Metal single-atom modified two-dimensional defect type tungsten oxide nanomaterial, preparation method thereof and ultrasonic dynamic antitumor treatment application

Abstract

The application discloses a kind of metal single atom modified two-dimensional defect type tungsten oxide nanomaterial and its preparation method and ultrasonic power antitumor treatment application.Prepare two-dimensional WO 3‑x Nanosheet with oxygen defect under nitrogen atmosphere by calcination method, then uniformly deposit metal single atom on its surface by atomic deposition technology, prepare different metal single atom modified defect type tungsten oxide nanomaterial.The synergistic effect between metal single atom on the surface of the material and oxygen defect has superior catalytic activity for ultrasonic wave induced reactive oxygen species (ROS) generation, and the generated ROS can quickly kill cancer cells.Compared with traditional ultrasonic sensitizers, the synergistic effect between metal single atom and oxygen defect rich tungsten oxide in the material of the application can construct more active ultrasonic sensitizer, so as to be applied to high-performance ultrasonic power tumor treatment.

Description

Technical Field

[0001] This invention relates to the field of nanomaterials technology, specifically to a two-dimensional defective tungsten oxide nanomaterial modified with a single metal atom, its preparation method, and its application in ultrasonic-driven antitumor therapy. Background Technology

[0002] Currently, sonicated ultrasound therapy (SDT) is an emerging cancer treatment method. Its principle is based on the generation of reactive oxygen species (ROS) with strong oxidizing capabilities by an ultrasound sensitizer under the influence of ultrasound waves, thereby inducing apoptosis and death in cancer cells. Due to the advantages of ultrasound, such as strong penetration into deep tissues, low cost, non-invasiveness, and minimal side effects, SDT has shown great potential in clinical cancer treatment. However, in practical applications, the lack of highly efficient ultrasound sensitizers to continuously generate large amounts of ROS limits means that SDT remains in the early stages of research.

[0003] Research has found that while inorganic materials are promising acoustic sensitizers for cancer-specific electron-hole recombination (SDT), most inorganic sensitizers exhibit wide band gaps and high electron-hole recombination rates, resulting in low SDT efficiency. Recent studies have shown that defect engineering is a powerful strategy for tuning the band gap of metal oxide nanomaterials, as defects can hinder electron-hole recombination. - ) and holes (h + The composite of inorganic nanomaterials with active sites can be added to enhance the performance of SDT (Sound Sensitive Technology). Although some progress has been made in the structural engineering of inorganic nanomaterials as sound sensitive agents, there is still much room for improvement in achieving efficient and high-volume production of ROS in SDT.

[0004] In recent years, metal single atoms have attracted widespread attention due to their quantum size, boundary effects, and high activity sites. Studies have shown that metal single atoms can serve as nanozymes for cancer treatment, exhibiting catalytic activity and kinetics similar to natural enzymes. Research has demonstrated the preparation of zinc single-atom-doped carbon nanomaterials using zinc carbide-based metal-organic frameworks, followed by the attachment of molybdenum disulfide quantum dots to their surface via electrostatic interactions, constructing molybdenum disulfide-modified zinc single-atom catalysts for sonodynamic and ion therapy in osteomyelitis. However, currently developed metal single-atom sonosensitizers exhibit poor kinetic activity and require synergistic effects with other therapeutic approaches (such as ion or chemokinetic therapy) to achieve effective tumor treatment. Therefore, developing highly efficient metal single-atom-based sonosensitizers and applying them to high-performance SDT remains a significant challenge. Summary of the Invention

[0005] The purpose of this invention is to address the problem that existing metal single-atom sonosensitive agents have poor kinetic activity and require synergy with other treatment methods to achieve effective tumor treatment. This invention provides a two-dimensional defective tungsten oxide nanomaterial modified with a metal single atom, its preparation method, and its application in ultrasound-driven antitumor therapy.

[0006] To address the shortcomings of the aforementioned technical problems, the present invention employs the following technical solution: a method for preparing two-dimensional defect-type tungsten oxide nanomaterials modified with single metal atoms. First, tungsten oxide hydrate nanosheets are prepared using a hydrothermal method, and then defect-rich WO3 nanosheets are prepared by calcination under a nitrogen atmosphere. 3-x Nanosheet materials were finally deposited on WO3 using atomic deposition technology. 3-x Metal single-atom deposition was performed on nanosheets to obtain two-dimensional defective tungsten oxide nanomaterials modified with metal single atoms.

