A bifunctional core-shell microsphere and a preparation method thereof

Abstract

The application discloses a kind of bifunctional core-shell microspheres and its preparation method, the core of this microsphere is its "core-shell partition, function synergy" structure: with biodegradable polymer load osteogenic active factor as core, realize bone conduction and bone induction;With the biohydrogel of metal organic framework synthesized in situ by metal ion crosslinking as shell, provide instant and long-acting antibacterial capacity;The structure is formed in one step by coaxial microfluidic technology, and the function of shell layer is strengthened by combining in-situ synthesis technology after processing, finally obtain uniform size, stable structure, controllable performance core-shell microspheres.

Description

Technical Field

[0001] This invention belongs to the field of preparation and application technology of biomedical materials—microsphere materials. It designs and constructs a core-shell structure with the characteristics of "functional spatial isolation" and "temporal synergistic control", and realizes the structure through a sophisticated microfluidic synergistic assembly and in-situ synthesis technology. Specifically, it relates to a bifunctional core-shell microsphere and its preparation method, which is particularly suitable for the repair and treatment of infectious bone defects. Background Technology

[0002] Infected bone defects pose a significant challenge to clinical orthopedics, commonly seen in open fractures, postoperative debridement for osteomyelitis, and postoperative infections following total joint replacement. Repairing and reconstructing these defects requires addressing two core issues simultaneously: effectively preventing and controlling bacterial infection at the implantation site, and promoting the regeneration and reconstruction of new bone tissue. Ideal bone repair materials should possess both long-lasting antibacterial activity and highly efficient osteogenic induction capabilities, with these two aspects working synergistically and without interference.

[0003] To address these issues, researchers have developed various functional microsphere materials. For example, patent CN104096263A discloses a submicron core-shell microsphere material that instantly releases active factors. Its core layer is poly(racemic lactate) loaded with bone morphogenetic protein-2 (BMP-2), and its shell layer is poly(lactide-glycolic acid) loaded with vascular endothelial growth factor (VEGF). This core-shell structure enables the sequential release of osteogenic and angiogenic factors. However, this material lacks antibacterial function and cannot address bacterial infection in infectious bone defects. Furthermore, its functional factors are all protein growth factors, which suffer from poor stability, easy inactivation, and high cost.

[0004] To address the need for combined antibacterial and osteogenic functions, patent CN115581799A discloses a composite microsphere. This microsphere uses a mesoporous material (such as mesoporous calcium silicate) as a carrier, loading silver ions (Ag⁺) and tissue regeneration inducing factors (such as BMP-2) into the pores. A water-soluble polymer layer and a biodegradable polymer layer are sequentially coated onto the surface of the mesoporous material. This technology protects the active factors through the mesoporous material and achieves controlled release through multilayer coating. However, this approach has the following drawbacks: First, although silver ions have broad-spectrum antibacterial properties, their release behavior is difficult to control precisely, and they are prone to initial burst release, leading to local cytotoxicity (Comparative Example 1 of this patent shows that silver ions are released too quickly without mesoporous material loading, resulting in cytotoxicity); Second, silver ions and tissue regeneration inducing factors are co-loaded in the same mesoporous channel, and their release behaviors are similar, making it impossible to achieve temporal synergy, and high concentrations of silver ions may have an adverse effect on osteoblast activity; Third, the shell is only a physical barrier and does not have active antibacterial function, and the antibacterial performance depends entirely on the silver ions in the nucleus.

[0005] Furthermore, patent CN116251229B discloses a composite gel microsphere, which disperses magnesium silicate nanoparticles loaded with benidipine in a methacrylamide gelatin (GelMA) / sodium alginate hydrogel, and prepares the microspheres by photopolymerization 3D printing. This material utilizes the magnesium silicate nanoparticles to release magnesium ions to promote osteoogenesis and loads drugs to enhance biological effects. However, this technology also suffers from the problem of lacking antibacterial function; at the same time, its nanoparticles are directly dispersed in the gel matrix without forming a core-shell structure, making it difficult to independently regulate the release behavior of functional factors; and the mechanical strength of the gel matrix itself is limited, and the matching between the degradation rate and the osteogenic process needs to be improved.

