CN114209875A

CN114209875A - Bioactive glass nano composite particles with antibacterial effect and high-efficiency hemostatic membrane-like structure camouflage and preparation method thereof

Info

Publication number: CN114209875A
Application number: CN202111462909.5A
Authority: CN
Inventors: 卢婷利; 郑彩云
Original assignee: Northwestern Polytechnical University
Current assignee: Shaanxi Liyun Zhicai Medical Biotechnology Co.,Ltd.
Priority date: 2021-12-02
Filing date: 2021-12-02
Publication date: 2022-03-22
Anticipated expiration: 2041-12-02
Also published as: CN114209875B

Abstract

The invention relates to a high-efficiency hemostatic membrane-like structure camouflage bioactive glass nano composite particle with an antibacterial effect and a preparation method thereof. The method comprises the steps of carrying out layer-by-layer self-assembly on raw materials of aminated macroporous bioactive glass, deionized water, bovine serum albumin and chitosan according to a layer-by-layer self-assembly sequence and obtaining the bioactive glass nano composite particles with the pseudo-membrane structure camouflage through an electrostatic adsorption principle. The preparation process is simple, and no secondary product is generated. The porous nano-particles with the membrane-like structure camouflage can simultaneously activate the intrinsic and extrinsic coagulation ways by being wrapped by bovine serum albumin with the platelet generation promoting effect and chitosan with the erythrocyte aggregation and antibacterial effects, so that a hemostatic plug can be quickly formed on a wound surface, the wound can be plugged, the bleeding amount can be reduced, and the bleeding time can be shortened. Improves the coagulation effect and biocompatibility of the bioactive glass, endows the bioactive glass with antibacterial performance, and has great clinical application value.

Description

Bioactive glass nano composite particles with antibacterial effect and high-efficiency hemostatic membrane-like structure camouflage and preparation method thereof

Technical Field

The invention belongs to the field of biomedical materials, and relates to bioactive glass nano composite particles with an antibacterial effect and a high-efficiency hemostatic membrane-like structure camouflage function and a preparation method thereof.

Background

The powdery hemostatic particles have wide application prospect in incompressible wounds such as large-area open wounds, internal organ injuries, body penetrating injuries and the like. Bleeding is a serious accident that occurs in people's daily life or war. Statistically, about 30% of the deaths are due to excessive blood loss when the body is severely damaged. Mortality rates of over 85% result from uncontrolled bleeding. Massive bleeding from the human body may lead to complications or death, and effective hemostasis is an important means to prevent death from trauma.

Therefore, a high-efficiency hemostatic material without side effects suitable for irregular wounds of limbs, trunk, internal organs and the like is the focus of current research. In recent years, porous hemostatic particles have been the focus of research on hemostatic materials due to their large specific surface area, high porosity, excellent liquid absorption rate, and the like. The porous material has a molecular sieve-like porous structure, and achieves the purpose of rapid hemostasis by absorbing water and concentrating blood coagulation factors by utilizing the contact activation principle of platelets. The nano-scale porous particle-based hemostatic agent has potential application prospects in the treatment of penetrating injuries and irregular bleeding. Due to their small size, access to the bleeding channels and access to hidden bleeding sites is facilitated.

Simple hemostatic materials cannot exert a good hemostatic effect because of single hemostatic route. The development trend of the hemostatic material is to exert the synergistic hemostatic effect among different materials through compounding. The preparation method of the prior hemostatic powder performs hemostasis by a single way, and has poor hemostatic efficiency. Chinese patent 201510868844.2 discloses a starch composite hemostatic dressing of mesoporous silica microspheres, which performs hemostasis through strong liquid absorption of composite hemostatic auxiliary materials, and the hemostatic approach is simple. Chinese patent 201810146389.9 discloses a mesoporous silica composite microsphere with high-efficiency blood coagulation function, which is compounded with polysaccharide to improve the hemostatic property, but has no antibacterial property and general hemostatic effect. Chinese patent 201922184318.0 discloses a wound composite dressing with bioactive glass base, which has antibacterial function and no wound hemostasis effect by attaching enzyme disinfectant to bioactive glass layerAnd is limited in the type of applicable wound. It is known that porous bioactive glass containing silicate components similar to those of hemostatic materials such as zeolite, having an ultra-high specific surface area capable of rapidly absorbing blood, and containing Ca²⁺The (blood coagulation factors IV and IVV) participate in blood coagulation cascade reaction to promote the conversion of fibrinogen into fibrin, and are widely applied to the preparation of hemostatic materials. The bovine serum albumin has negative charges on the surface, and can promote the generation of platelet factors and accelerate the formation of a fibrin network. The chitosan has positive charges on the surface, has broad-spectrum antibacterial function, and can gather red blood cells to accelerate the formation of the hemostatic suppository. The common macroporous bioactive glass has a common hemostatic effect and a single hemostatic mechanism, and the membrane-like structure camouflage nano particles prepared by wrapping bovine serum albumin and chitosan by a layer-by-layer self-assembly method can cooperate with various blood coagulation mechanisms to realize rapid hemostasis and inhibit the bacterial growth of a wound surface to achieve the antibacterial and anti-inflammatory effects.

Therefore, the development of a nano-scale composite hemostatic particle which has an antibacterial effect, is biodegradable, has good biocompatibility, can efficiently and rapidly coagulate blood and has a small amount of bleeding, and is synergistic with multiple blood coagulation mechanisms is urgently needed in the field.

Disclosure of Invention

Technical problem to be solved

In order to avoid the defects of the prior art, the invention provides the bioactive glass nano composite particles with the antibacterial high-efficiency hemostatic membrane-like structure camouflage and the preparation method thereof, and provides the nano composite hemostatic particles with the membrane-like structure camouflage, which have the antibacterial function in cooperation with a plurality of blood coagulation mechanisms, are biodegradable, have good biocompatibility, can perform high-efficiency and rapid blood coagulation, reduce the amount of bleeding and shorten the bleeding time, and the preparation method and the application thereof.

Technical scheme

A high-efficiency hemostatic membrane-like structure camouflage bioactive glass nano composite particle with an antibacterial effect is characterized by comprising aminated macroporous bioactive glass, bovine serum albumin and chitosan, wherein the diameter of the composite particle is 300-800 nm; the weight ratio of the macroporous bioactive glass to the bovine serum albumin to the chitosan is 100:200-400: 100-200; the average pore diameter of the macroporous bioactive glass is 50-70 nm.

The diameter of the bioactive glass nano composite particles disguised by the film-like structure is preferably 400-700 nm.

