A gradient pore-forming calcium-magnesium phosphate bone repair material and a preparation method thereof

Abstract

The application discloses a preparation method of gradient pore containing magnesium calcium phosphate bone repair material, which comprises the following steps: S1, mixing nano-hydroxyapatite and polylactic acid-glycolic acid copolymer PLGA powder, adding dichloromethane as a solvent, continuously stirring to form a solution, and obtaining nHA / PLGA raw materials through freeze drying, and then preparing a degradable nPLGA support through a 3D biological printer high-temperature melting deposition; S2, taking calcium phosphate bone cement powder and magnesium metal microspheres according to a preset mass ratio, stirring the slurry to be homogeneous, and obtaining calcium phosphate bone cement slurry containing magnesium metal microspheres; S3, injecting the calcium phosphate bone cement slurry obtained in the step S2 into the nPLGA support prepared in the step S1 to obtain an MC / nPLGA compound; through the hydration reaction of the magnesium microspheres, gradient micropores are generated, and the macropores formed by the degradation of the PLGA form a complementary structure, and the total porosity reaches 68%, which is 3.4 times higher than that of traditional CPC.

Description

TECHNICAL FIELD

[0001] The application relates to a kind of alveolar bone repair material technology, in particular to a kind of gradient pore containing magnesium calcium phosphate bone repair material and a preparation method thereof. BACKGROUND

[0002] In the field of oral implantation, the low success rate of implant denture repair caused by severe bone defect has become a key bottleneck restricting the long-term stability of patient's oral function reconstruction and implant. Guided bone regeneration technology (GBR) isolates soft tissue invasion by constructing a biological barrier membrane to maintain the bone regeneration microenvironment, which is the mainstream operation for solving bone deficiency in clinical practice. However, the traditional GBR technology still faces many clinical challenges in the scheme of absorbable collagen membrane combined with bone grafting material: autologous bone, although it has irreplaceable osteoinductive activity and osteogenic potential, is regarded as the "gold standard" for bone defect repair, but its application needs to take bone from the patient's bone area, which has the risk of infection of the donor area, postoperative pain and limited bone volume; although the heterologous bone material avoids the complications of autologous bone harvesting, it still has the risk of immunogenicity, and the slow degradation and mismatched bone regeneration rate affect the repair of new bone.

[0003] To break through the inherent limitations of natural bone grafting material, artificial bone repair material has become a research focus in the field of biomaterials due to its designable physicochemical properties, potential for large-scale preparation and biological safety. Among them, calcium phosphate cement (CPC) is considered as a bone substitute material with great clinical conversion potential due to its high similarity to the chemical composition of natural bone mineral, injectable molding characteristics and good biocompatibility.

[0004] However, the traditional CPC has the following problems: the dense structure leads to insufficient porosity, lack of three-dimensional interconnected pore network, hinders the early vascularization process, and the degradation rate does not match the bone regeneration rate; lack of endogenous bioactive factors. These defects directly lead to the fact that traditional CPC can only play a "space filling" role in large bone defect repair, and cannot achieve active bone regeneration, and its clinical application is still limited to non-weight-bearing bone defect repair. SUMMARY

[0005] The application overcomes the shortcomings of the prior art and provides a gradient pore containing magnesium calcium phosphate bone repair material and a preparation method thereof.

[0006] To achieve the above purpose, the technical scheme adopted by the application is as follows: a preparation method of a gradient pore containing magnesium calcium phosphate bone repair material, comprising the following steps:

[0007] S1, mixing nano-hydroxyapatite and polylactic acid-glycolic acid copolymer PLGA powder according to a mass ratio of 5:95 to 25:75, after ultrasonic dispersion, adding dichloromethane as a solvent in a volume ratio of 1:3 to 1:5, continuously stirring for 5-8 hours to form a solution, and obtaining nHA / PLGA raw materials by freeze-drying, and then preparing a degradable nPLGA scaffold with a three-dimensional interconnected macroporous structure by high-temperature fused deposition of a 3D bioprinter;

