Tissue adhesive material as well as preparation method and application thereof

Abstract

The invention discloses a tissue adhesive material as well as a preparation method and application thereof. The tissue binding material disclosed by the invention is bi-crosslinked network hydrogel formed by temperature-sensitive crosslinking and Schiff base reaction; the temperature-sensitive crosslinking is generated by a modified polylactide caprolactone-polyethylene glycol-polylactide caprolactone copolymer, the copolymer is subjected to gel transition when the temperature is close to the body temperature, and the hydrogel is formed through crosslinking; the Schiff base reaction is generated by the polyamino polymer and the aldehyde group of the modified polylactide caprolactone-polyethylene glycol-polylactide caprolactone copolymer. Hydrogel crosslinked with a multi-amino polymer is formed in situ on the basis of modified polylactide caprolactone-polyethylene glycol-polylactide caprolactone copolymer hydrogel, so that bi-crosslinked network hydrogel is formed, and the tissue bonding material is good in biocompatibility, has biodegradability, is non-toxic, and can be applied to the field of tissue bonding. And firm adhesion with tissues can be generated, so that gas leakage and blood leakage of damaged tissue parts are prevented.

Description

Technical Field

[0001] The present application relates to the technical field of medical gels, and particularly to a tissue adhesive material, a preparation method thereof, and an application thereof. Background Art

[0002] Tens of millions of people suffer from various tissue traumas every year, including skin cuts, chronic wounds, surgical incisions, etc. Suturing and stapling are traditional surgical methods for dealing with traumas, but they will cause additional tissue damage and cannot effectively prevent the leakage of body fluids and gases. Tissue adhesive materials can seal the wound site, provide mechanical support and hemostasis while preventing leakage, and provide an effective solution for surgical intervention of traumas.

[0003] Currently, existing tissue adhesive materials have obvious chemical property disadvantages, and their clinical applications are still limited. For example, although fibrin adhesives are considered the gold standard, they have low viscosity and anastomotic leakage is likely to occur. Cyanoacrylate adhesives have strong stability, but are considered toxic, can cause obvious inflammatory reactions, and require an additional curing process, which is inconvenient for clinical use. Monocrosslinked polyethylene glycol tissue adhesives have good biocompatibility, but have low strength. An ideal tissue adhesive material should have biocompatibility, biodegradability, non-toxicity, and be able to form a firm adhesion with tissues.

[0004] Therefore, the research and development of tissue adhesive materials with better performance remains the research focus and difficulty in this field. Summary of the Invention

[0005] The purpose of the present application is to provide a new tissue adhesive material, a preparation method thereof, and an application thereof.

[0006] In order to achieve the above purpose, the present application adopts the following technical solutions:

[0007] One aspect of the present application discloses a tissue adhesive material, which is a double-crosslinked network hydrogel formed by temperature-sensitive crosslinking and Schiff base reaction; wherein, the temperature-sensitive crosslinking is generated by a modified poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer, and the copolymer undergoes a gel transition when approaching body temperature and crosslinks to form a hydrogel; the Schiff base reaction is generated by the aldehyde group of a multi-amino polymer and the modified poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer. In the present application, "approaching body temperature" refers to a temperature of 15 - 37°C. In one implementation manner of the present application, heating the copolymer to 30 - 37°C can ensure the occurrence of the gel transition.

[0008] It should be noted that in the tissue adhesive material of the present application, the modified poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer is in a liquid form at a temperature lower than body temperature, and undergoes a gel transition when approaching body temperature to form a hydrogel. The present application creatively combines the hydrogel formed by the modified poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer itself with the hydrogel formed by the Schiff base reaction of the multi-amino polymer and the modified poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer, and in-situ forms a Schiff base reaction hydrogel on the basis of the copolymer hydrogel, thereby forming a double-crosslinked network hydrogel, which can not only improve the tissue adhesiveness and strength of the tissue adhesive material of the present application, but also has the advantages of good biocompatibility and biodegradability. In short, the tissue adhesive material of the present application has good biocompatibility, is biodegradable, non-toxic, and can produce a firm adhesion with tissues, effectively preventing gas leakage and blood leakage at the damaged tissue site of the human body.

[0009] In one implementation manner of the present application, the modified poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer is composed of three segments of polymers, namely poly(lactide-co-caprolactone) (PLACL), polyethylene glycol (PEG), and poly(lactide-co-caprolactone) (PLACL), and the molecular weight ratio of the three segments of polymers is in sequence (1000 - 4000):(1000 - 4000):(1000 - 4000).

