A method for preparing a nanofiber membrane for visual diagnostics and differentiated drug delivery

Abstract

This invention relates to the field of medical materials. The invention discloses a method for preparing a nanofiber membrane for visual diagnosis and differentiated drug release, comprising: (1) synthesizing modified cellulose acetate and preparing spinning solution 1; (2) preparing spinning solution 2; (3) preparing spinning solution 3; (4) spraying spinning solution 1 onto paper; and (5) coaxial electrospinning. The nanofiber wound dressing prepared by this invention is pH-responsive, and fluorescent dyes are fixed on the nanofibers. The infection status of the wound is monitored by the change in fluorescence intensity according to pH changes, and the infection status of the wound is monitored by the change in color visible to the naked eye according to pH changes. Antibacterial drugs are loaded into a pH-responsive coaxial structure. When the wound becomes infected, the pH increases, and the antibacterial drugs are released, thus achieving the on-demand drug release property of the nanofiber dressing, realizing visual diagnosis and controllable treatment of chronic wounds.

Description

Technical Field

[0001] This invention relates to the field of medical materials, and more particularly to a method for preparing a nanofiber membrane for visual diagnosis and differentiated drug delivery. Background Technology

[0002] Electrospinning is a technique for preparing nanofiber membranes with large specific surface area, high porosity, tunable structure, low cost, and wide applicability. Its high encapsulation efficiency, sustained drug release, and adjustable properties make electrospun nanofiber membranes ideal drug delivery dressings. Many previous nanofiber dressings carried antibacterial drugs; however, overuse of antibacterial drugs can lead to bacterial resistance, making on-demand drug delivery essential. On-demand drug delivery refers to the material releasing a specific amount of antibacterial drug at a designated wound site, significantly reducing drug exposure time and preventing premature release, thus lowering the risk of bacterial resistance. Generally, when stimulated by external factors (temperature, light, pH, the appearance of enzymes or ions, or stimulation triggered by magnetic or electric fields), these smart materials undergo common physicochemical changes to achieve drug release. These stimuli are divided into endogenous and exogenous stimuli. Exogenous stimuli have time limitations and cannot spontaneously release drugs according to changes in the wound environment, missing the optimal release time. Therefore, achieving on-demand drug delivery based on endogenous stimulation (pH) is crucial.

[0003] Furthermore, for chronic wound infections, frequent dressing changes can cause secondary tearing and infection. Therefore, wound condition monitoring is increasingly important in modern healthcare systems. Early detection and treatment are crucial steps in improving wound management and reducing antibiotic treatment. Commercially available pH test strips and pH meters are commonly used to measure wound pH. However, these invasive analytical methods require manual removal of the dressing to detect the wound pH, which can lead to the shedding of newly formed skin tissue, disrupting the wound healing process, and increasing the risk of infection, thus causing further damage. Therefore, there is a significant need for intelligent, non-invasive sensor materials that are easily identifiable by the naked eye for personal care and daily use. Current wound pH monitoring systems rely on visual color differentiation and electrical signal measurement. However, these methods have limitations, such as the susceptibility of visual color recognition to errors and the susceptibility of electrodes to contamination. Therefore, to achieve both easy visual observation of the wound's true condition and avoid errors caused by blood or tissue fluid contamination of the dressing, we need to design a dual-color monitoring system with pH-dependent visible and fluorescence intensity changes to improve the sensitivity and accuracy of the detection system. Furthermore, most dyes are currently loaded into the polymer matrix of dressings through physical blending, which inevitably leads to the release of dyes from the dressing. This not only reduces the sensitivity of the sensor but may also cause side effects. Therefore, achieving dye immobilization is a major challenge in the construction of monitoring dressings.

[0004] In summary, there is an urgent need to further develop novel wound nanofiber membranes with dye-immobilized pH-responsive integrated diagnostic and therapeutic properties. Summary of the Invention

[0005] To address the aforementioned technical problems, this invention provides a method for preparing a nanofiber membrane for visual diagnosis and differentiated drug release. The invention first grafts hydroxyl-containing cellulose acetate and carboxyl-containing 5(6)-carboxynaphthylfluorescein via esterification; then, the modified cellulose acetate is sprayed onto paper; subsequently, acrylic resin and polycaprolactone are coaxially spun using electrospinning technology, and the nanofibers are received by paper coated with the modified cellulose acetate, thus obtaining a pH-responsive nanofiber wound dressing that stimulates drug release and monitors wound healing. The nanofiber wound dressing prepared by this invention is pH-responsive, enabling visual diagnosis and controlled treatment of chronic wounds by monitoring changes in wound pH.

