CN111450309A

CN111450309A - Anti-infection silicon dioxide biological tissue adhesive and application thereof

Info

Publication number: CN111450309A
Application number: CN201910045956.6A
Authority: CN
Inventors: 高亦鲲; 王思玲; 林伟康; 刘迎春
Original assignee: Shenyang Pharmaceutical University
Current assignee: Shenyang Pharmaceutical University
Priority date: 2019-01-18
Filing date: 2019-01-18
Publication date: 2020-07-28
Anticipated expiration: 2039-01-18
Also published as: CN111450309B

Abstract

The invention relates to a preparation method of a drug-loaded antibacterial nano mesoporous silica biological tissue adhesive and application of the adhesive in tissue adhesion. The preparation method of the drug-loaded mesoporous silica comprises the following steps: preparing oil-in-water emulsion by using n-octane, hexadecyl trimethyl ammonium bromide and water, polymerizing a styrene monomer and an ethyl orthosilicate monomer in oil phase droplets, and removing styrene through demulsification, centrifugation and high-temperature calcination to prepare mesoporous silica nanoparticles; adding the prepared silicon dioxide into lysozyme solution to load lysozyme, and centrifugally drying to prepare the anti-infection silicon dioxide biological tissue adhesive. The invention aims to solve the defects of secondary injury caused by surgical suture in wound treatment, the limitation that certain organs and tissues are not suitable for suture, poor effect of the traditional organic biological adhesive and the like. Has the advantages of rapid action, no secondary wound and the like, and has wide application prospect in the field of wound treatment.

Description

Anti-infection silicon dioxide biological tissue adhesive and application thereof

Technical Field

The invention belongs to the field of biomedicine, relates to a preparation method and application of a novel biological adhesive, and particularly relates to preparation and application of lysozyme-loaded anti-infection nano mesoporous silica.

Background

Chronic intractable wounds and injuries such as visceral surgery incisions place a heavy burden on the patient and the health care system. At present, surgical suture and polymer adhesive are generally used as treatment means for wounds caused by liver, lung, heart and other viscera operations. However, for soft tissues such as liver and lung, the surgical suture can cause certain damage to organs and leave incision gaps; polymeric binders have also limited their range of application due to their limited utility in a wet in vivo environment and the complexity of the conditions necessary to control the polymerization or crosslinking reaction in vivo. Research shows that under the condition of room temperature, one drop of silicon oxide nano particle solution can generate rapid and strong adhesion between two independent hydrogels through a 'nano bridging' effect, and the effect can still be verified on skin wound and visceral surgery wound models.

The lysozyme is also called muramidase, is an alkaline enzyme capable of hydrolyzing mucopolysaccharide in pathogenic bacteria, can be directly combined with viral protein with negative charge by destroying β -1,4 glycosidic bonds between N-acetylmuramic acid and N-acetylglucosamine in cell walls to decompose insoluble mucopolysaccharide of the cell walls into soluble glycopeptides so as to further cause the cell walls to break and release contents to dissolve bacteria, can be directly combined with DNA, RNA and deglycoprotein to inactivate viruses, has antibacterial, anti-inflammatory and antiviral effects, is a relatively stable protein, has relatively high heat resistance and relatively low production cost, and has a relatively wide antibacterial and antibacterial spectrum.

In conclusion, the research establishes a novel biological adhesion treatment system based on the mesoporous silicon dioxide nano material, and the double functions of the mesoporous silicon nano particles on the nano bridging effect of the wound tissues and the antibacterial activity of lysozyme loaded on the wound tissues are utilized to promote incision and tissue adhesion and anti-infection repair.

Disclosure of Invention

The invention aims to provide an anti-infection silicon dioxide biological tissue adhesive, which is lysozyme-loaded mesoporous silicon dioxide nanoparticles with antibacterial activity, and adopts the following preparation technical scheme:

(1) cetyl Trimethyl Ammonium Bromide (CTAB) is stirred and dissolved in water at the temperature of 20-60 ℃, then n-octane is dropped in, and the stirring is continued to prepare an oil-in-water heterogeneous system.

