A method of surface modification of a biomaterial and a modified pericardial material

Abstract

This invention discloses a method for surface-modified biomaterials and a modified pericardial material, relating to the field of biomedical materials technology. The method includes the following steps: S1, providing animal pericardial material cross-linked with glutaraldehyde; providing a bridging molecule containing a first bridging group and a second bridging group, and a phosphorylcholine polymer with a third bridging group at its end; binding the first bridging group to an aldehyde group on the surface of the pericardial material via a first covalent reaction; binding the second bridging group to the third bridging group via a second covalent reaction, thereby achieving grafting of the phosphorylcholine polymer onto the material surface; the first covalent reaction and / or the second covalent reaction include Schiff base formation and reductive amination processes. The modified pericardial material prepared in this application introduces a phosphorylcholine biomimetic coating, which can improve blood compatibility on the material surface, reduce activation of the coagulation system and platelet adhesion, and inhibit the initiation of calcium ion deposition on the material surface, while exhibiting high safety.

Description

Technical Field

[0001] This invention relates to the field of biomedical materials technology, and in particular to a method for surface-modified biomaterials and modified pericardial materials. Background Technology

[0002] Artificial bio-heart valves, as key medical devices for treating valvular heart disease, have been widely used in clinical practice due to their excellent biocompatibility and hemodynamic properties. Among them, bovine pericardium and porcine pericardium, with their superior mechanical properties, suitable collagen fiber structure, and good processability, have become the mainstream natural biomaterials for preparing artificial bio-heart valves. Their performance directly determines the implantation effect and service life of the artificial valve.

[0003] To enhance the mechanical strength, structural stability, and biodegradability of bovine and porcine pericardial materials and meet the long-term use requirements for clinical implantation, glutaraldehyde (GA) crosslinking is currently the standard processing technology widely adopted in the industry. Glutaraldehyde crosslinks with the amino groups of collagen fibers in pericardial materials to form a stable covalent network, thereby enhancing the material's mechanical load-bearing capacity and resistance to enzymatic degradation, extending the service life of the implanted valve. This process has long dominated the industrial production of artificial biological heart valves due to its advantages such as ease of operation, controllable cost, and significant crosslinking effect. However, glutaraldehyde crosslinking treatment has two inherent and difficult-to-overcome technical defects that severely limit the clinical application efficacy and safety of artificial biological heart valves:

[0004] Firstly, there are the issues of toxicity and calcification caused by residual aldehyde groups. After the glutaraldehyde cross-linking reaction, a certain amount of free aldehyde groups inevitably remain on the surface of the pericardial material. These residual aldehyde groups are highly cytotoxic and will continuously stimulate surrounding tissues and cause chronic inflammatory reactions after implantation. More importantly, the residual aldehyde groups can act as initiation sites for pathological calcification, inducing calcium ions to deposit on the material surface and form calcified nodules, leading to valve stiffness, mechanical performance deterioration, and ultimately valve failure. This is one of the core reasons for mid-term failure of artificial biological heart valves after implantation.

[0005] Secondly, there is insufficient blood compatibility due to thrombosis. Even after glutaraldehyde cross-linking modification, the blood compatibility of the bovine and porcine pericardial surfaces is not ideal. The chemical properties and microstructure of the material surface easily activate the human coagulation system, inducing platelet adhesion, aggregation, and thrombus formation. This can not only block the valve passages and affect hemodynamics but also potentially cause serious complications such as thromboembolism. Therefore, patients who receive this type of valve implantation need to take anticoagulants post-surgery. However, anticoagulant therapy not only increases the patient's medication burden but may also increase the risk of bleeding, significantly impacting the patient's quality of life and treatment safety.

[0006] In summary, while existing processing techniques for natural pericardial materials based on glutaraldehyde crosslinking can meet the basic mechanical performance requirements of artificial biological heart valves, the calcification failure caused by residual aldehyde toxicity and the insufficient blood compatibility due to thrombosis have become key bottlenecks restricting the safety and long-term effectiveness of artificial biological heart valves in clinical applications. Therefore, developing a modification technology that can address these inherent defects while retaining the excellent mechanical properties of natural pericardial materials is of great significance for improving the clinical application value of artificial biological heart valves and is also a technical problem that urgently needs to be solved in this field. Summary of the Invention

[0007] This invention discloses a method for surface-modified biomaterials and a modified pericardial material, in order to solve the aforementioned technical problems existing in the processing technology of natural pericardial materials crosslinked with glutaraldehyde in related technologies.

