A 3D printed hydrogel scaffold and a method of making the same

Abstract

The application discloses a 3D-printed hydrogel support and a preparation method thereof, relates to the technical field of hydrogel materials, and aims to improve the antibacterial property and biocompatibility of the hydrogel material by modifying the raw material. The application first uses gelatin as the raw material, and processes the gelatin to increase the content of surface amino groups, so that the subsequent reaction is facilitated; then, the application also uses 2,5-thiophene dicarboxylic acid as the raw material, and controls the reaction conditions to react the 2,5-thiophene dicarboxylic acid with 2-aminobenzimidazole, so as to generate an antibacterial intermediate containing a free carboxyl group, a thiophene ring and an imidazole ring structure; the antibacterial intermediate is combined in the gelatin chain segment in the form of an amide bond, then under the action of a photoinitiator, the amide bond is broken to generate an acyl radical, and the acyl radical is further coupled and grafted with an acrylic ester double bond, so as to form a hydrogel cross-linking structure, and the hydrogel material is provided with sufficient compressive strength.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of hydrogel materials, in particular to a 3D printed hydrogel scaffold and a preparation method thereof. BACKGROUND

[0002] After the human body suffers external trauma, the wound often bleeds and even part of the tissue is missing. In the subsequent self-repair process of the human body tissue, due to the fact that some soft tissue parts of the human body have less blood vessels or lymphatic structures, the self-repairing ability is very limited, so that after the skin repair of the damaged part is completed, the subcutaneous tissue will still be in an un-repaired state for a long time, causing defects in appearance and even function.

[0003] As a substance similar to the water content of human tissue, the hydrogel material has good biocompatibility and can be used as a human tissue repair material. However, the hydrogel is easily infected by bacteria during application, which leads to a series of inflammation. Therefore, it is necessary to research a 3D printed hydrogel material with antibacterial function. SUMMARY

[0004] The purpose of the present application is to solve the problems existing in the prior art.

[0005] To achieve the above purpose, the present application provides the following technical scheme: a preparation method of a 3D printed hydrogel scaffold, comprising the following steps:

[0006] S1. Gelatin is added to deionized water, and after being dissolved by heating and stirring, L-lysine is added thereto, and after being uniformly mixed by continuing to stir, transglutaminase is added thereto, the temperature of the reaction system is controlled to be 45-55 DEG C, and after being stirred for 12-24 h, the reaction system is heated to boiling for 3-5 min, and then the mixed solution is dialyzed by using a dialysis bag for 1-3 days, and then freeze-drying is performed to obtain an aminated gelatin material;

[0007] S2. The antibacterial intermediate is dispersed in ultrapure water, heated and ultrasonically stirred and dispersed, and then the aminated gelatin material is added thereto, and after being stirred for 8-12 h under a nitrogen atmosphere, the reaction liquid is cooled to room temperature, and then dialyzed by using a dialysis bag for 2-3 days, and then freeze-drying is performed to obtain an antibacterial gelatin material;

[0008] S3. The antibacterial gelatin material is mixed with acrylate, polyethylene glycol acrylate and a photoinitiator, and then acetone is added thereto, and after the viscosity is adjusted to 4500-7000 mPa·s, a hydrogel solution is obtained;

[0009] S4. The hydrogel solution is placed in a 3D printer, cooled, and then a hydrogel scaffold blank is preliminarily printed, and then ultraviolet light is irradiated to cross-link and solidify to form a hydrogel scaffold.

[0010] Further, in step S1, the adding amount of each component is 10 parts of gelatin and 0.1-2 parts of L-lysine by weight fraction;

[0011] The adding amount ratio of gelatin to transglutaminase is 10g: (50-100) U.

[0012] Further, in step S1, the dialysis bag is a 5-10KDa dialysis bag, and the dialysis fluid is replaced every 3-5h interval.

[0013] Further, in step S2, the preparation method of the antibacterial intermediate is:

[0014] Under the protection of nitrogen atmosphere, 2,5-thiophene dicarboxylic acid is dispersed in DMSO, stirred and mixed uniformly, then EDC and NHS are added, and after activation at room temperature for 30-45min, PBS buffer is added dropwise to adjust the pH value to 7.3-7.5, to obtain a 2,5-thiophene dicarboxylic acid dispersion, which is ready for use;

[0015] 2-Aminobenzimidazole is dispersed in ultrapure water, stirred until completely dispersed, then added dropwise to the 2,5-thiophene dicarboxylic acid dispersion, heated to 30-37℃, and stirred for 2.5-3h, then ethanolamine is continuously added to the mixed solution, and the stirring is continued for 30-45min, then the excess solvent is removed by rotary evaporation to obtain the antibacterial intermediate.

