Dynamic oxygen release microsphere capable of eliminating active oxygen as well as preparation method and application of dynamic oxygen release microsphere

Abstract

The invention discloses a dynamic oxygen release microsphere capable of eliminating active oxygen and a preparation method and application thereof, the microsphere sequentially comprises an outer layer shell, an inner layer shell and a core from outside to inside, the core is calcium peroxide, the inner layer shell is gelatin, the outer layer shell is also crosslinked on the outer side of the inner layer shell, and nano cerium oxide is also uniformly dispersed in the outer layer shell. According to the oxygen deficit rule in the remediation process, the oxygen release rate can be dynamically changed by changing the mass ratio of carboxymethyl chitosan to sodium alginate, proper oxygen is supplied in all stages of remediation, meanwhile, possible active oxygen is removed, and oxidative stress is prevented. Compared with the prior art, not only has the characteristics of good stability, biocompatibility, degradability and the like, but also the outer shell has a certain hemostatic effect after being crosslinked, the oxygen release rate is matched with the requirement of damaged tissues, and a good microenvironment is provided for skin repair, so that the normal proceeding of the repair process is promoted, and the repair efficiency is improved. And the material is a very potential ideal material for tissue repair.

Description

Technical Field

[0001] The invention relates to the technical field of tissue engineering materials, and in particular to a dynamic oxygen-releasing microsphere capable of eliminating active oxygen, and a preparation method and application thereof. Background Art

[0002] As the largest organ in the human body and the outermost barrier of human tissue, the skin is the easiest and most frequently damaged. Common skin injuries include ulcers, scratches from foreign objects, burns, etc. Skin self-repair is one of the most complex repair processes in human tissue repair. Damaged skin can be in direct contact with the outside world, which increases the risk of wound infection and the difficulty of skin repair. Skin injuries are divided into three levels: mild, moderate, and severe. Mild skin damage can be healed through tissue self-repair, while severe skin damage, such as burns, often requires medical intervention. Current research on repairing severe skin damage focuses on the improvement of skin replacement materials, stem cell and stem cell-derived product treatments, etc. However, no matter which treatment method is used, its effectiveness will be greatly reduced in the hypoxic environment of the injured tissue.

[0003] When the skin is injured, the vascular tissue at the wound is destroyed, and oxygen cannot reach the wound through the blood vessels, resulting in a local hypoxic environment in the tissue. Short-term hypoxia can promote cell and tissue functions, such as angiogenesis, cell proliferation, migration, differentiation and tissue regeneration. However, long-term hypoxia in severely injured wounds can impair the healing activity at the cell and tissue levels, leading to slow wound healing, tissue necrosis and inflammatory response. In addition, the oxygen status of the damaged tissue changes dynamically. In the early stage of injury, the oxygen content of the tissue is low; as the wound heals, the repaired blood vessels restore part of the oxygen supply, and the oxygen content of the tissue increases. Correspondingly, the oxygen demand of the damaged tissue decreases from high to low. In addition, oxygen is consumed by inflammatory cells (mainly neutrophils), and reactive oxygen species (ROS) are generated during the respiratory burst. Excessive oxygen supply can also generate reactive oxygen species, thereby aggravating wound non-healing. Therefore, it is necessary to prepare an oxygen release system with dynamic oxygen supply and reactive oxygen scavenging function, which has good development prospects. Studies have found that after skin injury, the pH of the damaged area changes. Under normal circumstances, the skin is weakly acidic, with a pH between 4.5 and 6.5. Depending on the degree of damage, the pH increases and eventually becomes alkaline. As tissue repair proceeds, the pH gradually falls back to normal. Therefore, overall, the pH of the skin during the repair process is from high to low, which forms a certain corresponding relationship with the changing law of oxygen demand during the repair process. At present, oxygen-releasing materials have also become a hot topic of research in the field of tissue repair. For example, the invention patent CN115490927A provides an injectable oxygen-releasing hydrogel, which has a double cross-linked network, fast and controllable molding speed, good shape adaptability, self-healing, adhesion and antibacterial properties. However, the oxygen release law of the hydrogel is consistent with the degradation rate of the hydrogel, which does not fully meet the need for oxygen in the wound healing process. The invention patent CN111116942A provides a self-cross-linked oxygen-releasing hydrogel using a hydrogen peroxide source as an oxygen release agent, wherein the hydrogel releases oxygen by catalyzing the hydrogen peroxide source, and the controllable release of oxygen is achieved by changing the ratio of the hydrogel raw materials. However, the oxygen-releasing agent hydrogen peroxide used therein is cytotoxic, and if the catalysis is incomplete, it will cause cell necrosis. Invention patent CN108310470B discloses a dynamic oxygen-releasing microsphere and its preparation method and use. The dynamic oxygen-releasing microsphere is a microsphere with gelatin as the matrix, and contains inorganic peroxide particles inside the microsphere. However, since gelatin is very easily soluble in water, simply wrapping calcium peroxide with gelatin will quickly decompose within one or two days and thus cannot achieve slow oxygen release. In summary, the oxygen release amount of the above-mentioned oxygen-releasing materials does not have the characteristics that match the needs of damaged tissues, and does not take into account the damage to cells caused by reactive oxygen species generated by excessive oxygen release.

