Titanium dioxide-based composite material, method for preparing same and use thereof

Abstract

The application belongs to the technical field of photocatalysts, and particularly relates to a titanium dioxide-based composite material and a preparation method and application thereof. The application provides a titanium dioxide-based composite material, which comprises titanium dioxide nanoflowers and sheet-like carbon nitride loaded on the titanium dioxide nanoflowers. By introducing carbon nitride into the composite material, visible light absorption and catalytic activity can be further improved, so that the consumption of glucose as a sacrificial agent in a high glucose microenvironment is promoted. The results of a photocatalytic glucose depletion experiment show that the titanium dioxide-based composite material provided by the application can effectively degrade glucose and produce a large amount of hydrogen under visible light irradiation, can reduce the pro-apoptotic effect of a high sugar environment on cells, promote cell proliferation and migration, and thus support the healing of a diabetic wound.

Description

Technical Field

[0001] This invention belongs to the field of photocatalyst technology, specifically relating to a titanium dioxide-based composite material, its preparation method, and its application. Background Technology

[0002] Diabetes mellitus is a common chronic metabolic disease characterized by persistent hyperglycemia and related complications, including impaired wound healing. It is estimated that approximately 30% of diabetic patients develop chronically impaired wounds, primarily located in the lower extremities, known as diabetic foot ulcers (DFU). This has become a persistent medical challenge, leading to amputations and increasing healthcare costs. Hyperglycemia is a major cause of impaired wound healing in diabetic patients, including the production of pro-inflammatory cytokines such as interleukin (IL)-1β and tumor necrosis factor-α (TNF-α). While these cytokines help clear dead cells and bacteria and lay the foundation for tissue repair, excessive or prolonged immune responses can also lead to chronic inflammation and angiogenesis disorders, thus affecting wound healing in diabetic patients. Furthermore, hyperglycemia is also associated with dysregulation of myofibroblast differentiation and insufficient ECM production, the latter being crucial for promoting wound contraction and accelerating the healing process.

[0003] Currently, typical methods for treating chronic diabetic wounds include debridement of ulcerated necrotic tissue, topical antibiotics, and wound dressings (such as membranes, fibers, and hydrogels). However, these methods have limited therapeutic efficacy and numerous side effects (including kidney and liver toxicity), restricting their application. Although negative pressure therapy, hyperbaric oxygen therapy (HBOT), and growth factor therapy have been proposed as effective strategies for combating diabetic wounds, these methods are time-consuming and expensive. Therefore, there is an urgent need for new treatment methods that can both lower local glucose levels and reduce inflammation and promote wound healing to meet the needs of diabetic patients.

[0004] Hydrogen is considered a safe and effective medical gas with anti-inflammatory and antioxidant properties. Recent studies have shown that hydrogen has potential in treating inflammation-related diseases, including accelerating wound healing in diabetic patients. Through mechanisms such as inhibiting inflammatory responses, reducing oxidative stress, and promoting cell proliferation, hydrogen can promote wound healing and improve the quality of wound healing in diabetic patients. Furthermore, hydrogen can improve blood circulation and enhance microcirculation, thereby delivering more nutrients and oxygen to the wound and promoting healing. Clinical trials have confirmed the feasibility of delivering hydrogen to patients via inhalation or injection. While traditional titanium dioxide (TiO2) possesses good stability and high photocatalytic activity, it only exhibits good catalytic activity under ultraviolet light and has weak absorption under visible light, thus limiting its application in the visible light region. Summary of the Invention

[0005] The purpose of this invention is to provide a titanium dioxide-based composite material, its preparation method, and its application. The titanium dioxide-based composite material provided by this invention has highly efficient photocatalytic activity.

[0006] To achieve the above objectives, the present invention provides the following technical solution:

[0007] The present invention provides a titanium dioxide-based composite material comprising titanium dioxide nanoflowers and sheet-like carbon nitride supported on the titanium dioxide nanoflowers.

[0008] Preferably, the loading percentage of flaky carbon nitride on the titanium dioxide-based composite material is 10-70% by mass.

[0009] This invention also provides a method for preparing the titanium dioxide-based composite material described in the above technical solution, comprising the following steps:

[0010] Amorphous titanium dioxide is obtained by mixing a titanium source and an acidic reagent and carrying out a hydrothermal reaction.