[0010] It should be noted that the PEG segment has good water solubility. In the molecular weight range of 1000 - 4000 Da, the hydrogen bond interaction between it and water molecules is relatively strong, and it can better reflect hydrophilicity. When combined with a suitable poly(lactide-co-caprolactone) segment (i.e., PLACL with a molecular weight of 1000 - 4000 Da), it is conducive to the hydrophilic-hydrophobic transition at a certain temperature, so that the copolymer aqueous solution exhibits thermosensitivity. When the molecular weight of the PLACL segment is too low, it may lead to insufficient hydrophobicity and it is difficult to form an obvious temperature response. While when the molecular weight is too high, the overall water solubility of the copolymer is too poor, which is also not conducive to the manifestation of thermosensitivity.

[0011] In one implementation manner of the present application, the molar ratio of lactide to caprolactone in poly(lactide-co-caprolactone) is 1:(0.5 - 2).

[0012] It should be noted that lactide has relatively strong hydrophobicity, while caprolactone is relatively weak; increasing the proportion of lactide will enhance the hydrophobicity of the copolymer, and it is easy to have hydrophobic aggregation and cause phase transition at a higher temperature; increasing the proportion of caprolactone will weaken the overall hydrophobicity of the copolymer, and hydrophobic aggregation will occur at a lower temperature, thus affecting the temperature range of thermosensitivity; therefore, the present application preferably adopts a molar ratio of lactide to caprolactone of 1:(0.5 - 2).

[0013] In one implementation of the present application, the modification is to modify the hydroxyl end groups of the poly(lactide-caprolactone)-polyethylene glycol-poly(lactide-caprolactone) copolymer into aldehyde groups.

[0014] In one implementation of the present application, the multi-amino polymer is polyethyleneimine.

[0015] In one implementation of the present application, the molecular weight of polyethyleneimine is 600 - 20000 Da.

[0016] In one implementation of the present application, the weight of the modified poly(lactide-caprolactone)-polyethylene glycol-poly(lactide-caprolactone) copolymer is 1 - 400 times the weight of the multi-amino polymer.

[0017] On the other hand, the present application discloses the use of the tissue adhesive material of the present application in the preparation of a material for preventing gas leakage or liquid leakage at the damaged tissue site of the human body.

[0018] On yet another aspect, the present application discloses a preparation method of the tissue adhesive material of the present application, which includes the following steps:

[0019] S1. Dissolve the modified poly(lactide-caprolactone)-polyethylene glycol-poly(lactide-caprolactone) copolymer in a solvent at 2 - 8 °C to form a solution, labeled as component one;

[0020] S2. Dissolve the multi-amino polymer in a solvent at 2 - 8 °C to form a solution, labeled as component two;

[0021] S3. Mix component one and component two, stir evenly to obtain a mixed solution, perform in-situ crosslinking, and when the temperature rises to body temperature, or when the mixed solution rises to body temperature during use, a double-crosslinked network hydrogel is formed.

[0022] In one implementation of the present application, the solvent is water or phosphate buffer solution.

[0023] In one implementation of the present application, the same solvent is used in step S1 and step S2.

[0024] In one implementation of the present application, the time for in-situ crosslinking is 3 - 600 seconds.

[0025] Among them, in-situ crosslinking means that after mixing component one and component two evenly, let it stand for 3 - 600 seconds to react.

[0026] In one implementation of the present application, the total concentration of the solute in component one is 1 - 400 mg / mL.

[0027] In one implementation of the present application, the concentration of the solute in component two is 1 - 100 mg / mL.

[0028] In one implementation of the present application, the preparation method of the modified poly(lactide-caprolactone)-polyethylene glycol-poly(lactide-caprolactone) copolymer includes:

[0029] Heat polyethylene glycol, lactide, caprolactone, and stannous octoate to 90 - 150 °C under the protection of an inert atmosphere and react for 18 - 30 h. Then dissolve it in dichloromethane at a concentration of 0.2 - 0.8 g / mL. Pour the solution into petroleum ether with a volume 5 - 20 times that of the solution, precipitate, remove the supernatant. After redissolving and precipitating 3 - 5 times, collect the precipitate and dry it to obtain the poly(lactide-caprolactone)-polyethylene glycol-poly(lactide-caprolactone) copolymer;

[0030] Dissolve the poly(lactide-caprolactone)-polyethylene glycol-poly(lactide-caprolactone) copolymer, 4-formylbenzoic acid, 4-dimethylaminopyridine, and dicyclohexylcarbodiimide in dichloromethane at a concentration of 0.05 - 0.5 g / mL at 0 - 5 °C. After reacting at room temperature for 18 - 30 h, pour the solution into methyl tert-butyl ether with a volume 5 - 20 times that of the solution, precipitate, remove the supernatant. After redissolving and precipitating 3 - 5 times, collect the precipitate and dry it to obtain the modified poly(lactide-caprolactone)-polyethylene glycol-poly(lactide-caprolactone) copolymer. Among them, redissolving and precipitating 3 - 5 times means dissolving the precipitate in dichloromethane, then pouring the solution into petroleum ether or methyl tert-butyl ether to precipitate, removing the supernatant, and repeating this process 3 - 5 times.

