阅读说明：本技术 一种多功能生物活性微晶玻璃纳米材料及其制备方法和应用 (Multifunctional bioactive glass ceramic nano material and preparation method and application thereof ) 是由 雷波 牛雯 于 2021-07-30 设计创作，主要内容包括：本发明公开了一种多功能生物活性微晶玻璃纳米材料及其制备方法和应用,方法包括以下步骤：基于溶胶凝胶模板法以TEOS、TEP、四水硝酸钙分别作为硅源、磷源和钙源合成BGN；将BGN与乙酰丙酮钼混合并通过水热反应得到钼掺杂的生物活性微晶玻璃纳米材料。本发明使用的溶胶-凝胶模板法和水热法的方法环保、操作方便、原料成本低；制备的Mo-BGN具有优异的光热性能、抗氧化性能、生物相容性、抗炎性质和促血管能力,同时,在近红外激光照射下Mo-BGN也表现出抗菌和抗肿瘤性质,因此该纳米材料在感染/肿瘤术后组织再生中有着很好的应用前景。(The invention discloses a multifunctional bioactive glass ceramics nano material and a preparation method and application thereof, wherein the method comprises the following steps: synthesizing BGN by respectively taking TEOS, TEP and calcium nitrate tetrahydrate as a silicon source, a phosphorus source and a calcium source based on a sol-gel template method; mixing BGN and molybdenum acetylacetonate, and carrying out hydrothermal reaction to obtain the molybdenum-doped bioactive glass ceramic nano material. The sol-gel template method and the hydrothermal method used in the invention are environment-friendly, convenient to operate and low in raw material cost; the prepared Mo-BGN has excellent photo-thermal property, oxidation resistance, biocompatibility, anti-inflammatory property and blood vessel promoting capability, and simultaneously, the Mo-BGN also shows antibacterial and anti-tumor properties under the irradiation of near-infrared laser, so the nano material has good application prospect in tissue regeneration after infection/tumor operation.)

1. The preparation method of the multifunctional bioactive glass ceramic nano material is characterized by comprising the following steps:

synthesizing bioactive glass nanoparticles by taking tetraethoxysilane, triethyl phosphate and calcium nitrate tetrahydrate as a silicon source, a phosphorus source and a calcium source respectively based on a sol-gel template method;

mixing the bioactive glass nano particles with molybdenum acetylacetonate, and carrying out hydrothermal reaction to obtain the molybdenum-doped bioactive glass ceramic nano material.

2. The preparation method of the multifunctional bioactive glass ceramic nano material according to claim 1, wherein the specific method for synthesizing bioactive glass nano particles comprises the following steps:

adding a template agent into a solvent, stirring until the template agent is fully dissolved to obtain a mixed solution, adding ethyl orthosilicate, fully reacting, adding an aqueous solution of triethyl phosphate and calcium nitrate tetrahydrate, and fully reacting until the reaction is finished; and centrifuging and washing the reaction product, and calcining after freeze drying to obtain the bioactive glass nano-particles.

3. The method for preparing the multifunctional bioactive glass ceramic nano material as claimed in claim 2, wherein the template agent is dodecylamine or bromohexadecylpyridine.

4. The method for preparing the multifunctional bioactive glass ceramic nano material as claimed in claim 3, wherein the template agent is dodecylamine, and the dodecylamine is added into the mixed solution of the absolute ethyl alcohol and the deionized water, and stirred until the mixed solution is fully dissolved to obtain the mixed solution.

5. The preparation method of the multifunctional bioactive glass-ceramic nano material as claimed in claim 3, wherein the template agent is cetylpyridinium bromide, the cetylpyridinium bromide and urea are completely dissolved in a mixed solution containing cyclohexane and deionized water, isopropyl alcohol is added after full stirring, and the mixed solution is obtained after full stirring.

6. The preparation method of the multifunctional bioactive glass ceramic nano material according to claim 1, wherein the hydrothermal reaction comprises the following specific steps:

completely dispersing molybdenum acetylacetonate and bioactive glass nano particles in an ethanol solution according to the mass ratio of (3-18) to 1, and performing ultrasonic treatment to uniformly mix the particles; carrying out hydrothermal reaction at 160-200 ℃ to obtain a precipitate, and freeze-drying the precipitate to obtain the molybdenum-doped bioactive glass ceramic nano material.

7. The method for preparing the multifunctional bioactive glass-ceramic nano material according to claim 1, wherein the chemical composition of the bioactive glass nano particles is 80SiO₂-16CaO-4P₂O₅And 60SiO₂-36CaO-4P₂O₅。

8. A multifunctional bioactive glass ceramic nano material is characterized by being prepared by the method of any one of claims 1 to 7.

9. The multifunctional bioactive microcrystalline glass nanomaterial as claimed in claim 8, wherein the microcrystalline glass nanomaterial has a crystalline structure and an amorphous structure, and the valence state of Mo exists in two forms, namely +4 and + 6.

10. The multifunctional bioactive glass ceramics nano material prepared by the method of any one of claims 1 to 7 is applied to a repairing material for tissue regeneration after infection/tumor operation.

Technical Field

The invention belongs to the technical field of degradable biomedical materials, and particularly relates to a multifunctional bioactive glass ceramic nano material, and a preparation method and application thereof.

Background

The skin, as the first line of defense against viral entry, inevitably faces serious cancer and trauma threats. Although there are established treatment regimens for both skin tumor therapy and wound repair, the synergistic treatment of tumor and wound remains a significant clinical challenge. Particularly for the postoperative treatment of local solid tumors, not only is there a risk of tumor recurrence, but the chronic inflammatory microenvironment at the site of tumor residual also has a negative impact on the healing of the numerous tissue defects resulting from surgical resection of the tumor. In recent years, with the continuous and deep research of biological materials, the development of degradable biological platforms with the purpose of treatment and repair is the key for the successful treatment of skin-related diseases. Although the currently developed multifunctional biomaterials have application advantages in the fields of tumor treatment and tissue repair, the superposition of multiple functional components increases the difficulty of preparation and degradation, and the efficiency of treatment and repair is still to be improved.

