Citric acid-based biological material as well as preparation method and application thereof

Abstract

The invention relates to a citric acid-based particle injection material as well as a preparation method and application thereof. The preparation method comprises the following steps: melting and heating citric acid and 1, 8-octylene glycol to form a POC prepolymer, dissolving the POC prepolymer with an organic solvent to obtain a dissolved solution, and slowly pouring the dissolved solution into ultrapure water for multiple times for purification to obtain a purified POC degradable prepolymer; and dissolving the POC degradable prepolymer with a solvent, adding inorganic salt particles, and carrying out a cross-linking reaction to obtain the POC injectable particle material. Medicines, active molecules, various metal ions and the like are loaded on POC in a physical compounding or chemical crosslinking mode, and the citric acid-based particle injection material is obtained. The citric acid-based biological material provided by the invention, as a novel medical beauty raw material and a related particle injection filler thereof, can effectively promote regeneration, proliferation, migration and chemotaxis of skin cells, promote the production of collagen and achieve a medical beauty effect. Meanwhile, the immune response of the organism can be activated, inflammation is reduced, the product after degradation is citric acid which can be metabolized randomly, and obvious immunogenicity and toxic and side effects are avoided.

Description

Technical Field

[0001] The present invention relates to the technical field of biomaterials, and in particular to a citric acid-based biomaterial and a preparation method and application thereof. Background Art

[0002] Skin aging is a health problem that everyone has to face. With the increase of age and the accumulation of environmental factors, the phenomenon of dryness, wrinkles, loss of elasticity and other phenomena on the skin surface gradually intensifies, and may even lead to skin dysfunction and increase the risk of skin diseases and skin malignancies. In the pursuit of youth, beauty and health, skin aging brings psychological and physiological burdens to people. At present, anti-skin aging strategies such as retinoic acid, radio frequency technology and dermal fillers are aimed at stimulating the production of collagen in the dermis to restore the function of aging skin. Skin aging is a health problem that everyone needs to face, and with the increase of age, skin aging will become more serious, and the apparent symptoms will become more obvious, which greatly causes people to worry about changes in appearance.

[0003] Skin aging is closely related to changes in collagen. The amino acid with the highest content in collagen is hydroxyproline. Compared with other tissues, hydroxyproline is mainly present in collagen in the skin. Therefore, it can be used as an indicator to evaluate skin aging. Hyaluronic acid is an acidic mucopolysaccharide with water-retaining effect, which is secreted by fibroblasts in the skin. Hyaluronic acid can improve nutrient metabolism, increase elasticity, and prevent skin aging. Its content gradually decreases with age. Halogenated tyrosine increases with age in both light-exposed and light-protected skin, indicating that it may be a useful biomarker for skin aging.

[0004] Dermal filling technology has always been an important means of treating skin aging. At present, dermal filling materials mainly use polymer materials, such as hyaluronic acid, polylactic acid, etc. These materials have good biocompatibility and degradability, and can be integrated with human tissue to achieve the purpose of repairing and filling the skin. At the same time, these materials also have good moisturizing and skin rejuvenation effects, which can improve the texture and appearance of the skin.

[0005] However, existing injectable medical beauty needles have toxic side effects and are prone to a series of problems such as wound infection. Once they accidentally enter the blood, they can easily cause blood clots. Summary of the invention

[0006] In view of the fact that existing injectable medical cosmetic materials have toxic side effects and are prone to wound infection and other problems, the present invention provides a citric acid-based biomaterial and a preparation method and application thereof.

[0007] The purpose of the present invention can be achieved by the following technical solutions:

[0008] In a first aspect, the present invention provides a method for preparing a POC injectable particulate material, comprising the following steps:

[0009] The POC degradable prepolymer is freeze-dried for later use;

[0010] The freeze-dried POC degradable prepolymer is dissolved in a solvent, inorganic salt particles are added, cross-linked, and then ground, the inorganic salt is removed, and dried to obtain the POC injectable particulate material.

[0011] In one embodiment of the present invention, the solvent is selected from one or a combination of dioxane, anhydrous ethanol, acetone or dichloromethane.

[0012] In one embodiment of the present invention, the amount of the freeze-dried POC degradable prepolymer and the solvent is in the ratio of 10 g: 10-200 mL.