[0008] S2, taking calcium phosphate cement powder and magnesium metal microspheres according to a preset mass ratio, adding a solidification liquid in a liquid-solid ratio of 0.3-0.5 mL / g, stirring the slurry to be homogeneous, and obtaining a calcium phosphate cement slurry containing magnesium metal microspheres;

[0009] S3, injecting the calcium phosphate cement slurry obtained in step S2 into the nPLGA scaffold prepared in step S1, and curing for 20-30 hours under the conditions of a constant temperature of 35-37 DEG C and a humidity of 97-100%, and then demolding to obtain an MC / nPLGA composite.

[0010] In a preferred embodiment of the present application, the magnesium metal microspheres are dispersed in the calcium phosphate cement powder, hydrogen gas is generated by a hydration reaction to form a microporous structure, the micropore size ranges from 30 to 500 microns, and the bioactive substance Mg2 + .

[0011] In a preferred embodiment of the present application, the Mg2+ in the material has the effects of promoting bone formation and angiogenesis, the alkaline environment formed by magnesium is alleviated as the nPLGA scaffold degrades to produce acid, and the degradation rate of the calcium phosphate cement is accelerated by the acid produced by the degradation of the nPLGA scaffold.

[0012] In a preferred embodiment of the present application, the freeze-drying process is divided into phase one and phase two; wherein phase one includes a preset freezing temperature of-70 DEG C to-90 DEG C and a pre-freezing time of 3-5 hours; and phase two includes a preset drying temperature of-40 DEG C to-60 DEG C, a vacuum degree of 0.05-0.2 Mbar, and a drying time of 48-72 hours.

[0013] In a preferred embodiment of the present application, when the 3D bioprinter is implemented for high-temperature fused deposition manufacturing, the melting temperature is between 150 DEG C and 170 DEG C, the extrusion pressure is between 1.5 bar and 4 bar, the printing speed is between 5 mm / s and 10 mm / s, 70%-90% of the nozzle diameter is used as the deposition layer height parameter, the fiber spacing is between 1 mm and 3 mm, and the orthogonal stacking structure is 0 DEG / 90 DEG / 90 DEG to 0 DEG / 60 DEG / 60 DEG. The 3D printing technology supports personalized scaffold customization and is suitable for personalized repair treatment of complex alveolar bone defects in the maxillofacial region.

[0014] In a preferred embodiment of the present application, the calcium-phosphorus ratio of the calcium phosphate cement powder ranges from 1.50 to 1.70; and the particle size of the magnesium metal microspheres ranges from 10 to 150 microns.

[0015] In a preferred embodiment of the present application, the repair material is composed of calcium phosphate cement, magnesium metal microspheres and 3D bioprinting high polymer polymer scaffold, and has a multi-level pore structure complementary to immediate pore formation and delayed pore formation, with a total porosity of 60% to 75%, and the overall strength of the repair material is 8 to 25 MPa, which can resist the pressure of oral mucosa and chewing.

[0016] In a preferred embodiment of the present application, the multi-level pore structure of the repair material is used to simulate the multi-level pore structure of natural bone.

[0017] The present application solves the defects in the background art, and has the following advantages:

[0018] (1) The magnesium microspheres hydration reaction produces 30-500 micron gradient micropores accounting for 75%, which is complementary to the 300-500 micron macropores formed by the degradation of the PLGA scaffold, with a total porosity of 68%, which is 3.4 times higher than that of traditional CPC. This structure makes the body fluid penetration rate increase by 3 times, shortens the vascularization time by 40%, and increases the bone integration efficiency by 55%. Moreover, the three-dimensional interconnected macropore structure and the micropore network form a "macro-micro" dual-level transport channel, promoting nutrient exchange and metabolic waste discharge.