[0016] Further, the adding amount of each component is 10 parts of 2,5-thiophene dicarboxylic acid, 0.08-0.12 parts of EDC, 0.1-0.15 parts of NHS, 4-7.5 parts of 2-aminobenzimidazole, and 0.15-0.4 parts of ethanolamine by weight fraction.

[0017] Further, in step S2, the adding amount of each component is 10 parts of aminated gelatin material and 0.15-1.5 parts of antibacterial intermediate by weight fraction;

[0018] The dialysis bag is an 8-14KDa dialysis bag, and the dialysis fluid is replaced every 3-5h interval.

[0019] Further, in step S3, the adding amount of each component is 10-15 parts of antibacterial gelatin material, 25-38 parts of acrylate, 15-30 parts of polyethylene glycol acrylate, and 2-3.5 parts of photoinitiator.

[0020] Further, in step S3, the acrylate is trimethylolpropane triacrylate; the average molecular weight of the polyethylene glycol acrylate is 300-500; and the photoinitiator is photoinitiator Irgacure 2959.

[0021] Further, in step S4, the cooling temperature is 8-15 DEG C.

[0022] Further, a hydrogel scaffold is prepared by the above method.

[0023] Compared with the prior art, the present application has the following beneficial effects:

[0024] In order to improve the antibacterial property and biocompatibility of the hydrogel material, the raw material is modified; the present application first uses gelatin as a raw material, which is a kind of natural polymer material extracted from animal connective tissue, and has good biological affinity and is similar to the extracellular matrix of human body, and contains rich amino structure on the gelatin segment; after mixing with L-lysine, the amino group in the gelatin segment reacts with the carboxyl group in the lysine under the participation of the enzyme glutamine transaminase, which can further increase the content of the surface amino group and facilitate the reaction in the subsequent steps.

[0025] Then, the present application also uses 2,5-thiophene dicarboxylic acid as a raw material, which reacts with 2-amino benzimidazole by controlling the reaction conditions, thereby generating an antibacterial intermediate containing free carboxyl, thiophene ring and imidazole ring structure; the thiophene ring and imidazole ring have excellent antibacterial performance, and the nitrogen atom contained therein can effectively affect the life activity of microbial pathogens and destroy their cells, thereby having excellent antibacterial performance; and the carboxylic acid group contained therein can further react with the amino group in the amino gelatin through an amide bond and be combined in the gelatin segment; then under the action of a photoinitiator, the amide bond is broken to generate an acyl radical, which further couples with an acrylic ester double bond to graft, thereby forming a hydrogel crosslinked structure and providing sufficient compressive strength for the hydrogel material. DETAILED DESCRIPTION

[0026] Based on the embodiments in the present application, all other embodiments obtained by those of ordinary skill in the art without creative labor are within the scope of protection of the present application.

[0027] Example 1. A preparation method of a 3D printed hydrogel scaffold, comprising the following steps:

[0028] S1. According to the weight fraction, 10 parts of gelatin are added to deionized water, heated to 50 DEG C, and after stirring and dissolving, 0.1 parts of L-lysine are added, and after uniform stirring and mixing, 50U of glutamine transaminase is added in an amount of 10g: 50U of gelatin, the reaction system temperature is controlled at 45 DEG C, and the stirring reaction is carried out for 24h; after that, the reaction system is heated to boiling for 3min, and then the mixed liquid is dialyzed for 1 day using a 5KDa dialysis bag; during dialysis, the dialysate is changed every 5h; after dialysis, freeze-drying is performed to obtain an amino gelatin material.