[0004] Therefore, how to prepare an oxygen supply product whose oxygen release rate matches the needs of damaged tissue and has good biocompatibility has become a technical problem that needs to be solved urgently. Summary of the invention

[0005] In view of the above-mentioned deficiencies in the prior art, the technical problem to be solved by the present invention is: how to provide a dynamic oxygen-releasing microsphere that can eliminate active oxygen and a preparation method and application thereof, so as to solve the problems that the oxygen release rate of the existing oxygen-releasing materials does not match the needs of damaged tissues, and excessive oxygen supply easily generates active oxygen and is toxic to cells.

[0006] In order to solve the above technical problems, the present invention adopts the following technical solutions: a dynamic oxygen-releasing microsphere that can eliminate active oxygen, the microsphere includes an outer shell, an inner shell and a core from the outside to the inside, the core is calcium peroxide, the inner shell is gelatin, the outer side of the inner shell is cross-linked with an outer shell, the outer shell is formed by cross-linking carboxymethyl chitosan and sodium alginate, and nano-cerium oxide is evenly dispersed in the outer shell; the microsphere dynamically changes the oxygen release rate by responding to changes in environmental pH. In this way, gelatin is a product obtained by hydrolysis of natural collagen, and is a very important natural biopolymer material with good biocompatibility and tissue compatibility, good biodegradability, and the degradation product is non-toxic. Carboxymethyl chitosan is a natural polysaccharide that contains both cations (-NH 3+ ) group, and contains an anion (-COO - ) group, is an amphoteric polyelectrolyte with special pH sensitivity. The gelatin in the inner shell contains a large number of amine groups, which will undergo ion exchange with the carboxyl groups in the sodium alginate in the outer shell, thereby forming physical crosslinks; at the same time, both contain a large number of hydroxyl groups, which can interact with amine groups or hydroxyl groups in other molecules through hydrogen bonds. These two effects can further strengthen the crosslinking, making the microsphere shell structure more stable and tighter, degrading more slowly, and better meeting the oxygen release requirements for a longer time. The nano-cerium oxide particles evenly mixed in the outer shell can achieve active oxygen scavenging.

[0007] Another object of the present invention is to provide a method for preparing the above-mentioned dynamic oxygen-releasing microspheres capable of eliminating active oxygen, comprising the following steps:

[0008] 1) adding calcium peroxide to a gelatin solution to obtain a gelatin / calcium peroxide suspension, and then preheating the gelatin / calcium peroxide suspension;

[0009] 2) adding the preheated gelatin / calcium peroxide suspension dropwise into the oil phase, maintaining a temperature of 45-60° C. and a stirring speed of 600-1000 rpm, until emulsification forms a stable W / O type emulsion, rapidly cooling in an ice bath, adding a crosslinking agent to solidify the crosslinking when the temperature drops to 4-6° C., washing it after sufficient reaction, and lyophilizing it to obtain gelatin / calcium peroxide microspheres;

[0010] 3) mixing carboxymethyl chitosan and sodium alginate and dissolving them in water to prepare a shell solution, and then adding nano-cerium oxide and the gelatin / calcium peroxide microspheres obtained in step 2) and mixing them evenly to obtain a mixed solution;

[0011] 4) dropping the mixed solution obtained in step 3) into the oil phase, maintaining a temperature of 45-60° C. and a stirring speed of 600-1000 rpm until the solution is completely emulsified, then adding a crosslinking agent, continuing stirring, and after sufficient reaction, washing and drying, the dynamic oxygen-releasing microspheres capable of eliminating active oxygen are obtained.