Currently, in the use of synthetic biodegradable materials, Bioactive Glass Nanoparticles (BGNs) have significant advantages in biomedical applications including drug/gene delivery, tumor treatment, and tissue repair due to their controllable structure, excellent biodegradability, simple synthesis technology, and low preparation cost. However, the research and application of BGN are still in the primary stage at present, and difficulties and challenges of easy agglomeration, lack of modification sites and the like exist. However, with the complication of clinical requirements, by controlling the components of the BGN, designing multifunctional BGN containing other functional ions is an effective method for solving the application problem.

Disclosure of Invention

The invention aims to provide a multifunctional bioactive microcrystalline glass nano material and a preparation method thereof, the method is simple in process, and the obtained nano material has good biocompatibility, oxidation resistance and photo-thermal property and shows great application advantages in infection/tumor treatment and postoperative tissue repair.

In order to achieve the purpose, the invention adopts the following technical scheme to realize the purpose:

a preparation method of a multifunctional bioactive glass ceramic nano material comprises the following steps:

synthesizing bioactive glass nanoparticles by taking tetraethoxysilane, triethyl phosphate and calcium nitrate tetrahydrate as a silicon source, a phosphorus source and a calcium source respectively based on a sol-gel template method;

mixing the bioactive glass nano particles with molybdenum acetylacetonate, and carrying out hydrothermal reaction to obtain the molybdenum-doped bioactive glass ceramic nano material.

As a further improvement of the invention, the specific method for synthesizing the bioactive glass nanoparticles comprises the following steps:

adding a template agent into a solvent, stirring until the template agent is fully dissolved to obtain a mixed solution, adding ethyl orthosilicate, fully reacting, adding an aqueous solution of triethyl phosphate and calcium nitrate tetrahydrate, and fully reacting until the reaction is finished; and centrifuging and washing the reaction product, and calcining after freeze drying to obtain the bioactive glass nano-particles.

As a further improvement of the invention, the template agent is dodecylamine or bromohexadecylpyridine.

As a further improvement of the invention, the template agent is dodecylamine, and the dodecylamine is added into the mixed solution of the absolute ethyl alcohol and the deionized water and stirred until the dodecylamine is fully dissolved to obtain a mixed solution.

As a further improvement of the invention, the template agent is cetylpyridinium bromide, the cetylpyridinium bromide and urea are completely dissolved in a mixed solution containing cyclohexane and deionized water, isopropanol is added after full stirring, and the mixed solution is obtained after full stirring until full dissolution.

As a further improvement of the invention, the specific method of the hydrothermal reaction is as follows:

completely dispersing molybdenum acetylacetonate and bioactive glass nano particles in an ethanol solution according to the mass ratio of (3-18) to 1, and performing ultrasonic treatment to uniformly mix the particles; carrying out hydrothermal reaction at 160-200 ℃ to obtain a precipitate, and freeze-drying the precipitate to obtain the molybdenum-doped bioactive glass ceramic nano material.

As a further improvement of the invention, the chemical composition of BGN is 80SiO₂-16CaO-4P₂O₅And 60SiO₂-36CaO-4P₂O₅。

A multifunctional bioactive glass ceramics nano material is prepared by the method.

The microcrystalline glass nano material has a crystalline state and an amorphous state structure, and the valence state of Mo exists in two forms of +4 and + 6.

An application of multifunctional bioactive glass ceramics nano material as a repairing material for tissue regeneration after infection/tumor operation.

Compared with the prior art, the invention has the following beneficial effects:

the invention provides a preparation method of a multifunctional bioactive glass ceramics nano material for tissue regeneration after infection/tumor operation, aiming at the defects of single function, limited surface modification sites and the like of the traditional bioactive glass material, the method firstly synthesizes BGN particles by a classical sol-gel template method or an improved template method, and then obtains a multivalent molybdenum doped bioactive glass ceramics material (Mo-BGN) by taking human microelement molybdenum as doping ions and through hydrothermal reaction; the microcrystalline glass material acts on an infected wound or a postoperative wound of a tumor in an in-situ coating mode to play the multifunctional functions of antibiosis, anti-tumor and tissue healing. The ion doping technology used by the invention has the advantages of rich doped ions, simple doping process, controllable product morphology and the like, and the related template synthesis technology and hydrothermal synthesis method are safe and environment-friendly, convenient to operate and low in cost. The experimental results prove that: the molybdenum-doped bioactive material prepared by the method is calcium molybdate (CaMoO)₄) Molybdenum dioxide (MoO)₂) The microcrystalline glass composed of the amorphous glass has stronger photo-thermal heating capacity and excellent oxidation resistance, and the good biocompatibility, photo-thermal anti-tumor and photo-thermal antibacterial effects can effectively kill tumor cells and bacteria, promote the migration of endothelial cells and show a certain effect of promoting tissue repair. The BGN used in the invention has good biocompatibility and compatibilityThe degradation property is realized, the BGN is endowed with additional antibacterial and antitumor functions through the doping of molybdenum, and the synthesized Mo-BGN not only can not cause immunogenic reaction, but also can play corresponding functions in the inflammation stage and the proliferation stage related to wound healing through antibacterial and vascularization promoting effects.

The invention also has the following advantages:

1) the BGN synthesis method used in the invention is a sol-gel template method, and the template-catalyst dodecylamine (DDA) or bromohexadecylpyridine (CPB) belongs to a surfactant, has good solubility and is cheap and easy to obtain; particularly, BGN synthesized by using a template-catalyst CPB has a dendritic morphology, and a large specific surface area of the BGN provides abundant sites for ion doping.

2) According to the invention, the BGN is modified by utilizing the hydrothermal reaction of molybdenum acetylacetonate, the traditional bioactive material shows multifunctional bioactivity through a simple synthesis process, and compared with a nano composite material, the composite material has the advantages of simple process and stable performance.