The weight ratio of the aminated macroporous bioactive glass, the bovine serum albumin and the chitosan is preferably 100:300-400: 200.

The average specific surface area of the macroporous bioactive glass is 250-700m²g^-1。

The average pore diameter of the macroporous bioactive glass is 30-60 nm.

The average specific surface area of the macroporous bioactive glass is 400-700m²g^-1。

The chitosan has deacetylation degree of 95% or more and viscosity of 10-200.

In another preferred embodiment, the viscosity of the chitosan is 100-200.

The method for preparing the bioactive glass nano composite particles with the antibacterial high-efficiency hemostatic membrane-like structure camouflage is characterized by comprising the following steps:

step 1: mixing aminated macroporous bioactive glass, deionized water and bovine serum albumin according to the weight ratio of 100:100-200:300-400 to obtain a solution required by the first-step reaction of layer-by-layer self-assembly; the pH value of the mixture is 7-8

Step 2: carrying out rotary centrifugation on the reaction solution with 8000-;

and step 3: mixing the macroporous bioactive glass coated with the bovine serum albumin, the deionized water and the chitosan according to the weight ratio of 100: 100-;

and 4, step 4: and (3) centrifuging the solution obtained in the step (3), collecting precipitates, washing and drying the precipitates to obtain the bioactive glass nano composite particles which are coated with chitosan outside the bovine serum albumin and have the membrane-like structure camouflage.

In another preferred example, the granulation solution comprises aminated macroporous bioactive glass, water, bovine serum albumin and chitosan in a weight ratio of 100:100-200:100-300: 10-100.

The macroporous bioactive glass is prepared by adopting a sol-gel method, and the raw material sources are cetyl ammonium bromide, ethyl orthosilicate and calcium nitrate tetrahydrate, wherein: the relative mass ratio of Si to Ca is 13: 0.5-1; the solvent is ethanol, diethyl ether, ammonia water and water in a weight ratio of 20:40:4: 150; preparing bioactive glass wet gel at 16-40 deg.C and pH of solution of 7-13, hydrolyzing and gelling with silicon source; then the macroporous bioactive glass powder is obtained by centrifugal collection, drying and calcination.

In another preferred embodiment, the pH of the aqueous ammonia solution is preferably 10 to 12.

In another preferred embodiment, the silicon source, i.e. tetraethoxysilane, is added dropwise at a rate of 2 ml/min.

In another preferred embodiment, the preparation temperature of the macroporous bioactive glass is preferably 20-30 ℃.

In another preferred embodiment, in the preparation process of the macroporous bioactive glass, the silicon source is hydrolyzed and gelated to prepare the bioactive glass wet gel at 400-.

In another preferred embodiment, the preparation method of the macroporous bioactive glass further comprises the following steps: centrifuging the prepared wet gel, drying in vacuum at 30-60 ℃ for 12-24 hours, removing water in the prepared wet gel, fully grinding, and calcining in a muffle furnace to obtain the macroporous bioactive glass.

In another preferred embodiment, the rotational speed for centrifugal collection of the prepared wet gel is 8000-.

In another preferred example, the relative mass ratio of the macroporous bioactive glass Si to Ca is 13: 0.5-1.

The preparation method of the aminated macroporous bioactive glass comprises the following steps: dispersing macroporous bioactive glass powder in isopropanol, dropwise adding 3-aminopropyltriethoxysilane, stirring, cooling, refluxing, centrifuging, collecting, washing with deionized water, and drying to obtain aminated macroporous bioactive glass; the mass ratio of the macroporous bioactive glass to the 3-aminopropyltriethoxysilane is 1: 5-10.

In another preferred embodiment, the preparation temperature of the aminated macroporous bioactive glass is 70-90 deg.C, preferably 80 deg.C.

In another preferred embodiment, the preparation process of the aminated macroporous bioactive glass is performed at 200-.

In another preferred embodiment, the average surface charge of the aminated macroporous bioactive glass is 30-35 mV, and the average diameter is 500-600 nm.

In another preferred embodiment, the average surface charge of the aminated macroporous bioactive glass coated with bovine serum albumin is 9-16 mV, and the average diameter is 560-600 nm.

In another preferred embodiment, the bioactive glass nanocomposite particles camouflaged on a film-like structure have an average surface charge of 22-34 mV and an average diameter of 560-600 nm.

In another preferred example, the pH value of the buffer solution in the process of coating the bioactive glass nano composite particles with the membrane-like structure camouflage is 7-8.

In another preferred example, the buffer solution in the process of coating the bioactive glass nano composite particles with the membrane-like structure camouflage can also be 0.5 mol/ml sodium chloride aqueous solution.

In another preferred example, the method further comprises the step of washing the bioactive glass nano composite particles with deionized water, wherein the bioactive glass nano composite particles are obtained by the step (2) and are in a membrane-like structure camouflage mode.

In another preferred example, the bioactive glass nano composite particles with the membrane-like structure camouflage obtained in the step (3) are washed by deionized water and then dried in an oven, for example, at 20-30 ℃ for 6-12 hours.

In another preferred example, the bioactive glass nanocomposite particles camouflaged in a membrane-like structure obtained in step (3) are washed with deionized water and then lyophilized in a lyophilizer, such as lyophilization for 8 to 12 hours.

In step 1, 0.5 mol/ml aqueous sodium chloride solution was used as a buffer to adjust the pH.

The use method of the bioactive glass nano composite particles with the antibacterial high-efficiency hemostatic membrane-like structure camouflage is characterized by comprising the following steps: can be used as rapid hemostatic material, skin repairing material or tissue engineering material.

Advantageous effects

The invention provides a high-efficiency hemostatic membrane-like structure camouflage bioactive glass nano composite particle with an antibacterial effect and a preparation method thereof, and the membrane-like structure camouflage bioactive glass nano composite particle comprises bioactive glass, bovine serum albumin and chitosan, the diameter distribution is 400-700nm, and the pore size distribution is 50-70 nm. Raw materials of aminated macroporous bioactive glass, deionized water, bovine serum albumin (polyanion) and chitosan (polycation) in a weight ratio of 100:100-200:400:200 are sequentially dispersed in aqueous solutions of bovine serum albumin and chitosan according to a layer-by-layer self-assembly sequence, and the bioactive glass nano composite particles with the pseudo-membrane structure camouflage are obtained by an electrostatic adsorption principle. The preparation process is simple, and no secondary product is generated. The porous nano-particles with the membrane-like structure camouflage can simultaneously activate the intrinsic and extrinsic coagulation ways by being wrapped by bovine serum albumin with the platelet generation promoting effect and chitosan with the erythrocyte aggregation and antibacterial effects, so that a hemostatic plug can be quickly formed on a wound surface, the wound can be plugged, the bleeding amount can be reduced, and the bleeding time can be shortened. The membrane-like structure camouflage not only improves the coagulation effect and biocompatibility of the bioactive glass, but also endows the bioactive glass with antibacterial performance, can quickly stop bleeding, can be used in the fields of skin repair after wound closure and the like, and has great clinical application value.