[0012] Preferably, the concentration of the gelatin is 0.2-0.8 g / ml; the mass ratio of the calcium peroxide to the gelatin is 1:2-8.

[0013] Preferably, the oil phase is prepared by adding liquid paraffin into Span 80 and stirring the mixture at 50-60°C.

[0014] Preferably, the mass ratio of the liquid paraffin to Span80 is 100:1-5; the stirring speed is 600-1000 rpm, and the time is 20-40 min.

[0015] Preferably, the mass ratio of the carboxymethyl chitosan to sodium alginate is 1:1 to 1:3; the mass volume ratio of the nano-cerium oxide to the shell solution is 5 to 20 μg: 1 mL. In this way, by changing the mass ratio of carboxymethyl chitosan to sodium alginate, the oxygen release rate of the microspheres at different pH values ​​is adjusted to achieve the controlled release of oxygen. Through comparative experiments, the oxygen release rate in this ratio range is most consistent with the law of natural healing and hypoxia of the skin. The oxygen release rate of the microspheres is positively correlated with the pH value, that is, the larger the pH value, the greater the oxygen release rate.

[0016] Preferably, the cross-linking agent in step 2) is glutaraldehyde or genistein; and the cross-linking agent in step 4) is a 10% calcium chloride solution or a mixture of a calcium chloride solution and glutaraldehyde.

[0017] Another object of the present invention is to provide the use of the dynamic oxygen-releasing microspheres or the dynamic oxygen-releasing microspheres prepared by the above method in skin repair engineering.

[0018] Another object of the present invention is to provide a skin repair material, the active ingredients of which include the above-mentioned dynamic oxygen-releasing microspheres or the dynamic oxygen-releasing microspheres prepared by the above-mentioned method.

[0019] Compared with the prior art, the present invention has the following beneficial effects:

[0020] 1. The dynamic oxygen-releasing microspheres SA / CMCS_@CaO2 provided by the present invention that can eliminate active oxygen are based on calcium peroxide as the microsphere core, and a two-layer shell structure is wrapped outside the core, and nano-cerium oxide particles are evenly distributed in the outer shell to achieve active oxygen removal. On the one hand, the gelatin in the inner shell of the structure contains a large number of amine groups, which will undergo ion exchange with the carboxyl groups in the sodium alginate of the outer shell, thereby forming physical crosslinks; at the same time, both contain a large number of hydroxyl groups, which can interact with amine groups or hydroxyl groups in other molecules through hydrogen bonds. These two effects can further strengthen the crosslinking, making the microsphere structure more stable, tighter, and degrading more slowly, which can better meet the oxygen release for a longer time. On the other hand, the structure isolates calcium peroxide from cells to the greatest extent, preventing calcium peroxide and conversion products such as hydrogen peroxide from contacting cells to produce toxicity, and at the same time, it can also isolate calcium peroxide from sodium alginate, preventing sodium alginate and calcium ions from crosslinking and directly gelling, which is not conducive to the formation of microspheres. The microsphere components of the present invention are more uniform, the structure is stable, it is not easy to deform, the particle size is uniform, and the oxygen release rate matches the needs of damaged tissues. In addition, the structural similarity between sodium alginate and the extracellular matrix provides a 3D environment similar to ECM. Therefore, the dynamic oxygen-releasing microspheres of the present invention provide a good microenvironment for cells, which is conducive to cell proliferation and differentiation, and can further promote skin repair.

[0021] 2. The dynamic oxygen-releasing microspheres prepared by the present invention can adjust the oxygen release rate of the microspheres at different pH values ​​by changing the mass ratio of carboxymethyl chitosan and sodium alginate, thereby achieving a controlled release of oxygen. Through experiments, the ratio that best conforms to the law of hypoxia in the natural healing process can be obtained. The oxygen release rate of the oxygen-releasing microspheres prepared by the present invention is positively correlated with the pH value. The oxygen release rate can be dynamically changed according to the law of hypoxia in the repair process, and oxygen can be supplied in appropriate amounts at each stage of the repair. At the same time, the possible active oxygen can be removed to prevent oxidative stress. By regulating the ambient oxygen, a good microenvironment is provided for skin repair, thereby promoting the normal progress of the repair process. The present invention can be used for the repair of injured tissues such as skin in a hypoxic environment, expanding the scope of application, and is a very potential tissue repair material. In addition, the present invention has no complex process and operation, does not require expensive instruments, has simple and easy-to-obtain raw materials, has a low preparation cost, is convenient for mass preparation, and has a very broad application prospect in the field of tissue repair engineering.