3) The Mo-BGN has excellent photo-thermal property and oxidation resistance due to the doping of molybdenum ions; the reason for the photo-thermal property of the material is probably MoO in Mo-BGN₂Due to the oxygen vacancy structure of Mo-BGN, the oxidation resistance of the Localized Surface Plasmon Resonance (LSPR) effect can effectively capture free radicals.

4) The doping of Mo ions in the invention can not affect the inherent blood compatibility and cell compatibility of BGN, mainly because Mo element as a trace element in human body can not affect the normal metabolism of life.

5) The Mo-BGN prepared by the invention has biocompatible photo-thermal temperature (42 ℃) under laser irradiation, and the temperature can selectively generate irreversible damage to bacteria and tumor cells and does not influence the cell activity of normal tissue cells; the in vitro cell migration, anti-inflammation and angiogenesis promotion experiments show that the antioxidant activity and bioactive glass components (Si and Ca) of the Mo-BGN have continuous endothelial cell migration, anti-inflammation and angiogenesis promotion effects on the wound surface environment, and provide favorable conditions for repairing infected wound surfaces and postoperative wound surfaces of tumors.

Drawings

FIG. 1 is a schematic structural diagram of a nanomaterial of the present invention;

FIG. 2 is a 15Mo-BGN (molybdenum acetylacetonate: BGN ═ 15:1) TEM morphology photograph of the multifunctional bioactive glass ceramics nano material synthesized by the invention and used for tissue regeneration after infection/tumor operation;

FIG. 3 is the physicochemical characterization results of the 15Mo-BGN produced. Wherein, FIG. 3A is FTIR spectra; FIG. 3B is an XRD pattern; FIG. 3C is an XPS spectrum; FIG. 3D is a UV-Vis-NIR spectrum;

FIG. 4 shows the photo-thermal and oxidation resistance properties of 15Mo-BGN prepared in the invention. FIG. 4A is a graph of temperature versus irradiation time for a sample irradiated with a 808nm laser; FIG. 4B is a cyclical stability curve; FIG. 4C is a UV-Vis spectrum of a DPPH solution after incubation of the solution with the material for 30 minutes; FIG. 4D is the oxidation resistance;

fig. 5 shows the toxicity test results of the bioactive glass ceramics nano-material prepared by the invention on staphylococcus aureus (s. aureus, fig. 5A), methicillin-resistant staphylococcus aureus (MRSA, fig. 5B), human umbilical vein endothelial cells (HUVEC, fig. 5C) and human malignant melanoma cells (a375, fig. 5D) before and after laser action, and "+" represents laser irradiation;

FIG. 6 is a graph showing the anti-inflammatory and vasogenic effects of 15Mo-BGN prepared according to the invention. Wherein, FIG. 6A shows the expression result of inflammatory factor TNF-alpha; FIG. 6B shows the expression result of IL-1 β; FIG. 6C shows the results of IL-10 expression; FIG. 6D shows the relative vascular factor (CD31) expression results;

fig. 7 shows the result of the tissue regeneration after infection/tumor operation of the bioactive glass-ceramic nanomaterial prepared by the invention. Wherein, fig. 7A is a macroscopic result of MRSA-infected skin wound repair; fig. 7B is a macroscopic result of tumor-partially resected skin wound repair.

Detailed Description

The invention is described in further detail below with reference to the accompanying drawings:

the invention aims to prepare the bioactive glass ceramics nano material with good biocompatibility, antibiosis and anti-tumor properties, and realizes tissue regeneration after infection/tumor operation by utilizing the photo-thermal and oxidation resistance of the material.

Bioactive Glass Nanoparticles (BGNs) have been widely used in the biomedical field because of their controlled biodegradation, good biocompatibility, and low cost. However, BGN has no antibacterial and antitumor properties, so that the biological function of the BGN is limited; the ion doping technology is one of important means for modifying BGN, and the main strategy is to introduce functional ions into BGN to realize the regulation and control of the characteristics of the BGN, such as light, heat, electricity, magnetism and the like.

Molybdenum (Mo) is an important trace element in human bodies and has good biological effect on different tissues. Molybdenum is a trace element essential to living body, is very important for normal metabolism of life, and is in high valence state (Mo)⁶⁺) Molybdenum MoO in low valence state, associated with bone and tooth growth₂Can be used as a photo-thermal agent to show controllable temperature rising effect under Near Infrared (NIR) stimulation. On the other hand, Mo is a typical valence-variable element, Mo⁶⁺And Mo⁴⁺Respectively has the tissue growth promoting ability and the photo-thermal property. Can be prepared by mixing Mo⁶⁺And Mo⁴⁺Single-component multifunctional Mo-BGN is designed by introducing BGN, and material support is provided for tumor/infected photothermal therapy and tissue repair. The Mo-BGN is synthesized by BGN and molybdenum acetylacetonate through hydrothermal reaction, has the advantages of strong antioxidant activity, high photothermal conversion efficiency, low toxicity, biodegradability and the like, and if the morphology of the initial BGN can be controlled and the dosage of the molybdenum acetylacetonate can be adjusted in the synthesis process, the efficiency of the Mo-BGN material on infection/tumor treatment and postoperative tissue repair can be greatly improved.

If the multivalent Mo is doped into the BGN network, the BGN can be endowed with photothermal treatment and repair promotion functions. Therefore, the invention realizes the doping of the multivalent Mo by utilizing the hydrothermal reaction, designs and synthesizes the Mo-BGN with strong antioxidant activity, high photothermal conversion efficiency and low toxicity. And the physical and chemical properties, biocompatibility, in-vivo and in-vitro photo-thermal properties and the functions and mechanisms of the synthesized multivalent Mo-doped bioactive glass nanoparticles in the wound healing process caused by bacterial infection and tumor excision are researched.