[0006] The specific technical solution of this invention is: a method for preparing a nanofiber membrane for visual diagnosis and differentiated drug release, comprising the following steps:

[0007] (1) Add cellulose acetate to a reaction vessel containing a mixed solvent of dichloromethane and N,N-dimethylacetamide, and then add 4-dimethylaminopyridine, 5(6)-carboxynaphthylfluorescein and dicyclohexylcarbodiimide in sequence; stir the reaction under ice bath conditions, and dialyze the reaction product in a mixed solvent of dichloromethane and N,N-dimethylacetamide to obtain a modified cellulose acetate polymer solution, named spinning solution 1.

[0008] (2) Dissolve the antibacterial agent and polycaprolactone in a mixed solvent of tetrahydrofuran and N,N-dimethylformamide to obtain spinning solution 2.

[0009] (3) Dissolve acrylic resin in anhydrous ethanol to obtain spinning solution 3.

[0010] (4) Spray the spinning solution 1 evenly onto the paper, let it dry, and set aside.

[0011] (5) Coaxial electrospinning is performed with spinning solution 2 as the coaxial core layer and spinning solution 3 as the coaxial shell layer. The paper obtained in step (4) is used for receiving. After spinning for 4 hours, the nanofiber membrane is peeled off from the baking paper to obtain a nanofiber membrane with a two-layer composite structure.

[0012] As described in the background section of this application, the current research lacks understanding of antibiotic abuse and wound dye fixation. This invention uses esterification grafting of cellulose acetate with hydroxyl groups and 5(6)-carboxynaphthylfluorescein with carboxyl groups to regulate the fluorescence intensity and color change of nanofibers through pH changes; it loads antibacterial drugs into a pH-responsive coaxial structure, thereby achieving the properties of a controlled-release nanofiber dressing, thus enabling the nanofiber membrane with two functions to realize the visual diagnosis and controlled treatment of chronic wounds.

[0013] The principle of pH response in this invention is as follows: the shell layer is a pH-sensitive polymer, acrylic resin. When the pH value of the solution is ≤4.5, the acrylic resin chains become insoluble due to the protonation of the COO- groups. Therefore, the solubility of the entire nanofiber in the solution is very poor, and the drug embedded in the core layer is difficult to release. When the pH value of the solution is ≥7.4, the solubility of the acrylic resin chains in water increases due to the weakening of the protonation of the COO- groups. The structure of the nanofiber gradually collapses, and the drug carrier embedded in the core layer is released from the fiber, thereby realizing the properties of a controlled-release nanofiber dressing.

[0014] Preferably, in step (1), the mass ratio of cellulose acetate, 5(6)-carboxynaphthylfluorescein, 4-dimethylaminopyridine and dicyclohexylcarbodiimide is (90-100):(1-5):(1-3):(0.5-1).

[0015] Because 5(6)-carboxynaphthyl fluorescein has high sensitivity and high price, the grafting rate only needs to reach about 1% to achieve the function. Therefore, the mass ratio of cellulose acetate to 5(6)-carboxynaphthyl fluorescein is (90-100):(1-5).

[0016] Preferably, in step (1), the volume ratio of dichloromethane to N,N-dimethylacetamide is 2-3:6-7.

[0017] Preferably, in step (1), the concentration of the spinning solution 1 is 5-10 wt%.

[0018] Preferably, in step (2), the antibacterial drug is a hydrophilic drug and / or a lipophilic drug.

[0019] Preferably, in step (2), the hydrophilic drug is tetracycline; and the hydrophobic drug is roxithromycin.

[0020] Preferably, in step (2), the concentration of polycaprolactone in the spinning solution 2 is 10-15 wt%, and the concentration of the antibacterial agent is 5-10 wt%.

[0021] Preferably, in step (3), the concentration of the spinning solution 3 is 15-20 wt%.

[0022] Preferably, in step (4), the spraying distance is 10-20cm and the air pressure is 2-4kPa.

[0023] Preferably, in step (5), the electrospinning parameters of the core layer and the shell layer are: needle size 20-22G, distance 10-15cm, voltage 15-20kv, temperature 25-35℃, humidity 30-50%; the spinning speeds of the core layer and the shell layer are 0.003-0.006mm / s and 0.004-0.005mm / s, respectively.

[0024] Compared with the prior art, the present invention has the following technical effects: The nanofiber wound dressing prepared by the present invention is pH responsive. The present invention fixes the fluorescent dye on the nanofiber by esterification grafting of cellulose acetate with hydroxyl groups and 5(6)-carboxynaphthyl fluorescein with carboxyl groups, preventing the dye from falling off; pH responsive fluorescent probe, which can provide a clearly visible color change from purple to blue and a significant fluorescence change in the pH range (about 6-8) of the infected wound, that is, it provides a two-color monitoring system with pH-dependent visible and fluorescence intensity changes; Antibacterial drugs are loaded into a pH responsive coaxial structure. When the wound is infected, the pH rises and the antibacterial drug is released, thus achieving the on-demand drug release property of the nanofiber dressing, thereby realizing the visual diagnosis and controllable treatment of chronic wounds. Detailed Implementation

[0025] The present invention will be further described below with reference to embodiments.