[0031] In one implementation of the present application, the dosage of stannous octoate is 0.0005 - 0.001 times the total molar amount of lactide and caprolactone.

[0032] In one implementation of the present application, the inert atmosphere is argon.

[0033] In one implementation of the present application, the molar ratio of the poly(lactide-caprolactone)-polyethylene glycol-poly(lactide-caprolactone) copolymer, 4-formylbenzoic acid, 4-dimethylaminopyridine, and dicyclohexylcarbodiimide is 1:(6 - 10):(0.5 - 2):(5 - 15).

[0034] Due to the above technical solutions, the beneficial effects of the present application are as follows:

[0035] The tissue adhesive material of the present application in-situ forms a hydrogel crosslinked with a polyamino polymer on the basis of the modified poly(lactide-caprolactone)-polyethylene glycol-poly(lactide-caprolactone) copolymer hydrogel, thereby forming a double-crosslinked network hydrogel. The tissue adhesive material of the present application has good biocompatibility, biodegradability, non-toxicity, and can form a firm adhesion with tissues, preventing gas leakage and blood leakage at the damaged tissue site, providing a new tissue adhesive material for damaged tissues. Description of the Drawings

[0036] Figure 1 It is the statistical result of the bursting strength test of the hydrogels prepared in the examples and comparative examples of this application;

[0037] Figure 2 It is the statistical result of the lap shear tensile strength of the hydrogels prepared in the examples and comparative examples of this application;

[0038] Figure 3 It is the statistical result of the in vitro degradation test of the hydrogel prepared in the examples of this application. Detailed implementation manners

[0039] Existing tissue adhesives have obvious chemical property drawbacks and their clinical applications are still limited. For example, although fibrin adhesives are considered the gold standard, they have low viscosity and anastomotic leakage is likely to occur; cyanoacrylate adhesives have strong stability, but are considered toxic and can cause obvious inflammatory reactions, and an additional curing process is required, which is inconvenient for clinical use; single-crosslinked polyethylene glycol tissue adhesives have good biocompatibility, but have low strength.

[0040] This application has developed a dual-crosslinked tissue adhesive absorbable material based on thermosensitivity and Schiff base reaction and its preparation method. The temperature sensitivity of this material is generated by the modified poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer. The copolymer solution shows a liquid form below body temperature and undergoes a gel transition when approaching body temperature to form a hydrogel; the Schiff base reaction is generated by the aldehyde group of the multi-amino polymer and the modified poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer. The tissue adhesive of this application has good biocompatibility, is biodegradable, non-toxic and can produce a firm adhesion with tissues, and can effectively prevent gas leakage and blood leakage at the damaged tissue site of the human body.

[0041] The following further elaborates on this application through specific examples. The following examples only further illustrate this application and should not be construed as a limitation of this application.

[0042] Unless otherwise specified, the technical means used in the examples are conventional means well-known to those skilled in the art.

[0043] Example 1

[0044] S1. The dry PEG (molecular weight 1000 Da), lactide, caprolactone, and stannous octoate were heated to 110 °C under argon protection and reacted for 18 h in a molar ratio of 1:7.74:7.74:0.012. Then, they were dissolved in dichloromethane at a concentration of 0.4 g / mL. The solution was slowly poured into 10 times the volume of petroleum ether to precipitate. The supernatant was removed. After repeating the dissolution and precipitation process 4 times, the precipitate was collected and dried to obtain a poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer (PLACL-PEG-PLACL) with a molecular weight of 3000 Da. The molecular weight of each of the three segments of the polymer was 1000 Da.

[0045] Among them, the structural formula of PLACL-PEG-PLACL is as follows:

[0046]

[0047] S2. The dry PLACL-PEG-PLACL, 4-formylbenzoic acid, 4-dimethylaminopyridine, and dicyclohexylcarbodiimide were dissolved in dichloromethane at a concentration of 0.2 g / mL at 0 - 5 °C in a molar ratio of 1:8:1:10. After reacting at room temperature for 24 h, the solution was slowly poured into 10 times the volume of methyl tert-butyl ether to precipitate. The supernatant was removed. After repeating the dissolution and precipitation process 4 times, the precipitate was collected and dried to obtain a modified poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer (CHO-PLACL-PEG-PLACL-CHO).

[0048] The structural formula of CHO-PLACL-PEG-PLACL-CHO is as follows:

[0049]

[0050] S3. Weigh CHO-PLACL-PEG-PLACL-CHO into a phosphate buffer solution at 2 - 8 °C with a concentration of 200 mg / mL, which is Component One; add polyethyleneimine to another phosphate buffer solution with a concentration of 30 mg / mL, and stir the solution at room temperature to form a homogeneous solution, which is Component Two. Mix the two-component solutions in equal volumes and crosslink in situ for 30 seconds. After raising the temperature to body temperature (about 36.5 °C), the double-crosslinked network hydrogel of this example is obtained.