Therefore, the first purpose of the invention is to provide a preparation method of a multifunctional bioactive microcrystalline glass nano material, which comprises the following steps:

1) synthesizing BGN by a sol-gel template method: adding anhydrous ethanol and deionized water into a 250mL round-bottom flask according to the molar ratio of 3:1, fully mixing, then adding 10g of template-catalyst dodecylamine (DDA), stirring in an environment at 40 ℃ until the DDA is fully dissolved, then dropping 4mL of Tetraethoxysilane (TEOS) into the mixed solution under the condition of continuous stirring, dropping triethyl phosphate (TEP) into the reaction system after reacting for 30 minutes, continuing to stir for 15 minutes, adding 1mL of 5mol L^-1Reacting the aqueous solution of calcium nitrate tetrahydrate for 3 hours at the temperature of 40 ℃; centrifugally collecting reaction products, washing the reaction products by absolute ethyl alcohol and deionized water, and calcining the reaction products for 3-10 hours in a muffle furnace at 600-650 ℃ after freeze drying to obtain BGN products;

2) preparing Mo-BGN by a hydrothermal method: molybdenum acetylacetonate and BGN were completely dispersed in 10mL of a 75% ethanol solution in a mass ratio of (3-18): 1. Secondly, the solution is subjected to ultrasonic treatment for 20-40 minutes, and the molybdenum acetylacetonate and BGN are uniformly mixed. And then directly transferring the solution into a 25mL high-pressure reaction kettle, and preserving the temperature in an oven at 160-200 ℃ for 8-12 hours. Cooling the high-pressure reaction kettle to room temperature, taking out the suspension in the polytetrafluoroethylene lining, centrifuging at high speed to obtain a precipitate, and freeze-drying the precipitate to obtain a molybdenum-doped bioactive glass ceramic nano material (Mo-BGN);

the invention further improves the following steps:

the chemical composition of BGN in the step 1) can be 80SiO₂-16CaO-4P₂O₅And 60SiO₂-36CaO-4P₂O₅。

Dodecylamine (DDA) or bromohexadecylpyridine (CPB) may be used as a templating agent in step 1). If CPB is used, the corresponding synthesis process is modified as follows: completely dissolving 1.25g of CPB and 0.75g of urea in a mixed solution containing 37.5mL of cyclohexane and 37.5mL of deionized water, adding 1.15g of isopropanol after vigorously stirring for 15 minutes, and continuing stirring for 2 hours at the environment of 25 ℃; subsequently, 3.40mL of TEOS was added dropwise to the mixed solution while maintaining stirring, and the temperature was raised to 70 ℃ after 30 minutes of reaction. Stirring was continued for 7.5 hours, and after the reaction temperature stabilized, 0.77mL of TEP and 1mL of an aqueous solution of calcium nitrate tetrahydrate were added in that order at 30-minute intervals. After further stirring for 16 hours, a white bioactive glass sol was obtained. And then washing the gel separated by centrifugation with acetone, ethanol and deionized water for three times respectively to prepare the BGN precursor. And after the precursor is freeze-dried, calcining for 3-10 hours at 600-650 ℃ to remove residual organic matters and obtain the BGN with the radial morphology.

The molar ratio of molybdenum acetylacetonate to BGN in step 2) is 3:1, 9:1, 15:1 and 18: 1.

The second purpose of the invention is to provide the multivalent molybdenum-doped bioactive material prepared based on the method, which is a multifunctional bioactive glass ceramic nanomaterial, and the reaction mechanism and the product structure of the nanomaterial are shown in fig. 1.

The multifunctional bioactive microcrystalline glass nano material prepared by the method has Mo⁶⁺And Mo⁴⁺The tissue growth promoting and photo-thermal heating performance. When the Mo-BGN is applied to the body in an in-situ coating mode, the Mo-BGN not only can promote the healing of various wounds caused by bacterial infection and surgical tumor excision, but also can effectively inhibit the recurrence of tumors. Particularly in post-tumor resection treatment, laser-mediated warming of the material can bring the wound environment to 42 ℃, which is a temperature sufficient to kill tumor cells and bacteria, but promote endothelial cell migration. Meanwhile, the oxidation resistance caused by a large number of oxygen vacancies in the Mo-BGN has an obvious promotion effect on tissue repair.

The third purpose of the invention is to provide the application of the multifunctional bioactive glass ceramics nano material as a repairing material for tissue regeneration after infection/tumor operation.

The result shows that the multifunctional microcrystalline glass with bioactivity is synthesized by an in-situ hydrothermal method. The Mo-BGN has a nanoparticle shape with good dispersibility; the physical and chemical structure characterization result shows that the Mo-BGN has CaMoO₄-MoO₂Microcrystalline glass structure of-BGN, Mo ions in Mo⁴⁺/Mo⁶⁺Is in the form of Mo-BGN; due to the Mo-BGN structurePresence of MoO₂The crystal form and rich oxygen vacancy, Mo-BGN has selective photo-thermal anticancer, antibacterial, free radical scavenging, antioxidant, anti-inflammatory and good angiogenesis activity. Mo-BGN not only can effectively improve tissue reconstruction of infected wound and tumor resection defect, but also can inhibit tumor recurrence, and has good tissue safety. The work provides a good strategy for designing multifunctional bioactive nano materials with simple components to treat the tissue regeneration of complex disease injury.

For better understanding of the present invention, the present invention will be described in detail with reference to the following embodiments, but the present invention is not limited to the following examples.

