Method for compositing inorganic mineral and protein

Abstract

The present invention provides a method for compositing an inorganic mineral and a protein. A uniformly mixed protein solution and an inorganic mineral solution are injected into a shear stress loading chamber, and a dehydrating agent is added into a dialysis chamber. The shear stress loading chamber and the dialysis chamber are separated by means of a dialysis membrane. The applied shear stress is 1-10 Pa, the action time is 1-8 days, and the reaction temperature is 10-50 °C. The molecular weight cut-off of the dialysis membrane is in a range of 3-50 kDa. The resulting composite material has excellent mechanical properties, including high strength and toughness, exhibits good biocompatibility, and thus can support cell adhesion, proliferation, and differentiation, thereby effectively promoting bone tissue regeneration. The composite material prepared by the present invention has broad application prospects in bone repair and other biomedical fields.

Description

A method for combining inorganic minerals and proteins Technical Field

[0001] This invention belongs to the field of biomedical material preparation technology, and in particular relates to a method for preparing inorganic mineral and protein composites. This method is based on shear stress and dialysis concentration technology to prepare inorganic mineral and protein-based composite materials, which are applicable to the field of tissue engineering, such as bone defect repair. Background Technology

[0002] With the rapid development of bone repair and tissue engineering, the development of biomedical materials with good mechanical properties and biocompatibility has become a research focus. Bone tissue has a complex hierarchical structure, and its regeneration and repair require materials with high mechanical strength, excellent biocompatibility, and the ability to support cell adhesion, proliferation, and differentiation. Although traditional bone repair materials possess certain bioactivity, they still have shortcomings in terms of mechanical properties, tissue integration, and material processability. Therefore, the development of composite materials that simultaneously possess excellent mechanical properties and biological functions is particularly important.

[0003] In bone repair and tissue engineering applications, proteins (such as collagen, serum proteins, silk fibroin, and fibronectin) have been widely used in the preparation of biomedical materials due to their excellent biocompatibility and ability to support cell growth. These proteins, at high concentrations, can form three-dimensional fibrous structures similar to natural tissues, which is crucial for mimicking the mechanical properties and biological functions of bone tissue. However, because protein solutions are typically at low concentrations and the fibers are unevenly arranged, their mechanical properties and structural stability are limited, making it difficult to meet the needs of clinical applications. Existing bone repair materials have shortcomings in terms of mechanical properties, biocompatibility, and tissue integration, making it difficult to simultaneously meet the multiple requirements of high strength, toughness, and cell support. Summary of the Invention

[0004] In view of this, the present invention aims to propose a method for composite materials of inorganic minerals and proteins to overcome the shortcomings of existing technologies. This invention is a method for preparing inorganic mineral and protein-based composite materials based on shear stress and dialysis concentration technology. It is particularly suitable for the field of biomedical materials, such as bone defect repair and tissue engineering applications. Through the method of this invention, the concentration of the protein solution can be effectively increased, the orderliness of the fiber arrangement can be improved, and its composite with inorganic minerals can be promoted, ultimately producing a composite scaffold material with excellent mechanical properties and biocompatibility.

[0005] To achieve the above objectives, the technical solution of the present invention is implemented as follows:

[0006] A method for combining inorganic minerals and proteins involves injecting a uniformly mixed protein solution and an inorganic mineral solution into a shear stress loading chamber, adding a dehydrating agent to a dialysis chamber, separating the shear stress loading chamber and the dialysis chamber by a dialysis membrane, applying a shear stress of 1-10 Pa, acting for 1-8 days, and reacting at a temperature of 10-50°C; the dialysis membrane has a molecular weight cutoff range of 3-50 kDa.

[0007] The magnitude, duration, and temperature of shear stress are adjustable. Shear stress can improve the fluidity of proteins and cause them to align in an orderly manner along the shear direction, thereby forming a uniform three-dimensional fibrous structure.