[0006] In summary, existing technologies still have the following common limitations in addressing the repair of infected bone defects:

[0007] 1. Interference between functional factors: When antibacterial components and osteogenic factors are co-loaded in the same space (such as mesoporous channels or polymer matrix), their release behaviors are similar and cannot achieve temporal synergy; high-concentration release of antibacterial components may be toxic to osteoblasts and inhibit bone regeneration.

[0008] 2. Uncontrollable antibacterial function: Existing materials mostly rely on traditional antibacterial agents such as antibiotics or silver ions, which pose risks of drug resistance, burst release effects, and cytotoxicity; the shell design is mostly a physical barrier and does not have active antibacterial function, and the antibacterial performance depends entirely on the antibacterial agent loaded in the core.

[0009] 3. Poor stability of osteogenic factors: Protein growth factors (such as BMP-2 and VEGF) are expensive, easily inactivated, and have short in vivo half-lives, making it difficult to meet the economic and stability requirements for clinical translation.

[0010] 4. The preparation process is complex and the structure is difficult to control: Traditional emulsification methods are difficult to prepare core-shell microspheres with uniform size; physical blending methods are prone to uneven distribution and shedding of functional factors; post-synthesis modification methods are cumbersome and have harsh conditions, which can easily damage the bioactivity of materials.

[0011] 5. Insufficient microenvironment regulation capability: Existing materials do not fully consider the impact of local microenvironment changes after implantation (such as acid production from PLGA degradation and alkali production from MgO hydrolysis) on material stability and cell activity, making it difficult to achieve long-term stable function.

[0012] Therefore, there is an urgent need in this field to develop a biodegradable microsphere material that combines osteogenic and antibacterial functions, has spatially isolated functional factors with independently controllable release behavior, a simple preparation process, and a uniform and stable structure, in order to solve the clinical challenges in the repair of infected bone defects.

[0013] The information disclosed in this background section is intended only to enhance the understanding of the overall background of the invention and should not be construed as an admission or in any way implying that the information constitutes prior art known to those skilled in the art. Summary of the Invention

[0014] To address the shortcomings of existing technologies, this invention provides a bifunctional core-shell microsphere and its preparation method.

[0015] To achieve the above objectives, the present invention adopts the following technical solution:

[0016] A bifunctional core-shell microsphere, the microsphere comprising the following:

[0017] The core layer is composed of a biodegradable polymer material and magnesium oxide (MgO) nanoparticles dispersed therein;

[0018] The shell is composed of a hydrogel matrix, which is cross-linked with metal ions and in situ generates metal-organic framework (MOF) crystals in its network.

[0019] The mass fraction of MgO nanoparticles in the core layer is 1% to 10%.

[0020] Preferably, the hydrogel matrix is ​​sodium alginate.

[0021] Preferably, the metal ion is copper ion Cu. 2+ .

[0022] Preferably, the biodegradable polymer material is polylactic acid-glycolic acid copolymer (PLGA).

[0023] Preferably, the metal-organic framework crystal is HKUST-1.

[0024] Preferably, the particle size of the microspheres is 100–500 μm.

[0025] Unlike existing technologies that simply physically mix osteogenic and antibacterial components (functional hybridization), this invention strictly confines the osteogenic function (PLGA+MgO) to the core and precisely constructs the potent, contact antibacterial function (SA-Cu-HKUST-1) in the shell, thus achieving physical spatial isolation of the functions and avoiding adverse interference between components.

[0026] Unlike inert shells that merely act as physical diffusion barriers, the shell of this invention is itself an active and long-lasting antibacterial functional body. The in-situ formation of HKUST-1 allows it to tightly bind with the alginate network, realizing the antibacterial component (Cu... 2+ The stable, high-load, and slow, continuous release of the shell overcomes the burst release problem caused by simple mixing or physical adsorption.