[0011] The amorphous titanium dioxide was subjected to a first annealing to obtain titanium dioxide nanoflowers;

[0012] A suspension is obtained by ultrasonically mixing flake carbon nitride and an organic solvent;

[0013] The titanium dioxide nanoflowers and suspension are mixed and subjected to a second annealing to obtain the titanium dioxide-based composite material.

[0014] Preferably, the titanium source includes one or more of tetrabutyl titanate, titanium trichloride, titanium tetrachloride, and titanium isopropoxide; the acidic reagent includes one or more of acetic acid, hydrochloric acid, and lactic acid.

[0015] The volume ratio of the titanium source to the acidic reagent is 1:10 to 50.

[0016] Preferably, the temperature of the hydrothermal reaction is 100-150°C, and the holding time is 8-12 hours.

[0017] Preferably, the temperature of the first annealing is 300-400℃, and the holding time is 2-4h.

[0018] Preferably, the ratio of the flake carbon nitride to the organic solvent is 0.05–0.35 g: 70 mL;

[0019] The mass ratio of the sheet-like carbon nitride to titanium dioxide nanoflowers is 0.05–0.35:0.5.

[0020] Preferably, the temperature of the second annealing is 300-400°C, and the holding time is 1-2 hours.

[0021] The present invention also provides the application of the titanium dioxide-based composite material described in the above technical solution or the titanium dioxide-based composite material prepared by the preparation method described in the above technical solution in the preparation of a catalyst for treating slow wound healing in diabetic patients.

[0022] Preferably, the slow healing of diabetic wounds includes diabetic incision healing disorder, diabetic chronic ulcer, and diabetic abnormal scar hyperplasia.

[0023] This invention provides a titanium dioxide-based composite material comprising titanium dioxide nanoflowers and sheet-like carbon nitride supported on the titanium dioxide nanoflowers. By introducing carbon nitride into the composite material, this invention further enhances visible light absorption and catalytic activity, thereby promoting the consumption of glucose as a sacrificial agent in a high-glucose microenvironment. This invention aims to explore the application of the composite material in photocatalytic glucose depletion and hydrogen production reactions, and to investigate its potential application in treating diabetic wounds. Results of photocatalytic glucose depletion experiments show that the titanium dioxide-based composite material provided by this invention can effectively degrade glucose and generate a large amount of hydrogen gas under visible light irradiation, mitigating the pro-apoptotic effects of a high-glucose environment, promoting cell proliferation and migration, and thus supporting the healing of diabetic wounds, providing a new and effective method for treating diabetic wounds. Furthermore, the hydrogen gas produced as a byproduct has anti-inflammatory effects, providing an important therapeutic mechanism for research. To further explore its potential application in treating diabetic wounds, this invention coated the composite material onto an artificially prepared diabetic wound model and subjected it to light treatment. Observations show that the photocatalytically generated hydrogen gas can promote wound healing and reduce the risk of infection. Attached Figure Description

[0024] Figure 1 The images shown are scanning electron microscope (SEM) and transmission electron microscope (TEM) images of the CNT50 obtained in Example 1.

[0025] Figure 2 This is a statistical graph showing the effect of CNT50 obtained in Example 1 on HSF proliferation under light conditions;

[0026] Figure 3 This is a comparison of the effects of CNT50 obtained in Example 1 on HSF cell viability under high glucose conditions;

[0027] Figure 4 This is a comparison of the effects of CNT50 obtained in Example 1 on HSF cell viability under high glucose conditions;

[0028] Figure 5 The figure shows the effect of CNT50 obtained in Example 1 on the mRNA expression levels of COL1A1, TGF-β and α-SMA under high glucose environment;

[0029] Figure 6 This is a comparison of the effects of CNT50 obtained in Example 1 on the viability of HUVEC cells under high glucose conditions;

[0030] Figure 7 This is a comparison of the effects of CNT50 obtained in Example 1 on HUVEC cell apoptosis under high glucose conditions;

[0031] Figure 8 This is a comparison of the effects of CNT50 obtained in Example 1 on HUVEC angiogenesis under high glucose conditions;

[0032] Figure 9 The figure shows the effect of CNT50 obtained in Example 1 on the expression levels of IL-1β, TNF-α and VEGF under high glucose environment;

[0033] Figure 10 The following is a comparative and statistical graph showing the effect of CNT50 obtained in Example 1 on wound healing in diabetic mice;

[0034] Figure 11 The above is a comparison and statistical chart of the wound area of ​​CNT50 in diabetic mice obtained in Example 1.