[0008] The rotational speed of the rotating cone plate in the shear stress loading chamber is calculated based on the following three formulas to apply a fixed amount of shear stress:

[0009]

[0010] Shear rate (s) -1 ); Shear stress (Pa); Dynamic viscosity (Pa·s); : Cone angle (radian); Ω: Angular velocity (rad / s);

[0011] Preferably, the initial concentration of the protein solution is 1~10 mg / mL; the concentration of the inorganic mineral solution is 1~100 mM; and the volume ratio of the protein solution to the inorganic mineral solution is 1:1~1:10.

[0012] Preferably, the inorganic mineral solution is a solution containing one or more of the following: calcium ions, phosphate ions, strontium ions, iron ions, zinc ions, silver ions, magnesium ions, citrate, insoluble strontium salt, and insoluble magnesium salt.

[0013] Preferably, the inorganic mineral is one or more of hydroxyapatite, calcium carbonate, calcium phosphate, amorphous calcium phosphate, and amorphous strontium carbonate solution.

[0014] Preferably, the inorganic mineral solution contains an organic additive, wherein the organic additive is one or more of polyaspartic acid, sodium citrate, polyacrylic acid, and carboxymethyl chitosan.

[0015] The functions of the organic additives include, but are not limited to: a) stabilizing the amorphous calcium phosphate during the composite process, slowing down its phase transformation and maintaining it in a metastable liquid state, thus facilitating its entry into the organic matrix (such as the interior of collagen fibers) and achieving superior composite properties; b) providing additional functions, such as imparting antibacterial properties to the material by adding specific organic matrices (such as antibacterial materials like carboxymethyl chitosan). Through the synergistic effect of the aforementioned inorganic minerals and organic additives, the resulting composite material exhibits superior mineralization ability, biocompatibility, and functionality in biomedical applications.

[0016] Preferably, the protein is one or more of the following: collagen, silk fibroin, keratin, elastin, fibrin, lysozyme, fibronectin, serum albumin, lactoferrin, transferrin, insulin, myoglobin, hemoglobin, egg white albumin, β-lactoglobulin, α-lactalbumin, or amyloid proteins generated during the phase transition of the above proteins.

[0017] Structural proteins: These provide mechanical support and structural stability for composite materials and include collagen, silk fibroin, keratin, elastin, and fibroin. Collagen imparts good biocompatibility and mechanical strength to materials; silk fibroin and keratin enhance strength and toughness; elastin is suitable for tissue repair materials requiring flexibility; and fibroin contributes to gelation and structural stability.

[0018] Functional proteins: These enhance the bioactivity and antibacterial properties of composite materials and promote cell adhesion, growth, and differentiation. They include lysozyme, fibronectin, serum proteins (such as human serum albumin and bovine serum albumin), lactoferrin, transferrin, insulin, myoglobin, hemoglobin, egg white albumin, β-lactoglobulin, and α-lactalbumin. Lysozyme and lactoferrin possess antibacterial activity; fibronectin promotes cell adhesion and proliferation; serum proteins improve the biocompatibility of materials; transferrin and insulin promote cell differentiation and tissue healing; myoglobin and hemoglobin provide local oxygen delivery support in repair materials with high oxygen requirements; and egg white albumin, β-lactoglobulin, and α-lactalbumin provide nutritional support.

[0019] The amyloid protein generated during the phase transition of the aforementioned proteins is also included in the protein solution to enhance the mechanical strength, bioactivity, and structural stability of the composite material.

[0020] Preferably, the dehydrating agent is one or more of polyethylene glycol solution, salt solution, sodium polyacrylate, sodium alginate, sodium carboxymethyl cellulose, or silica gel. The salt solution is one or more of NaCl, KCl, Na₂HPO₄, and KH₂PO₄ solution.

[0021] Preferably, the concentration of the polyethylene glycol solution is 20~400 mg / mL, more preferably 250~350 mg / mL; the salt concentration of the salt-containing solution is 3~30 g / L.

