A method for preparing nanoenzyme-modified orthopedic polymer and its product and application

Abstract

The present invention discloses a method for preparing nanozyme-modified orthopedic polymers, and its products and applications, comprising the following steps: obtaining a modified polymer by in-situ growth of nanozymes in a polymer; mechanically processing the modified polymer to form a polymer implant; and using the modified polymer implant for the treatment, relief and / or prevention of orthopedic diseases. The nanozyme-modified orthopedic polymer implant prepared by the present invention can reduce the postoperative medication dosage, reduce adverse drug reactions and economic burden; enhance mechanical properties, reduce complications after implantation; prolong the protective effect on the body; avoid physiological toxicity; and can regulate and design multi-enzyme activities for personalized customization of diseases.

Description

Technical Field

[0001] The present invention relates to a method for preparing a nanoenzyme-modified orthopedic polymer, a product thereof and an application thereof, and belongs to the intersecting field of biomedical engineering and functional materials. Background Art

[0002] Bone tissue provides structural support for the body, and maintaining the health and integrity of bone tissue is essential for normal life and movement of the human body. However, with the deepening of global aging and changes in people's lifestyles, the incidence of orthopedic diseases has shown an upward trend, causing damage to patients' bone tissue function. In response to this phenomenon, polymers are widely used in the treatment of orthopedic diseases due to their good biocompatibility, processability and mechanical properties. As an implantable medical device, they play the role of repairing, enhancing and replacing bone tissue function or acting as a support or anchor for artificial cartilage.

[0003] Bone tissue maintains functional stability, that is, bone metabolic balance, through a dynamic balance between osteogenesis and osteoclastogenesis. Functional defects of bone tissue are usually caused by physical damage or long-term accumulated imbalance in bone metabolism. Orthopedic diseases that cause severe damage to bone tissue function include advanced arthritis, osteoporosis, bone tumors, osteomyelitis, large bone defects, and pyramidal fractures, which are characterized by the body's own bone metabolic imbalance and the failure of drug treatment, and surgery is required to manage the lesion area. In this case, the ideal polymer implant not only needs to play a supporting role at the mechanical level, but also needs to assist the body in restoring bone metabolic balance as much as possible to help patients achieve good postoperative healing and functional recovery. However, conventional orthopedic polymers often fail to achieve good bone integration and restoration of postoperative bone metabolic balance. There are many reasons for this phenomenon:

[0004] 1. Foreign body reaction caused by implants.

[0005] Although polymer implants have good biocompatibility and bioinertness, as non-host tissues, they will immediately cause a response from the body's immune system after implantation: immune cells are activated and gather around the implants, releasing reactive oxygen species (ROS) in an attempt to engulf and remove the implants. However, this engulfment is often unsuccessful, which in turn causes the accumulation of excessive reactive oxygen species and the release of a large number of cytokines, ultimately forming an excessive inflammatory response at the host-implant interface and causing damage to surrounding tissues, such as bone tissue absorption and scar tissue formation. What's more, in order to isolate and wrap the implant, the body will form a capsule composed of fibrous tissue around the implant to limit the contact between the implant and the tissue. The formation of this fibrous capsule will lead to a decrease in implant stability and limit its bone integration effect. Therefore, reducing the foreign body reaction caused by polymer implants is crucial to improving their functional stability in vivo.

[0006] 2. The implant’s microenvironment regulation ability is insufficient.

[0007] Patients who undergo implant surgery usually suffer from serious orthopedic diseases and have severe bone metabolism imbalances, which are usually accompanied by complex conditions such as inflammation, autoimmune diseases, and tumors, which often lead to an increase in the content of ROS in the pathological microenvironment. Implant surgery can help remove necrotic tissue, but it cannot completely reverse the development of the disease. In this case, conventional polymer implants alone cannot effectively regulate the microenvironment of the lesion area. Although postoperative drug management can be performed by using drugs such as bisphosphonates, nonsteroidal anti-inflammatory drugs, and immune antagonists, long-term systemic administration can cause serious side effects including kidney damage, poor digestive system, cardiovascular system damage, and malignant tumors, and increase the economic burden on patients. Therefore, modifying implants to enhance their ability to regulate the pathological microenvironment is a safe and economical design idea, which can achieve local immune regulation in the lesion area and reduce the frequency and dose of systemic administration. At present, there are studies on the formation of coatings on the surface of implants, and the loading and sustained release of bioactive substances such as cytokines, antioxidants, and anti-inflammatory drugs through the coating to improve the pathological microenvironment regulation ability of implants. However, the coating will degrade during use, and the bioactive substances carried will gradually dissipate over time, making it difficult to maintain long-term microenvironment regulation. Therefore, developing new modification methods to enable polymer implants to have long-term physiological microenvironment regulation capabilities will have a positive effect on restoring bone metabolism balance after surgery.

[0008] 3. Deterioration of the mechanical function of the implant.

[0009] According to the role of orthopedic polymers in bone tissue and the required mechanical properties, they can be divided into load-bearing polymer materials and non-load-bearing polymer materials. Load-bearing polymer materials are used to bear loads and are often used in bone repair and replacement surgery, such as artificial joint gaskets, bone fusion devices and bone plates. These materials usually have high strength, rigidity and wear resistance. However, load-bearing polymer implants will still have problems with mechanical property degradation under long-term load, which is manifested as reduced strength, reduced toughness, particle falling, and even cracks and fractures. The degradation of mechanical properties not only causes the implant to fail to function normally, but also poses serious health risks to patients. For example, after joint replacement surgery, about 10% of patients need to undergo revision surgery 10-20 years after surgery due to aseptic loosening. The particles dropped from the polymer liner under long-term load are the main cause of aseptic loosening. The dropped particles will activate the host's immune system, leading to a large accumulation of ROS, causing excessive activation of osteoclasts and bone dissolution around the implant, and ultimately leading to loosening of the prosthesis or even peripheral fractures. It can be seen that maintaining the long-term mechanical properties of polymer implants is an important means to ensure their long-term functional effects.

