An antibacterial transparent dressing

Abstract

The invention discloses a kind of antibacterial transparent dressing and its preparation method and application, belong to the field of antibacterial drug design and synthesis technology.First, using p-carboxylbenzaldehyde, 2,4-dimethylpyrrole etc. as raw materials to prepare the fluoroboron dipyrrole parent nucleus substituted by monocarboxyl group;Then two heavy atom iodine are introduced at its 2,6 position to increase its triplet and singlet oxygen quantum yield;Further with 1-(2-aminoethyl) piperidine, 4-methyl-1-piperazine ethylamine and N, N-dimethylethylenediamine respectively carry out condensation reaction, finally compound is made into aqueous solution and C10-30 acrylate cross-linked polymer is loaded on transparent PU film, and antibacterial transparent dressing is prepared.The photosensitizer synthesis cost of the present invention is low, side reaction is few, and yield is higher, and it is easy to purify, and transparent PU film is easy to obtain, is conducive to industrialized production, and can be used to prepare medical antibacterial dressing.

Description

Technical Field

[0001] The invention belongs to the technical field of antibacterial drug design and synthesis, and particularly relates to an antibacterial transparent dressing and a preparation method and application thereof. Background Art

[0002] Bacterial wound infection is a major challenge worldwide. For example, currently over 400 million people worldwide are diagnosed with diabetes. The incidence of diabetic foot disease is approximately 6.3%, with a higher incidence in men than in women. Clinical manifestations of diabetic foot disease include wound infection, tissue damage, and bacterial inflammation, leading to edema, pain, and purulent exudate. This persistent inflammatory state produces a series of tissue reactions that make wound healing difficult. In fact, approximately 20% of moderate or severe diabetic foot infections result in some degree of amputation, which can be life-threatening and imposes a significant socioeconomic burden. There is currently no effective treatment for diabetic foot disease, resulting in global diabetes healthcare costs reaching $727 billion in 2017. Therefore, there is an urgent need to develop effective, safe, and easily applicable methods to prevent and treat skin infections.

[0003] BODIPY photosensitizers are acid and alkali resistant, have good photostability, and a high molar extinction coefficient. By introducing an iodine atom into the parent ring of BODIPY, the intersystem crossing efficiency is improved, allowing the transition of singlet excitation to triplet excited states and increasing the production of singlet oxygen. The introduction of tertiary amine groups into BODIPY compounds allows them to be easily protonated in aqueous solution, forming cationic quaternary ammonium, which enhances binding to bacteria on negatively charged surfaces. Furthermore, these compounds are not prone to aggregation and have good solubility. Their maximum absorption wavelength is 538nm, close to the maximum emission wavelength of sunlight, allowing them to exert antibacterial activity without the need for additional light source irradiation, making them particularly suitable for use in transparent antibacterial patches. Summary of the Invention

[0004] The present invention aims to overcome the limitation of traditional bandages that lack light transmission when photosensitizers are introduced into them, and to provide a method for preparing a transparent antibacterial dressing. This transparent dressing effectively kills most microorganisms, including Gram-negative bacteria, Gram-negative bacteria, and drug-resistant bacteria. Due to its excellent antibacterial activity, this transparent dressing significantly promotes wound healing.

[0005] To achieve the above object, the present invention adopts the following technical solutions:

[0006] A method for preparing an antibacterial transparent dressing comprises the following steps:

[0007] (1) Preparation of photosensitizer.

[0008] (2) The photosensitizer pure product is prepared into a mother solution with DMSO, which is then diluted with physiological saline and mixed with C10-30 acrylate cross-linked polymer, and then loaded on a transparent PU film to obtain a transparent antibacterial dressing.