[0008] To solve the above problems, the present invention adopts the following technical solution:

[0009] In a first aspect, embodiments of this application provide a method for surface-modified biomaterials, comprising the following steps:

[0010] Animal pericardial material treated with glutaraldehyde is provided, with residual active aldehyde groups on its surface;

[0011] A bridging molecule comprising a first primary amino group and a second primary amino group is provided.

[0012] Provides phosphorylcholine polymers with aldehyde-terminated ends;

[0013] The first primary amino group is combined with the aldehyde group on the surface of the pericardial material through a first covalent reaction. The first covalent reaction includes: immersing the animal pericardial material in a bridging molecule solution to react and form a first Schiff base bond, then performing a first wash, then placing it in a first reducing agent solution to reduce the first Schiff base bond to a first secondary amine bond, and then performing a second wash.

[0014] The second primary amino group is bonded to the aldehyde group at the end of the phosphorylcholine polymer through a second covalent reaction. The second covalent reaction includes: immersing the animal pericardial material treated with the first covalent reaction in a phosphorylcholine polymer solution to react and form a second Schiff base bond, and then placing it in a second reducing agent solution to reduce the second Schiff base bond to a second secondary amine bond.

[0015] This enables the grafting of phosphorylcholine polymers onto the surface of animal pericardial materials.

[0016] Secondly, embodiments of this application provide a modified pericardial material prepared by the above method.

[0017] The technical solutions adopted in the embodiments of the present invention can achieve the following beneficial effects:

[0018] (1) The modified pericardial material prepared by the method of surface modification of biomaterials provided in the embodiments of this application is prepared by grafting phosphorylcholine polymer onto the surface of pericardial material after glutaraldehyde crosslinking. By taking advantage of the biomimetic properties of phosphorylcholine groups, a phosphorylcholine biomimetic coating is introduced. On the one hand, it can improve the blood compatibility of the material surface and reduce the activation of the coagulation system and platelet adhesion. On the other hand, it can inhibit the initiation of calcium ion deposition on the material surface. From the mechanism of action, it addresses the two core problems of glutaraldehyde crosslinked pericardial material being prone to thrombosis and calcification, and achieves simultaneous improvement of anticoagulation and anticalcification performance.

[0019] (2) The modified pericardial material prepared by the method of surface modification of biomaterials provided in the embodiments of this application firstly forms covalent connections with the pericardial material and phosphorylcholine polymer through bridging molecules, and then reduces the generated Schiff base bond to a secondary amine bond, and finally forms a stable covalent bond connection structure between "animal pericardial material-bridging molecule-phosphorylcholine polymer", which avoids the situation of phosphorylcholine biomimetic coating falling off due to weak connection during long-term service in the body, and ensures that the material performance continues to be performed.

[0020] (3) The modified pericardial material prepared by the method of surface modification of biomaterials provided in the embodiments of this application uses bridging molecules containing bis-primary amino groups for directional connection, which can guide the phosphorylcholine polymer to be precisely grafted onto the surface of the animal pericardial material, avoiding the problem that the active groups of phosphorylcholine may be masked due to random grafting, ensuring the effective performance of the biomimetic function of phosphorylcholine polymer, ensuring the high activity of phosphorylcholine biomimetic coating, and making the anticoagulation and anticalcification properties of pericardial material stable and reliable.

[0021] (4) The modified pericardial material prepared by the method of surface modification of biomaterials provided in the embodiments of this application can not only play an anticoagulant role by grafting phosphorylcholine polymer, but also eliminate the free aldehyde groups remaining on the surface of the pericardial material through the reaction process. This solves the problem of toxic stimulation of residual aldehyde groups, further optimizes the biocompatibility of the material, reduces the risk of inflammatory response after the pericardial material is implanted into the human body, and ensures the safety of the final pericardial material when used in artificial heart valves. Attached Figure Description

[0022] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0023] Figure 1 This is a flowchart of the preparation process of Examples 1-11 of this application;

[0024] Figure 2 This is a schematic diagram illustrating the preparation principle of Examples 1-11 of this application;