The bioactive glass nano composite particles with the membrane-like structure camouflage have the functions of coordinating with various coagulation mechanisms, simultaneously having an antibacterial effect, being biodegradable, good in biocompatibility, capable of efficiently and quickly coagulating blood, reducing bleeding amount, shortening bleeding time and the like. Can be used in the fields of emergency hemostasis and skin repair, and has higher clinical application value.

Meanwhile, according to the preparation method of the bioactive glass nano composite particles with the pseudo-membrane structure camouflage, the preparation process is simplified according to the existing layer-by-layer self-assembly preparation principle, and the problem of single hemostatic route of the existing silicic acid hemostatic material is solved by wrapping bovine serum albumin with platelet factor activating and chitosan with erythrocyte aggregation and antibacterial effects on the surface of the aminated macroporous bioactive glass. In the aspect of hemostasis mechanism, bioactive glass nano composite particles disguised by a membrane-like structure are contacted with a wound surface, the chitosan surface at the outermost layer is provided with positive charges to quickly gather red blood cells, and the bovine serum albumin activates platelets along with the dissolution of chitosan; calcium ions are released in the process of in vivo degradation of the macroporous bioactive glass, and endogenous and exogenous coagulation pathways can be activated simultaneously; comprehensively, the bioactive glass nano composite particles with the similar membrane structure camouflage can quickly start a blood coagulation path, wherein chitosan wrapped at the outermost layer has an antibacterial function.

The invention has the advantages that:

(1) the macroporous bioactive glass composite particles with the camouflage membrane structure have the functions of quickly stopping bleeding by coordinating with endogenous and exogenous coagulation ways, being biodegradable, good in biocompatibility, capable of efficiently and quickly coagulating blood, reducing bleeding amount, shortening bleeding time and the like;

(2) in the process of promoting rapid blood coagulation, the blood platelet and the aggregated red blood cell can be obviously activated, and the blood coagulation cascade reaction can be amplified.

(3) Has remarkable antibacterial effect on escherichia coli and staphylococcus aureus while promoting hemostasis.

(4) The production process is simple, complex large-scale equipment is not needed, and no by-product is generated;

drawings

Fig. 1 is an SEM picture of macroporous bioactive glass, bioactive glass nano composite particles camouflaged with a membrane-like structure:

a: macroporous bioactive glass; b: bovine serum albumin membrane-like structure disguising macroporous bioactive glass; c: bovine serum albumin and chitosan membrane structures disguise macroporous bioactive glass.

Fig. 2 is a TEM image of macroporous bioactive glass, bioactive glass nano-composite particles camouflaged with membrane-like structure:

FIG. 3 shows the nitrogen adsorption and desorption and pore size distribution of macroporous bioactive glass.

FIG. 4 shows the charge and particle size variation of the bioactive glass nanocomposite particle disguised with macroporous bioactive glass and membrane-like structures.

FIG. 5 is a Fourier transform infrared absorption spectrum of a bioactive glass nanocomposite particle disguised with macroporous bioactive glass and a membrane-like structure.

Fig. 6 shows the in vitro coagulation time and coagulation effect of the macroporous bioactive glass and bioactive glass nano composite particles disguised in a membrane-like structure:

the external blood coagulation time and blood coagulation effect of the macroporous bioactive glass and the bioactive glass nano composite particles with membrane-like structures camouflage. Note: and indicates that P <0.05 and P <0.001 for the samples and blanks. The ^ A represents that the P of the sample is less than or equal to 0.001 compared with Yunnan white drug powder.

FIG. 7 is the in vitro coagulation results of macroporous bioactive glass and bioactive glass nanocomposite particles camouflaged with a membrane-like structure.

Fig. 8 is a hemolysis result of macroporous bioactive glass and membrane-like structure camouflaged bioactive glass nanocomposite particles:

SEM results (A) and activated platelet pictures (B) of aggregated erythrocytes and fibrin formation by macroporous bioactive glass and bioactive glass nanocomposite particles camouflaged with membrane-like structures.

Fig. 9 is a photograph and SEM picture of activated platelets, aggregated red blood cells and fibrin formation after contacting blood with macroporous bioactive glass and bioactive glass nanocomposite particles camouflaged with a membrane-like structure:

aptt (a) and pt (b) of macroporous bioactive glass and membrane-like structure camouflaged bioactive glass nanocomposite particles. Note that x, xand x indicate that P <0.01, P <0.001 and P <0.0001 compared to the blank group for the samples. ^ A represents that the P of the sample is less than 0.0001 compared with Yunnan Baiyao powder.

Fig. 10 is the APTT and PT results for macroporous bioactive glass and bioactive glass nanocomposite particles camouflaged with a membrane-like structure:

the hemostatic effect of the macroporous bioactive glass and the bioactive glass nano composite particles with membrane-like structure camouflage in a rat liver injury model. Note that x, and x represent P <0.05, P <0.01, P <0.001, and P <0.0001 compared to the blank. ^ and ^ represent that P <0.001 and P <0.0001 for the sample compared with Yunnan Baiyao powder.

Fig. 11 shows the antibacterial evaluation results of the bioactive glass nanocomposite particles camouflaged with macroporous bioactive glass and membrane-like structures:

the macroporous bioactive glass and the bioactive glass nano composite particles with the membrane-like structure camouflage have the bacteriostatic effect on staphylococcus aureus (left) and escherichia coli (right). Note: 1: negative (LB + bacterial solution); 2: 26.38 μ g/mL; 3: 32.14. mu.g/mL; 4: 38.6 μ g/mL; 5: positive (2. mu.g/mL cefuroxime)

Fig. 12 is a result of cytotoxicity evaluation of macroporous bioactive glass and bioactive glass nanocomposite particles camouflaged with a membrane-like structure.