[0051] Example 2

[0052] S1. The dry PEG (molecular weight 1500 Da), lactide, caprolactone, and stannous octoate were heated to 120 °C under argon protection and reacted for 24 h in a molar ratio of 1:7.74:7.74:0.012. Then, it was dissolved in dichloromethane at a concentration of 0.3 g / mL. The solution was slowly poured into 10 times the volume of petroleum ether to precipitate. The supernatant was removed. After repeating the dissolution and precipitation 4 times, the precipitate was collected and dried to obtain a poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer (PLACL-PEG-PLACL) with a molecular weight of 3500 Da. The molecular weights of the three segments of the polymer were 1000 Da, 1500 Da, and 1000 Da, and the structural formula was the same as that in Example 1.

[0053] S2. The dry PLACL-PEG-PLACL, 4-formylbenzoic acid, 4-dimethylaminopyridine, and dicyclohexylcarbodiimide were dissolved in dichloromethane at a concentration of 0.2 g / mL at 0 - 5 °C in a molar ratio of 1:8:1:10. After reacting at room temperature for 24 h, the solution was slowly poured into 10 times the volume of methyl tert-butyl ether to precipitate. The supernatant was removed. After repeating the dissolution and precipitation 4 times, the precipitate was collected and dried to obtain a modified poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer (CHO-PLACL-PEG-PLACL-CHO), and the structural formula was the same as that in Example 1.

[0054] S3. Weigh CHO-PLACL-PEG-PLACL-CHO into a phosphate buffer solution at 2 - 8 °C with a concentration of 200 mg / mL; add polyethyleneimine to another phosphate buffer solution with a concentration of 30 mg / mL, and the mixed solution was stirred at room temperature to form a homogeneous solution. The two-component solutions were mixed evenly in equal volumes and crosslinked in situ for 30 seconds. After raising the temperature to body temperature (about 36.5 °C), the double-crosslinked network hydrogel of this example was obtained.

[0055] Example 3

[0056] S1. The dry PEG (molecular weight 1500 Da), lactide, caprolactone, and stannous octoate were heated to 120 °C under argon protection and reacted for 24 h in a molar ratio of 1:11.6:11.6:0.017. Then, it was dissolved in dichloromethane at a concentration of 0.3 g / mL. The solution was slowly poured into 10 times the volume of petroleum ether to precipitate. The supernatant was removed. After repeating the dissolution and precipitation 4 times, the precipitate was collected and dried to obtain a poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer (PLACL-PEG-PLACL) with a molecular weight of 4500 Da. The molecular weights of the three segments of the polymer were all 1500 Da, and the structural formula was the same as that in Example 1.

[0057] S2. Dissolve the dried PLACL-PEG-PLACL, 4-formylbenzoic acid, 4-dimethylaminopyridine, and dicyclohexylcarbodiimide in dichloromethane at a concentration of 0.2 g / mL at a molar ratio of 1:8:1:10 at 0 - 5 °C. After reacting at room temperature for 24 h, slowly pour the solution into 10 times the volume of methyl tert-butyl ether to precipitate. Remove the supernatant, and after redissolving and precipitating 4 times repeatedly, collect and dry the precipitate to obtain the modified poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer (CHO-PLACL-PEG-PLACL-CHO), whose structural formula is the same as that in Example 1.

[0058] S3. Weigh CHO-PLACL-PEG-PLACL-CHO into a phosphate buffer solution at 2 - 8 °C with a concentration of 200 mg / mL; add polyethyleneimine to another phosphate buffer solution with a concentration of 30 mg / mL, and stir the mixed solution at room temperature to form a homogeneous solution. Mix the two-component solutions in equal volumes and crosslink in situ for 30 seconds, and then heat to body temperature (about 36.5 °C) to obtain the double-crosslinked network hydrogel of this example.

[0059] Example 4

[0060] S1. Heat PEG (molecular weight 1500 Da), lactide, caprolactone, and stannous octoate at a molar ratio of 1:15.48:15.48:0.023 to 120 °C under argon protection and react for 24 h. Then dissolve it in dichloromethane at a concentration of 0.3 g / mL. Slowly pour the solution into 10 times the volume of petroleum ether to precipitate. Remove the supernatant, and after redissolving and precipitating 4 times repeatedly, collect and dry the precipitate to obtain the poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer (PLACL-PEG-PLACL) with a molecular weight of 5500 Da. The molecular weights of the three segments of the polymer are 2000 Da, 1500 Da, and 2000 Da, and the structural formula is the same as that in Example 1.

