Fish scale collagen hydrogel and preparation method thereof

Abstract

The invention relates to the technical field of biological materials, in particular to fish scale collagen hydrogel and a preparation method thereof.The preparation method comprises the following steps that fish scale collagen, polymeric monomers, a photoinitiator, a cross-linking agent, polydopamine nanoparticles and water are added into a reactor, the mixture is cooled to the room temperature after being heated and stirred, then the mixture is subjected to ultraviolet irradiation, and the fish scale collagen hydrogel is obtained. The fish scale collagen hydrogel is obtained. The preparation process is simple, fish scale collagen, polymeric monomers and polydopamine nanoparticles are used as raw materials, heat-induced protein denaturation and three-network concepts are combined, the fish scale collagen is used as a first-layer network, monomer polymers are used as a second-layer network, and the mechanical property is improved; and then adding polydopamine nano particles to form the three-network hydrogel. According to the fish scale collagen hydrogel disclosed by the invention, the hydrogel is endowed with antibacterial property while the mechanical property is further improved, and the introduced polydopamine can convert light energy into heat energy, so that the prepared fish scale collagen hydrogel can be used for thermal physiotherapy.

Description

Technical Field

[0001] The invention relates to the technical field of biomaterials, and in particular to a fish scale collagen hydrogel and a preparation method thereof. Background Art

[0002] Hydrogel is a cross-linked polymer network material formed by the reaction of one or more monomers. It can absorb water and swell, has good biocompatibility, high water content and adjustable physical and chemical properties, and shows broad application prospects in the biomedical field. However, due to problems such as low cross-linking density, uneven network structure or insufficient toughness of molecular chains, the mechanical properties of traditional hydrogels are generally poor, making it difficult to meet the needs of complex mechanical environments in practical applications.

[0003] Currently, mechanical properties are usually improved by introducing inorganic or organic nanomaterials such as nanoclay and cellulose nanocrystals as reinforcing phases. However, nanofillers are prone to cause stress concentration due to uneven dispersion, leading to local fracture, and excessive addition may sacrifice the swelling properties of the hydrogel, resulting in poor results.

[0004] In view of this, how to improve the mechanical properties of hydrogel without sacrificing its swelling properties is a technical problem that needs to be solved urgently. Summary of the invention

[0005] In order to solve the problem of poor mechanical properties of hydrogels in the prior art, the present invention provides a method for preparing fish scale collagen hydrogel, which combines heat-induced protein denaturation and the concept of three networks, with fish scale collagen as the first network layer, N,N'-dimethylacrylamide as the second network layer, and polydopamine nanoparticles cross-linked therewith to form a third network layer. The three-network hydrogel based on fish scale collagen is prepared by a one-pot method, which improves the mechanical properties and solves the problem of poor mechanical properties of hydrogels in the prior art.

[0006] The technical solution adopted by the present invention to solve its technical problem is: A method for preparing a fish scale collagen hydrogel comprises the following steps: adding fish scale collagen, a polymerization monomer, a photoinitiator, a cross-linking agent, polydopamine nanoparticles and water into a reactor, heating and stirring, cooling to room temperature, and irradiating with ultraviolet light to obtain the fish scale collagen hydrogel.

[0007] Optionally, heating the mixed solution includes: heating the mixed solution to 60-90°C.

[0008] Optionally, the polymerizable monomer is N,N'-dimethylacrylamide.

[0009] Optionally, the photoinitiator is 2959.

[0010] Optionally, the cross-linking agent is N,N-methylenebisacrylamide.

[0011] Optionally, the usage ratio of the fish scale collagen, the polymerized monomer, and the polydopamine nanoparticles is (5-12) g: (2-4) mL: (0.01-0.1) g.

[0012] Optionally, the fish scale collagen is prepared according to the following method: placing fish scales in an EDTA standard solution, reacting in a normal temperature water bath under stirring conditions, washing, and obtaining decalcified fish scales; adding the decalcified fish scales to purified water, heating at 60-100°C to obtain a crude solution; centrifuging the crude solution, taking the supernatant, evaporating, freeze-drying, and pulverizing to obtain the fish scale collagen.