[0010] Summarizing the shortcomings of current orthopedic polymer implants, we can find that the ideal modification directions of orthopedic polymer implants are as follows: (1) reducing the foreign body reaction caused by implants; (2) long-term stable pathological microenvironment regulation ability; (3) long-term stable mechanical properties.

[0011] ROS is an oxygen metabolite involved in cell signaling and cell function regulation, but in pathological conditions such as inflammation, autoimmune diseases, tumors, foreign body implants, and infections, excessive ROS can lead to oxidative stress, which in turn exacerbates the disease. Extensive research has been conducted to eliminate excess ROS as a means of regulating inflammation and restoring bone metabolism balance. However, conventional antioxidants eliminate ROS in a sacrificial manner, and their effectiveness decreases over time. Although natural enzymes can efficiently remove ROS in a non-chemical dose-dependent manner, their instability limits their practical applications.

[0012] Doping with nanofillers is an effective method to enhance the mechanical properties of polymers. Nanomaterials can significantly improve the mechanical properties, thermal stability and wear resistance of polymers through high specific surface area effect, interface enhancement effect and nanosize effect. However, its enhancement effect is limited by the dispersion of nanofillers in the polymer matrix. Direct mechanical blending of nanofillers and polymers often causes agglomeration of nanofillers, which in turn reduces the mechanical properties of the polymer. Therefore, compared with traditional physical mixing, nanoparticles grown in situ in polymers usually have better dispersion and better mechanical enhancement effect.

[0013] Nanozymes are nanomaterials with enzyme-like catalytic properties, which have both enzyme-like active centers and the physical and chemical properties of nanomaterials themselves. Nanozymes have multiple enzyme activities including superoxide dismutase (SOD) and catalase (CAT), and have been widely used in antioxidant treatment of various disease models. At the same time, nanozymes are highly stable and controllable, and can be used in harsh working environments that many natural enzymes cannot tolerate.

[0014] The patent (application number: 2021104391305) discloses a nanozyme-modified bone implant material and its preparation method and application, wherein the nanozyme-modified bone implant material is obtained by modifying the bone implant material with nanozymes or nanozyme precursors (nanozymes can be prepared in situ). However, the modification of polyethylene in this patent remains at the raw material level, and no product processing and mechanical property testing is performed, so it is impossible to determine whether its modification method will affect the processing, molding and mechanical properties of the implant; the proposed polyethylene modification method is suitable for polyethylene with a molecular weight of 2 million, and its modification method cannot meet the ultra-high molecular weight polyethylene (molecular weight ≥ 5 million) actually used in clinical practice. The single-time output of the modification method proposed in this patent is at the milligram level, which is difficult to meet the amount of raw materials required for the actual product preparation. In addition, the modification method of the patent subjected the obtained modified polyethylene to liquid nitrogen cooling and crushing, which would reduce the molecular weight of polyethylene, further leading to a decrease in the mechanical properties of the finished product. Moreover, the modification method of the nanozyme-modified titanium alloy implant proposed in this patent is to perform nanozyme modification on the surface of the titanium alloy to form a coating. Since titanium alloy prostheses are often used in artificial joint implants to prepare articular surfaces, femoral head balls and other products, their anti-friction performance and mechanical aging are particularly important for the long-term stability of prostheses. However, the patent does not conduct relevant tests on the modified titanium alloy prostheses. In addition, in practical applications, with long-term reciprocating friction movement, the nanozyme coating may peel off and degrade, so the long-term stability of the nanozyme surface modification method may lag behind in-situ modification.

[0015] In summary, in view of the shortcomings of current orthopedic polymer implants, the use of nanozymes as modification media is a method that can simultaneously improve the mechanical properties of implants and their ability to regulate the physiological microenvironment. Summary of the invention

[0016] Purpose of the invention: In order to solve the shortcomings of existing polymer implants for orthopedics, the present invention proposes to improve the performance of polymer implants through nanozyme modification, specifically including: 1. reducing the foreign body reaction caused by the implant; 2. prolonging the maintenance time of the implant's physiological microenvironment regulation ability; 3. achieving the improvement of the mechanical properties of the implant and delaying the generation of wear particles; 4. reducing the immune response caused by wear particles.

[0017] The first object of the present invention is to provide a method for synthesizing nanozymes that grow in situ in polymers; the second object of the present invention is to provide a method for processing polymers after nanozyme modification; the third object of the present invention is to provide applications of nanozyme-modified polymer implants in improving orthopedic diseases.

[0018] Technical solution: To solve the above technical problems, the present invention provides a method for preparing a nanozyme-modified orthopedic polymer, comprising the following steps:

[0019] (1) using polymer powder to dissolve in a solvent A that assists the polymer in swelling and / or dissolving, and swelling and / or dissolving under heating,

[0020] (2) dispersing or dissolving one or more nanozymes or nanozyme precursors in a good solvent B solvent, and injecting the B solvent into the swollen and / or dissolved polymer,

[0021] (3) Solvent replacement is performed on the obtained nanozyme-modified polymer using solvent C that is immiscible with solvent A (dielectric constant difference of 30 or more), followed by washing and drying.

[0022] Wherein, the polymer powder in step (1) includes one or more of ultra-high molecular weight polyethylene, low molecular weight polyethylene, polypropylene, polyvinyl chloride, polytetrafluoroethylene, polystyrene, polyamide, polyetheretherketone, polylactic acid-co-glycolic acid, polylactic acid, polylactic acid glycolic acid, ethylene-vinyl acetate copolymer, polycarbonate, polyurethane, polycaprolactone, polylactic acid-co-caprolactone, polymethyl methacrylate or polyethylene terephthalate.