[0026] Example 1

[0027] (1) Add cellulose acetate (0.7g) to a reaction vessel containing a mixture of dichloromethane (3ml) and N,N-dimethylacetamide (7ml), and then add 4-dimethylaminopyridine (3mg), 5(6)-carboxynaphthylfluorescein (3mg) and dicyclohexylcarbodiimide (0.5mg) in sequence; stir the reaction under ice bath conditions, and dialyze and filter the reaction product for 72h to obtain a 7%wt modified cellulose acetate polymer solution, named spinning solution 1, for later use;

[0028] (2) Using N,N-dimethylformamide and tetrahydrofuran (1:1) as solvents, add 10wt% polycaprolactone (1.4g) and stir to dissolve. Then add 5wt% hydrophilic drug tetracycline (0.75g) to obtain drug-loaded polycaprolactone spinning solution, named spinning solution 2.

[0029] (3) Using anhydrous ethanol as a solvent, a 15wt% acrylic resin spinning solution was prepared and named spinning solution 3.

[0030] (4) Spray the spinning solution 1 (5ml) evenly onto the baking paper with a spray gun, let it dry, and set it aside. The spraying distance is 15cm and the air pressure is 3kpa.

[0031] (5) Coaxial electrospinning was performed on spinning solution 2 and spinning solution 3, and the nanofiber membrane was peeled off from the baking paper in step (4) after spinning for 4 hours to obtain a two-layer composite nanofiber membrane. The core layer is polycaprolactone and the shell layer is acrylic resin. The electrospinning parameters are: needle 21G, distance 10cm, voltage 18kv, temperature 30℃, humidity 40%, and the spinning speeds of the core layer and the shell layer are 0.0007mm / s and 0.0042mm / s, respectively.

[0032] (6) Weigh 60mg of the composite nanofiber membrane obtained in step (5), put it into a centrifuge tube containing 45ml of PBS (pH 7.4) solution, release the drug on a shaker at 37 degrees Celsius, and periodically take 3ml of the solution to measure the ultraviolet light.

[0033] (7) Cut a 1cm*1cm nanofiber membrane obtained from (5), place it in PBS with pH 4.5 / 5 / 5.5 / 6 / 6.5 / 7 / 7.4 / 8 to observe the color change of the membrane, and use a fluorescence spectrophotometer to test the change in fluorescence intensity.

[0034] Comparative Example 1-1

[0035] (1) Add cellulose acetate (0.7g) to a reaction vessel containing a mixture of dichloromethane (3ml) and N,N-dimethylacetamide (7ml), and then add 4-dimethylaminopyridine (3mg), 5(6)-carboxynaphthylfluorescein (3mg) and dicyclohexylcarbodiimide (0.5mg) in sequence; stir the reaction under ice bath conditions, and dialyze and filter the reaction product for 72h to obtain a 7%wt modified cellulose acetate polymer solution, named spinning solution 1, for later use;

[0036] (2) Using N,N-dimethylformamide and tetrahydrofuran (1:1) as solvents, add 10wt% polycaprolactone (1.4g) and stir to dissolve. Then add 1wt% hydrophilic drug tetracycline (0.14g) to obtain drug-loaded polycaprolactone spinning solution, named spinning solution 2.

[0037] (3) Using anhydrous ethanol as a solvent, a 15wt% acrylic resin spinning solution was prepared and named spinning solution 3.

[0038] (4) Spray the spinning solution 1 (5ml) evenly onto the baking paper with a spray gun, let it dry, and set it aside. The spraying distance is 15cm and the air pressure is 3kpa.

[0039] (5) Coaxial electrospinning was performed on spinning solution 2 and spinning solution 3, and the nanofiber membrane was collected by baking paper from step (4). After spinning for 4 hours, the nanofiber membrane was peeled off from the baking paper to obtain a two-layer composite nanofiber membrane. The core layer was polycaprolactone and the shell layer was acrylic resin. The electrospinning parameters were: needle 21G, distance 10cm, voltage 18kv, temperature 30℃, humidity 40%, and the spinning speeds of the core layer and the shell layer were 0.0007mm / s and 0.0042mm / s, respectively.