(2) Tetraethoxysilane (TEOS) was dropped into the above system, and lysine was added.

(3) And (3) dripping a styrene monomer into the system in the step (2), and adding azodiisobutyramidine hydrochloride (AIBA) under the protection of nitrogen.

(4) After the reaction was stirred, the reaction mixture was cooled to room temperature and allowed to stand overnight.

(5) The reaction solution was mixed with absolute ethanol and centrifuged.

(6) And calcining the precipitate in a muffle furnace and grinding to obtain the mesoporous silica nanoparticles.

(7) The prepared mesoporous silica nanoparticles are dissolved in phosphate buffer solution of lysozyme with pH of 6.8 to obtain lysozyme-loaded mesoporous silica nanoparticles.

Wherein the content of the first and second substances,

n-octane in step (1): the mass ratio of the hexadecyl trimethyl ammonium bromide aqueous solution is 0.2-0.4;

the temperature in the step (1) is 20-60 ℃;

the concentration of the hexadecyl trimethyl ammonium bromide aqueous solution in the step (1) is 1-5mg/m L.

The weight-volume ratio of lysine to tetraethoxysilane in the step (2) (g/m L) is 1:25-1:80, preferably 1:25-1: 50.

Cetyl trimethyl ammonium bromide in step (3): the mass ratio of styrene monomer is 1: 15.15-40.45.

The final concentration of azobisisobutyramidine hydrochloride added in step (3) was 0.4-1.5mg/m L.

The reaction time in the step (4) is 3-6 hours;

the volume ratio of the reaction liquid to the absolute ethyl alcohol in the step (5) is 1: 1; the centrifugal rotating speed is as follows: 5000-.

And (6) putting the precipitate into a muffle furnace, and calcining at 500-600 ℃ for 5-6 hours.

In the step (7), the concentration of the lysozyme is 1-25mg/m L, and the concentration of the mesoporous silica nanoparticles is 0.5-15mg/m L.

Specifically, the method comprises the following steps:

(1) adding Cetyl Trimethyl Ammonium Bromide (CTAB) serving as a surfactant into distilled water, stirring the mixture for 2 hours at the temperature of 20-60 ℃ (preferably 60 ℃), then dripping a certain amount of n-octane at the speed of 2 drops/second, wherein the mass ratio of the n-octane to a cetyl trimethyl ammonium bromide aqueous solution is 0.2-0.4 (preferably 0.4), continuously stirring the mixture for 2 hours at the temperature to form a water-CTAB-n-octane oil-in-water heterogeneous system, and the concentration of the cetyl trimethyl ammonium bromide aqueous solution is 2.13 × 10^-3g/mL。

(2) Tetraethoxysilane (TEOS) was dropped into the above system at a rate of 2 drops/sec, and lysine was added in a weight-to-volume ratio of lysine to tetraethoxysilane (g/m L): 1: 48.5.

(3) Dropping a styrene monomer into the system at a speed of 2 drops/second until the concentration of the styrene monomer in the system is 0.219-0.585 mg/m L (preferably 0.585mg/m L), adding azodiisobutyramidine hydrochloride AIBA, ethyl orthosilicate, lysine, hexadecyl trimethyl ammonium bromide and the styrene monomer under the condition of nitrogen protection, wherein the mass ratio of the azodiisobutyramidine hydrochloride AIBA, the ethyl orthosilicate to the lysine to the hexadecyl trimethyl ammonium bromide to the styrene monomer is 10: 0.22: 1: 15.15-40.45, preferably 10: 0.22: 1: 40.45, and the weight-volume ratio of the added azodiisobutymidine hydrochloride in volume is 0.84mg/m L

(4) Stirring for reaction for 3-6 hours (preferably 3 hours), cooling the reaction solution to room temperature, and standing overnight.

(5) And (3) mixing the reaction solution in the step (5) with absolute ethyl alcohol at a ratio of 1:1, centrifuging once at 12000rpm for 15 minutes, mixing the precipitate with 50m L absolute ethyl alcohol, ultrasonically dispersing for 20 minutes, centrifuging once again at 12000rpm for 15 minutes, and drying the precipitate at room temperature.