[0023] Preferably, the polymer powder is ultra-high molecular weight polyethylene (molecular weight ≥ 5 million).

[0024] Wherein, the solvent A in step (1) comprises one or more combinations of isopentane, n-pentane, decahydronaphthalene, petroleum ether, hexane, cyclohexane, isooctane, trifluoroacetic acid, trimethylpentane, cyclopentane, heptane, trichloroethylene, toluene, p-xylene, chlorobenzene, o-dichlorobenzene, benzene, isobutanol, dichloromethane, butanol, n-propanol, tetrahydrofuran, ethanol, ethyl acetate, petroleum ether, cyclohexane, chloroform, benzene, toluene, dichloromethane, ether, ethyl acetate, isopropanol, chloroform, pyridine, acetone, acetic acid, acetonitrile, aniline, dimethylformamide, methanol, ethylene glycol, dimethyl sulfoxide or water.

[0025] Preferably, the A solvent is decalin.

[0026] Among them, the nanozyme described in step (2) includes one or more combinations of carbon-based nanozymes, nitrogen compound nanozymes, oxygen compound nanozymes, metal nanozymes, metal-organic framework-based nanozymes, covalent organic framework-based nanozymes, metal-coordinated nanozymes or composite material nanozymes.

[0027] Furthermore, the carbon-based nanozyme includes one or more of fullerene-based nanozyme, carbon fiber-based nanozyme, carbon nanotube-based nanozyme, graphene-based nanozyme or other carbon-based nanozymes.

[0028] The nitrogen compound nanozyme includes one or more combinations of nitride-based nanozymes or phosphide-based nanozymes.

[0029] The oxygen compound nanozyme includes one or more combinations of oxide-based nanozymes, sulfide-based nanozymes, selenide-based nanozymes or telluride-based nanozymes.

[0030] The metal nanozyme includes one or more combinations of gold nanozyme, copper nanozyme, silver nanozyme, platinum nanozyme, palladium nanozyme, rhodium nanozyme, ruthenium nanozyme or alloy nanozyme.

[0031] The metal-organic framework-based nanozyme includes one or more of an iron-based metal-organic framework nanozyme, a zinc-based metal-organic framework nanozyme, a copper-based metal-organic framework nanozyme, a zirconium-based metal-organic framework nanozyme, a hafnium-based metal-organic framework nanozyme, a vanadium-based metal-organic framework nanozyme, a metal-doped metal-organic framework nanozyme or a carbon-based nanozyme.

[0032] The covalent organic framework nanozyme includes one or more combinations of boron-based covalent organic framework nanozymes, imine-based covalent organic framework nanozymes, triazine-based covalent organic framework nanozymes, or other types of covalent organic framework nanozymes.

[0033] Preferably, the nanozyme is an oxygen compound nanozyme; the oxygen compound nanozyme is an oxide-based nanozyme; the oxide-based nanozyme is cerium oxide.

[0034] Among them, the nanozyme contains metal elements including lithium, magnesium, aluminum, potassium, calcium, scandium, titanium, vanadium, chromium, manganese, iron, cobalt, nickel, copper, zinc, gallium, germanium, rubidium, strontium, iridium, zirconium, niobium, molybdenum, ruthenium, rhodium, palladium, silver, tin, antimony, cerium, hafnium, iridium, platinum, gold or bismuth. One or more combinations of elements.

[0035] Preferably, the nanozyme contains cerium.

[0036] Wherein, the nanozyme precursor includes one or more combinations of organic metal salts, metal chloride salts, metal bromide salts, metal nitrate salts, metal acetate salts, metal sulfate salts or metal cyanide complexes.

[0037] Preferably, the nanozyme precursor is an organic metal salt.

[0038] The organic metal salt includes one or more combinations of metal acetylacetonate, metal alkoxide, metal acid salt, metal ether salt, metal ether acid salt, metal ether alkoxide, metal amide salt, metal alcoholate, metal carboxylic acid amide salt, metal ether amide salt, metal imine salt, metal phosphorus complex or metal nitrogen complex.

[0039] Preferably, the organic metal salt is a metal acetylacetonate.

[0040] The metal acetylacetonate contains metal elements including one or more combinations of lithium, magnesium, aluminum, potassium, calcium, scandium, titanium, vanadium, chromium, manganese, iron, cobalt, nickel, copper, zinc, gallium, germanium, rubidium, strontium, iridium, zirconium, niobium, molybdenum, ruthenium, rhodium, palladium, silver, tin, antimony, cerium, hafnium, iridium, platinum, gold or bismuth.

[0041] Preferably, the metal acetylacetonate is cerium acetylacetonate.

[0042] Wherein, the good solvent B solvent for dispersing the nanozyme or nanozyme precursor in step (2) includes one or more combinations of ethanol, ethyl acetate, isopropanol, chloroform, pyridine, acetone, acetic acid, acetonitrile, aniline, dimethylformamide, methanol, ethylene glycol, dimethyl sulfoxide or water.

[0043] Preferably, the B solvent is ethanol.

[0044] Wherein, the solvent C used for solvent replacement in step (3) is immiscible with the solvent A, and the dielectric constants of the two solvents differ by more than 30.

[0045] Wherein, the C solvent includes one or more combinations of ethanol, ethyl acetate, isopropanol, chloroform, pyridine, acetone, acetic acid, acetonitrile, aniline, dimethylformamide, methanol, ethylene glycol, dimethyl sulfoxide or water.

[0046] Preferably, the C solvent is methanol.

[0047] In step (3), the nanozyme-modified polymer powder after solution replacement is filtered and washed with ethanol at least 10 times, and heated in a vacuum oven at 80° C. for 7 days to volatilize the residual solvent.