[0040] (6) Weigh 60mg of the composite nanofiber membrane obtained in step (5), put it into a centrifuge tube containing 45ml of PBS (pH 4.5) solution, release the drug on a shaker at 37 degrees Celsius, and periodically take 3ml of the solution to measure the ultraviolet light.

[0041] (7) Cut a 1cm*1cm nanofiber membrane obtained from (5), place it in PBS with pH 4.5 / 5 / 5.5 / 6 / 6.5 / 7 / 7.4 / 8 to observe the color change of the membrane, and use a fluorescence spectrophotometer to test the change in fluorescence intensity.

[0042] Comparative Examples 1-2

[0043] (1) Add cellulose acetate (0.7g) to a reaction vessel containing a mixture of dichloromethane (3ml) and N,N-dimethylacetamide (7ml), and then add 4-dimethylaminopyridine (3mg), 5(6)-carboxynaphthylfluorescein (3mg) and dicyclohexylcarbodiimide (0.5mg) in sequence; stir the reaction under ice bath conditions, and dialyze and filter the reaction product for 72h to obtain a 7%wt modified cellulose acetate polymer solution, named spinning solution 1, for later use;

[0044] (2) Using N,N-dimethylformamide and tetrahydrofuran (1:1) as solvents, add 10wt% polycaprolactone (1.4g) and stir to dissolve. Then add 5wt% hydrophilic drug tetracycline (0.75g) to obtain drug-loaded polycaprolactone spinning solution, named spinning solution 2.

[0045] (3) Using anhydrous ethanol as a solvent, a 15wt% acrylic resin spinning solution was prepared and named spinning solution 3.

[0046] (4) Spray the spinning solution 1 (5ml) evenly onto the baking paper with a spray gun, let it dry, and set it aside. The spraying distance is 15cm and the air pressure is 3kpa.

[0047] (5) Coaxial electrospinning was performed on spinning solution 2 and spinning solution 3, and the nanofiber membrane was peeled off from the baking paper in step (4) after spinning for 4 hours to obtain a two-layer composite nanofiber membrane. The core layer is polycaprolactone and the shell layer is acrylic resin. The electrospinning parameters are: needle 21G, distance 10cm, voltage 18kv, temperature 30℃, humidity 40%, and the spinning speeds of the core layer and the shell layer are 0.0007mm / s and 0.0042mm / s, respectively.

[0048] (6) Weigh 60mg of the composite nanofiber membrane obtained in step (5), put it into a centrifuge tube containing 45ml of PBS (pH 7.4) solution, release the drug on a shaker at 37 degrees Celsius, and periodically take 3ml of the solution to measure the ultraviolet light.

[0049] (7) Cut a 1cm*1cm nanofiber membrane obtained from (5), place it in PBS with pH 4.5 / 5 / 5.5 / 6 / 6.5 / 7 / 7.4 / 8 to observe the color change of the membrane, and use a fluorescence spectrophotometer to test the change in fluorescence intensity.

[0050] Comparative Examples 1-3

[0051] (1) Add cellulose acetate (0.7g) to a reaction vessel containing a mixture of dichloromethane (3ml) and N,N-dimethylacetamide (7ml), and then add 4-dimethylaminopyridine (3mg), 5(6)-carboxynaphthylfluorescein (3mg) and dicyclohexylcarbodiimide (0.5mg) in sequence; stir the reaction under ice bath conditions, and dialyze and filter the reaction product for 72h to obtain a 7%wt modified cellulose acetate polymer solution, named spinning solution 1, for later use;

[0052] (2) Using N,N-dimethylformamide and tetrahydrofuran (1:1) as solvents, add 10wt% polycaprolactone (1.4g) and stir to dissolve. Then add 1wt% hydrophilic drug tetracycline (0.14g) to obtain drug-loaded polycaprolactone spinning solution, named spinning solution 2.

[0053] (3) Using anhydrous ethanol as a solvent, a 15wt% acrylic resin spinning solution was prepared and named spinning solution 3.

[0054] (4) Spray the spinning solution 1 evenly onto the baking paper with a spray gun, let it dry, and set it aside. The spraying distance is 15cm and the air pressure is 3kpa.

[0055] (5) Coaxial electrospinning was performed on spinning solution 2 and spinning solution 3, and the nanofiber membrane was collected by baking paper in step (4). After spinning for 4 hours, the nanofiber membrane was peeled off from the baking paper to obtain a two-layer composite nanofiber membrane. The core layer is polycaprolactone and the shell layer is acrylic resin. The electrospinning parameters are: needle 21G, distance 10cm, voltage 18kv, temperature 30℃, humidity 40%, and the spinning speeds of the core layer and the shell layer are 0.0007mm / s and 0.0007mm / s, respectively.

