CN113694243B

CN113694243B - Photosensitizer/clay composite material and preparation method and application thereof

Info

Publication number: CN113694243B
Application number: CN202110960883.0A
Authority: CN
Inventors: 高玉婷; 张家鑫; 严春杰; 潘玉凤
Original assignee: China University of Geosciences
Current assignee: China University of Geosciences
Priority date: 2021-08-20
Filing date: 2021-08-20
Publication date: 2022-05-13
Anticipated expiration: 2041-08-20
Also published as: CN113694243A

Abstract

The invention provides a photosensitizer/clay composite material and a preparation method and application thereof. The composite material is prepared by intercalating a photosensitizer in clay, namely intercalating positively charged aggregation-induced emission micromolecule organic matters into the interlayer of the clay. The composite material has the advantages of photodynamic and chemical power synergistic antibacterial, can rapidly generate hydroxyl free radicals and singlet oxygen under illumination, rapidly kill bacteria, has small damage to normal cells, and has better biocompatibility; the preparation method provided by the invention has the characteristics of simple process, convenience in operation, low cost and the like. The composite material also has the effects of antibiosis and hemostasis, has good biological safety and has huge application prospect in the field of medical dressings.

Description

Photosensitizer/clay composite material and preparation method and application thereof

Technical Field

The invention relates to the field of inorganic/organic composite sterilization materials, in particular to a photosensitizer/clay composite material and a preparation method and application thereof.

Background

With the improvement of the living conditions of people, the development of medical health and health career is also receiving wide attention. Bacterial infection is not only a problem requiring attention in the early stages of wound emergency, but also a challenge in the later stages of wound healing. Bacterial infections can cause severe tissue damage because microorganisms can compete with the host's immune system, and thus invade living tissue, especially staphylococcus aureus, not only slowing the wound healing process, but even causing tissue necrosis. Antibiotics are widely applied to the treatment of bacterial infection and other problems, but the long-term administration of antibiotics not only accelerates the evolution and formation of antibiotic-resistant bacteria, but also threatens the health of people through various ways such as food, water, livestock and the like. Therefore, an antibacterial material with high efficiency and low toxic and side effects is urgently needed to be researched to solve the problem of bacterial infection.

Photodynamic therapy (PDT) is an emerging method for treating bacterial infections because of its rapid, broad-spectrum, non-invasive nature. The sterilization mechanism of the photodynamic therapy is based on the super strong oxidation reaction of active oxygen molecules such as hydroxyl free radicals, superoxide anions, singlet oxygen, hydrogen peroxide and the like on bacteria. The active oxygen molecules effectively kill bacteria by destroying the integrity of the cell membrane structure, increasing the ion permeability of the cell membrane or by directly destroying unsaturated lipids, polypeptides, enzymes and other components in the cell. However, the generation of active oxygen is closely related to the optical properties of the photosensitizer and the energy of light, and the lifetime and release distance of singlet oxygen molecules generated by the photosensitizer are short, resulting in great limitations. In addition, the photosensitizer has certain cytotoxicity, and the photosensitizer needs to be modified to reduce the toxicity. Clay is a water-containing aluminosilicate product, is generated from feldspar rock in the earth crust through long-term weathering and geological effects, is widely divided, various and abundant in reserves in nature, and is a precious natural resource. Layered aluminosilicate minerals are one of the clay minerals and have a long history of use as pharmaceuticals. The layered aluminosilicate mineral has excellent adsorbability, cation exchange property, dispersion suspension property and the like. Since the layered aluminosilicate mineral has better cation exchange property, Fe with Fenton reaction can be used²⁺Or Fe³⁺The interlayer can generate a large amount of OH under the action of hydrogen peroxide, and the interlayer can also generate a small-molecule photosensitizer under the action of lightA large number of¹O₂OH and¹O₂has strong bactericidal action, and the intercalated composite material has the effect of synergistic sterilization of chemical power and photodynamic.

Therefore, it is very significant to develop an inorganic/organic composite sterilization material with photodynamic and chemodynamic synergistic sterilization effects.

Disclosure of Invention

The invention aims to provide a photosensitizer/clay composite material and a preparation method and application thereof aiming at the defects in the prior art.

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

the invention provides a photosensitizer/clay composite material, which comprises a photosensitizer and layered clay, wherein the photosensitizer is intercalated in the clay.