[0013] Optionally, the concentration of the EDTA standard solution is 0.3 mol / L, and the material ratio of the fish scale to the EDTA standard solution is 1 g: (40-60) mL.

[0014] Optionally, the material ratio of the decalcified fish scale to the purified water is 1 g: (30-50) mL.

[0015] Another object of the present invention is to provide a fish scale collagen hydrogel, which is prepared by the method for preparing the fish scale collagen hydrogel as described above.

[0016] The beneficial effects of the present invention are: The method for preparing fish scale collagen hydrogel provided by the invention has a simple preparation process, takes fish scale collagen, polymerized monomers and polydopamine nanoparticles as raw materials, combines heat-induced protein denaturation and the concept of three networks, takes fish scale collagen as a first-layer network, and monomer polymers as a second-layer network, adopts a one-pot method to prepare the fish scale collagen-based hydrogel, thereby improving the mechanical properties; and then by adding polydopamine nanoparticles, the mechanical properties are further improved while giving the hydrogel antibacterial properties, and the introduced polydopamine can convert light energy into heat energy, so that the prepared fish scale collagen hydrogel can be used for thermal therapy. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] The present invention is further described below in conjunction with the accompanying drawings and embodiments.

[0018] Figure 1 This is a schematic diagram of the preparation process of fish scale collagen hydrogel in the present invention; Figure 2 It is a schematic diagram of the preparation process of the fish scale collagen solution in the present invention. DETAILED DESCRIPTION

[0019] The present invention is now further described in detail. The embodiments described below are exemplary and intended to be used to explain the present invention, but cannot be understood as limiting the present invention. All other embodiments obtained by ordinary technicians in this field without creative work based on the embodiments of the present invention belong to the scope of protection of the present invention.

[0020] In order to solve the problem of poor mechanical properties of hydrogels in the prior art, the present invention provides a method for preparing a fish scale collagen hydrogel, which comprises the following steps: firstly extracting collagen from fish scales by water extraction, then adding polymerization monomers, a photoinitiator, a cross-linking agent, polydopamine nanoparticles and water, heating and stirring, cooling to room temperature, and then irradiating with ultraviolet light to obtain the fish scale collagen hydrogel.

[0021] During this preparation process, see Figure 1 As shown, fish scale collagen is first added, and then a polymerization monomer, a photoinitiator, a cross-linking agent, polydopamine nanoparticles and an appropriate amount of deionized water are added, and the mixed solution is heated to denature FCP by thermal induction and polymerize to form a first layer of network; by ultraviolet irradiation, N,N'-dimethylacrylamide forms free radicals and further polymerizes to form a double network; at the same time, polydopamine is dispersed in the network, cross-linked with the first layer and the second layer of network, to form a third layer of network entangled with the first layer and the second layer of network, thereby preparing a fish scale collagen hydrogel with a three-layer network structure, and the three-layer network structure is entangled with each other.

[0022] The method for preparing fish scale collagen hydrogel provided by the invention has a simple preparation process. The method uses fish scale collagen, polymerized monomers and polydopamine nanoparticles as raw materials, combines heat-induced protein denaturation, uses fish scale collagen as a first-layer network, monomer polymers as a second-layer network, and polydopamine nanoparticles as a third-layer network, and adopts a one-pot method to prepare a three-network hydrogel based on fish scale collagen, thereby improving the mechanical properties and giving the hydrogel antibacterial properties. In addition, the introduced polydopamine can convert light energy into heat energy, so that the prepared fish scale collagen hydrogel can be used for thermal therapy.

[0023] The present invention adds PDA NPs on the basis of the double network. Since PDA NPs contain amino functional groups and catechol groups, after ultraviolet light irradiation, free radicals can be generated after hydrogen is removed from the amino or hydroxyl functional groups, thereby initiating a series of monomer polymerizations. In addition, the phenolic hydroxyl groups of PDA NPs can be covalently cross-linked with DMAAm through Michael addition, and there is also a Michael addition reaction between the catechol and amide groups in the PDANPs molecules, as well as reversible non-covalent bonding such as hydrogen bonding, and finally the fish scale collagen hydrogel is prepared.