(6) And placing the precipitate dried at room temperature into a muffle furnace, calcining at 500-600 ℃ for 5-6 hours, and grinding to obtain the mesoporous silica nanoparticles.

(7) The mesoporous silica nanoparticles are dissolved in 5mg/m L lysozyme phosphate buffer solution with pH of 6.8, the concentration of the mesoporous silica nanoparticles is 2.5mg/m L, the mesoporous silica nanoparticles are stirred for 3 hours at the temperature of 4 ℃ and at the rpm of 200, and finally the mesoporous silica nanoparticles loaded with lysozyme are obtained by centrifugation at the rpm of 8000 for 3 minutes.

The mesoporous silica nanoparticles prepared by the method are monodisperse, the particle size is 97-550nm, the pore diameter is 5-11nm, and the specific surface area is 397-1059 m²Per g, preferably with a prescribed pore size of 10.49nm and a specific surface area of 610.69m²/g。

The drug loading rate of the mesoporous silica nanoparticle loaded with lysozyme protein prepared by the invention is 15-30%, the drug release rate reaches 20-50% within 5 hours, and meanwhile, the released drug still maintains higher activity. The maximum average anti-pulling force of the self-made PDMA bionic gel and the Wistar rat liver after adhesion respectively reaches 0.065 and 0.049N/cm²Mg, about 11 times and 8.5 times the average tensile resistance after adhesion of the commercially available non-porous silica nanoparticles TM-50. The bonding effect is obviously improved.

Drawings

FIG. 1 is a TEM micrograph of sample 3.

FIG. 2 is an SEM micrograph of sample 3.

Fig. 3 shows the results of

samples

2, 3, and 4 when the concentrations of lysozyme to the samples are 1:2, the drug loading temperature is 4 ℃, and the calculated loading curve is obtained within 3 hours under the condition of the rotation speed of 200 rpm.

Figure 4 is a graph of the percent drug release over 5 hours for

samples

2, 3, and 4 after loading in 25m L pH6.8 phosphate buffer in a water bath at 37 ℃.

Fig. 5 is a schematic diagram of an apparatus for testing the tensile resistance using a tensile tester after bonding two PDMA hydrogels using sample 3.

Figure 6 is the average anti-pull force after bonding PDMA hydrogel or Wistar rat liver using sample 2, sample 3, sample 4, respectively.

FIG. 7 is a schematic diagram of an apparatus for testing anti-pull force using a tensile testing machine after adhering two Wistar rat livers using sample 3.

FIG. 8 shows the activity of lysozyme released within five hours from three drug-loaded samples in 25m L pH6.8 phosphate buffer in a 37 ℃ water bath.

Detailed Description

EXAMPLE 1 preparation of mesoporous silica

Sample 1

Weighing 0.3g of CTAB into a 500m L three-necked bottle, adding 96m L of distilled water, protecting the interior of a reaction device with nitrogen, magnetically stirring for 1 hour at 70 ℃ to fully dissolve the CTAB, weighing 45m L of n-octane, dripping the CTAB solution system at the speed of 2 drops/second, continuously stirring for 20 minutes after dripping to form uniform emulsion, weighing 3.2m L of tetraethyl orthosilicate TEOS, dripping the TEOS into the solution at the speed of 2 drops/second, weighing 0.066g of lysine into the solution, then weighing 8.9m L of methyl methacrylate monomer, dripping the methyl methacrylate into the solution at the speed of 2 drops/second, weighing 113.4mg of azodiisobutyl hydrochloride, adding the obtained product into BA reaction liquid, magnetically stirring in a 70 ℃ water bath, continuously reacting for 4 hours, stopping heating after the reaction is finished, cooling the reaction liquid to room temperature, transferring the reaction liquid to a beaker, sealing the beaker, standing overnight, standing for 15 minutes, and preparing a precipitate which fails.