[0025] Figure 3 This is a comparison chart of the anticoagulation test results of Examples 1-2 and Comparative Example 1 of this application;

[0026] Figure 4 This is a comparison chart of the qualitative results of the amount of calcium attached to the body in the anti-calcification experimental bodies of Examples 1-2 and Comparative Example 1 of this application. Detailed Implementation

[0027] To make the objectives, technical solutions, and advantages of this invention clearer, the technical solutions of this invention will be described in detail below. Obviously, the described embodiments are merely some embodiments of this invention, and not all embodiments. Based on the embodiments of this invention, all other implementation methods obtained by those skilled in the art without creative effort are within the scope of protection of this invention.

[0028] Collection, handling, storage and transportation of fresh beef and pork heart capsules: They can generally be collected and harvested from slaughterhouses that comply with GB 12694-2016 "Hygienic Standards for Livestock and Poultry Slaughtering and Processing" and are registered with the Ministry of Agriculture and Rural Affairs and the State Food and Drug Administration, and in accordance with the procedures described in YY / T 0287-2017, GB / T 44353.1-2024, GB / T 44353.2-2024, YY-T0771.3-2009, and YY-T0771.4 2015.

[0029] Fresh beef or pork pericardium obtained from local slaughterhouses was immediately placed in refrigerated phosphate-buffered saline (PBS, 0.1 M, pH 7.4) at a temperature between 2°C and 8°C and then immediately transferred to the laboratory for further processing.

[0030] Please see Figure 1 and Figure 2 :

[0031] I. Example:

[0032] Example 1:

[0033] A method for surface-modifying pericardial materials includes the following steps:

[0034] S1. Provides glutaraldehyde-crosslinked calf heart material:

[0035] In a clean environment at 15-25℃, connective tissue, fat, blood vessels, mucosa, fascia, and visible foreign matter adhering to fresh bovine pericardium were removed within 2 hours after collection. During the cleaning process, the pericardium was continuously rinsed with sterile saline. The cleaned and rinsed bovine pericardium material was then immersed in glutaraldehyde phosphate buffer solution with a pH of 7.4 and a mass percentage concentration of 0.8% for crosslinking at room temperature for 72 hours. After that, it was thoroughly washed with sterile saline to obtain bovine pericardium material crosslinked with glutaraldehyde.

[0036] S2, grafting of phosphorylcholine polymers; specifically including the following sub-steps:

[0037] S21. Bridging treatment: The glutaraldehyde-crosslinked bovine pericardium material was immersed in a 0.1M hexamethylenediamine aqueous solution at pH 6.5, ensuring complete immersion. The reaction was carried out at 37°C for 24 hours, allowing a primary amino group of hexamethylenediamine to react with the residual aldehyde group on the surface of the bovine pericardium material to form a Schiff base bond. Then, the material was thoroughly washed with sterile physiological saline to remove the physically adsorbed hexamethylenediamine.

[0038] S22, First Reduction: The bovine pericardium material treated in step S21 is transferred to a 0.1 M sodium cyanoborohydride solution with a pH of 3.5 and a concentration of 0.1 M for immersion, ensuring complete submersion in the sodium cyanoborohydride solution. The reaction is carried out at room temperature with shaking for 4 hours. The Schiff base bond generated in step S21 is reduced to a stable secondary amine bond. Then, it is thoroughly washed with a 0.1 M sodium cyanoborohydride solution.

[0039] S23. Phosphorylcholine polymer grafting: The pericardium material treated in step S22 is immersed in an aqueous solution of aldehyde-terminated poly(2-methacryloyloxyethyl) phosphorylcholine homopolymer (PC-CHO, Mn=10000g / mol) with a concentration of 10mg / mL, ensuring complete immersion in the aqueous solution of the polymer. The reaction is carried out at 37℃ for 24h, so that the aldehyde group of the phosphorylcholine polymer reacts with another primary amino group of the bridging molecule to form a Schiff base bond.

[0040] S24. Second reduction: The bovine pericardial material treated in step S23 is transferred to a 0.1 M sodium cyanoborohydride solution with a pH of 3.5 and is immersed completely in the sodium cyanoborohydride solution. The reaction is carried out at room temperature with shaking for 6 hours. The Schiff base bonds generated in step S23 are reduced to stable secondary amine bonds. Then, the material is thoroughly washed with sterile physiological saline to obtain the modified pericardial material with surface covalently grafted phosphorylcholine polymer.