[0035] Figure 12 The image shows a comparison and statistical diagram of the expression of CD31 and α-SMA in the wounds of diabetic mice by CNT50 obtained in Example 1. Detailed Implementation

[0036] The present invention provides a titanium dioxide-based composite material comprising titanium dioxide nanoflowers and sheet-like carbon nitride supported on the titanium dioxide nanoflowers.

[0037] In this invention, the crystal phase of the titanium dioxide nanoflowers is preferably anatase. In this invention, the mass percentage of plate-like carbon nitride loaded on the titanium dioxide-based composite material is preferably 10-70%, more preferably 30-50%.

[0038] This invention also provides a method for preparing the titanium dioxide-based composite material described in the above technical solution, comprising the following steps:

[0039] Amorphous titanium dioxide is obtained by mixing a titanium source and an acidic reagent and carrying out a hydrothermal reaction.

[0040] The amorphous titanium dioxide was subjected to a first annealing to obtain titanium dioxide nanoflowers;

[0041] A suspension is obtained by ultrasonically mixing flake carbon nitride and an organic solvent;

[0042] The titanium dioxide nanoflowers and suspension are mixed and subjected to a second annealing to obtain the titanium dioxide-based composite material.

[0043] In this invention, unless otherwise specified, all raw materials used in the preparation are commercially available products well known to those skilled in the art.

[0044] This invention involves mixing a titanium source and an acidic reagent, and then performing a hydrothermal reaction to obtain amorphous titanium dioxide.

[0045] In this invention, the titanium source preferably includes one or more of tetrabutyl titanate, titanium trichloride, titanium tetrachloride and titanium isopropoxide, and more preferably tetrabutyl titanate; the acidic reagent preferably includes one or more of acetic acid, hydrochloric acid and lactic acid, and more preferably acetic acid; the volume ratio of the titanium source and the acidic reagent is preferably 1:10 to 50, and more preferably 1:30.

[0046] In this invention, the mixing process is preferably as follows: adding a titanium source to an acidic reagent and stirring. The stirring time is preferably 5–20 min. The hydrothermal reaction temperature is preferably 100–150°C, more preferably 110–140°C; the holding time is preferably 8–15 h, more preferably 10–12 h. The hydrothermal reaction is preferably carried out in a high-pressure reactor with a polytetrafluoroethylene liner.

[0047] After the hydrothermal reaction, the present invention preferably includes cooling the obtained reaction system to room temperature, collecting the product by centrifugation, and washing and drying the product; the washing preferably includes washing with deionized water and anhydrous ethanol in sequence; the drying process is preferably: drying overnight at 50-90°C, and the temperature is further preferably 70°C.

[0048] After obtaining the amorphous titanium dioxide, the present invention performs a first annealing on the amorphous titanium dioxide to obtain titanium dioxide nanoflowers.

[0049] In this invention, the temperature of the first annealing is preferably 300–500°C, more preferably 400°C; the holding time is preferably 2–4 hours. In this invention, the first annealing is preferably carried out in a tube furnace.

[0050] This invention involves ultrasonically mixing flake carbon nitride and an organic solvent to obtain a suspension.

[0051] In this invention, the flake-shaped carbon nitride is preferably obtained by preparation, and the preparation method preferably includes: calcining melamine to obtain carbon nitride; mixing the carbon nitride with methanol and then annealing to obtain the flake-shaped carbon nitride.

[0052] In this invention, the calcination temperature is preferably 450–550°C, and the holding time is preferably 1–3 hours, more preferably 2 hours; the calcination is preferably carried out in an air atmosphere; the crucible used for calcination is preferably an alumina crucible. After calcination, this invention also preferably includes pulverizing the obtained product. In this invention, the carbon nitride is preferably light yellow. In this invention, the mixing of carbon nitride and methanol is preferably carried out under stirring conditions, and the stirring time is preferably 24–48 hours. After mixing, this invention also preferably includes drying the obtained mixture, and the drying temperature is preferably 50–90°C. In this invention, the annealing temperature is preferably 300–400°C, and the holding time is preferably 2 hours.