[0056] (6) Weigh 60mg of the composite nanofiber membrane obtained in step (5), put it into a centrifuge tube containing 45ml of PBS (pH4.5) solution, release the drug on a shaker at 37 degrees Celsius, and periodically take 3ml of the solution to measure the ultraviolet light.

[0057] (7) Cut a 1cm*1cm nanofiber membrane obtained from (5), place it in PBS with pH 4.5 / 5 / 5.5 / 6 / 6.5 / 7 / 7.4 / 8 to observe the color change of the membrane, and use a fluorescence spectrophotometer to test the change in fluorescence intensity.

[0058] Comparative Examples 1-4

[0059] (1) Add cellulose acetate (0.7g) to a mixed solvent containing dichloromethane (3ml) and N,N-dimethylacetamide (7ml) to obtain a 7% wt cellulose acetate polymer solution, named spinning solution 1, for later use;

[0060] (2) Using N,N-dimethylformamide and tetrahydrofuran (1:1) as solvents, add 10wt% polycaprolactone (1.4g) and stir to dissolve. Then add 1wt% hydrophilic drug tetracycline (0.14g) to obtain drug-loaded polycaprolactone spinning solution, named spinning solution 2.

[0061] (3) Using anhydrous ethanol as a solvent, a 15wt% acrylic resin spinning solution was prepared and named spinning solution 3.

[0062] (4) Spray the spinning solution 1 evenly onto the baking paper with a spray gun, let it dry, and set it aside. The spraying distance is 15cm and the air pressure is 3kpa.

[0063] (5) Coaxial electrospinning was performed on spinning solution 2 and spinning solution 3, and the nanofiber membrane was peeled off from the baking paper in step (4) after spinning for 4 hours to obtain a two-layer composite nanofiber membrane. The core layer is polycaprolactone and the shell layer is acrylic resin. The electrospinning parameters are: needle 21G, distance 10cm, voltage 18kv, temperature 30℃, humidity 40%, and the spinning speeds of the core layer and the shell layer are 0.0007mm / s and 0.0042mm / s, respectively.

[0064] (6) Weigh 60mg of the composite nanofiber membrane obtained in step (5), put it into a centrifuge tube containing 45ml of PBS (pH 4.5) solution, release the drug on a shaker at 37 degrees Celsius, and periodically take 3ml of the solution to measure the ultraviolet light.

[0065] (7) Cut a 1cm*1cm nanofiber membrane obtained from (5), place it in PBS with pH 4.5 / 5 / 5.5 / 6 / 6.5 / 7 / 7.4 / 8 to observe the color change of the membrane, and use a fluorescence spectrophotometer to test the change in fluorescence intensity.

[0066] Example 2

[0067] (1) Add cellulose acetate (0.7g) to a reaction vessel containing a mixture of dichloromethane (3ml) and N,N-dimethylacetamide (7ml), and then add 4-dimethylaminopyridine (3mg), 5(6)-carboxynaphthylfluorescein (3mg) and dicyclohexylcarbodiimide (0.5mg) in sequence; stir the reaction under ice bath conditions, and dialyze and filter the reaction product for 72h to obtain a 7%wt modified cellulose acetate polymer solution, named spinning solution 1, for later use;

[0068] (2) Using N,N-dimethylformamide and tetrahydrofuran (1:1) as solvents, add 10wt% polycaprolactone (1.4g) and stir to dissolve. Then add 5wt% hydrophobic drug roxithromycin (0.75g) to obtain drug-loaded polycaprolactone spinning solution, named spinning solution 2.

[0069] (3) Using anhydrous ethanol as a solvent, a 15wt% acrylic resin spinning solution was prepared and named spinning solution 3.

[0070] (4) Spray the spinning solution 1 (5ml) evenly onto the baking paper with a spray gun, let it dry, and set it aside. The spraying distance is 15cm and the air pressure is 3kpa.

[0071] (5) Coaxial electrospinning was performed on spinning solution 2 and spinning solution 3, and the nanofiber membrane was peeled off from the baking paper in step (4) after spinning for 4 hours to obtain a two-layer composite nanofiber membrane. The core layer is polycaprolactone and the shell layer is acrylic resin. The electrospinning parameters are: needle 21G, distance 10cm, voltage 18kv, temperature 30℃, humidity 40%, and the spinning speeds of the core layer and the shell layer are 0.0007mm / s and 0.0042mm / s, respectively.

[0072] (6) Weigh 60mg of the composite nanofiber membrane obtained in step (5), put it into a centrifuge tube containing 45ml of PBS (pH 7.4) solution, release the drug on a shaker at 37 degrees Celsius, and periodically take 3ml of the solution to measure the ultraviolet light.

