Alendronic acid modified ZIF-8 encapsulated TPP-carbon dot bone targeting nano material as well as preparation method and application thereof

Abstract

The invention relates to the technical field of biomedical materials, in particular to an alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone targeting nano-material and a preparation method and application thereof.The alendronate-modified ZIF-8 serves as a carrier, TPP-modified carbon dots are encapsulated in the carrier, citric acid, urea and cerium salt serve as raw materials of the TPP-modified carbon dots, and the alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone targeting nano-material is prepared through a one-step method. After cerium-doped carbon quantum dots are synthesized through a hydrothermal method, TPP is grafted through an amide method, alendronate endows the material with a bone targeting function, and TPP modified carbon dots endow the material with antioxidant and mitochondrial targeting functions. According to the invention, the bone targeting property is excellent, and bone focuses are efficiently enriched; the anti-oxidation and anti-inflammatory capacities are high, and the infected microenvironment can be improved; zIF-8 packaging is stable, and functional components can be controllably released; multiple components cooperate to promote bone repair, accumulation of non-target organs is little, and the composition is safe, reliable and suitable for treatment of various bone-related diseases.

Description

Technical Field

[0001] This invention relates to the field of biomedical materials technology, specifically to an alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterial, its preparation method, and its application. Background Technology

[0002] Bone defects, bone infections, and osteoporosis-related bone diseases are common challenges in orthopedic clinics. Their onset is often related to factors such as trauma, metabolic disorders, and infection. These conditions not only lead to bone structural damage and functional loss but can also cause complications such as infection spread, delayed healing, and malunion, severely impacting patients' quality of life. Among traditional treatments, autologous bone transplantation, while effective, suffers from limitations such as donor shortages and significant bone harvesting trauma. Allogeneic bone transplantation is prone to immune rejection and carries a high risk of infection. Repair materials such as metal implants and bioceramics have limited biocompatibility, lack of targeting, and difficulty adapting to the infection microenvironment, resulting in low utilization of effective components and potential side effects from accumulation in non-target organs.

[0003] With the development of nanomedicine, carbon quantum dots have attracted widespread attention in the field of bone repair due to their good biocompatibility, excellent antioxidant properties, and modifiability. However, carbon quantum dots alone lack tissue-specific targeting capabilities, making it difficult to accurately accumulate at bone lesion sites. Furthermore, their function is singular, failing to simultaneously address multiple needs such as infection suppression, inflammation regulation, and improved bone repair efficiency. ZIF-8, as a typical metal-organic framework material, possesses high specific surface area, controllable pore structure, and good biocompatibility, and is often used as a nanocarrier to achieve the encapsulation and controlled release of functional components. However, unmodified ZIF-8 lacks bone targeting, is easily taken up by non-target organs in vivo, and the encapsulated components may suffer from problems such as excessively rapid release and insufficient stability.

[0004] The application of targeted modification technology offers a solution to the above problems: alendronate can achieve bone-targeted enrichment through specific chelation with hydroxyapatite in bone tissue, but its use alone is insufficient to address cellular-level inflammation regulation; triphenylphosphine (TPP), as a mitochondrial-targeting molecule, can mediate the entry of functional components into mitochondria and inhibit oxidative stress and inflammatory responses, but it lacks bone tissue specificity, resulting in low enrichment efficiency at lesion sites. Currently, there are no reports on the organic integration of alendronate-modified ZIF-8 carriers, TPP-targeted carbon dots, and cerium-doped antioxidant functions. Existing materials often suffer from insufficient targeting precision, poor synergy between antioxidant and bone-repairing effects, and poor stability of encapsulated components, making it difficult to meet the comprehensive treatment needs of complex bone diseases. Therefore, there is an urgent need to develop a nanomaterial that combines precise bone targeting, efficient antioxidant activity, stable encapsulation, and synergistic repair functions. Summary of the Invention

[0005] (a) Technical problems to be solved To address the shortcomings of existing technologies, this invention provides an alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterial, its preparation method, and its application.

[0006] (II) Technical Solution A bone-targeting nanomaterial with alendronate-modified ZIF-8 encapsulated TPP-carbon dots is disclosed. The nanomaterial uses an alendronate-modified zeolite imidazole ester framework material-8 (ZIF-8) as a carrier, with TPP-modified carbon dots encapsulated within the carrier. The TPP-modified carbon dots are cerium-doped carbon quantum dots synthesized via a hydrothermal method using citric acid, urea, and cerium salts. TPP with mitochondrial-targeting function is then grafted onto the surface of the cerium-doped carbon quantum dots using an amide method. The alendronate imparts bone-targeting functionality to the nanomaterial, while the TPP-modified carbon dots impart antioxidant and mitochondrial-targeting functions.

[0007] Preferably, the nanomaterial has a particle size of 80-150 nm, and after being placed in a phosphate buffer solution at pH 7.4 for 72 h, the dispersion index (PDI) is ≤0.25, with no obvious agglomeration or precipitation; the cerium salt includes one or more of cerium nitrate, cerium chloride, cerium sulfate, cerium acetate, cerium iodide, and cerium bromide.

[0008] Preferably, the method for preparing the bone-targeting nanomaterial of ZIF-8 encapsulated with TPP-carbon dots modified with alendronate involves synthesizing ZIF-8 and simultaneously encapsulating TPP-modified carbon dots; mixing and stirring the obtained ZIF-8 encapsulated with TPP-modified carbon dots with sodium alendronate trihydrate to graft alendronate onto the surface of the ZIF-8 encapsulated with TPP-modified carbon dots, forming ZIF-8 encapsulated with TPP-carbon dots modified with alendronate; wherein, the preparation process of the TPP-modified carbon dots is as follows: cerium-doped carbon quantum dots are synthesized by hydrothermal method using citric acid, urea, and cerium salt as raw materials, and then TPP is grafted onto the surface of the cerium-doped carbon quantum dots by amide method.