[0009] The chemical structural formula of the photosensitizer is:

[0010] Where X is

[0011] The preparation method of the photosensitizer is to use compound 4-carboxybenzaldehyde, 2,4-dimethylpyrrole, 2,3-dichloro-5,6-dicyano-p-benzoquinone (DDQ), triethylamine (Et3N), boron trifluoride etherate (BF3·Et2O) and trifluoroacetic acid (CF3COOH) as starting materials to synthesize compound A Then, compound A, I2 and HIO3 were used as starting materials to synthesize iodinated fluoroborane dipyrrole derivative B. Then, iodinated fluoroborane dipyrrole derivative B, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, 1-hydroxybenzotriazole, triethylamine and any one of 4-methyl-1-piperazineethylamine, N-(2-aminoethyl)piperidine and N,N-dimethylethylenediamine are reacted to synthesize tertiary amino group-containing fluoroborane dipyrrole derivative type photosensitizer.

[0012] The obtained amino-substituted fluoroboron dipyrrole derivative can be used as an antibacterial photosensitizer in photodynamic therapy.

[0013] Furthermore, the above-mentioned fluoroboron dipyrrole derivatives are used to inhibit Gram-negative bacteria and Gram-positive bacteria.

[0014] Furthermore, the above-mentioned fluoroboron dipyrrole derivatives are used to kill antibiotic-resistant bacteria.

[0015] Furthermore, the fluoroboron dipyrrole derivative is used to prepare a transparent antibacterial dressing for use in a drug for promoting the healing of diabetic wound infections caused by drug-resistant bacteria.

[0016] The beneficial effects of the present invention are:

[0017] (1) This type of fluoroborane dipyrrole derivative contains a tertiary amino group that is easily protonated in aqueous solution, which facilitates the binding of the compound to bacterial cells. It does not require the introduction of fixed cationic groups and has low toxicity to mammalian cells.

[0018] (2) The maximum absorption of the compound is around 538 nm, which is close to the maximum absorption wavelength of sunlight and is very suitable for making a transparent antibacterial dressing.

[0019] (3) The fluoroborane dipyrrole derivative has undergone simple chemical modification, and its maximum absorption and emission are located in the green light region. It has weak tissue penetration ability and only treats the skin surface during photodynamic therapy without causing toxicity to deep tissues. It is an ideal photosensitizer.

[0020] (4) The target compound has a single structure, no isomers exist, and the product is easy to purify;

[0021] (5) The synthesis method is simple and can be completed in just a few steps. The reaction conditions are mild, there are few side reactions, the raw materials are easily available, and the cost is low, which is conducive to industrial production.

[0022] (6) It has broad-spectrum antibacterial properties and a good killing effect on drug-resistant bacteria. It is highly effective against bacteria and environmentally friendly. It has a positive effect on reducing the accumulation of drugs in the environment and weakening the drug resistance of pathogenic microorganisms. It overcomes the defects of ordinary active fungicides such as high residue, high pollution, and easy resistance. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 This is a preparation route for photosensitizer fluoroboron dipyrrole derivatives.

[0024] Figure 2 This is a diagram showing the antibacterial effect of photosensitizer fluoroboron dipyrrole derivatives on luminescent Escherichia coli and luminescent Staphylococcus aureus.

[0025] Figure 3 This is a diagram showing the hemolytic safety effect of the photosensitizer fluoroborane dipyrrole derivative Compound 3.

[0026] Figure 4 This is a picture of the effect of preparing a transparent patch using the photosensitizer fluoroborane dipyrrole derivative Compound 3.

[0027] Figure 5 This is a schematic diagram of the experiment on the transparent dressing prepared from the photosensitizer fluoroboron dipyrrole derivative Compound 3 to treat drug-resistant bacterial wound infection in diabetic mice and its effect in promoting wound healing.

[0028] Figure 6 This is the hydrogen spectrum of the photosensitizer fluoroboranedipyrrole derivative Compound 1.

[0029] Figure 7 This is the hydrogen spectrum of the photosensitizer fluoroboranedipyrrole derivative Compound 2.