[0013] In one embodiment of the present invention, the inorganic salt particles are selected from one or a combination of NaCl, KCl, MgCl2, etc. The inorganic salt particles serve as pore-forming agents and hardening materials to facilitate subsequent grinding treatment.

[0014] In one embodiment of the present invention, the amount of the inorganic salt particles added is 1-20 times the amount of the freeze-dried POC degradable prepolymer.

[0015] In one embodiment of the present invention, the cross-linking reaction conditions are: >50°C, cross-linking for 12-72 hours under vacuum or without vacuum conditions. Preferably, cross-linking for 12-72 hours at 60-80°C. More preferably, cross-linking for 72 hours at 80°C.

[0016] In one embodiment of the present invention, the inorganic salt particle size before grinding is 50-150 microns. The POC degradable prepolymer and the inorganic salt are cross-linked and then ground, and then sieved to obtain a POC injectable particulate material, and the particle size of the POC injectable particulate material is preferably 30-100 microns or <30 microns. The method for removing inorganic salts from the sieved POC injectable particulate material is: adding pure water to the POC injectable particulate material to dissolve the inorganic salts, and then freeze-drying the POC injectable particulate material.

[0017] In one embodiment of the present invention, the method for preparing the POC degradable prepolymer comprises the following steps:

[0018] The citric acid and 1,8-octanediol are mixed and melted, and the hydroxyl groups of the 1,8-octanediol can be covalently cross-linked with the carboxyl groups of the citric acid, and then heated and stirred under vacuum conditions to react to form a POC prepolymer;

[0019] The POC prepolymer is dissolved by an organic solvent to obtain a solution, and the solution is slowly poured into ultrapure water for purification multiple times by utilizing the property that the prepolymer is immiscible in water, so as to obtain a purified POC degradable prepolymer.

[0020] In one embodiment of the present invention, the amount of citric acid and 1,8-octanediol used is in a weight ratio of 1:(1-10) to (1-10):1.

[0021] In one embodiment of the present invention, the mixing and melting of citric acid and 1,8-octanediol is carried out under the protection of an inert gas, and the temperature of the mixed dissolution is 120-160° C., preferably 160° C. The inert gas includes nitrogen and the like.

[0022] In one embodiment of the present invention, the stirring reaction under vacuum conditions is: 120-140°C, 300-1000 rpm stirring speed, react for 1-48 hours, then reduce the speed until stirring cannot be continued, and the reaction stops. The stirring reaction under vacuum conditions is preferably 140°C, 300 rpm stirring speed, and react for 12 hours. The operation of reducing the speed can be first to 150 rpm stirring speed, and then to 60 rpm stirring speed, until stirring cannot be continued.

[0023] In one embodiment of the present invention, the volume mass ratio of the organic solvent to the POC prepolymer is 1-100:1.

[0024] In one embodiment of the present invention, the organic solvent is selected from one or a combination of dioxane, anhydrous ethanol, acetone or dichloromethane.

[0025] In one embodiment of the present invention, the dissolution of the POC prepolymer by the organic solvent is carried out under stirring conditions, and the stirring condition is 50-1000 rpm, preferably 300 rpm.

[0026] In one embodiment of the present invention, when the dissolved solution is slowly poured into ultrapure water for purification multiple times, the volume ratio of the dissolved solution to the ultrapure water is 1:10-1000.

[0027] The POC degradable prepolymer can be stored in a refrigerated environment.

[0028] The second aspect of the present invention provides a POC injectable particulate material prepared by the method according to the first aspect of the present invention.

[0029] In a third aspect of the present invention, a citric acid-based microparticle injection material is provided, comprising the POC injectable microparticle material, and a drug, an active molecule, a bioglass or a metal ion loaded on the POC injectable microparticle material. The metal ion includes but is not limited to Mg 2+ ,Ca 2+ ,Zn 2+ ,Cu 2+ wait.

[0030] In one embodiment of the present invention, the drug or active molecule is selected from adenosine triphosphate and glutamine.

[0031] In one embodiment of the present invention, the loading amount of the adenosine triphosphate relative to 1 gram of the POC injectable microparticle material is 5-50 mg, and the loading amount of the glutamine relative to 1 gram of the POC injectable microparticle material is 10-100 mg.