Further, the photosensitizer is a positively charged aggregation-induced emission micromolecule organic matter.

Further, the positively charged aggregation-induced emission small molecule organic matter comprises one or more of BPCI, TPCI, and TPCB.

Further, the clay includes one or more of kaolinite, montmorillonite, illite, and chlorite.

The invention also provides a preparation method of the photosensitizer/clay composite material, which comprises the steps of mixing clay and buffer solution to prepare uniform slurry, and dripping the photosensitizer into the slurry for light-shielding reaction to obtain the photosensitizer/clay composite material.

Further, the preparation method comprises the following steps:

step S1, adding clay with certain mass into buffer solution to prepare suspension with w/v of 0.2-0.5%;

step S2, carrying out ultrasonic crushing on the suspension liquid obtained in the step S1 to obtain uniform slurry;

step S3, dropwise adding 0.015-0.6 mg/mL of photosensitizer into the slurry obtained in the step S2, stirring, and reacting for 6-48 h in a dark place at the temperature of 20-60 ℃;

and step S4, carrying out freeze drying on the solution reacted in the step S3 to obtain the photosensitizer/clay composite material.

Further, the buffer solution in step S1 is a phosphoric acid buffer solution, and the pH value of the phosphoric acid buffer solution is 6.5 to 7.4.

Further, the photosensitizer in the step S3 is TPCI, and the concentration of TPCI is 0.15 mg/mL-6.0 mg/mL.

Further, the clay is montmorillonite, and the mass concentration ratio of the montmorillonite to the TPCI is in the range of: 13.0 to 6.5

The invention also provides the application of the photosensitizer/clay composite material in the antibacterial hemostatic dressing.

Further, the applied conditions comprise illumination conditions and hydrogen peroxide conditions, wherein the illumination conditions comprise illumination wavelength of 400-600 nm and illumination light source power of 1mW/cm²～50mW/cm²(ii) a The hydrogen peroxide concentration in the hydrogen peroxide condition is 10-200 mu mol/L.

The technical scheme provided by the invention has the beneficial effects that:

(1) the invention provides a photosensitizer/clay composite material, which is prepared by intercalating positively charged aggregation-induced emission micromolecule organic matters into the interlayer of clay. The composite material has the advantages of photodynamic and chemical power synergistic antibiosis, can rapidly generate hydroxyl free radicals and singlet oxygen under illumination to rapidly kill bacteria, has small damage to normal cells and better biocompatibility compared with a single photosensitizer or clay, and also shows better capability of promoting wound healing in animal experiments;

(2) the preparation method of the photosensitizer/clay composite material has the characteristics of simple process, convenient operation, low cost and the like;

(3) the photosensitizer/clay composite material has the effects of resisting bacteria and stopping bleeding, has good biological safety and has huge application prospect in the field of medical dressings.

Drawings

FIG. 1 is an XRD overlay of MMT and TPCI/MMT in example 1;

FIG. 2 is a FTIR overlay of MMT, TPCI and TPCI/MMT in example 1;

FIG. 3a is an SEM photograph of the MMT of example 1;

FIG. 3b is an SEM photograph of TPCI/MMT of example 1;

FIG. 4a is a BET plot of MMT in example 1;

FIG. 4b is a BET plot of TPCI/MMT in example 1;

FIG. 5a is a plot of the OH production rate of TPCI/MMT in example 1;

FIG. 5b is a graph of ESR detection. OH yield of TPCI/MMT in example 1;

FIG. 6a shows TPCI/MMT in example 1¹O₂Generating a rate map;

FIG. 6b is ESR detection of TPCI/MMT in example 1¹O₂Generating a quantity map;

FIG. 7 is a graph comparing the phototoxicity of TPCI/MMT, TPCI and MMT in example 1;

fig. 8a is a graph comparing the anti-e.coli effects of TPCI/MMT and MMT in example 1;

FIG. 8b is a graph comparing the anti-S.a μ re μ s effects of TPCI/MMT and MMT in example 1

FIG. 9 is a comparison of hemolysis experiments for TPCI/MMT, TPCI and MMT in example 6;

FIG. 10 plot of the antimicrobial efficiency of TPCI/MMT composite of example 1;

FIG. 11 plot of the antimicrobial efficacy of TPCI/MMT composites of example 2;

FIG. 12 plot of the antimicrobial efficacy of TPCI/MMT composites of example 3;

FIG. 13 plot of the antimicrobial efficiency of TPCI/MMT composite of example 4;

FIG. 14a is a photograph of TPCI/MMT and MMT in example 1 taken at different times during the course of wound healing in mice in an animal experiment;

FIG. 14b is an assessment of wound area at different times during wound healing in the mice in the animal experiments with TPCI/MMT and MMT of example 1.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, embodiments of the present invention will be further described with reference to the accompanying drawings and examples.