[0030] Figure 8 This is the hydrogen spectrum of the photosensitizer fluoroboranedipyrrole derivative Compound 3. DETAILED DESCRIPTION

[0031] The present invention will be described in detail below with reference to specific embodiments. The following embodiments are not intended to limit the scope of protection of the present invention. Any improvements and changes made on the basis of the present invention are within the scope of protection of the present invention.

[0032] Example 1: Preparation of photosensitizer fluoroboron dipyrrole derivatives

[0033] The synthetic route of fluoroboranedipyrrole derivatives is shown in Figure 1 .

[0034] (1) 4-Carboxybenzaldehyde and 2,4-dimethylpyrrole were added to anhydrous tetrahydrofuran in a molar ratio of 1:2, and then 1 to 2 drops of trifluoroacetic acid were added. The mixture was reacted at room temperature in the dark overnight. Then, 1 equivalent (based on the molar amount of 4-carboxybenzaldehyde) of 2,3-dichloro-5,6-dicyano-p-benzoquinone was added. After the reaction was continued for 4 hours, triethylamine and boron trifluoride ether were added to the reaction system in an equivalent ratio of 1:1 (based on the molar amount of 4-carboxybenzaldehyde) under ice bath conditions. The reaction was allowed to proceed in the dark overnight and the reaction was terminated. The black solid was removed by filtration using diatomaceous earth. The crude product was then washed with water, saturated sodium bicarbonate solution and saturated brine. The orange compound A was separated by silica gel column chromatography using dichloromethane and methanol as eluents. 0.81 g, yield 33%. Structural characterization: 1 H NMR (500MHz, CDCl3): δ = 8.24 (d, J = 8.7Hz, 2H, ArH), 7.45 (d, J = 8.7Hz, 2H, ArH), 6.00 (s, 2H, pyrrole-H), 2.57 (s, 6H, CH3), 1.37 (s, 6H, CH3).

[0035] (2) Compound A, I2, and HIO3 were added to anhydrous ethanol in a molar ratio of 1:2.5:2, and the mixture was reacted at 60°C under nitrogen for 1 to 2 hours. After the reaction, the solvent was evaporated under reduced pressure, and then the mixture was separated by silica gel column chromatography using dichloromethane-methanol as the eluent to obtain a red solid compound B, whose structural formula is: 0.32 g, yield 95%; structural characterization: 1 H NMR (500MHz, CDCl3): δ=8.27 (d, J=8.0Hz, 2H, ArH), 7.42 (d, J=8.0Hz, 2H, ArH), 2.66 (s, 6H, CH3), 1.38 (s, 6H, CH3).

[0036] (3) Compound B, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, 1-hydroxybenzotriazole, and triethylamine were reacted in an ice bath in the dark for 1 hour, and then 4-methyl-1-piperazineethylamine was added and the reaction was continued in the dark for 20 hours under nitrogen protection. After the reaction, the reaction solution was poured into water and extracted three times with dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, and then dried under reduced pressure. The pink solid compound 1 was obtained after purification by silica gel chromatography using methanol-dichloromethane as the developing solvent. Its structural formula is Structural characterization: 1 HNMR (400MHz, CDCl3): δ = 8.05 (d, J = 7.6Hz, 2H), 7.38 (d, J = 7.6Hz, 2H), 3.71 (s, 2H), 2.9 1(dd,J=48.4,18.5Hz,10H),2.66(d,J=9.5Hz,6H),2.48(s,3H),1.40(d,J=21.2Hz,6H);