[0024] In the preparation process of the fish scale collagen hydrogel provided by the present invention, the components are all connected to the system through chemical bonds, so that the swelling properties of the hydrogel are not sacrificed while the mechanical properties are improved.

[0025] In addition, the polydopamine introduced in the present invention is connected to the double network hydrogel through the free catechol groups and the reversible non-covalent bonds of the first layer network and the second layer network, and the catechol groups of the polydopamine chains form reversible non-covalent bonds, so that after the prepared fish scale collagen hydrogel is damaged, the fluidity of the molecular chains at the cross section of the hydrogel is improved by near-infrared light irradiation or heating, and the reversible chemical bonds between the polymer chains are reformed, thereby realizing the repair of the hydrogel network and giving the hydrogel certain self-healing properties.

[0026] Moreover, based on the efficient photothermal conversion performance of polydopamine, the fish scale collagen hydrogel is more suitable for self-healing through near-infrared irradiation.

[0027] The present invention preferably heats the mixed solution including: heating the mixed solution to 60-90° C., preferably the heating temperature is 80° C., and the heating time is 15-25 min.

[0028] In order to ensure the mechanical properties of fish scale collagen hydrogel, the preferred polymerization monomer in the present invention is N, N'-dimethylacrylamide, the preferred photoinitiator is 2959, the preferred cross-linking agent is N, N-methylenebisacrylamide, and further preferably the usage ratio of fish scale collagen, polymerization monomer, and polydopamine nanoparticles is (5-12) g: (2-4) mL: (0.01-0.1) g.

[0029] Compared with other collagens, fish scale collagen contains higher levels of hydroxyproline, hydroxylysine and proline, making the triple helix structure tighter, so that fish scale collagen has higher mechanical properties than other collagens, so that the hydrogen bonds and electrostatic effects between the networks of the triple network hydrogel formed by fish scale collagen are stronger. Therefore, the hydrogel prepared by the fish scale collagen in the present invention has better mechanical properties; and, since fish scale collagen contains more hydroxyproline and hydroxylysine than other collagens, it can promote cell growth and tissue repair; fish scale collagen peptide (FSCP) can prevent CoCl2-induced cytotoxicity and TNF α-induced inflammatory response of human HaCaT keratinocytes, giving the fish scale collagen hydrogel the ability to relieve inflammation, and can be used in burns and other aspects.

[0030] See also Figure 2As shown, the preferred fish scale collagen solution of the present invention is prepared according to the following method: placing fish scales in an EDTA standard solution, reacting in a normal temperature water bath under stirring conditions, washing, and obtaining decalcified fish scales; adding the decalcified fish scales to purified water, heating at 60-100° C., preferably at 80° C., and preferably heating for 25 minutes to obtain a crude solution; centrifuging the crude solution and taking the supernatant.

[0031] The supernatant is passed through a water bath and slowly heated at 40-60°C to evaporate some of the water. Then the concentrated collagen solution is poured into a freeze-drying tray and spread into a thin layer. The freeze-drying tray is placed in a freeze dryer and pre-frozen to -40°C to -80°C. The freeze-drying program is started to sublimate the water under vacuum conditions within 24-48 hours. After drying, the freeze-drying tray is removed to obtain a porous collagen solid. Finally, the dried collagen solid is crushed with a grinder and sieved to obtain a uniform fish scale collagen powder.

[0032] Fish scale collagen is extracted from fish scales. Since collagen is highly conserved during biological evolution, the molecular structure of FCP is highly similar to that of mammalian collagen, which provides a basis for FCP to replace mammalian collagen in tissue engineering.

[0033] Specifically, the present invention preferably has a concentration of 0.3 mol / L of EDTA standard solution, and a material ratio of fish scale to EDTA standard solution of 1 g: (40-60) mL; preferably, a material ratio of decalcified fish scale to purified water of 1 g: (30-50) mL.

[0034] Another object of the present invention is to provide a fish scale collagen hydrogel, which is prepared by the method for preparing the fish scale collagen hydrogel as described above.