[0053] In this invention, the organic solvent preferably includes methanol. In this invention, the ratio of the flake carbon nitride to the organic solvent is preferably 0.05–0.35 g: 70 mL, more preferably 0.25 g: 70 mL. In this invention, the ultrasonic mixing time is preferably 0.5–2 h, more preferably 1 h.

[0054] After obtaining the titanium dioxide nanoflowers and the suspension, the present invention mixes the titanium dioxide nanoflowers and the suspension and performs a second annealing to obtain the titanium dioxide-based composite material.

[0055] In this invention, the mass ratio of the flake-shaped carbon nitride to the titanium dioxide nanoflowers is preferably 0.05–0.35:0.5, more preferably 0.25:0.5. In this invention, the mixing method is preferably stirring; the stirring time is preferably 2 hours. After mixing, this invention further preferably includes drying the resulting mixture; the drying temperature is preferably 50–90°C, more preferably 70°C. In this invention, the second annealing temperature is preferably 300–400°C, and the holding time is preferably 1–2 hours; the second annealing is preferably carried out in a tube furnace.

[0056] The present invention also provides the application of the titanium dioxide-based composite material described in the above technical solution or the titanium dioxide-based composite material prepared by the preparation method described in the above technical solution in the preparation of a catalyst for treating slow wound healing in diabetic patients.

[0057] In this invention, the slow healing of diabetic wounds preferably includes diabetic incision healing disorders, diabetic chronic ulcers, and diabetic abnormal scar hyperplasia. This invention does not impose any specific limitations on the application method; any method well-known to those skilled in the art can be used.

[0058] To further illustrate the present invention, a titanium dioxide-based composite material, its preparation method, and its application are described in detail below with reference to the accompanying drawings and embodiments. However, these descriptions should not be construed as limiting the scope of protection of the present invention.

[0059] Example 1

[0060] 1 mL of tetrabutyl titanate was added to 30 mL of acetic acid and stirred vigorously for 10 minutes. The mixture was then sealed in a 50 mL high-pressure vessel lined with polytetrafluoroethylene and heated at 140 °C for 12 h for hydrothermal reaction. After cooling to room temperature, the product was collected by centrifugation and then thoroughly washed with deionized water and anhydrous ethanol. The product was dried overnight at 70 °C to obtain amorphous titanium dioxide (TNF).

[0061] The obtained amorphous titanium dioxide was placed in a tube furnace and annealed at 400℃ for 2 hours to obtain anatase-type amorphous titanium dioxide nanoflowers (TNFas).

[0062] 5g of melamine was placed in an alumina crucible under ambient air conditions, then heated at 550℃ for 2 hours, and subsequently crushed to obtain carbon nitride powder.

[0063] Carbon nitride powder was stirred in methanol for 24 hours, dried in an oven at 70°C, and the resulting powder was annealed in a tube furnace at 400°C for 2 hours to obtain flake carbon nitride.

[0064] 0.25 g of flake carbon nitride was added to 70 mL of methanol and sonicated for 1 h to obtain a uniform suspension; 0.5 g of amorphous titanium dioxide nanoflowers were added to the suspension and stirred for 2 h; the mixture was dried in an oven at 70 °C, and the resulting powder was annealed in a tube furnace at 400 °C for 2 h to obtain the titanium dioxide-based composite material (denoted as CNT50).

[0065] Performance testing

[0066] Test Example 1

[0067] Figure 1 The scanning electron microscope (SEM) image and transmission electron microscope (TEM) image of the CNT50 obtained in Example 1 are shown below. Figure 1 It can be seen that the synthesized CNT50 samples exhibit different spherical and flower-like structures, with diameters ranging from micrometers. Figure 1 A and Figure 1 B). The synthesized pCN nanosheets exhibit a typical layered structure similar to graphene, indicating the formation of a graphitic phase after calcination. Further TEM observation revealed the two-dimensional (2D) folded sheet structure of the pCN photocatalyst and showed that calcination did not significantly affect the morphology of TNFas. Figure 1 C and Figure 1D). Furthermore, HRTEM images show lattice fringes in the CNT50 sample with a lattice spacing of approximately 0.35 nm, consistent with the (101) plane of anatase TiO2. Figure 1 E).