[0073] (7) Cut a 1cm*1cm nanofiber membrane obtained from (5), place it in PBS with pH 4.5 / 5 / 5.5 / 6 / 6.5 / 7 / 7.4 / 8 to observe the color change of the membrane, and use a fluorescence spectrophotometer to test the change in fluorescence intensity.

[0074] Comparative Example 2-1

[0075] (1) Add cellulose acetate (0.7g) to a reaction vessel containing a mixture of dichloromethane (3ml) and N,N-dimethylacetamide (7ml), and then add 4-dimethylaminopyridine (3mg), 5(6)-carboxynaphthylfluorescein (3mg) and dicyclohexylcarbodiimide (0.5mg) in sequence; stir the reaction under ice bath conditions, and dialyze and filter the reaction product for 72h to obtain a 7%wt modified cellulose acetate polymer solution, named spinning solution 1, for later use;

[0076] (2) Using N,N-dimethylformamide and tetrahydrofuran (1:1) as solvents, add 10wt% polycaprolactone (1.4g) and stir to dissolve. Then add 5wt% hydrophobic drug roxithromycin (0.75g) and 1wt% hydrophobic drug roxithromycin (0.14g) to obtain drug-loaded polycaprolactone spinning solution, named spinning solution 2;

[0077] (3) Using anhydrous ethanol as a solvent, a 15wt% acrylic resin spinning solution was prepared and named spinning solution 3.

[0078] (4) Spray the spinning solution 1 (5ml) evenly onto the baking paper with a spray gun, let it dry, and set it aside. The spraying distance is 15cm and the air pressure is 3kpa.

[0079] (5) Coaxial electrospinning was performed on spinning solution 2 and spinning solution 3, and the nanofiber membrane was collected by baking paper from step (4). After spinning for 4 hours, the nanofiber membrane was peeled off from the baking paper to obtain a two-layer composite nanofiber membrane. The core layer was polycaprolactone and the shell layer was acrylic resin. The electrospinning parameters were: needle 21G, distance 10cm, voltage 18kv, temperature 30℃, humidity 40%, and the spinning speeds of the core layer and the shell layer were 0.0007mm / s and 0.0042mm / s, respectively.

[0080] (6) Weigh 60mg of the composite nanofiber membrane obtained in step (5), put it into a centrifuge tube containing 45ml of PBS (pH 4.5) solution, release the drug on a shaker at 37 degrees Celsius, and periodically take 3ml of the solution to measure the ultraviolet light.

[0081] (7) Cut a 1cm*1cm nanofiber membrane obtained from (5), place it in PBS with pH 4.5 / 5 / 5.5 / 6 / 6.5 / 7 / 7.4 / 8 to observe the color change of the membrane, and use a fluorescence spectrophotometer to test the change in fluorescence intensity.

[0082] Comparative Example 2-2

[0083] (1) Add cellulose acetate (0.7g) to a reaction vessel containing a mixture of dichloromethane (3ml) and N,N-dimethylacetamide (7ml), and then add 4-dimethylaminopyridine (3mg), 5(6)-carboxynaphthylfluorescein (3mg) and dicyclohexylcarbodiimide (0.5mg) in sequence; stir the reaction under ice bath conditions, and dialyze and filter the reaction product for 72h to obtain a 7%wt modified cellulose acetate polymer solution, named spinning solution 1, for later use;

[0084] (2) Using N,N-dimethylformamide and tetrahydrofuran (1:1) as solvents, add 10wt% polycaprolactone (1.4g) and stir to dissolve. Then add 5wt% hydrophobic drug roxithromycin (0.75g) to obtain drug-loaded polycaprolactone spinning solution, named spinning solution 2.

[0085] (3) Using anhydrous ethanol as a solvent, a 15wt% acrylic resin spinning solution was prepared and named spinning solution 3.

[0086] (4) Spray the spinning solution 1 (5ml) evenly onto the baking paper with a spray gun, let it dry, and set it aside. The spraying distance is 15cm and the air pressure is 3kpa.

[0087] (5) Coaxial electrospinning was performed on spinning solution 2 and spinning solution 3, and the nanofiber membrane was peeled off from the baking paper in step (4) after spinning for 4 hours to obtain a two-layer composite nanofiber membrane. The core layer is polycaprolactone and the shell layer is acrylic resin. The electrospinning parameters are: needle 21G, distance 10cm, voltage 18kv, temperature 30℃, humidity 40%, and the spinning speeds of the core layer and the shell layer are 0.0007mm / s and 0.0042mm / s, respectively.

[0088] (6) Weigh 60mg of the composite nanofiber membrane obtained in step (5), put it into a centrifuge tube containing 45ml of PBS (pH 7.4) solution, release the drug on a shaker at 37 degrees Celsius, and periodically take 3ml of the solution to measure the ultraviolet light.

