which is difficult to treat with CAZ.

[0081] Example 6: In vivo antibacterial effect of BPT-ICG@Bd against osteomyelitis

[0082] The in vivo antibacterial effect of the composite sonosensitizer targeted live bacteria delivery system BPT-ICG@Bd prepared in Example 3 against osteomyelitis was studied in this example.

[0083] SD rats (200-250 g) were purchased from Zhuhai Baitishun Biotechnology Co., Ltd. and used in accordance with the guidelines approved by the GIBH Experimental Animal Welfare and Ethics Committee. After the rats were anesthetized, the hair on the hind legs was removed, and the skin was disinfected with iodophor. The tibial plateau was exposed with a scalpel, and a 1.6 mm diameter cortical bone defect was formed with a skull drill. Then, 100 μL of Pseudomonas aeruginosa suspension (10 8 CFU / mL) was injected into the medullary canal through the bone defect to establish a model of osteomyelitis. For different groups of treatment, PBS or BPT-ICG@Bd was injected into the medullary canal, or cefazolin (CAZ) was injected intravenously. Finally, the bone defect was sealed with bone wax, disinfected and then sutured. The Con group was injected with 100 μL of PBS; the US group was injected with 100 μL of PBS and treated with ultrasound for 5 minutes after 2 hours; the CAZ group was injected intravenously with CAZ (30 mg / kg); and the BPT-ICG@Bd group was injected with 100 μL of BPT-ICG@Bd (10 7 PFU / mL Bd, 50 μg / mL BPT-ICG), and treated with ultrasound for 5 minutes after 2 hours. All ultrasound parameters were 1.5 W cm -2 , 1.0 MHz, 50% duty cycle.

[0084] After treatment, bone marrow tissue was collected for 10-fold gradient dilution, and 100 μL of homogenate was plated on CN medium solid plates to evaluate the in vivo antibacterial activity.

[0085] Figure 7 The treatment of BPT-ICG@Bd on Pseudomonas aeruginosa-induced rat osteomyelitis in Example 6 is shown, wherein, Figure 7 (a) is a schematic diagram of the modeling and treatment of Pseudomonas aeruginosa-induced rat osteomyelitis; Figure 7 (b) and Figure 7 (c) are, respectively, the colony pictures in the bone marrow of infected rats after treatment with different samples (Con: no treatment; US: injection of PBS in situ, and treatment with ultrasound for 5 minutes after 2 hours; CAZ: intravenous injection of cefazolin (CAZ) 30 mg / kg; BPT-ICG@Bd: injection of BPT-ICG@Bd (10 7 PFU / mL Bd, 50 μg / mL BPT-ICG) in situ, and treatment with ultrasound for 5 minutes after 2 hours) and the CFU number change diagram; Figure 7 (d) is a diagram of the body weight change of rats in each group during the treatment period.

[0086] To evaluate the therapeutic effect of BPT-ICG@Bd on P. aeruginosa biofilm-related infection in vivo, we further constructed a P. aeruginosa-induced osteomyelitis SD rat model Figure 7 (a)) First, 1 x 10 7 CFU of P. aeruginosa (100 μL, 10 8 CFU / mL) was inoculated into the tibial plateau of each mouse, and the mice were fed for 120 h to establish the osteomyelitis infection model. Subsequently, PBS or BPT-ICG@Bd was added to the intramedullary canal, and cefazidime (CAZ) was injected intravenously as a positive control. The US group was injected with 100 μL of PBS, and the BPT-ICG@Bd group was injected with 100 μL of BPT-ICG@Bd (10 7 PFU / mL, 50 μg / mL of BPT-ICG), and 2 h later, the mice were treated with ultrasound for 5 min (1.5 W cm -2 , 50% duty cycle, 1.0 MHz). The CAZ group was injected intravenously with cefazidime (30 mg / kg). After treatment, the bone marrow tissue was collected and gradient diluted, and 100 μL of the homogenate was plated on CN medium solid plates. As shown in Figs. Figure 7 (b)- ​ (c), after two treatments on days 0 and 7, the number of bacteria treated with BPT-ICG@Bd was reduced by three orders of magnitude compared with the control, and the antibacterial effect was significantly stronger than that of the positive control CAZ group. ​ (d) The weight change of the rats since the treatment period was recorded, and the weight of the BPT-ICG@Bd group showed a healthy growth trend, close to that of the normal group. However, the growth rate of the other groups was significantly lower than that of the normal group, indicating that bacterial infection had a significant inhibitory effect on their growth.

[0087] It is apparent for those skilled in the art that the present application is not limited to the details of the foregoing exemplary embodiments, and that the present application can be implemented in other particular forms without departing from the spirit or essential characteristics of the present application. Therefore, the embodiments should be considered in all respects as illustrative and not restrictive, and the scope of the present application should be defined by the appended claims rather than the above description, and it is intended to include all changes falling within the meaning and range of equivalents of the claims.

[0088] In addition, it should be understood that although the present specification is described in terms of embodiments, not every embodiment contains only one independent technical solution, and the specification is described in this way only for the sake of clarity, and those skilled in the art should consider the specification as a whole, and the technical solutions in each embodiment can be appropriately combined to form other embodiments that those skilled in the art can understand.

Claims

1. A bacterial-targeted delivery system for a composite acoustic sensitizer, characterized in that, The composite acoustic sensor is composed of inorganic and organic acoustic sensories, wherein, The inorganic acoustic sensitizer is selected from titanium dioxide nanoparticles, and the organic acoustic sensitizer is selected from indocyanine green. The mass ratio of the inorganic acoustic sensitizer to the organic acoustic sensitizer is 1:1; The titanium dioxide nanoparticles are black hollow mesoporous titanium dioxide modified with platinum nanoparticles. The preparation method of the composite acoustic sensor includes the following steps: S1. Ethanol, polyvinylpyrrolidone aqueous solution, hydrochloric acid and titanium tetrafluoride aqueous solution were mixed and reacted, heated and collected by centrifugation, and washed to obtain white hollow mesoporous titanium dioxide. S2. Disperse the white hollow mesoporous titanium dioxide powder in water, add chloroplatinic acid hexahydrate, stir and react, then add sodium borohydride, and after the reaction, obtain white hollow mesoporous titanium dioxide modified with platinum nanoparticles. S3. The white hollow mesoporous titanium dioxide modified with platinum nanoparticles is heated in an inert gas atmosphere to obtain black hollow mesoporous titanium dioxide modified with platinum nanoparticles. S4. Disperse the platinum nanoparticle-modified black hollow mesoporous titanium dioxide in water, sonicate, add the indocyanine green, and react to obtain the composite sound-sensing agent. In step S2, the mass ratio of white hollow mesoporous titanium dioxide powder to chloroplatinic acid hexahydrate is 1:(0.16~0.3); The bacterial-targeted delivery system for the composite acoustic sensitizer involves modifying the composite acoustic sensitizer on the surface of Bdellovibrio bacteria via in-situ polymerization of surface dopamine.

2. The bacterial targeted delivery system according to claim 1, characterized in that, In step S3, the heating temperature is 300-700 ℃ and the heating time is 0.1-3 h.

3. The bacterial targeted delivery system according to claim 1, characterized in that, In step S4, the reaction time is 8-24 h.

Images