[0007] As a further optimization of the preparation method of the metal single-atom modified two-dimensional defective tungsten oxide nanomaterial of the present invention, the preparation method specifically includes the following steps: S1. Dissolve Na2WO4·H2O, citric acid and glucose in deionized water under ultrasonic stirring, then add hydrochloric acid and stir further. Transfer the mixture to a reactor for heating reaction. The precipitate after reaction is washed and dried to obtain tungsten oxide hydrate nanosheets. S2. Calcining tungsten oxide hydrate nanosheets under a nitrogen atmosphere yields defective WO3. 3-x Nanosheets; S3, Defective WO 3-x Nanosheets were dispersed on quartz wafers containing ethanol, dried at room temperature, and then transferred to a closed atomic deposition reactor for atomic deposition, depositing single metal atoms onto defect-type WO3. 3-x Nanosheets are two-dimensional defect-type tungsten oxide nanomaterials modified with single metal atoms.

[0008] As a further optimization of the preparation method of the metal single-atom modified two-dimensional defective tungsten oxide nanomaterial of the present invention: the molar ratio of Na2WO4·H2O, citric acid and glucose in step S1 is 2:3:10.

[0009] As a further optimization of the preparation method of the metal single-atom modified two-dimensional defective tungsten oxide nanomaterial of the present invention: the heating reaction temperature in step S1 is 100-120℃, and the holding time is not less than 12 h.

[0010] As a further optimization of the preparation method of the two-dimensional defective tungsten oxide nanomaterial modified with a single metal atom according to the present invention, the calcination process conditions in step S2 are as follows: the heating rate is 2-5℃ / min, the temperature is raised to 300-500℃, and the carbonization time is 1-3h.

[0011] As a further optimization of the preparation method of the metal single-atom modified two-dimensional defective tungsten oxide nanomaterial of the present invention: the metal single-atom precursor used for atomic deposition in step S3 is (2,2,6,6-tetramethyl-3,5-heptanoic acid)copper(II).

[0012] As a further optimization of the preparation method of the two-dimensional defective tungsten oxide nanomaterial modified with metal single atoms of the present invention: the metal single atom precursor used for atomic deposition in step S3 is ferrocene, bis(cyclopentadienyl)cobalt or bis(cyclopentadienyl)nickel.

[0013] As a further optimization of the preparation method of the metal single-atom modified two-dimensional defective tungsten oxide nanomaterial of the present invention: the reaction gas of the atomic deposition is O3, and the deposition temperature is 200-300℃.

[0014] A two-dimensional defective tungsten oxide nanomaterial modified with a single metal atom was prepared by the above-described method.

[0015] The application of the aforementioned two-dimensional defective tungsten oxide nanomaterials modified with metal single atoms in high-performance ultrasound-driven therapy utilizes ultrasound to promote the induction and enhancement of reactive oxygen species generation and production, thereby improving the activity of ultrasound sensitizers.

[0016] The present invention has the following beneficial effects: 1. This invention uses a combination of hydrothermal method and atomic deposition technology to prepare two-dimensional defect-type WO3 modified with single atoms of metals such as Cu, Fe, Co and Ni. 3-x Nanomaterials. In this material, WO3 3-x It possesses single-crystal properties and nanoscale size similar to WO3, while exhibiting abundant nanopores and defects, and strong coordination with metal single atoms. Furthermore, Cu-WO3... 3-x Nanomaterials can enhance the yield of reactive oxygen species (ROS) under ultrasound stimulation, and these materials possess advantages such as good biocompatibility, low hemolysis, and high biosafety. Under ultrasound stimulation, this material can kill almost all tumor cells. Compared with traditional ultrasound sensitizers, this material exhibits superior therapeutic effects in high-performance SDT (sound-assisted targeted therapy) for cancer treatment.

[0017] 2. This invention utilizes atomic deposition technology to uniformly deposit different types of metal single atoms onto a two-dimensional defect-type WO3. 3-x The preparation process of nanomaterials is simple, green, and mild, and does not affect the morphology of the carrier. Attached Figure Description

