Selenium-loaded photocrosslinked alginate hydrogel and preparation method thereof

Abstract

The present invention discloses a selenium-loaded photocross-linked alginate hydrogel and a preparation method thereof, and relates to the technical field of polymer gels. The preparation method is as follows: first, alginate is dissolved in deionized water, sodium chloride and a buffer are added and stirred until completely dissolved, the pH is adjusted to weak acidity, N-hydroxy-sulfosuccinimide and a condensing agent are added, and after stirring evenly, 2-aminoethyl methacrylate hydrochloride is added to carry out a methacrylate reaction to obtain methacrylated alginate. The alginate and a photoinitiator are separately dissolved in deionized water to prepare a storage solution, and selenomethionine is dissolved in the photoinitiator storage solution. Finally, a photoinitiator solution containing selenomethionine and a methacrylated alginate storage solution are mixed, and a photocross-linking reaction is initiated by ultraviolet irradiation to obtain a selenium-loaded photocross-linked alginate hydrogel. The present invention introduces selenomethionine into a photocross-linked alginate hydrogel scaffold, which has good cell biocompatibility and promotes osteogenesis.

Description

Technical Field

[0001] The invention relates to the technical field of polymer gels, in particular to a selenium-loaded photocrosslinked alginate hydrogel and a preparation method thereof. Background Art

[0002] Amidst the ongoing development of biomedical materials, hydrogels have become a research hotspot due to their unique properties. Hydrogels possess high hydration capacity, allowing them to absorb large amounts of water while maintaining their structural stability. This property contributes to their excellent biocompatibility and effectively reduces irritation to surrounding tissues. Their three-dimensional network structure not only provides an extracellular matrix-like microenvironment for cell adhesion, proliferation, and differentiation, but also serves as a carrier for the loading and release of drugs, bioactive molecules, and other substances. Hydrogels exhibit significant potential for application in a variety of medical fields, including drug delivery, wound dressings, wound repair, and tissue engineering.

[0003] Alginate, a natural polymer extracted from marine algae, plays an important role in hydrogel preparation due to its outstanding biodegradability and biocompatibility. Alginate hydrogels gradually degrade in the body through enzymatic or hydrolytic processes. The degradation products are non-toxic small molecules that are naturally metabolized and excreted by the human body without accumulating in the body and causing adverse effects. Furthermore, the abundant active groups on alginate molecules can interact with receptors on the cell surface, promoting cell adhesion and growth, and providing physical support and bioactive signals for tissue repair.

[0004] In recent years, research on the trace element selenium in the biomedical field has yielded fruitful results. Selenium is an essential trace element for the human body and participates in numerous important physiological processes within the body. From an antioxidant perspective, selenium is a key component of antioxidant enzymes such as glutathione peroxidase. It catalyzes reduction reactions, scavenges excess reactive oxygen species and free radicals in the body, protects cells from oxidative damage, and maintains intracellular redox balance. In terms of immunomodulation, selenium has a significant impact on the development, maturation, and function of immune cells. It can enhance the activity of T lymphocytes, B lymphocytes, and macrophages, regulate the synthesis and secretion of immunoglobulins, and enhance the body's immune response. During inflammation, selenium can inhibit the release of inflammatory cytokines such as tumor necrosis factor and interleukins, reduce the infiltration of inflammatory cells, and thus exert an anti-inflammatory effect. Specifically in bone metabolism, selenium regulates the activity of osteoblasts and osteoclasts, promoting osteoblast proliferation and differentiation, inhibiting excessive osteoclast resorption, and maintaining normal bone metabolism and remodeling. It plays an indispensable role in bone growth, repair, and maintenance of bone mass.