[0037] Compound B, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, 1-hydroxybenzotriazole, and triethylamine were reacted in an ice bath in the dark for 1 hour, and then 1-(2-aminoethyl)piperidine was added and the reaction was continued in the dark for 20 hours under nitrogen protection. After the reaction, the reaction solution was poured into water and extracted three times with dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, and then dried under reduced pressure. After purification by silica gel chromatography using methanol-dichloromethane as the developing solvent, a pink solid compound 2 was obtained, whose structural formula is Structural characterization: 1 HNMR (500MHz, CDCl3): δ = 8.03 (d, J = 8.6Hz, 2H), 7.62 (s, 1H), 7.36 (d, J = 8.6Hz, 2H), 3.65 (dd, J = 11.7, 5. 6Hz,2H),2.73(t,J=5.8Hz,2H),2.65(s,6H),2.60(s,3H),1.75-1.65(m,4H),1.53(s,2H),1.38(s,7H);

[0038] Compound B, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, 1-hydroxybenzotriazole, and triethylamine were reacted in an ice bath in the dark for 1 hour, and then N,N-dimethylethylenediamine was added and the reaction was continued in the dark for 20 hours under nitrogen protection. After the reaction, the reaction solution was poured into water and extracted three times with dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, and then dried under reduced pressure. After purification by silica gel chromatography using methanol-dichloromethane as the developing solvent, a pink solid compound 3 was obtained, whose structural formula is Structural characterization: 1HNMR (500MHz, CDCl3): δ = 8.01 (d, J = 8.3Hz, 2H), 7.36 (d, J = 8.3Hz, 2H), 3.62 (dt ,J=8.0,5.1Hz,2H),2.65(d,J=1.3Hz,8H),2.36(s,6H),1.37(d,J=1.4Hz,6H);

[0039] Example 2: Antibacterial properties of photosensitizer fluoroboron dipyrrole derivatives

[0040] The experimental strains used were Gram-positive luminescent Staphylococcus aureus, Gram-negative luminescent Escherichia coli, Gram-positive Staphylococcus aureus (S. aurues, ATCC 6358), Gram-negative Escherichia coli (E. coli, ATCC 8739), and methicillin-resistant Staphylococcus aureus (MRSA, ATCC 33591). The specific steps are as follows:

[0041] (1) Dissolve the fluoroborane dipyrrole derivative in DMSO as a stock solution (10 mmol / L). Use PBS solution to dilute the stock solution 3-fold to different concentrations and add it to a 96-well black board. Add three wells to each concentration as a parallel group, add 20 μL to each well, and set up a group with the same volume of PBS as a control group.

[0042] (2) The experimental group was divided into two groups, namely the light group and the dark group. The dark group was protected from light throughout the experiment.

[0043] (3) Pipette 180 μL of bacterial solution (10 6 CFU / mL) into each well and incubate for 30 min at a wavelength of 520 nm and a power of 4 mW / cm 2 The light source was irradiated for 4 min (the total light dose was 1 J / cm 2 ), and the luminescence value was measured using an enzyme-labeled instrument.

[0044] (4) The experiment was repeated 3 times, and the bacterial survival rate was calculated as follows: S l =(Le / Lc)×100%; where: S1—bacterial survival rate (%); Le—luminescence intensity of the test sample; Lc—luminescence intensity of the control sample.

[0045] (5) Use the two-fold dilution method to determine the MIC of the compound. Dilute the compound solution with LB medium in a concentration gradient. Take 100 μL of LB medium containing the compound and add 100 μL of bacterial solution (final concentration is 10 6CFU / mL). Three parallel wells were set up for each group. After incubation, the mixture was illuminated. The mixture was incubated at 37°C for 20 hours, and bacterial growth was observed (based on the clarity of the liquid and the absence of flocculent or lumpy white precipitates at the bottom). The concentration at which no bacterial growth was observed was designated as the MIC.

[0046] (6) The results showed the half-maximal inhibitory concentration (IC50) of the fluoroborane dipyrrole series derivatives against luminescent Escherichia coli and luminescent Staphylococcus aureus under light or no light conditions, such as Figure 2 As shown. Under light-free conditions, the IC50 values ​​of the three fluoroboron dipyrrole derivatives against luminescent Escherichia coli were in the micromolar range, and the IC50 values ​​against luminescent Staphylococcus aureus were in the nanomolar range. Table 1 shows the MIC values ​​of the fluoroboron dipyrrole derivatives against E. coli, S. aureus, and methicillin-resistant S. aureus. Among them, Compound 3 exhibited the best overall antibacterial activity.