[0018] Figure 1 These are transmission electron microscopy (TEM) images (a), transmission electron microscopy (HR-TEM) images (b), selected area electron diffraction (SAED) images (c), and atomic force microscopy (AFM) images (d) of the WO3 nanosheets prepared in Example 1. Figure 2 WO prepared in Example 1 3-x TEM images (a), SAED images (b), HR-TEM images (c), and high-angle annular dark-field scanning transmission electron microscopy (HAADE-STEM) images (d, e, and f) of nanosheets; Figure 3 The Cu-WO prepared in Example 1 3-x High-angle annular dark-field scanning transmission electron microscopy (HAADE-STEM) images of nanosheets (a and b) and energy-dispersive X-ray (EDX) elemental mapping image (c). Figure 4 The Cu-WO prepared in Example 1 3-x X-ray absorption near-edge structure (XANES) spectra of nanosheets and blank copper foil (a) and extended X-ray absorption fine structure (EXAFS) spectra (b). Figure 5 The Cu-WO prepared in Example 1 3-x Nanosheets and WO 3-x XRD patterns of nanosheets and WO3 nanosheets; Figure 6 The Cu-WO prepared in Example 1 3-x Nanosheets and WO 3-x W4f XPS spectrum (a) and O1s XPS spectrum (b) and ESR spectrum (c) of nanosheets and WO3 nanosheets; Figure 7 The PBS, TiO2, WO3, and WO3 prepared in Example 1 are... 3-x Cu-WO3, Cu-WO 3-x and M-WO 3-x Fluorescence intensity (ah) at different ultrasound times, and PBS, WO3, WO 3-x Cu-WO3 and Cu-WO 3-x ESR spectrum under ultrasound (i); Figure 8 The WO3(a) and WO3(a) prepared in Example 1 are... 3-x (b) Cu-WO 3-x(c) UV-Vis-NIR diffuse reflectance spectra of nanosheets; Figure 9 The WO3(a) and WO3(a) prepared in Example 1 are... 3-x (b) Cu-WO 3-x (c) Band gap of nanosheets; Figure 10 The WO3(a) and WO3(a) prepared in Example 1 are... 3-x (b) Cu-WO 3-x (c) Mott-Schottky curves of nanosheets; Figure 11 The WO3(a) and WO3(a) prepared in Example 1 are... 3-x (b) Cu-WO 3-x (c) Relative fluorescence intensity and photoluminescence spectrum of nanosheets under ultrasonic treatment using a hydroacetylimine 123 (DHR 123) probe (d). Figure 12 Different concentrations of Cu-WO3 prepared in Example 1 3-x - Survival rate of 4T1 cells treated with PEG nanosheets (a); compared with Cu-WO 3-x -Hemolysis rate of blood after 4 h of co-cultivation with PEG nanosheets (b); using RhB@Cu-WO 3-x - Fluorescence intensity of RhB in 4T1 cells cultured with PEG at different time points (c); different doses of WO3-PEG, WO 3-x -PEG, Cu-WO3-PEG and Cu-WO 3-x - Cytotoxicity of PEG nanosheets to 4T1 cells (d); Calcein-AM / PI staining positivity rate of 4T1 cells after sonication treatment (e); Quantitative analysis of apoptosis in 4T1 cells (f). Detailed Implementation

[0019] To better understand the present invention, the following embodiments further illustrate the content of the present invention, but the content of the present invention is not limited to the following embodiments.

[0020] A method for preparing two-dimensional defective tungsten oxide nanomaterials modified with single metal atoms involves first preparing tungsten oxide hydrate nanosheets using a hydrothermal method, and then preparing defect-rich WO3 nanosheets by calcination under a nitrogen atmosphere. 3-x Nanosheet materials were finally deposited on WO3 using atomic deposition technology. 3-x Metal single-atom deposition was performed on nanosheets to obtain two-dimensional defective tungsten oxide nanomaterials modified with metal single atoms.

[0021] As a further optimization of the preparation method of the metal single-atom modified two-dimensional defective tungsten oxide nanomaterial of the present invention, the preparation method specifically includes the following steps: S1. Dissolve Na2WO4·H2O, citric acid and glucose in deionized water under ultrasonic stirring, then add hydrochloric acid and stir further. Transfer the mixture to a reactor for heating reaction. The precipitate after reaction is washed and dried to obtain tungsten oxide hydrate nanosheets. S2. Calcining tungsten oxide hydrate nanosheets under a nitrogen atmosphere yields defective WO3. 3-x Nanosheets; S3, Defective WO 3-x Nanosheets were dispersed on quartz wafers containing ethanol, dried at room temperature, and then transferred to a closed atomic deposition reactor for atomic deposition, depositing single metal atoms onto defect-type WO3. 3-x Nanosheets are two-dimensional defect-type tungsten oxide nanomaterials modified with single metal atoms.

[0022] As a further optimization of the preparation method of the metal single-atom modified two-dimensional defective tungsten oxide nanomaterial of the present invention: the molar ratio of Na2WO4·H2O, citric acid and glucose in step S1 is 2:3:10.

[0023] As a further optimization of the preparation method of the metal single-atom modified two-dimensional defective tungsten oxide nanomaterial of the present invention: the heating reaction temperature in step S1 is 100-120℃, and the holding time is not less than 12 h.