[0068] Test Example 2

[0069] The CNT50 obtained in Example 1 was added to a 20 mM glucose solution, placed in a sealed reactor under vacuum, and its hydrogen production performance under light conditions was tested. Glucose consumption was measured using a spectrophotometer. A full-thickness skin wound with a diameter of 10 mm was created on the back of each mouse using a circular skin biopsy puncture machine. On days 1, 3, 5, and 7 post-surgery, CNT50 (10 mg / kg) or its petroleum jelly carrier was applied topically to the wound. Mice in the sun-treated group were exposed to sunlight for 3 × 15 min daily. Wound healing and observation were performed.

[0070] Experiment 1: The effect of CNT50-mediated photocatalytic therapy on accelerating HSF proliferation

[0071] Human skin fibroblasts (HSF) were cultured in DMEM medium containing 10% fetal bovine serum (FBS). HSF were incubated with PBS (100 μL) and CNT50 (100 μg / mL) for 30 minutes, followed by stimulation with glucose (25 mM) or water as a control. For light treatment, xenon lamps (100 mW / cm²) were used every 8 hours. 2 Cells were irradiated for 10 minutes. After culturing for 24 hours, the supernatant and cells were collected for further study. HSF proliferation was measured by CCK-8 assay; HSF was collected and incubated with CCK-8 solution in the dark at 37°C for 2 hours. Then, 100 μL of the reaction mixture was transferred to a 96-well plate, and the OD value was detected at 450 nm using a spectrophotometer. Cell viability was calculated using the following formula: Cell viability (%) = (OD 测试 -OD 对照 ) / (OD 对照 -OD 空白 The result is as follows Figure 2 As shown in the figure. The results showed that, compared with the corresponding control group, hyperglycemic conditions significantly inhibited HSF proliferation. However, CNT50 significantly promoted HSF cell proliferation under light conditions compared with dark conditions, demonstrating that CNT50 has the effect of improving HSF proliferation in a hyperglycemic microenvironment.

[0072] Experiment 2: The effect of CNT50-mediated photocatalytic therapy on accelerating HSF activation

[0073] The HSF culture method was the same as in Experiment 1. Cultured HSF was collected, and its activation status was determined using EdU. BeyoClick was used.TM EdU was measured using an EdU cell proliferation kit and an Alexa Fluor 647. Fluorescence intensity was detected using confocal microscopy, and quantitative analysis was performed using ImageJ software. The results are shown below. Figure 3 As shown in the figure. By observing the number and degree of EdU staining, it was found that CNT50 significantly enhanced the cell viability of HSF under light conditions compared to dark conditions after glucose stimulation, proving that CNT50 has the effect of accelerating the activation of HSF cells in a high glucose microenvironment.

[0074] Experiment 3: The role of CNT50-mediated photocatalytic therapy in accelerating HSF migration in a high-glucose environment

[0075] The HSF culture method was the same as in Experiment 1. Cultured HSF was collected, and the effect of CNT50 on HSF migration was determined using a cell scratch assay. HSF was seeded in 6-well culture plates for 48 hours to form a cell monolayer. The cell monolayer was then scratched with a 10 μL pipette, washed twice with PBS, and incubated with PBS (100 μL) and CNT50 (100 μg / mL) for 30 minutes, followed by treatment with glucose (25 mM) or water as a control. For VIS treatment, cells were exposed to a xenon lamp (100 mW / cm²) every 8 hours. 2 Irradiate for 10 minutes. After 24 hours of incubation, capture cell images using an inverted fluorescence microscope. The results are as follows: Figure 4 As shown. Scratch repair rate (%) = (S0-S24) / S0×100%, where S0 and S24 represent the scratch areas before and after intervention, respectively. The results showed that in a high-glucose environment, CNT50 significantly promoted HSF cell migration under light conditions compared to dark conditions, demonstrating that CNT50 has the effect of accelerating HSF cell migration.