[0005] However, despite some progress in the research and application of hydrogels, such as the existing preparation methods of drug-loaded hydrogels, which have opened up new avenues for hydrogel drug delivery, the introduction of selenium has not been effectively studied in many studies of drug-loaded hydrogels. Summary of the Invention

[0006] The present invention aims to provide a selenium-loaded photocrosslinked alginate hydrogel and its preparation method to address the technical problems raised by the aforementioned background art. The present invention introduces selenomethionine into the photocrosslinked alginate hydrogel scaffold, facilitating cell adhesion and expansion on its surface, enhancing the osteogenic activity of osteoblasts, and exhibiting a significant osteogenesis-promoting effect.

[0007] To achieve the above object, the present invention provides the following technical solutions:

[0008] A method for preparing selenium-loaded photocrosslinked alginate hydrogel comprises the following steps:

[0009] S1. Dissolving alginate in deionized water, adding sodium chloride and a buffer, stirring until completely dissolved, adjusting the pH of the solution to weak acidity, then adding N-hydroxy-sulfosuccinimide and a condensing agent, stirring evenly, adding 2-aminoethyl methacrylate hydrochloride, and performing a methacrylate reaction to obtain methacrylated alginate;

[0010] S2, dissolving methacrylated alginate in deionized water to obtain a methacrylated alginate storage solution;

[0011] S3, dissolving the photoinitiator in deionized water to obtain a photoinitiator storage solution, and dissolving selenomethionine in the photoinitiator storage solution to obtain a selenomethionine-containing photoinitiator solution;

[0012] S4. Mixing a selenomethionine-containing photoinitiator solution with a methacrylated alginate storage solution to obtain a precursor solution, irradiating the precursor solution with ultraviolet light to initiate a photocrosslinking reaction, and preparing a selenium-loaded photocrosslinked alginate hydrogel.

[0013] Preferably, in step S1, the alginate is selected from one or more of sodium alginate, potassium alginate, and ammonium alginate.

[0014] Preferably, in step S1, the buffer is 2-(N-morpholino)ethanesulfonic acid.

[0015] Preferably, in step S1, the condensing agent is 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride.

[0016] Preferably, in step S1, the mass ratio of alginate to 2-aminoethyl methacrylate hydrochloride is 6:2-4.

[0017] Preferably, in step S3, the photoinitiator is 2-hydroxy-4-(2-hydroxyethoxy)-2-methylpropiophenone.

[0018] In technical solution of the present invention, use alginate as basic raw material, for hydrogel builds basic skeleton structure, utilize its good biodegradability and biocompatibility to build hydrogel basic skeleton, in vivo, hydrogel can be gradually degraded, and degradation products are nontoxic and harmless, can be discharged by human metabolism, can not produce long-term adverse effects to body, and simultaneously with surrounding tissue affinity high, is conducive to tissue repair and regeneration, reduces the generation of immune rejection reaction. Sodium chloride is used to regulate the ionic strength of solution, maintains the stability of reaction system. Buffer 2-(N-morpholine) ethanesulfonic acid, the pH value of solution is maintained in weakly acidic range, to ensure that subsequent carboxyl activation reaction can be carried out under suitable pH. N-hydroxy-sulfosuccinimide and condensing agent 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride synergistically, activate the carboxyl on the alginate molecule, improve its reactive behavior, be convenient to react with 2-aminoethyl methacrylate hydrochloride, make alginate molecular chain introduce methacrylate group, possess photocrosslinking ability. Photoinitiators such as 2-hydroxy-4-(2-hydroxyethoxy)-2-methylpropiophenone decompose under ultraviolet irradiation to produce free radicals, which trigger the polymerization reaction of double bonds on the methacrylated alginate molecular chain to achieve photocrosslinking. Selenomethionine is a selenium-containing amino acid with multiple biological activities such as antioxidant, immunomodulatory, and osteogenesis promotion. Through the crosslinking reaction of methacrylated alginate, selenomethionine is loaded inside the crosslinked alginate hydrogel to obtain selenium-loaded photocrosslinked alginate hydrogel. The reaction flow chart for the preparation of selenium-loaded photocrosslinked alginate hydrogel is shown in Figure 1 .