[0024] As a further optimization of the preparation method of the two-dimensional defective tungsten oxide nanomaterial modified with a single metal atom according to the present invention, the calcination process conditions in step S2 are as follows: the heating rate is 2-5℃ / min, the temperature is raised to 300-500℃, and the carbonization time is 1-3h.

[0025] As a further optimization of the preparation method of the two-dimensional defective tungsten oxide nanomaterial modified with metal single atoms of the present invention: the metal single atom precursor used for atomic deposition in step S3 is (2,2,6,6-tetramethyl-3,5-heptanoic acid)copper(II), ferrocene, bis(cyclopentadienyl)cobalt or bis(cyclopentadienyl)nickel.

[0026] As a further optimization of the preparation method of the metal single-atom modified two-dimensional defective tungsten oxide nanomaterial of the present invention: the reaction gas of the atomic deposition is O3, and the deposition temperature is 200-300℃.

[0027] Example 1

[0028] First, NaWO4H2O (0.68 g, 2 mmol), citric acid (0.63 g, 3 mmol), and glucose (1.98 g, 10 mmol) were dissolved in 60 mL of deionized water under stirring and sonication. Then, 6 mL of hydrochloric acid (6 M) was added, and the mixture was stirred for another 0.5 h before being transferred to a 100 mL stainless steel reactor and heated to 120 °C for 24 h. The resulting precipitate was washed with deionized water (3 times) and ethanol (2 times), and dried at 60 °C for 12 h to obtain blackish-green tungsten trioxide hydrate nanosheets.

[0029] The 500 mg of tungsten trioxide hydrate nanosheets prepared above were placed flat in a ceramic crucible and placed in a tube furnace. Under a nitrogen atmosphere, the temperature was increased to 400 °C at a rate of 3 °C / min and held for 2 h to obtain deep blue defective WO3 nanosheets. 3-x Nanosheets.

[0030] Defective WO 3-x The sample was dispersed on a quartz wafer containing ethanol, then dried at room temperature and transferred to an ALD chamber for atomic layer deposition. Using (2,2,6,6-tetramethyl-3,5-heptadecanoic acid)copper(II) as the copper precursor and O3 as the reaction gas, deposition was performed at 250 °C to prepare copper-modified defective tungsten oxide (Cu-WO3). 3-x Nanomaterials.

[0031] The single atoms obtained by atomic deposition technology in this invention exist not only at defects but also at other locations on the surface, allowing them to cover the surface of the carrier more uniformly and to better synergize with the carrier. Therefore, they can more easily sensitize surrounding oxygen to generate reactive oxygen species under the action of ultrasound, thus making them useful for tumor treatment.

[0032] Example 2

[0033] First, NaWO4H2O (0.34 g, 1 mmol), citric acid (0.315 g, 1.5 mmol), and glucose (0.99 g, 5 mmol) were dissolved in 40 mL of deionized water under stirring and sonication. Then, 6 mL of hydrochloric acid (6 M) was added, and the mixture was stirred for 1 h before being transferred to a 100 mL stainless steel reactor and heated to 100 °C for 18 h. The resulting precipitate was washed with deionized water (3 times) and ethanol (2 times), and dried at 60 °C for 15 h to obtain blackish-green tungsten trioxide hydrate nanosheets.

[0034] The 500 mg of tungsten trioxide hydrate nanosheets prepared above were placed flat in a ceramic crucible and placed in a tube furnace. Under a nitrogen atmosphere, the temperature was increased to 400 °C at a rate of 2 °C / min and held for 2 h to obtain deep blue defective WO3 nanosheets. 3-x Nanosheets.

[0035] Defective WO 3-x Nanosheets were dispersed on quartz wafers containing ethanol. The samples were then dried at room temperature and transferred to an ALD chamber for atomic layer deposition. Using ferrocene as the iron precursor and O3 as the reaction gas, deposition was performed at 250 °C to prepare iron-monoatom modified defective tungsten oxide (Fe-WO3). 3-x Nanomaterials.

[0036] Example 3

[0037] First, NaWO4H2O (0.34 g, 1 mmol), citric acid (0.315 g, 1.5 mmol), and glucose (0.99 g, 5 mmol) were dissolved in 40 mL of deionized water under stirring and sonication. Then, 6 mL of hydrochloric acid (6 M) was added, and the mixture was stirred for 1 h before being transferred to a 100 mL stainless steel reactor and heated to 100 °C for 18 h. The resulting precipitate was washed with deionized water (3 times) and ethanol (2 times), and dried at 60 °C for 15 h to obtain blackish-green tungsten trioxide hydrate nanosheets.