[0035] The fish scale collagen hydrogel provided by the present invention uses fish scale collagen, polymerized monomers and polydopamine nanoparticles as raw materials, combines heat-induced protein denaturation, uses fish scale collagen as a first-layer network, monomer polymers as a second-layer network, and polydopamine nanoparticles as a third-layer network, and adopts a one-pot method to prepare a three-network hydrogel based on fish scale collagen, which improves the mechanical properties and imparts antibacterial properties to the hydrogel. In addition, the introduced polydopamine can convert light energy into heat energy, so that the prepared fish scale collagen hydrogel can be used for thermal therapy.

[0036] In order to make the above-mentioned objects, features and advantages of the present invention more obvious and easy to understand, specific embodiments of the present invention are described in detail below with reference to the accompanying drawings.

[0037] Unless otherwise specified, the fish scale collagen in each embodiment of the present invention and the comparative example is prepared according to the following method: Add the washed fish scales to a 0.3 mol / L EDTA standard solution at a solid-liquid ratio of 1:50 (g:mL) for decalcification. Fix the beaker in a water bath at room temperature and stir with a magnetic stirrer. After the reaction is complete, take out the fish scales and wash off the EDTA residue on the fish scales with purified water. Then heat the washed and dried fish scales and purified water at a temperature of 80 °C and a solid-liquid ratio of 1:40 (g:mL) for 90 min to obtain a relevant solution. After cooling the above solution at room temperature, pour it into a centrifuge tube, centrifuge it at 8000 rpm for 15 min, take the supernatant, pour it into a beaker for later use.

[0038] The supernatant is passed through a water bath and slowly heated at 50°C to evaporate some of the water. Then the concentrated collagen solution is poured into a freeze-drying tray and spread into a thin layer. The freeze-drying tray is placed in a freeze dryer and pre-frozen to -40°C to -80°C. The freeze-drying program is started to sublimate the water under vacuum conditions within 36 hours. After drying, the freeze-drying tray is removed to obtain a porous collagen solid. Finally, the dried collagen solid is crushed with a grinder and sieved to obtain a uniform powder.

[0039] Example 1 This embodiment provides a method for preparing fish scale collagen hydrogel, which comprises the following steps: Weigh 8.8g of fish scale collagen, add 2 mL of N,N'-dimethylacrylamide, 0.08 g of photoinitiator 2959, 0.004 g of MBA, and then add 0.01g of polydopamine nanoparticles, add appropriate amount of deionized water, and prepare 100ml solution. Heat and stir at 80℃ for 20min, cool to room temperature, and then place it under a 365nm ultraviolet lamp for ten minutes to obtain fish scale collagen tri-network hydrogel.

[0040] Example 2 This embodiment provides a method for preparing fish scale collagen hydrogel, which comprises the following steps: Weigh 5g of fish scale collagen, add 3 mL of N,N'-dimethylacrylamide, 0.08 g of photoinitiator 2959, and 0.004 g of MBA to make a solution, then add 0.05g of polydopamine nanoparticles and appropriate amount of deionized water to make a 100ml solution. Heat and stir at 80℃ for 20min, cool to room temperature, and then place it under a 365nm ultraviolet lamp for ten minutes to obtain fish scale collagen tri-network hydrogel.

[0041] Example 3 This embodiment provides a method for preparing fish scale collagen hydrogel, which comprises the following steps: Weigh 12g of fish scale collagen, add 4 mL of N,N'-dimethylacrylamide, 0.1 g of photoinitiator 2959, and 0.008 g of MBA to make a solution, then add 0.1g of polydopamine nanoparticles and appropriate amount of deionized water to make a 100ml solution. Heat and stir at 80℃ for 20min, cool to room temperature, and then place it under a 365nm ultraviolet lamp for ten minutes to obtain fish scale collagen tri-network hydrogel.

[0042] Each comparative example of the present invention is compared with Example 1.

[0043] Comparative Example 1 This comparative example provides a method for preparing fish scale collagen hydrogel, which comprises the following steps: Weigh 8.8 g of fish scale collagen, add deionized water to make a 100 ml solution, heat and stir at 80°C for 20 min, and cool to room temperature to form fish scale collagen hydrogel.

[0044] Comparative Example 2 This comparative example provides a method for preparing a poly N,N'-dimethylacrylamide hydrogel, which comprises the following steps: 2 mL of N,N'-dimethylacrylamide and 8 mL of distilled water were measured successively, 0.08 g of photoinitiator 2959 and 0.004 g of N,N-methylenebisacrylamide were weighed, mixed and irradiated with 365 nm ultraviolet light for ten minutes to obtain poly N,N'-dimethylacrylamide hydrogel.