[0019] The introduction of selenomethionine into the hydrogel changes the surface properties of the hydrogel. Selenomethionine not only has biological activity itself, but also can work synergistically with other ingredients to promote cell adhesion and extension on the surface of the hydrogel. The lack of bone inductivity is a key problem that traditional photo-cross-linked alginate hydrogels are difficult to meet the clinical needs of bone defect repair. The present invention hopes to effectively solve this problem with the help of the biological activity of selenomethionine. Selenomethionine can enhance the antioxidant and anti-inflammatory responses of osteoblasts through signal pathways, thereby promoting the osteogenic differentiation and mineralization ability of osteoblasts. Both in vivo and in vitro experiments have confirmed that selenomethionine-loaded photo-cross-linked alginate hydrogels can significantly increase the osteogenic activity of osteoblasts. In the rat skull defect experiment, the hydrogel of the present invention was implanted, and Micro-CT detection found that the bone density at the bone defect was significantly increased, and the amount of new bone tissue generated was significantly more than that of the control group, which fully demonstrated its excellent effect in promoting osteogenesis.

[0020] Preferably, in step S4, the mass of selenomethionine is 1-6% of the mass of methacrylated alginate.

[0021] In the technical solution of the present invention, as described above, by introducing selenomethionine into alginate hydrogel, the mechanical strength of the hydrogel and the osteogenesis-promoting effect of the hydrogel can be significantly improved. In order to achieve the above technical effects, alginate hydrogel must be combined with a sufficient amount of selenomethionine. Therefore, the invention controls the mass of selenomethionine to be more than 1% of the mass of methacrylated alginate. However, as the amount of selenomethionine continues to increase, the present invention team unexpectedly discovered that when an excess of selenomethionine is combined with the alginate hydrogel, that is, the mass of selenomethionine is more than 6% of the mass of methacrylated alginate, the mechanical properties (tensile strength) of the alginate hydrogel scaffold suddenly drop significantly, making it difficult to withstand the physiological stress during bone tissue repair, and easily deforming and breaking in the body, and unable to provide a stable support structure for bone tissue regeneration. This may be because selenomethionine has a certain molecular structure and size, and too much selenomethionine will occupy a larger space in the reaction system. In the cross-linking reaction, the molecules participating in the reaction need to be close to each other and reach a suitable spatial orientation to react. Too much selenomethionine molecules will spatially hinder other substances participating in the cross-linking reaction from being close to each other, increasing the steric hindrance of the reaction, and then affecting the normal formation of the cross-linked network, causing its mechanical properties to decline. Therefore, the present invention simultaneously controls the quality of selenomethionine to be less than 6% of the quality of methacrylated alginate. The osteogenesis promoting effect and good mechanical properties of the alginate hydrogel scaffold are balanced by controlling the addition of selenomethionine.

[0022] A selenium-loaded photocrosslinked alginate hydrogel is prepared by the above method.

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

[0024] 1. This invention introduces selenomethionine into photocross-linked alginate hydrogels, effectively addressing the lack of osteoinductivity found in conventional gels. Selenomethionine enhances the antioxidant and anti-inflammatory responses of osteoblasts through specific signaling pathways, significantly promoting osteoblast differentiation and mineralization, and exhibiting a potent osteogenesis-promoting effect.

[0025] 2. Using alginate as the base material, the hydrogel's basic framework is constructed, leveraging its excellent biodegradability and biocompatibility. In the body, the hydrogel gradually degrades, and the degradation products are non-toxic and harmless, metabolized and excreted by the body without causing long-term adverse effects. At the same time, it has a high affinity with surrounding tissues, facilitating tissue repair and regeneration, and reducing the occurrence of immune rejection reactions.