[0027] This invention also provides a method for preparing bifunctional core-shell microspheres, comprising the following steps:

[0028] 1) Preparation of internal and external phase fluids: The internal phase fluid contains biodegradable polymer materials and MgO nanoparticles dispersed therein; the external phase fluid is an aqueous solution of a hydrogel matrix;

[0029] 2) Using a coaxial microfluidic device, the inner and outer phase fluids are extruded together to form core-shell structured droplets;

[0030] 3) The droplets are collected in a cross-linked solution containing metal ions, causing the shell hydrogel matrix to undergo ionic cross-linking and solidification to obtain solidified microspheres;

[0031] 4) The solidified microspheres are brought into contact with an organic ligand solution, so that the metal ions in the shell react with the organic ligand in situ to generate metal-organic framework crystals, thus obtaining bifunctional core-shell microspheres.

[0032] Preferably, the metal ion in step 3) is a copper ion, and the crosslinking solution is a copper salt solution; the organic ligand in step 4) is pyromellitic acid (BTC).

[0033] Preferably, the in-situ reaction time in step 4) is 1 to 24 hours, and the reaction temperature is 20 to 40°C.

[0034] By adopting the above technical solution, the method divides the formation of the core-shell structure and the construction of the functional shell into two closely connected but independent stages: the first is the "physical forming" stage, which uses microfluidic technology to precisely control the size and structure of microspheres, and the second is the "chemical synthesis" stage, which drives the functionalization of the shell.

[0035] Unlike the traditional method of blending pre-synthesized HKUST-1 powder with polymer materials, the "in-situ generation" technology of this invention ensures that HKUST-1 crystals are uniformly and stably anchored in the alginate gel network, forming an organic-inorganic composite reinforcing shell, which effectively prevents HKUST-1 from falling off and agglomerating, and optimizes its release behavior.

[0036] The third invention provides the use of bifunctional core-shell microspheres in the preparation of medical materials or drug delivery systems for the repair of infectious bone defects and with antibacterial function.

[0037] This invention provides a biodegradable antibacterial-osteogenic bifunctional core-shell microsphere for bone tissue engineering. Through precise core-shell structure design and ingenious in-situ synthesis process, it achieves precise spatial isolation and temporal synergy between osteogenic and antibacterial functions. This fundamentally solves the prominent problems in existing technologies, such as mutual interference of functional factors, uncontrollable release behavior, complex preparation process, and poor structural stability. It has the following beneficial effects:

[0038] 1. This invention strictly confines the osteogenic functional component to the core layer, which is composed of biodegradable polymer PLGA and uniformly dispersed MgO nanoparticles, while precisely constructing the antibacterial functional component in the shell layer, which is composed of a metal ion cross-linked hydrogel network and metal-organic framework crystals generated in situ within it. This functional spatial isolation design avoids direct contact and mutual interference between the antibacterial components and osteogenic factors, allowing the copper ions released from the shell layer to first form a highly efficient antibacterial microenvironment outside the microspheres, clearing bacteria around the implantation area and creating a clean and suitable tissue regeneration space for the magnesium ions subsequently released from the core layer. Both functions perform their respective roles without interference, truly achieving a synergistic effect of antibacterial and osteogenic functions.

[0039] 2. Regarding release behavior, this invention significantly improves the burst release effect commonly found in traditional materials through a multi-layered controlled-release design of the shell. The hydrogel network formed by metal ion crosslinking in the shell itself acts as the first diffusion barrier, while the in-situ generated metal-organic framework crystals are uniformly anchored within this network. The release of antibacterial copper ions is kinetically controlled by crystal dissolution and coordination bond breaking, rather than simple free diffusion, thus achieving a stable and long-lasting release of copper ions. This ensures a sustained antibacterial concentration while effectively reducing local cytotoxicity that may result from excessively high initial concentrations. Simultaneously, the degradation of the core layer PLGA is a slow bulk erosion process. Combined with the shell acting as an additional physical barrier, this makes the release of magnesium ions more gentle and prolonged, matching the long-term process of bone tissue regeneration. This creates an ideal sequential effect of "antibacterial first, then osteogenic, with long-lasting synergy," perfectly meeting the clinical needs of repairing infected bone defects.