[0061] S2. Dissolve the dried PLACL-PEG-PLACL, 4-formylbenzoic acid, 4-dimethylaminopyridine, and dicyclohexylcarbodiimide in dichloromethane at a concentration of 0.2 g / mL at a molar ratio of 1:8:1:10 at 0 - 5 °C. After reacting at room temperature for 24 h, slowly pour the solution into 10 times the volume of methyl tert-butyl ether to precipitate. Remove the supernatant, and after redissolving and precipitating 4 times repeatedly, collect and dry the precipitate to obtain the modified poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer (CHO-PLACL-PEG-PLACL-CHO), whose structural formula is the same as that in Example 1.

[0062] S3. Weigh CHO-PLACL-PEG-PLACL-CHO into a phosphate buffer solution at 2-8 °C with a concentration of 200 mg / mL; add polyethyleneimine to another phosphate buffer solution with a concentration of 30 mg / mL, stir the mixed solution at room temperature to form a homogeneous solution. Mix the two-component solutions in equal volumes evenly and crosslink in situ for 30 seconds, and then raise the temperature to body temperature (about 36.5 °C) to obtain the double-crosslinked network hydrogel of this example.

[0063] Example 5

[0064] S1. Dry PEG (molecular weight 2000 Da), lactide, caprolactone, and stannous octoate were heated to 130 °C under argon protection and reacted for 20 h in a molar ratio of 1:15.48:15.48:0.023, and then dissolved in dichloromethane at a concentration of 0.3 g / mL. The solution was slowly poured into 10 times the volume of petroleum ether, and a precipitate was formed. The supernatant was removed, and the precipitate was dissolved and precipitated 4 times repeatedly. After that, the precipitate was collected and dried to obtain a poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer (PLACL-PEG-PLACL) with a molecular weight of 6000 Da. The molecular weights of the three segments of the polymer were all 2000 Da, and the structural formula was the same as that in Example 1.

[0065] S2. Dry PLACL-PEG-PLACL, 4-formylbenzoic acid, 4-dimethylaminopyridine, and dicyclohexylcarbodiimide were dissolved in dichloromethane at a concentration of 0.2 g / mL at 0-5 °C in a molar ratio of 1:8:1:10, and reacted at room temperature for 24 h. Then, the solution was slowly poured into 10 times the volume of methyl tert-butyl ether, and a precipitate was formed. The supernatant was removed, and the precipitate was dissolved and precipitated 4 times repeatedly. After that, the precipitate was collected and dried to obtain a modified poly(lactide-co-caprolactone)-polyethylene glycol-poly(lactide-co-caprolactone) copolymer (CHO-PLACL-PEG-PLACL-CHO), and the structural formula was the same as that in Example 1.

[0066] S3. Weigh CHO-PLACL-PEG-PLACL-CHO into a phosphate buffer solution at 2-8 °C with a concentration of 200 mg / mL; add polyethyleneimine to another phosphate buffer solution with a concentration of 30 mg / mL, stir the mixed solution at room temperature to form a homogeneous solution. Mix the two-component solutions in equal volumes evenly and crosslink in situ for 30 seconds, and then raise the temperature to body temperature (about 36.5 °C) to obtain the double-crosslinked network hydrogel of this example.

[0067] Comparative Example 1

[0068] Based on Example 1, dissolve CHO-PLACL-PEG-PLACL-CHO (the molecular weights of the three segments of the polymer are 1000 Da, 1000 Da, and 1000 Da) in phosphate buffer at 2 - 8 °C with a concentration of 100 mg / mL. After raising the temperature to body temperature (about 36.5 °C), a single-crosslinked network hydrogel of this example is obtained.

[0069] Comparative Example 2

[0070] Based on Example 2, dissolve CHO-PLACL-PEG-PLACL-CHO (the molecular weights of the three segments of the polymer are 1000 Da, 1500 Da, and 1000 Da) in phosphate buffer at 2 - 8 °C with a concentration of 100 mg / mL. After raising the temperature to body temperature (about 36.5 °C), a single-crosslinked network hydrogel of this example is obtained.

[0071] Comparative Example 3

[0072] Based on Example 3, dissolve CHO-PLACL-PEG-PLACL-CHO (the molecular weights of the three segments of the polymer are 1500 Da, 1500 Da, and 1500 Da) in phosphate buffer at 2 - 8 °C with a concentration of 100 mg / mL. After raising the temperature to body temperature (about 36.5 °C), a single-crosslinked network hydrogel of this example is obtained.

[0073] Comparative Example 4

[0074] Based on Example 4, dissolve CHO-PLACL-PEG-PLACL-CHO (the molecular weights of the three segments of the polymer are 2000 Da, 1500 Da, and 2000 Da) in phosphate buffer at 2 - 8 °C with a concentration of 100 mg / mL. After raising the temperature to body temperature (about 36.5 °C), a single-crosslinked network hydrogel of this example is obtained.