[0038] The 500 mg of tungsten trioxide hydrate nanosheets prepared above were placed flat in a ceramic crucible and placed in a tube furnace. Under a nitrogen atmosphere, the temperature was increased to 300 °C at a rate of 3 °C / min and held for 2 h to obtain deep blue defective WO3 nanosheets. 3-x Nanosheets.

[0039] Will WO 3-x The sample was dispersed on a quartz wafer containing ethanol, then dried at room temperature and transferred to an ALD chamber for atomic layer deposition. Using bis(cyclopentadienyl)cobalt as the cobalt precursor and O3 as the reaction gas, deposition was performed at 250°C to produce cobalt single-atom modified defective tungsten oxide (Co-WO3). 3-x Nanomaterials.

[0040] Example 4

[0041] First, NaWO4H2O (0.34 g, 1 mmol), citric acid (0.315 g, 1.5 mmol), and glucose (0.99 g, 5 mmol) were dissolved in 40 mL of deionized water under stirring and sonication. Then, 6 mL of hydrochloric acid (6 M) was added, and the mixture was stirred for 1 h before being transferred to a 100 mL stainless steel reactor and heated to 100 °C for 18 h. The resulting precipitate was washed with deionized water (3 times) and ethanol (2 times), and dried at 60 °C for 15 h to obtain blackish-green tungsten trioxide hydrate nanosheets.

[0042] The 500 mg of tungsten trioxide hydrate nanosheets prepared above were placed flat in a ceramic crucible and placed in a tube furnace. Under a nitrogen atmosphere, the temperature was increased to 500 °C at a rate of 5 °C / min and held for 1 h to obtain deep blue defective WO3 nanosheets. 3-x Nanosheets.

[0043] Will WO 3-x The sample was dispersed on a quartz wafer containing ethanol, then dried at room temperature and transferred to an ALD chamber for atomic layer deposition. Using bis(cyclopentadienyl)nickel as the nickel precursor and O3 as the reaction gas, deposition was performed at 250°C to deposit nickel-modified defective tungsten oxide (Ni-WO3). 3-x Nanomaterials.

[0044] <WO3、WO 3-x Cu-WO 3-x Morphological characteristics> Take WO3 and WO3 obtained in Example 1 3-x 1 mg of each nanomaterial was added to a centrifuge tube, diluted with 10 mL of deionized water, and 10 μL of each was dropped onto a copper grid. The samples were observed under a transmission electron microscope, and the thickness of the WO3 nanosheets was characterized by an atomic force microscope (AFM).

[0045] like Figure 1 As shown, transmission scanning electron microscopy (TEM) images reveal that the prepared WO3 exists as uniform nanosheets with a size of 100-200 nm. High-resolution TEM (HR-TEM) images of the WO3 nanosheets show distinct lattice fringes, confirming its single-crystal nature. Bright rectangular diffraction spots are observed in the SAED electron diffraction pattern of the WO3 nanosheets, further confirming the single-crystal nature of this material. Simultaneously, AFM images show that the thickness of the WO3 nanosheets is in the range of 15-20 nm. Figure 2 As shown, via WO 3-x TEM images of the nanosheets show that the material has a similar shape and size to WO3. The SAED pattern reveals that WO...3-x It also exhibits bright, rectangular diffraction spots, indicating its single-crystal nature. Furthermore, WO 3-x HR-TEM images of the nanosheets revealed continuous lattice fringes, abundant nanoscale pores, and defects. High-angle annular dark-field scanning transmission electron microscopy (HAADE-STEM) images further illustrate the WO 3-x Nanosheets contain abundant nanopores and defects. ALD technology was used to fabricate nanosheets in WO4. 3-x Cu single atoms are deposited on the surface of nanosheets to obtain Cu-WO 3-x Nanosheets. The Cu loading, determined by inductively coupled plasma atomic emission spectrometry (ICP-AES), is approximately 0.04 wt%. Figure 3 As shown, Cu-WO 3-x HAADF-STEM images of nanosheets show their relationship with WO 3-x Similar characteristics were observed, namely, the display of continuous lattice fringes with abundant nanopores and defects. Because the atomic number of Cu is smaller than that of the support W, Cu atoms could not be directly identified from TEM. However, the presence of Cu was confirmed by energy-dispersive X-ray (EDX) spectroscopy, and its uniform distribution on the nanosheet surface was demonstrated. Figure 4 As shown, to further prove the existence of Cu single atoms, we conducted X-ray absorption near-edge structure spectroscopy (XANES) studies on Cu-WO3. 3-x Cu-WO3 exhibits a higher energy absorption threshold than copper foil, indicating that Cu-WO3... 3-x The copper in Cu carries a positive charge. The coordination structure of single Cu atoms was further revealed by extended X-ray absorption fine structure (EXAFS) spectroscopy. In Cu-WO3... 3-x In the R-space of the WO3 matrix, there is only a peak around 1.5 Å, which is absent in the copper foil. This indicates that the peak is due to the contribution of the Cu-O bond and not the presence of a Cu-Cu metallic bond. In contrast, the R-space curve of the copper foil shows only a strong Cu-Cu metallic bond peak. This result clearly demonstrates the role of Cu single atoms in WO3. 3-x Successful deposition on nanosheets, and strong coordination between Cu single atoms and defect sites on the nanosheet surface.