Sample 2

Weighing 0.3g of CTAB in a 500m L three-necked bottle, adding 93m L distilled water, magnetically stirring for 2 hours at 20 ℃ to fully dissolve the CTAB, weighing 42m L n-octane, dropping the CTAB solution system at a speed of 2 drops/second, continuously stirring for 2 hours after dropping to form a uniform emulsion, weighing 3.2m L tetraethyl orthosilicate TEOS at a speed of 2 drops/second to drop into the solution, weighing 0.066g of lysine to the solution, then weighing 8.9m L styrene monomer at a speed of 2 drops/second to drop into the solution, after dropping, making nitrogen protection in the reaction device, weighing 113.4mg of azodiisobutymidine hydrochloride AII to add into the reaction liquid, magnetically stirring for continuous reaction for 6 hours at 60 ℃, stopping heating after the reaction is finished, after the reaction liquid is cooled to room temperature, transferring the reaction liquid to a preservative film, sealing, next day, adding 1:1 night, fully stirring for continuous reaction for 6 hours at 60 ℃, placing the supernatant after the reaction liquid is cooled to room temperature, placing the precipitate, centrifuging for 15 minutes, adding 12000rpm, and drying the precipitate, and centrifuging the precipitate for 15 minutes, thus obtaining the nano-particle.

The specific surface area of the sample is 1058.73m by adopting a specific surface area and pore size distribution tester²The pore diameter per gram was 7.03 nm. The particle size of the sample was determined to be 550.3nm using a dynamic laser light scattering apparatus.

Sample 3

Weighing 0.3g of Cetyl Trimethyl Ammonium Bromide (CTAB) in a 500m L three-necked bottle, adding 93m L distilled water, magnetically stirring for 2 hours in a water bath at 60 ℃ to fully dissolve the CTAB, weighing 42m L n-octane, dripping into the CTAB solution system at the speed of 2 drops/second, continuously stirring for 2 hours after dripping, measuring 3.2m L tetraethyl orthosilicate (TEOS) to drip into the solution at the speed of 2 drops/second, weighing 0.066g of lysine to add into the solution, measuring 13.35m L styrene monomer to drip into the solution at the speed of 2 drops/second, stopping heating after the reaction is finished, cooling the reaction solution to room temperature, transferring the reaction solution to a beaker, sealing the beaker, adding 113.4mg of azodiisobutyl hydrochloride (AIBA) into the reaction solution, continuously reacting for 3 hours at 60 ℃, continuously stirring for 3 hours after stirring, continuously stirring for 3 hours at 60 ℃, after the reaction solution is cooled to room temperature, transferring the reaction solution to a beaker, precipitating by using a centrifugal precipitation method, adding 1205 rpm, centrifuging, precipitating by adding 1-year centrifugal precipitation, centrifuging for 15 minutes, adding 1205 rpm, and precipitating silica, and then adding the supernatant to obtain a supernatant, precipitating, centrifuging, precipitating, and sintering.

The specific surface area of the sample is 610.69m by adopting a specific surface area and pore size distribution tester²Per g, pore size 10.49 nm. The particle size of the sample was determined to be 170.4nm using a dynamic laser light scattering apparatus. The clear mesoporous channel structure can be seen by observing the sample by adopting a transmission electron microscope and a scanning electron microscope. (see attached FIG. 1 and FIG. 2)

Sample No. 4

Weighing 0.3g of CTAB in a 500m L three-necked bottle, adding 93m L distilled water, magnetically stirring for 2 hours at 20 ℃ to fully dissolve the CTAB, weighing 42m L n-octane, dropping the CTAB solution system at a speed of 2 drops/second, continuously stirring for 2 hours after dropping to form a uniform emulsion, weighing 3.2m L tetraethyl orthosilicate TEOS at a speed of 2 drops/second to drop into the solution, weighing 0.066g of lysine to the solution, then weighing 5.0m L styrene monomer at a speed of 2 drops/second to drop into the solution, after dropping, making nitrogen protection in the reaction device, weighing 113.4mg of azodiisobutymidine hydrochloride AII to add into the reaction liquid, magnetically stirring for continuous reaction for 6 hours at 60 ℃, stopping heating after the reaction is finished, after the reaction liquid is cooled to room temperature, transferring the reaction liquid to a preservative film, sealing, next day, fully stirring at a volume ratio of 1:1, continuously reacting for 6 hours at 60 ℃, placing the supernatant in a 60 ℃ water bath, centrifuging for 15 minutes, drying, and then placing the supernatant after the supernatant is dried by centrifugal stirring, centrifuging for 15 minutes at 1205 rpm, and sintering the supernatant after the supernatant of the supernatant, and sintering the supernatant of the precipitated silica is obtained by using 15 minutes.