[0032] When the citric acid-based microparticle injection material includes a POC injectable microparticle material, and adenosine triphosphate and glutamine loaded on the POC injectable microparticle material, the citric acid-based microparticle injection material is recorded as ATP-Gln@ / POC.

[0033] In a fourth aspect of the present invention, a method for preparing a citric acid-based microparticle injection material is provided, wherein a drug or active molecule is combined with the POC injectable microparticle material by incubation or chemical cross-linking.

[0034] The incubation method is as follows: dissolving the drug or active molecule and the POC injectable microparticle material in physiological saline or culture medium, incubating them for 12-48 hours by physical adsorption through a micro-oscillator, and then separating, purifying, and drying them by centrifugation at 1000-2000 rpm for multiple times;

[0035] The chemical cross-linking method is that the drug or active molecule is connected to the POC injectable microparticle material through a chemical reaction.

[0036] In another embodiment of the present invention, the chemical cross-linking method is: directly heating adenosine triphosphate with glutamine and POC injectable microparticle material and polyol to cross-link; or directly heating ATP, glutamine, citric acid, and 1,8-pentanediol during the reaction to form cross-links; or the above-mentioned POC degradable citric acid-based material, in the case of DCC / NHS as a catalyst, is coupled with adenosine triphosphate and glutamine to form bionic microparticles or scaffolds.

[0037] The fifth aspect of the present invention provides a citric acid-based biomimetic material, comprising the POC injectable particulate material provided in the second aspect of the present invention, and one or a combination of collagen, chitosan, hyaluronic acid or hydroxyapatite loaded on the POC injectable particulate material. The loading amount of collagen, chitosan and / or hyaluronic acid and / or hydroxyapatite is 0%-50wt%, and is not 0.

[0038] The preparation method of the citric acid-based biomimetic material can be: a combination of POC injectable microparticle material and one or more of collagen, chitosan, hyaluronic acid or hydroxyapatite is used to form biomimetic microparticles or scaffolds by fixing and cross-linking using DCC / DMAP as a catalyst.

[0039] In one embodiment of the present invention, the citric acid-based biomimetic material comprises the POC injectable particulate material provided in the second aspect of the present invention, and one or a combination of collagen, chitosan, hyaluronic acid or hydroxyapatite loaded on the POC injectable particulate material, and a drug or active molecule loaded on the POC injectable particulate material.

[0040] In one embodiment of the present invention, the citric acid-based biomimetic material specifically includes the POC injectable particulate material provided in the second aspect of the present invention, and one or a combination of collagen, chitosan, hyaluronic acid or hydroxyapatite loaded on the POC injectable particulate material, and adenosine triphosphate and glutamine loaded on the POC injectable particulate material.

[0041] The present invention further provides an injection material, which is composed of the following formula components in weight percentage: 1%-50% of the POC injectable microparticle material, 0.5-10% of sodium carboxymethyl cellulose CMC, and 40-95% of sterile physiological saline;

[0042] Alternatively, the injectable material is obtained by dispersing the POC injectable microparticle material with inorganic microparticles such as collagen, hyaluronic acid, bioglass or hydroxyapatite in sterile saline.

[0043] In a sixth aspect of the present invention, the citric acid-based microparticle injection material or the citric acid-based biomimetic material is provided for use in preparing biomedical cosmetic materials, especially as a biomedical cosmetic filling material, and more specifically, it can be used for skin tissue dermal filling and breast augmentation prosthesis.

[0044] The present application scheme uses citric acid as the main material. Citric acid is a natural organic acid with good biocompatibility. Therefore, the nanocarrier based on citric acid can reduce the immune rejection reaction to the body when used in vivo. At the same time, it has good drug release performance. Citric acid nanocarriers can be used as drug carriers to improve the efficacy of drugs and reduce side effects by controlling the release rate and release site of drugs in the body. Biomaterials based on citric acid can be obtained by simple chemical synthesis and preparation methods, and the preparation process is relatively simple and low in cost. Citric acid-based biomaterials have good stability and degradability, can stay in the body for a long time and slowly release drugs, and are gradually metabolized and excreted from the body over time.