Detailed Description

The invention will now be further described with reference to the following examples and drawings:

the inventor of the application researches extensively and deeply, and researches a novel high-efficiency hemostatic bioactive glass nano composite particle disguised by a membrane-like structure and a preparation method thereof by wrapping bovine serum albumin and a chitosan membrane-like structure on the surface of bioactive glass by utilizing a layer-by-layer self-assembly synthesis principle for the first time. The particles comprise macroporous bioactive glass, bovine serum albumin and chitosan, and the diameter of the bioactive glass nano composite particles disguised by the membrane-like structure prepared by the weight ratio of the aminated macroporous bioactive glass, water, bovine serum albumin and chitosan is 100:100-200:400:200 is 400-700 nm. The preparation method of the bioactive glass nano composite particles with the pseudo-membrane structure camouflage does not need complex large-scale equipment, and has simple and efficient process; after the sample preparation is finished, only vacuum drying or freeze drying is needed for hours; in addition, in the coating process of the membrane-like structure, the consumed time and the energy are less, the electrostatic adsorption effect can be realized only by simple stirring, the preparation period of the product is obviously shortened while the energy consumption is reduced, the utilization rate of equipment is improved, the time cost is reduced, and the membrane-like structure is friendly to the environment and does not generate toxic waste water and waste gas. On the basis of this, the present invention has been completed.

In the first aspect of the invention, the macroporous bioactive glass nano composite particles comprise macroporous bioactive glass, bovine serum albumin and chitosan, wherein the diameter of the macroporous bioactive glass nano composite particles is mesoporous silica and chitosan, and the diameter of the mesoporous silica and chitosan is 400-700 nm.

Preparation method

The invention discloses a novel macroporous bioactive glass nano composite particle with a hemostatic function and a pseudo-membrane structure camouflage function, and discloses a preparation process thereof.

The specific preparation process comprises the steps of preparation of macroporous bioactive glass and amination thereof and synthesis of composite particles with membrane-like structure camouflage.

Preparing macroporous bioactive glass: dissolving 1% of surfactant cetyl ammonium bromide and 3.5% of tetraethoxysilane in a mixed ammonia water solution of ethanol and ether (water: ethanol: ether: ammonia water: 150:20:40:4), controlling the pH value to be 10-12, controlling the temperature to be 20-30 ℃, stirring for 4 hours at 800-.

Preparing aminated macroporous bioactive glass: dispersing 50% of macroporous bioactive glass particles in isopropanol, dropwise adding 3-aminopropyltriethoxysilane at the speed of 1 ml/min, stirring at 500 revolutions per minute for 200-36 hours, cooling and refluxing for 24-36 hours, centrifuging at 12000 revolutions per minute for 5 minutes for collection, washing with deionized water, and vacuum drying at 30-60 ℃ for 12-24 hours to obtain the aminated macroporous bioactive glass.

Preparing the nano composite particles with the pseudo-membrane structure camouflage: according to the layer-by-layer self-assembly synthesis principle, 1.5-4 mg/ml of bovine serum albumin is dissolved in an aqueous solution containing aminated macroporous bioactive glass at the pH value of 7-8, wherein the weight ratio of the aminated macroporous bioactive glass, deionized water and the bovine serum albumin is 100:100-200: 300-400. Stirring for 30 minutes at 20-30 ℃ and 600 revolutions and centrifuging for 5 minutes at 8000-12000 revolutions, collecting the first-step composite particles with the pseudo-membrane structure, and vacuum drying for 4-8 hours at 20-30 ℃. 0.5-2 mg/ml of chitosan is dissolved in the aqueous solution containing the first-step composite particles with the membrane-like structure camouflage, the pH value is 7-8, and the weight ratio of the first-step composite particles with the membrane-like structure camouflage, the deionized water and the chitosan is 100: 200-. Stirring for 30 minutes at 20-30 ℃ and 600 revolutions and centrifuging for 5 minutes at 8000-12000 revolutions, collecting the final product nano composite particles with the membrane-like structure camouflage, and drying for 4-8 hours in vacuum at 20-30 ℃.

In another preferred example, the diameter of the bioactive glass nano composite particles disguised by the film-like structure is in the range of 400-700 nm.

In another preferred embodiment, the chitosan is selected from the group consisting of: the deacetylation degree is more than or equal to 95 percent, and the viscosity is 100-200.

In another preferred embodiment, the weight ratio of the aminated macroporous bioactive glass, the bovine serum albumin and the chitosan is 100:200-400:100-200, preferably 100:300-400: 200.

In another preferred embodiment, the preparation method of the bioactive glass nano composite particles with the membrane-like structure camouflage comprises the following steps:

(1) providing a solution required by the first step of layer-by-layer self-assembly reaction, wherein the solution comprises aminated macroporous bioactive glass, deionized water and bovine serum albumin with the weight ratio of 100:100-200:300-400, and the pH value is 7-8.

(2) Centrifuging the reaction solution in the step (1) at 10000rpm for 5 minutes, and collecting the precipitate for the second step of layer-by-layer self-assembly reaction.

(3) Providing a solution required by the layer-by-layer self-assembly second-step reaction, wherein the solution comprises the precipitate, deionized water and chitosan collected in the step (2) with the weight ratio of 100: 100-200.

In another preferred example, the composite particle solution with the camouflage-like membrane structure comprises aminated macroporous bioactive glass, water, bovine serum albumin and chitosan in a weight ratio of 100:100-200:100-300: 10-100.

In another preferred embodiment, the mass ratio of the macroporous bioactive glass to the 3-aminopropyltriethoxysilane is 1: 5-10.

Use of

The macroporous bioactive glass composite particles with the membrane-like structure camouflage have the functions of quickly stopping bleeding by cooperating with various blood coagulation mechanisms, simultaneously have an antibacterial effect, are biodegradable, have good biocompatibility, can efficiently and quickly coagulate blood, reduce the amount of bleeding, shorten the bleeding time and the like. Can be applied to the fields of emergency hemostasis and skin repair independently or in combination with other medicines, and has higher clinical application value.

The features mentioned above with reference to the invention, or the features mentioned with reference to the embodiments, can be combined arbitrarily. All the features disclosed in this specification may be combined in any combination, and each feature disclosed in this specification may be replaced by alternative features serving the same, equivalent or similar purpose. Thus, unless expressly stated otherwise, the features disclosed are merely generic examples of equivalent or similar features.

The invention will be further illustrated with reference to the following specific examples. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention. The experimental procedures, in which specific conditions are not noted in the following examples, are generally carried out according to conventional conditions or according to conditions recommended by the manufacturers.

Percentages and fractions are weight percentages and weight fractions unless otherwise indicated. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. In addition, any methods and materials similar or equivalent to those described herein can be used in the methods of the present invention. The preferred embodiments and materials described herein are intended to be exemplary only.