The specific surface area of the sample is 397.53m by adopting a specific surface area and pore size distribution tester²The pore diameter per gram was 5.25 nm. Sample particle size determination using dynamic laser scatteringIt was 97.4 nm.

The differences in specific surface area, pore size and particle size of the different samples are mainly determined by the amount of styrene monomer and the reaction time after addition of styrene and AIBA when the molar ratio of hexaalkyltrimethylammonium bromide: the mass ratio of styrene monomer is 1: 15.15-40.45, when the reaction time is 3-6h, the aperture of the prepared sample can reach 5-11nm, the specific surface area can reach 397-1059nm, and the particle size can reach 97-550 nm.

EXAMPLE 2 preparation of lysozyme-loaded mesoporous silica nanoparticles

Weighing 40mg of lysozyme, placing the lysozyme in a 10m L penicillin bottle, adding a PBS solution with the pH value of 8m L of 6.8, magnetically stirring the lysozyme for 1 hour at the temperature of 4 ℃ at 200rpm to fully dissolve the lysozyme, then weighing 20mg of mesoporous silica nanoparticles prepared in the example 1, placing the mesoporous silica nanoparticles into the lysozyme solution, continuously magnetically stirring the mesoporous silica nanoparticles for 3 hours at the temperature of 4 ℃ at the speed of 200rpm, centrifuging the mesoporous silica nanoparticles for 3 minutes at 8000rpm after stirring, and removing supernatant to obtain the mesoporous silica nanoparticles loaded with lysozyme protein.

In the preparation process of the mesoporous silica nanoparticle loaded with lysozyme, samples are taken at 15 minutes, 30 minutes, 45 minutes, 1 hour, 1.5 hours, 2 hours and 3 hours respectively, the sampling mode is that 500 mu L stirring liquid is sucked and centrifuged for 3 minutes at 8000rpm, then 200 mu L supernatant is taken to be stored at 4 ℃ to be tested, Coomassie brilliant blue G-250 staining solution is prepared, 100mg Coomassie brilliant blue G-250 is weighed and placed in a 1000m L brown measuring flask, 95% ethanol of 50m L is added, ultrasonic dissolution is carried out, 85% phosphoric acid of 100m L is added, distilled water is used for fixing the volume to 1000m L, the mixture is mixed evenly and stored at 4 ℃ for later use.

The supernatant obtained at each time point of 7 mu L was precisely aspirated, diluted to 500 mu L with phosphate buffered saline PBS (pH6.8), mixed with the above Coomassie brilliant blue staining solution of 3m L, shaken well, and after 3 minutes of dark staining, the UV-visible absorbance (500 mu L pH6.8 phosphate buffered saline which was stained by the UV-visible absorptiometer in the same way was used to correct the zero point) was measured at 595nm, and the UV-visible absorbance was substituted into the previously drawn absorbance-lysozyme concentration standard curve to calculate the actual protein concentration, the mass of lysozyme not loaded in the solution system, and the mass of lysozyme loaded was reversely deduced.

After 3 hours of drug loading, the loading ratios of sample 2 and sample 3 to lysozyme reached 27.0% and 28.0%, respectively (the drug loading curves are shown in fig. 3).

Example 3 lysozyme-loaded mesoporous silica nanoparticle drug release assay

Centrifuging the residual solution of the lysozyme loaded in the example 2 for 3 hours at 8000rpm for 3 minutes, transferring the solid precipitate into a 50m L flat-bottomed flask, adding 25m L phosphate buffer solution with pH6.8, magnetically stirring in a water bath at 37 ℃, continuously releasing the drug for 24 hours, sampling at 15 minutes, 30 minutes, 1 hour, 1.5 hours, 2 hours, 3 hours, 4 hours and 5 hours respectively, sucking 1000 mu L stirring liquid, centrifuging at 8000rpm for 3 minutes, taking 500 mu L supernatant, storing the supernatant at 4 ℃ to be tested, adding 500 mu L PBS with pH6.8 into the residual liquid and the solid, blowing uniformly, and adding the mixture back into a drug release system.