[0041] Example 2:

[0042] A method for surface-modifying pericardial materials includes the following steps:

[0043] S1. Provides glutaraldehyde-crosslinked calf heart material;

[0044] In a clean environment at 15-25℃, connective tissue, fat, blood vessels, mucosa, fascia, and visible foreign matter adhering to fresh bovine pericardium were removed within 2 hours after collection. During the cleaning process, the pericardium was continuously rinsed with sterile saline. The cleaned and rinsed bovine pericardium material was then immersed in glutaraldehyde phosphate buffer solution with a pH of 7.4 and a mass percentage concentration of 0.8% for crosslinking at room temperature for 72 hours. After that, it was thoroughly washed with sterile saline to obtain bovine pericardium material crosslinked with glutaraldehyde.

[0045] S2, grafting of phosphorylcholine polymers; specifically including the following sub-steps:

[0046] S21. Bridging treatment: The glutaraldehyde-crosslinked bovine pericardium material was immersed in an L-lysine aqueous solution with a pH of 7.4 and a concentration of 0.1 M, and reacted at 37°C for 48 hours to allow a primary amino group of the bridging molecule to react with the residual aldehyde group on the surface of the bovine pericardium material to form a Schiff base bond; then, it was thoroughly washed with sterile physiological saline to remove the physically adsorbed L-lysine.

[0047] S22, First reduction: The bovine pericardium material treated in step S21 is transferred into a 0.1 M sodium cyanoborohydride solution with a pH of 4.0 and shaken at room temperature for 6 hours. The Schiff base bond generated in step S21 is reduced to a stable secondary amine bond. Then, it is thoroughly washed with a 0.1 M sodium cyanoborohydride solution.

[0048] S23. Phosphorylcholine polymer grafting: The pericardium material treated in step S22 is immersed in an aqueous solution of aldehyde-terminated poly(2-methacryloyloxyethyl phosphorylcholine) (PC-CHO, Mn=10,000 g / mol) with a concentration of 10 mg / mL and reacted at 37℃ for 24 h, so that the aldehyde group of the phosphorylcholine polymer reacts with another primary amino group of the bridging molecule to form a Schiff base bond.

[0049] S24. Second reduction: The bovine pericardial material treated in step S23 is transferred into a sodium cyanoborohydride solution with a pH of 4.0 and a concentration of 0.1 M. The reaction is carried out with shaking at room temperature for 6 hours. The Schiff base bond generated in step S23 is reduced to a stable secondary amine bond. Then, the material is thoroughly washed with sterile physiological saline to obtain the modified pericardial material with surface covalently grafted phosphorylcholine polymer.

[0050] Example 3:

[0051] The difference between this embodiment and Example 1 is that the bridging molecule used is a 0.01M aqueous solution of hexamethylenediamine.

[0052] Everything else is the same as in Example 1;

[0053] A modified pericardial material with surface covalently grafted phosphorylcholine polymer was obtained.

[0054] Example 4:

[0055] The difference between this embodiment and Example 1 is that the bridging molecule used is a 2M aqueous solution of hexamethylenediamine.

[0056] Everything else is the same as in Example 1;

[0057] A modified pericardial material with surface covalently grafted phosphorylcholine polymer was obtained.

[0058] Example 5:

[0059] The difference between this embodiment and Example 1 is that the bridging molecule used is a 0.1M aqueous solution of ethylenediamine.

[0060] Everything else is the same as in Example 1;

[0061] A modified pericardial material with surface covalently grafted phosphorylcholine polymer was obtained.

[0062] Example 6:

[0063] The difference between this embodiment and Example 1 is that the bridging molecule used is a 0.1M aqueous solution of 1,3-propanediamine.

[0064] Everything else is the same as in Example 1;

[0065] A modified pericardial material with surface covalently grafted phosphorylcholine polymer was obtained.

[0066] Example 7:

[0067] The difference between this embodiment and Example 1 is that the bridging molecule used is a 0.1M aqueous solution of 1,4-butanediamine.

[0068] Everything else is the same as in Example 1;

[0069] A modified pericardial material with surface covalently grafted phosphorylcholine polymer was obtained.