[0040] 3. Regarding structural stability and mechanical properties, the shell of this invention is not a single polymer network, but an organic-inorganic composite structure formed through a dual mechanism of ionic crosslinking and coordination bonding. The in-situ generation of metal-organic framework crystals within the alginate gel network allows them to act as a nano-reinforcing phase, tightly bonding with the gel matrix and significantly enhancing the shell's mechanical strength, toughness, and resistance to degradation. This reinforced composite shell enables the microspheres to maintain structural integrity over long periods in the complex mechanical environments of implantation and vivo, preventing breakage or collapse, thus ensuring the durability and reliability of its function.

[0041] 4. This invention also demonstrates significant advantages in terms of biocompatibility and microenvironment regulation. The acidic products generated by PLGA degradation in the core layer and the process of MgO degradation consuming hydrogen ions mutually restrain each other, forming a local acid-base buffer system. This effectively reduces the aseptic inflammatory response that may be caused by pure PLGA materials, providing a more suitable microenvironment for cell adhesion, proliferation, and differentiation. At the same time, the high concentration of copper ions with potential cytotoxicity is strictly confined to the shell layer and released slowly, maintaining an effective antibacterial concentration locally in the microspheres while minimizing the risk of toxicity to surrounding normal tissue cells, thus cleverly balancing the contradiction between antibacterial efficacy and biosafety.

[0042] 5. Furthermore, the coaxial microfluidic preparation technology employed in this invention enables precise control over the microsphere size and core-shell structure, resulting in microspheres with uniform particle size, good monodispersity, and high batch stability. Uniform size and structure mean that each microsphere exhibits consistent drug loading and release behavior, which not only ensures the reliability and reproducibility of in vitro and in vivo experimental results but also lays a solid quality foundation for subsequent clinical translation and industrial production.

[0043] In summary, this invention is not a simple combination or improvement of existing technologies, but rather achieves functional integration and optimization at multiple levels—molecular, microscopic, and macroscopic—through systematic structural design and process innovation. The resulting synergistic effects, controllable release behavior, enhanced structural stability, high biosafety, and controllable preparation quality demonstrate its significant clinical application potential in the repair of infected bone defects, surpassing existing technologies. Attached Figure Description

[0044] Figure 1 This is a flowchart of the preparation process;

[0045] Figure 2 SEM images of microspheres at different magnifications;

[0046] Figure 3 Characterization diagram of EDS elemental distribution in microspheres;

[0047] Figure 4 This is the EDS full spectrum analysis diagram of the microspheres;

[0048] Figure 5 This is a plate colony count diagram showing the antibacterial properties of microspheres. Detailed Implementation

[0049] The present invention will be further described below through specific embodiments. To make the inventive objectives, technical solutions, and beneficial technical effects of the present invention clearer, the present invention will be further described in detail below with reference to the embodiments. It should be understood that the embodiments described in this specification are merely for explaining the present invention and are not intended to limit the present invention.

[0050] Unless otherwise stated, all instruments and reagents used in the examples are commercially available or synthesized using conventional methods and can be used directly without further processing, and all instruments used in the examples are commercially available.

[0051] Example 1: Preparation as microsphere material

[0052] Core-shell microspheres were prepared using the following steps:

[0053] 1) Preparation of internal and external phase fluids: The internal phase fluid comprises polylactic acid-glycolic acid copolymer (PLGA) and MgO nanoparticles dispersed therein (MgO nanoparticles have a mass fraction of 5%); the external phase fluid is an aqueous solution of sodium alginate matrix;

[0054] 2) Using a coaxial microfluidic device, the inner and outer phase fluids are extruded together to form core-shell structured droplets;

[0055] 3) The droplets are collected in a cross-linking solution containing copper ions, causing the shell hydrogel matrix to undergo ionic cross-linking and solidification to obtain solidified microspheres;

[0056] 4) The solidified microspheres are brought into contact with BTC solution, and the copper ions in the shell react with BTC in situ for 12 hours at a reaction temperature of 30°C to generate HKUST-1, thus obtaining bifunctional core-shell microspheres (microspheres with a particle size of 300 μm).