[0022] 3. The oxygen-releasing microspheres prepared by the present invention all use natural biomaterials as raw materials, and not only have good biocompatibility, degradability and other properties, but also the outer shell of the microspheres uses carboxymethyl chitosan / sodium alginate, which has a certain hemostatic effect after cross-linking, and can also promote skin repair. The oxygen release rate matches the needs of damaged tissues, providing a new option for tissue repair. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1This is the macroscopic observation of the dynamic oxygen-releasing microspheres prepared by the present invention.

[0024] Figure 2 The microscopic morphology of the dynamic oxygen-releasing microspheres prepared in the present invention.

[0025] Figure 3 This is a statistical diagram of the particle size of the dynamic oxygen-releasing microspheres prepared in the present invention.

[0026] Figure 4 This is the infrared spectrum of the dynamic oxygen-releasing microspheres prepared by the present invention.

[0027] Figure 5 The swelling rates of the dynamic oxygen-releasing microspheres prepared in Examples 1 to 3 of the present invention under different pH conditions.

[0028] Figure 6 The water absorption rate of the dynamic oxygen-releasing microspheres prepared in Examples 1 to 3 of the present invention under different pH conditions.

[0029] Figure 7 The degradation rate of the dynamic oxygen-releasing microspheres prepared in Example 2 of the present invention under different pH conditions.

[0030] Figure 8 The oxygen release rate of the dynamic oxygen-releasing microspheres prepared in Example 2 of the present invention under different pH conditions. DETAILED DESCRIPTION

[0031] The present invention is further described in detail below with reference to the examples. The reagents and experimental methods used in the examples are commercially available and conventionally operated unless otherwise specified.

[0032] 1. A method for preparing dynamic oxygen-releasing microspheres capable of eliminating active oxygen

[0033] Embodiment 1: This embodiment is prepared by the following steps:

[0034] 1) Weigh 1-4 g of calcium peroxide and add 10 ml of a gelatin solution with a concentration of 0.2%-0.8% g / ml, stir and mix to obtain a gelatin / calcium peroxide suspension, and then preheat the gelatin / calcium peroxide suspension to 50° C.; measure 50 ml of liquid paraffin, add it to 1 mL of Span80, and then stir at 50° C. and 1000 rpm for 20 min to obtain an oil phase.

[0035] 2) The preheated gelatin / calcium peroxide suspension was added dropwise to the oil phase, stirred at 50°C and 1000 rpm for 30 min until a stable W / O emulsion was formed, and then quickly cooled in an ice bath. When the temperature dropped to 5°C, 4 ml of glutaraldehyde was added for cross-linking. After sufficient reaction for 2 h, it was washed and freeze-dried to obtain gelatin / calcium peroxide microspheres.

[0036] 3) Carboxymethyl chitosan and sodium alginate are mixed in a mass ratio of 1:3 and dissolved in water to prepare a carboxymethyl chitosan / sodium alginate shell solution with a concentration of 0.5% to 1%, and then 10 μg / ml nano cerium oxide and gelatin / calcium peroxide microspheres obtained in step 2) are added according to a mass volume ratio of nano cerium oxide to shell solution of 15 μg:1 mL, and the mixture is evenly mixed to obtain a mixed solution.

[0037] 4) The mixed solution obtained in step 3) is dropped into the oil phase (50 ml of liquid paraffin plus 1 mL of Span80), stirred at 50° C. and 1000 rpm for 2 to 3 h until the solution is completely emulsified, and then 5 ml of 10% calcium chloride is added, and stirring is continued for 0.5 to 1 h. After washing and drying, the dynamic oxygen-releasing microspheres SA / CMCS_@CaO2 that can eliminate active oxygen are obtained.