[0070] Example 8:

[0071] The difference between this embodiment and Embodiment 1 is that the animal pericardium material used is: fresh pig pericardium;

[0072] Everything else is the same as in Example 1;

[0073] A modified pericardial material with surface covalently grafted phosphorylcholine polymer was obtained.

[0074] Example 9:

[0075] A method for surface-modifying pericardial materials includes the following steps:

[0076] S1. Provides glutaraldehyde-crosslinked calf heart material;

[0077] In a clean environment at 15-25℃, connective tissue, fat, blood vessels, mucosa, fascia, and visible foreign matter adhering to fresh bovine pericardium were removed within 2 hours after collection. During the cleaning process, the pericardium was continuously rinsed with sterile saline. The cleaned and rinsed bovine pericardium was then immersed in glutaraldehyde phosphate buffer solution with a pH of 7.4 and a mass percentage concentration of 0.8% for crosslinking at room temperature for 72 hours. After that, it was thoroughly washed with sterile saline to obtain bovine pericardium treated with glutaraldehyde.

[0078] S2, grafting of phosphorylcholine polymers; specifically including the following sub-steps:

[0079] S21. Bridging treatment: The glutaraldehyde-crosslinked bovine pericardium material was immersed in an aqueous solution of ethylenediamine at pH 7.0 and a concentration of 1.5 M, ensuring complete immersion. The reaction was carried out at 10°C for 72 hours, allowing a primary amino group of the bridging molecule to react with the residual aldehyde group on the surface of the bovine pericardium material to form a Schiff base bond. Then, the material was thoroughly washed with sterile physiological saline to remove the physically adsorbed ethylenediamine.

[0080] S22, First reduction: The bovine pericardium material treated in step S21 is transferred to a 1.5 M sodium cyanoborohydride solution with a pH of 3.0 and is immersed in the solution, ensuring complete immersion. The reaction is carried out at room temperature with shaking for 72 hours. The Schiff base bond generated in step S21 is reduced to a stable secondary amine bond. Then, it is thoroughly washed with a 1.5 M sodium cyanoborohydride solution.

[0081] S23. Phosphorylcholine polymer grafting: The bovine pericardium material treated in step S22 is immersed in an aqueous solution of aldehyde-terminated poly(2-methacryloyloxyethyl) phosphorylcholine homopolymer (PC-CHO, Mn=150000g / mol) with a concentration of 0.1mg / mL, ensuring complete immersion in the aqueous solution of the polymer. The reaction is carried out at 10℃ for 72h, allowing the aldehyde group of the phosphorylcholine polymer to react with another primary amino group of the bridging molecule to form a Schiff base bond.

[0082] S24. Second reduction: The bovine pericardial material treated in step S23 is transferred into a 1.5 M sodium cyanoborohydride solution with a pH of 4.0, ensuring complete immersion in the sodium cyanoborohydride solution. The reaction is carried out at room temperature with shaking for 72 hours. The Schiff base bonds generated in step S23 are reduced to stable secondary amine bonds. Then, the material is thoroughly washed with sterile physiological saline to obtain the modified pericardial material with surface covalently grafted phosphorylcholine polymer.

[0083] Example 10:

[0084] A method for surface-modifying pericardial materials includes the following steps:

[0085] S1. Provides glutaraldehyde-crosslinked calf heart material;

[0086] In a clean environment at 15-25℃, connective tissue, fat, blood vessels, mucosa, fascia, and visible foreign matter adhering to fresh bovine pericardium were removed within 2 hours after collection. During the cleaning process, the pericardium was continuously rinsed with sterile saline. The cleaned and rinsed bovine pericardium material was then immersed in glutaraldehyde phosphate buffer solution with a pH of 7.4 and a mass percentage concentration of 0.8% for crosslinking at room temperature for 72 hours. After that, it was thoroughly washed with sterile saline to obtain bovine pericardium material crosslinked with glutaraldehyde.

[0087] S2, grafting of phosphorylcholine polymers; specifically including the following sub-steps:

[0088] S21. Bridging treatment: The glutaraldehyde-crosslinked bovine pericardium material was immersed in a 1M 1,3-propanediamine aqueous solution at pH 8.0, ensuring complete immersion. The reaction was carried out at 40°C for 4 hours, allowing a primary amino group of the bridging molecule to react with the residual aldehyde group on the surface of the bovine pericardium material to form a Schiff base bond. Then, the material was thoroughly washed with sterile physiological saline to remove the physically adsorbed 1,3-propanediamine.