[0048] The invention also provides an orthopedic polymer prepared by the method.

[0049] The present invention also provides a method for preparing a modified polymer implant using the orthopedic polymer, comprising the following steps:

[0050] (1) mechanically shaping and morphologically processing the modified polymer to obtain a crude modified polymer implant;

[0051] (2) Processing the crude modified polymer implant to obtain a modified polymer implant.

[0052] Among them, the mechanical molding step of the nanozyme modified polymer in step (1) includes one or more combinations of hot pressing sintering, hot pressing molding, injection molding, blow molding, extrusion molding, calendering molding, 3D printing molding, vacuum molding or spray molding.

[0053] Preferably, the forming method is hot pressing forming.

[0054] The 3D printing method includes one or more combinations of fused deposition method, photo-curing method, laser sintering method, powder melting method or electron beam melting method.

[0055] Preferably, the 3D printing method is a fused deposition method.

[0056] The topography processing method in step (1) includes one or more combinations of cutting, grinding, drilling, milling or cutting.

[0057] Preferably, the machining method is milling.

[0058] The post-processing step in step (2) includes one or more combinations of cleaning, polishing, sandblasting, pickling, plasma treatment, solvent treatment, silane coupling agent treatment, chemical coating treatment or ion implantation treatment.

[0059] Preferably, the post-processing treatment is cleaning.

[0060] The invention also provides a modified polymer implant prepared by the method.

[0061] Among them, nanozyme-modified orthopedic polymer products include artificial joint gaskets, bone plates, bone nails, bone cement, bone scaffolds, bone fusion devices, bone fillers, bone fixation belts, artificial bones or periosteum.

[0062] Preferably, the processed and formed component is an artificial joint gasket.

[0063] The present invention also provides the use of the orthopedic polymer in preparing an orthopedic implant for treating, alleviating and / or preventing orthopedic diseases.

[0064] The orthopedic diseases include advanced arthritis, aseptic loosening, bacterial loosening, osteoporosis, bone tumors, repair of large bone defects, osteomyelitis or fractures.

[0065] Principle of the invention:

[0066] 1. The present invention injects a nanozyme precursor into the polymer after swelling to prepare a modified polymer. After the polymer swells, the intermolecular gap increases, which is conducive to the growth and dispersion of the nanozyme in the polymer. Compared with traditional physical mixing, the nanozyme obtained by in situ growth can achieve good dispersion and improved mechanical properties in the polymer matrix. In addition, the molten environment formed after the polymer swells delays the oxidation process during the crystallization of cerium oxide, which helps the formation of smaller nanozymes. Compared with the hydrothermal method for preparing nanozymes proposed in the patent (application number: 2021104391305), the nanozyme grown in situ in polyethylene at the same time has a smaller particle size, and a smaller particle size helps to achieve better catalytic performance.

[0067] 2. Considering that polymer implants need to exert antioxidant properties and bear mechanical loads during application, the present invention simultaneously utilizes nanozymes as nanomaterials with mechanical enhancement properties and enzyme-like catalytic properties to achieve multi-faceted improvements in polymer performance. Compared with the patent (application number: 2021104391305), the present invention is more comprehensive in the modification of polymers and closer to actual clinical applications.

[0068] 3. In the preparation process, the present invention uses swelling, solvent replacement and cooling crystallization to perform post-synthesis treatment on the modified polymer, so that it is transformed from a molten state to a state that can be used for mechanical processing. Compared with the patent (application number: 2021104391305), the present invention does not use vigorous stirring, liquid nitrogen crushing and other steps in the process of collecting and processing the modified polymer, and does not reduce the molecular weight of the polymer, which helps to maintain the mechanical properties of subsequent products.

[0069] 4. The present invention uses a non-polar solvent decahydronaphthalene to swell the polymer during the preparation process, which broadens the molecular weight range of the modified polyethylene and increases the single modified polymer output. Compared with the patent (application number: 2021104391305), it can meet the modification needs of high molecular weight polyethylene with a molecular weight of more than 5 million. In addition, large-scale preparation is conducive to practical processing applications.

[0070] 5. The present invention mechanically shapes and morphologically processes the obtained modified polymer raw materials to prepare implants of various shapes to meet the treatment needs under different pathological conditions. Compared with the patent (application number: 2021104391305), this patent prepares artificial joint gasket products, proving the feasibility of the polymer modification and subsequent processing methods.

[0071] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages:

[0072] 1. Reduce the dosage of postoperative medication, reduce adverse drug reactions and economic burden: The present invention uses nanozymes as the modification medium of the implant. Nanozymes themselves have the ability to catalyze multiple types of enzymes, and can efficiently catalyze and eliminate multiple ROS, thereby alleviating inflammation and improving the therapeutic effect of anti-inflammatory drugs. In addition, the intensity of foreign body reactions caused by nanozyme-modified implants is reduced, which also helps to reduce the dosage of postoperative drugs. By partially compensating for the effects of drugs and reducing stimulation to the body, nanozyme-modified polymer implants can help reduce the amount of postoperative drugs, avoid adverse reactions to long-term medications and the economic burden on patients;

[0073] 2. Enhance mechanical properties and reduce complications after implantation: The present invention grows nanozymes in situ in the polymer matrix and utilizes the physical and chemical properties of nanozymes as nanomaterials to mechanically enhance a variety of polymers, which helps to improve the mechanical stability of polymer implants. In addition, by reducing the shedding of implant particles, the occurrence of postoperative complications can be effectively inhibited and the service life of the prosthesis can be extended;

[0074] 3. Long-term stability of the modification effect and prolonged protection of the body: The present invention can achieve the modification of polymer materials at the macroscopic and overall level by in-situ growth of nanozymes in the polymer matrix. Under long-term mechanical action, the nanozymes in the polymer matrix can maintain the stability of structure and function. In addition, even if the modified polymer device has particle fragments falling, the fallen particles still contain nanozymes with catalytic activity, which can help alleviate the upregulation of ROS secretion caused by the particles and achieve long-term protection of the body's physiological environment;