[0022] Preferably, the shear stress loading chamber also contains a polysaccharide compound or a polyphenol compound; the concentration of the added polysaccharide compound or polyphenol compound is 0~10 mg / mL; it is added when it is necessary to improve the bioactivity, biocompatibility or mechanical properties of the material; preferably, the polysaccharide compound is one or more of chondroitin sulfate and hyaluronic acid, and the polyphenol compound is proanthocyanidins.

[0023] Preferably, the flow rate of the liquid in the shear stress chamber and the dialysis chamber is 1 mL / min to 15 mL / min, to maintain the pressure balance on both sides of the dialysis membrane, remove water and impurities from the solution, and achieve the purpose of concentration.

[0024] High-concentration inorganic mineral-protein composite materials were collected under dialysis concentration and shear stress. The resulting material not only exhibited high concentration but also a uniform fibrous structure, demonstrating excellent mechanical properties and biocompatibility. The concentrated organic-inorganic composite material can be further processed according to application requirements, for example: transforming the composite gel into a porous scaffold material through freeze-drying; forming a dense scaffold through mechanical compression for high-load bone repair applications; preparing the freeze-dried material into powder form for easy storage or use in different environments, or using it to prepare other composite materials.

[0025] The shear stress loading chamber described in the above method is implemented by a hydraulic shear loading device, which has a rotating impeller or piston structure capable of applying constant shear stress, or the device is a cone-plate viscometer, injection pump, or flat plate flow chamber circulation device capable of providing constant directional fluid shear force.

[0026] This invention also provides the application of inorganic mineral and protein-based composite materials prepared by the method described above in the preparation of bone repair or other biomedical products.

[0027] Shear thinning refers to the phenomenon that the viscosity of a solution decreases with increasing shear rate under shear stress. Appropriate application of fluid shear force can significantly improve the physical properties of protein solutions (such as collagen solutions). Shear thinning enhances the fluidity of solutions, making operation at high concentrations more feasible, which facilitates protein concentration. Under shear stress, proteins not only reduce flow resistance but also achieve a more ordered arrangement along the shear direction, forming a three-dimensional fibrous matrix with uniform structure and excellent mechanical properties. This matrix not only mimics the fibrous arrangement in natural bone tissue but also enhances the overall mechanical properties of the material.

[0028] Besides shearing techniques, dialysis concentration also plays a crucial role in the concentration of protein solutions and the preparation of composite materials with inorganic minerals. By removing excess water and small molecule impurities, dialysis concentration effectively increases protein concentration, thereby enhancing its binding with inorganic minerals (such as hydroxyapatite and calcium phosphate). The concentrated, high-concentration protein solution can tightly bind with inorganic minerals to form a high-strength composite material. This composite material possesses excellent mechanical properties and bioactivity, supporting cell growth and bone tissue regeneration.

[0029] This invention combines shearing and dialysis concentration techniques. Shear stress enhances the fluidity of protein solutions and the ordered arrangement of fibers. By concentrating the protein through dialysis and combining it with inorganic minerals, a composite scaffold material with excellent mechanical properties is prepared. These composite materials not only retain the hardness and compressive strength of inorganic minerals but also possess the flexibility and biocompatibility of biomatrix materials, making them highly suitable for bone repair. By rationally controlling shear stress and concentration conditions, the composite material exhibits good fluidity and processability, while displaying high strength and high toughness after final curing, making it suitable for tissue engineering applications such as bone repair and other biomedical fields.

[0030] Compared with existing technologies, the method for compounding inorganic minerals and proteins described in this invention has the following advantages:

[0031] This invention, by combining shear stress and dialysis concentration techniques, not only effectively increases the concentration of protein solutions but also improves the arrangement of protein fibers and the uniformity of the material. This technology promotes the tight binding of proteins with inorganic minerals, forming high-strength composite materials with excellent biological functions and mechanical properties. These materials support cell adhesion, proliferation, and differentiation, thereby accelerating bone tissue regeneration.