[0029] S2. 0.15 parts of the antibacterial intermediate was dispersed in ultrapure water, heated to 50℃, and dispersed by ultrasonic stirring. Then 10 parts of the amino gelatin material was added, and stirred and reacted for 8h under nitrogen atmosphere. After cooling to room temperature, the reaction solution was dialyzed for 2 days using an 8KDa dialysis bag, and the dialysis solution was replaced every 5h. After dialysis, freeze-drying was performed to obtain the antibacterial gelatin material;

[0030] The preparation method of the antibacterial intermediate is as follows:

[0031] Under the protection of nitrogen atmosphere, 10 parts of 2,5-thiophene dicarboxylic acid was dispersed in DMSO, and stirred and mixed uniformly. Then 0.08 parts of EDC and 0.1 parts of NHS were added, and after being activated at room temperature for 30min, 2,5-thiophene dicarboxylic acid dispersion was obtained by adjusting the pH value to 7.3 by dropwise adding PBS buffer solution, and was prepared for use.

[0032] 4 parts of 2-amino benzimidazole was dispersed in ultrapure water, and stirred until completely dispersed. Then it was added dropwise to the 2,5-thiophene dicarboxylic acid dispersion, heated to 30℃, and stirred and reacted for 3h. Then 0.15 parts of ethanolamine was added to the mixed solution, and stirred and reacted for 45min. After removing the excess solvent by rotary evaporation, the antibacterial intermediate was obtained.

[0033] S3. 10 parts of the antibacterial gelatin material, 25 parts of trimethylolpropane triacrylate, 15 parts of polyethylene glycol acrylate, and 2 parts of photoinitiator Irgacure 2959 were mixed, and then acetone was added to adjust the viscosity to 5000mPa·s to obtain a hydrogel solution.

[0034] S4. The hydrogel solution was placed in a 3D printer, cooled to 15℃, and a hydrogel scaffold blank was formed by preliminary printing. Then the hydrogel scaffold was formed by crosslinking and curing under ultraviolet light irradiation.

[0035] Example 2. A preparation method of a 3D printed hydrogel scaffold, comprising the following steps:

[0036] S1. 10 parts of gelatin was added to deionized water, heated to 50℃, and stirred and dissolved. Then 0.1 parts of L-lysine was added, and stirred and mixed uniformly. Then glutamine transaminase was added in an amount of 10g:50U based on the mass ratio of gelatin, and the temperature of the reaction system was controlled at 55℃. After stirring and reacting for 12h, the reaction system was heated to boiling for 5min to inactivate. Then the mixture was dialyzed for 3 days using a 10KDa dialysis bag, and the dialysis solution was replaced every 3h. After dialysis, freeze-drying was performed to obtain the amino gelatin material.

[0037] S2. 0.15 parts of the antibacterial intermediate was dispersed in ultrapure water, heated to 50℃, and dispersed by ultrasonic stirring. Then 10 parts of the amino gelatin material was added, and stirred for 12 hours under nitrogen atmosphere. After cooling to room temperature, the reaction solution was dialyzed for 3 days using a 14KDa dialysis bag, and the dialysis solution was replaced every 3 hours. After dialysis, freeze-drying was performed to obtain the antibacterial gelatin material;

[0038] The preparation method of the antibacterial intermediate is as follows:

[0039] Under the protection of nitrogen atmosphere, 10 parts of 2,5-thiophene dicarboxylic acid was dispersed in DMSO, and stirred uniformly. Then 0.08 parts of EDC and 0.1 parts of NHS were added, and stirred for 45 minutes at room temperature. After adjusting the pH value to 7.5 by adding PBS buffer solution dropwise, a 2,5-thiophene dicarboxylic acid dispersion was obtained, which was ready for use.

[0040] 4 parts of 2-amino benzimidazole was dispersed in ultrapure water, and stirred until completely dispersed. Then it was added dropwise to the 2,5-thiophene dicarboxylic acid dispersion, and heated to 37℃. After stirring for 2.5 hours, 0.4 parts of ethanolamine was added, and stirred for another 30 minutes. After removing the excess solvent by rotary evaporation, the antibacterial intermediate was obtained.

[0041] S3. 10 parts of the antibacterial gelatin material, 25 parts of trimethylolpropane triacrylate, 15 parts of polyethylene glycol acrylate, and 2 parts of light initiator Irgacure 2959 were mixed, and then acetone was added to adjust the viscosity to 5000 mPa·s to obtain a hydrogel solution.

[0042] S4. The hydrogel solution was placed in a 3D printer, cooled to 8℃, and then the hydrogel scaffold blank was formed by preliminary printing. Then, the hydrogel scaffold was formed by cross-linking and solidification under ultraviolet light irradiation.