All chemicals used in the study were analytical grade reagents and the bacteria used in the experiment were E.coli and S.aureus. The photosensitizer TPCI is self-made and synthesized by the applicant, and the structural formula of the photosensitizer TPCI is shown as the formula (I):

example 1:

8mL of PBS (pH 6.5) was weighed into a centrifuge tube, 16Mg of Montmorillonite (MMT) was weighed into the centrifuge tube, and disrupted in an ultrasonic cell disruptor for 60min to obtain a uniform slurry, at which time the concentration (w/v) of MMT was 0.2%. 15.42mg of TPCI was weighed into a centrifuge tube, and 1mL of dimethyl sulfoxide (DMSO) was added thereto, and dissolved by sonication, at which time the concentration of TPCI was 10 mmol/L. 100. mu.L of 10mmol/L TPCI was measured and diluted into 1mL of an aqueous solution, at which time the concentration of TPCI was 1 mmol/L. Weighing 2.7mL and 2mg/mL MMT solution in a centrifuge tube, slowly dripping 1mmol/L TPCI solution with the volume ratio of MMT to TPCI being 9:1 under the stirring at the temperature of 20 ℃, and reacting for 6 hours in a dark place to obtain TPCI/MMT composite material solution, namely the photosensitizer/clay composite material 1.

Example 2:

4mL of PBS (pH 6.5) was weighed into a centrifuge tube, 8Mg of Montmorillonite (MMT) was weighed into the centrifuge tube, and disrupted in an ultrasonic cell disruptor for 15min to obtain a uniform slurry, at which time the concentration (w/v) of MMT was 0.2%. 15.42mg of TPCI was weighed into a centrifuge tube, and 1mL of dimethyl sulfoxide (DMSO) was added thereto, and dissolved by sonication, at which time the concentration of TPCI was 10 mmol/L. 100. mu.L of 10mmol/L TPCI was measured and diluted into 1mL of an aqueous solution, at which time the concentration of TPCI was 1 mmol/L. Weighing 2.1mL and 2mg/mL MMT solution in a centrifuge tube, slowly dropwise adding 1mmol/L TPCI solution with the volume ratio of MMT to TPCI being 9:1 under stirring at 30 ℃, and reacting for 12 hours in a dark place to obtain TPCI/MMT composite material solution, namely the photosensitizer/clay composite material 2.

Example 3:

10mL of PBS (pH 6.5) is weighed into a centrifuge tube, 20Mg of Montmorillonite (MMT) is weighed into the centrifuge tube, and the mixture is crushed for 120min in an ultrasonic cell crusher to obtain uniform slurry, wherein the concentration (w/v) of the MMT is 0.2%. 15.42mg of TPCI was weighed into a centrifuge tube, and 1mL of dimethyl sulfoxide (DMSO) was added thereto, and dissolved by sonication, at which time the concentration of TPCI was 10 mmol/L. 100. mu.L of 10mmol/L TPCI was measured and diluted into 1mL of an aqueous solution, at which time the concentration of TPCI was 1 mmol/L. Weighing 2.1mL and 2mg/mL MMT solution in a centrifuge tube, slowly dropwise adding 1mmol/L TPCI solution with the volume ratio of MMT to TPCI being 9:1 under stirring at 60 ℃, and reacting for 48h in a dark place to obtain TPCI/MMT composite material solution, namely the photosensitizer/clay composite material 3.