Example 1

1) Synthesizing BGN by a classical sol-gel template method: adding absolute ethyl alcohol and deionized water into a 250mL round-bottom flask according to the molar ratio of 3:1, fully mixing, then adding 10g of template-catalyst DDA, stirring at 40 ℃ until the DDA is fully dissolved, then dripping 4mL of Tetraethoxysilane (TEOS) into the mixed solution under the condition of continuous stirring, after reacting for 30 minutes, dripping TEP into the reaction system, continuing stirring for 15 minutes, adding 1mL of 5mol L^-1Reacting the aqueous solution of calcium nitrate tetrahydrate for 3 hours at the temperature of 40 ℃; centrifugally collecting reaction products, washing the reaction products by absolute ethyl alcohol and deionized water, and calcining the reaction products for 3 hours in a muffle furnace at 650 ℃ after freeze drying to obtain a BGN product;

2) preparing Mo-3BGN by a hydrothermal method: molybdenum acetylacetonate and BGN were completely dispersed in 10mL of 75% ethanol solution at a mass ratio of 3: 1. Secondly, the solution is treated by ultrasonic for 30 minutes to ensure that the molybdenum acetylacetonate and BGN are uniformly mixed. The solution was then transferred directly to a 25mL autoclave and incubated in an oven at 180 ℃ for 10 hours. And (3) cooling the high-pressure reaction kettle to room temperature, taking out the suspension in the polytetrafluoroethylene lining, centrifuging at a high speed to obtain a precipitate, and freeze-drying the precipitate to obtain the molybdenum-doped bioactive glass ceramic nano material (Mo-3 BGN).

Example 2

1) Synthesizing BGN by a classical sol-gel template method: absolute ethyl alcohol and deionized water according to a molar ratioAdding the mixture into a 250mL round-bottom flask in a ratio of 3:1, fully mixing, then adding 10g of template-catalyst DDA, stirring in an environment at 40 ℃ until the DDA is fully dissolved, then dropping 4mL of Tetraethoxysilane (TEOS) into the mixed solution under the condition of continuous stirring, dropping TEP into the reaction system after reacting for 30 minutes, continuing stirring for 15 minutes, adding 1mL of 5mol L^-1Reacting the aqueous solution of calcium nitrate tetrahydrate for 3 hours at the temperature of 40 ℃; centrifugally collecting reaction products, washing the reaction products by absolute ethyl alcohol and deionized water, and calcining the reaction products for 5 hours in a muffle furnace at 630 ℃ after freeze drying to obtain a BGN product;

2) preparing Mo-3BGN by a hydrothermal method: molybdenum acetylacetonate and BGN were completely dispersed in 10mL of 75% ethanol solution at a mass ratio of 9: 1. Secondly, the solution is treated by ultrasonic for 30 minutes to ensure that the molybdenum acetylacetonate and BGN are uniformly mixed. The solution was then transferred directly to a 25mL autoclave and incubated in an oven at 180 ℃ for 10 hours. And (3) cooling the high-pressure reaction kettle to room temperature, taking out the suspension in the polytetrafluoroethylene lining, centrifuging at a high speed to obtain a precipitate, and freeze-drying the precipitate to obtain the molybdenum-doped bioactive glass ceramic nano material (Mo-9 BGN).

Example 3

1) Synthesizing BGN by a classical sol-gel template method: adding absolute ethyl alcohol and deionized water into a 250mL round-bottom flask according to the molar ratio of 3:1, fully mixing, then adding 10g of template-catalyst DDA, stirring at 40 ℃ until the DDA is fully dissolved, then dripping 4mL of Tetraethoxysilane (TEOS) into the mixed solution under the condition of continuous stirring, after reacting for 30 minutes, dripping TEP into the reaction system, continuing stirring for 15 minutes, adding 1mL of 5mol L^-1Reacting the aqueous solution of calcium nitrate tetrahydrate for 3 hours at the temperature of 40 ℃; centrifugally collecting reaction products, washing the reaction products by absolute ethyl alcohol and deionized water, and calcining the reaction products for 6 hours in a muffle furnace at the temperature of 620 ℃ after freeze drying to obtain a BGN product;

2) preparing Mo-15BGN by a hydrothermal method: molybdenum acetylacetonate and BGN were completely dispersed in 10mL of 75% ethanol solution at a mass ratio of 15: 1. Secondly, the solution is treated by ultrasonic for 30 minutes to ensure that the molybdenum acetylacetonate and BGN are uniformly mixed. The solution was then transferred directly to a 25mL autoclave and incubated in an oven at 180 ℃ for 10 hours. And (3) cooling the high-pressure reaction kettle to room temperature, taking out the suspension in the polytetrafluoroethylene lining, centrifuging at a high speed to obtain a precipitate, and freeze-drying the precipitate to obtain the molybdenum-doped bioactive glass ceramic nano material (Mo-15 BGN).

Example 4

1) Synthesizing BGN by a classical sol-gel template method: adding absolute ethyl alcohol and deionized water into a 250mL round-bottom flask according to the molar ratio of 3:1, fully mixing, then adding 10g of template-catalyst DDA, stirring at 40 ℃ until the DDA is fully dissolved, then dripping 4mL of Tetraethoxysilane (TEOS) into the mixed solution under the condition of continuous stirring, after reacting for 30 minutes, dripping TEP into the reaction system, continuing stirring for 15 minutes, adding 1mL of 5mol L^-1Reacting the aqueous solution of calcium nitrate tetrahydrate for 3 hours at the temperature of 40 ℃; centrifugally collecting reaction products, washing the reaction products by absolute ethyl alcohol and deionized water, and calcining the reaction products for 3 hours in a muffle furnace at 650 ℃ after freeze drying to obtain a BGN product;

2) preparing Mo-18BGN by a hydrothermal method: molybdenum acetylacetonate and BGN were completely dispersed in 10mL of 75% ethanol solution at a mass ratio of 18: 1. Secondly, the solution is treated by ultrasonic for 30 minutes to ensure that the molybdenum acetylacetonate and BGN are uniformly mixed. The solution was then transferred directly to a 25mL autoclave and incubated in an oven at 180 ℃ for 10 hours. And (3) cooling the high-pressure reaction kettle to room temperature, taking out the suspension in the polytetrafluoroethylene lining, centrifuging at a high speed to obtain a precipitate, and freeze-drying the precipitate to obtain the molybdenum-doped bioactive glass ceramic nano material (Mo-18 BGN).