[0026] 3. By precisely controlling the amount of selenomethionine added, the osteogenesis-promoting effect and mechanical properties of the hydrogel scaffold are effectively balanced. This ensures that selenomethionine exerts its osteogenesis-promoting bioactivity while also ensuring that the hydrogel possesses sufficient mechanical strength to withstand the physiological stresses of bone tissue repair, providing a stable support structure for bone tissue regeneration. This meets diverse clinical needs and improves the adaptability and effectiveness of the hydrogel in bone tissue engineering applications. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 The present invention is a reaction flow chart for preparing the selenium-loaded photocrosslinked alginate hydrogel.

[0028] Figure 2 This is a microscope image of rat bone marrow stromal stem cells encapsulated in the hydrogel of Comparative Example 1 stained with ALP staining solution after being cultured in vitro for 14 days.

[0029] Figure 3 This is a microscope image of rat bone marrow stromal stem cells encapsulated in the hydrogel according to Example 1 stained with ALP staining solution after being cultured in vitro for 14 days.

[0030] Figure 4 This is a microscope image of rat bone marrow stromal stem cells encapsulated in hydrogel according to comparative example 1 stained with ARS staining solution after being cultured in vitro for 21 days.

[0031] Figure 5 This is a microscope image of rat bone marrow stromal stem cells encapsulated in the hydrogel according to Example 1 stained with ARS staining solution after being cultured in vitro for 21 days.

[0032] Figure 6 Micro-CT images of the blank control group hydrogel repairing rat skull.

[0033] Figure 7 This is a Micro-CT image of the rat skull repaired by the hydrogel in comparative example 1.

[0034] Figure 8 This is a Micro-CT image of the rat skull repaired by the hydrogel in Example 1. DETAILED DESCRIPTION

[0035] The following is a clear and complete description of the technical solutions in the embodiments of the present invention. Obviously, the implementation regulations described are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts are within the scope of protection of the present invention.

[0036] Example 1

[0037] A method for preparing selenium-loaded photocrosslinked alginate hydrogel comprises the following steps:

[0038] Step S1: At room temperature, 4 g of sodium alginate was dissolved in 400 mL of deionized water, 11.69 g of sodium chloride and 3.91 g of 2-(N-morpholine)ethanesulfonic acid buffer were added, and the mixture was magnetically stirred overnight until completely dissolved. The solution was titrated with 5 mol / L sodium hydroxide to a pH of 6.5, and then 1.06 g of N-hydroxy-sulfosuccinimide and 3.5 g of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride condensation agent were added respectively. After stirring for 5 min, 2.28 g of 2-aminoethyl methacrylate hydrochloride was added. The mixture was reacted in the dark for 24 h, and then the reaction solution was poured into 2000 mL of acetone to precipitate the product methacrylated alginate, and filtered through a Buchner funnel. The product was redissolved in 400 ml of deionized water. After the product was completely dissolved, the solution was aliquoted into dialysis bags (molecular weight 3500 Da) and dialyzed against deionized water for three days, with the deionized water replaced every 12 hours. After dialysis, the solution was filtered twice using a filter membrane (pore size 0.22 μm) and then freeze-dried using a freeze dryer to obtain methacrylated alginate.

[0039] Step S2: Weigh 12 g of methacrylated alginate, add it to 480 mL of deionized water, and stir until it is completely dissolved to obtain a methacrylated alginate stock solution.

[0040] Step S3: Dissolve 0.5 g of the photoinitiator 2-hydroxy-4-(2-hydroxyethoxy)-2-methylpropiophenone in 100 mL of deionized water and stir until completely dissolved to obtain a photoinitiator stock solution. Dissolve 0.63 g of selenomethionine in the photoinitiator stock solution and stir evenly to obtain a selenomethionine-containing photoinitiator solution.

[0041] Step S4: A selenomethionine-containing photoinitiator solution is mixed with a methacrylated alginate stock solution to obtain a precursor solution. The precursor solution is poured into a specific mold and irradiated with an ultraviolet lamp (wavelength 365 nm, intensity 1.7 mW / cm²) for 15 minutes to initiate a photocrosslinking reaction, thereby producing a selenium-loaded photocrosslinked alginate hydrogel.