[0046] <WO3、WO 3-x Cu-WO 3-x Material characterization Take the WO3 and WO3 prepared in Example 1 3-x Cu-WO 3-x Five mg of each nanomaterial was placed on a glass slide for XRD testing, and then three mg of each sample was taken for X-ray photoelectron spectroscopy (XPS) testing.

[0047] like Figure 5As shown, various technologies are used to analyze WO3 and WO 3-x Cu-WO 3-x The nanosheets were further characterized. The X-ray powder diffraction (XRD) peaks of the WO3 nanosheets were consistent with the standard PDF card (No. 83-0951) for monoclinic WO3. 3-x and Cu-WO 3-x The nanosheets all exhibited similar peaks, but with slightly lower intensity, indicating that WO 3-x The defects in the nanosheets and the deposited Cu single atoms do not affect their crystallinity.

[0048] like Figure 6 As shown, XPS was used to analyze WO3 and WO 3-x Cu-WO 3-x The nanosheets were analyzed. The high-resolution XPS W4f spectrum of the WO3 nanosheets showed two main peaks at 37.8 and 35.7 eV. In contrast, the high-resolution XPS W4f spectrum of WO3 nanosheets... 3-x and Cu-WO 3-x The two pairs of peaks displayed by the nanosheets and W 5+ This is more pronounced at 36.7 and 34.7 eV, indicating that WO 3-x and Cu-WO 3-x All nanosheets contain W 5+ WO 3-x and Cu-WO 3-x The high-resolution XPS O 1s spectrum of the nanosheets peaked at 530.5 eV for the WO bonds and at 531.6 eV for the oxygen vacancies. We further characterized the oxygen vacancies using electron spin resonance (ESR) spectroscopy. Compared to WO3 nanosheets, WO... 3-x and Cu-WO 3-x The nanosheets exhibit a symmetrical ESR signal at g = 2.003, proving that WO 3-x and Cu-WO 3-x The nanosheets all had more oxygen vacancies than the WO3 nanosheets, which is consistent with the results of TEM and XPS.

[0049] <WO3、WO 3-x Cu-WO 3-x Characterization of catalytic performance in inducing ROS formation The singlet oxygen green fluorescent probe (SOSG) was used to study WO3 and WO3. 3-x Cu-WO 3-x The performance of the sample in generating ROS under ultrasonic treatment. 200 μL of WO3 and WO4 were added. 3-x Cu-WO 3-x (1 mg·mL) -1200 μL of SOSG (10 μM) and 200 μL of SOSG (10 μM) were added to 1600 μL of deionized water, and then the solution was irradiated with ultrasound (3 W / cm²). -2 (5 min). Detection was performed by continuously recording the fluorescence intensity of SOSG at 525 nm. 1 The generation of O2.

[0050] like Figure 7 As shown, the fluorescence intensity of each group of SOSG increased with increasing sonication time, indicating successful generation. 1 O2. Compared to WO3 nanosheets, defective WO3 nanosheets... 3-x The nanosheets exhibited higher SOSG fluorescence intensity after 5 min of sonication, indicating that WO 3-x Nanosheets have stronger properties than WO3 nanosheets. 1 O2 generation capacity. Cu-WO 3-x The SOSG fluorescence intensity of the nanosheets was significantly enhanced, and its ROS generation was 3.1 times that of WO3 nanosheets and 6.7 times that of commercial TiO2 ultrasound sensitizers. Even when Cu single atoms were deposited on WO3 nanosheets, the ROS generation capacity was still improved, demonstrating that the synergistic effect of oxygen vacancies and Cu single atoms can significantly improve the SDT performance of this ultrasound sensitizer. After 5 minutes of ultrasonic treatment, Cu-WO3 nanosheets showed significantly better ROS generation compared to other metal nanocomposites. 3-x It exhibits the highest fluorescence intensity at 525 nm, which is Fe-WO3. 3-x twice as much ( Figure 7 h). This result indicates that Cu-WO 3-x Nanosheets modified with numerous metal single atoms in WO3 3-x Among nanomaterials, its SDT effect is the best. Meanwhile, from WO3 and WO... 3-x Cu-WO3 and Cu-WO 3-x TEMP / nanosheets 1 The ESR spectrum of O2 shows that Cu-WO 3-x The nanosheet peak intensity is stronger than that of other materials, which also confirms that Cu-WO 3-x It has strong ROS generation capabilities and huge SDT potential.