[0045] Adenosine triphosphate (ATP) is an energy molecule in cells that can provide energy for cells and promote cell metabolism and growth. In the process of tissue regeneration, ATP can promote cell division and proliferation, accelerate tissue repair and regeneration. ATP can also promote vasodilation, increase blood flow, and improve blood supply and oxygen supply to the myocardium, thereby providing a better environment for tissue regeneration.

[0046] Glutamine is an amino acid that has multiple functions in cells. During tissue regeneration, glutamine can promote the regeneration of muscle tissue. Glutamine can act as a sensor molecule to control the regeneration program of muscle tissue. At the same time, glutamine can also promote protein synthesis and decomposition, providing the energy and substances required for muscle regeneration.

[0047] Compared with the prior art, the present invention designs a POC injectable microparticle material loaded with ATP and glutamine (Gln) to obtain a citric acid-based microparticle injection material, which can release bioactive factors (glutamine) and stimulate the production and continuous supply of endogenous ATP in body cells, thereby activating epidermal cell regeneration (such as Figure 5 ), proliferation, and significantly promoted the migration of human epidermal fibroblasts HSF (as shown in Figure 6 ) and chemotaxis (as shown Figure 7 As shown), it promotes the production of collagen and achieves medical beauty effects.

[0048] In the present application scheme, after the POC injectable microparticle material is used to load ATP and glutamine (Gln) to obtain a citric acid-based microparticle injection material, it can also be combined with collagen and / or chitosan and / or hyaluronic acid and / or hydroxyapatite to form a composite bionic microparticle or scaffold material to extend its degradation time; at the same time, by loading and slowly releasing ATP and / or glutamine, the effect of delaying skin aging and promoting the regeneration of multifunctional human tissue is further enhanced, and it can be applied to medical beauty fields such as skin tissue dermal filling and breast augmentation prosthesis.

[0049] The citric acid-based biomaterial provided by the present invention is a new type of medical and aesthetic raw material, which can promote the regeneration, proliferation, migration and chemotaxis of skin cells, promote the production of collagen, and achieve medical and aesthetic effects. At the same time, it can activate the body's immune response to reduce inflammation. The degradation product is citric acid, which can be metabolized randomly by the body without obvious immunogenicity and toxic side effects. BRIEF DESCRIPTION OF THE DRAWINGS

[0050] Figure 1 Development and preparation of ATP-Gln@ / POC injectable medical aesthetic materials combined with hydroxyapatite, chitosan and collagen injectable materials;

[0051] Figure 2 The effects of different concentrations of citric acid on HSF cell proliferation. Under the condition of 2000μM concentration, it can still significantly promote the proliferation of HSF cells.

[0052] Figure 3 The effect of different concentrations of adenosine triphosphate (ATP) on the proliferation of HSF cells. 10mM concentration can significantly promote the proliferation of HSF cells.

[0053] Figure 4 The effects of different concentrations of glutamine Gln on the proliferation of HSF cells. 60mM concentration can significantly promote the proliferation of HSF cells.

[0054] Figure 5 Detection of aging marker expression;

[0055] Figure 6 Effects of different groups on HSF cell migration;

[0056] Figure 7 Effects of different groups on HSF cell chemotaxis;

[0057] Figure 8 The production of collagen. DETAILED DESCRIPTION

[0058] The present invention is described in detail below with reference to the accompanying drawings and specific embodiments.

[0059] Example 1

[0060] Preparation and characterization of ATP / Gln@POC injectable material based on citric acid

[0061] 1. First, prepare the POC degradable prepolymer (POC) as follows:

[0062] Take 20g of citric acid and 15g of 1,8-octanediol and place them in a 50mL round-bottom flask, dissolve them at 160℃ under nitrogen protection to form a POC prepolymer for 60 minutes. Stir the reaction at 140℃ and 300 rpm for 12 hours under vacuum conditions, first reduce the stirring speed to 150 rpm, then reduce the stirring speed to 60 rpm until the stirring cannot be continued to stop the reaction. Dissolve with 150mL of dioxane reagent under stirring at 300 rpm. Slowly pour the dissolved solution into an appropriate amount of 1.5L ultrapure water again to obtain a purified POC degradable prepolymer, pre-freeze it in a -80℃ refrigerator, and freeze-dry it in vacuum for use.