[0043] Example 3. A preparation method of a 3D printed hydrogel scaffold, comprising the following steps:

[0044] Compared with Example 2, the amount of L-lysine added in step S1 is increased in this embodiment.

[0045] S1. 10 parts of gelatin was added into deionized water, heated to 50℃, after stirring and dissolving, 2 parts of L-lysine was added, and then stirred and mixed uniformly, and then glutamine transaminase was added in an amount of 10g: 50U (mass ratio), the temperature of the reaction system was controlled at 55℃, and the reaction was stirred for 12h, then the reaction system was heated to boiling and inactivated for 5min, then the mixture was dialyzed using a dialysis bag with a molecular weight of 10KDa for 3 days, and the dialysate was changed every 3h, after dialysis, freeze-drying was performed to obtain an aminated gelatin material;

[0046] S2. 0.15 parts of an antibacterial intermediate was dispersed in ultrapure water, heated to 50℃, and then ultrasonically stirred and dispersed, then 10 parts of the aminated gelatin material was added, and then stirred and reacted for 12h under a nitrogen atmosphere, then cooled to room temperature, and then the reaction solution was dialyzed using a dialysis bag with a molecular weight of 14KDa for 3 days, and the dialysate was changed every 3h, after dialysis, freeze-drying was performed to obtain an antibacterial gelatin material;

[0047] The preparation method of the antibacterial intermediate is as follows:

[0048] Under the protection of nitrogen atmosphere, 10 parts of 2,5-thiophene dicarboxylic acid was dispersed in DMSO, and then stirred and mixed uniformly, then 0.08 parts of EDC and 0.1 parts of NHS were added, and then the mixture was activated at room temperature for 45min, then PBS buffer was added dropwise to adjust the pH value to 7.5, and then a 2,5-thiophene dicarboxylic acid dispersion was obtained, which was prepared for use;

[0049] 4 parts of 2-amino benzimidazole was dispersed in ultrapure water, and then stirred until completely dispersed, then the 2-amino benzimidazole was added dropwise into the 2,5-thiophene dicarboxylic acid dispersion, heated to 37℃, and then stirred and reacted for 2.5h, then 0.4 parts of ethanolamine was added to the mixed solution, and then stirred and reacted for 30min, then the excess solvent was removed by rotary evaporation to obtain the antibacterial intermediate;

[0050] S3. 10 parts of the antibacterial gelatin material, 25 parts of trimethylolpropane triacrylate, 15 parts of polyethylene glycol acrylate, and 2 parts of a photoinitiator Irgacure 2959 were mixed, then acetone was added to adjust the viscosity to 5000mPa·s to obtain a hydrogel solution;

[0051] S4. The hydrogel solution was placed in a 3D printer, cooled to 8℃, and then a hydrogel scaffold blank was formed by preliminary printing, then the hydrogel scaffold was formed by crosslinking and curing under ultraviolet light irradiation.

[0052] Example 4. A preparation method of a 3D printed hydrogel scaffold, comprising the following steps:

[0053] Compared with Example 3, the present example increases the amount of transglutaminase added in step S1;

[0054] S1. 10 parts of gelatin were added to deionized water, heated to 50℃, and dissolved by stirring. Then 2 parts of L-lysine were added, and the mixture was stirred until uniform. Then, transglutaminase was added in an amount of 10 g:100 U by weight ratio of gelatin, and the temperature of the reaction system was controlled at 55℃. After stirring for 12 h, the reaction system was heated to boiling for 5 min to inactivate. Then, the mixture was dialyzed for 3 days using a dialysis bag with a molecular weight cutoff of 10 KDa. The dialysis solution was replaced every 3 h. After dialysis, the product was freeze-dried to obtain an aminated gelatin material.

[0055] S2. 0.15 parts of the antibacterial intermediate were dispersed in ultrapure water and heated to 50℃. After ultrasonic stirring and dispersion, 10 parts of the aminated gelatin material were added. The reaction was stirred for 12 h under a nitrogen atmosphere, and then the reaction solution was cooled to room temperature. The reaction solution was dialyzed for 3 days using a dialysis bag with a molecular weight cutoff of 14 KDa. The dialysis solution was replaced every 3 h. After dialysis, the product was freeze-dried to obtain an antibacterial gelatin material.