[0032] Compared with existing technologies, this invention significantly improves the concentration and composite efficiency of proteins and inorganic minerals by combining shear stress and dialysis concentration, and forms a composite scaffold material with excellent mechanical properties and biological functions under high concentration conditions. This material can be widely used in bone repair, tissue engineering, and other biomedical fields, providing a new method for efficiently preparing high-performance biomedical materials. Attached Figure Description

[0033] The accompanying drawings, which form part of this invention, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an undue limitation of the invention. In the drawings:

[0034] Figure 1 is a schematic diagram of the preparation of inorganic mineral and protein-based composite materials based on shear stress and dialysis concentration technology;

[0035] Figure 2 shows scanning electron microscope (SEM) images of concentrated mineralized collagen scaffolds and statically unconcentrated mineralized collagen scaffolds prepared according to the method of Example 1; where low magnification is 3000x and high magnification is 10000x.

[0036] Figure 3 shows the elemental distribution (EDS Mapping) of the mineralized collagen scaffold prepared based on the method of Example 1;

[0037] Figure 4 shows the elemental analysis of the mineralized collagen scaffold prepared based on the method of Example 1;

[0038] Figure 5 is a comparison of thermogravimetric analysis of concentrated mineralized collagen scaffolds prepared based on the method of Example 1 and static unconcentrated mineralized collagen scaffolds;

[0039] Figure 6 is a comparison of the Young's modulus of concentrated mineralized collagen scaffolds prepared according to the method of Example 1 and static unconcentrated mineralized collagen scaffolds.

[0040] Figure 7 is a scanning electron microscope image of the concentrated amorphous calcium phosphate and lysozyme composite material prepared based on the method of Example 3; where low magnification is 200x and high magnification is 1000x.

[0041] Figure 8 is a scanning electron microscope image of cells on the surface of the concentrated mineralized collagen scaffold prepared according to the method of Example 1; where the magnification is 1000x.

[0042] Figure 9 shows a scanning electron microscope image of the concentrated mineralized collagen scaffold prepared based on the method of Example 4; the magnification is 3000x.

[0043] Figure 10 shows a scanning electron microscope image of the concentrated mineralized collagen scaffold prepared based on the method of Example 5;

[0044] The magnification is 5000x. Detailed Implementation

[0045] Unless otherwise defined, the technical terms used in the following embodiments have the same meanings as commonly understood by those skilled in the art. Unless otherwise specified, the experimental reagents used in the following embodiments are conventional biochemical reagents; and the experimental methods described are conventional methods.

[0046] The present invention will be described in detail below with reference to embodiments.

[0047] Example 1: Preparation of high-concentration inorganic mineral and collagen composite materials based on shear stress and dialysis concentration

[0048] A 3 mg / mL acidic type I collagen solution was mixed with an amorphous calcium phosphate solution at a 1:1 volume ratio. The amorphous calcium phosphate solution contained 240 µg / mL polyaspartic acid, 1.67 mM calcium ions, 9.5 mM phosphate ions, 150 mM sodium chloride, and 10 mM sodium citrate. The mixed solution was placed in the upper shear stress loading chamber, and the shear loading device was activated to apply a shear stress of 1 Pa, with the temperature set to 37°C.

[0049] A 300 mg / mL polyethylene glycol (PEG) solution was placed in the lower dialysis chamber as a dehydrating agent. The molecular weight cutoff of the dialysis membrane was 8000-14000 Da. The solutions on both sides of the dialysis membrane were continuously injected into the upper stress-loading chamber and the lower dialysis chamber at a flow rate of 1 mL / min, respectively.

[0050] Under the influence of dialysis and shear stress, after 5 days of treatment, the collagen-amorphous calcium phosphate composite material gradually concentrated, with the collagen fibers arranged in an orderly manner along the shear direction. After treatment, the composite material in the upper chamber was collected. This material exhibited high concentration, good fiber arrangement, and excellent mechanical properties, making it suitable for bone repair and tissue engineering applications.