Example 4:

6mL of PBS (pH 6.5) was weighed into a centrifuge tube, 12Mg of Montmorillonite (MMT) was weighed into the centrifuge tube, and disrupted in an ultrasonic cell disruptor for 30min to obtain a uniform slurry, at which time the concentration (w/v) of MMT was 0.2%. 15.42mg of TPCI was weighed into a centrifuge tube, and 1mL of dimethyl sulfoxide (DMSO) was added thereto, and dissolved by sonication, at which time the concentration of TPCI was 10 mmol/L. 100. mu.L of 10mmol/L TPCI was measured and diluted into 1mL of an aqueous solution, at which time the concentration of TPCI was 1 mmol/L. Weighing 2.1mL and 2mg/mL MMT solution in a centrifuge tube, slowly dripping 1mmol/L TPCI solution with the volume ratio of MMT to TPCI being 9:1 under the stirring of 40 ℃, and reacting for 18 hours in a dark place to obtain TPCI/MMT composite material solution, namely the photosensitizer/clay composite material 4.

Example 5:

6mL of PBS (pH 7.4) was weighed into a centrifuge tube, 12Mg of Montmorillonite (MMT) was weighed into the centrifuge tube, and disrupted in an ultrasonic cell disruptor for 30min to obtain a uniform slurry, at which time the concentration (w/v) of MMT was 0.2%. 15.42mg of TPCI was weighed into a centrifuge tube, and 1mL of dimethyl sulfoxide (DMSO) was added thereto, and dissolved by sonication, at which time the concentration of TPCI was 10 mmol/L. 100. mu.L of 10mmol/L TPCI was measured and diluted into 1mL of an aqueous solution, at which time the concentration of TPCI was 1 mmol/L. 880 mu L of PBS (pH 7.4) is weighed into a centrifuge tube, 100 mu L of MMT solution with 2mg/mL is dripped into the centrifuge tube under the stirring at the temperature of 20 ℃, then 20 mu L of TPCI solution with 1mmol/L is dripped slowly, and the TPCI/MMT composite material solution is obtained after the dark reaction for 48 hours, namely the photosensitizer/clay composite material 5.

Example 6:

6mL of PBS (pH 7.4) was weighed into a centrifuge tube, 12Mg of Montmorillonite (MMT) was weighed into the centrifuge tube, and disrupted in an ultrasonic cell disruptor for 30min to obtain a uniform slurry, at which time the concentration of MMT was 2 mg/mL. 15.42mg of TPCI was weighed into a centrifuge tube, and 1mL of dimethyl sulfoxide (DMSO) was added thereto, and dissolved by sonication, at which time the concentration of TPCI was 10 mmol/L. 100. mu.L of 10mmol/L TPCI was measured and diluted into 1mL of an aqueous solution, at which time the concentration of TPCI was 1 mmol/L. Measuring 860 mu L of PBS (pH 7.4) in a centrifuge tube, dropwise adding 100 mu L of 2mg/mL MMT solution under stirring at 20 ℃, then slowly dropwise adding 40 mu L of 1mmol/L of TPCI solution, and reacting for 48h in a dark place to obtain TPCI/MMT composite material solution, namely the photosensitizer/clay composite material 6.

To better illustrate the beneficial effects of the photosensitizer/clay composite of the present invention, it will be illustrated below by structural characterization, phototoxicity testing, antibacterial testing, and hemostatic testing.

1. The photosensitizer/clay composite 1 prepared in example 1 was subjected to structural characterization

In order to accurately judge the feasibility of the intercalation of the photosensitizer in clay, a sample of the photosensitizer/clay composite material 1 obtained in this example 1 was subjected to an X-ray diffractometer, an infrared spectrometer, a scanning electron microscope, BET pore size analysis and ESR measurement, and the results are shown in fig. 1, 2, 3, 4 and 5:

FIG. 1 is an XRD pattern of MMT and TPCI/MMT, obtained by XRD analysis, wherein the interlayer spacing of TPCI/MMT is enlarged to that of TPCI introduced between MMT layers

FIG. 2 is an FTIR plot of MMT, TPCI and TPCI/MMT, which is analyzed by FTIR and has distinct MMT and TPCI characteristic peaks on TPCI/MMT, the positions of the characteristic peaks are basically the same, and MMT and TPCI are conjectured by electrostatic force;

the SEM image of the MMT in FIG. 3a and the SEM image of the TPCI/MMT in FIG. 3b show that the MMT is a compact structure, and the interlayer spacing is enlarged and the structure is loose after the TPCI is intercalated;

the BET plot of FIG. 4a MMT and the BET plot of FIG. 4b TPCI/MMT, as determined by BET analysis, are of type III adsorption isotherm, are of polymolecular layer adsorption, and have a specific surface area of 28.13m for TPCI/MMT²A specific surface area per gram (5.22 m) much larger than MMT²The/g is favorable for adsorption;