[0076] Experiment 4: CNT50-mediated photocatalytic therapy promotes ECM deposition in a high-glucose environment

[0077] Based on Experiment 3, total RNA was extracted from HSF using TRIzol. Following the FastQuant RT kit instructions, 50 ng of RNA was reverse transcribed into cDNA. Quantitative real-time PCR was performed using SYBR Green dye and a 7300 real-time quantitative PCR system. The results are as follows: Figure 5As shown in the figure. The results showed that, compared with the control group, high glucose incubation significantly reduced the mRNA levels of COL1A1, TGF-β, and α-SMA in cells. Under VIS irradiation, CNT50 significantly reversed this reduction, while the mRNA levels in the dark group did not show any significant change. COL1A1, TGF-β, and α-SMA have long been considered sources of extracellular matrix (ECM) deposition, suggesting that CNT50 can promote the generation and deposition of ECM in a high glucose environment under light conditions.

[0078] Experiment 5: CNT50-mediated photocatalytic therapy enhances the viability of HUVEC cells in a high-glucose environment

[0079] Since endothelial cells play a crucial role in angiogenesis and diabetic wound healing, we evaluated the efficacy of CNT50 against human umbilical vein endothelial cells (HUVECs). HUVECs were cultured in Ham's F12K medium containing 10% FBS. HUVECs were incubated with PBS (100 μL) and CNT50 (100 μg / mL) for 30 min, followed by stimulation with glucose (25 mM) or water as a control. For light treatment, xenon lamps (100 mW / cm²) were used every 8 hours. 2 Cells were irradiated for 10 minutes. After culturing for 24 hours, the supernatant and cells were collected for further study. HUVEC proliferation was measured using CCK8 assay (method same as in Example 1), and the results are as follows. Figure 6 As shown in the figure. The results showed that more apoptotic HUVECs were found in the high glucose environment compared with the control group, and CNT50 significantly enhanced the cell viability of HUVECs under photocatalytic conditions compared with dark conditions, indicating that CNT50 has the effect of accelerating the activation of HUVEC cells in a high glucose microenvironment.

[0080] Experiment 6: CNT50-mediated photocatalytic therapy inhibits apoptosis in HUVEC cells in a high-glucose environment

[0081] The culture method for HUVECs was the same as in Example 5. Cultured cells were collected, and TUNEL assays were performed using a TUNEL kit according to the manufacturer's instructions. Images were examined using a confocal microscope, and quantitative analysis was performed using ImageJ software. The results are as follows: Figure 7 As shown, the more cells stained by TUNEL and the more pronounced the color, the higher the degree of apoptosis. Observation of the number and degree of TUNEL staining revealed that CNT50 significantly inhibited apoptosis in HUVECs in a high-glucose environment under light conditions compared to dark conditions, demonstrating that photocatalytic conditions enhance the inhibitory effect of CNT50 on apoptosis in HUVECs in a high-glucose environment.

[0082] Experiment 7: CNT50-mediated photocatalytic therapy promotes angiogenesis in HUVECs in a high-glucose environment

[0083] The HUVEC culture method was the same as in Experiment 5. Cultured cells were seeded into 96-well plates pre-coated with Matrigel, and the drug (the same as in Example 5) was added and incubated with the HUVECs for 30 minutes. Then, glucose (25 mM) was added to stimulate the cells. After incubating the 96-well plates at 37°C and 5% CO2 for 6 hours, the cells were visualized using a confocal microscope. Capillary network images of at least three random regions were captured, and the grid number of the tubes was calculated using ImageJ software. The results are shown below. Figure 8 As shown in the figure. The results show that CNT50 can promote angiogenesis of HUVECs more significantly under light conditions than under dark conditions, proving that photocatalytic conditions can enhance the role of CNT50 in promoting angiogenesis of HUVECs in high-glucose environments.

[0084] Experiment 8: CNT50-mediated photocatalytic therapy inhibits the inflammatory response of HUVECs in a high-glucose environment.