[0063] 2. Preparation of POC injectable microparticle material, the method is as follows:

[0064] 10 g of purified POC degradable prepolymer was dissolved in 10 mL of dioxane, and then a sufficient amount of ground NaCl inorganic salt particles with a particle size of 50-150 microns were added, and cross-linked at 80° C. for 72 hours. After grinding, a POC injectable microparticle material was obtained.

[0065] 3. Preparation of POC-loaded ATP / glutamine (ATP-Gln@ / POC) injectable granule material, the method is as follows:

[0066] ATP, glutamine, and ATP / glutamine were prepared in 1 mol / L DMEM high-glucose medium solutions respectively, and completely dissolved, so that the concentration of the POC injectable microparticle material was 10 mM (calculated by the amount of citric acid), the ATP concentration was 10 mM, and the Gln concentration was 20 mM. The micro-oscillator was used for physical adsorption and incubated for 24 hours. Injectable ATP@POC (ATP 10 mM, POC 10 mM), Gln@POC (Gln 20 mM, POC 10 mM), and ATP-Gln@POC (ATP 10 mM, Gln 20 mM, POC 10 mM) were constructed respectively.

[0067] 4. Injectable ATP-Gln@ / POC promotes the proliferation of skin cells

[0068] Injectable ATP@POC, Gln@POC, and ATP-Gln@POC were co-incubated with skin cells, and cell proliferation was detected by CCK-8 and flow cytometry. HSF cells were stimulated with D-galactose at a concentration of 20 mg / mL for 24 hours to establish a cell senescence model.

[0069] The Cellular Senescence Detection Kit-SPiDER-βGal was used to detect the effect of ATP-Gln@ / POC in delaying HSF cell senescence. Bafilomycin A1 working solution (dilute Bafilomycin A1 DMSO stock solution 1,000 times with culture medium or HBSS), SPiDER-βGal working solution (mix SPiDERβGalDMSO stock solution and Bafilomycin A1 DMSO stock solution, and dilute the mixture 1,000 times with DMEM high glucose medium), and storage solution (dilute 1 mL of SPiDER-βGal working solution with 1 mL of culture medium or 1 mL of HBSS). After inoculating HSF cells in a 35 mm culture dish, a control group (PBS phosphate buffer treatment group, no active ingredient), a hyaluronic acid (HA, 10 mM) group, a citric acid group (10 mM), a free ATP group (10 mM), a free Gln group (20 mM), ATP@POC (ATP10 mM, POC 10 mM), Gln@POC (Gln 20 mM, POC 10 mM) and ATP-Gln@ / POC (ATP10 mM, Gln 20 mM, POC 10 mM) were set up and cultured overnight in a 37°C, 5% CO2 incubator in the culture medium; after removing the culture medium, wash once with 2 ml of culture medium or 2 mL of HBSS; after adding 1 mL of Bafilomycin A1 working solution, culture in a 37°C, 5% CO2 incubator for 1 h. After adding 1 mL of SPiDER-βGal working solution, the cells were cultured in a 37°C, 5% CO2 incubator for 30 min. The supernatant was removed, and the cells were washed twice with 2 mL of culture medium or 2 mL of HBSS, and then observed under a fluorescence microscope.

[0070] 5. ATP-Gln@ / POC injectable filling material (i.e., ATP-Gln@ / POC in the above embodiment) promotes the migration of skin cells

[0071] Cell migration can be detected by cell scratching combined with live and dead cell fluorescence staining. HSF cells were digested with trypsin, centrifuged and counted, and then 1×10 6 Cells / well were inoculated into 6-well plates and cultured overnight in an incubator. After the cells adhered to the wall, three parallel lines were drawn in the wells perpendicular to the wall with a sterilized toothpick. PBS was slowly injected along the wall of the wells, rinsed three times, and the drawn cells were removed. A control group, a citric acid group, a free ATP group, a free Gln group, ATP@POC, Gln@POC, and ATP-Gln@ / POC were set up respectively. The cells were observed and photographed under a microscope at 0h, 24h, and 48h, and the cell spacing was analyzed using Image J software.