[0089] S22, First reduction: The bovine pericardium material treated in step S21 is transferred to a 1.0 M sodium borohydride solution with a pH of 4.0 and is immersed in the solution, ensuring complete immersion. The reaction is carried out at room temperature with shaking for 1 hour. The Schiff base bond generated in step S21 is reduced to a stable secondary amine bond. Then, it is thoroughly washed with a 0.05 M sodium borohydride solution.

[0090] S23. Phosphorylcholine polymer grafting: The pericardium material treated in step S22 is immersed in an aqueous solution of aldehyde-terminated poly(2-methacryloyloxyethyl) phosphorylcholine homopolymer (PC-CHO, Mn=600g / mol) with a concentration of 150mg / mL, ensuring complete immersion in the aqueous solution of the polymer. The reaction is carried out at 40℃ for 1h, so that the aldehyde group of the phosphorylcholine polymer reacts with another primary amino group of the bridging molecule to form a Schiff base bond.

[0091] S24. Second reduction: The bovine pericardial material treated in step S23 is transferred into a sodium cyanoborohydride solution with a pH of 4.0 and a concentration of 1.0M. The mixture is shaken at room temperature for 1 hour. The Schiff base bonds generated in step S23 are reduced to stable secondary amine bonds. Then, the material is thoroughly washed with sterile physiological saline to obtain the modified pericardial material with surface covalently grafted phosphorylcholine polymer.

[0092] Example 11:

[0093] A method for surface-modifying pericardial materials includes the following steps:

[0094] S1. Provides glutaraldehyde-crosslinked calf heart material;

[0095] In a clean environment at a temperature of 15-25℃, connective tissue, fat, blood vessels, mucosa, fascia, and visible foreign matter adhering to the fresh bovine pericardium were removed within 2 hours after collection. During the cleaning process, the pericardium was continuously rinsed with sterile saline. The cleaned and rinsed bovine pericardium material was then immersed in 0.8% glutaraldehyde phosphate buffer solution at pH 7.4 and crosslinked at room temperature for 72 hours to obtain glutaraldehyde-crosslinked bovine pericardium, which was then washed with sterile saline for later use.

[0096] S2, grafting of phosphorylcholine polymers; specifically including the following sub-steps:

[0097] S21. Bridging treatment: The glutaraldehyde-crosslinked bovine pericardium material was immersed in a 1.2 M L-lysine aqueous solution at pH 6.5, ensuring complete immersion. The reaction was carried out at 20°C for 24 hours, allowing a primary amino group of the bridging molecule to react with the residual aldehyde group on the surface of the bovine pericardium material to form a Schiff base bond. Then, the material was thoroughly washed with sterile physiological saline to remove the physically adsorbed L-lysine.

[0098] S22, First reduction: The bovine pericardium material treated in step S21 is transferred to a 1.2M sodium cyanoborohydride solution with a pH of 5.5 and a concentration of 1.2M for immersion, ensuring complete submersion in the sodium cyanoborohydride solution. The reaction is carried out at room temperature with shaking for 24 hours, and the Schiff base bond generated in step S21 is reduced to a stable secondary amine bond; then, it is thoroughly washed with a 1.2M sodium cyanoborohydride solution.

[0099] S23. Phosphorylcholine polymer grafting: The bovine pericardium material treated in step S22 is immersed in an aqueous solution of aldehyde-terminated poly(2-methacryloyloxyethyl) phosphorylcholine homopolymer (PC-CHO, Mn=10000g / mol) with a concentration of 100mg / mL, ensuring complete immersion in the aqueous solution of the polymer. The reaction is carried out at 20℃ for 24h, allowing the aldehyde group of the phosphorylcholine polymer to react with another primary amino group of the bridging molecule to form a Schiff base bond.