Example 5

1) Synthesizing BGN by an improved sol-gel template method: completely dissolving 1.25g of CPB and 0.75g of urea in a mixed solution containing 37.5mL of cyclohexane and 37.5mL of deionized water, adding 1.15g of isopropanol after vigorously stirring for 15 minutes, and continuing stirring for 2 hours at the environment of 25 ℃; subsequently, 3.40mL of TEOS was added dropwise to the mixed solution while maintaining stirring, and the temperature was raised to 70 ℃ after 30 minutes of reaction. Stirring was continued for 7.5 hours, and after the reaction temperature stabilized, 0.77mL of TEP and 1mL of an aqueous solution of calcium nitrate tetrahydrate were added in that order at 30-minute intervals. After further stirring for 16 hours, a white bioactive glass sol was obtained. And then washing the gel separated by centrifugation with acetone, ethanol and deionized water for three times respectively to prepare the BGN precursor. After the precursor is freeze-dried, calcining for 10 hours at 600 ℃ to remove residual organic matters and obtain BGN with radial morphology;

2) preparing Mo-3BGN by a hydrothermal method: molybdenum acetylacetonate and BGN were completely dispersed in 10mL of 75% ethanol solution at a mass ratio of 3: 1. Secondly, the solution is treated by ultrasonic for 30 minutes to ensure that the molybdenum acetylacetonate and BGN are uniformly mixed. The solution was then transferred directly to a 25mL autoclave and incubated in an oven at 180 ℃ for 10 hours. And (3) cooling the high-pressure reaction kettle to room temperature, taking out the suspension in the polytetrafluoroethylene lining, centrifuging at a high speed to obtain a precipitate, and freeze-drying the precipitate to obtain the molybdenum-doped bioactive glass ceramic nano material (Mo-3 BGN).

Example 6

1) Synthesizing BGN by an improved sol-gel template method: completely dissolving 1.25g of CPB and 0.75g of urea in a mixed solution containing 37.5mL of cyclohexane and 37.5mL of deionized water, adding 1.15g of isopropanol after vigorously stirring for 15 minutes, and continuing stirring for 2 hours at the environment of 25 ℃; subsequently, 3.40mL of TEOS was added dropwise to the mixed solution while maintaining stirring, and the temperature was raised to 70 ℃ after 30 minutes of reaction. Stirring was continued for 7.5 hours, and after the reaction temperature stabilized, 0.77mL of TEP and 1mL of an aqueous solution of calcium nitrate tetrahydrate were added in that order at 30-minute intervals. After further stirring for 16 hours, a white bioactive glass sol was obtained. And then washing the gel separated by centrifugation with acetone, ethanol and deionized water for three times respectively to prepare the BGN precursor. After the precursor is freeze-dried, calcining for 10 hours at 600 ℃ to remove residual organic matters and obtain BGN with radial morphology;

2) preparing Mo-9BGN by a hydrothermal method: molybdenum acetylacetonate and BGN were completely dispersed in 10mL of 75% ethanol solution at a mass ratio of 9: 1. Secondly, the solution is treated by ultrasonic for 30 minutes to ensure that the molybdenum acetylacetonate and BGN are uniformly mixed. The solution was then transferred directly to a 25mL autoclave and incubated in an oven at 180 ℃ for 10 hours. And (3) cooling the high-pressure reaction kettle to room temperature, taking out the suspension in the polytetrafluoroethylene lining, centrifuging at a high speed to obtain a precipitate, and freeze-drying the precipitate to obtain the molybdenum-doped bioactive glass ceramic nano material (Mo-9 BGN).

Example 7

1) Synthesizing BGN by an improved sol-gel template method: completely dissolving 1.25g of CPB and 0.75g of urea in a mixed solution containing 37.5mL of cyclohexane and 37.5mL of deionized water, adding 1.15g of isopropanol after vigorously stirring for 15 minutes, and continuing stirring for 2 hours at the environment of 25 ℃; subsequently, 3.40mL of TEOS was added dropwise to the mixed solution while maintaining stirring, and the temperature was raised to 70 ℃ after 30 minutes of reaction. Stirring was continued for 7.5 hours, and after the reaction temperature stabilized, 0.77mL of TEP and 1mL of an aqueous solution of calcium nitrate tetrahydrate were added in that order at 30-minute intervals. After further stirring for 16 hours, a white bioactive glass sol was obtained. And then washing the gel separated by centrifugation with acetone, ethanol and deionized water for three times respectively to prepare the BGN precursor. After the precursor is freeze-dried, calcining for 10 hours at 600 ℃ to remove residual organic matters and obtain BGN with radial morphology;

2) preparing Mo-15BGN by a hydrothermal method: molybdenum acetylacetonate and BGN were completely dispersed in 10mL of 75% ethanol solution at a mass ratio of 15: 1. Secondly, the solution is treated by ultrasonic for 30 minutes to ensure that the molybdenum acetylacetonate and BGN are uniformly mixed. The solution was then transferred directly to a 25mL autoclave and incubated in an oven at 180 ℃ for 10 hours. And (3) cooling the high-pressure reaction kettle to room temperature, taking out the suspension in the polytetrafluoroethylene lining, centrifuging at a high speed to obtain a precipitate, and freeze-drying the precipitate to obtain the molybdenum-doped bioactive glass ceramic nano material (Mo-15 BGN).

Example 8

1) Synthesizing BGN by an improved sol-gel template method: completely dissolving 1.25g of CPB and 0.75g of urea in a mixed solution containing 37.5mL of cyclohexane and 37.5mL of deionized water, adding 1.15g of isopropanol after vigorously stirring for 15 minutes, and continuing stirring for 2 hours at the environment of 25 ℃; subsequently, 3.40mL of TEOS was added dropwise to the mixed solution while maintaining stirring, and the temperature was raised to 70 ℃ after 30 minutes of reaction. Stirring was continued for 7.5 hours, and after the reaction temperature stabilized, 0.77mL of TEP and 1mL of an aqueous solution of calcium nitrate tetrahydrate were added in that order at 30-minute intervals. After further stirring for 16 hours, a white bioactive glass sol was obtained. And then washing the gel separated by centrifugation with acetone, ethanol and deionized water for three times respectively to prepare the BGN precursor. After the precursor is freeze-dried, calcining for 10 hours at 600 ℃ to remove residual organic matters and obtain BGN with radial morphology;

2) preparing Mo-18BGN by a hydrothermal method: molybdenum acetylacetonate and BGN were completely dispersed in 10mL of 75% ethanol solution at a mass ratio of 18: 1. Secondly, the solution is treated by ultrasonic for 30 minutes to ensure that the molybdenum acetylacetonate and BGN are uniformly mixed. The solution was then transferred directly to a 25mL autoclave and incubated in an oven at 180 ℃ for 10 hours. And (3) cooling the high-pressure reaction kettle to room temperature, taking out the suspension in the polytetrafluoroethylene lining, centrifuging at a high speed to obtain a precipitate, and freeze-drying the precipitate to obtain the molybdenum-doped bioactive glass ceramic nano material (Mo-18 BGN).