[0072] 6. Effect of ATP-Gln@ / POC injectable filling material on cell chemotaxis

[0073] At the same time, the effect on cell chemotaxis was detected by Tanswell assay. HSF cells with a confluence of about 90% and in the logarithmic growth phase were selected and cultured overnight in serum-free DMEM medium to starve the cells. The cells were then digested with trypsin and the concentration of the cell suspension was adjusted to 1×10 5 / mL. Complete culture medium and 600μL of prepared culture medium with different substances were added to the 24-well plate, 200μL of cell suspension was added to the Tanswell chamber, and the cells were evenly distributed by shaking gently. Be careful not to generate bubbles at the bottom of the chamber. The Tanswell chambers were placed in 24-well plates and cultured in a cell culture incubator for 24 hours. The liquid in the upper and lower chambers was discarded, and the cells were washed with PBS 2-3 times. 600μL of 4% paraformaldehyde was added to the lower chamber to fix the cells for 15min (shaking table, 50rpm), the cell fixative was removed, and 600μL of crystal violet staining solution was added to each lower chamber for staining for 10min. Excess crystal violet dye was washed with PBS, and the unmigrated cells in the chamber were wiped with a cotton swab, and the chamber was inverted to air dry naturally at room temperature. Observe under a microscope and take 3 fields of view in each well. Count the cells.

[0074] 7. Effect of ATP-Gln@ / POC injectable filler material on collagen production in HSF cells

[0075] After 24 hours of treatment with the control group, citric acid group, free ATP group, free Gln group, ATP@POC, Gln@POC and ATP-Gln@ / POC, the cells were washed 3 times with PBS to remove the cell culture medium, fixed the cells with 600 μL of 4% paraformaldehyde for 15 min (shaking incubator, 50 rpm), washed 3 times with PBS to remove the cell fixative, and stained with Sirius red staining solution for 10-60 min; rinsed slightly with running water, washed 3 times with PBS to remove the surface staining solution; added Mayer's hematoxylin staining solution to stain for 8-10 min, washed 3 times with PBS to remove the surface staining solution and examined under a microscope.

[0076] Results and discussion:

[0077] Through the above method, the citric acid-based ATP / Gln@POC injectable material was successfully constructed. Figure 1 As shown in the top picture, it has excellent injectability and stability, and can be used for further research. At the same time, it can form a composite hydrogel with collagen, hydroxyapatite, chitosan, etc., to realize the development and preparation of injectable hydrogel medical beauty materials.

[0078] By testing the effects of different release and degradation products of ATP-Gln@ / POC on the proliferation of human epidermal fibroblasts HSF, the results showed that 2000μM citric acid, 10mM ATP and 60mM Gln can significantly promote the proliferation of HSF cells. Figure 2-4 The subsequent ATP-Gln@ / POC concentration was determined based on this result.

[0079] in, Figure 2 The figure shows the effect of different concentrations of citric acid on the proliferation of HSF cells. The concentration of 2000 μM can still significantly promote the proliferation of HSF cells.

[0080] Figure 3 The figure shows the effect of different concentrations of adenosine triphosphate (ATP) on the proliferation of HSF cells. Under the condition of 10mM concentration, it can significantly promote the proliferation of HSF cells.

[0081] Figure 4 The figure shows the effect of different concentrations of glutamine Gln on the proliferation of HSF cells. 60mM concentration can significantly promote the proliferation of HSF cells.

[0082] Delaying cell aging

[0083] The expression of SA-β-Gal increases with cell aging, and the progress of cell aging can be monitored by the change in the number of stained cells. The qualitative and quantitative analysis of the SPiDER-βGal kit revealed that ( Figure 5 ), the negative control group can see that the HSF cells treated with D-gal show stained senescent cells, while the positive group (HA, 10mM) effectively reduced the number of senescent cells by about 50% compared with the model group after incubation for 24 hours. Compared with the positive control group (D-galactose), the number of senescent cells in the injectable ATP-Gln@POC medical beauty material experimental group did not differ significantly from that in the positive control group (D-galactose), and was significantly better than the positive control (HA) group. The results show that the injectable ATP-Gln@POC medical beauty material can effectively delay the aging of HSF cells, and has a better anti-aging effect.