[0009] Preferably, ZIF-8 is synthesized by a solvothermal method, with zinc nitrate and 2-methylimidazole as the reaction raw materials, methanol as the solvent, a molar ratio of zinc nitrate to 2-methylimidazole of 1:9-1:11, a reaction temperature of 25-30℃, and a reaction time of 10 minutes.

[0010] Preferably, ZIF-8 is purified by centrifugation at 12000 r / min for 10 min, washing with methanol 3-4 times, and vacuum drying at -80℃ for 5-6 h.

[0011] Preferably, the molar ratio of ZIF-8 to sodium alendronate trihydrate is 1:2-1:4. During mixing, deionized water is used as the dispersant, and the grafting is achieved by stirring at 25-28℃ and 250-350 r / min for 0.5 h.

[0012] Preferably, before encapsulation, TPP-modified carbon dots are dispersed by ultrasonic assistance and mixed with 2-methylimidazole until homogeneous. The ultrasonic power is 300-450W and the ultrasonic time is 10min. During the ultrasonic process, the system temperature is maintained at 28-32℃ by water bath temperature control.

[0013] Preferably, the product is further processed by centrifugation at 12000 r / min for 10 min, washing with deionized water 3 times, and vacuum freeze-drying at -80℃ for 5-6 h.

[0014] Preferably, the alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterial is used in bone tissue repair, treatment of bone infection, or targeted therapy for bone-related diseases.

[0015] Preferably, in the treatment of bone infection, the nanomaterial can be targeted and enriched at the site of bone infection, and the antioxidant function of TPP-modified carbon dots can inhibit the inflammatory response and help improve the infection microenvironment.

[0016] (iii) Beneficial technical effects Compared with existing technologies, the beneficial effects of this invention are: 1. This material uses alendronic acid-modified ZIF-8 as a carrier. By leveraging the specific chelation between alendronic acid and hydroxyapatite in bone tissue, the material's targeted accumulation ability at bone lesion sites is significantly enhanced, while the accumulation in non-target organs is significantly reduced, effectively reducing potential side effects and improving biosafety.

[0017] 2. The high specific surface area and controllable pore size structure of the ZIF-8 carrier enable efficient encapsulation of TPP-modified carbon dots. This not only prevents the functional components from being degraded and inactivated during in vivo transport but also regulates their release rate, ensuring sustained efficacy at the lesion site. This solves the problems of excessively rapid release or insufficient stability associated with traditional nanomaterials. Simultaneously, the TPP-modified carbon dots retain the excellent antioxidant function brought by cerium doping, and through the mitochondrial targeting action of TPP, they can precisely enter the mitochondria, inhibiting oxidative stress and the release of inflammatory factors, effectively improving the infection microenvironment and creating favorable conditions for bone repair.

[0018] 3. A synergistic effect of "bone-targeted enrichment - mitochondrial inflammation regulation - stable and controllable release" is formed among the components: bone-targeting function ensures the material accurately reaches the lesion, antioxidant function regulates the microenvironment, and the ZIF-8 carrier ensures functional stability and controllable release. The organic combination of these three elements achieves integrated satisfaction of multiple therapeutic needs. This nanomaterial exhibits good biocompatibility, a simple and controllable preparation method, high batch-to-batch reproducibility, and a wide range of applications. It can be used for bone tissue repair, treatment of bone infections, and targeted therapy for various bone-related diseases. Compared with existing technologies, its functions are more comprehensive, its targeting is more precise, and its application is more flexible, significantly expanding the application scenarios of nanomaterials in the field of orthopedics and possessing significant clinical translational value. Attached Figure Description

[0019] Figure 1 Transmission electron microscopy (TEM) image of alendronic acid-modified ZIF-8-encapsulated TPP-carbon dot bone-targeting nanomaterials; Figure 2 Particle size distribution curve of TPP-carbon dot bone-targeting nanomaterials encapsulated with alendronic acid-modified ZIF-8; Figure 3 X-ray photoelectron spectroscopy (XPS) of alendronic acid-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials; Figure 4 A comparison of Fourier transform infrared (FT-IR) spectra of pure ZIF-8 and alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials (P-ZIF). Figure 5 The 1H NMR spectrum of pure ZIF-8, sodium alendronate (ALN), and alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials (P-ZIF) 1 H NMR comparison image. Detailed Implementation

[0020] according to Figures 1 to 5 The specific embodiments of the present invention are as follows: Example 1: Preparation of alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials using cerium nitrate as the cerium salt 1. Preparation of TPP-modified carbon dots (TPP-Ce@CDs) Raw material preparation: Weigh 5.0g of analytical grade citric acid, 10.0g of urea, and cerium nitrate hexahydrate with a purity of 98%, and strictly control the mass ratio of the three to be 5:10:1; take triphenylphosphine (TPP) with a purity of 98.5%, place it in a vacuum distillation apparatus, set the distillation temperature to 225℃ and the vacuum degree to 0.09MPa, distill for 30min to remove low-boiling-point organic impurities, cool to room temperature, and then seal and store in the dark.

[0021] Hydrothermal synthesis of cerium-doped carbon quantum dots (Ce@CDs): Citric acid, urea, and cerium nitrate hexahydrate were added to 50 mL of deionized water and placed on a magnetic stirrer. The stirring speed was adjusted to 350 r / min and stirred continuously for 45 min until the raw materials were completely dissolved to form a transparent solution without visible particles. The pH of the system was slowly adjusted to 6.0 with 0.1 mol / L hydrochloric acid solution and monitored in real time with a precision pH meter with an accuracy of 0.01. The solution was then transferred to a 100 mL polytetrafluoroethylene-lined hydrothermal reactor, with the solution volume controlled at 70 mL and the filling degree at 70% to prevent the solution from boiling over during the reaction. The reactor was then sealed and placed in a forced-air drying oven, and the temperature was increased to 180 °C at a rate of 5 °C / min and kept at a constant temperature for 6 h. After the reaction was completed, the drying oven was closed, and the reactor was allowed to cool naturally to below 80 °C. The reactor was then slowly opened, and the pale yellow reaction solution was collected. The solution was filtered under reduced pressure using a 0.22 μm aqueous filter membrane to remove unreacted solid particles and impurities, yielding a pure Ce@CDs solution.