The 500 mu L supernatant obtained at each time point was mixed with the Coomassie brilliant blue staining solution described in example 2, shaken well, and after 3 minutes of dark staining, the UV-visible absorbance was measured at 595nm (the UV-visible absorptiometry had used a 500 mu L pH6.8 phosphate buffer solution for zero calibration, which had been stained in the same way), and was substituted into the previously drawn absorbance-lysozyme concentration standard curve to calculate the actual protein concentration and the mass of released protein in the solution system. after 5 hours of drug release, the release ratios of sample 2 and sample 3 to the loaded lysozyme reached 48.4% and 51.1%, respectively (the drug release curve is shown in FIG. 4).

EXAMPLE 4 mesoporous silica nanoparticle bonded biomimetic hydrogel PDMA anti-pulling test

The preparation process of the poly N, N ' -dimethylacrylamide hydrogel PDMA comprises the steps of mixing 42.5m L distilled water with 6.8m L N, N ' -dimethylacrylamide monomer, adding 9.2mg of N, N ' -methylenebisacrylamide MBA and 164mg of potassium persulfate KPS under the magnetic stirring at 25 ℃ and the nitrogen protection, fully stirring for 5 minutes, measuring 90 mu L of tetramethylethylenediamine TEMED, adding the mixture into a reaction liquid system, rapidly stirring for 20 seconds, transferring the reaction liquid into a 10cm × 10cm plastic culture dish, and horizontally standing overnight under the nitrogen protection.

The next day, the PDMA hydrogel was diced at a length (× width × thickness) of 5cm × 1.5.5 cm × 0.5.5 cm and weighedSamples 2 and 3 of fungal enzyme at 2mg/cm²Uniformly spreading the mixture on the surface of the PDMA cut block, covering a rectangular surface (with the length of × width) of 0.7cm × 1.5.5 cm, overlapping and attaching another PDMA with the covered rectangular surface, pressing for 30 seconds, vertically fixing the PDMA bonded by the method on a tension tester (shown in figure 5), and testing the maximum anti-pulling force until slipping, wherein the actual anti-pulling force is obtained by adding the maximum anti-pulling force and the gravity of a single PDMA hydrogel.

The maximum tensile resistance after PDMA bonding of sample 2 and sample 3 reaches 0.050 and 0.065N/cm²Mg. (FIG. 6)

Example 5 mesoporous silica nanoparticle-bonded liver anti-drag test

Male Wistar rats weighing 300g were fasted for 12 hours, the right liver lobes were harvested after cervical dislocation, the width and thickness of the liver lobes were recorded after hemostasis using gauze, the cross-sectional area was estimated as half the thickness of the width ×, the liver was bisected by cutting longitudinally at the length 1/2 of the liver, and

samples

2 and 3 were bisected at 2mg/cm²Spreading on the cut surface of the liver. The two sections were superimposed and pressed for 1 minute. The adhered liver lobes were mounted vertically in a tensile tester (see fig. 7) and tested for maximum tensile resistance until breakage. And after the test is finished, weighing and recording the mass of the half liver lobe positioned below the liver lobe during the test, wherein the actual anti-pulling force is obtained by adding the maximum anti-pulling force and the gravity of the half liver lobe.

The maximum anti-pulling force of sample 2 and sample 3 carrying lysozyme after being adhered to the right side liver lobe of male Wistar rat reaches 0.026 and 0.049N/cm respectively²Mg. (FIG. 6)

EXAMPLE 6 measurement of antimicrobial Activity of lysozyme released from mesoporous silica nanoparticles

10.4g of sodium dihydrogen phosphate, 7.86g of disodium hydrogen phosphate and 0.37g of ethylenediaminetetraacetic acid were weighed out and dissolved in 1000m L of distilled water to obtain phosphate buffered saline PBS having a pH of 6.2.