[0085] The HUVEC cell culture method was the same as in Example 6. The cells were incubated with the same drug (as in Example 6) for 30 minutes, followed by stimulation with glucose (25 mM). HUVECs were collected, and the levels of IL-1β, TNF-α, and VEGF in the HUVEC cell culture supernatant were measured according to the ELISA kit instructions. The results are as follows: Figure 9 As shown in the figure. The results showed that CNT50 could more significantly inhibit the production of inflammatory factors (IL-1β, TNF-α) in HUVECs under light conditions compared with dark conditions, while increasing the expression level of angiogenesis-related growth factor (VEGF). This indicates that photocatalysis can enhance the role of CNT50 in alleviating the inflammatory response of HUVECs in a high-glucose environment, while also promoting angiogenesis.

[0086] Experiment 9: CNT50-mediated photocatalytic therapy promotes wound healing in diabetic mice

[0087] Following previous experimental methods, a streptozotocin (STZ)-induced diabetic mouse model was used. Male ICR mice (20–25 g) were intraperitoneally injected with STZ (80 mg / kg, dissolved in 100 μL citrate buffer, pH 4.2–4.5) once daily for 5 consecutive days. After 4 weeks, mice with fasting blood glucose concentrations exceeding 16.6 mM were classified as diabetic mice. After anesthesia with 3% isoflurane and hair removal, a full-thickness skin wound with a diameter of 10 mm was created on the back of each mouse using a circular skin biopsy punch. CNT50 (10 mg / kg) or its petroleum jelly carrier was applied topically to the wound on days 1, 3, 5, and 7 postoperatively. For the light exposure group, mice were exposed to sunlight for 3 × 15 minutes daily. Wound healing was observed and photographed on days 1, 5, and 11 postoperatively, and the wound area was measured and analyzed using ImageJ software. The results are shown below. Figure 10 As shown in the figure. We then investigated the effect of CNT50-mediated photocatalytic therapy on wound healing in diabetic mice. The results showed that under sunlight, CNT50 significantly promoted wound healing in diabetic mice compared to darkness, indicating that CNT50 has a role in promoting wound healing in diabetic mice under light conditions.

[0088] Experiment 10: CNT50-mediated photocatalytic therapy promotes granulation tissue formation in diabetic mice

[0089] After modeling, skin specimens were collected from the backs of diabetic mice and fixed in 10% neutral pH buffered formalin at 4°C. The specimens were then embedded in paraffin and sectioned to obtain sections with a thickness of 5 μm. The sections were stained with hematoxylin and eosin (H&E), and finally observed and analyzed using an optical microscope. The results are as follows: Figure 11 As shown in the figure. The results showed that, compared with the dark group, the CNT50 light-exposed group had more abundant granulation tissue, smaller scars, and thicker new epidermis in the wound space on day 11. This indicates that CNT50 has the effect of promoting the formation of new granulation tissue and promoting wound healing in diabetic mice under light conditions.

[0090] Experiment 11: CNT50-mediated photocatalytic therapy alleviates wound scarring in diabetic mice

[0091] Paraffin sections of prepared skin specimens from the backs of diabetic mice were dewaxed, rehydrated, and then stained with Masson's stain according to the Solarbio manufacturer's instructions. Finally, the sections were observed and analyzed using an optical microscope. The results are as follows: Figure 11As shown in the figure. The results showed that CNT50 significantly promoted collagen deposition and reduced wound area under light conditions compared to dark conditions, indicating that CNT50 can alleviate wound scarring and promote wound healing in diabetic mice under light conditions.

[0092] Experiment 12: CNT50-mediated photocatalytic therapy promotes collagen deposition in wounds of diabetic mice

[0093] Paraffin sections of prepared skin specimens from the backs of diabetic mice were dewaxed, rehydrated, and then subjected to immunofluorescence staining. After dewaxing and antigen retrieval, slides were incubated overnight at 4°C with primary antibodies (anti-CD31 and α-SMA), respectively. Following incubation, the sections were washed with 0.1M PBS and incubated for 1 hour at room temperature with either Alexa Fluor 488 or IgG-Alexa Fluor 555. After washing, the sections were mounted with DAPI and observed under a confocal microscope. Fluorescence intensity was measured using ImageJ software. The results are shown below. Figure 12 As shown, activated fibroblasts are an important source of collagen deposition, and the number of cells expressing α-SMA significantly increased in mice under light exposure after CNT50 was introduced. CD31 is an endogenous angiogenic factor, mainly expressed by immune cells and endothelial cells, and is particularly important for maintaining skin vascular homeostasis. The sunlight group of CNT50 showed higher CD31 expression in wound tissue. CNT50 significantly promoted collagen deposition and angiogenesis under light conditions compared to dark conditions, indicating that CNT50 has a role in promoting wound healing in diabetic mice under light conditions.