[0022] Amide grafting of TPP: Take 50 mL of the above Ce@CDs solution, weighing it to calculate that it contains approximately 1.0 g of Ce@CDs. Transfer it to a 250 mL three-necked flask and add 50 mL of anhydrous N,N-dimethylformamide (DMF), with a water content ≤0.05%, as the reaction solvent. Purge one port of the three-necked flask with high-purity nitrogen gas (99.999% purity) at a flow rate of 10 mL / min for 20 min to remove oxygen from the system and prevent oxygen from interfering with amide bond formation. Weigh 0.14 g of purified TPP according to a TPP to Ce@CDs mass ratio of 1:7 and add it to the flask through the other port. Turn on the magnetic stirrer and adjust the speed to 300 r / min. Simultaneously, the oil bath temperature was raised to 70℃ and the reaction was maintained at this temperature for 3 hours, with nitrogen continuously purging for protection during the reaction. After the reaction, the oil bath was turned off, and the reaction solution was allowed to cool to room temperature. It was then transferred to a dialysis bag with a molecular weight cutoff of 2000 Da and dialyzed at room temperature for 30 hours using deionized water as the dialysis medium. Fresh deionized water was replaced every 7 hours, with each replacement being twice the volume of the original medium in the dialysis container, to ensure that unreacted TPP and DMF were fully removed. After dialysis, the TPP-Ce@CDs dispersion in the dialysis bag was transferred to a freeze-drying bottle and placed in a vacuum freeze dryer at -80℃ for 24 hours to obtain a pale yellow TPP-Ce@CDs powder, which was then sealed and stored away from light.

[0023] 2. Synthesize ZIF-8 and simultaneously encapsulate TPP-Ce@CDs Ultrasonic dispersion of TPP-Ce@CDs: Weigh 0.2g of the above TPP-Ce@CDs powder, add 20mL of analytical grade methanol (99.5% purity), transfer to an ultrasonic cleaner, set the ultrasonic power to 350W, and simultaneously turn on the water bath temperature control device to keep the system temperature stable at 30℃ to prevent overheating and particle agglomeration. Continue ultrasonic treatment for 10min. After ultrasonic treatment, add 4.86g of 2-methylimidazole (98% purity) to the dispersion, place it on a magnetic stirrer and stir at 250r / min for 10min until the 2-methylimidazole is completely dissolved to obtain a 2-methylimidazole methanol solution containing TPP-Ce@CDs.

[0024] Solvent-thermal synthesis of encapsulated ZIF-8: Weigh 1.48 g of zinc nitrate hexahydrate (98% purity), dissolve it in 10 mL of methanol, and stir until completely dissolved to obtain a zinc nitrate methanol solution. Slowly add the zinc nitrate methanol solution dropwise through a constant-pressure dropping funnel to the above-mentioned 2-methylimidazole methanol solution containing TPP-Ce@CDs, controlling the dropping rate at 1 mL / min to ensure uniform reaction. At this point, the molar ratio of zinc nitrate to 2-methylimidazole is 1:10. After the addition is complete, allow the reaction to stand at room temperature (28℃) for 10 min. During this period, the solution gradually becomes white and turbid, indicating that ZIF-8 crystals have formed and TPP-Ce@CDs are simultaneously encapsulated. The reaction results... After washing, the mixture was transferred to a 50 mL centrifuge tube and centrifuged at 12000 r / min for 10 min in a high-speed centrifuge. The supernatant containing unreacted zinc nitrate and 2-methylimidazole was discarded, and the white precipitate was collected. 20 mL of methanol was added to the precipitate, and the mixture was vortexed and centrifuged again at 12000 r / min for 10 min. The washing was repeated 4 times to ensure the removal of residual raw materials. The washed precipitate was transferred to a vacuum drying bottle and placed in a vacuum drying oven at -80℃ with a vacuum degree of 0.09 MPa for 5.5 h to obtain ZIF-8 powder for encapsulating TPP-Ce@CDs (ZIF-8@TPP-Ce@CDs).

[0025] 3. Alendronate modification Weigh 1.0 g of ZIF-8 powder encapsulating TPP-Ce@CDs, add it to 50 mL of deionized water, and stir on a magnetic stirrer at 200 r / min for 10 min to ensure uniform dispersion of the powder. Weigh alendronate sodium trihydrate at a molar ratio of ZIF-8 to alendronate sodium trihydrate of 1:3, add it to the above dispersion, adjust the stirring speed to 300 r / min, and set the constant temperature water bath temperature to 26℃, and continue stirring for 0.5 h. After the reaction is complete, transfer the mixture to a centrifuge tube and centrifuge at 12000 r / min for 10 min to collect the precipitate. Add 30 mL of deionized water to the precipitate, sonicate at 200 W for 1 min, centrifuge again, and repeat the washing process 3 times. Transfer the washed precipitate to a freeze-drying bottle, place it in a vacuum freeze dryer at -80℃ with a vacuum degree of 0.09 MPa, and dry for 5.5 h to obtain alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials.

[0026] Product characterization Transmission electron microscopy revealed that the product exhibited uniform dispersion at different scales of 1μm, 500nm, 200nm, and 100nm, with no obvious agglomeration. The particles were generally spherical with regular morphology, and the ZIF-8 carrier encapsulation layer was clearly visible. The TPP-Ce@CDs particles inside were uniformly dispersed without any aggregated blocks, confirming the effectiveness of the encapsulation process.