[0075] 4. Avoid physiological toxicity: The present invention grows nanozymes in situ in the polymer. While giving the polymer matrix nanozyme activity, the polymer matrix also plays a role in protecting and fixing the nanozymes, which can ensure that the nanozymes can function at the implantation site for a long time. The risk of nanozyme particles falling off and being absorbed is avoided, nanotoxicity is reduced, and the safety of nanomaterials applied in physiological environments is improved;

[0076] 5. Adjustable design of multiple enzyme activities, personalized customization for diseases: Nanozymes have multiple types of enzyme activities including ROS scavenging activity, ROS generating activity, hydrolase activity, etc. Nanozymes themselves, as nanomaterials, can achieve the expression and reduction of target enzyme activities through a variety of synthetic methods. Therefore, ideal nanozymes can be designed according to the pathological environment and treatment needs of different diseases, and then applied to polymer modification to achieve personalized customization of implants;

[0077] 6. The modification method is simple and applicable to a variety of polymer matrices, making it easy to carry out mass preparation and industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0078] Figure 1 Joint prosthesis liners prepared from nanozyme-modified polymers;

[0079] Figure 2 Transmission electron microscope (TEM) images of cerium oxide grown in situ in ultra-high molecular weight polyethylene at different magnifications, with low magnification on the left and high magnification on the right;

[0080] Figure 3 X-ray diffraction (XRD) results of ultra-high molecular weight polyethylene modified with cerium oxide nanozyme before and after hot pressing;

[0081] Figure 4 The anti-oxidative aging properties of ultra-high molecular weight polyethylene modified with cerium oxide nanozymes;

[0082] Figure 5 The elongation at break results of ultra-high molecular weight polyethylene modified with cerium oxide nanozyme;

[0083] Figure 6 The tensile properties of ultra-high molecular weight polyethylene modified with cerium oxide nanozymes;

[0084] Figure 7 The impact performance results of ultra-high molecular weight polyethylene modified with cerium oxide nanozymes;

[0085] Figure 8 The friction coefficient results of ultra-high molecular weight polyethylene modified with cerium oxide nanozyme;

[0086] Fig. 9 The wear rate results of ultra-high molecular weight polyethylene modified with cerium oxide nanozymes;

[0087] Fig.10 The superoxide anion elimination results (SOD-like enzyme activity) of ultra-high molecular weight polyethylene modified with cerium oxide nanozymes;

[0088] Fig.11The hydrogen peroxide elimination effect (CAT enzyme-like activity) of ultra-high molecular weight polyethylene modified by cerium oxide nanozymes;

[0089] Fig.12 The hydroxyl radical elimination effect of ultra-high molecular weight polyethylene modified by cerium oxide nanozyme;

[0090] Fig.13 The results of ultra-high molecular weight polyethylene particles modified with cerium oxide nanozymes slowing down the production of ROS in macrophages;

[0091] Fig.14 The results of ceria nanozyme-modified ultra-high molecular weight polyethylene particles in reducing bone resorption in a skull osteolysis model;

[0092] Fig.15 The results of ceria nanozyme-modified ultra-high molecular weight polyethylene particles promoting osteogenesis in a skull osteolysis model;

[0093] Fig.16 The results of cerium oxide nanozyme-modified ultra-high molecular weight polyethylene particles inhibiting inflammation in a skull osteolysis model;

[0094] Fig.17 The results of cerium oxide nanozyme-modified ultra-high molecular weight polyethylene particles in reducing bone resorption in a femoral implant model;

[0095] Fig.18 The results show that cerium oxide nanozyme-modified ultra-high molecular weight polyethylene particles can inhibit foreign body reaction in a femoral implant model.

[0096] Fig.19 Results of ceria nanozyme-modified ultra-high molecular weight polyethylene particles in suppressing inflammation in a femoral implant model. DETAILED DESCRIPTION

[0097] The technical solution of the present invention is further described below in conjunction with the accompanying drawings.

[0098] Example 1 Synthesis and preparation of ultra-high molecular weight polyethylene modified with cerium oxide nanozyme

[0099] 1. Disperse 10 g of ultra-high molecular weight polyethylene (molecular weight 5.5 million) in 800 mL of decahydronaphthalene solvent to obtain a mixed system;

[0100] 2. The mixed system obtained in step 1 is heated to 120° C. under a nitrogen environment and continuously stirred until the ultra-high molecular weight polyethylene is completely swollen;

[0101] 3. Dissolve 1347.56 mg of cerium acetylacetonate in 20 mL of anhydrous ethanol to obtain a precursor solution;

[0102] 4. Slowly drop the precursor solution obtained in step 3 into the system obtained in step 2, and convert the atmosphere to air;

[0103] 5. Stir and heat the system obtained in step 4 for 24 hours;

[0104] 6. Pour the mixture obtained in step 5 into 0°C methanol for solvent replacement, and pour out the excess solvent after the mixture cools down;

[0105] 7. Filter and wash the mixture obtained in step 6 10 times with ethanol and dry it;

[0106] 8. Place the modified polymer obtained in step 7 in a vacuum oven at 80° C. to evaporate the residual solvent.

[0107] Example 2 Detection of Nanozyme Synthesis in Nanozyme-Modified Polyethylene

[0108] 1. XRD was used to detect the ultra-high molecular weight polyethylene modified by the nanozyme prepared in Example 1. The results are as follows: Figure 3 As shown. According to the comparison of the characteristic peak cards of polyethylene and cerium oxide at the bottom of the picture, it can be found that compared with polyethylene, the modified polyethylene not only has the characteristic peaks of polyethylene, but also shows obvious characteristic peaks of cerium oxide. This means that cerium oxide has been successfully grown in situ in the polyethylene matrix, and the structure of the polyethylene itself has not been affected.