[0094] The above findings collectively demonstrate that CNT50-mediated photocatalytic therapy can promote cell proliferation and migration, increase collagen deposition, and promote angiogenesis by releasing hydrogen gas and locally degrading glucose, ultimately accelerating the healing of diabetic wounds. The results of this invention demonstrate the potential clinical application of the titanium dioxide-based composite material provided by this invention in accelerating the healing process of diabetic wounds through photocatalytic hydrogen production and glucose consumption. This novel treatment strategy offers a new and effective treatment option for diabetic patients, potentially improving their quality of life and reducing the risk of complications.

[0095] Although the above embodiments have provided a detailed description of the present invention, they are only some embodiments of the present invention, and not all embodiments. Other embodiments can be obtained based on these embodiments without creative effort, and these embodiments all fall within the protection scope of the present invention.

Claims

1. A titanium dioxide-based composite material, characterized in that, Includes titanium dioxide nanoflowers and sheet-like carbon nitride supported on the titanium dioxide nanoflowers; The preparation method of the titanium dioxide-based composite material includes the following steps: Tetrabutyl titanate and an acidic reagent are mixed and subjected to a hydrothermal reaction to obtain amorphous titanium dioxide; The amorphous titanium dioxide was subjected to a first annealing to obtain titanium dioxide nanoflowers; the temperature of the first annealing was 300-500℃, and the holding time was 2-4h. A suspension is obtained by ultrasonically mixing flake carbon nitride and an organic solvent; The titanium dioxide nanoflowers and the suspension were mixed and subjected to a second annealing to obtain the titanium dioxide-based composite material; the temperature of the second annealing was 300-400℃ and the holding time was 1-2h. The method for preparing the flake-shaped carbon nitride includes: calcining melamine to obtain carbon nitride; mixing the carbon nitride with methanol and then annealing it to obtain the flake-shaped carbon nitride.

2. The titanium dioxide-based composite material according to claim 1, characterized in that, The loading percentage of flaky carbon nitride on the titanium dioxide-based composite material is 10-70%.

3. The method for preparing the titanium dioxide-based composite material according to claim 1 or 2, characterized in that, Includes the following steps: Tetrabutyl titanate and an acidic reagent are mixed and subjected to a hydrothermal reaction to obtain amorphous titanium dioxide; The amorphous titanium dioxide was subjected to a first annealing to obtain titanium dioxide nanoflowers; the temperature of the first annealing was 300-500℃, and the holding time was 2-4h. A suspension is obtained by ultrasonically mixing flake carbon nitride and an organic solvent; The titanium dioxide nanoflowers and the suspension were mixed and subjected to a second annealing to obtain the titanium dioxide-based composite material; the temperature of the second annealing was 300-400℃ and the holding time was 1-2h. The method for preparing the flake-shaped carbon nitride includes: calcining melamine to obtain carbon nitride; The carbon nitride and methanol were mixed and annealed to obtain the flake-shaped carbon nitride.

4. The preparation method according to claim 3, characterized in that, The acidic reagent includes one or more of acetic acid, hydrochloric acid, and lactic acid; The volume ratio of the tetrabutyl titanate to the acidic reagent is 1:10 to 50.

5. The preparation method according to claim 3, characterized in that, The hydrothermal reaction is carried out at a temperature of 100–150°C for 8–15 hours.

6. The preparation method according to claim 3, characterized in that, The ratio of the flake carbon nitride to the organic solvent is 0.05–0.35 g: 70 mL; The mass ratio of the sheet-like carbon nitride to titanium dioxide nanoflowers is 0.05–0.35:0.

5.

7. The application of the titanium dioxide-based composite material according to claim 1 or 2 or the titanium dioxide-based composite material prepared by the preparation method according to any one of claims 3 to 6 in the photocatalytic glucose depletion and hydrogen production reaction.

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