[0027] The particle size of the product was 172.95±2.83 nm, and the particle size distribution range was concentrated in the range of 80-420 nm, with the main peak corresponding to about 170 nm. The dispersion index (PDI) was 0.21, indicating that the product has good dispersion stability in aqueous solution.

[0028] X-ray photoelectron spectroscopy (XPS) analysis detected characteristic peaks for Ce 3d, N 1s, P 2p, Zn 2p, and O 1s, confirming the success of cerium doping, TPP grafting, ZIF-8 support, and alendronate modification. The Zn 2p characteristic peaks, located near 1024.8 eV and 1047.6 eV, correspond to zinc in ZIF-8. The Ce 3d characteristic peak confirms successful cerium doping. The P 2p characteristic peak originates from phosphorus in TPP and alendronate, exhibiting symmetrical peak shape and stable intensity, indicating strong phosphorus binding. The N 1s characteristic peak further corroborates the presence of nitrogen in TPP and ZIF-8. The absence of significant impurity peaks in the characteristic peaks indicates good product purity.

[0029] Fourier transform near-infrared spectroscopy was used to compare the spectral curves of pure ZIF-8 and the product (P-ZIF). The product's spectral curves were found to be in the range of 1000-1300 cm⁻¹. -1The region exhibits distinct P=O stretching vibration peaks and POC characteristic absorption peaks, corresponding to the phosphorus-oxygen bond in alendronic acid; at 1600-1700 cm⁻¹... -1 The presence of characteristic absorption peaks for amide bonds in the region confirms that alendronate was successfully grafted onto ZIF-8@TPP-Ce@CDs via amide bonds.

[0030] By comparing the chemical shift curves of pure ZIF-8, sodium alendronate (ALN), and the product (P-ZIF) using 1H NMR spectroscopy, the product showed characteristic hydrogen peaks unique to sodium alendronate in the 2.0-4.0 ppm range, with stable peak shape and intensity, indicating that sodium alendronate had been successfully grafted onto the support surface. At the same time, the characteristic hydrogen peaks of ZIF-8 were retained, confirming that the grafting process did not damage the framework structure of ZIF-8, and the product structure was well intact.

[0031] Example 2: Preparation of alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials using cerium chloride as the cerium salt. 1. Preparation of TPP-modified carbon dots (TPP-Ce@CDs) Raw material preparation: Weigh 5.0g of citric acid, 9.5g of urea, and cerium chloride hexahydrate with a purity of 98%, in a mass ratio of 5:9.5:0.95; take TPP, purify it by vacuum distillation (220℃, 0.08MPa) for 30min, and cool it for later use.

[0032] Hydrothermal synthesis of Ce@CDs: The above raw materials were dissolved in 50 mL of deionized water and stirred on a magnetic stirrer at 300 r / min for 40 min until completely dissolved. The pH of the system was adjusted to 5.5 with 0.1 mol / L sodium hydroxide solution. The solution was then transferred to a 100 mL polytetrafluoroethylene-lined hydrothermal reactor, with the solution volume controlled at 65 mL and the filling degree at 65%. The reactor was sealed and placed in a forced-air drying oven, and the temperature was increased to 170 °C at a rate of 5 °C / min and kept at a constant temperature for 7 h. After the reaction was completed, the drying oven was closed and the reactor was allowed to cool naturally to below 80 °C. The reactor was then opened to collect the reaction solution, which was filtered through a 0.22 μm aqueous filter membrane to remove unreacted solid particles, yielding the Ce@CDs solution.

[0033] Amide grafting of TPP: Take a solution containing 1.0 g Ce@CDs, add 50 mL of anhydrous DMF, and purge with high-purity nitrogen (99.999% purity) at a flow rate of 10 mL / min for 15 min; weigh 0.2 g of purified TPP into the system at a TPP to Ce@CDs mass ratio of 1:5, raise the oil bath temperature to 60℃, turn on the magnetic stirrer and adjust the speed to 250 r / min, and react at a constant temperature for 4 h; after the reaction is completed, transfer the solution to a dialysis bag with a molecular weight cutoff of 1500 Da, dialyze with deionized water at room temperature for 24 h, and change the deionized water every 6 h; after dialysis, freeze-dry the dispersion to obtain TPP-Ce@CDs powder.

[0034] 2. Synthesize ZIF-8 and simultaneously encapsulate TPP-Ce@CDs Ultrasonic dispersion: Weigh 0.2g of TPP-Ce@CDs powder, add 20mL of methanol, transfer to an ultrasonic cleaner, set the ultrasonic power to 400W and the water bath temperature to 28℃, and sonicate for 10min; after sonication, add 4.37g of 2-methylimidazole to the dispersion, at which point the molar ratio of zinc nitrate to 2-methylimidazole is 1:9, and stir on a magnetic stirrer until completely dissolved.

[0035] Solvent-thermal synthesis: Weigh 1.48 g of zinc nitrate hexahydrate and dissolve it in 10 mL of methanol by stirring until dissolved to obtain a zinc nitrate methanol solution; slowly add this solution dropwise to the above 2-methylimidazolium methanol solution containing TPP-Ce@CDs, and after the addition is complete, let it stand at room temperature (25 °C) for 10 min; after the reaction is complete, transfer the mixture to a centrifuge tube and centrifuge at 12000 r / min for 10 min, discard the supernatant and collect the precipitate; wash the precipitate with 20 mL of methanol, vortex to mix and centrifuge, repeat 3 times; place the washed precipitate in a vacuum drying oven at -80 °C (0.08 MPa) and dry for 5 h to obtain ZIF-8@TPP-Ce@CDs powder.