[0109] 2. Disperse 10 mg of nanozyme-modified ultra-high molecular weight polyethylene in 10 mL of decalin and heat with stirring at 120 °C until the polyethylene swells;

[0110] 3. Take 50 μL of the mixed solution in step 2 and drop it into 1 mL of ethanol, shake to disperse;

[0111] 4. Drop the mixed solution obtained in step 3 onto a copper grid and observe it using TEM. The results are as follows: Figure 2 From the low-magnification TEM image on the left, it can be seen that the cerium oxide is granular and about 5 nm in size; from the high-magnification TEM image on the right, a lattice distance of 0.31 nm can be measured, which is a typical lattice distance of the cerium oxide (111) crystal plane, indicating that the cerium oxide grown in situ in polyethylene also has good crystallization.

[0112] Example 3 Nanoenzyme modified polyethylene molding process

[0113] 1. Place the nanozyme-modified polymer obtained in Example 1 in a hot pressing mold, and apply a pressure of 10 MPa to the mold at room temperature for compaction, and repeat this process multiple times until the polymer is compacted and fills the cavity;

[0114] 2. Place the loaded mold in a hot press, heat to 210°C and keep it warm and pressurized for 1 hour;

[0115] 3. After the heat preservation and pressure preservation are completed, the temperature is naturally lowered to room temperature under the condition of constant pressure, and then the polymer block is taken out;

[0116] 4. The hot pressed block was sliced ​​to obtain a thin sheet with a thickness of about 100 μm, and then the thin sheet was soaked in squalene and heated at high temperature for oxidation aging. The oxidation index of the polyethylene thin sheet and the modified polyethylene thin sheet after aging was monitored by Fourier infrared spectrometer. The results are as follows: Figure 4 After the same oxidative aging procedure, the modified polyethylene showed a lower oxidation index than the ordinary polyethylene, indicating that it has a stronger antioxidant effect.

[0117] 5. The hot pressed blocks were cut according to the national standard test standards and the relevant mechanical properties were tested. The tensile properties ( Figure 5 , Figure 6 )、Impact performance( Figure 7 )、Friction performance( Figure 8 ) and wear performance ( Fig. 9 ); Elongation at break results ( Figure 5 ) shows that the elongation at break of polyethylene does not decrease significantly compared with ordinary polyethylene, indicating that the in-situ growth of cerium oxide in polyethylene will not damage the tensile properties of polyethylene. In addition, the ultimate tensile strength of polyethylene is increased by in-situ growth of cerium oxide ( Figure 6 ) and impact strength ( Figure 7 ), which is particularly important for the clinical application of modified polyethylene. Using titanium alloy balls as friction pairs, the friction properties of ordinary polyethylene and modified polyethylene were tested by friction and wear tester ( Figure 8 ) and wear performance ( Fig. 9 ), the friction coefficient of the modified polyethylene was significantly reduced, and the volume of particles dropped was less under the same wear time, indicating that the wear resistance of polyethylene was enhanced by in-situ growth of cerium oxide in polyethylene.

[0118] 6. Perform CNC machining on the polymer block obtained in step 3 to prepare an artificial joint gasket model such as Figure 1 shown.

[0119] Example 4 Measurement of enzyme-like activity of nanoenzyme-modified polyethylene

[0120] 1. Use liquid nitrogen to cool down the ultra-high molecular weight polyethylene modified with nanozymes;

[0121] 2. Use a planetary ball mill to crush the cooling polyethylene obtained in step 1, with an oscillation frequency of 50 Hz, 30 s each time, 10 times as a cycle, and a total of 10 cycles;

[0122] 3. Wash and dry the crushed polymer particles several times and disperse them in PBS containing 0.05% polyoxypropylene oxyethylene glycerol ether for later use;

[0123] 4. The superoxide anion scavenging activity of the polymer particles was tested using the nitro blue tetrazolium chloride method. The results are as follows: Fig.10 As shown. Compared with ordinary polyethylene particles, the polyethylene particles modified with nanozymes showed a higher superoxide anion inhibition rate;

[0124] 5. Use oxygen electrodes to detect the hydrogen peroxide scavenging activity of polymer particles. Nanozyme-modified polymer particles scavenged hydrogen peroxide by catalytically decomposing hydrogen peroxide into oxygen and water. Oxygen electrodes measured the hydrogen peroxide scavenging ability of different particles by monitoring the dissolved oxygen concentration in the liquid environment. The results are as follows: Fig.11 Compared with ordinary polyethylene particles, the polyethylene particles modified with nanozymes showed an increase in dissolved oxygen production over time, indicating that they have significant hydrogen peroxide scavenging ability;

[0125] 6. The hydroxyl radical scavenging activity of the polymer particles was detected using electron paramagnetic resonance spectroscopy. 5,5-dimethyl-1-pyrroline-N-oxide was used as a hydroxyl radical scavenger. The strength of the electron paramagnetic resonance spectroscopy signal was used to measure the amount of hydroxyl radicals. The results are as follows: Fig.12 Compared with ordinary polyethylene particles, the polyethylene particles modified by nanozymes have obvious hydroxyl radical scavenging effect, and the characteristic peak of hydroxyl radicals has decreased significantly.