[0019] (2) The magnesium microspheres are embedded in the CPC matrix in a gradient distribution, and slowly release Mg2+ through hydration reaction, avoiding local pH mutation. This release mode can sustainably activate VEGF expression for 28 days, which is 2 times longer than traditional magnesium-doped materials. The magnesium element doping optimizes the calcium-phosphorus ratio of the material to 1.50-1.70, which is closer to the inorganic components of natural bone, thereby promoting bone-like apatite deposition.

[0020] (3) The PLGA scaffold and the CPC matrix form a rigid-flexible coupling system, and the overall strength of the composite material is maintained at 10-20 MPa, so that it can withstand a maximum chewing pressure of about 15 MPa, improving the fatigue resistance, thereby being significantly superior to pure PLGA scaffold. At the same time, the acid produced by the degradation of PLGA and the neutralization reaction of CPC form a pH buffering effect, thereby avoiding the risk of local acidification or alkalization.

[0021] (4) The variable fiber arrangement is realized by high-temperature fused deposition process, and the pore gradient of the scaffold is accurately matched with the stress distribution of the bone defect site, which improves the pore control accuracy by 60% compared with traditional process. At the same time, the freeze-drying process makes nHA uniformly dispersed in the PLGA matrix, and the interfacial bonding strength of the composite material reaches 2.1 MPa, which is 3 times higher than that of traditional blending process. BRIEF DESCRIPTION OF DRAWINGS

[0022] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the accompanying drawings needed to be used in the embodiments or prior art description will be briefly introduced. Obviously, the accompanying drawings in the following description only represent some embodiments of the present application, and for those skilled in the art, other drawings can be obtained from these drawings without any creative effort.

[0023] Figure 1 is a preparation flow chart of the preferred embodiment of the present application;

[0024] Figure 2 is a scanning chart of the different quality ratio of the magnesium-containing microspheres MC bone cement of the preferred embodiment of the present application;

[0025] Figure 3 is a real object chart of the nHA-containing PLGA scaffold of different shapes of the preferred embodiment of the present application;

[0026] Figure 4 is a scanning electron microscope chart of the CPC / nPLGA and MC / nPLGA composite after the nPLGA scaffold of the preferred embodiment of the present application is degraded. DETAILED DESCRIPTION

[0027] The technical solutions in the embodiments of the present application will be described clearly and completely in the following description with reference to the accompanying drawings in the embodiments of the present application. Obviously, the described embodiments only represent some of the embodiments of the present application, but not all the embodiments. Based on the embodiments in the present application, all the other embodiments obtained by those skilled in the art without any creative effort fall within the protection scope of the present application.

[0028] In the following description, many specific details are set forth in order to provide a thorough understanding of the present application, but the present application can also be implemented in other ways different from those described herein, therefore, the protection scope of the present application is not limited by the specific embodiments disclosed below.

[0029] It needs to be pointed out that at present, the preparation technology of porous calcium phosphate ceramic materials mainly includes sacrificial template method, gas foaming method and 3D printing method. The sacrificial template method can improve the mechanical strength of CPC while forming reserved pores in the CPC matrix by introducing degradable pore-forming agents (degradable polymer microspheres, gelatin microspheres, glucose, polymer short fibers and short fiber network, etc.), although the porosity can be increased to 60%-70%, but the problems of poor pore connectivity and organic solvent residue exist. The gas foaming method uses hydrogen peroxide thermal decomposition or acid-base reaction of sodium bicarbonate to generate gas to construct pores, among which the sodium bicarbonate foaming method is widely used due to its simple operation and low cost, and connected pores with a pore size of 100-500 μm can be obtained, but the controllability of the foaming process is poor, and the mechanical properties of the bone cement are significantly degraded. The 3D printing technology can realize the controllable adjustment of pore size and porosity by precisely designing the scaffold pore structure, but high temperature sintering is needed to enhance the mechanical properties, and toxic solvents may be left in the printing slurry. The contradiction between the porous structure and the mechanical properties of the material restricts the application of calcium phosphate bone cement.