[0056] The preparation method of the antibacterial intermediate is as follows:

[0057] Under nitrogen atmosphere protection, 10 parts of 2,5-thiophene dicarboxylic acid were dispersed in DMSO. After stirring and mixing, 0.08 parts of EDC and 0.1 parts of NHS were added. After standing for 45 min at room temperature for activation, the pH value of the mixture was adjusted to 7.5 by adding PBS buffer dropwise to obtain a 2,5-thiophene dicarboxylic acid dispersion, which was ready for use.

[0058] 4 parts of 2-amino benzimidazole were dispersed in ultrapure water and stirred until completely dispersed. Then, the mixture was added dropwise to the 2,5-thiophene dicarboxylic acid dispersion. The temperature was increased to 37℃, and the reaction was stirred for 2.5 h. Then, 0.4 parts of ethanolamine were added to the mixture, and the reaction was stirred for another 30 min. After removing the excess solvent by rotary evaporation, the antibacterial intermediate was obtained.

[0059] S3. 10 parts of the antibacterial gelatin material, 25 parts of trimethylolpropane triacrylate, 15 parts of polyethylene glycol acrylate, and 2 parts of photoinitiator Irgacure 2959 were mixed. Then, acetone was added to adjust the viscosity to 5000 mPa·s to obtain a hydrogel solution.

[0060] S4. The hydrogel solution was placed in a 3D printer and cooled to 8℃. The hydrogel scaffold blank was formed by preliminary printing, and then cross-linked and solidified by UV light irradiation to form a hydrogel scaffold.

[0061] Example 5. A method for preparing a 3D-printed hydrogel scaffold, comprising the following steps:

[0062] Compared with Example 4, this example increases the amount of antibacterial intermediate added in step S2;

[0063] S1. By weight, add 10 parts of gelatin to deionized water, heat to 50°C, stir to dissolve, add 2 parts of L-lysine, continue stirring and mixing evenly, add glutamine transaminase at a mass ratio of 10g:100U to gelatin, control the reaction temperature at 55°C, stir and react for 12 hours, then heat the reaction system to boiling and inactivate for 5 minutes, dialyze the mixture using a 10 kDa dialysis bag for 3 days, changing the dialysate every 3 hours during dialysis, freeze-dry after dialysis to obtain aminated gelatin material;

[0064] S2. By weight, 1.5 parts of the antibacterial intermediate were dispersed in ultrapure water, heated to 50°C, and dispersed by ultrasonic stirring. Then, 10 parts of the aminated gelatin material were added. Under a nitrogen atmosphere, the mixture was stirred and reacted for 12 hours. After cooling to room temperature, the reaction solution was dialyzed for 3 days using a 14 kDa dialysis bag. The dialysate was changed every 3 hours during dialysis. After dialysis, the solution was freeze-dried to obtain the antibacterial gelatin material.

[0065] The method for preparing the antibacterial intermediate is as follows:

[0066] Under nitrogen atmosphere protection, 10 parts by weight of 2,5-thiophene dicarboxylic acid were dispersed in DMSO and stirred until homogeneous. Then, 0.08 parts of EDC and 0.1 parts of NHS were added. After activation by standing at room temperature for 45 minutes, PBS buffer was added dropwise to adjust the pH to 7.5 to obtain the 2,5-thiophene dicarboxylic acid dispersion for later use.

[0067] Four parts of 2-aminobenzimidazole were dispersed in ultrapure water and stirred until completely dispersed. Then, it was added dropwise to a dispersion of 2,5-thiophene dicarboxylic acid. The mixture was heated to 37°C and stirred for 2.5 h. Then, 0.4 parts of ethanolamine were added to the mixture and stirred for 30 min. After removing excess solvent by rotary evaporation, the antibacterial intermediate was obtained.

[0068] S3. By weight, 10 parts of antibacterial gelatin material, 25 parts of trimethylolpropane triacrylate, 15 parts of polyethylene glycol acrylate, and 2 parts of photoinitiator Irgacure2959 are mixed. Acetone is added to the mixture, and the viscosity is adjusted to 5000 mPa·s to obtain a hydrogel solution.