[0036] 3. Alendronate modification Weigh 1.0 g of ZIF-8@TPP-Ce@CDs powder, add 50 mL of deionized water, and stir on a magnetic stirrer for 10 min to disperse it. Weigh alendronate sodium trihydrate (purity 98%) according to the molar ratio of ZIF-8 to alendronate sodium trihydrate of 1:2 and add it to the system. Stir at 250 r / min for 0.5 h at 25 °C. After the reaction is complete, centrifuge at 12000 r / min for 10 min to collect the precipitate, wash with 30 mL of deionized water, sonicate for 1 min and centrifuge, repeat 3 times. Place the precipitate in a vacuum freeze dryer at -80 °C and dry for 5 h to obtain the target nanomaterial.

[0037] Example 3: Preparation of alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials using cerium sulfate as the cerium salt. 1. Preparation of TPP-modified carbon dots (TPP-Ce@CDs) Raw material preparation: Weigh 5.0g of citric acid, 10.5g of urea, and cerium sulfate octahydrate with a purity of 98%, in a mass ratio of 5:10.5:1.05; take TPP, purify it by vacuum distillation (230℃, 0.1MPa) for 30min, cool it, and seal it for later use.

[0038] Hydrothermal synthesis of Ce@CDs: The above raw materials were dissolved in 50 mL of deionized water and stirred on a magnetic stirrer at 450 r / min for 50 min until completely dissolved. The pH of the system was adjusted to 6.5 with 0.1 mol / L hydrochloric acid solution. The solution was then transferred to a 100 mL polytetrafluoroethylene-lined hydrothermal reactor, with the solution volume controlled at 75 mL and the filling degree at 75%. The reactor was sealed and placed in a forced-air drying oven, and the temperature was increased to 190 °C at a rate of 5 °C / min and kept at a constant temperature for 4 h. After the reaction was completed, the temperature was cooled to below 80 °C, and the reaction solution was collected and filtered through a 0.22 μm aqueous filter membrane to obtain the Ce@CDs solution.

[0039] Amide grafting of TPP: Take a solution containing 1.0 g Ce@CDs, add 50 mL of anhydrous DMF, and purge with high-purity nitrogen (99.999% purity) at a flow rate of 10 mL / min for 25 min; weigh 0.11 g of purified TPP into the system at a TPP to Ce@CDs mass ratio of 1:9, raise the oil bath temperature to 80℃, adjust the stirring speed to 350 r / min, and react at a constant temperature for 2 h; after the reaction, transfer the solution to a dialysis bag with a molecular weight cutoff of 2500 Da, dialyze at room temperature for 36 h, and replace the deionized water every 8 h; after dialysis, freeze-dry to obtain TPP-Ce@CDs powder.

[0040] 2. Synthesize ZIF-8 and simultaneously encapsulate TPP-Ce@CDs Ultrasonic dispersion: Weigh 0.2g of TPP-Ce@CDs powder, add 20mL of methanol, transfer to an ultrasonic cleaner, set the ultrasonic power to 450W and the water bath temperature to 32℃, and sonicate for 10min; after sonication, add 5.35g of 2-methylimidazole to the dispersion, at which point the molar ratio of zinc nitrate to 2-methylimidazole is 1:11, and stir until completely dissolved.

[0041] Solvent-thermal synthesis: Weigh 1.48 g of zinc nitrate hexahydrate and dissolve it in 10 mL of methanol by stirring to obtain a zinc nitrate methanol solution; slowly add this solution dropwise to the above dispersion, and after the addition is complete, let it stand at room temperature (30 °C) for 10 min; after the reaction, centrifuge the mixture and collect the precipitate; wash the precipitate four times with 20 mL of methanol, centrifuging for 10 min each time; place the precipitate in a vacuum drying oven at -80 °C (0.1 MPa) and dry for 6 h to obtain ZIF-8@TPP-Ce@CDs powder.

[0042] 3. Alendronate modification Weigh 1.0 g of ZIF-8@TPP-Ce@CDs powder and add 50 mL of deionized water to disperse it. Weigh alendronate sodium trihydrate into the system according to the molar ratio of ZIF-8 to alendronate sodium trihydrate of 1:4. Stir at 350 r / min for 0.5 h at 28 °C. After the reaction, collect the precipitate by centrifugation, wash it three times with deionized water, and centrifuge after ultrasonic dispersion for 1 min each time. Place the precipitate in a vacuum freeze dryer at -80 °C and dry it for 6 h to obtain the target nanomaterial.

[0043] Example 4: Preparation of alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials using cerium acetate as the cerium salt 1. Preparation of TPP-modified carbon dots (TPP-Ce@CDs) Raw material preparation: Weigh 5.0g of citric acid, 10.2g of urea, and cerium acetate trihydrate with a purity of 98%, in a mass ratio of 5:10.2:1.02; take TPP, purify it by vacuum distillation (225℃, 0.09MPa) for 30min, and cool it for later use.

[0044] Hydrothermal synthesis of Ce@CDs: The above raw materials were dissolved in 50 mL of deionized water and stirred at 320 r / min for 42 min on a magnetic stirrer. After complete dissolution, the pH was adjusted to 6.2 with 0.1 mol / L hydrochloric acid. The solution was transferred to a 100 mL hydrothermal reactor, and the solution volume was controlled to be 68 mL with a filling degree of 68%. After sealing, the reactor was placed in a forced-air drying oven and heated to 175 °C at 5 °C / min. The reaction was kept at this temperature for 5 h. After cooling, the reaction solution was collected and filtered through a 0.22 μm aqueous filter membrane to obtain the Ce@CDs solution.

[0045] Amide grafting of TPP: Take a solution containing 1.0 g Ce@CDs, add 50 mL of anhydrous DMF, and purge with high-purity nitrogen for 18 min; weigh 0.17 g TPP into the system at a TPP to Ce@CDs mass ratio of 1:6, raise the oil bath temperature to 65℃, stir at 280 r / min, and react for 3.5 h; after the reaction, transfer the solution to a dialysis bag with a molecular weight cutoff of 1800 Da, dialyze at room temperature for 28 h, and replace the deionized water every 7 h; after dialysis, freeze dry to obtain TPP-Ce@CDs powder.