[0057] Example 2:

[0058] Core-shell microspheres were prepared using the following steps:

[0059] 1) Preparation of internal and external phase fluids: The internal phase fluid comprises polylactic acid-glycolic acid copolymer (PLGA) and MgO nanoparticles dispersed therein (MgO nanoparticles have a mass fraction of 10%); the external phase fluid is an aqueous solution of sodium alginate matrix;

[0060] 2) Using a coaxial microfluidic device, the inner and outer phase fluids are extruded together to form core-shell structured droplets;

[0061] 3) The droplets are collected in a cross-linking solution containing copper ions, causing the shell hydrogel matrix to undergo ionic cross-linking and solidification to obtain solidified microspheres;

[0062] 4) The solidified microspheres are brought into contact with a BTC solution, and the copper ions in the shell react with BTC in situ for 5 hours at a reaction temperature of 40°C to generate HKUST-1, thus obtaining bifunctional core-shell microspheres (microspheres with a particle size of 500 μm).

[0063] Example 3:

[0064] Core-shell microspheres were prepared using the following steps:

[0065] 1) Preparation of internal and external phase fluids: The internal phase fluid comprises polylactic acid-glycolic acid copolymer (PLGA) and MgO nanoparticles dispersed therein (MgO nanoparticles have a mass fraction of 1%); the external phase fluid is an aqueous solution of sodium alginate matrix;

[0066] 2) Using a coaxial microfluidic device, the inner and outer phase fluids are extruded together to form core-shell structured droplets;

[0067] 3) The droplets are collected in a cross-linking solution containing copper ions, causing the shell hydrogel matrix to undergo ionic cross-linking and solidification to obtain solidified microspheres;

[0068] 4) The solidified microspheres are brought into contact with a BTC solution, and the copper ions in the shell react with BTC in situ for 24 hours at a reaction temperature of 20°C to generate HKUST-1, thus obtaining bifunctional core-shell microspheres (microspheres with a particle size of 100 μm).

[0069] Example 4: The microspheres from Example 1 were selected as the experimental subject for antibacterial testing.

[0070] 1. Experimental strain

[0071] Escherichia coli (ATCC 25922).

[0072] 2. Experimental Grouping

[0073] a. Blank control group: Original bacterial solution, without any antibacterial materials;

[0074] b. Negative control: Pure PLGA / MgO blank microspheres;

[0075] c. Positive control: PLGA / MgO@Cu-Alg core-shell microspheres;

[0076] d. Experimental group: PLGA / MgO@Cu-MOF-Alg core-shell microspheres.

[0077] 1.2.3 represent serial dilutions of 10⁻¹, 10⁻², and 10⁻³, respectively.

[0078] 3. Experimental Methods

[0079] Each group of materials was mixed with 1×10⁴ CFU / mL bacterial suspension at a ratio of 1:9 (v / v) and co-cultured at 37℃ and 150 rpm for 12 h. Subsequently, the culture was serially diluted by 10⁻¹, 10⁻², and 10⁻³, and 10 μL of each was spread onto LB agar plates and incubated at 37℃ for 16 h before counting the colonies.

[0080] 4. Evaluation Indicators

[0081] The antibacterial rate (R, %) is calculated according to formula (1): R = (N0-N) / N0×100%

[0082] Where N0 is the average colony count (CFU) of the blank control group, and N is the average colony count (CFU) of the experimental group.

[0083] 5. Experimental Results

[0084] Compared with the blank control group, at a dilution of 10⁻², the antibacterial rate of the negative control was 10%, the antibacterial rate of the positive control was 100%, and the antibacterial rate of the experimental group was 97.5%. At a dilution of 10⁻³, the antibacterial rate of the positive control group was 100%, the antibacterial rate of the experimental group was 100%, and the negative control group had no significant antibacterial activity (P > 0.05).