[0042] Example 2

[0043] A method for preparing selenium-loaded photocrosslinked alginate hydrogel comprises the following steps:

[0044] Step S1: At room temperature, 4 g of sodium alginate was dissolved in 400 mL of deionized water, 11.69 g of sodium chloride and 3.91 g of 2-(N-morpholine)ethanesulfonic acid buffer were added, and the mixture was magnetically stirred overnight until completely dissolved. The solution was titrated with 5 mol / L sodium hydroxide to a pH of 6.5, and then 1.06 g of N-hydroxy-sulfosuccinimide and 3.5 g of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride condensation agent were added respectively. After stirring for 5 min, 2.28 g of 2-aminoethyl methacrylate hydrochloride was added. The mixture was reacted in the dark for 24 h, and then the reaction solution was poured into 2000 mL of acetone to precipitate the product methacrylated alginate, and filtered through a Buchner funnel. The product was redissolved in 400 ml of deionized water. After the product was completely dissolved, the solution was aliquoted into dialysis bags (molecular weight 3500 Da) and dialyzed against deionized water for three days, with the deionized water replaced every 12 hours. After dialysis, the solution was filtered twice using a filter membrane (pore size 0.22 μm) and then freeze-dried using a freeze dryer to obtain methacrylated alginate.

[0045] Step S2: Weigh 12 g of methacrylated alginate, add it to 480 mL of deionized water, and stir until it is completely dissolved to obtain a methacrylated alginate stock solution.

[0046] Step S3: Dissolve 0.5 g of the photoinitiator 2-hydroxy-4-(2-hydroxyethoxy)-2-methylpropiophenone in 100 mL of deionized water and stir until completely dissolved to obtain a photoinitiator stock solution. Dissolve 0.26 g of selenomethionine in the photoinitiator stock solution and stir evenly to obtain a selenomethionine-containing photoinitiator solution.

[0047] Step S4: A selenomethionine-containing photoinitiator solution is mixed with a methacrylated alginate stock solution to obtain a precursor solution. The precursor solution is poured into a specific mold and irradiated with an ultraviolet lamp (wavelength 365 nm, intensity 1.7 mW / cm²) for 15 minutes to initiate a photocrosslinking reaction, thereby producing a selenium-loaded photocrosslinked alginate hydrogel.

[0048] Example 3

[0049] A method for preparing selenium-loaded photocrosslinked alginate hydrogel comprises the following steps:

[0050] Step S1: At room temperature, 4 g of sodium alginate was dissolved in 400 mL of deionized water, 11.69 g of sodium chloride and 3.91 g of 2-(N-morpholine)ethanesulfonic acid buffer were added, and the mixture was magnetically stirred overnight until completely dissolved. The solution was titrated with 5 mol / L sodium hydroxide to a pH of 6.5, and then 1.06 g of N-hydroxy-sulfosuccinimide and 3.5 g of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride condensation agent were added respectively. After stirring for 5 min, 2.28 g of 2-aminoethyl methacrylate hydrochloride was added. The mixture was reacted in the dark for 24 h, and then the reaction solution was poured into 2000 mL of acetone to precipitate the product methacrylated alginate, and filtered through a Buchner funnel. The product was redissolved in 400 ml of deionized water. After the product was completely dissolved, the solution was aliquoted into dialysis bags (molecular weight 3500 Da) and dialyzed against deionized water for three days, with the deionized water replaced every 12 hours. After dialysis, the solution was filtered twice using a filter membrane (pore size 0.22 μm) and then freeze-dried using a freeze dryer to obtain methacrylated alginate.

[0051] Step S2: Weigh 12 g of methacrylated alginate, add it to 480 mL of deionized water, and stir until it is completely dissolved to obtain a methacrylated alginate stock solution.