[0051] <WO3、WO 3-x M-WO 3-x ROS generation mechanism of materials > like Figure 8 As shown, the UV diffuse reflectance curves of WO3 nanosheets exhibit strong UV-Vis absorption (220-500 nm), but weak absorption in the NIR region (>1000 nm), due to its wide band gap. Conversely, WO... 3-xand Cu-WO 3-x The nanosheets exhibit strong absorption in both the visible and NIR regions, indicating that oxygen vacancies can effectively reduce the band gap of the WO3 nanosheets. Furthermore, calculations of WO3 and WO... 3-x Cu-WO 3-x The band gaps are 3.14, 1.87, and 1.70 eV, respectively. Figure 9 ), further indicating that based on WO x The presence of oxygen vacancies in the ultrasonic sensitizer can significantly reduce the band gap of WO3, while deposited metal single atoms can further reduce the band gap. Subsequently, based on the Mott-Schotky diagram, the CB positions of the above-mentioned ultrasonic sensitizer were determined to be -0.17 eV, -0.12 eV, and -0.16 eV, respectively. Figure 10 Accordingly, WO3 and WO are calculated. 3-x Cu-WO 3-x The VB positions are 2.97, 1.75, and 1.54 eV, respectively. Based on the above results, compared with WO3 and Cu-WO3 nanosheets, Cu-WO3... 3-x e in nanosheet VB - -h + It is easier to be excited and separated. WO 3-x Cu single atoms on the surface of nanosheets are more conducive to capturing e. - Promote its interaction with O 2 The reaction produces O 2- Thus further with h + Combine to generate the final 1 O2.

[0052] Meanwhile, the presence of O was verified using the dihydroacetylimine 123 (DHR 123) probe. 2- The generation of. In Figure 11 In AC, compared to WO3, based on WO 3-x The ultrasound sensitizer showed increased fluorescence intensity within 5 minutes of ultrasound. Furthermore, Cu-WO3... 3-x The nanosheets exhibited the strongest fluorescence intensity, indicating the O generated during their formation. 2- The largest quantity. For example... Figure 11 As shown in d, WO3 nanosheets exhibit a strong photoluminescence (PL) emission peak, while defective WO3 nanosheets show a strong photoluminescence (PL) emission peak. 3-x The PL intensity of this peak is significantly reduced in the nanosheets. Most importantly, Cu-WO 3-x Nanosheets exhibited the lowest PL intensity among all ultrasound sensitizers, indicating that they possess the highest e - -h + Separation efficiency.

[0053] <WO3、WO 3-x Cu-WO3-x In vitro cell testing of nanomaterials > Due to Cu-WO 3-x Nanosheets exhibit good ROS generation performance under ultrasound, and Cu-WO3 is synthesized using polyethylene glycol (PEG), which has good biocompatibility. 3-x The material was modified, and the Cu-WO4 model was explored in vitro. 3-x -The SDT effect of PEG. Prior to this, Cu-WO3 was used to treat mouse lung cancer (4T1) cells with diphenyltetrazolium bromide (MTT). 3-x -In vitro biocompatibility analysis of PEG.

[0054] Depend on Figure 12 As shown in Figure a, different concentrations of Cu-WO3 were used. 3-x -PEG (0-100 μg mL) -1 After cultivation, the viability of 4T1 cells remained close to 100%, indicating good biocompatibility. The hemolysis method was used to further confirm its biocompatibility. After culturing mouse erythrocytes with different concentrations of the material for 4 hours, Cu-WO4... 3-x -PEG at different concentrations (12.5, 25, 50, 100 and 200 μg mL) -1 All are below the international standard of 5% ( Figure 12 (b) It has low hemolytic activity and high biosafety. Secondly, by reacting 4T1 cells with RhB-labeled Cu-WO3... 3-x -PEG(RhB@Cu-WO 3-x -PEG) nanomaterials were cultured at different times (0-24 h) to study the cell response to Cu-WO3. 3-x -The uptake of PEG materials. Observed in fluorescence microscopy images, the uptake of Cu-WO3... 3-x PEG-treated 4T1 cells showed a strong RhB red fluorescence signal, reaching its maximum intensity after 8 h of culture. Figure 12 c), indicating that 4T1 cells respond to Cu-WO4 3-x -PEG has a highly efficient endocytic effect.