[0045] Comparative Example 3 This comparative example provides a method for preparing fish scale collagen hydrogel, which comprises the following steps: Weigh 8.8g of fish scale collagen, add 2 mL of N,N'-dimethylacrylamide, 0.08 g of photoinitiator 2959, 0.004 g of MBA, and appropriate amount of deionized water to make a 100ml solution. Heat and stir at 80℃ for 20min, cool to room temperature, and then place it under a 365nm ultraviolet lamp for ten minutes to obtain fish scale collagen hydrogel.

[0046] The hydrogels prepared in the above embodiments and comparative examples were tested for performance, and the testing method was as follows: The tensile and compression tests were carried out using a universal material testing machine. Each sample was tested at least 3 times.

[0047] Tensile test: We cut the samples into regular strips and recorded the thickness, set the gauge length to 15 mm, and performed the tensile test at a tensile rate of 100 mm / min.

[0048] Compression test: We cut the samples into regular blocks, set the gauge length to 20 mm, and performed compression tests at a compression rate of 50 mm / min.

[0049] The test results are shown in Table 1: Table 1 Tensile strength (KPa) Compression strength (MPa) Toughness (KJ / m3) Elongation at break (%) Example 1 971 2.94 161 881 Example 2 758 2.70 137 733 Example 3 844 2.41 106 719 Comparative Example 1 418 1.62 27 315 Comparative Example 2 354 1.73 80 448 Comparative Example 3 721 2.46 114 604 Based on the above ideal embodiments of the present invention, the relevant staff can make various changes and modifications without departing from the technical concept of the present invention through the above description. The technical scope of the present invention is not limited to the contents of the specification, and its technical scope must be determined according to the scope of the claims.

Claims

1. A method for preparing fish scale collagen hydrogel, characterized in that: The method comprises the following steps: adding fish scale collagen, polymerization monomer, photoinitiator, cross-linking agent, polydopamine nanoparticles and water into a reactor, heating and stirring, cooling to room temperature, and irradiating with ultraviolet light to obtain fish scale collagen hydrogel.

2. The method for preparing fish scale collagen hydrogel according to claim 1, characterized in that: Heating the mixed solution includes: heating the mixed solution to 60-90°C.

3. The method for preparing the fish scale collagen hydrogel according to claim 1, characterized in that: The polymerizable monomer is N,N'-dimethylacrylamide.

4. The method for preparing the fish scale collagen hydrogel according to claim 1, characterized in that: The photoinitiator is 2959.

5. The method for preparing the fish scale collagen hydrogel according to claim 1, characterized in that: The cross-linking agent is N,N-methylenebisacrylamide.

6. The method for preparing the fish scale collagen hydrogel according to claim 1, characterized in that: The usage ratio of the fish scale collagen, the polymerized monomer, and the polydopamine nanoparticles is (5-12) g: (2-4) mL: (0.01-0.1) g.

7. The method for preparing the fish scale collagen hydrogel according to any one of claims 1 to 6, characterized in that: The fish scale collagen is prepared according to the following method: placing fish scales in an EDTA standard solution, reacting in a normal temperature water bath under stirring conditions, washing, and obtaining decalcified fish scales; adding the decalcified fish scales into purified water, heating at 60-100° C., and obtaining a crude solution; The crude solution is centrifuged, and the supernatant is taken, evaporated, freeze-dried, and pulverized to obtain the fish scale collagen.

8. The method for preparing fish scale collagen hydrogel according to claim 7, characterized in that: The concentration of the EDTA standard solution is 0.3 mol / L, and the material ratio of the fish scale to the EDTA standard solution is 1 g: (40-60) mL.

9. The method for preparing the fish scale collagen hydrogel according to claim 7, characterized in that: The material ratio of the decalcified fish scale to the purified water is 1 g: (30-50) mL.

10. A fish scale collagen hydrogel, characterized in that: The method is used to prepare the fish scale collagen hydrogel according to any one of claims 1 to 9.

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