[0052] Step S3: Dissolve 0.5 g of the photoinitiator 2-hydroxy-4-(2-hydroxyethoxy)-2-methylpropiophenone in 100 mL of deionized water and stir until completely dissolved to obtain a photoinitiator stock solution. Dissolve 0.53 g of selenomethionine in the photoinitiator stock solution and stir evenly to obtain a selenomethionine-containing photoinitiator solution.

[0053] Step S4: A selenomethionine-containing photoinitiator solution is mixed with a methacrylated alginate stock solution to obtain a precursor solution. The precursor solution is poured into a specific mold and irradiated with an ultraviolet lamp (wavelength 365 nm, intensity 1.7 mW / cm²) for 15 minutes to initiate a photocrosslinking reaction, thereby producing a selenium-loaded photocrosslinked alginate hydrogel.

[0054] Example 4

[0055] A method for preparing selenium-loaded photocrosslinked alginate hydrogel comprises the following steps:

[0056] Step S1: At room temperature, 4 g of sodium alginate was dissolved in 400 mL of deionized water, 11.69 g of sodium chloride and 3.91 g of 2-(N-morpholine)ethanesulfonic acid buffer were added, and the mixture was magnetically stirred overnight until completely dissolved. The solution was titrated with 5 mol / L sodium hydroxide to a pH of 6.5, and then 1.06 g of N-hydroxy-sulfosuccinimide and 3.5 g of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride condensation agent were added respectively. After stirring for 5 min, 2.28 g of 2-aminoethyl methacrylate hydrochloride was added. The mixture was reacted in the dark for 24 h, and then the reaction solution was poured into 2000 mL of acetone to precipitate the product methacrylated alginate, and filtered through a Buchner funnel. The product was redissolved in 400 ml of deionized water. After the product was completely dissolved, the solution was aliquoted into dialysis bags (molecular weight 3500 Da) and dialyzed against deionized water for three days, with the deionized water replaced every 12 hours. After dialysis, the solution was filtered twice using a filter membrane (pore size 0.22 μm) and then freeze-dried using a freeze dryer to obtain methacrylated alginate.

[0057] Step S2: Weigh 12 g of methacrylated alginate, add it to 480 mL of deionized water, and stir until it is completely dissolved to obtain a methacrylated alginate stock solution.

[0058] Step S3: Dissolve 0.5 g of the photoinitiator 2-hydroxy-4-(2-hydroxyethoxy)-2-methylpropiophenone in 100 mL of deionized water and stir until completely dissolved to obtain a photoinitiator stock solution. Dissolve 0.72 g of selenomethionine in the photoinitiator stock solution and stir evenly to obtain a selenomethionine-containing photoinitiator solution.

[0059] Step S4: A selenomethionine-containing photoinitiator solution is mixed with a methacrylated alginate stock solution to obtain a precursor solution. The precursor solution is poured into a specific mold and irradiated with an ultraviolet lamp (wavelength 365 nm, intensity 1.7 mW / cm²) for 15 minutes to initiate a photocrosslinking reaction, thereby producing a selenium-loaded photocrosslinked alginate hydrogel.

[0060] Example 5

[0061] A method for preparing selenium-loaded photocrosslinked alginate hydrogel comprises the following steps:

[0062] Step S1: At room temperature, 4 g of sodium alginate was dissolved in 400 mL of deionized water, 11.69 g of sodium chloride and 3.91 g of 2-(N-morpholine)ethanesulfonic acid buffer were added, and the mixture was magnetically stirred overnight until completely dissolved. The solution was titrated with 5 mol / L sodium hydroxide to a pH of 6.5, and then 1.06 g of N-hydroxy-sulfosuccinimide and 3.5 g of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride condensation agent were added respectively. After stirring for 5 min, 2.28 g of 2-aminoethyl methacrylate hydrochloride was added. The mixture was reacted in the dark for 24 h, and then the reaction solution was poured into 2000 mL of acetone to precipitate the product methacrylated alginate, and filtered through a Buchner funnel. The product was redissolved in 400 ml of deionized water. After the product was completely dissolved, the solution was aliquoted into dialysis bags (molecular weight 3500 Da) and dialyzed against deionized water for three days, with the deionized water replaced every 12 hours. After dialysis, the solution was filtered twice using a filter membrane (pore size 0.22 μm) and then freeze-dried using a freeze dryer to obtain methacrylated alginate.