[0051] Comparative Example 1: Preparation of Inorganic Mineral and Collagen Composite Materials under Static Conditions

[0052] A 3 mg / mL acidic type I collagen solution was mixed with an amorphous calcium phosphate solution at a 1:1 volume ratio. The amorphous calcium phosphate solution contained 240 µg / mL polyaspartic acid, 1.67 mM calcium ions, 9.5 mM phosphate ions, 150 mM sodium chloride, and 10 mM sodium citrate. The mixed solution was placed in a beaker, and the reaction temperature was set to 37°C.

[0053] Under static conditions, after 5 days of processing, scanning electron microscopy revealed a relatively loose collagen fiber structure with obvious porous regions visible under low magnification. High-magnification images further showed gaps between fibers, and the three-dimensional network density was significantly lower than that of the shear-concentrated sample, exhibiting an overall porous and discontinuous morphology. The deposition of mineralized materials (such as calcium phosphate crystals) on the collagen fiber surface was uneven and poorly oriented. Compared to the mineralized collagen scaffold prepared by the method in Example 1, the mechanical properties of the material were reduced.

[0054] Figure 2 shows that after shear dialysis concentration, the collagen fibers exhibit significantly denser structure with no obvious pores or loose areas between fibers. Under low magnification, a uniform, continuous network structure is visible, while high magnification further reveals tightly packed fibers forming a high-density three-dimensional network. The fiber arrangement demonstrates a high degree of order; longitudinal section observation shows that the collagen fibers are distributed in parallel layers with clear hierarchies and consistent orientation. Hydroxyapatite crystals are uniformly deposited on the surface and in the interstices of the collagen fibers, forming a continuous mineralization layer. High-magnification SEM images show nanoscale mineralization particles tightly attached to the fiber surface, forming a typical "collagen-mineral" composite structure. The SEM results of concentrated mineralized collagen demonstrate that the shear dialysis process successfully achieved densification, ordering, and efficient mineralization of the collagen structure. Compared with concentrated mineralized collagen subjected to shear force, the sample without shear force showed significant deterioration in density, ordered arrangement, and mineralization uniformity.

[0055] In Figure 3, combined with energy dispersive spectroscopy analysis, calcium and phosphorus elements are evenly distributed on collagen fibers, further confirming the high degree of mineralization and uniform distribution.

[0056] In Figure 4, the elemental analysis results explain that the calcium-to-phosphorus ratio of mineralized collagen is 1.49, which is close to the calcium-to-phosphorus ratio of hydroxyapatite in natural bone.

[0057] Figure 5 shows that after 5 days of mineralization, the residual weight of inorganic minerals in the concentrated mineralized collagen scaffold was 65.53%, which was significantly higher than that in the static group (47.59%). This indicates that the concentrated mineralized collagen scaffold prepared by shear dialysis can significantly improve the deposition of inorganic minerals.

[0058] In Figure 6, the average DMT modulus (representing Young's modulus) of the shear force group was 2.5 GPa, significantly higher than the 1.2 GPa of the static group. The higher Young's modulus indicates that shear dialysis treatment significantly improved the stiffness of the mineralized collagen scaffold, reflecting a denser inorganic mineral-collagen composite structure. The Young's modulus comparison in Figure 6 also shows that the introduction of shear force can effectively increase the Young's modulus of the composite mineralized collagen scaffold.

[0059] In Figure 8, SEM images show that the cells exhibit numerous extended filamentous and plate-like pseudopodia, adhering well to the surface of the concentrated mineralized collagen scaffold. This demonstrates that the high-strength composite material prepared based on this technology possesses excellent biological functions and can support cell adhesion.

[0060] Example 2: Preparation of high-concentration inorganic mineral and hyaluronic acid / collagen composite materials based on shear stress and dialysis concentration

[0061] A 2 mg / mL hyaluronic acid solution and a 3 mg / mL acidic collagen solution were mixed at a volume ratio of 1:10, and then uniformly mixed with an amorphous strontium carbonate solution at a volume ratio of 1:2. The amorphous strontium carbonate solution contained 2 mg / mL carboxymethyl chitosan, 10 mM strontium ions, and 6 mM carbonate ions. The mixture was placed in a shear stress loading chamber, and a shear stress of 2 Pa was applied at a temperature of 37°C.