[0055] Based on the above results, we also subjected Cu-WO3 to ultrasonic treatment. 3-x - PEG in vitro cytotoxicity test of 4T1 cells ( Figure 12 d). Compared with blank and sonication alone, the use of WO3-PEG+US, WO 3-x The cell viability of materials with PEG+US and Cu-WO3-PEG+US decreased, but the cell-killing effect remained insufficient. In Cu-WO3... 3-x In the PEG+US experimental group, cell viability further decreased, indicating that Cu-WO3...3-x -PEG exhibits the best therapeutic effect. Furthermore, this result was visually verified through staining experiments using calcein-AM (green fluorescence in live cells) and propidium iodide (red fluorescence in dead cells). Figure 12 e). Strong green fluorescence was observed in both the control group and the ultrasound experimental group, therefore the number of cell deaths was negligible. It is worth noting that cells treated with WO3-PEG + US and WO... 3-x Treatment with PEG +US and Cu-WO3-PEG +US resulted in a sharp increase in cell death and enhanced red fluorescence. For Cu-WO3... 3-x In the PEG + US group, almost all cells died, indicating that Cu-WO 3-x -PEG exhibits superior therapeutic efficacy. Furthermore, apoptosis was analyzed by flow cytometry using the Annexin V-FITC / PI kit. Figure 12 f shows that Cu-WO 3-x The PEG+US experiment induced the highest apoptosis rate (9.8% in the early stage and 80.8% in the late stage).

[0056] The specific embodiments of the present invention have been described above. It should be understood that the present invention is not limited to the specific embodiments described above, and those skilled in the art can make various modifications or variations within the scope of the claims, which do not affect the essence of the present invention.

Claims

1. A method for preparing a two-dimensional defect-type tungsten oxide nanomaterial modified with a single metal atom, characterized in that: Specifically, the following steps are included: S1. Tungsten oxide hydrate nanosheets were prepared by hydrothermal method. Specifically, Na2WO4·H2O, citric acid and glucose were dissolved in deionized water under ultrasonic stirring. Then hydrochloric acid was added and stirred further. The mixture was transferred to a reactor for heating reaction. The precipitate after reaction was washed and dried to obtain tungsten oxide hydrate nanosheets. S2. Calcining tungsten oxide hydrate nanosheets under a nitrogen atmosphere yields defective WO3. 3-x Nanosheets; S3, Defective WO 3-x Nanosheets were dispersed on quartz wafers containing ethanol, dried at room temperature, and then transferred to a closed atomic deposition reactor for atomic deposition, depositing single metal atoms onto defect-type WO3. 3-x Nanosheets are two-dimensional defect-type tungsten oxide nanomaterials modified with single metal atoms. The single-atom metal precursors used in atomic deposition are (2,2,6,6-tetramethyl-3,5-heptanediol)copper(II), ferrocene, bis(cyclopentadienyl)cobalt or bis(cyclopentadienyl)nickel, and the deposition temperature is 200-300℃.

2. The method for preparing a two-dimensional defect-type tungsten oxide nanomaterial modified with a single metal atom as described in claim 1, characterized in that: In step S1, the molar ratio of Na2WO4·H2O, citric acid, and glucose is 2:3:

10.

3. The method for preparing a two-dimensional defect-type tungsten oxide nanomaterial modified with a single metal atom as described in claim 1, characterized in that: The heating reaction temperature in step S1 is 100-120℃, and the holding time is not less than 12 hours.

4. The method for preparing a two-dimensional defect-type tungsten oxide nanomaterial modified with a single metal atom as described in claim 1, characterized in that: The calcination process conditions in step S2 are as follows: heating rate of 2-5℃ / min, heating to 300-500℃, and carbonization time of 1-3h.

5. The method for preparing a two-dimensional defect-type tungsten oxide nanomaterial modified with a single metal atom as described in claim 1, characterized in that: The reaction gas for the atomic deposition is O3.

6. A two-dimensional defect-type tungsten oxide nanomaterial modified with a single metal atom, characterized in that: It is prepared by the preparation method described in any one of claims 1-5.

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