[0100] S24. Second reduction: The bovine pericardial material treated in step S23 is transferred to a 1.2 M sodium cyanoborohydride solution with a pH of 5.5 and ensured to be completely submerged in the sodium cyanoborohydride solution. The reaction is carried out at room temperature with shaking for 24 hours. The Schiff base bonds generated in step S23 are reduced to stable secondary amine bonds. Then, the material is thoroughly washed with sterile physiological saline to obtain the modified pericardial material with surface covalently grafted phosphorylcholine polymer.

[0101] II. Comparative Example:

[0102] Comparative Example 1

[0103] In a clean environment at a temperature of 15-25℃, connective tissue, fat, blood vessels, mucosa, fascia, and visible foreign matter adhering to the fresh bovine pericardium were removed within 2 hours after collection. During the cleaning process, the pericardium was continuously rinsed with sterile saline. The cleaned and rinsed bovine pericardium material was then immersed in glutaraldehyde phosphate buffer solution with a pH of 7.4 and a mass percentage concentration of 0.8%, and crosslinked at room temperature for 72 hours to obtain glutaraldehyde crosslinked bovine pericardium material.

[0104] III. Experimental Examples:

[0105] 1. Anticoagulant performance test:

[0106] The modified pericardial materials prepared in Examples 1 and 2, as well as the glutaraldehyde-crosslinked bovine pericardial material prepared in Comparative Example 1, were subjected to partial coagulation activating enzyme time (PTT) experiments. The negative group consisted of high-density polyethylene material, the positive group consisted of glass beads, and the plasma used in the experiment served as the blank group.

[0107] 1.1 Experimental Methods: The procedure was performed in accordance with the "Coagulation Test" method in the draft for comments completed in July 2024 - National Standard GB / T 14233.2-20XX "Test Methods for Medical Infusion, Transfusion and Injection Equipment Part 2: Biological Test Methods".

[0108] 1.2 Experimental Results: As shown in Table 1 and Figure 3 As shown.

[0109] 2. Anti-calcification effect experiment:

[0110] The modified pericardial materials prepared in Examples 1 and 2, as well as the glutaraldehyde-crosslinked bovine pericardial material prepared in Comparative Example 1, were subjected to in vitro calcification tests.

[0111] 2.1 Experimental Methods: The modified pericardial materials prepared in Examples 1 and 2, as well as the glutaraldehyde-crosslinked bovine pericardial material prepared in Comparative Example 1, were immersed in a solution containing high calcium and phosphorus elements for 72 hours. After removal, they were washed and dried, and then the amount of calcium adsorbed on the pericardial materials was measured. The tissue calcium content was determined by atomic absorption spectrometry. The high calcium and phosphorus element solution was prepared with deionized water and contained 20 mM calcium chloride. and 20mM sodium dihydrogen phosphate .

[0112] 2.2 Experimental Results: As shown in Table 1 and Figure 4 As shown

[0113] Table 1. Anticoagulant properties and anticalcification effects

[0114]

[0115] From Table 1 and Figure 3 It can be known that:

[0116] Comparative Example 1, lacking the biomimetic support of phosphorycholine groups due to the absence of phosphorycholine polymer grafting onto the glutaraldehyde-crosslinked pericardial material surface, exhibited a slightly lower PTT time than the blank and negative control groups, failing to demonstrate improved anticoagulant performance. Examples 1 and 2, however, employed directional bonding with bridging molecules containing bis-primary amino groups. This guided the precise grafting of phosphorycholine polymers onto the animal pericardial material surface, avoiding the masking of active phosphorycholine groups that might occur with random grafting. This ensured the effective functioning of the phosphorycholine polymer's biomimetic properties and guaranteed the high activity of the phosphorycholine biomimetic coating. This coating, in turn, improved the material's surface blood compatibility and inhibited the activation of the coagulation system and platelet adhesion, significantly extending the PTT time and demonstrating excellent anticoagulant effects. The modified pericardial materials in Examples 3 to 11 also employed directional bonding with bridging molecules containing bis-primary amino groups, thus exhibiting similarly excellent anticoagulant effects.

[0117] From Table 1 and Figure 4 It can be known that:

[0118] In Examples 1 and 2, the calcium content was significantly reduced compared to Comparative Example 1 due to the use of bis-primary amino bridging molecules for directional linking, which blocked the residual aldehyde sites caused by glutaraldehyde crosslinking. Simultaneously, a phosphorylcholine polymer with excellent biocompatibility was grafted onto the materials. This resulted in superior anti-calcification properties. The modified pericardial materials in Examples 3 to 11 also employed bis-primary amino bridging molecules for directional linking, thus exhibiting similarly excellent anti-calcification properties.