FIG. 5a is a plot of OH production rate for TPCI/MMT and FIG. 5b is a plot of ESR detection OH production for TPCI/MMT, which was analyzed to generate a large amount of OH in light and faster than in the dark, while a strong characteristic OH peak was detected for ESR;

FIG. 6a TPCI/MMT¹O₂ESR detection to generate rate map and FIG. 6b TPCI/MMT¹O₂The generated quantity map can be analyzed, and the TPCI/MMT can generate a large quantity under illumination¹O₂While strong ESR was detected¹O₂A characteristic peak;

from the above, photosensitizers are successfully intercalated in clays and exhibit potential biological activity.

2. The photosensitizer/clay composite 1 prepared in example 1 was subjected to phototoxicity test

The specific content of the test is as follows:

cells were first incubated in 96-well plates at a density of 5000 cells per well for 24 hours, then the medium was aspirated, followed by addition of DMEM medium containing different concentrations of the materials MMT (0.01mg/mL, 0.1mg/mL,0.5mg/mL), TPCI (0.5. mu. mol/L, 5. mu. mol/L, 25. mu. mol/L), TPCI/MMT (0.01+0.5,0.1+5,0.5+25), incubation was continued for 4 hours, then the light group was illuminated under a white light for 1 hour, the dark group was shielded from light for 1 hour, incubation was continued for 12 hours, then 100. mu.L of MTT solution (0.5mg/mL) was added to each well for 4 hours, the medium was aspirated, and 100. mu.L of dimethyl sulfoxide solution was added to each well to dissolve crystal violet. After sufficient shaking, the absorbance at 570nm was read using a microplate reader.

The results are shown in FIG. 7, where MMT is more toxic at high concentrations; TPCI has larger cytotoxicity under illumination; the TPCI/MMT has less cytotoxicity under the light and the dark, which shows that the TPCI/MMT has less damage to normal cells and better biological safety under the same concentration no matter under the light or dark condition

3. The details of the antibacterial comparative test of the photosensitizer/clay composite 1 prepared in example 1 and clay are as follows:

3-5 independent colonies are picked up to a fresh LB liquid culture medium and cultured for 12h at 37 ℃ to grow to a stable phase, and the concentration of the bacteria liquid is calibrated. The bacterial suspension was diluted with PBS to OD 0.0001 (about 1mL with 2X 10)⁵Individual bacteria) TPCI/MMT and MMT are diluted by PBS, 1mL of diluted liquid medicine and 1mL of bacterial liquid are added into a glass bottle to be used as an experimental group, 1mL of bacterial liquid are added into the glass bottle to be used as a blank group, the blank group is illuminated by a white light lamp for 1h, and the blank group is processed to be a control experimental group in a dark place. Untreated bacterial blank served as a 100% control for bacterial growth, with each inhibition experiment performed in triplicate.

The results are shown in fig. 8a and 8b, the antibacterial rate of MMT under the condition of illumination, e.coli and S.a μ re μ s is 62% and 60%, respectively, and the antibacterial rate of TPCI/MMT under the condition of illumination is 100%, which shows that TPCI/MMT has better bactericidal effect than MMT, and may help to eliminate bacteria at wound infection and recover wound.

4. Comparative hemostasis test of photosensitizer/clay composite 1 prepared in example 6 with clay

Collecting 1mL of anticoagulated mouse blood, centrifuging, washing, diluting with PBS, mixing 150 μ L of diluted blood with the liquid medicine, adding 30 μ L of TRITON as a positive control group, adding 150 μ L of PBS as a negative control group, setting each group in 3 parallels, centrifuging at 3000r for 10min after 3h, and sampling 150 μ L of samples to a 96-well plate to determine the absorbance at 540 nm.

The result is shown in fig. 9, the MMT hemolysis is very obvious, the hemolysis rate reaches about 80%, TPCI/MMT has better blood compatibility, and the hemolysis rate is only about 2%, which shows that after TPCI is introduced into MMT layers, the hemolysis problem of MMT is solved, and the TPCI/MMT has better biological safety.