The multifunctional bioactive glass ceramics nano material (Mo-BGN) for tissue regeneration after infection/tumor operation, which is prepared by the invention, has inherent photothermal antioxidant antibacterial anti-inflammatory activity due to the special microcrystalline structure, oxygen vacancy and bioactive components. When Mo-BGN acts on an infected wound or a postoperative wound of a tumor, the material has a biocompatible photothermal temperature (42 ℃) under the mediation of 808nm laser, and the temperature can selectively kill tumor cells and various bacteria, but shows excellent cell compatibility to normal cells around the wound. In addition, the photothermal, antioxidant and bioactive glass components (bioactive Si and Ca) of Mo-BGN have a sustained anti-inflammatory and angiogenic effect on the wound environment, which makes it a significant advantage in applications for tissue regeneration after infection/tumor surgery, as analyzed in detail below in combination with experimental data.

Fig. 2 is a TEM topography photograph of the multifunctional bioactive glass ceramics nano material 15Mo-BGN (molybdenum acetylacetonate: BGN: 15:1) which can be used for tissue regeneration after infection/tumor operation and is synthesized by the present invention. As can be seen from TEM photographs with different magnifications, the hydrothermal reaction does not affect the dispersibility of the nanoparticles, and the original spherical morphology and particle size distribution range (200-300nm) of BGN can be maintained.

FIG. 3 is the physicochemical characterization results of the 15Mo-BGN produced. As can be seen from the FTIR spectra shown in FIG. 3A, 1632, 1100 and 801cm^-1The infrared absorption peak is attributed to the characteristic absorption peak of BGN, 945cm^-1The absorption band at (b) corresponds to the shear vibration of Mo-O-Mo. As can be seen from fig. 3B, the diffraction packet at 23.0 ° corresponds to amorphous silica, and the diffraction peaks at 25.3, 37.0, 38.6, and 48.1 ° correspond to CaMoO₄Diffraction peaks of (101), (112), (204), and (220) crystal planes of (JCPDS 29-0351); the diffraction peaks at 27.5, 36.1 and 54.4 ° 2 θ correspond to MoO₂(JCPDS No.29-0351) and (311). The product prepared by Mo doping was proved to be crystalline (CaMoO)₄With MoO₂) And amorphous state. This product, which has both crystalline and amorphous structures, is called a glass-ceramic. FIG. 3C shows 4 characteristic peaks, binding energies of 233.9eV and 231.8eV for Mo 3d_3/2Can be respectively assigned to Mo⁶⁺And Mo⁴⁺While Mo 2d at 229.6eV and 228.1eV_5/2Peaks correspond to Mo⁶⁺And Mo⁴⁺It is shown that Mo in 15Mo-BGN exists in two forms, namely +4 and + 6. The UV-Vis-NIR absorption spectrum shown in FIG. 3D shows that the material has an absorbance in the near infrared wavelength range of 400-1000 nm. Therefore, the photo-thermal temperature rise performance of the 15Mo-BGN material can be researched by using 808nm near-infrared laser.

FIG. 4 shows the photo-thermal and oxidation resistance properties of 15Mo-BGN prepared in the invention. As can be seen from FIG. 4A, the sample pairs are 808nm (0.8W cm)^-210 minutes) the absorption of the laser depends on the concentration of the sample. After laser irradiation, 100-^-1The temperature of 15Mo-BGN can reach 42 ℃. The photothermal cycling stability results shown in fig. 4B also illustrate the ability of the sample to maintain a stable and sustained temperature increase after undergoing five cycles of "laser on-laser off". The controllable photothermal capacity of 15Mo-BGN determines the application value of the compound in the fields of tumor treatment and efficient sterilization. As shown in FIG. 4C, 15Mo-BGN showed similarities to ascorbic acid (VC, positive control)And (4) oxidation resistance. Comparing the oxidation resistance of the materials in each group (FIG. 4D) found that the material was at 5. mu.g mL^-1The DPPH & ltSUB & gt can be eliminated by 50.9% under the concentration of the (D & ltSUB & gt), which indicates that 15Mo-BGN has strong free radical capture capacity. The presence of oxygen vacancies is presumed to be the main cause of its high antioxidant activity.

Fig. 5 shows the results of the toxicity measurements of the bioactive glass ceramics nano-material prepared by the invention on s.aureus bacteria, MRSA bacteria, HUVEC cells and a375 cells before and after the laser action. From the antibacterial results (FIGS. 5A and 4B), it can be seen that after 12 hours of exposure to the 15Mo-BGN sample, neither strain was significantly inhibited compared to the control without laser (Blank group). The cytocompatibility results shown in FIGS. 5D and 4E indicate that the material was between 0-250 μ g mL without laser irradiation^-1There was no significant cytotoxicity on both a375 and HUVEC cells in the concentration range, but laser irradiation was sufficient to kill more than 80% of a375 tumor cells, whereas near-infrared laser had no effect on HUVEC cell viability. The results show that the 15Mo-BGN can effectively kill bacteria and skin tumor cells under laser irradiation without affecting normal tissue cells.