[0119] The above description is merely a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any changes or substitutions that can be easily conceived by those skilled in the art within the scope of the technology disclosed in the present invention should be included within the scope of protection of the present invention.

Claims

1. A method for surface modification of biomaterials, characterized in that, Includes the following steps: Animal pericardial material treated with glutaraldehyde is provided, with residual active aldehyde groups on its surface; A bridging molecule comprising a first primary amino group and a second primary amino group is provided; the bridging molecule is selected from any one of ethylenediamine, hexamethylenediamine, 1,3-propanediamine, 1,4-butanediamine, and lysine. Provided is a phosphorylcholine polymer with aldehyde-terminated ends; said phosphorylcholine polymer is a homopolymer of aldehyde-terminated poly(2-methacryloyloxyethyl phosphorylcholine) with a number-average molecular weight of 600 g / mol to 150,000 g / mol. The first primary amine group is bonded to the aldehyde group on the surface of the pericardial material through a first covalent reaction. The first covalent reaction includes: immersing the animal pericardial material in a bridging molecule solution to react and form a first Schiff base bond, followed by a first wash to remove the physically adsorbed bridging molecules; then placing it in a first reducing agent solution to reduce the first Schiff base bond to a first secondary amine bond, followed by a second wash. The second primary amino group is bonded to the aldehyde group at the end of the phosphorylcholine polymer through a second covalent reaction. The second covalent reaction includes: immersing the animal pericardial material treated with the first covalent reaction in a phosphorylcholine polymer solution to react and form a second Schiff base bond, and then placing it in a second reducing agent solution to reduce the second Schiff base bond to a second secondary amine bond. This enables the grafting of phosphorylcholine polymers onto the surface of animal pericardial materials.

2. The method for surface-modified biomaterials according to claim 1, characterized in that, In the first covalent reaction, the animal pericardial material is immersed in a bridging molecule solution to form the first Schiff base bond. Specifically, the animal pericardial material after glutaraldehyde cross-linking treatment is immersed in a bridging molecule aqueous solution with a pH of 5.0 to 8.0 and a concentration of 0.01M to 2M, and reacted at a temperature of 10℃ to 40℃ for 4h to 72h; then a first washing is performed.

3. The method for surface-modified biomaterials according to claim 1, characterized in that, In the first covalent reaction, the first Schiff base bond is reduced to the first secondary amine bond in the first reducing agent solution. Specifically, the animal pericardial material is transferred into a reducing agent solution with a pH of 3.0 to 8.5 and a concentration of 0.01M to 2M, and the reaction is carried out with shaking at room temperature for 1 to 72 hours; then a second washing is performed.

4. The method for surface-modified biomaterials according to claim 1, characterized in that, In the first covalent reaction, the reducing agent is selected from at least one of sodium cyanoborohydride and sodium borohydride.

5. The method for surface-modified biomaterials according to claim 1, characterized in that, In the second covalent reaction, the animal pericardial material treated in the first covalent reaction is immersed in a phosphorylcholine polymer solution to form a second Schiff base bond. Specifically, the animal pericardial material treated in the first covalent reaction is immersed in a phosphorylcholine polymer solution with a concentration of 0.1 mg / mL to 150 mg / mL and reacted at a temperature of 10°C to 40°C for 1 h to 72 h.

6. The method for surface-modified biomaterials according to claim 1, characterized in that, In the second covalent reaction, the second Schiff base bond is reduced to the second secondary amine bond in the second reducing agent solution. Specifically, the animal pericardial material is transferred into a reducing agent solution with a pH of 3.0 to 8.5 and a concentration of 0.01M to 2M, and reacted with shaking at room temperature for 1 to 72 hours, followed by washing.

7. The method for surface-modified biomaterials according to claim 1, characterized in that, In the second covalent reaction, the reducing agent is selected from at least one of sodium cyanoborohydride and sodium borohydride.

8. The method for surface-modified biomaterials according to claim 1, characterized in that, The animal pericardial material is derived from cattle, pigs, or sheep.

9. A modified pericardial material prepared by the method according to any one of claims 1-8.

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