[0063] Step S2: Weigh 12 g of methacrylated alginate, add it to 480 mL of deionized water, and stir until it is completely dissolved to obtain a methacrylated alginate stock solution.

[0064] Step S3: Dissolve 0.5 g of the photoinitiator 2-hydroxy-4-(2-hydroxyethoxy)-2-methylpropiophenone in 100 mL of deionized water and stir until completely dissolved to obtain a photoinitiator stock solution. Dissolve 0.12 g of selenomethionine in the photoinitiator stock solution and stir evenly to obtain a selenomethionine-containing photoinitiator solution.

[0065] Step S4: A selenomethionine-containing photoinitiator solution is mixed with a methacrylated alginate stock solution to obtain a precursor solution. The precursor solution is poured into a specific mold and irradiated with an ultraviolet lamp (wavelength 365 nm, intensity 1.7 mW / cm²) for 15 minutes to initiate a photocrosslinking reaction, thereby producing a selenium-loaded photocrosslinked alginate hydrogel.

[0066] Comparative Example 1

[0067] The difference between Comparative Example 1 and Example 1 is that selenomethionine is not loaded in the cross-linked alginate hydrogel, and the other steps are the same.

[0068] Comparative Example 2

[0069] The difference between Comparative Example 2 and Example 5 is that the mass of selenomethionine is 0.5% of the mass of methacrylated alginate, and the other steps are the same.

[0070] Comparative Example 3

[0071] The difference between Comparative Example 3 and Example 4 is that the mass of selenomethionine is 7% of the mass of methacrylated alginate, and the other steps are the same.

[0072] Comparative Example 4

[0073] The difference between Comparative Example 4 and Example 4 is that the mass of selenomethionine is 8% of the mass of methacrylated alginate, and the other steps are the same.

[0074] Performance testing:

[0075] 1. Test of promoting osteogenesis performance:

[0076] (1) The rat bone marrow stromal stem cells were encapsulated with the hydrogels prepared in Example 1 and Comparative Example 1. After 14 days of in vitro culture, the cells were stained with ALP staining solution for 2 hours and observed and recorded under a microscope. After 21 days of in vitro culture, the cells were stained with ARS staining solution for 30 minutes, rinsed gently with pure water twice, and observed and recorded under a microscope. Microscopic observation images are shown in Figure 2-5 .from Figure 2-5 It can be seen that the ALP and ARS staining areas and quantities in Example 1 are significantly greater than those in Comparative Example 1, demonstrating that the hydrogel of the present invention has good osteogenesis-promoting performance.

[0077] (2) A 5mm dental trephine was used to create bone defects in rat skulls. The bone defects were rinsed with physiological saline and the hydrogels of Example 1 and Comparative Example 1 were implanted. A group without hydrogel implantation was set up as a blank control group. After 8 weeks of culture, the experimental animals were killed and skull specimens were obtained. Micro-CT was used to detect the bone density of each group of bone defects to evaluate and compare the repair effect of the defect. Micro-CT images are shown in Figure 6-8 .from Figure 6-8 It can be seen that the repair area of ​​rat skull by the hydrogel in Example 1 is significantly larger than that in Comparative Example 1 and the blank control group, which proves that the hydrogel of the present invention has good osteogenesis-promoting performance.