[0085] 6. Conclusion

[0086] PLGA / MgO@Cu-MOF crosslinked sodium alginate hydrogel core-shell microspheres exhibited rapid and efficient antibacterial properties against Escherichia coli within a 12-hour contact time, meeting the core technical requirements for antibacterial medical devices or dressings.

[0087] Example 5: Microstructure and elemental composition characterization of bifunctional core-shell microspheres (SEM-EDS full spectrum analysis)

[0088] 1. Test materials

[0089] The PLGA / MgO@Cu-MOF-Alg bifunctional core-shell microspheres (particle size 300 μm, MgO mass fraction 5%) prepared in Example 1 were selected as the test sample; the negative control was pure PLGA / MgO blank microspheres.

[0090] 2. Test instruments

[0091] Field emission scanning electron microscope (SU8600, equipped with EDS energy dispersive spectroscopy module); vacuum freeze dryer; ion sputtering instrument (for gold sputtering).

[0092] 3. Experimental Procedure

[0093] S1. Sample pretreatment: The microspheres to be tested were placed in a vacuum freeze dryer and dried for 24 hours to remove moisture from the sample and avoid morphological collapse; the dried microspheres were then evenly spread on conductive adhesive and sputtered with gold for 60 seconds to enhance the conductivity of the sample.

[0094] S2. SEM Microscopic Morphology Observation: The pretreated sample was placed on the SU8600 scanning electron microscope sample stage, and the accelerating voltage was set to 3.00kV. The overall morphology and shell surface details of the microspheres were observed at magnifications of ×100, ×4.00k, and ×20.0k, respectively. Secondary electron morphology images were acquired, and the particle size, sphericity, shell crystal distribution, and structural integrity of the microspheres were recorded.

[0095] S3. EDS full spectrum and elemental quantitative analysis: Based on SEM observation, a single microsphere with complete morphology was selected as the test area. The EDS energy spectrum analysis module was turned on to perform a full spectrum scan (energy range 0~18keV), and the elemental composition full spectrum of the microsphere was collected. The detected elements were quantitatively analyzed by weight percentage (Wt%). The test was repeated 3 times and the average value was taken.

[0096] 4. Test Results

[0097] 1) SEM morphology results: Under a low magnification of ×100 (scale bar 500 μm), the microspheres are regular spherical with uniform particle size (average 300 μm), without adhesion or breakage, and have good monodispersity; Under a medium magnification of ×4.00k (scale bar 10.0 μm), triangular / plate-like crystals are uniformly distributed on the surface of the microsphere shell, which are HKUST-1 crystals generated in situ, and the crystals are tightly bonded to the sodium alginate gel matrix; Under a high magnification of ×20.0k (scale bar 2.00 μm), the HKUST-1 crystals have clear facets, complete structure, no dissolution or detachment, the microsphere shell is dense, and the core-shell bonding is crack-free.

[0098] 2) EDS full spectrum and quantitative results: The characteristic energy peaks of C, O, Mg and Cu were clearly visible in the EDS full spectrum of the microspheres. Trace amounts of Ca (as environmental impurities during reagent or sample preparation) were detected, and no other impurity peaks were observed. The elemental quantitative analysis results were consistent with those in Table 1 of Example 5, namely C: 61.51±0.19%, O: 29.16±0.19%, Mg: 0.05±0.02%, Ca: 0.12±0.01%, Cu: 9.17±0.09%, with a total percentage of 100%, which is completely consistent with the material composition design of the microspheres "PLGA / MgO core + sodium alginate-Cu-HKUST-1 shell".

[0099] Table 1 is as follows:

[0100]

[0101] Example 6: Elemental spatial distribution characterization of bifunctional core-shell microspheres (EDS surface scan mapping analysis) 1. The experimental materials were the same as in Example 5, namely PLGA / MgO@Cu-MOF-Al bifunctional core-shell microspheres prepared in Example 1.

[0102] 2. Test instruments

[0103] Same as Example 6, SU8600 field emission scanning electron microscope (equipped with EDS surface scanning module).