[0046] 2. Synthesize ZIF-8 and simultaneously encapsulate TPP-Ce@CDs Weigh 0.2g of TPP-Ce@CDs powder, add 20mL of methanol, transfer to an ultrasonic cleaner, set the ultrasonic power to 380W and the water bath temperature to 29℃, and sonicate for 10min. After sonication, add 5.11g of 2-methylimidazole, at which point the molar ratio of zinc nitrate to 2-methylimidazole is 1:10.5, and stir until dissolved. Weigh 1.48g of zinc nitrate hexahydrate and dissolve it in 10mL of methanol, and slowly add it dropwise to the above dispersion. Let it stand at 27℃ for 10min. After the reaction, centrifuge (12000r / min, 10min), collect the precipitate and wash it 3 times with methanol. Place the precipitate in a vacuum drying oven at -80℃ with a vacuum degree of 0.09MPa and dry for 5.5h to obtain ZIF-8@TPP-Ce@CDs powder.

[0047] 3. Alendronate modification Weigh 1.0 g of ZIF-8@TPP-Ce@CDs powder and add 50 mL of deionized water to disperse by stirring. Weigh alendronate sodium trihydrate into the system according to the molar ratio of ZIF-8 to alendronate sodium trihydrate of 1:3.5, and stir at 320 r / min for 0.5 h at 27 °C. After the reaction, collect the precipitate by centrifugation and wash it three times with deionized water. Place the precipitate in a vacuum freeze dryer at -80 °C and dry it for 5.5 h to obtain the target nanomaterial.

[0048] Example 5: Preparation of alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials using cerium iodide as the cerium salt. 1. Preparation of TPP-modified carbon dots (TPP-Ce@CDs) Raw material preparation: Weigh 5.0g of citric acid, 9.8g of urea, and cerium iodide hexahydrate with a purity of 97%, in a mass ratio of 5:9.8:0.98; take TPP, purify it by vacuum distillation (228℃, 0.095MPa) for 30min, and cool it for later use.

[0049] Hydrothermal synthesis of Ce@CDs: The above raw materials were dissolved in 50 mL of deionized water and stirred at 380 r / min for 48 min on a magnetic stirrer. After complete dissolution, the pH was adjusted to 5.8 with 0.1 mol / L hydrochloric acid. The solution was transferred to a 100 mL hydrothermal reactor, and the solution volume was controlled to be 72 mL with a filling degree of 72%. After sealing, the reactor was placed in a forced-air drying oven and heated to 185 °C at 5 °C / min. The reaction was kept at this temperature for 5.5 h. After cooling, the reaction solution was collected and filtered through a 0.22 μm aqueous filter membrane to obtain the Ce@CDs solution.

[0050] Amide grafting of TPP: Take a solution containing 1.0 g Ce@CDs, add 50 mL of anhydrous DMF, and purge with high-purity nitrogen for 22 min (flow rate 10 mL / min); weigh 0.13 g TPP into the system according to the mass ratio of TPP to Ce@CDs 1:7.5, raise the oil bath temperature to 75℃, stir at 320 r / min, and react for 2.5 h; after the reaction, transfer the solution to a dialysis bag with a molecular weight cutoff of 2200 Da, dialyze at room temperature for 32 h, and change the deionized water every 7 h; after dialysis, freeze dry to obtain TPP-Ce@CDs powder.

[0051] 2. Synthesize ZIF-8 and simultaneously encapsulate TPP-Ce@CDs Weigh 0.2g of TPP-Ce@CDs powder, add 20mL of methanol, transfer to an ultrasonic cleaner, set the ultrasonic power to 420W and the water bath temperature to 30℃, and sonicate for 10min. After sonication, add 4.61g of 2-methylimidazole, at which point the molar ratio of zinc nitrate to 2-methylimidazole is 1:9.5, and stir until dissolved. Weigh 1.48g of zinc nitrate hexahydrate and dissolve it in 10mL of methanol, and slowly add it dropwise to the above dispersion. Let it stand at 26℃ for 10min. After the reaction, centrifuge (12000r / min, 10min), collect the precipitate and wash it 4 times with methanol. Place the precipitate in a vacuum drying oven at -80℃ with a vacuum degree of 0.095MPa and dry for 6h to obtain ZIF-8@TPP-Ce@CDs powder.

[0052] 3. Alendronate modification Weigh 1.0 g of ZIF-8@TPP-Ce@CDs powder and add 50 mL of deionized water to disperse by stirring. Weigh alendronate sodium trihydrate (purity 98%) into the system according to the molar ratio of ZIF-8 to alendronate sodium trihydrate of 1:2.5, and stir at 300 r / min for 0.5 h at 26 °C. After the reaction, collect the precipitate by centrifugation and wash it three times with deionized water. Place the precipitate in a vacuum freeze dryer at -80 °C and dry it for 6 h to obtain the target nanomaterial.

[0053] Example 6: Preparation of alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials using cerium bromide as the cerium salt 1. Preparation of TPP-modified carbon dots (TPP-Ce@CDs) Raw material preparation: Weigh 5.0g of citric acid, 10.1g of urea, and cerium bromide hexahydrate with a purity of 97%, in a mass ratio of 5:10.1:1.01; take TPP, purify it by vacuum distillation (222℃, 0.085MPa) for 30min, and cool it for later use.

[0054] Hydrothermal synthesis of Ce@CDs: The above raw materials were dissolved in 50 mL of deionized water and stirred at 400 r / min for 45 min on a magnetic stirrer. After complete dissolution, the pH was adjusted to 6.1 with 0.1 mol / L hydrochloric acid. The solution was transferred to a 100 mL hydrothermal reactor, and the solution volume was controlled to be 70 mL with a filling degree of 70%. After sealing, the reactor was placed in a forced-air drying oven and heated to 180 °C at 5 °C / min. The reaction was kept at this temperature for 6 h. After cooling, the reaction solution was collected and filtered through a 0.22 μm aqueous filter membrane to obtain the Ce@CDs solution.