[0038] Embodiment 2 This embodiment is prepared by the following steps:

[0039] 1) Weigh 1-4 g of calcium peroxide and add 10 ml of a gelatin solution with a concentration of 0.2%-0.8% g / ml, stir and mix to obtain a gelatin / calcium peroxide suspension, and then preheat the gelatin / calcium peroxide suspension to 50° C.; measure 50 ml of liquid paraffin, add it to 1 mL of Span80, and then stir at 50° C. and 1000 rpm for 20 min to obtain an oil phase.

[0040] 2) The preheated gelatin / calcium peroxide suspension was added dropwise to the oil phase, stirred at 50°C and 1000 rpm for 30 min until a stable W / O emulsion was formed, and then quickly cooled in an ice bath. When the temperature dropped to 5°C, 4 ml of glutaraldehyde was added for cross-linking. After sufficient reaction for 2 h, it was washed and freeze-dried to obtain gelatin / calcium peroxide microspheres.

[0041] 3) Carboxymethyl chitosan and sodium alginate are mixed in a mass ratio of 1:1 and dissolved in water to prepare a carboxymethyl chitosan / sodium alginate shell solution with a concentration of 0.5% to 1%, and then 10 μg / ml nano cerium oxide and gelatin / calcium peroxide microspheres obtained in step 2) are added according to a mass volume ratio of nano cerium oxide to shell solution of 15 μg:1 mL, and the mixture is evenly mixed to obtain a mixed solution.

[0042] 4) dropping the mixed solution obtained in step 3) into the oil phase (50 ml of liquid paraffin plus 1 ml of Span80), stirring at 50° C. and 1000 rpm for 2 to 3 hours until the solution is completely emulsified, then adding 5 ml of 10% calcium chloride, stirring for 0.5 to 1 hour, washing and drying, and obtaining the dynamic oxygen-releasing microspheres capable of eliminating active oxygen.

[0043] SA / CMCS_@CaO2.

[0044] Embodiment 3 This embodiment is prepared by the following steps:

[0045] 1) Weigh 1-4 g of calcium peroxide and add 10 ml of a gelatin solution with a concentration of 0.2%-0.8% g / ml, stir and mix to obtain a gelatin / calcium peroxide suspension, and then preheat the gelatin / calcium peroxide suspension to 50° C.; measure 50 ml of liquid paraffin, add it to 1 mL of Span80, and then stir at 50° C. and 1000 rpm for 20 min to obtain an oil phase.

[0046] 2) The preheated gelatin / calcium peroxide suspension was added dropwise to the oil phase, stirred at 50°C and 1000 rpm for 30 min until a stable W / O emulsion was formed, and then quickly cooled in an ice bath. When the temperature dropped to 5°C, 4 ml of glutaraldehyde was added for cross-linking. After sufficient reaction for 2 h, it was washed and freeze-dried to obtain gelatin / calcium peroxide microspheres.

[0047] 3) Carboxymethyl chitosan and sodium alginate are mixed in a mass ratio of 3:1 and dissolved in water to prepare a carboxymethyl chitosan / sodium alginate shell solution with a concentration of 0.5% to 1%, and then 10 μg / ml nano cerium oxide and gelatin / calcium peroxide microspheres obtained in step 2) are added according to a mass volume ratio of nano cerium oxide to shell solution of 15 μg:1 mL, and the mixture is evenly mixed to obtain a mixed solution.

[0048] 4) dropping the mixed solution obtained in step 3) into the oil phase (50 ml of liquid paraffin plus 1 ml of Span80), stirring at 50° C. and 1000 rpm for 2 to 3 hours until the solution is completely emulsified, then adding 5 ml of 10% calcium chloride, stirring for 0.5 to 1 hour, washing and drying, and obtaining the dynamic oxygen-releasing microspheres capable of eliminating active oxygen.

[0049] SA / CMCS_@CaO2.

[0050] 2. Performance Verification

[0051] 1. Observe the macroscopic and microscopic morphologies of the gelatin / calcium peroxide microspheres, SA / CMCS_@CaO2 dynamic oxygen-releasing microspheres and gelatin prepared in Example 2 and count the particle sizes. The results are as follows: Figures 1 to 3 shown.