[0104] 3. Experimental Procedure

[0105] Sample pretreatment: Same as step 1) in Example 6, the microspheres were freeze-dried, sputter-coated with gold and then fixed on conductive adhesive.

[0106] EDS elemental surface scan test: The sample was placed on the SEM sample stage, and a single microsphere with a complete morphology was selected as the surface scan area (scanning area 5μm×5μm). The accelerating voltage was set to 3.00kV, and the EDS surface scan module was turned on. Characteristic X-ray signals of C(Kα1,2), O(Kα1), Cu(Kα1), Mg(Kα1,2), and Ca(Kα1) were collected respectively. Surface scan distribution maps (mapping maps) of each element were generated, and the maps of each element were superimposed with the secondary electron morphology map of the microsphere to obtain the EDS elemental layered superimposed map.

[0107] 4. Experimental results: C and O element distribution: The surface scan spectra of C and O elements show that they are continuously and uniformly distributed throughout the entire microsphere region, and completely overlap with the secondary electron morphology of the microsphere. This proves that C and O, as the core elements of the organic matrix (PLGA, sodium alginate, HKUST-1 ligand) and inorganic components (MgO, HKUST-1 coordinated oxygen), constitute the overall framework of the microsphere, and the framework structure is complete and continuous.

[0108] Cu element distribution: The surface scan spectrum of Cu element shows that the Cu signal is concentrated only in the surface shell region of the microsphere, which corresponds perfectly to the distribution position of the HKUST-1 crystal observed by SEM. There is no obvious Cu signal in the core region, which proves that the Cu element is strictly confined to the shell and is the core component of the HKUST-1 crystal.

[0109] Mg element distribution: The surface scan spectrum of Mg element shows that the Mg signal is concentrated only in the core layer region of the microsphere, and there is no Mg signal in the shell region. The Mg signal in the core layer is uniformly distributed, which proves that the MgO nanoparticles are successfully confined in the core layer of PLGA and do not diffuse into the shell layer.

[0110] Ca element distribution: The surface scan spectrum of Ca element shows that the Ca signal is distributed in trace amounts and discretely on the surface of the microspheres, without obvious concentrated areas, proving that Ca is an undesigned impurity and its content is extremely low, which does not affect the structure and function of the microspheres.

[0111] Elemental layer overlay diagram: The overlay results of the elemental spectra and secondary electron morphology diagrams show that the core-shell structure of the microspheres has clear boundaries, and the Cu (shell) and Mg (core) elements are completely spatially isolated without any interpenetration.

[0112] 5. Experimental Conclusions

[0113] The bifunctional core-shell microspheres prepared by this invention achieve precise spatial isolation of functional elements, with the antibacterial element Cu concentrated in the shell layer and the osteogenic element Mg concentrated in the core layer, which is in perfect agreement with the structural design of "core-shell partitioning and functional synergy", effectively avoiding mutual interference between antibacterial and osteogenic components.

[0114] Example 7: Structural stability verification test of bifunctional core-shell microspheres

[0115] 1. Test materials

[0116] Bifunctional core-shell microspheres prepared in Examples 1-3 (particle sizes of 300 μm, 500 μm, and 100 μm, respectively); the control group consisted of pure calcium alginate gel microspheres (synthesized in situ without HKUST-1).

[0117] 2. Test instruments

[0118] Constant temperature shaker; phosphate buffered saline (PBS, pH=7.4); field emission scanning electron microscope (SU8600); electronic balance.

[0119] 3. Experimental Procedure

[0120] In vitro degradation treatment: Microspheres of each experimental group and control group were placed in centrifuge tubes containing PBS buffer at a solid-liquid ratio of 1:100 (g / mL) and placed in a constant temperature shaker at 37℃ and 150rpm for in vitro degradation. Samples were taken at 7d, 14d and 28d.

[0121] Structural integrity observation: After sampling, the microspheres were washed with deionized water, freeze-dried, and then characterized by SEM (test parameters are the same as in Example 6). The sphericity, shell integrity, and whether cracking / breakage occurred were observed.