[0084] Effects on HSF cell migration and chemotaxis

[0085] The migration and chemotaxis of HSF cells are of great significance for the regeneration of aging skin. Therefore, the Transwell experiment was used to detect the effect of injectable ATP-Gln@POC medical aesthetic materials on cell migration. Live and dead cell staining was used to observe cell morphology and count the cell scratch distance. Crystal violet can bind to DNA in the cell nucleus, making the cell nucleus appear purple. After cell staining, the counting results are as follows: Figure 6 and Figure 7As shown in the figure, compared with the control group and the positive control HA group, the number of cells passing through the chamber increased significantly in the ATP-Gln@POC experimental group, and the number of migrated cells also increased, and was significantly better than the positive control (HA) group. The results show that injectable ATP-Gln@POC medical beauty materials can effectively promote the migration and chemotaxis of HSF cells.

[0086] Effects on collagen expression in HSF cells

[0087] Many factors cause the aging of skin HSF cells, which ultimately affect the expression of collagen in HSF cells. As HSF cell aging increases, the level of collagen precursors in the cells decreases, further exacerbating aging. Figure 8 As shown in the figure, after D-gal treatment, the expression of collagen precursor was significantly reduced. Sirius red staining was used to explore the changes in collagen fibers. Collagen fibers are orange-yellow in color, with higher collagen fibers in young skin and lower collagen fibers in aged skin. After incubation with ATP-Gln@POC, the production of collagen fibers in human skin fibroblasts was effectively promoted, and was significantly better than the positive control (HA) group. In summary, ATP-Gln@POC can stimulate the generation of collagen fibers in the aged dermis to reshape the extracellular matrix, thereby achieving the effect of delaying skin aging.

[0088] It can be seen that the present invention develops and designs POC injectable medical aesthetic materials, which can load biologically active substances such as ATP, glutamine, or active metal ions through mesoporous adsorption or chemical reaction, and can be combined with collagen, chitosan, hydroxyapatite, etc. for synergistic injection. The present invention can provide new materials and methods for skin tissue filling, promoting skin tissue regeneration and anti-aging, and for medical aesthetics, anti-aging or tissue regeneration.

[0089] The above description of the embodiments is to facilitate the understanding and use of the invention by those skilled in the art. It is obvious that those skilled in the art can easily make various modifications to these embodiments and apply the general principles described herein to other embodiments without creative work. Therefore, the present invention is not limited to the above embodiments, and improvements and modifications made by those skilled in the art based on the disclosure of the present invention without departing from the scope of the present invention should be within the scope of protection of the present invention.

Claims

1. A method for preparing a POC injectable particulate material, characterized in that: The following steps are involved: The POC degradable prepolymer is freeze-dried for later use; The freeze-dried POC degradable prepolymer is dissolved in a solvent, and inorganic salt particles are added. After a cross-linking reaction, the POC injectable particulate material is obtained by grinding, removing the inorganic salt, and drying.

2. The method for preparing a POC degradable prepolymer according to claim 1, characterized in that: The solvent is selected from one or a combination of dioxane, anhydrous ethanol, acetone or dichloromethane; The dosage relationship between the freeze-dried POC degradable prepolymer and the solvent is 10g:10-200mL.

3. The method for preparing a POC degradable prepolymer according to claim 1, characterized in that: The inorganic salt particles are selected from one or a combination of NaCl, KCl, and MgCl2; the added amount of the inorganic salt particles is 1-20 times the mass of the freeze-dried POC degradable prepolymer.

4. The method for preparing a POC degradable prepolymer according to claim 1, characterized in that: The conditions of the cross-linking reaction are: >50° C., cross-linking for 12-72 hours under vacuum or no vacuum conditions.

5. The method for preparing a POC degradable prepolymer according to claim 1, characterized in that: The preparation method of the POC degradable prepolymer comprises the following steps: The citric acid and 1,8-octanediol are mixed and melted, and then heated and stirred under vacuum conditions to react to form a POC prepolymer; The POC prepolymer is dissolved by an organic solvent to obtain a solution, and the solution is slowly poured into ultrapure water for purification multiple times to obtain a purified POC degradable prepolymer.