[0030] Therefore, the application quickly generates instant porous bone cement through the hydration reaction of magnesium metal microspheres and CPC, and synchronously releases Mg2 + ; The 3D bioprinting technology is used to construct a nano-hydroxyapatite modified high molecular polymer degradable scaffold, so that it can provide bone grafting space support for the porous bone cement in the early stage of implantation, gradually degrade in the later stage of implantation, and form a three-dimensional interconnected large pore in situ in the bone cement, so that a bone repair material with a bionic porous structure, high porosity and coordinated mechanical properties, and simultaneously releasing osteogenic active ingredients is obtained. This two-stage pore forming strategy not only realizes the formation and regulation of the pore structure, the coordination of the porosity and the mechanical properties, but also plays the biological activity effect of Mg2 + , and improves the osteogenesis and angiogenesis performance of the composite scaffold. The method of combining instant pore formation of magnesium microspheres in the calcium phosphate bone cement matrix and degradation of the degradable high molecular polymer scaffold to form interconnected large pores is used to form a three-dimensional porous interconnected structure in the calcium phosphate bone cement.

[0031] Specifically, as shown in Figure 1 : A preparation method of a gradient pore-forming magnesium-containing calcium phosphate bone repair material, comprising the following steps:

[0032] S1, nano-hydroxyapatite and polylactic acid-glycolic acid copolymer PLGA powder are mixed in a mass ratio of 5:95 to 25:75, ultrasonic dispersion is performed, dichloromethane is added as a solvent in a volume ratio of 1:3 to 1:5, continuous stirring is performed for 5-8 hours to form a solution, and nHA / PLGA raw materials are obtained by freeze-drying, and a degradable nPLGA scaffold with a three-dimensional interconnected large pore structure is prepared by a 3D bioprinter high-temperature fused deposition.

[0033] Further, the magnesium metal microspheres are dispersed in the calcium phosphate cement powder to generate hydrogen gas by hydration reaction to form a microporous structure with a pore size ranging from 30 to 500 μm, and release a bioactive substance Mg2 + . The Mg2 + has the effects of promoting osteogenesis and angiogenesis, the alkaline environment formed by magnesium is alleviated as the nPLGA scaffold degrades to produce acid, and the acid produced by the degradation of the nPLGA scaffold accelerates the degradation rate of the calcium phosphate cement.

[0034] S2, the calcium phosphate cement powder and magnesium metal microspheres are weighed according to a predetermined mass ratio, a solidification liquid is added at a liquid-solid ratio of 0.3-0.5 mL / g, the slurry is stirred and homogenized to obtain a calcium phosphate cement slurry containing magnesium metal microspheres.

[0035] Further, the freeze-drying process is divided into stages one and two; stage one includes a preset freezing temperature of-70 to-90°C and a pre-freezing time of 3-5 hours; stage two includes a preset drying temperature of-40 to-60°C, a vacuum degree of 0.05-0.2 Mbar, and a drying time of 48-72 hours.

[0036] When the 3D bioprinter implements high-temperature fused deposition manufacturing, the melting temperature is between 150 and 170°C, the extrusion pressure is between 1.5 and 4 bar, the printing speed is between 5 and 10 mm / s, 70%-90% of the nozzle diameter is used as the deposition layer height parameter, the fiber spacing is between 1 and 3 mm, and the orthogonal stacking structure is 0° / 90° / 90° to 0° / 60° / 60°. The 3D printing technology supports personalized scaffold customization and is suitable for personalized repair treatment of complex alveolar bone defects in the maxillofacial region.

[0037] The calcium-phosphorus ratio of the calcium phosphate cement powder ranges from 1.50 to 1.70, and the particle size of the magnesium metal microspheres ranges from 10 to 150 μm.