5. The series of photosensitizer/clay composites prepared in examples 1 to 5 were tested for antibacterial effect

The specific content of the test is as follows:

sample 1: 2mL of the photosensitizer/clay composite 1 solution prepared in example 1 was measured and diluted to 4mL of PBS (pH 6.5) to obtain a 1-fold diluted photosensitizer/clay composite 1. Uniformly mixing the bacterial liquid, the hydrogen peroxide solution and the photosensitizer/clay composite material 1 solution diluted by 1 time in the same volume, then keeping out of the sun in a constant temperature shaking table for 1 hour, and then absorbing the mixed solution with the same volume for illumination for 1 hour and dark treatment for 1 hour respectively.

Sample 2: 400. mu.L of the photosensitizer/clay composite 2 solution prepared in example 2 was measured and diluted to 4mL of a PBS (pH 6.5) solution to obtain a 10-fold diluted photosensitizer/clay composite 2 solution. Uniformly mixing the bacterial liquid, the hydrogen peroxide solution and the photosensitizer/clay composite material 2 diluted by 10 times in the same volume, then keeping out of the sun in a constant temperature shaking table for 1h, and then absorbing the mixed solution with the same volume for illumination for 1h and dark treatment for 1h respectively.

Sample 3: 40. mu.L of the reacted solution of the photosensitizer/clay composite 3 prepared in example 3 was measured and diluted to 4mL of a PBS (pH 6.5) solution to obtain a 100-fold diluted photosensitizer/clay composite 3 solution. Uniformly mixing the bacterial liquid, the hydrogen peroxide solution and the photosensitizer/clay composite material 3 solution diluted by 100 times in the same volume, then keeping out of the sun in a constant temperature shaking table for 1 hour, and then absorbing the mixed solution with the same volume for illumination for 1 hour and dark treatment for 1 hour respectively.

Sample 4: 80. mu.L of the reacted solution of the photosensitizer/clay composite 4 solution prepared in example 4 was measured and diluted to 4mL of a PBS (pH 6.5) solution to obtain a 20-fold diluted photosensitizer/clay composite 4 solution. Uniformly mixing the same volume of bacterial liquid, hydrogen peroxide solution and 20 times diluted photosensitizer/clay composite material 4 solution, then performing light-shielding action for 1h in a constant-temperature shaking table, and then absorbing the equal volume of mixed solution to perform illumination for 1h and dark treatment for 1h respectively.

The illumination condition is that the wavelength of illumination is 400 nm-600 nm and the power of the light source of illumination is 1mW/cm²～50mW/cm²。

The results are shown in FIGS. 10, 11, 12 and 13.

Sample 1 prepared in example 1 was used to resist 100% E.coli under light; 54.4% of escherichia coli resistance in the dark, 100% of staphylococcus aureus resistance in the light and 57.4% of staphylococcus aureus resistance in the dark.

Sample 2 prepared in example 2 was used to be resistant to 79.9% of E.coli under light; 48.7 percent of escherichia coli resistance in the dark, 62 percent of staphylococcus aureus resistance in the light and 55.7 percent of staphylococcus aureus resistance in the dark.

Sample 3 prepared in example 3 was used to protect against e.coli 45.7% under light; anti-escherichia coli 37.9% in dark, anti-staphylococcus aureus 48.8% in light and anti-staphylococcus aureus 45.3% in dark.

Sample 4 prepared in example 4 was used to be 57.5% resistant to e.coli under light; 46.6 percent of escherichia coli resistance in the dark, 51.4 percent of staphylococcus aureus resistance in the light and 49.1 percent of staphylococcus aureus resistance in the dark.

Obviously, the bacteriostatic effect of TPCI/MMT is obvious, mainly because the photosensitizer TPCI generates a large amount of active oxygen molecules under the action of illumination, and the active oxygen molecules effectively kill bacteria by destroying the structural integrity of cell membranes, increasing the ion permeability of the cell membranes or directly destroying unsaturated lipids, polypeptides, enzymes and other components in cells. Moreover, after 10-200 mu mol/L of hydrogen peroxide is added, Fe ions in the montmorillonite react with the hydrogen peroxide to generate hydroxyl free radicals, so that partial sterilization effect is provided, and the sterilization effect is improved to 100% after TPCI is introduced.

5. The series of photosensitizer/clay composites prepared in examples 5 to 6 were tested for hemostatic effect

The specific content of the test is as follows:

sample 5: the same volume of mouse red blood cells was mixed with the solution of photosensitizer/clay composite 5 prepared in example 5 for 3 hours.