Example 1

This example relates to the synthesis of macroporous bioactive glass

210mL of a 9.5% ethanol/19% diethyl ether mixed aqueous solution, 2g of CTAB as a surfactant, and 4mL of aqueous ammonia dropwise to adjustAdjusting pH to 10-12, adding 7.5mL TEOS into syringe pump, and weighing 1.4g Ca (NO)₃)₂·4H₂Adding O into the solution, stirring for 4h at 20-30 ℃ at 1000 r/min, centrifuging for 5min at 10000 r/min after the reaction is finished, collecting the precipitate, washing for 3 times by using absolute ethyl alcohol and deionized water respectively, drying for 24-36h in vacuum, fully grinding, and roasting for 5h at 550 ℃ to obtain the macroporous bioactive glass. The diameter size distribution is 400-700nm, the pore size distribution is 50-70nm, and the specific surface area is 242.84m²g^-1。

The morphology and the macroporous structure of the prepared macroporous bioactive glass were observed using a scanning electron microscope ((SEM, JSM-7500F, JEOL, Japan) and a transmission electron microscope (TEM, Talos F200X, FEI, USA), and as a result, as shown in fig. 1A and 2A, it was confirmed that the prepared bioactive glass particles had a macroporous structure.

Example 2

This example relates to the synthesis of macroporous bioactive glass

210mL of a mixed aqueous solution of 9.5% ethanol and 19% ethylene glycol ethyl ether, 2g of CTAB as a surfactant, 4mL of ammonia water dropwise added to adjust the pH value to 10-12, 7.5mL of tetraethoxysilane TEOS (tetraethyl orthosilicate) added by a syringe pump, and 1.4g of Ca (NO) (NO: 1) weighed₃)₂·4H₂Adding O into the solution, stirring for 4h at 20-30 ℃ at 1000 r/min, centrifuging for 5min at 10000 r/min after the reaction is finished, collecting the precipitate, washing for 3 times by using absolute ethyl alcohol and deionized water respectively, drying for 24-36h in vacuum, fully grinding, and roasting for 5h at 550 ℃ to obtain the macroporous bioactive glass. The diameter size distribution is 300-400nm, the pore size distribution is 20-40nm, and the specific surface area is 698.36m²g^-1。

Example 3

This example relates to the synthesis of aminated macroporous bioactive glass

0.5g of the prepared macroporous bioactive glass is weighed and dispersed in 200mL of isopropanol, 5mL of APTES is dripped, and then the mixture is cooled and refluxed for 24h at 80 ℃ and 400 r/min. The product is collected after 10000-rotation centrifugation for 5 minutes, washed 3 times by deionized water and absolute ethyl alcohol respectively, and dried for 24 hours in vacuum at 50 ℃. The aminated macroporous bioactive glass is obtained, and the surface charge is 37.5 millivolts.

Example 4

This example relates to the synthesis of aminated macroporous bioactive glass

0.1g of the prepared macroporous bioactive glass is weighed and dispersed in 200mL of isopropanol, 0.5mL of APTES is dripped, and the mixture is cooled and refluxed for 36 hours at 80 ℃ at 400 r/min. The product is collected after 10000-rotation centrifugation for 5 minutes, washed 3 times by deionized water and absolute ethyl alcohol respectively, and dried for 24 hours in vacuum at 50 ℃. The aminated macroporous bioactive glass is obtained, and the surface charge is 29.8 millivolts.

Example 5

The embodiment relates to the synthesis of membrane-like structure camouflage macroporous bioactive glass composite particles

0.1g of aminated macroporous bioactive glass particles are weighed and dispersed in 200mL of bovine serum albumin aqueous solution with the final concentration of 4mg/mL, the temperature is 20-30 ℃, 300 r/min, and the stirring is carried out for 30 min. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times with deionized water. The washed precipitate was dispersed in 200mL of a chitosan aqueous solution having a final concentration of 2mg/mL at 20-30 ℃ at 300 rpm, and stirred for 30 minutes. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times each with deionized water. It is known that the surface charge of the aminated macroporous bioactive glass is 37.5 millivolts, the diameter size is 560nm, and the PDI is 0.147. After the first layer of bovine serum albumin membrane-like structure is wrapped, the surface charge of the obtained composite particles is 10.7 millivolts, the diameter is 568nm, and the PDI is 0.236; after the second layer of chitosan film structure is wrapped, the surface charge of the obtained composite particles is 30.8 millivolts, the diameter is 575nm, and the PDI is 0.258.

Example 6

0.1g of aminated macroporous bioactive glass particles are weighed and dispersed in 200mL of bovine serum albumin aqueous solution with the final concentration of 3mg/mL, the temperature is 20-30 ℃, 300 r/min, and the stirring is carried out for 30 min. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times with deionized water. The washed precipitate was dispersed in 200mL of a chitosan aqueous solution having a final concentration of 2mg/mL at 20-30 ℃ at 300 rpm, and stirred for 30 minutes. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times each with deionized water. It is known that the surface charge of the aminated macroporous bioactive glass is 37.5 millivolts, the diameter size is 560nm, and the PDI is 0.147. After the first layer of bovine serum albumin membrane-like structure is wrapped, the surface charge of the obtained composite particles is 11.2 millivolts, the diameter is 565nm, and the PDI is 0.262; after the second layer of chitosan-like membrane structure is wrapped, the surface charge of the obtained composite particles is 33.6 millivolts, the diameter size is 580nm, and the PDI is 0.298.

Example 7

0.1g of aminated macroporous bioactive glass particles are weighed and dispersed in 200mL of bovine serum albumin aqueous solution with the final concentration of 2mg/mL, the temperature is 20-30 ℃, 300 r/min, and the stirring is carried out for 30 min. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times with deionized water. The washed precipitate was dispersed in 200mL of a chitosan aqueous solution having a final concentration of 2mg/mL at 20-30 ℃ at 300 rpm, and stirred for 30 minutes. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times with deionized water. It is known that the surface charge of the aminated macroporous bioactive glass is 37.5 millivolts, the diameter size is 560nm, and the PDI is 0.147. After the first layer of bovine serum albumin membrane-like structure is wrapped, the surface charge of the obtained composite particles is 15.2 millivolts, the diameter is 563nm, and the PDI is 0.297; after the second layer of chitosan film structure is wrapped, the surface charge of the obtained composite particles is 32.1 millivolts, the diameter is 572nm, and the PDI is 0.301.