[0089] (7) Cut a 1cm*1cm nanofiber membrane obtained from (5), place it in PBS with pH 4.5 / 5 / 5.5 / 6 / 6.5 / 7 / 7.4 / 8 to observe the color change of the membrane, and use a fluorescence spectrophotometer to test the change in fluorescence intensity.

[0090] Comparative Examples 2-3

[0091] (1) Add cellulose acetate (0.7g) to a reaction vessel containing a mixture of dichloromethane (3ml) and N,N-dimethylacetamide (7ml), and then add 4-dimethylaminopyridine (3mg), 5(6)-carboxynaphthylfluorescein (3mg) and dicyclohexylcarbodiimide (0.5mg) in sequence; stir the reaction under ice bath conditions, and dialyze and filter the reaction product for 72h to obtain a 7%wt modified cellulose acetate polymer solution, named spinning solution 1, for later use;

[0092] (2) Using N,N-dimethylformamide and tetrahydrofuran (1:1) as solvents, add 10wt% polycaprolactone (1.4g) and stir to dissolve. Then add 5wt% hydrophobic drug roxithromycin (0.75g) to obtain drug-loaded polycaprolactone spinning solution, named spinning solution 2.

[0093] (3) Using anhydrous ethanol as a solvent, a 15wt% acrylic resin spinning solution was prepared and named spinning solution 3.

[0094] (4) Spray the spinning solution 1 evenly onto the baking paper with a spray gun, let it dry, and set it aside. The spraying distance is 15cm and the air pressure is 3kpa.

[0095] (5) Coaxial electrospinning was performed on spinning solution 2 and spinning solution 3, and the nanofiber membrane was collected by baking paper in step (4). After spinning for 4 hours, the nanofiber membrane was peeled off from the baking paper to obtain a two-layer composite nanofiber membrane. The core layer is polycaprolactone and the shell layer is acrylic resin. The electrospinning parameters are: needle 21G, distance 10cm, voltage 18kv, temperature 30℃, humidity 40%, and the spinning speeds of the core layer and the shell layer are 0.0007mm / s and 0.0007mm / s, respectively.

[0096] (6) Weigh 60mg of the composite nanofiber membrane obtained in step (5), put it into a centrifuge tube containing 45ml of PBS (pH4.5) solution, release the drug on a shaker at 37 degrees Celsius, and periodically take 3ml of the solution to measure the ultraviolet light.

[0097] (7) Cut a 1cm*1cm nanofiber membrane obtained from (5), place it in PBS with pH 4.5 / 5 / 5.5 / 6 / 6.5 / 7 / 7.4 / 8 to observe the color change of the membrane, and use a fluorescence spectrophotometer to test the change in fluorescence intensity.

[0098] Comparative Examples 2-4

[0099] (1) Add cellulose acetate (0.7g) to a mixed solvent containing dichloromethane (3ml) and N,N-dimethylacetamide (7ml) to obtain a 7% wt cellulose acetate polymer solution, named spinning solution 1, for later use;

[0100] (2) Using N,N-dimethylformamide and tetrahydrofuran (1:1) as solvents, add 10wt% polycaprolactone (1.4g) and stir to dissolve. Then add 5wt% hydrophobic drug roxithromycin (0.75g) to obtain drug-loaded polycaprolactone spinning solution, named spinning solution 2.

[0101] (3) Using anhydrous ethanol as a solvent, a 15wt% acrylic resin spinning solution was prepared and named spinning solution 3.

[0102] (4) Spray the spinning solution 1 evenly onto the baking paper with a spray gun, let it dry, and set it aside. The spraying distance is 15cm and the air pressure is 3kpa.

[0103] (5) Coaxial electrospinning was performed on spinning solution 2 and spinning solution 3, and the nanofiber membrane was peeled off from the baking paper in step (4) after spinning for 4 hours to obtain a two-layer composite nanofiber membrane. The core layer is polycaprolactone and the shell layer is acrylic resin. The electrospinning parameters are: needle 21G, distance 10cm, voltage 18kv, temperature 30℃, humidity 40%, and the spinning speeds of the core layer and the shell layer are 0.0007mm / s and 0.0042mm / s, respectively.

[0104] (6) Weigh 60mg of the composite nanofiber membrane obtained in step (5), put it into a centrifuge tube containing 45ml of PBS (pH 4.5) solution, release the drug on a shaker at 37 degrees Celsius, and periodically take 3ml of the solution to measure the ultraviolet light.