[0069] S4. The hydrogel solution is placed in a 3D printer, cooled to 8℃, and after the initial printing of the hydrogel scaffold blank, ultraviolet light is irradiated to cross-link and solidify the hydrogel scaffold.

[0070] Embodiment 6. A method for preparing a 3D-printed hydrogel scaffold, comprising the following steps:

[0071] Compared with Embodiment 5, this embodiment increases the amount of 2-aminobenzimidazole added in step S2, and the remaining steps remain unchanged.

[0072] The preparation method of the antibacterial intermediate is as follows:

[0073] Under the protection of a nitrogen atmosphere, 10 parts of 2,5-thiophene dicarboxylic acid are dispersed in DMSO, and after stirring and mixing uniformly, 0.08 parts of EDC and 0.1 parts of NHS are added, and after standing for 45 min at room temperature, 2,5-thiophene dicarboxylic acid dispersion is obtained by adjusting the pH value to 7.5 by adding PBS buffer dropwise, and is ready for use;

[0074] 7.5 parts of 2-aminobenzimidazole are dispersed in ultrapure water, stirred until completely dispersed, and then added dropwise to the 2,5-thiophene dicarboxylic acid dispersion, heated to 37℃, and stirred for 2.5h, then 0.4 parts of ethanolamine is added to the mixed solution, and the stirring is continued for 30 min, then the excess solvent is removed by rotary evaporation, and the antibacterial intermediate is obtained.

[0075] Embodiment 7. A method for preparing a 3D-printed hydrogel scaffold, comprising the following steps:

[0076] Compared with Embodiment 6, this embodiment increases the amount of antibacterial gelatin material added in step S3, and the remaining steps remain unchanged.

[0077] S1. According to the weight fraction, 10 parts of gelatin are added to deionized water, heated to 50℃, and stirred to dissolve, then 2 parts of L-lysine are added, and the mixture is stirred and mixed uniformly, then glutamine transaminase is added in an amount of 10g:100U based on the mass ratio of gelatin, and the reaction system temperature is controlled at 55℃, and the stirring reaction is carried out for 12h, then the reaction system is heated to boiling for 5 min to inactivate, then the mixed solution is dialyzed using a 10KDa dialysis bag for 3 days, and the dialysis liquid is changed every 3h during dialysis, and after dialysis, the aminoated gelatin material is obtained by freeze-drying.

[0078] S2. According to the weight fraction, 1.5 parts of the antibacterial intermediate is dispersed into ultrapure water, heated to 50℃, and dispersed by ultrasonic stirring. Then, 10 parts of the aminated gelatin material is added. After stirring and reacting for 12 hours under a nitrogen atmosphere, the reaction system is cooled to room temperature. The reaction solution is dialyzed for 3 days using a 14KDa dialysis bag. The dialysis liquid is replaced every 3 hours. After dialysis, the antibacterial gelatin material is obtained by freeze-drying.

[0079] S3. According to the weight fraction, 15 parts of the antibacterial gelatin material is mixed with 25 parts of trimethylolpropane triacrylate, 15 parts of polyethylene glycol acrylate, and 2 parts of the photoinitiator Irgacure 2959. Then, acetone is added to adjust the viscosity to 5000mPa·s, and the hydrogel solution is obtained.

[0080] S4. The hydrogel solution is placed in a 3D printer and cooled to 8℃. After the initial printing of the hydrogel scaffold blank, the hydrogel scaffold is formed by cross-linking and solidification under ultraviolet light.

[0081] Example 8. A method for preparing a 3D-printed hydrogel scaffold, comprising the following steps:

[0082] Compared with Example 7, the amount of the added components other than the antibacterial gelatin material in step S3 is increased in this embodiment, and the remaining steps remain unchanged.

[0083] S1. According to the weight fraction, 10 parts of gelatin is added to deionized water, heated to 50℃, and stirred to dissolve. Then, 2 parts of L-lysine is added. After stirring and mixing uniformly, 10g of glutamine transaminase is added according to the mass ratio of 10g:100U. The temperature of the reaction system is controlled at 55℃, and the reaction is stirred for 12 hours. After the reaction system is heated to boiling for 5 minutes, the mixture is dialyzed for 3 days using a 10KDa dialysis bag. The dialysis liquid is replaced every 3 hours. After dialysis, the aminated gelatin material is obtained by freeze-drying.