[0055] Amide grafting of TPP: Take a solution containing 1.0 g Ce@CDs, add 50 mL of anhydrous DMF, and purge with high-purity nitrogen for 20 min (flow rate 10 mL / min); weigh 0.125 g TPP into the system at a TPP to Ce@CDs mass ratio of 1:8, raise the oil bath temperature to 72℃, stir at 300 r / min, and react for 3 h; after the reaction, transfer the solution to a dialysis bag with a molecular weight cutoff of 2000 Da, dialyze at room temperature for 30 h, and change the deionized water every 7 h; after dialysis, freeze dry to obtain TPP-Ce@CDs powder.

[0056] 2. Synthesize ZIF-8 and simultaneously encapsulate TPP-Ce@CDs Weigh 0.2g of TPP-Ce@CDs powder, add 20mL of methanol, transfer to an ultrasonic cleaner, set the ultrasonic power to 430W and the water bath temperature to 31℃, and sonicate for 10min. After sonication, add 4.98g of 2-methylimidazole, at which point the molar ratio of zinc nitrate to 2-methylimidazole is 1:10.2, and stir until dissolved. Weigh 1.48g of zinc nitrate hexahydrate and dissolve it in 10mL of methanol, and slowly add it dropwise to the above dispersion. Let it stand at 29℃ for 10min. After the reaction, centrifuge (12000r / min, 10min), collect the precipitate and wash it 4 times with methanol. Place the precipitate in a vacuum drying oven at -80℃ with a vacuum degree of 0.085MPa and dry for 5.5h to obtain ZIF-8@TPP-Ce@CDs powder.

[0057] 3. Alendronate modification Weigh 1.0 g of ZIF-8@TPP-Ce@CDs powder and add 50 mL of deionized water to disperse by stirring. Weigh alendronate sodium trihydrate (purity 98%) into the system according to the molar ratio of ZIF-8 to alendronate sodium trihydrate of 1:3.2, and stir at 330 r / min for 0.5 h at 27 °C. After the reaction, centrifuge to collect the precipitate, wash it three times with deionized water, and place the precipitate in a vacuum freeze dryer at -80 °C for 5.5 h to obtain the target nanomaterial.

[0058] Comparative Example 1: ZIF-8 encapsulated TPP-carbon dot nanomaterials without alendronate modification 1. Preparation process Except for omitting the "alendronic acid modification" step, the rest of the operation is the same as in Example 1.

[0059] Preparation of TPP-Ce@CDs: Following the same steps as in Example 1, the same raw materials were weighed and the same process parameters were controlled to obtain TPP-Ce@CDs powder.

[0060] ZIF-8 Synthesis and Encapsulation: Following the same steps as in Example 1, TPP-Ce@CDs were encapsulated into ZIF-8 to obtain ZIF-8@TPP-Ce@CDs powder. Without alendronic acid modification, the powder was directly placed in a vacuum freeze dryer at -80℃ and dried for 3.5 hours to obtain the product of Comparative Example 1.

[0061] Comparative Example 2: Alendronate-modified ZIF-8 encapsulated carbon dot nanomaterials without TPP modification 1. Preparation process Except for the step of “TPP-modified carbon dots” being changed to “preparation of unmodified TPP Ce@CDs”, the rest of the operation is the same as in Example 2.

[0062] Ce@CDs preparation: Following the same steps as in Example 2, citric acid, urea, and cerium chloride were weighed, and the same mass ratio and hydrothermal process were controlled to obtain a Ce@CDs solution; the step of grafting TPP with amide was omitted, and the Ce@CDs solution was directly freeze-dried to obtain Ce@CDs powder.

[0063] ZIF-8 Synthesis and Encapsulation: Following the same steps as in Example 2, Ce@CDs were encapsulated into ZIF-8 to obtain ZIF-8@Ce@CDs powder.

[0064] Alendronate modification: Following the same steps as in Example 2, ZIF-8@Ce@CDs were modified with alendronate to obtain the product of Comparative Example 2.

[0065] Comparative Example 3: Alendronate-modified ZIF-8 nanomaterials without encapsulated TPP-carbon dots 1. Preparation process Except for omitting TPP-Ce@CDs in the “ZIF-8 synthesis” step, the rest of the operation is the same as in Example 3.

[0066] Synthesis of pure ZIF-8: Following the same steps as in Example 3, without adding TPP-Ce@CDs powder, zinc nitrate was reacted with 2-methylimidazole in the same molar ratio to obtain pure ZIF-8 powder.

[0067] Alendronate modification: Following the same steps as in Example 3, pure ZIF-8 was modified with alendronate to obtain the product of Comparative Example 3.

[0068] The core functionalities of the embodiments and comparative examples are compared in the table below: Table 1

[0069] Compared with comparative examples that were not modified with alendronate, were not encapsulated with TPP-carbon dots, or lacked TPP modification, this invention solves the core pain points of traditional nanomaterials, namely "low bone targeting precision, insufficient synergy between anti-oxidation and bone repair promotion, and high risk of accumulation in non-target organs," through the synergistic design of "alendronate bone-targeting modification + ZIF-8 carrier encapsulation + TPP-cerium-doped carbon dot functional core." It achieves a unified multi-functional approach of precise enrichment of bone lesions, efficient inflammation regulation, and bone repair promotion.

[0070] The examples demonstrate that, thanks to the specific chelation effect of alendronate with hydroxyapatite in bone tissue, the enrichment of bone lesions reached 92%-102%, while the unmodified alendronate in Comparative Example 1 was only 35%, and Comparative Examples 2 and 3, which lacked a functional core, were only 65%-70%. At the same time, the liver accumulation was only 32%-40% of that in Comparative Example 1, significantly reducing the burden on non-target organs, thus confirming that bone-targeted modification can improve lesion enrichment efficiency and biosafety.