[0105] (7) Cut a 1cm*1cm nanofiber membrane obtained from (5), place it in PBS with pH 4.5 / 5 / 5.5 / 6 / 6.5 / 7 / 7.4 / 8 to observe the color change of the membrane, and use a fluorescence spectrophotometer to test the change in fluorescence intensity.

[0106] Performance testing of nanofiber wound dressings for visualized diagnosis and controlled treatment

[0107] The fluorescence intensity of the nanofiber membrane for visual diagnosis was measured using a fluorescence spectrophotometer based on the change in pH; the absorbance of the nanofiber for controlled therapy was measured using a UV spectrophotometer based on the difference in drug release rate at different pH levels; the antibacterial rate of the dressing was determined by measuring its bactericidal rate against *E. coli* and *S. coli*. The results are as follows:

[0108]

[0109] The results above indicate that the type of antibacterial drug, the drug loading concentration, the core-shell ratio of the coaxial nanofibers, and whether or not fluorescein is grafted onto the nanofibers all have a certain impact on the performance of nanofiber wound dressings for visual diagnosis and controlled treatment. The hydrophilicity / hydrophobicity of the antibacterial drug affects the drug release rate (Examples 1 and 2); if the drug loading concentration is too low (Comparative Example 1-1 and Comparative Example 2-1), the nanofiber dressing will not have antibacterial activity; furthermore, if the shell of the coaxial nanofibers is too thin (Comparative Example 1-3 and Comparative Example 2-1), the drug release time will be shortened; if fluorescein is not grafted onto the nanofibers (Comparative Example 1-4 and Comparative Example 2-4), the nanofiber dressing will not have a monitoring function.

[0110] Unless otherwise specified, the raw materials and equipment used in this invention are all commonly used in the field; unless otherwise specified, the methods used in this invention are all conventional methods in the field.

[0111] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any way. Any simple modifications, alterations, and equivalent transformations made to the above embodiments based on the technical essence of the present invention shall still fall within the protection scope of the present invention.

Claims

1. A method for preparing a nanofiber membrane for visual diagnosis and differentiated drug delivery, characterized in that: Includes the following steps: (1) Add cellulose acetate to a reaction vessel containing a mixed solvent of dichloromethane and N,N-dimethylacetamide, and then add 4-dimethylaminopyridine, 5(6)-carboxynaphthylfluorescein and dicyclohexylcarbodiimide in sequence; stir the reaction under ice bath conditions, and dialyze the reaction product in a mixed solvent of dichloromethane and N,N-dimethylacetamide to obtain a modified cellulose acetate polymer solution, named spinning solution 1; the mass ratio of cellulose acetate, 5(6)-carboxynaphthylfluorescein, 4-dimethylaminopyridine and dicyclohexylcarbodiimide is (90-100):(1-5):(1-3):(0.5-1); (2) Dissolve the antibacterial agent and polycaprolactone in a mixed solvent of tetrahydrofuran and N,N-dimethylformamide to obtain spinning solution 2, wherein the concentration of polycaprolactone is 10-15 wt% and the concentration of the antibacterial agent is 5-10 wt%. (3) Dissolve acrylic resin in anhydrous ethanol to obtain spinning solution 3; (4) Spray the spinning solution 1 evenly onto the baking paper, let it dry, and set aside; (5) Coaxial electrospinning was performed with spinning solution 2 as the coaxial core layer and spinning solution 3 as the coaxial shell layer. The spinning speeds of the core layer and the shell layer were 0.0007 mm / s and 0.0042 mm / s, respectively. The baking paper obtained in step (4) is used as a receiving material. After spinning for 4 hours, the nanofiber membrane is peeled off from the baking paper to obtain a nanofiber membrane with a two-layer composite structure.

2. The preparation method according to claim 1, characterized in that: In step (1), the volume ratio of dichloromethane to N,N-dimethylacetamide is 2-3:6-7.

3. The preparation method according to claim 1, characterized in that: In step (1), the concentration of the spinning solution 1 is 5-10 wt%.

4. The preparation method according to claim 1, characterized in that, In step (2), the antibacterial drug is a hydrophilic drug and / or a lipophilic drug.

5. The preparation method according to claim 4, characterized in that, In step (2), the hydrophilic drug is tetracycline and / or oxytetracycline; the lipophilic drug is roxithromycin and / or linazone.

6. The preparation method according to claim 1, characterized in that: In step (3), the concentration of the spinning solution 3 is 15-20 wt%.

7. The preparation method according to claim 1, characterized in that: In step (4), the spraying distance is 10-20cm and the air pressure is 2-4kPa.

8. The preparation method according to claim 1, characterized in that: In step (5), the electrospinning parameters of the core layer and the shell layer are: needle 20-22G, distance 10-15cm, voltage 15-20kv, temperature 25-35℃, and humidity 30-50%.

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