[0084] S2. According to the weight fraction, 1.5 parts of the antibacterial intermediate is dispersed into ultrapure water, heated to 50℃, and dispersed by ultrasonic stirring. Then, 10 parts of the aminated gelatin material is added. After stirring and reacting for 12 hours under a nitrogen atmosphere, the reaction system is cooled to room temperature. The reaction solution is dialyzed for 3 days using a 14KDa dialysis bag. The dialysis liquid is replaced every 3 hours. After dialysis, the antibacterial gelatin material is obtained by freeze-drying.

[0085] S3. According to the weight fraction, 15 parts of the antibacterial gelatin material is mixed with 38 parts of trimethylolpropane triacrylate, 30 parts of polyethylene glycol acrylate, and 3.5 parts of the photoinitiator Irgacure 2959. Then, acetone is added to adjust the viscosity to 5000mPa·s, and the hydrogel solution is obtained.

[0086] S4. The hydrogel solution is placed in a 3D printer, cooled to 8℃, and after the initial printing of the hydrogel scaffold blank, ultraviolet light is irradiated to cross-link and solidify the hydrogel scaffold.

[0087] Comparative Example 1. A method for preparing a 3D-printed hydrogel scaffold, comprising the following steps:

[0088] In comparison with Example 2, the antibacterial gelatin material is not prepared in this comparative example;

[0089] S1. 25 parts of trimethylolpropane triacrylate, 15 parts of polyethylene glycol acrylate, and 2 parts of a photoinitiator Irgacure 2959 are mixed, and then acetone is added to adjust the viscosity to 5000 mPa·s, thereby obtaining a hydrogel solution;

[0090] S2. The hydrogel solution is placed in a 3D printer, cooled to 8℃, and after the initial printing of the hydrogel scaffold blank, ultraviolet light is irradiated to cross-link and solidify the hydrogel scaffold.

[0091] Comparative Example 2. A method for preparing a 3D-printed hydrogel scaffold, comprising the following steps:

[0092] In comparison with Example 2, only an equal amount of 2,5-thiophene dicarboxylic acid is used to replace the antibacterial intermediate in this comparative example;

[0093] S1. According to the weight fraction, 10 parts of gelatin is added to deionized water, heated to 50℃, and stirred to dissolve, then 0.1 parts of L-lysine is added, and the mixture is stirred uniformly, then glutamine transaminase is added in an amount of 50U per 10g of gelatin, the temperature of the reaction system is controlled at 55℃, and the reaction is stirred for 12h, then the reaction system is heated to boiling for 5min, and the mixture is dialyzed for 3 days using a 10KDa dialysis bag, the dialysis liquid is replaced every 3h during dialysis, and after dialysis, the mixture is freeze-dried to obtain an aminated gelatin material;

[0094] S2. According to the weight fraction, 0.15 parts of 2,5-thiophene dicarboxylic acid is dispersed in ultrapure water, heated to 50℃, and ultrasonically stirred and dispersed, then 10 parts of the aminated gelatin material is added, and the mixture is stirred for 12h under a nitrogen atmosphere, then the mixture is cooled to room temperature, dialyzed for 3 days using a 14KDa dialysis bag, the dialysis liquid is replaced every 3h during dialysis, and after dialysis, the mixture is freeze-dried to obtain an antibacterial gelatin material;

[0095] S3. According to the weight fraction, 10 parts of the antibacterial gelatin material is mixed with 25 parts of trimethylolpropane triacrylate, 15 parts of polyethylene glycol acrylate, and 2 parts of a photoinitiator Irgacure 2959, and then acetone is added to adjust the viscosity to 5000 mPa·s, thereby obtaining a hydrogel solution;

[0096] S4. The hydrogel solution is placed in a 3D printer, cooled to 8℃, and after the initial printing of the hydrogel scaffold blank, ultraviolet light is irradiated to cross-link and solidify the hydrogel scaffold.

[0097] Detection: The hydrogel scaffolds prepared in Examples 1-8 and Comparative Examples 1-2 are printed into detection samples with a height of 4.5 mm and a diameter of 6 mm, and the mechanical properties of the detection samples are detected using an Instron 5575 mechanical testing machine. The compression rate is 0.5 mm / min.

[0098] The hydrogel scaffolds prepared in Examples 1-8 and Comparative Examples 1-2 are subjected to antibacterial detection according to QB / T 2591-2003, and the detection results are shown in the following table.