[0038] The repair material is composed of calcium phosphate cement, magnesium metal microspheres, and a 3D bioprinted high-molecular polymer scaffold, and has a multi-level porous structure that complements immediate pore formation and delayed pore formation, with a total porosity of 60%-75% and an overall strength of 8-25 MPa, which can resist oral mucosa and masticatory pressure.

[0039] The multi-level porous structure of the repair material is used to simulate the multi-level pore structure of natural bone.

[0040] The 50-300 μm micropores generated by the magnesium microspheres provide rapid perfusion of body fluids and nutrients, while the 300-500 μm macropores formed by the degradation of PLGA promote the growth of blood vessels and bone tissue, with a total porosity of 68%, which is significantly higher than the 30% of traditional CPC materials.

[0041] The initial compressive strength of the composite PLGA scaffold calcium phosphate cement is 17.8 MPa, which offsets the strength loss of CPC due to the immediate porous structure, and the overall strength of the composite scaffold is 10-20 MPa, which can resist the pressure of oral mucosa and chewing without collapse of the material.

[0042] S3, inject the calcium phosphate cement slurry obtained in step S2 into the nPLGA scaffold prepared in step S1, and cure at a constant temperature of 35-37°C and a humidity of 97-100% for 20-30 hours, and after demolding, obtain the MC / nPLGA composite.

[0043] Embodiment

[0044] S1, prepare porous magnesium-containing microsphere cement.

[0045] Take tetracalcium phosphate and dicalcium phosphate dihydrate as the main components, and the calcium-phosphorus ratio is 1.67. The particle size of the magnesium metal microspheres is 50 μm, and the curing liquid is selected as a phosphate buffer solution to prepare calcium phosphate cement (CPC) powder.

[0046] The specific preparation steps include:

[0047] S11, weigh 10 g of CPC powder and 0.2 g of magnesium microspheres, add a curing liquid with a liquid-solid ratio of 0.4 mL / g, that is, add 4.8 mL of phosphate buffer solution, and stir the slurry until it is homogeneous.

[0048] S12, inject the slurry into a Teflon mold with a size of 6x2 mm, and cure at 37°C and 95% humidity for 24 hours.

[0049] S13, after demolding, dry at 60°C under vacuum conditions for 12 hours to obtain an MC cement sample containing 2% magnesium microspheres by mass ratio.

[0050] S2, prepare nHA / PLGA degradable scaffold

[0051] Prepare an nHA / PLGA degradable scaffold using nano-hydroxyapatite (nHA), poly(lactic-co-glycolic acid) (PLGA) with a monomer ratio of PLA:PGA=85:15 and a molecular weight of 300 kDa, and dichloromethane as a solvent.

[0052] The specific preparation steps include:

[0053] S21, weigh the nHA and PLGA powders according to a mass ratio of 10:90 using an analytical balance to ensure the accuracy of the ratio.

[0054] S22, place the mixed powder in a 50 mL glass beaker and put it into an ultrasonic cleaning machine, adjust the power to 300 W and the frequency to 30 kHz, and continue processing for 30 minutes.

[0055] S23, the mixed powder after ultrasonic treatment was transferred to a 250 mL round-bottom flask, and 40 mL of dichloromethane was added in a volume ratio of 1:4.

[0056] S24, a magnetic stirrer was used, the stirring speed was set to 500 rpm, and stirring was continued for 6 hours until a milky white solution was formed.

[0057] S25, the completely dissolved mixed slurry was slowly poured into a flat glass dish with a diameter of 10 cm, placed in a fume hood, and the surrounding temperature was controlled to 25°C and the relative humidity was 30%, and left to stand for 24 hours to allow the solvent to evaporate naturally.

[0058] S26, after the solvent was completely volatilized, the preliminarily solidified material was taken out of the glass dish and cut into appropriate size blocks with a scalpel. It was placed in a freeze dryer and pre-frozen at -80°C for 4 hours, then dried at -50°C under a vacuum of 0.1 Mbar for 48 hours to completely remove the residual solvent. The nHA / PLGA raw material was obtained.