[0062] A 300 mg / mL polyethylene glycol (PEG) solution was placed in the dialysis chamber as a dehydrating agent. The molecular weight cutoff of the dialysis membrane was 8000-14000 Da. The mixed solution and the PEG solution were injected into the shear stress chamber and the dialysis chamber, respectively, at a flow rate of 1 mL / min.

[0063] After 5 days of processing, the fibers of hyaluronic acid and collagen were orderly arranged along the shear direction and combined with amorphous strontium carbonate. The composite material in the upper chamber was finally collected, exhibiting excellent mechanical properties and fiber arrangement, making it suitable for bone repair and tissue engineering.

[0064] Example 3: Preparation of a high-concentration amorphous calcium phosphate and lysozyme composite material based on shear stress and dialysis concentration

[0065] A 3 mg / mL lysozyme solution was mixed with an amorphous calcium phosphate solution at a 1:1 volume ratio. The amorphous calcium phosphate solution contained 240 µg / mL polyacrylic acid, 1.67 mM calcium ions, 9.5 mM phosphate ions, and 150 mM sodium chloride. This mixture was then placed in a shear stress loading chamber, where a shear stress of 5 Pa was applied at a temperature of 37°C.

[0066] A 300 mg / mL polyethylene glycol (PEG) solution was placed in the dialysis chamber as a dehydrating agent. The molecular weight cutoff of the dialysis membrane was 8000-14000 Da. The mixed solution and the PEG solution were injected into the shear stress chamber and the dialysis chamber, respectively, at a rate of 1 mL / min.

[0067] After 5 days of processing, lysozyme and amorphous calcium phosphate were gradually concentrated under shear stress. The composite material in the upper chamber was collected; this material exhibits good mechanical properties and is suitable for bone repair and tissue engineering applications.

[0068] As shown in Figure 7, scanning electron microscopy results reveal that after shear dialysis concentration, the surface of the amorphous calcium phosphate and lysozyme composite material exhibits a continuous and dense composite network. The amorphous calcium phosphate particles are uniformly embedded in the lysozyme matrix, without obvious pores or cracks. This indicates that shear force and concentration promote the tight interweaving of amorphous calcium phosphate and lysozyme, forming a hierarchical structure similar to natural biomineralization.

[0069] Example 4: Preparation of high-concentration inorganic mineral and collagen composite materials based on shear stress and dialysis concentration

[0070] A 3 mg / mL acidic type I collagen solution was mixed with an amorphous calcium phosphate solution at a 1:1 volume ratio. The amorphous calcium phosphate solution contained 240 µg / mL polyaspartic acid, 1.67 mM calcium ions, 9.5 mM phosphate ions, 150 mM sodium chloride, and 10 mM sodium citrate. The mixed solution was placed in the upper shear stress loading chamber, and the shear loading device was activated to apply a shear stress of 5 Pa, with the temperature set to 37°C.

[0071] A 300 mg / mL polyethylene glycol (PEG) solution was placed in the lower dialysis chamber as a dehydrating agent. The molecular weight cutoff of the dialysis membrane was 8000-14000 Da. The solutions on both sides of the dialysis membrane were continuously injected into the upper stress-loading chamber and the lower dialysis chamber at a flow rate of 1 mL / min, respectively.

[0072] Under the influence of dialysis and shear stress, after 5 days of treatment, the collagen-amorphous calcium phosphate composite material gradually concentrated, with the collagen fibers arranged in an orderly manner along the shear direction. After treatment, the composite material in the upper chamber was collected. This material exhibited high concentration, good fiber arrangement, and excellent mechanical properties, making it suitable for bone repair and tissue engineering applications.

[0073] As shown in Figure 9, scanning electron microscopy reveals that after dialysis concentration under a shear stress of 5 Pa, the collagen fiber structure becomes significantly denser, with no obvious pores or loose areas between fibers. The fiber arrangement exhibits a high degree of order, with hydroxyapatite crystals uniformly deposited on the surface and in the gaps between the collagen fibers, forming a continuous mineralized layer.