[0075] Comparative Example 5

[0076] Based on Example 5, dissolve CHO-PLACL-PEG-PLACL-CHO (the molecular weights of the three segments of the polymer are 2000 Da, 2000 Da, and 2000 Da) in phosphate buffer at 2 - 8 °C with a concentration of 100 mg / mL. After raising the temperature to body temperature (about 36.5 °C), a single-crosslinked network hydrogel of this example is obtained.

[0077] Comparative Example 6

[0078] Weigh para - formylbenzoate PEG with a molecular weight of 3000 Da into phosphate buffer with a concentration of 200 mg / mL; add polyethyleneimine to another phosphate buffer with a concentration of 30 mg / mL, and stir the mixed solution at room temperature to form a homogeneous solution. In - situ mix the two - component solutions in equal volume for 30 seconds, and after raising the temperature to body temperature, Comparative Example 6 is obtained.

[0079] Comparative Example 7

[0080] Weigh para - formylbenzoate PEG with a molecular weight of 4500 Da into phosphate buffer at a concentration of 200 mg / mL; add polyethyleneimine to another phosphate buffer at a concentration of 30 mg / mL. The mixed solution is stirred at room temperature to form a homogeneous solution. The two - component solutions are mixed in situ in equal volumes for 30 seconds and then heated to body temperature to obtain Comparative Example 7.

[0081] Comparative Example 8

[0082] Weigh para - formylbenzoate PEG with a molecular weight of 6000 Da into phosphate buffer at a concentration of 200 mg / mL; add polyethyleneimine to another phosphate buffer at a concentration of 30 mg / mL. The mixed solution is stirred at room temperature to form a homogeneous solution. The two - component solutions are mixed in situ in equal volumes for 30 seconds and then heated to body temperature to obtain Comparative Example 8.

[0083] The gels prepared in the above examples and comparative examples were tested as follows:

[0084] 1. Gel transition temperature test

[0085] At 4 °C, the sample is dissolved in phosphate buffer and then transferred to a small tube and placed in a water bath capable of precisely controlling the temperature. The heating rate during the gel transition temperature test is 1 °C / min, and it is placed at each temperature for 10 min. The test temperature range is 10 °C - 50 °C. If the copolymer solution does not flow within 30 seconds when the test tube is inverted, it is considered that gel transition behavior has occurred, and this temperature is the gel transition temperature of the copolymer solution at this concentration. Table 1 shows the gel transition temperature test results of Comparative Examples 1 - 5.

[0086] Table 1 Gel transition temperatures of Comparative Examples 1 - 5

[0087] Sample Gel transition temperature Comparative Example 1 18℃ Comparative Example 2 23℃ Comparative Example 3 27℃ Comparative Example 4 32℃ Comparative Example 5 35℃

[0088] The results in Table 1 show that the gel transition temperatures of the 5 block copolymers with different molecular weight ratios are all above 8 °C and below body temperature, indicating that the CHO - PLACL - PEG - PLACL - CHO material provided by the present invention indeed has temperature sensitivity.

[0089] 2. Burst strength test

[0090] The burst strength measurement was carried out based on the standard test method for surgical sealants (ASTM F2392-04). A hole with a diameter of 3 mm was made in the collagen casing, and 400 μL of the in-situ cured gel of the example or comparative example was sealed above the hole. The burst strength was measured 5 minutes after the hydrogel was stabilized in-situ at the puncture site, that is, pressure was applied to the normal saline in the test device at a constant rate of 2 mL / min, and the pressure in the system was measured using a pressure gauge. The peak pressure during the pressure measurement was regarded as the burst strength.

[0091] The burst strength data of Examples 1-5 and Comparative Examples 1-8 are as Figure 1 shown. Figure 1 The results show that compared with the single-crosslinked network hydrogels of Comparative Examples 1 to 8, the double-crosslinked network hydrogels provided in Examples 1 to 5 have higher tissue adhesiveness and strength, both exceeding 150 mmHg, higher than the normal human blood pressure of 120 mmHg. A large amount of collagen is contained on the surface of human tissues such as the skin, lungs, stomach, intestines, and pancreas. The double-crosslinked materials provided in Examples 1 to 5 can adhere to collagen, thereby effectively preventing gas leakage and blood leakage at the damaged tissue sites of the human body.

[0092] 3. Lap shear tensile test

[0093] The test scheme was carried out with reference to the industry standard YY / T 0729.1-2009 of the People's Republic of China. Fresh pigskin was selected as the test substrate. The length of the two pieces of pigskin was 5 ± 0.2 cm, and the width was 2.5 ± 0.1 cm. The treatment of pigskin was referred to Appendix B of YY / T 0729.1-2009.