Taking the supernatant with the drug release time of 1 hour, 2 hours, 3 hours, 4 hours and 5 hours respectively, and diluting the supernatant until the protein concentration is 50 mu g/m L to be tested.

Precisely weighing 2.5mg of lysozyme L ys, placing the lysozyme in a 50m L measuring flask, adding phosphate buffer solution to dissolve the lysozyme, fixing the volume, weighing 200mg of the wall-dissolving micrococcus, adding phosphate buffer solution (pH is 6.2) to about 3m L, grinding for about 3min, adding phosphate buffer solution to the total volume of 200m L, carrying out equilibrium heat preservation in a water bath at 25 ℃, precisely measuring 3m L in a 1cm colorimetric cell, measuring absorbance at the wavelength of 450nm, a, precisely measuring 0 second reading a, measuring 0.15m L of a sample solution which is subjected to equilibrium heat preservation in the water bath at 25 ℃, adding the sample solution into the colorimetric cell, rapidly mixing the sample solution and measuring the absorbance A by using a second meter, measuring the absorbance A at 60 seconds, simultaneously, precisely measuring 0.15m L of the phosphate buffer solution (pH 6.2), operating the same way, measuring a sampling value a 'of 0s reading a and a reading A' of 60s as a blank, calculating the titer by dividing the reduction value of 60 seconds, and correcting the enzyme activity by the following formula:

the lysozyme released by the two drug-loaded mesoporous silica nanoparticles still keeps higher bacteriostatic activity (figure 7).

Claims

1. An anti-infective mesoporous silica biological tissue adhesive, characterized in that: the preparation method comprises the following steps:

(1) stirring and dissolving cetyl trimethyl ammonium bromide in water at the temperature of 20-60 ℃, then dripping n-octane, and continuously stirring to prepare an oil-in-water heterogeneous system;

(2) dropping ethyl orthosilicate into the system, and adding lysine;

(3) dripping a styrene monomer into the system in the step (2), and adding azodiisobutymidine hydrochloride under the protection of nitrogen;

(4) after stirring and reacting, cooling the reaction solution to room temperature, and standing overnight;

(5) mixing the reaction solution with absolute ethyl alcohol, and performing centrifugal separation;

(6) calcining the precipitate in a muffle furnace and grinding to obtain mesoporous silica nanoparticles;

2. The anti-infective mesoporous silica biological tissue adhesive as claimed in claim 1, wherein the mass ratio of n-octane to the aqueous solution of cetyltrimethyl ammonium bromide is 0.2-0.4, the temperature is 20-60 ℃, and the concentration of the aqueous solution of cetyltrimethyl ammonium bromide is 1-5mg/m L.

3. The anti-infective mesoporous silica biological tissue adhesive according to claim 1, wherein the weight to volume ratio of lysine to tetraethoxysilane in step (2) is (g/m L): 1:25-1: 80.

4. The anti-infective mesoporous silica biological tissue adhesive according to claim 1, wherein the ratio of cetyl trimethylammonium bromide: the mass ratio of styrene monomer is 1: 15.15-40.45.

5. The anti-infective mesoporous silica biological tissue adhesive according to claim 1, wherein the final concentration of azodiisobutyramidine hydrochloride added in step (3) is 0.4-1.5mg/m L.

6. The anti-infective mesoporous silica biological tissue adhesive of claim 1, wherein: the reaction time in the step (4) is 3 to 6 hours.

7. The anti-infective mesoporous silica biological tissue adhesive of claim 1, wherein: the volume ratio of the reaction liquid to the absolute ethyl alcohol in the step (5) is 1: 1; the centrifugal rotating speed is as follows: 5000-.

8. The anti-infective mesoporous silica biological tissue adhesive according to claim 1, wherein the concentration of lysozyme in step (7) is 1-25mg/m L, and the concentration of mesoporous silica nanoparticles is 0.5-15mg/m L.

9. Use of the anti-infective mesoporous silica biological tissue adhesive of any one of claims 1 to 8 in the preparation of a biological adhesive for promoting incision, tissue adhesion, and anti-infective repair.