6. The method for preparing a POC degradable prepolymer according to claim 5, characterized in that: The amount of citric acid and 1,8-octanediol used is in a weight ratio of 1:(1-10) to (1-10):1; the mixed melting of citric acid and 1,8-octanediol is carried out under the protection of an inert gas, and the temperature of the mixed melting is 120-160° C.; The stirring reaction under vacuum conditions is carried out at 120-140° C. and a stirring speed of 300-1000 rpm for 1-48 hours, and then the speed is reduced until continuous stirring is impossible, and the reaction is stopped.

7. The method for preparing a POC degradable prepolymer according to claim 5, characterized in that: The volume mass ratio of the organic solvent to the POC prepolymer is 1-100:1; The organic solvent is selected from one or a combination of dioxane, anhydrous ethanol, acetone or dichloromethane; Dissolving the POC prepolymer by an organic solvent is carried out under stirring conditions, wherein the stirring conditions are 50-1000 rpm; When the dissolved solution is slowly poured into ultrapure water for purification multiple times, the volume ratio of the dissolved solution to the ultrapure water is 1:10-1000.

8. A POC injectable particulate material, characterized in that: The POC injectable particulate material is prepared by the method according to any one of claims 1 to 7.

9. A citric acid-based microparticle injection material, characterized in that: The method comprises the POC injectable particulate material according to claim 8, and drugs, active molecules, bioglass or metal ions loaded on the POC injectable particulate material.

10. A citric acid-based microparticle injection material according to claim 9, characterized in that: The drug or active molecule is selected from adenosine triphosphate and glutamine.

11. A citric acid-based microparticle injection material according to claim 10, characterized in that: The loading amount of the adenosine triphosphate relative to 1 gram of the POC injectable microparticle material is 5-50 mg, and the loading amount of the glutamine relative to 1 gram of the POC injectable microparticle material is 10-100 mg.

12. The method for preparing a citric acid-based microparticle injection material according to claim 9 or 10, characterized in that: The drug or active molecule is combined with the POC injectable microparticle material by incubation or chemical cross-linking. The incubation method is as follows: dissolving the drug or active molecule and the POC injectable microparticle material in physiological saline or culture medium, incubating them for 12-48 hours by physical adsorption through a micro-oscillator, and then separating, purifying, and drying them by centrifugation at 1000-2000 rpm for multiple times; The chemical cross-linking method is that the drug or active molecule is connected to the POC injectable microparticle material through a chemical reaction.

13. A citric acid-based biomimetic material, characterized in that: The invention comprises the POC injectable particulate material according to claim 8, and one or a combination of collagen, chitosan, hyaluronic acid or hydroxyapatite loaded on the POC injectable particulate material.

14. A citric acid-based biomimetic material according to claim 13, characterized in that: The loading amount of one or a combination of collagen, chitosan, hyaluronic acid or hydroxyapatite is 0%-50wt% and is not zero.

15. The citric acid-based biomimetic material according to claim 13, characterized in that: The POC injectable microparticle material is also loaded with drugs or active molecules.

16. The citric acid-based biomimetic material according to claim 15, characterized in that: The method comprises the POC injectable particulate material according to claim 8, adenosine triphosphate and glutamine loaded on the POC injectable particulate material, and one or a combination of collagen, chitosan, hyaluronic acid or hydroxyapatite loaded on the POC injectable particulate material.

17. An injection material, characterized in that: The injection material is composed of the following formula components in weight percentage: 1%-50% of the POC injectable microparticle material according to claim 8, 0.5-10% of sodium carboxymethyl cellulose CMC, and 40-95% of sterile physiological saline; Or the injectable material is obtained by dispersing the POC injectable microparticle material according to claim 8 with collagen, hyaluronic acid, bioglass or hydroxyapatite microparticles in sterile saline.

18. A use of the citric acid-based microparticle injection material according to claim 9 or the citric acid-based biomimetic material according to claim 12, characterized in that: The citric acid-based microparticle injection material or the citric acid-based biomimetic material is used to prepare biomedical cosmetic materials.

19. The use according to claim 17, characterized in that: The biomedical cosmetic materials include skin tissue dermal filling materials and breast augmentation prosthesis materials.

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