[0052] from Figure 1 It can be seen that the gelatin / calcium peroxide microspheres and SA / CMCS_@CaO2 dynamic oxygen-releasing microspheres prepared by the present invention are in the form of light yellow powder and white powder, respectively. Figure 2Microscopic observations show that the microscopic morphology of gelatin / calcium peroxide and SA / CMCS_@CaO2 are both spherical, indicating that both can successfully crosslink microspheres. Further, statistics show that the particle size of gelatin / calcium peroxide microspheres is between 20 and 100 μm; the particle size of SA / CMCS_@CaO2 dynamic oxygen-releasing microspheres is between 20 and 180 μm, with no irregular shapes and high particle size uniformity ( Figure 3 ).

[0053] 2. The gelatin / calcium peroxide microspheres and SA / CMCS_@CaO2 dynamic oxygen-releasing microspheres prepared in Example 2 were analyzed by infrared spectroscopy using a Fourier transform infrared spectrometer from Perkin Elmer, USA. Blank gelatin and calcium peroxide were used as control groups. The results are shown in Figure 2. Figure 4 shown.

[0054] from Figure 4 It can be seen that calcium peroxide has a wavelength of 1500-1000cm -1 There is an obvious absorption peak at the place where the peroxide bond is stretched and vibrated, and the characteristic absorption peak of the corresponding wavelength can also be seen in gelatin / calcium peroxide microspheres and SA / CMCS_@CaO2 oxygen-releasing microspheres, but the intensity and area of ​​the peaks are different between the groups, and there is a certain shift, which shows that calcium peroxide is indeed encapsulated in SA / CMCS_@CaO2 microspheres.

[0055] 3. The SA / CMCS_@CaO2 microspheres prepared in Examples 1 to 3 were placed under different pH conditions to test their swelling and water absorption properties. The results are as follows: Figures 5-6 shown.

[0056] from Figure 5 / 6, it can be seen that the swelling rate and water absorption rate of the SA / CMCS_@CaO2 dynamic oxygen-releasing microspheres of Examples 1 and 2 are positively correlated with the pH value, that is, the swelling rate and water absorption rate of the microspheres are larger under alkaline conditions, and smaller under acidic conditions, and the changes in the swelling and water absorption performance of Example 2 under different pH conditions are more significant. However, the changes in the swelling and water absorption performance of Example 3 and the pH value do not follow a specific pattern, which may be due to the excessive content of carboxymethyl chitosan, and the carboxymethyl chitosan molecular chain is regular and the molecular chains are closely arranged, which is not conducive to the passage of water molecules, but under neutral conditions, the large amount of -COO contained in the carboxymethyl chitosan - The strong electrostatic repulsion between them increases the water absorption capacity. Therefore, only when the carboxymethyl chitosan and sodium alginate in the outer shell are in a certain ratio, the swelling rate and water absorption rate of the microspheres are positively correlated with the pH value.

[0057] 4. The SA / CMCS_@CaO2 dynamic oxygen-releasing microspheres prepared in Example 2 were placed in PBS with different pH values ​​to simulate the human body fluid environment with a phosphate buffer solution. The microspheres were taken out at different time points by an in vitro degradation experiment and their degradation performance was tested. The results are as follows: Figure 7 shown.

[0058] from Figure 7 It can be seen that with the passage of time, SA / CMCS_@CaO2 microspheres under different pH conditions have a certain degree of degradation, but the degradation rate of the microspheres is positively correlated with the pH value, that is, the higher the pH value, the greater the degradation rate, which is consistent with the results of swelling rate and water absorption rate.

[0059] 5. The SA / CMCS_@CaO2 dynamic oxygen-releasing microspheres prepared in Example 2 were placed in PBS with different pH values, and the dissolved oxygen in PBS was measured using a JPB-607A portable dissolved oxygen meter produced by Shanghai Yidian Scientific Instrument Co., Ltd. The results are shown in the figure. Figure 8 shown.

[0060] from Figure 8 It can be seen that under different pH conditions, the dissolved oxygen content in PBS of SA / CMCS_@CaO2 dynamic oxygen-releasing microspheres decreased rapidly in the first two days, and the downward trend was relatively gentle from the 2nd to the 8th day, and the dissolved oxygen content of SA / CMCS_@CaO2 dynamic oxygen-releasing microspheres was positively correlated with the pH value, which shows that SA / CMCS_@CaO2 dynamic oxygen-releasing microspheres can release more oxygen in the alkaline environment of damaged skin. Therefore, the oxygen release rate of the oxygen-releasing microspheres prepared by the present invention matches the needs of damaged tissues, and can dynamically change the oxygen release rate according to the hypoxia law of the skin repair process, and supply oxygen in appropriate amounts at each stage of repair. At the same time, the nano-cerium oxide particles evenly mixed in the outer shell can remove possible active oxygen and prevent oxidative stress. By regulating the environmental oxygen, a good microenvironment is provided for skin repair, thereby promoting the normal progress of the repair process.