FIG. 6 is a graph showing the anti-inflammatory and vasogenic effects of 15Mo-BGN prepared according to the invention. FIG. 6A shows the results of the expression of the inflammatory factor TNF- α; FIG. 6B shows the expression result of IL-1 β; FIG. 6C shows the results of IL-10 expression; fig. 6D shows the relative vascular factor (CD31) expression results. As shown in FIGS. 6A-6C, 15Mo-BGN significantly inhibited the expression of TNF- α and IL-1 β, but increased the expression of IL-10, as compared to the blank control. This result demonstrates that 15Mo-BGN can effectively inhibit inflammation. This significant anti-inflammatory effect is attributed to the multi-valence Mo doping, the oxygen vacancy structure of 15Mo-BGN facilitates the capture of free radicals. While free radicals are effectors of inflammatory reactions, oxygen radicals can induce inflammatory reactions via PRRS and non-PRRS pathways. Therefore, the inflammation is effectively relieved by consuming free radicals through 15Mo-BGN, and the skin wound surface repair caused by infection is promoted. In addition, as shown in fig. 6D, the expression of CD31 was higher in the 15Mo-BGN group than in the control group, indicating that it had good angiogenic ability.

Fig. 7 shows the result of the tissue regeneration after infection/tumor operation of the bioactive glass-ceramic nanomaterial prepared by the invention. FIG. 7A is a macroscopic view of MRSA-infected skin wound repair, showing that 15Mo-BGN + group had the fastest wound recovery rate within 14 days compared to the control group; fig. 7B is a macroscopic result of the tumor partially resected skin wound repair, and it can be seen from the figure that the wound surface of the 15Mo-BGN + group irradiated by near infrared was gradually closed within 14 days, and no obvious tumor recurrence was observed. The results show that the 15Mo-BGN has the effects of resisting bacteria, inhibiting tumor recurrence and promoting wound repair, and the results prove that the Mo-BGN is expected to be used as a repair material for tissue regeneration after infection/tumor operation.

Example 9

1) Synthesizing BGN by an improved sol-gel template method: completely dissolving 1.25g of CPB and 0.75g of urea in a mixed solution containing 37.5mL of cyclohexane and 37.5mL of deionized water, adding 1.15g of isopropanol after vigorously stirring for 15 minutes, and continuing stirring for 2 hours at the environment of 25 ℃; subsequently, 3.40mL of TEOS was added dropwise to the mixed solution while maintaining stirring, and the temperature was raised to 70 ℃ after 30 minutes of reaction. Stirring was continued for 7.5 hours, and after the reaction temperature stabilized, 0.77mL of TEP and 1mL of an aqueous solution of calcium nitrate tetrahydrate were added in that order at 30-minute intervals. After further stirring for 16 hours, a white bioactive glass sol was obtained. And then washing the gel separated by centrifugation with acetone, ethanol and deionized water for three times respectively to prepare the BGN precursor. After the precursor is freeze-dried, calcining for 6 hours at the temperature of 620 ℃ to remove residual organic matters and obtain BGN with radial morphology;

2) preparing Mo-18BGN by a hydrothermal method: molybdenum acetylacetonate and BGN were completely dispersed in 10mL of 75% ethanol solution at a mass ratio of 6: 1. Secondly, the solution is treated by ultrasonic for 30 minutes to ensure that the molybdenum acetylacetonate and BGN are uniformly mixed. The solution was then transferred directly to a 25mL autoclave and incubated in an oven at 160 ℃ for 12 hours. And (3) cooling the high-pressure reaction kettle to room temperature, taking out the suspension in the polytetrafluoroethylene lining, centrifuging at a high speed to obtain a precipitate, and freeze-drying the precipitate to obtain the molybdenum-doped bioactive glass ceramic nano material (Mo-6 BGN).

Example 10

1) Improvement of the structureSynthesizing BGN by a sol-gel template method: adding absolute ethyl alcohol and deionized water into a 250mL round-bottom flask according to the molar ratio of 3:1, fully mixing, then adding 10g of template-catalyst DDA, stirring at 40 ℃ until the DDA is fully dissolved, then dripping 4mL of Tetraethoxysilane (TEOS) into the mixed solution under the condition of continuous stirring, after reacting for 30 minutes, dripping TEP into the reaction system, continuing stirring for 15 minutes, adding 1mL of 5mol L^-1Reacting the aqueous solution of calcium nitrate tetrahydrate for 3 hours at the temperature of 40 ℃; centrifugally collecting reaction products, washing the reaction products by absolute ethyl alcohol and deionized water, and calcining the reaction products for 5 hours in a muffle furnace at 640 ℃ after freeze drying to obtain a BGN product;

2) preparing Mo-18BGN by a hydrothermal method: molybdenum acetylacetonate and BGN were completely dispersed in 10mL of 75% ethanol solution at a mass ratio of 10: 1. Secondly, the solution is treated by ultrasonic for 30 minutes to ensure that the molybdenum acetylacetonate and BGN are uniformly mixed. The solution was then transferred directly to a 25mL autoclave and incubated in an oven at 200 ℃ for 8 hours. And (3) cooling the high-pressure reaction kettle to room temperature, taking out the suspension in the polytetrafluoroethylene lining, centrifuging at a high speed to obtain a precipitate, and freeze-drying the precipitate to obtain the molybdenum-doped bioactive glass ceramic nano material (Mo-10 BGN).

The multifunctional bioactive glass ceramic nano material (Mo-BGN) for tissue regeneration after infection/tumor operation, which is prepared by the invention, has the characteristics of simple preparation process, excellent photo-thermal performance, strong oxidation resistance and good biocompatibility, can effectively kill tumor cells and various bacteria under the action of laser, and can inhibit inflammation and promote vascularization, so the nano material has good application prospect in tissue regeneration after infection/tumor operation.

The above-mentioned contents are only for illustrating the technical idea of the present invention, and the protection scope of the present invention is not limited thereby, and any modification made on the basis of the technical idea of the present invention falls within the protection scope of the claims of the present invention.