[0094] The test steps include:

[0095] 1) Prepare test samples according to the product instruction manual and Sections 7.1-7.2 of YY / T 0729.1-2009, and the adhesive usage amount was 0.4 mL.

[0096] 2) After the adhesive solidified for 1 min, gently scrape off the adhesive overflowing from the lap edge with a scalpel, and measure the bonding area. The bonding area = bonding length × bonding width, accurate to 0.05 cm 2 .

[0097] 3) Place the specimen in the fixture of the testing machine, and make the force application direction be the long axis direction of the specimen. Make the crosshead load the specimen at a speed of 5 mm / min until failure.

[0098] 4) Record the load at the time of failure.

[0099] In this example, at least 10 groups of parallel samples were specifically tested, and the test was carried out at 23 ± 2 °C and 50 ± 5% relative humidity. The lap shear tensile strength data of Examples 1-5 and Comparative Examples 1-8 are asFigure 2 As shown Figure 2 The data statistically analyzed are the averages of duplicate samples.

[0100] Figure 2 The results show that, compared with the single-crosslinked network hydrogels of Comparative Examples 1 to 8, the double-network hydrogels provided in Examples 1 to 5 have higher shear strength. The double-crosslinked materials provided in Examples 1 to 5 can effectively adhere to the tissue surface, and thus can be effectively used to prevent gas leakage and blood leakage.

[0101] 4. Cytotoxicity Test

[0102] The cytotoxicity test was carried out based on the standard test method for the biology of medical devices (GB / T 16886.58.4.1 Agar diffusion test). The double-crosslinked tissue adhesive materials prepared in Examples 1-5 or the hydrogels prepared in the comparative examples were placed on a solidified agar layer containing cells, and cultured at 37 °C in an atmosphere of 5% carbon dioxide, 95% air and 95% humidity for 24 hours, and the near confluence and morphology of the cells before and after culture were observed. After 24-hour culture, the effects of Examples 1-5 on the cells are shown in Table 2.

[0103] Table 2 Test results of the effects of the hydrogels of Examples and Comparative Examples on cells

[0104]

[0105]

[0106] It can be seen from the experimental data in Table 2 that all Examples and Comparative Examples do not have obvious negative effects on the cells, indicating that the double-crosslinked tissue adhesive materials provided by the present invention have good cytocompatibility.

[0107] 5. Biocompatibility Test

[0108] The biocompatibility test was carried out on the double-crosslinked tissue adhesive materials prepared in Examples 1-5, and the test was carried out according to the relevant national standards; specifically, the sensitization test and intracutaneous irritation test were carried out with reference to the national standard GB / T16886.10-2005, and the acute systemic toxicity test was carried out with reference to the national standard GB / T16886.11-1997. The biocompatibility test results of Examples 1-5 are shown in Table 3.

[0109] Table 3 Test results of sensitization, intracutaneous irritation and acute systemic toxicity of the hydrogels of Examples 1-5

[0110] Example Allergenicity Intradermal irritation Acute systemic toxicity Example 1 Non-allergenic No intradermal irritation Negative Example 2 Non-allergenic No intradermal irritation Negative Example 3 Non-allergenic No intradermal irritation Negative Example 4 Non-allergenic No intradermal irritation Negative Example 5 Non-allergenic No intradermal irritation Negative

[0111] It can be seen from the results in Table 3 that the double-crosslinked tissue adhesive materials provided in Examples 1 to 5 have good biocompatibility, no toxicity, no allergy and no irritation.

[0112] 6. In vitro degradation test

[0113] The double-crosslinked tissue adhesive materials prepared in Examples 1-5 were separately immersed in PBS solution, the initial weight of the hydrogel was weighed and recorded, the pH value of the solution was set to 7.4, the temperature was 37 °C, and the solution was shaken at a speed of 150 rpm using a bench-top orbital shaker. The hydrogel was taken out regularly, dried with filter paper, and then weighed. The weight retention rate was calculated. The change of the weight retention rate of the samples in Examples 1-5 with time is as Figure 3 shown.

[0114] As Figure 3 can be seen, the double-crosslinked tissue adhesive materials prepared in Examples 1 to 5 have good degradation performance in PBS solution and can be degraded within 60 days.

[0115] As can be seen from the above experiments, the double-crosslinked tissue adhesive and absorbable material based on thermosensitivity and Schiff base reaction provided in this application has no cytotoxicity, can be degraded by organisms, has higher strength and tissue adhesiveness compared with single-crosslinked hydrogels, and can be widely used in the adhesion of tissue wounds.