Example 8

0.1g of aminated macroporous bioactive glass particles are weighed and dispersed in 200mL of bovine serum albumin aqueous solution with the final concentration of 4mg/mL, the temperature is 20-30 ℃, 300 r/min, and the stirring is carried out for 30 min. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times with deionized water. The washed precipitate was dispersed in 200mL of a chitosan aqueous solution with a final concentration of 1mg/mL at 20-30 ℃ at 300 rpm, and stirred for 30 minutes. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times with deionized water. It is known that the surface charge of the aminated macroporous bioactive glass is 37.5 millivolts, the diameter size is 560nm, and the PDI is 0.147. After the first layer of bovine serum albumin membrane-like structure is wrapped, the surface charge of the obtained composite particles is 10.7 millivolts, the diameter is 568nm, and the PDI is 0.236; after the second layer of chitosan-like membrane structure is wrapped, the surface charge of the obtained composite particles is 26.5 millivolts, the diameter is 568nm, and the PDI is 0.235.

Example 9

0.1g of aminated macroporous bioactive glass particles are weighed and dispersed in 200mL of bovine serum albumin aqueous solution with the final concentration of 3mg/mL, the temperature is 20-30 ℃, 300 r/min, and the stirring is carried out for 30 min. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times with deionized water. The washed precipitate was dispersed in 200mL of a chitosan aqueous solution with a final concentration of 1mg/mL at 20-30 ℃ at 300 rpm, and stirred for 30 minutes. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times with deionized water. It is known that the surface charge of the aminated macroporous bioactive glass is 37.5 millivolts, the diameter size is 560nm, and the PDI is 0.147. After the first layer of bovine serum albumin membrane-like structure is wrapped, the surface charge of the obtained composite particles is 11.2 millivolts, the diameter is 565nm, and the PDI is 0.262; after the second layer of chitosan-like membrane structure is wrapped, the surface charge of the obtained composite particles is 22.5 millivolts, the diameter is 570nm, and the PDI is 0.231.

Example 10

0.1g of aminated macroporous bioactive glass particles are weighed and dispersed in 200mL of bovine serum albumin aqueous solution with the final concentration of 2mg/mL, the temperature is 20-30 ℃, 300 r/min, and the stirring is carried out for 30 min. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times with deionized water. The washed precipitate was dispersed in 200mL of a chitosan aqueous solution with a final concentration of 1mg/mL at 20-30 ℃ at 300 rpm, and stirred for 30 minutes. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times with deionized water. It is known that the surface charge of the aminated macroporous bioactive glass is 37.5 millivolts, the diameter size is 560nm, and the PDI is 0.147. After the first layer of bovine serum albumin membrane-like structure is wrapped, the surface charge of the obtained composite particles is 15.2 millivolts, the diameter is 563nm, and the PDI is 0.297; after the second layer of chitosan film structure is wrapped, the surface charge of the obtained composite particles is 25.2 millivolts, the diameter is 571nm, and the PDI is 0.312.

Example 11

0.1g of aminated macroporous bioactive glass particles are weighed and dispersed in 100mL of bovine serum albumin aqueous solution with the final concentration of 4mg/mL, the temperature is 20-30 ℃, 300 r/min, and the stirring is carried out for 30 min. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times with deionized water. The washed precipitate was dispersed in 200mL of a chitosan aqueous solution having a final concentration of 2mg/mL at 20-30 ℃ at 300 rpm, and stirred for 30 minutes. The precipitate was collected by centrifugation at 10000rpm for 5 minutes and washed 3 times with deionized water. It is known that the surface charge of the aminated macroporous bioactive glass is 37.5 millivolts, the diameter size is 560nm, and the PDI is 0.147. After the first layer of bovine serum albumin membrane-like structure is wrapped, the surface charge of the obtained composite particles is 9.5 millivolts, the diameter is 579nm, and the PDI is 0.196; after the second layer of chitosan film structure is wrapped, the surface charge of the obtained composite particles is 31.5 millivolts, the diameter is 593nm, and the PDI is 0.247.

Example 12

This example relates to the morphological and structural characterization of macroporous bioactive glass and membrane-like structure camouflaged macroporous bioactive glass nanocomposite particles

Taking the macroporous bioactive glass prepared in the example 1 as an example, the macroporous structure of the bioactive glass can be observed from a scanning electron microscope image and a transmission electron microscope image, and the diameter size distribution is 400-700 nm. The results are shown in FIG. 1A and FIG. 2A. Measurement with a Nitrogen isothermal adsorption-desorption Analyzer (Best Instrument Technology (Beijing) Co., LTD,3H-2000PS2)The macroporous bioactive glass prepared in example 1 has a specific surface area of 242.84m²g^-1The average pore size is about 11.41 nm. The nitrogen adsorption and desorption curves and the pore size distribution of the sample are shown in fig. 3.

Taking the pseudo-porous bioactive glass nano composite particle with the membrane-like structure prepared in example 5 as an example, the diameter of the pseudo-porous bioactive glass nano composite particle is 575nm, and the observation of a scanning electron microscope shows that the surface of the prepared porous particle is covered with bovine serum albumin (shown in figure 1B) and chitosan (shown in figure 1C), and the observation of the surface of the macroporous bioactive glass with the projection electron microscope shows that the pseudo-porous structure exists, and the results are shown in figures 2B and 2C.

The composite particles prepared in example 5 were measured to have a change in surface charge of 37.5, 10.7, 30.8 mv and a change in diameter of 560, 568, 575nm, respectively, with the envelope of the film-like structure, as measured by laser particle size and Zeta-potentiometer doppler electrophoresis (Zetasizer Nano ZS, Malvern Instruments ltd., England). The particle size and diameter size variation curves for the samples are shown in FIG. 4.

The composite particles prepared in example 5 were measured at 4000-400cm using a Fourier transform Infrared Spectroscopy (FTIR, Tensor27, Bruker Optics)^-1As shown in fig. 5, bovine serum albumin and chitosan can be observed to wrap the surface of the macroporous bioactive glass through electrostatic adsorption.

Example 13

This example relates to characterization of macroporous bioactive glass and membranous-like structure camouflaged macroporous bioactive glass nanocomposite particles with added accelerated coagulation performance

Taking the macroporous bioactive glass prepared in examples 1 and 5 and the macroporous bioactive glass nano composite particles disguised in a membrane-like structure as examples, 1mL of fresh SD rat anticoagulated whole blood is added into 10mg of the sample to be tested, the centrifuge tube is taken out at intervals of 10s at 37 ℃ and tilted once, whether the blood is coagulated or flows is observed, and when the blood is completely coagulated, namely, the blood flow does not appear until the centrifuge tube is tilted to 90 degrees, the time is stopped, and the blood coagulation time of each group is recorded. Only blood in a blank centrifuge tube is used as a negative control, Yunnan white drug powder is used as a positive control, and each sample is repeated for 3 times. The result shows that the in vitro blood coagulation time of the macroporous bioactive glass nano composite particles with the pseudo-membrane structure is about 55s at the shortest, and the process and the result are shown in figure 6.