[0061] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. A dynamic oxygen-releasing microsphere capable of eliminating active oxygen, characterized in that: The microspheres include an outer shell, an inner shell and a core from the outside to the inside, the core is calcium peroxide, the inner shell is gelatin, the outer shell is cross-linked with an outer shell on the outside, the outer shell is formed by cross-linking carboxymethyl chitosan and sodium alginate, and nano cerium oxide is evenly dispersed in the outer shell; the microspheres dynamically change the oxygen release rate by responding to changes in environmental pH.

2. A method for preparing dynamic oxygen-releasing microspheres capable of eliminating active oxygen according to claim 1, characterized in that: The following steps are involved: 1) adding calcium peroxide to a gelatin solution to obtain a gelatin / calcium peroxide suspension, and then preheating the gelatin / calcium peroxide suspension; 2) adding the preheated gelatin / calcium peroxide suspension dropwise into the oil phase, maintaining a temperature of 45-60° C. and a stirring speed of 600-1000 rpm, until emulsification forms a stable W / O type emulsion, rapidly cooling in an ice bath, adding a crosslinking agent to solidify the crosslinking when the temperature drops to 4-6° C., washing it after sufficient reaction, and lyophilizing it to obtain gelatin / calcium peroxide microspheres; 3) mixing carboxymethyl chitosan and sodium alginate and dissolving them in water to prepare a shell solution, and then adding nano-cerium oxide and the gelatin / calcium peroxide microspheres obtained in step 2) and mixing them evenly to obtain a mixed solution; 4) dropping the mixed solution obtained in step 3) into the oil phase, maintaining a temperature of 45-60° C. and a stirring speed of 600-1000 rpm until the solution is completely emulsified, then adding a crosslinking agent, continuing stirring, and after sufficient reaction, washing and drying, the dynamic oxygen-releasing microspheres capable of eliminating active oxygen are obtained.

3. The method for preparing the dynamic oxygen-releasing microspheres capable of eliminating active oxygen according to claim 2, characterized in that: The concentration of the gelatin solution is 0.2-0.8 g / ml.

4. The method for preparing the dynamic oxygen-releasing microspheres capable of eliminating active oxygen according to claim 2, characterized in that: The mass ratio of calcium peroxide to gelatin is 1:2-8.

5. The method for preparing the dynamic oxygen-releasing microspheres capable of eliminating active oxygen according to claim 2, characterized in that: The oil phase is prepared by adding liquid paraffin into Span80 and stirring evenly at 50-60°C.

6. The method for preparing the dynamic oxygen-releasing microspheres capable of eliminating active oxygen according to claim 5, characterized in that: The mass ratio of the liquid paraffin to Span80 is 100:1-5; the stirring speed is 600-1000 rpm, and the time is 20-40 min.

7. The method for preparing the dynamic oxygen-releasing microspheres capable of eliminating active oxygen according to claim 2, characterized in that: The mass ratio of the carboxymethyl chitosan to the sodium alginate is 1:1 to 1:3; the mass volume ratio of the nano cerium oxide to the shell solution is 5 to 20 μg: 1 mL.

8. The method for preparing the dynamic oxygen-releasing microspheres capable of eliminating active oxygen according to claim 2, characterized in that: The cross-linking agent in step 2) is glutaraldehyde or genistein; the cross-linking agent in step 4) is a 10% calcium chloride solution or a mixture of a calcium chloride solution and glutaraldehyde.

9. Use of the dynamic oxygen-releasing microspheres according to claim 1 or the dynamic oxygen-releasing microspheres prepared by the method according to any one of claims 2 to 8 in skin repair engineering.

10. A skin repair material, characterized in that: The active ingredients include the dynamic oxygen-releasing microspheres described in claim 1 or the dynamic oxygen-releasing microspheres prepared by the method described in any one of claims 2 to 8.

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