[0074] Example 5: Preparation of high-concentration inorganic mineral and collagen composite materials based on shear stress and dialysis concentration

[0075] A 3 mg / mL acidic type I collagen solution was mixed with an amorphous calcium phosphate solution at a 1:1 volume ratio. The amorphous calcium phosphate solution contained 240 µg / mL polyaspartic acid, 1.67 mM calcium ions, 9.5 mM phosphate ions, 150 mM sodium chloride, and 10 mM sodium citrate. The mixed solution was placed in the upper shear stress loading chamber, and the shear loading device was activated to apply a shear stress of 10 Pa, with the temperature set to 37°C.

[0076] A 300 mg / mL polyethylene glycol (PEG) solution was placed in the lower dialysis chamber as a dehydrating agent. The molecular weight cutoff of the dialysis membrane was 8000-14000 Da. The solutions on both sides of the dialysis membrane were continuously injected into the upper stress-loading chamber and the lower dialysis chamber at a flow rate of 1 mL / min, respectively.

[0077] Under the influence of dialysis and shear stress, after 5 days of treatment, the collagen-amorphous calcium phosphate composite material gradually concentrated, with the collagen fibers arranged in an orderly manner along the shear direction. After treatment, the composite material in the upper chamber was collected. This material exhibited high concentration, good fiber arrangement, and excellent mechanical properties, making it suitable for bone repair and tissue engineering applications.

[0078] As shown in Figure 10, scanning electron microscopy reveals that after dialysis concentration under a shear stress of 10 Pa, the collagen fiber structure becomes significantly denser, with no obvious pores or loose areas between fibers. The fiber arrangement exhibits a high degree of order, with hydroxyapatite crystals uniformly deposited on the surface and in the gaps between the collagen fibers, forming a continuous mineralized layer.

[0079] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for compounding inorganic minerals and proteins, characterized in that: A uniformly mixed protein solution and inorganic mineral solution are injected into a shear stress loading chamber. A dehydrating agent is added to the dialysis chamber. The shear stress loading chamber and the dialysis chamber are separated by a dialysis membrane. The applied shear stress is 1-10 Pa, the reaction time is 5-8 days, the reaction temperature is 10-50℃, and the flow rate of the liquid in the shear stress loading chamber and the dialysis chamber is 1-15 mL / min. The molecular weight cutoff of the dialysis membrane is 3-14 kDa. The initial concentration of the protein solution is 1-10 mg / mL. The concentration of the inorganic mineral solution is 1-100 mM. The volume ratio of the protein solution to the inorganic mineral solution is 1:1-1:

10. The inorganic mineral is one or more of hydroxyapatite, calcium carbonate, calcium phosphate, amorphous calcium phosphate, and amorphous strontium carbonate solution; the inorganic mineral solution contains organic additives, which are one or more of polyaspartic acid, sodium citrate, polyacrylic acid, and carboxymethyl chitosan. The protein is one or more of the following proteins that are generated during phase transition: collagen, silk fibroin, keratin, elastin, fibrin, lysozyme, fibronectin, serum albumin, lactoferrin, transferrin, insulin, myoglobin, hemoglobin, egg white albumin, β-lactoglobulin, and α-lactalbumin. The dehydrating agent is a polyethylene glycol solution with a concentration of 250-350 mg / mL.

2. The method for compounding inorganic minerals and proteins according to claim 1, characterized in that: The shear stress loading chamber also contains polysaccharide or polyphenol compounds; the concentration of the added polysaccharide or polyphenol compounds is 0~10 mg / mL.

3. The method for compounding inorganic minerals and proteins according to claim 1, characterized in that: The polysaccharide compounds are one or more of chondroitin sulfate and hyaluronic acid, and the polyphenolic compounds are proanthocyanidins.

4. The use of inorganic mineral and protein-based composite materials prepared by the method of inorganic mineral and protein compounding as described in any one of claims 1-3 in the preparation of bone repair or other biomedical products.

Images