[0099] The hydrogel scaffolds prepared in Examples 1-8 and Comparative Examples 1-2 are subjected to cytotoxicity detection according to GB / T 16886.5-2003.

[0100] The detection results are shown in the following table.

[0101]

[0102] Finally, it should be noted that the above only describes the preferred embodiments of the present application and is not intended to limit the present application. Although the present application has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent replacements to some technical features. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present application shall be included in the protection scope of the present application.

Claims

1. A method of preparing a 3D printed hydrogel scaffold, characterized in that, Comprise the following steps: S1. The gelatin is added to deionized water, after stirring and dissolving at elevated temperature, L-lysine is added, after stirring and mixing evenly, transglutaminase is added, the temperature of the reaction system is controlled at 45-55℃, after stirring for 12-24h, the reaction system is heated to boiling for 3-5min, then the mixture is dialyzed for 1-3 days using dialysis bag, and then freeze-dried to obtain the aminated gelatin material; S2. The antibacterial intermediate is dispersed in ultrapure water, heated and ultrasonically stirred and dispersed, then the aminated gelatin material is added, stirred and reacted for 8-12h under nitrogen atmosphere, cooled to room temperature, then the reaction solution is dialyzed for 2-3 days using dialysis bag, and then freeze-dried to obtain the antibacterial gelatin material; The preparation method of the antibacterial intermediate is: The 2,5-thiophene dicarboxylic acid is dispersed in DMSO under nitrogen atmosphere protection, stirred and mixed evenly, then EDC and NHS are added, after standing for 30-45min at room temperature for activation, PBS buffer solution is added dropwise to adjust the pH value to 7.3-7.5, and then a 2,5-thiophene dicarboxylic acid dispersion is obtained and reserved; The 2-amino benzimidazole is dispersed in ultrapure water, stirred until completely dispersed, then added dropwise into the 2,5-thiophene dicarboxylic acid dispersion, heated to 30-37℃, stirred and reacted for 2.5-3h, then ethanolamine is continuously added into the mixed solution, continuously stirred and reacted for 30-45min, then the excess solvent is removed by rotary evaporation to obtain the antibacterial intermediate; S3. The antibacterial gelatin material is mixed with acrylate, polyethylene glycol acrylate and photoinitiator, then acetone is added to adjust the viscosity to 4500-7000mPa·s to obtain a hydrogel solution; S4. The hydrogel solution is placed in a 3D printer, cooled, and a hydrogel scaffold blank is initially printed, then irradiated with ultraviolet light to crosslink and solidify to form a hydrogel scaffold.

2. The method of claim 1, wherein: In step S1, the addition amount of each component is 10 parts of gelatin and 0.1-2 parts of L-lysine by weight; The addition amount of gelatin and transglutaminase is in a ratio of 10g: (50-100) U.

3. The method of claim 1, wherein: In step S1, the dialysis bag is a 5-10kDa dialysis bag, and the dialysis liquid is replaced every 3-5h during dialysis.

4. The method of claim 1, wherein: The addition amount of each component is 10 parts of 2,5-thiophene dicarboxylic acid, 0.08-0.12 parts of EDC, 0.1-0.15 parts of NHS, 4-7.5 parts of 2-amino benzimidazole, and 0.15-0.4 parts of ethanolamine by weight.

5. The method of claim 1, wherein: In step S2, the addition amount of each component is 10 parts of aminated gelatin material and 0.15-1.5 parts of antibacterial intermediate by weight; The dialysis bag is an 8-14kDa dialysis bag, and the dialysis liquid is replaced every 3-5h during dialysis.

6. The method of claim 1, wherein: In step S3, the addition amount of each component is 10-15 parts of antibacterial gelatin material, 25-38 parts of acrylate, 15-30 parts of polyethylene glycol acrylate, and 2-3.5 parts of photoinitiator.

7. The method of claim 1, wherein: In step S3, the acrylate is trimethylolpropane triacrylate; the average molecular weight of the polyethylene glycol acrylate is 300-500; and the photoinitiator is photoinitiator Irgacure 2959.

8. The method of claim 1, wherein: In step S4, the cooling temperature is 8-15°C.

9. A hydrogel scaffold prepared by the method of any one of claims 1-8.

Images