[0047] Table 1 Comparison of antibacterial effects of photosensitizer fluoroboron dipyrrole derivatives

[0048]

[0049] Example 3: Hemolytic Safety of Photosensitizer Fluoroboron Dipyrrole Derivative (Compound 3)

[0050] 450 μL of PBS, 450 μL of 2% red blood cell suspension and 100 μL of compound PBS solution (final concentrations were 0.1, 1, 10, 25, 50 and 100 μM, respectively) were mixed. After incubation in a 37°C incubator for 30 minutes, the cells were irradiated with an LED flat panel light source for 4 minutes (dose of 1 J / cm 2 A dark control group, a positive control group (containing 550 μL of ultrapure water and 450 μL of a 2% red blood cell suspension), and a negative control group (containing 550 μL of PBS and 450 μL of a 2% red blood cell suspension) were also designed. The red blood cells were centrifuged at 800 rpm for 10 minutes until they were completely precipitated, and the ultraviolet absorption of the supernatant was measured at 570 nm. The hemolysis rate, Z, can be calculated using the following formula.

[0051] Z(%)=(D t -D nc ) / (D pc -D nc )×100%.

[0052] Here, D t 、D nc and Dpc represent the absorbance of samples, negative controls, and positive controls at 570 nm.

[0053] Example 4: Construction of a transparent antibacterial patch using a photosensitizer fluoroboron dipyrrole derivative (Compound 3)

[0054] A C10-30 alkyl acrylate crosspolymer was selected as the antimicrobial carrier. This polymer is a film-forming agent found in various cosmetics and has no adverse reactions or irritation risks. Compound 3 was dissolved in DMSO and diluted with saline. The mixture was then added to a final concentration of 25 μM and loaded onto an adhesively bonded transparent polyurethane film (3.5 cm × 3 cm), constructing a transparent patch on the side containing the adhesive.

[0055] Example 5: Experiment on the promotion of wound infection of drug-resistant bacteria in diabetic mice by photosensitizer fluoroboron dipyrrole derivative (Compound 3)

[0056] Methicillin-resistant Staphylococcus aureus (MRSA, ATCC 33591) was used as the experimental strain. Adult male ICR mice (4 weeks old, 18 ± 2 g) were housed and handled according to the recommendations of the Institutional Animal Care and Use Committee (IACUC). The animals were divided into two groups, 6 in each group, and maintained at 21°C on a 12-h light-dark cycle. After 4 weeks of high-fat diet (12492M, purchased from Beijing Boaigang Biotechnology Co., Ltd.), the mice weighed approximately 35 g. Each mouse was injected with 120 mg / kg STZ (0.1 M citrate-trisodium citrate buffer, pH 4.5, as an adjuvant) to induce hyperglycemia. Fasting blood glucose (FBG) levels were monitored using a glucometer on days 3, 7, and 14. During this period, bedding was changed daily, and the mice's living conditions were observed and recorded. Sufficient food and water were provided. Mice with fasting blood glucose levels stable above ≥11.2 mM and symptoms such as binge drinking, overeating, overdrinking, and weight loss were considered diabetic mice and selected for subsequent experiments. The general schematic diagram of the experiment is shown in the figure, and the specific steps are as follows:

[0057] (1) Wounds were established on diabetic mice and infected with MRSA bacteria. After the mice were anesthetized with isoflurane, the dorsal surface was shaved and a square piece of skin (10 mm × 10 mm) was excised at a depth of approximately 2.0 mm using sterile scissors. There was no obvious bleeding from the wound. A solution of MRSA growing in the logarithmic phase (50 μL) was evenly inoculated onto the wound surface of the mice and allowed to stand for 3 minutes to be absorbed by the wound tissue.