[0059] S27, high-temperature fused deposition manufacturing was carried out using a Bio-Architect WS 3D bioprinter. Digital modeling of the scaffold was completed by Materialise Magics 25.0 engineering software. The deposition layer height parameter was set to 80% of the nozzle diameter using the layering software provided with the printer, for example, if the nozzle diameter is 0.4 mm, the deposition layer height is 0.32 mm. The fiber spacing was adjusted to 2 mm, and the orthogonal stacking structure was set to 0° / 90°.

[0060] The preset melting temperature was 160°C, the extrusion pressure was 2.5 bar, and the printing speed was 7 mm / s. The PLGA particles with LA:GA=85:15 and Mw=300 kDa were loaded into the high-temperature stainless steel cartridge, and after 30 minutes of constant temperature melting, layer-by-layer deposition was carried out, and finally a PLGA scaffold with regular porous structure was obtained.

[0061] S3, construction of MC / nPLGA composite

[0062] S31, the prepared MC bone cement slurry was stirred uniformly and injected into the nPLGA scaffold to obtain the MC / nPLGA composite.

[0063] Comparative example

[0064] Calcium phosphate cement (CPC) was used as raw material, combined with 3D printing technology to obtain CPC scaffold with interconnected large pores, and dopamine solution was used for surface modification of the scaffold, and BMP-2 was loaded to promote the proliferation and osteogenic differentiation of mesenchymal stem cells.

[0065] The specific preparation process comprises: mixing the CPC solid phase powder and the solidification liquid, and stirring uniformly. The CPC slurry is printed into a scaffold with interconnected large pores through a 3D printing technology. The scaffold is placed in a dopamine alkaline aqueous solution to form a polydopamine layer. The polydopamine modified scaffold is immersed in a BMP-2 solution, and after standing, freeze-drying is performed to finally obtain a calcium phosphate-based bone repair scaffold.

[0066] The comparison results of the examples and the comparative examples are shown in Table 1.

[0067] Table 1

[0068] Comparative item Embodiment Comparative example Advantages of the embodiment Material composition CPC, magnesium metal microspheres, PLGA scaffold CPC only The embodiment introduces magnesium metal microspheres and PLGA scaffolds to construct composite materials, with more abundant functionality and biological activity. Pore structure Multistage pore structure (micropores 30~500μm, macropores 200~600μm), total porosity 60%~75% Connected macropores, but porosity and pore size range are not specified The multistage pore structure of the embodiment is more conducive to cell ingrowth and nutrient perfusion, with higher total porosity. Mechanical properties Overall strength of the composite scaffold 8~25MPa, able to withstand oral mucosa and chewing pressure The scaffold has connected macropores, meeting the mechanical strength requirements of cancellous bone, but the specific values are not given The embodiment provides specific mechanical property data, indicating that its strength is sufficient to cope with the pressure in the oral environment. Biological activity Mg2 + , which has osteoconductive and angiogenic effects; and the acid produced by PLGA degradation accelerates CPC degradation Loading of BMP-2 promotes cell adhesion and proliferation The magnesium microspheres and PLGA degradation characteristics in the embodiment provide dual biological activity, more conducive to bone repair. Personalized customization 3D bioprinting technology is used to support personalized scaffold customization 3D printing technology, but no mention of personalized customization The embodiment emphasizes the application of 3D bioprinting technology in personalized customization, more suitable for the repair of complex bone defects.

[0069] It can be seen from the comparison that the examples have significant advantages in material composition, pore structure, mechanical properties, biological activity and individual customization. The examples introduce magnesium metal microspheres and PLGA scaffolds to construct a composite material with a multi-level pore structure and excellent mechanical properties, and at the same time, utilize the biological activity of magnesium and the degradation characteristics of PLGA to achieve better bone repair effect. In addition, the application of 3D bioprinting technology also makes the examples have significant advantages in individual customization, and are more suitable for the repair treatment of complex bone defects.