[0116] The above content is a further detailed description of this application in combination with specific implementation manners. It cannot be determined that the specific implementation of this application is only limited to these descriptions. For those of ordinary skill in the technical field to which this application belongs, without departing from the concept of this application, several simple deductions or substitutions can still be made.

Claims

1. A tissue adhesive material, characterized in that: It is a double cross-linked network hydrogel formed by temperature-sensitive cross-linking and Schiff base reaction; The temperature-sensitive crosslinks are produced by a modified polylactide-caprolactone-polyethylene glycol-polylactide-caprolactone copolymer, which undergoes a gel transition when close to body temperature and crosslinks to form a hydrogel; The Schiff base reaction is generated by the polyamino polymer and the aldehyde group of the modified polylactide caprolactone-polyethylene glycol-polylactide caprolactone copolymer.

2. The tissue adhesive material according to claim 1, characterized in that: The modified polylactide caprolactone-polyethylene glycol-polylactide caprolactone copolymer consists of three polymer segments, namely polylactide caprolactone, polyethylene glycol and polylactide caprolactone, and the molecular weight ratios of the three polymer segments are (1000-4000):(1000-4000):(1000-4000) in sequence.

3. The tissue adhesive material according to claim 2, characterized in that: The molar ratio of lactide to caprolactone in the polylactide-caprolactone is 1:(0.5-2).

4. The tissue adhesive material according to claim 1, characterized in that: The modification is to modify the hydroxyl end groups of the polylactide caprolactone-polyethylene glycol-polylactide caprolactone copolymer into aldehyde groups.

5. The tissue adhesive material according to claim 1, characterized in that: The polyamino polymer is polyethyleneimine; Preferably, the molecular weight of the polyethyleneimine is 600-20000 Da.

6. The tissue adhesive material according to any one of claims 1 to 5, characterized in that: The weight of the modified polylactide caprolactone-polyethylene glycol-polylactide caprolactone copolymer is 1-400 times the weight of the polyamino polymer.

7. Use of the tissue adhesive material according to any one of claims 1 to 6 in the preparation of a material for preventing gas leakage or liquid leakage in damaged tissues of the human body.

8. The method for preparing the tissue adhesive material according to any one of claims 1 to 6, characterized in that: The following steps are included: S1. Dissolving the modified polylactide caprolactone-polyethylene glycol-polylactide caprolactone copolymer in a solvent at 2-8°C to prepare a solution, which is marked as component one; S2. dissolving the polyamino polymer in a solvent at 2-8°C to prepare a solution, which is marked as component 2; S3. Component one and component two are mixed and stirred evenly to obtain a mixed solution, and a mixed solution is cross-linked in situ. When the mixed solution is heated to body temperature, or when the mixed solution is heated to body temperature during use, a double cross-linked network hydrogel is formed.

9. The preparation method according to claim 8, characterized in that: The solvent is water or phosphate buffer; Preferably, step S1 and step S2 use the same solvent; Preferably, the in-situ cross-linking time is 3-600 seconds; Preferably, the total concentration of solutes in component 1 is 1-400 mg / mL; Preferably, the concentration of the solute in the component 2 is 1-100 mg / mL.

10. The preparation method according to claim 8 or 9, characterized in that: The preparation method of the modified polylactide caprolactone-polyethylene glycol-polylactide caprolactone copolymer comprises: Polyethylene glycol, lactide, caprolactone, and stannous octoate are heated to 90-150° C. under inert atmosphere protection for reaction for 18-30 hours, and then dissolved in dichloromethane at a concentration of 0.2-0.8 g / mL, and the solution is poured into 5-20 times the volume of petroleum ether to precipitate, and the supernatant is removed. After repeatedly dissolving and precipitating the precipitate 3-5 times, the precipitate is collected and dried to obtain a polylactide caprolactone-polyethylene glycol-polylactide caprolactone copolymer; Dissolve polylactide caprolactone-polyethylene glycol-polylactide caprolactone copolymer, 4-formylbenzoic acid, 4-dimethylaminopyridine, and dicyclohexylcarbodiimide in dichloromethane at a concentration of 0.05-0.5 g / mL at 0-5° C., react at room temperature for 18-30 hours, pour the solution into 5-20 times the volume of methyl tert-butyl ether, precipitate, remove the supernatant, repeatedly dissolve and precipitate 3-5 times, collect the precipitate, and dry to obtain the modified polylactide caprolactone-polyethylene glycol-polylactide caprolactone copolymer; Preferably, the amount of stannous octoate used is 0.0005-0.001 times the total molar amount of lactide and caprolactone; Preferably, the inert atmosphere is argon; Preferably, the molar ratio of polylactide caprolactone-polyethylene glycol-polylactide caprolactone copolymer, 4-formylbenzoic acid, 4-dimethylaminopyridine and dicyclohexylcarbodiimide is 1:(6-10):(0.5-2):(5-15).

Images