Sample 6: the same volume of murine erythrocytes was mixed homogeneously with the photosensitizer/clay composite 6 solution prepared in example 6 for 3 h.

The results showed that the hemolysis rate of sample 5 was 3.84%; the hemolysis rate of sample 6 is 2.53%, which shows that the hemolysis rate of TPCI/MMT is reduced and the biological safety is good under the condition of the same MMT concentration and the loading amount of TPCI is improved to a certain extent.

6. The photosensitizer/clay composite 1 prepared in example 1 was subjected to an animal test with clay

The specific content of the test is as follows:

BALB/c mice were anesthetized, dehaired and the excision wound of 10mm diameter was generated on the skin of the back of the mice. Pseudomonas aeruginosa (10) is then treated⁸CFU/mL, 20 μ L) were inoculated on the wound site and left to stand for 10 minutes to stimulate infection, then the mice were divided into 4 groups, pbs (light), mmt (dark), TPCI/mmt (dark), and TPCI/mmt (light), 20 μ L of the drug solution was respectively dropped on the wound site of the mice, the mice in the light group were placed under a white light lamp for 30min, the mice in the dark group were treated for 30min in the dark, and the wound area and infection were observed and analyzed with ImageJ software.

As shown in fig. 14a and 14b, the results indicated that after 14 days, the wounds healed significantly, in particular, the wounds of the TPCI/mmt (light) group healed substantially completely; the TPCI/MMT (light) group healed the fastest wound by quantitative assessment of wound area.

In conclusion, the TPCI/MMT composite is more biologically safe than MMT and TPCI under the same concentration conditions. The MMT contains Fe²⁺And Fe³⁺In H₂O₂With the action of (2), OH is generated, and TPCI generates a large amount of light¹O₂These ROS (. OH and)¹O₂) The cell components (such as unsaturated lipid, polypeptide, enzyme and the like) are oxidized and damaged, so that bacteria are killed, and the bacteria at the wound can be killed; the MMT also has strong water absorption, can absorb a large amount of water, activate blood coagulation factors, stimulate fibrinogen to be converted into fibrin, promote blood coagulation, block vascular damage and achieve the effect of quickly stopping bleeding.

The features of the embodiments and embodiments described herein above may be combined with each other without conflict.

The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents, improvements and the like that fall within the spirit and principle of the present invention are intended to be included therein.

Claims

1. A photosensitizer/clay composite characterized by: comprises photosensitizer and layered clay, wherein the photosensitizer is intercalated in the clay, the photosensitizer is TPCI, the clay is montmorillonite, the structural formula of TPCI is shown as formula (I),

formula (I).

2. A process for preparing the photosensitizer/clay composite of claim 1, wherein: mixing clay and buffer solution to obtain uniform slurry, and dripping photosensitizer into the slurry to react in dark to obtain the photosensitizer/clay composite material.

3. The method of claim 2, wherein: the method comprises the following steps:

s1, adding a certain mass of clay into the buffer solution to prepare suspension with w/v of 0.2-0.5%

Liquid;

s2, carrying out ultrasonic crushing on the suspension obtained in the step S1 to obtain uniform slurry;

s3, dropwise adding 0.015-0.6 mg/mL of photosensitizer into the slurry obtained in the step S2 under the conditions of a certain temperature and magnetic stirring, and reacting for 6-48 h in a dark place;

s4, freeze-drying the solution reacted in the step S3 to obtain the photosensitizer/clay composite material.

4. A process for preparing a photosensitizer/clay composite according to claim 3, wherein: in step S1, the buffer solution is a phosphoric acid buffer solution, and the pH value of the phosphoric acid buffer solution is 6.5-7.4.

5. A process for preparing a photosensitizer/clay composite according to claim 3, wherein: the photosensitizer is TPCI, the clay is montmorillonite, and the mass ratio of the montmorillonite to the TPCI is as follows: 13.0 to 6.5.

6. Use of a photosensitizer/clay composite obtained by the process of any one of claims 2 to 5 in the preparation of an antimicrobial hemostatic dressing.

7. The use of claim 6, wherein: the application conditions comprise illumination conditions and hydrogen peroxide conditions, wherein the illumination conditions comprise illumination wavelength of 400-600 nm and illumination light source power of 1mW/cm²~50 mW/cm²(ii) a The concentration of the hydrogen peroxide in the hydrogen peroxide condition is 10-200 mu mol/L.