Example 14

This example relates to the evaluation of the in vitro hemolytic properties of macroporous bioactive glass and membranous-like camouflaged macroporous bioactive glass nanocomposite particles

Taking the macroporous bioactive glass prepared in examples 1 and 5 and the macroporous bioactive glass nano composite particles disguised in a membrane-like structure as examples, the samples are dispersed in 3.5mL of PBS according to different concentration gradients (50, 100, 200 and 400 mu g/mL) and incubated at 37 ℃ for 30 min; 0.1mL of rat anticoagulated diluted blood (rat anticoagulated: PBS 4: 5) was added to the above PBS suspension, and the mixture was kept at 37 ℃ for 30min, followed by 10000rpm, 2 min. The absorbance of the supernatant at 540nm was measured. PBS solution and deionized water are respectively used as negative and positive control groups, and the hemolytic effect of the Yunnan white drug powder is considered as a control. The hemolysis rate was calculated according to the formula hemolysis rate (%) - (As-Ap)/(Ad-Ap) × 100%. As, Ap, and Ad represent the absorbance at 540nm after hemolysis experiments for the sample, PBS, and water, respectively. The results show that the hemolysis rate of each sample is below 5% of that specified for the hemostatic samples, and the results are shown in FIG. 7.

Example 15

This example relates to characterization of macroporous bioactive glass and membrane-like structure camouflaged macroporous bioactive glass nanocomposite particles to promote platelet activation, red blood cell aggregation, and fibrin formation during in vitro activated coagulation.

Characterization of red blood cell aggregation and fibrin formation, using macroporous bioactive glass and membrane-like structure disguised macroporous bioactive glass nanocomposite particles prepared in examples 1 and 5 as examples: after the blood was coagulated, 10mg of the sample was added to 1mL of SD rat anticoagulated whole blood, and the sample was washed 3 times with PBS to remove excess red blood cells. Fixing in 2.5% glutaraldehyde at 4 deg.C for 2.5h, sequentially dehydrating with gradient ethanol for three times, each for 10min, and observing in vitro erythrocyte adhesion condition with scanning electron microscope. The results show that the membrane-like structure is disguised of macroporous bioactive glass particles, and the results are shown in FIG. 8A. Platelet activation, SD rats anticoagulated whole blood was centrifuged at 1500 rpm for 15min and Platelet Rich Plasma (PRP) was aspirated with a pipette. Approximately 200. mu.L of PRP was added to 10mg of sample and incubated at 37 ℃ for 1 h. The sample is dehydrated and washed for three times by a series of gradient ethanol to eliminate the adhesion of the blood platelets on the surface of the material, each concentration is soaked for 10min, the sample is soaked in 2.5% glutaraldehyde fixing solution overnight at 4 ℃, and after drying, observation is carried out under a common optical microscope, and the result shows that the macroporous bioactive glass nano composite particles with the pseudo-membrane structure can activate the blood platelets to the maximum extent under the same condition, as shown in figure 8B.

Example 16

This example relates to characterization of macroporous bioactive glass and membrane-like structure camouflaged macroporous bioactive glass particles to promote platelet activation, red blood cell aggregation, and fibrin formation during in vitro activated coagulation.

Using the macroporous bioactive glass prepared in examples 1 and 5 and the macroporous bioactive glass nanocomposite particles disguised in a membrane-like structure as examples, 10mg of a sample was added to 1mL of SD rat anticoagulated whole blood. After blood coagulation, excess red blood cells were removed by washing 3 times with PBS. Fixing in 2.5% glutaraldehyde at 4 deg.C for 2.5h, sequentially dewatering and washing with gradient ethanol for three times, soaking for 15min at each concentration, drying, and observing in vitro erythrocyte adhesion set and fibrin network formation under scanning electron microscope, as shown in FIG. 8A. SD rats are anticoagulated with whole blood for 1500 rpm, centrifuged for 15min, and the upper layer Platelet Rich Plasma (PRP) is aspirated with a pipette. Approximately 200. mu.L of PRP was added to 10mg of sample and incubated at 37 ℃ for 1 h. The samples were washed three times with gradient ethanol dehydration, soaking for 15min at each concentration. The platelet activation status was recorded by soaking in 2.5% glutaraldehyde fixing solution at 4 ℃ overnight, drying, observation under a normal optical microscope and photographing, and the results are shown in FIG. 8B. The result shows that the macroporous bioactive glass nano composite particles with the pseudo-membrane structure can activate platelets to the maximum extent, form a fibrin network and aggregate red blood cells to form the hemostatic plug under the same condition.

Example 17

This example relates to coagulation pathway testing involving macroporous bioactive glass and membrane-like structure camouflaged macroporous bioactive glass nanocomposite particles.

Taking the macroporous bioactive glass prepared in examples 1 and 5 and the macroporous bioactive glass nano composite particles disguised in a membrane-like structure as examples, APTT: activated partial thromboplastin time, citrate anticoagulated whole blood of SD rats, 3000 rpm, centrifugation for 15min, preparation of Platelet Poor Plasma (PPP). Adding 50 μ L of APTT reagent into 50 μ L of PPP, incubating at 37 deg.C for 5min, adding 100 μ L of 0.025mol/L CaCl into the test tube₂And a sample to be tested (2mg), APTT was determined. PT: taking 50 mu L of PPP, 100 mu L of PT reagent and 2mg of sample, respectively incubating for 5min at 37 ℃, adding the sample to be detected and the PT reagent into plasma in a test tube, and determining PT; in the APTT and PT determination process, a sample is not added as a control group, Yunnan white drug powder hemostatic powder is used as a positive control, and the test result is expressed as the time percentage of the sample group and the control group. The results are shown in fig. 9A and 9B, and indicate that the macroporous bioactive glass nanocomposite particles with a camouflaged membrane-like structure can participate in activating intrinsic and extrinsic coagulation pathways simultaneously.

Example 18

This example relates to the evaluation of hemostatic effect of macroporous bioactive glass and membrane-like structure camouflaged macroporous bioactive glass nanocomposite particles in the liver bleeding model of SD rats.