[0078] 2. Mechanical properties test:

[0079] In order to accurately evaluate the mechanical properties of selenium-loaded photocross-linked alginate hydrogels, 5 rectangular specimens with a size of 20 mm in length, 5 mm in width, and 2 mm in thickness were cut from the hydrogels of Examples 1-5 and Comparative Examples 1-4. The test was carried out using an electronic universal material testing machine with an accuracy of 0.01N. The specimens were mounted on the fixtures of the testing machine, the fixture spacing was adjusted to 10 mm, and the stretching speed was set to 10 mm / min. During the stretching process, the testing machine automatically records the force-displacement data until the specimen breaks. Each sample was tested 5 times and the average value was taken. The tensile strength is calculated by the formula σ=Fmax / S (where σ is the tensile strength (MPa), Fmax is the maximum force (N) when the specimen breaks, and S is the initial cross-sectional area of ​​the specimen. In this experiment, S=5mm×2mm=10mm 2 ). The tensile strength (MPa) of the samples was calculated according to the above formula. The calculation results are shown in Table 1.

[0080] Table 1:

[0081]

[0082] The data show that in Examples 1-5, where the mass of selenomethionine accounted for 1%-6% of the mass of the methacrylated alginate, the hydrogel tensile strength ranged from 1.50 to 1.68 MPa, showing relative stability. These values ​​were higher than the 1.41 MPa achieved in Comparative Example 1 (unloaded with selenomethionine), demonstrating that the appropriate addition of selenomethionine can improve the mechanical strength of the hydrogel. In Comparative Examples 3 and 4, where the mass of selenomethionine accounted for 7% and 8%, respectively, the tensile strength dropped significantly to 1.03 MPa and 0.82 MPa, demonstrating that when the mass of selenomethionine exceeds 6% of the mass of the methacrylated alginate, the mechanical properties of the hydrogel are significantly reduced.

[0083] Although the present invention has been described in detail with reference to the aforementioned embodiments, it is still possible for those skilled in the art to modify the technical solutions described in the aforementioned embodiments, or to make equivalent substitutions for some of the technical features therein. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. A method for preparing selenium-loaded photocrosslinked alginate hydrogel, characterized in that: The following steps are involved: S1. Dissolve alginate in deionized water, add sodium chloride and a buffer, stir until completely dissolved, adjust the pH of the solution to weak acidity, then add N-hydroxy-sulfosuccinimide and a condensing agent, stir evenly, add 2-aminoethyl methacrylate hydrochloride, and carry out methacrylate esterification reaction, wherein the mass ratio of alginate to 2-aminoethyl methacrylate hydrochloride is 6:2-4, to obtain methacrylated alginate; S2, dissolving methacrylated alginate in deionized water to obtain a methacrylated alginate storage solution; S3, dissolving the photoinitiator in deionized water to obtain a photoinitiator storage solution, and dissolving selenomethionine in the photoinitiator storage solution to obtain a selenomethionine-containing photoinitiator solution; S4. Mix a photoinitiator solution containing selenomethionine with a methacrylated alginate storage solution, wherein the mass of selenomethionine is 1-6% of the mass of methacrylated alginate, to obtain a precursor solution. Irradiate the precursor solution with ultraviolet light to initiate a photocrosslinking reaction, thereby preparing a selenium-loaded photocrosslinked alginate hydrogel.

2. The method for preparing a selenium-loaded photocrosslinked alginate hydrogel according to claim 1, characterized in that: In step S1, the alginate is selected from one or more of sodium alginate, potassium alginate, and ammonium alginate.

3. The method for preparing a selenium-loaded photocrosslinked alginate hydrogel according to claim 1, characterized in that: In step S1, the buffer is 2-(N-morpholino)ethanesulfonic acid.

4. The method for preparing a selenium-loaded photocrosslinked alginate hydrogel according to claim 1, characterized in that: In the step S1, the condensing agent is 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride.

5. The method for preparing a selenium-loaded photocrosslinked alginate hydrogel according to claim 1, characterized in that: In step S3, the photoinitiator is 2-hydroxy-4-(2-hydroxyethoxy)-2-methylpropiophenone.

6. A selenium-loaded photocrosslinked alginate hydrogel, characterized in that: The method is prepared by any one of claims 1 to 5.

Images