[0058] (2) Wound-infected diabetic mice were randomly divided into three groups, with 5 mice in each group. Two groups were given a patch with a concentration twice that of the MIC (25 μM), and one group was given a patch and LED light irradiation (520 nm, 4 mW / cm 2 ,8min, total light dose 2J / cm 2), and another group was not irradiated. A third group was treated with normal saline light therapy. 24 hours after wound infection, the light-exposed group covered the wound with a patch containing Compound 3 and was exposed to light once daily for the first three days. Throughout the observation period, all mice were free to move around, provided with adequate water and food, and exposed to natural ambient light in the room. The wound area and body weight of the mice were measured and recorded daily, and the average value for each group was taken. The mice were observed for 12 consecutive days, and their blood glucose levels were monitored weekly.

[0059] (3) Compared with the control group using normal saline, the wound healing rate in the treatment group was significantly accelerated ( Figure 5 (b)). The experimental results confirmed the bactericidal effect of Compound 3.

[0060] (4) Moreover, under 520nm LED light, wound healing is more obvious ( Figure 5 (b)). Wounds without Compound 3 treatment showed suppuration, but those in the experimental group did not.

[0061] (5) Compound 3 did not affect the normal weight gain of mice during the wound treatment process, as shown in ( Figure 5 (d)) shows that Compound 3 has no obvious biological toxicity.

[0062] The above description is only a preferred embodiment of the present invention. All equivalent changes and modifications made according to the scope of the patent application of the present invention should fall within the scope of the present invention.

Claims

1. An antibacterial transparent dressing, characterized by: The antibacterial transparent patch is loaded with an antibacterial photosensitizer, the chemical structure of which is: Where X is Any of .

2. A method for preparing the antibacterial transparent dressing according to claim 1, characterized in that: The following steps are involved: (1) At room temperature, a carboxy BODIPY core was synthesized using 4-carboxybenzaldehyde, 2,4-dimethylpyrrole, 2,3-dichloro-5,6-dicyanobenzoquinone, triethylamine, boron trifluoride etherate, and trifluoroacetic acid as starting materials. Iodocarboxyfluoroboron dipyrrole derivatives were then synthesized using the carboxy BODIPY core, I2, and HIO3 as starting materials. (2) dissolving an iodocarboxyl fluoroboron dipyrrole derivative in solution A, causing an activation reaction, adding an amino small molecule, stirring, spin-drying, and removing impurities to obtain the photosensitizer; the solution A is a mixed solution of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, 1-hydroxybenzotriazole, and triethylamine; (3) The photosensitizer is dissolved in dimethyl sulfoxide, diluted and mixed evenly with a C10-30 acrylate cross-linked polymer, and loaded on a transparent polyurethane film to obtain the antibacterial transparent dressing.

3. The method according to claim 2, wherein: In step (1), the molar ratio of 4-carboxybenzaldehyde, 2,4-dimethylpyrrole, 2,3-dichloro-5,6-dicyanobenzoquinone, triethylamine, and boron trifluoride etherate is 1:2:1:1:1; the molar ratio of carboxyl BODIPY core, I2, and HIO3 is 1:2.5:

2.

4. The method according to claim 2, wherein: The amino small molecule described in step (2) is any one of 4-methyl-1-piperazineethylamine, 1-(2-aminoethyl)piperidine, and N,N-dimethylethylenediamine; and the molar ratio of the iodinated carboxyl fluoroboron dipyrrole derivative to the amino small molecule is 1:

10.

5. The method according to claim 2, wherein: The concentration of the photosensitizer in the application of step (3) is 25 μM.

6. Use of the antibacterial transparent dressing according to claim 1 in the preparation of medicines for bacterial skin infections.

7. The use according to claim 6, characterized in that: The antibacterial transparent patch is applied on the skin and illuminated with a 520nm LED flat panel light source.

Images