[0122] Mass retention rate determination: Weigh the dry weight of the microspheres before degradation and at each time point after degradation, and calculate the mass retention rate according to the formula: Mass retention rate (%) = (dry weight after degradation / dry weight before degradation) × 100%. Repeat each group 3 times and take the average value.

[0123] 4. Test Results

[0124] Structural integrity: After 28 days of degradation, the bifunctional core-shell microspheres of Examples 1-3 still maintained a regular spherical shape, with a dense shell without cracks, and the HKUST-1 crystals were tightly bonded to the sodium alginate gel matrix without falling off; the pure calcium alginate gel microspheres in the control group showed shell swelling after 7 days of degradation, shell cracking after 14 days, and spherical collapse after 28 days, with complete structural damage.

[0125] Mass retention rate: The mass retention rates of the microspheres in Examples 1-3 after 28 days of degradation were 78.2%, 75.6%, and 80.1%, respectively, all above 75%; while the mass retention rate of the pure calcium alginate gel microspheres in the control group after 28 days of degradation was only 32.5%.

[0126] 5. Experimental Conclusions

[0127] The bifunctional core-shell microspheres of this invention have an organic-inorganic composite shell formed by “Cu² + cross-linked sodium alginate + HKUST-1 in situ generation”, which significantly improves the mechanical strength and anti-degradation ability of the microspheres. They have excellent structural stability under physiological conditions, which can ensure that the structure remains intact for a long time after implantation in the body and achieve sustained function.

[0128] The above description of the disclosed embodiments enables those skilled in the art to make or use the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A bifunctional core-shell microsphere, characterized in that, The microspheres include the following: The core layer is composed of a biodegradable polymer material and magnesium oxide (MgO) nanoparticles dispersed therein; The shell is composed of a hydrogel matrix, which is cross-linked with metal ions and in situ generates metal-organic framework (MOF) crystals in its network. The mass fraction of MgO nanoparticles in the core layer is 1% to 10%.

2. The bifunctional core-shell microsphere according to claim 1, characterized in that, The hydrogel matrix is ​​sodium alginate.

3. The bifunctional core-shell microsphere according to claim 1, characterized in that, The metal ion is copper ion Cu. 2 + .

4. The bifunctional core-shell microsphere according to claim 1, characterized in that, The biodegradable polymer material is polylactic acid-glycolic acid copolymer (PLGA).

5. A bifunctional core-shell microsphere according to claim 1, characterized in that, The metal-organic framework crystal is HKUST-1.

6. A bifunctional core-shell microsphere according to claim 1, characterized in that, The microspheres have a particle size of 100–500 μm.

7. A method for preparing bifunctional core-shell microspheres according to any one of claims 1-6, characterized in that, Includes the following steps: 1) Preparation of internal and external phase fluids: The internal phase fluid contains biodegradable polymer materials and MgO nanoparticles dispersed therein; the external phase fluid is an aqueous solution of a hydrogel matrix; 2) Using a coaxial microfluidic device, the inner and outer phase fluids are extruded together to form core-shell structured droplets; 3) The droplets are collected in a cross-linked solution containing metal ions, causing the shell hydrogel matrix to undergo ionic cross-linking and solidification to obtain solidified microspheres; 4) The solidified microspheres are brought into contact with an organic ligand solution, so that the metal ions in the shell react with the organic ligand in situ to generate metal-organic framework crystals, thus obtaining bifunctional core-shell microspheres.

8. The method for preparing bifunctional core-shell microspheres according to claim 7, characterized in that, The metal ion mentioned in step 3) is a copper ion, and the crosslinking solution is a copper salt solution; the organic ligand mentioned in step 4) is pyromellitic acid (BTC).

9. The method for preparing bifunctional core-shell microspheres according to claim 7, characterized in that, The in-situ reaction time in step 4) is 1 to 24 hours, and the reaction temperature is 20 to 40°C.

10. Use of a bifunctional core-shell microsphere in the preparation of medical materials or drug delivery systems for the repair of infected bone defects and with antibacterial function.

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