Using the macroporous bioactive glass prepared in examples 1 and 5 and the macroporous bioactive glass particles camouflaged in a membrane-like structure as examples, SD rats were anesthetized with 10% chloral hydrate. The hair of the liver area was removed and the surgical area was disinfected with iodine and medical alcohol. The liver was debranned to expose the liver, a liver injury model was prepared, and a liver wound was incised 1cm with a scalpel. Immediately after blood absorption with conventional sterile gauze, hemostatic samples were applied to the wound site and gently lifted to observe the injury every 10 s. When bleeding stopped, the time to hemostasis and the amount of blood lost were determined. The Yunnan white drug powder as positive control, and the blank control of stopping bleeding without sample extraction, with hemostatic effect as shown in FIG. 10. The result shows that the macroporous bioactive glass nano composite particles with the pseudo-membrane structure can stop bleeding most quickly and reduce the blood loss.

Example 19

This example relates to the evaluation of the bacteriostatic effect of macroporous bioactive glass and membrane-like structure camouflaged macroporous bioactive glass nanocomposite particles on Staphylococcus aureus (s. aureus) and Escherichia coli (e.coli).

Using the membrane-like structure disguised macroporous bioactive glass particles prepared in example 5 as an example, s.aureus (ATCC 25923) and e.coil (ATCC 25922) were cultured to OD values of about 0.6-0.8, diluted to 5 x 10⁵CFU·mL^-1And (5) standby. 50. mu.L of diluted bacterial solution (5X 10) was added to each well of a 96-well plate⁵CFU·mL^-1) And respectively adding 50 mu L of nano particle suspension (sterilized) of samples to be detected with different concentrations (160, 133.32, 111.2, 138.6, 77.2, 64.28, 52.76 and 46.4 mu g/mL) as experimental groups, adding an antibacterial drug cefamycin as a positive control group, LB and bacterial liquid as a negative control group, using non-inoculated bacteria as a blank control group, and repeating 3 groups of groups to obtain the Minimum Inhibitory Concentration (MIC) of the bioactive glass nano composite particles with the similar membrane-like membrane structure camouflage. The results are shown in FIG. 11, where the MICs for S.aureus and E.coil were about 40 and 30 μ g/mL, respectively.

Example 19

This example relates to the evaluation of cellular compatibility of macroporous bioactive glass and membranous-like structure camouflaged macroporous bioactive glass nanocomposite particles with SD rat fibroblast L929.

Taking the macroporous bioactive glass prepared in the examples 1 and 5 and the macroporous bioactive glass nano composite particles with membrane-like structures as examples, the SRB staining method and the Calcein-AM/PI living/dead cell double staining kit are used for detecting the cell compatibility of the hemostatic particles. Mouse fibroblasts (L929) were seeded at a density of 2000/well in 96-well plates. Samples were diluted with DMEM complete medium to 1000, 500, 250, 125. mu.g/mL. The sample diluent and the cells were cultured for 24, 48 and 72 hours, and the DMEM complete culture solution group was used as a control group. Cells were stained with 0.4% SRB in 1% acetic acid. The absorbance of each well was measured at 540nm on a microplate reader and the cell viability of the samples was calculated as shown in FIG. 12A.

L929 was inoculated into 96-well plates at a density of 2000 cells/well, and the concentration of the sample group was measured by incubating the cells in DMEM complete medium at 1000. mu.g/mL for 24, 48, 72 hours, while the DMEM complete medium group was used as a control group. The dual staining labeling of live and dead cells was performed simultaneously using two dyes, Calcein-AM and PI. Analysis of the level of live and dead cells was performed using an inverted fluorescence microscope and the results are shown in FIG. 12B. The results show that the macroporous bioactive glass and the macroporous bioactive glass nano composite particles with the pseudo-membrane structure are co-cultured with the L929, the cell compatibility is good, and the cytotoxicity is avoided.

Claims

1. A high-efficiency hemostatic membrane-like structure camouflage bioactive glass nano composite particle with an antibacterial effect is characterized by comprising aminated macroporous bioactive glass, bovine serum albumin and chitosan, wherein the diameter of the composite particle is 300-800 nm; the weight ratio of the macroporous bioactive glass to the bovine serum albumin to the chitosan is 100:200-400: 100-200; the average pore diameter of the macroporous bioactive glass is 50-70 nm.

2. The bioactive glass nanocomposite particle with high-potency hemostatic membranous structure camouflage with antibacterial effect of claim 1, wherein: the average specific surface area of the macroporous bioactive glass is 250-700m²g^-1。

3. The bioactive glass nanocomposite particle with high-potency hemostatic membranous structure camouflage with antibacterial effect of claim 1, wherein: the average pore diameter of the macroporous bioactive glass is 30-60 nm.

4. The bioactive glass nanocomposite particle with high-potency hemostatic membranous structure camouflage with antibacterial effect of claim 2, wherein: the average specific surface area of the macroporous bioactive glass is 400-700m²g^-1。

5. The bioactive glass nanocomposite particle with high-potency hemostatic membranous structure camouflage with antibacterial effect of claim 1, wherein: the chitosan has deacetylation degree of 95% or more and viscosity of 10-200.

6. A method for preparing the highly effective hemostatic membrane-like structure camouflaged bioactive glass nanocomposite particles with antibacterial effect according to any one of claims 1 to 5, which is characterized by comprising the following steps:

Step 2: carrying out rotary centrifugation on the reaction solution with 8000-;

7. The method of claim 6, wherein: the macroporous bioactive glass is prepared by adopting a sol-gel method, and the raw material sources are cetyl ammonium bromide, ethyl orthosilicate and calcium nitrate tetrahydrate, wherein: the relative mass ratio of Si to Ca is 13: 0.5-1; the solvent is ethanol, diethyl ether, ammonia water and water in a weight ratio of 20:40:4: 150; preparing bioactive glass wet gel at 16-40 deg.C and pH of solution of 7-13, hydrolyzing and gelling with silicon source; then the macroporous bioactive glass powder is obtained by centrifugal collection, drying and calcination.

8. The method of claim 6, wherein: the preparation method of the aminated macroporous bioactive glass comprises the following steps: dispersing macroporous bioactive glass powder in isopropanol, dropwise adding 3-aminopropyltriethoxysilane, stirring, cooling, refluxing, centrifuging, collecting, washing with deionized water, and drying to obtain aminated macroporous bioactive glass; the mass ratio of the macroporous bioactive glass to the 3-aminopropyltriethoxysilane is 1: 5-10.

9. The method of claim 6, wherein: in step 1, 0.5 mol/ml aqueous sodium chloride solution was used as a buffer to adjust the pH.

10. A use method of the high-efficiency hemostatic membrane-like structure camouflage bioactive glass nano composite particles with the antibacterial effect according to any one of claims 1 to 5 is characterized in that: can be used as rapid hemostatic material, skin repairing material or tissue engineering material.