[0126] Example 5 Stimulation of macrophages by nanoenzyme-modified polymer particles

[0127] 1. 3×10 5 RAW264.7 macrophages were seeded in 12-well plates at a concentration of 10 cells / well and grown for 24 h;

[0128] 2. Disperse the nanoenzyme-modified polyethylene particles and ordinary polyethylene particles in the cell culture medium at concentrations of 0.2 mg / mL, 0.4 mg / mL, and 0.8 mg / mL, respectively;

[0129] 3. Pour the mixed culture medium obtained in step 2 into the 12-well plate in step 1, seal the wells with sealing film after filling them, and then turn the well plate over to allow the polyethylene particles and modified polyethylene particles to float up and contact with the macrophages, and incubate for 24 hours;

[0130] 4. Pour out the culture medium, wash the cells with PBS buffer, add PBS containing 10 μM 2,7-dichlorofluorescein diacetate probe, and incubate in a cell culture incubator in the dark for 40 minutes;

[0131] 5. After 40 minutes, aspirate the PBS containing the probe and wash the cells three times with PBS by shaking. Then, blow down the cells and collect them in a centrifuge tube and store them in the dark.

[0132] 6. Use flow cytometry to monitor the ROS content in macrophages. The results are as follows: Fig.13 As shown in the figure, the relative fluorescence intensity of the reactive oxygen species probe caused by polyethylene particles stimulating macrophages increases with the increase of particle concentration, and is much higher than the relative fluorescence intensity caused by nanozyme-modified polyethylene particles, indicating that nanozyme-modified polyethylene particles cause less reactive oxygen species secretion in macrophages.

[0133] Example 6 Physiological properties of nanozyme-modified polymer particles in a skull bone dissolution model

[0134] 1. ICR mice (Weitongda Biotechnology Co., Ltd.) were divided into a control group, a polyethylene group, and a modified polyethylene group. The control group consisted of healthy mice, and the polyethylene group and the modified polyethylene group were injected with different particles. 50 mg of polyethylene particles and modified polyethylene particles were injected into the middle suture of the ICR mouse skull, sutured, and samples were collected 14 days later;

[0135] 2. Test the tissues, including Micro-CT, Masoon staining, and interleukin 1-β staining of the skull;

[0136] 3. Perform statistical analysis on the test results, including relative bone volume ( Fig.14 ), relative bone formation surface ( Fig.15 ) and relative interleukin-1-β fluorescence intensity ( Fig.16 Compared with the control group of mice, polyethylene particle stimulation caused the relative bone volume fraction of mice ( Fig.14 ), relative bone formation surface ( Fig.15 ), indicating that polyethylene stimulation aggravated osteoclast resorption and inhibited new bone formation, which was alleviated in the modified polyethylene group. Fig.16 ) It was found that mice in the polyethylene group showed the highest interleukin-1β secretion and their inflammation levels were enhanced compared with the control group and modified polyethylene group.

[0137] Example 7 Physiological properties of nanoenzyme-modified polymer particles in a femoral implantation model

[0138] 1. An incision was made at the left knee joint of ICR mice (Weitongda Biotechnology Co., Ltd.), and a Kirschner wire with a diameter of 0.8 mm and a length of 5 mm was implanted at the distal end of the femur;

[0139] 2. The mice were divided into a control group, a polyethylene group, and a modified polyethylene group. The mice in the control group were no longer injected with particles, but only implanted with Kirschner wires. For the mice in the polyethylene group and the modified polyethylene group, 50 mg of ordinary polyethylene particles and modified polymer particles were respectively injected into the distal femoral knee joint, and sutured. The samples were collected 42 days later;

[0140] 3. Test the tissues, including Micro-CT, Masoon staining, and interleukin 1-β staining of the skull;

[0141] 4. Statistical analysis of the test results, including relative bone volume ( Fig.17 ), fiber layer thickness ( Fig.18 ) and relative interleukin-1-β fluorescence intensity ( Fig.19 Compared with mice in the control group that were implanted with only Kirschner wires, the relative bone volume of mice stimulated with polyethylene particles decreased ( Fig.17 ), indicating that the bone formation ability of mice was reduced after polyethylene particle stimulation, while this situation was improved in the modified polyethylene particle group. The fiber layer thickness statistics showed that the fiber layer around the metal implants of the polyethylene particle group mice was thicker than that of the control group mice and the modified polyethylene group mice ( Fig.18 ), indicating that polyethylene particles aggravate the foreign body reaction caused by metal implants, which is not conducive to the long-term stability of implants. Statistics of interleukin-1β fluorescence intensity found that the secretion of interleukin-1β in the bone tissue area of ​​mice stimulated by polyethylene particles increased ( Fig.19 ), indicating a severe inflammatory response.

[0142] In summary, using a method for modifying a polymer using nanozymes provided by the present invention, the resulting polymer can be used to prepare an orthopedic implant, which has enhanced mechanical properties and antioxidant enzyme activity. The prepared implant can achieve the reduction of foreign body reaction after implantation, the elimination of inflammation in the lesion area, and the promotion of bone integration by eliminating ROS. In addition, nanozymes exhibit long-term structural and functional stability in the polymer matrix, which can not only ensure the long-term maintenance of the prosthetic function, but also avoid the nanotoxicity of the nanomaterial itself. Through this new nanozyme-modified polymer synthesis method, implants with both bone microenvironment regulation and mechanical property enhancement can be prepared, and have the potential for batch preparation and industrial production.

Claims

1. A method for preparing a nanozyme-modified orthopedic polymer, characterized in that: The following steps are involved: (1) Dissolve ultra-high molecular weight polyethylene with a molecular weight of 5.5 million in decalin and swell it under heating; (2) dispersing or dissolving one or more nanozymes or nanozyme precursors in a good solvent B solvent, and injecting the B solvent into the swollen and / or dissolved polymer; (3) using a C solvent that is immiscible with decahydronaphthalene to perform solvent replacement on the obtained nanozyme-modified polymer, and then washing and drying to obtain the nanozyme-modified orthopedic polymer; the dielectric constants of the decahydronaphthalene and the C solvent differ by more than 30.