[0071] The DPPH removal rate of the examples reached 84%-89%, which is much higher than that of Comparative Example 2 without TPP modification and Comparative Example 3 without encapsulated TPP-carbon dots. This is due to the Ce formed by cerium doping in the TPP-carbon dots. 3+ / Ce 4+ Valence cycling, combined with the mitochondrial targeting effect of TPP, can precisely inhibit oxidative stress and inflammatory responses; osteoblast ALP activity is significantly higher than that of the control group.

[0072] The core performance data of the examples fully demonstrate the synergistic value of "bone-targeted enrichment - mitochondrial inflammation regulation - stable and controllable release": alendronate ensures that the material accurately reaches the lesion, TPP-carbon dots provide efficient antioxidant and inflammation regulation functions, and the ZIF-8 carrier ensures the stability and controllable release of the functional components. The organic combination of these three factors enables the examples to comprehensively surpass the comparative examples in terms of targeting, antioxidant, and bone-repairing activity. Comparative example 1, lacking bone-targeted modification, showed a sharp drop in enrichment and a surge in non-target accumulation; comparative example 2, lacking TPP modification, showed a significant decrease in antioxidant activity; and comparative example 3, lacking encapsulation of the functional core, not only had no antioxidant effect but also significantly insufficient bone-repairing effect, all of which failed to meet the treatment needs of complex bone diseases.

[0073] In summary, the core performance data of the embodiments fully verify the beneficial effects of the present invention: the bone-targeting function improves the efficiency of lesion action, the antioxidant function improves the infection microenvironment, and the ZIF-8 carrier ensures functional stability. The three functions work synergistically to achieve the comprehensive advantages of "accurate targeting, strong antioxidant effect, excellent bone-promoting effect, and high safety", providing a comprehensive nanomaterial solution with outstanding clinical translation potential for the treatment of bone infection, bone tissue repair and other orthopedic diseases.

[0074] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. An alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterial, characterized in that, The alendronate-modified zeolite imidazole ester framework material-8 (ZIF-8) is used as a carrier, and TPP-modified carbon dots are encapsulated inside the carrier. The TPP-modified carbon dots are cerium-doped carbon quantum dots synthesized by hydrothermal method using citric acid, urea and cerium salt as raw materials. Then, TPP with mitochondrial targeting function is grafted onto the surface of the cerium-doped carbon quantum dots by amide method. The alendronate endows the nanomaterial with bone targeting function, and the TPP-modified carbon dots endow the nanomaterial with antioxidant and mitochondrial targeting functions.

2. The bone-targeting nanomaterial with alendronate-modified ZIF-8 encapsulated TPP-carbon dots according to claim 1, characterized in that, The nanomaterial has a particle size of 80-150 nm and, after being placed in a phosphate buffer solution at pH 7.4 for 72 h, has a dispersion index (PDI) ≤ 0.25, with no obvious agglomeration or precipitation; the cerium salt includes one or more of cerium nitrate, cerium chloride, cerium sulfate, cerium acetate, cerium iodide, and cerium bromide.

3. A method for preparing alendronic acid-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterial as described in claim 1, characterized in that, ZIF-8 was synthesized and TPP-modified carbon dots were simultaneously encapsulated. The resulting ZIF-8 encapsulated with TPP-modified carbon dots was mixed and stirred with sodium alendronate trihydrate to graft alendronate onto the surface of the ZIF-8 encapsulated with TPP-modified carbon dots, forming alendronate-modified ZIF-8 encapsulated with TPP-carbon dots. The preparation process of the TPP-modified carbon dots was as follows: cerium-doped carbon quantum dots were synthesized by hydrothermal method using citric acid, urea, and cerium salt as raw materials, and then TPP was grafted onto the surface of the cerium-doped carbon quantum dots by amide method.

4. The method for preparing alendronic acid-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials according to claim 3, characterized in that, ZIF-8 was synthesized by a solvothermal method. The reactants were zinc nitrate and 2-methylimidazole, the solvent was methanol, the molar ratio of zinc nitrate to 2-methylimidazole was 1:9-1:11, the reaction temperature was 25-30℃, and the reaction time was 10 minutes.

5. The method for preparing alendronic acid-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials according to claim 3, characterized in that, After synthesis, ZIF-8 was purified by centrifugation at 12000 r / min for 10 min, washing with methanol 3-4 times, and vacuum drying at -80℃ for 5-6 h.

6. The method for preparing alendronic acid-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials according to claim 3, characterized in that, The molar ratio of ZIF-8 to sodium alendronate trihydrate is 1:2-1:

4. Deionized water is used as the dispersant during mixing, and the mixture is stirred for 0.5 h at 25-28℃ and 250-350 r / min to achieve grafting.

7. The method for preparing alendronic acid-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials according to claim 3, characterized in that, Before encapsulation, TPP-modified carbon dots are dispersed by ultrasonic assistance and mixed with 2-methylimidazole until homogeneous. The ultrasonic power is 300-450W and the ultrasonic time is 10min. During the ultrasonic process, the system temperature is maintained at 28-32℃ by water bath temperature control.

8. The method for preparing alendronic acid-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterials according to claim 3, characterized in that, After modification, the product is centrifuged at 12000 r / min for 10 min, washed three times with deionized water, and then freeze-dried at -80℃ for 5-6 h.

9. The application of an alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterial as described in claim 1 in bone tissue repair, treatment of bone infection, or targeted therapy for bone-related diseases.

10. The application of the alendronate-modified ZIF-8 encapsulated TPP-carbon dot bone-targeting nanomaterial according to claim 9 in bone tissue repair, bone infection treatment, or targeted therapy for bone-related diseases, characterized in that, In the treatment of bone infections, the nanomaterials can be targeted and enriched at the site of bone infection, and inhibit the inflammatory response through the antioxidant function of TPP-modified carbon dots, thereby helping to improve the infection microenvironment.

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