[0070] Based on the ideal embodiments of the present application, through the above description, relevant personnel can make various changes and modifications without deviating from the technical idea of the present application. The technical scope of the present application is not limited to the contents in the specification, and must be determined according to the scope of the claims.

Claims

1. A method for preparing a gradient-drilled magnesium-containing calcium phosphate bone repair material, characterized by, The method comprises the following steps: S1, mixing nano-hydroxyapatite and polylactic acid-glycolic acid copolymer PLGA powder according to a mass ratio of 5:95 to 25:75, ultrasonic dispersion, adding dichloromethane as a solvent in a volume ratio of 1:3 to 1:5, continuously stirring for 5-8 hours to form a solution, and obtaining nHA / PLGA raw materials by freeze-drying, and then obtaining a degradable nPLGA scaffold with a three-dimensional interconnected macroporous structure by high-temperature fused deposition of a 3D bioprinter; S2, taking calcium phosphate cement powder and magnesium metal microspheres according to a preset mass ratio, adding a solidification liquid in a liquid-solid ratio of 0.3-0.5 mL / g, stirring the slurry to be homogeneous, and obtaining a calcium phosphate cement slurry containing magnesium metal microspheres; S3, injecting the calcium phosphate cement slurry obtained in step S2 into the nPLGA scaffold prepared in step S1, and curing for 20-30 hours under the conditions of a constant temperature of 35-37℃ and a humidity of 97-100%, and then demolding to obtain an MC / nPLGA composite.

2. The preparation method of the gradient porous calcium phosphate bone repair material containing magnesium according to claim 1, characterized in that: The magnesium metal microspheres are dispersed in calcium phosphate bone cement powder, hydrogen gas is generated by hydration reaction to form a microporous structure, the micropore size ranges from 30 to 500 μm, and the bioactive substance Mg2 + is released.

3. The preparation method of the gradient porous calcium phosphate bone repair material containing magnesium according to claim 1, characterized in that: The freeze-drying process is divided into phase one and phase two; wherein phase one includes a preset freezing temperature of-70 to-90℃ and a pre-freezing time of 3-5 hours; and phase two includes a preset drying temperature of-40 to-60℃, a vacuum degree of 0.05-0.2 Mbar, and a drying time of 48-72 hours.

4. The preparation method of the gradient pore-containing calcium phosphate bone repair material containing magnesium according to claim 1, characterized in that: When the 3D bioprinter is implemented for high-temperature fused deposition manufacturing, the melting temperature is between 150-170℃, the extrusion pressure is between 1.5-4 bar, the printing speed is between 5-10 mm / s, 70-90% of the nozzle diameter is used as the deposition layer height parameter, the fiber spacing is between 1-3 mm, and the orthogonal stacking structure is 0° / 90° / 90° to 0° / 60° / 60°.

5. The preparation method of the gradient pore-containing magnesium-containing calcium phosphate bone repair material according to claim 1, characterized in that: The calcium phosphate cement powder has a calcium-phosphorus ratio in the range of 1.50-1.70, and the magnesium metal microspheres have a particle size in the range of 10-150 μm.

6. A gradient-porosity calcium phosphate bone repair material containing magnesium, produced according to the method of any one of claims 1 to 5, characterized in that, The repair material is composed of calcium phosphate cement, magnesium metal microspheres, and a 3D bioprinted high-molecular polymer scaffold, and has a multi-level pore structure with immediate pore formation and delayed pore formation complementation, a total porosity of 60-75%, and an overall strength of the repair material of 8-25 MPa, which can resist oral mucosa and chewing pressure.

7. The gradient porous magnesium-containing calcium phosphate bone repair material according to claim 6, characterized in that: The multi-level pore structure of the repair material is used to simulate the multi-level pore structure of natural bone.

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