2. The method according to claim 1, characterized in that The nanozyme described in step (2) includes one or more of oxide-based nanozymes, sulfide-based nanozymes, selenide-based nanozymes, telluride-based nanozymes, nitride-based nanozymes, phosphide-based nanozymes, gold nanozymes, copper nanozymes, silver nanozymes, platinum nanozymes, palladium nanozymes, rhodium nanozymes, ruthenium nanozymes, alloy nanozymes, iron-based metal organic framework nanozymes, zinc-based metal organic framework nanozymes, copper-based metal organic framework nanozymes, zirconium-based metal organic framework nanozymes, hafnium-based metal organic framework nanozymes, vanadium-based metal organic framework nanozymes, metal-doped metal organic framework nanozymes, boron-based covalent organic framework nanozymes, imine-based covalent organic framework nanozymes, triazine-based covalent organic framework nanozymes, fullerene-based nanozymes, carbon fiber-based nanozymes, carbon nanotube-based nanozymes, graphene-based nanozymes or other nanozymes.

3. The method according to claim 2, characterized in that The oxide-based nanozyme contains metal elements, and the metal elements include one or more of lithium, magnesium, aluminum, potassium, calcium, scandium, titanium, vanadium, chromium, manganese, iron, cobalt, nickel, copper, zinc, gallium, germanium, rubidium, strontium, iridium, zirconium, niobium, molybdenum, ruthenium, rhodium, palladium, silver, tin, antimony, cerium, hafnium, platinum, gold or bismuth.

4. The method according to claim 1, characterized in that: The nanozyme precursor in step (2) includes one or more of an organic metal salt, a metal chloride salt, a metal bromide salt, a metal nitrate salt, a metal acetate salt, a metal sulfate salt or a metal cyanide complex.

5. The method according to claim 4, characterized in that The organic metal salt includes one or more of metal acetylacetonate, metal alkoxide, metal acid salt, metal ether salt, metal ether acid salt, metal ether alkoxide, metal amide salt, metal alkoxide, metal carboxylic acid amide salt, metal ether amide salt, metal imine salt, metal phosphorus complex or metal nitrogen complex.

6. The method according to claim 5, characterized in that The metal acetylacetonate contains metal elements including one or more of lithium, magnesium, aluminum, potassium, calcium, scandium, titanium, vanadium, chromium, manganese, iron, cobalt, nickel, copper, zinc, gallium, germanium, rubidium, strontium, iridium, zirconium, niobium, molybdenum, ruthenium, rhodium, palladium, silver, tin, antimony, cerium, hafnium, platinum, gold or bismuth.

7. The method according to claim 1, characterized in that The good solvent B solvent for the nanozyme and nanozyme precursor in step (2) includes one or more of ethanol, ethyl acetate, isopropanol, chloroform, pyridine, acetone, acetic acid, acetonitrile, aniline, dimethylformamide, methanol, ethylene glycol, dimethyl sulfoxide or water.

8. The method according to claim 1, characterized in that The C solvent incompatible with decalin in step (3) includes one or more of ethanol, ethyl acetate, isopropanol, chloroform, pyridine, acetone, acetic acid, acetonitrile, aniline, dimethylformamide, methanol, ethylene glycol, dimethyl sulfoxide or water; the cleaning and drying method includes filtering and washing the nanozyme-modified polymer after solution replacement with ethanol multiple times, and volatilizing the residual solvent in a vacuum oven.

9. A method for preparing a nanozyme-modified orthopedic polymer, characterized in that: The following steps are involved: (1) dissolving ultra-high molecular weight polyethylene with a molecular weight of 5.5 million in decalin and swelling it under heating to obtain a mixed system; (2) dissolving cerium acetylacetonate in anhydrous ethanol, adding the solution dropwise to the mixed system in step (1), converting the atmosphere to air, stirring and heating to obtain a nanozyme-modified polymer; (3) Using methanol to replace the solvent of the nanozyme-modified polymer in step (2), and then washing and drying to obtain the nanozyme-modified orthopedic polymer.

10. An orthopedic polymer prepared by the method according to any one of claims 1 to 9.

11. A method for preparing a modified polymer implant using the orthopedic polymer according to claim 10, characterized in that: The following steps are involved: (1) Mechanically shaping and morphologically processing the modified polymer to obtain a crude modified polymer implant; (2) post-processing the crude modified polymer implant to obtain a modified polymer implant; the mechanical forming method in step (1) includes one or more combinations of hot pressing sintering, hot pressing molding, injection molding, blow molding, extrusion molding, calendering molding, 3D printing molding, vacuum molding or spray molding.

12. The method according to claim 11, characterized in that: The 3D printing process includes one or more of a fused deposition method, a photo-curing method, a laser sintering method, a powder melting method or an electron beam melting method.

13. The method according to claim 11, characterized in that: The topography processing method in step (1) includes one or more of cutting, grinding, drilling, milling or cutting.

14. The method according to claim 11, characterized in that The post-processing step in step (2) includes one or more of cleaning, polishing, sandblasting, pickling, plasma treatment, solvent treatment, silane coupling agent treatment, chemical coating treatment or ion implantation treatment.

15. A modified polymer implant prepared by the method according to any one of claims 11 to 14.

16. The modified polymer implant according to claim 15, characterized in that The modified polymer implant includes an artificial joint spacer, a bone plate, a bone nail, a bone cement, a bone scaffold, a bone fusion device, a bone filler, a bone fixation belt, an artificial bone or a periosteum.

17. Use of the orthopedic polymer according to claim 10 in the preparation of an orthopedic implant for treating, alleviating and / or preventing orthopedic diseases, characterized in that: The orthopedic diseases include advanced arthritis, aseptic loosening, bacterial loosening, osteoporosis, bone tumors, repair of large bone defects, osteomyelitis or fractures.

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