Preparation method of a double-layer structure biomimetic analgesic dressing

Abstract

The application relates to the technical field of biomedical materials, in particular to a preparation method of a double-layer structure biomimetic analgesic dressing, which comprises the following steps: step 1, a PCL / SF / GT electrospinning fiber membrane with an arrangement structure is prepared by adopting an electrospinning technology; step 2, oxidized sodium alginate is synthesized by reacting sodium periodate with sodium alginate, and then by dialysis and freeze-drying; step 3, a pain-relieving drug solution is dispersed in a Tris-HCl buffer solution, then hydrochloric acid dopamine is added, and a polymer precipitate of the polydopamine wrapped drug is obtained by centrifugation; step 4, a hydrogel is prepared; and step 5, a double-layer structure biomimetic composite dressing with the lower layer being the fiber membrane and the upper layer being the hydrogel is prepared by adopting a physical layer-by-layer stacking mode. The double-layer structure biomimetic analgesic dressing prepared by the application has excellent pain-relieving, hemostatic, antibacterial, tissue growth promoting and wound healing properties.

Description

Technical Field

[0001] This invention relates to the field of biomedical materials technology, and in particular to a method for preparing a double-layer biomimetic analgesic dressing. Background Technology

[0002] Medical dressings are indispensable basic consumables in the healthcare system, widely used in wound care, surgical protection, infection control, postoperative rehabilitation, and other scenarios to provide patients with basic medical care. Driven by the urgent clinical need for efficient management solutions for chronic, refractory wounds, wounds such as diabetic ulcers, pressure ulcers, and large-area burns have complex and lengthy healing processes. Patients not only suffer from continuous pain but also face the risks of high wound exudation and susceptibility to infection, which significantly reduces their quality of life and imposes a heavy social and medical burden. However, traditional dressings are too singular in function, failing to achieve synergy and balance in multiple key aspects such as moisturizing, antibacterial properties, analgesia, and promoting healing. Therefore, developing a novel functional dressing that can actively regulate the wound microenvironment and integrates efficient barrier function, continuous analgesia, and tissue regeneration promotion has extremely important clinical value and social significance.

[0003] Electrospinning technology can produce synthetic nanofibers that structurally mimic the extracellular matrix (ECM). These electrospun nanofibers, due to their high specific surface area and high porosity, offer new possibilities for tissue regeneration, making them an ideal choice for preparing wound dressings that meet biomedical requirements. Hydrogels are three-dimensional network structures formed by cross-linking hydrophilic polymers containing numerous polar functional groups. Their unique three-dimensional network can swell fully in water without dissolving, maintaining its initial morphology. Simultaneously, this structure possesses the ability to load and control drug release, exhibits excellent biocompatibility, and can even achieve biocompatibility similar to that of living tissues, without affecting normal human metabolic processes.

[0004] Electrospun nanofibers possess high specific surface area and porous structure, which are beneficial for cell adhesion, proliferation, migration, and differentiation. Hydrogels, due to their high water content, high porosity, softness, and high compatibility with human skin tissue, are ideal medical dressing materials. Therefore, combining electrospun nanofibers with hydrogels not only improves the mechanical properties of hydrogels but also forms a multilayered composite structure similar to biological tissue, thereby endowing the composite material with new biological functions and demonstrating greater advantages in wound healing. Currently, physically combining electrospun nanofibers and hydrogels to form composite materials is the most convenient and representative method. By stacking nanofibers and hydrogel plates layer by layer, integrated composite materials with layered structures can be obtained.

[0005] In conclusion, the preparation of biomimetic skin-like dressings using natural polymer materials shows great promise. By combining electrospun nanofibers with hydrogels and introducing drugs, dressings with multiple functions such as pain relief, hemostasis, antibacterial properties, and cell proliferation promotion can be prepared, providing new ideas for clinical wound treatment. Summary of the Invention

[0006] The purpose of this invention is to address the shortcomings of existing technologies by proposing a method for preparing a double-layer biomimetic analgesic dressing. This method simplifies the steps, uses milder conditions, has better biocompatibility, and exhibits superior overall performance, making it beneficial for clinical application.

[0007] To achieve the above objectives, the present invention adopts the following technical solution: a method for preparing a double-layer biomimetic analgesic dressing, comprising the following steps:

[0008] Step 1: Polycaprolactone (PCL), silk fibroin (SF), and gelatin (GT) were blended and dissolved in hexafluoroisopropanol (HFIP). After stirring at room temperature for 12 h, the mixture was spun at room temperature and dried under vacuum to prepare PCL / SF / GT electrospun fiber membranes with different orientations.

[0009] Step 2: Dissolve sodium alginate (SA) in deionized water, and slowly add sodium periodate (NaIO4) dissolved in deionized water to the SA solution dropwise. Stir the reaction under light-protected conditions. Finally, add ethylene glycol and stir to terminate the oxidation reaction. Place the solution in a dialysis bag and dialyze with deionized water for 3 days. After freeze-drying, obtain a white flocculent product of oxidized sodium alginate (OSA).

[0010] Step 3: First, dissolve the pain-relieving drug in DMSO solution, then add it to Tris-HCl buffer and stir to disperse. Add dopamine hydrochloride and adjust the pH to 8.5-9.5 at room temperature, and then magnetically stir to react. After centrifugation, the solution after reaction is obtained as a polymer precipitate of polydopamine-encapsulated drug (PDA@Drug).

[0011] Step 4: Dissolve sodium oxidized alginate (OSA), carboxymethyl chitosan (CMC), and polydopamine-encapsulated drug (PDA@Drug) in deionized water to obtain OSA solution, CMC solution, and PDA@Drug solution, respectively. Then, mix them uniformly in a certain volume ratio to prepare a hydrogel.

[0012] Step 5: Directly cover the surface of the hydrogel prepared in Step 4 with the nanofiber membrane spun in Step 1. Prepare a biomimetic composite dressing with a bilayer structure of electrospun fiber membrane / hydrogel, with the lower layer being a fiber membrane and the upper layer being a hydrogel, by physically stacking the layers one by one.

[0013] Preferably, in step 1, the PCL, SF and GT are blended and dissolved in HFIP at a mass ratio of 5:3:2-8:1:1 to obtain a spinning solution with a total mass fraction of 10-20 wt%, and the roller speed is 250-2500 rpm, thereby obtaining fiber membranes with different orientations (oriented arrangement (A) and random arrangement (R)).

[0014] Preferably, in step 1, the spinning needle size is 23 G, the spinning speed is 1-2 mL / h, the spinning voltage is 15-20 KV, the receiving distance is 10-20 cm, and the spinning time is 2-8 h.

[0015] Preferably, in step 2, the concentration of the SA solution is 0.5-2 wt%, the concentration of the NaIO4 solution is 0.5 mol / L, the reaction time is 24-48 h, 3 mL of ethylene glycol is added to terminate the oxidation reaction, the termination time is 0.5-2 h, and the dialysis bag is a molecular weight cutoff of 14000 Da.

[0016] Preferably, in step 3, the analgesic drug is first dissolved in DMSO solution at a concentration of 1-2 mg / mL, and then diluted to a concentration of 0.5 × 10⁻⁶ mg / mL. -6 - 1.5×10 -6 Solution M reacts with dopamine hydrochloride.

[0017] Preferably, in step 3, the pH is 8.5-9.5, the magnetic stirring reaction time is 24-48 h, the centrifuge speed is 5000-7000 rpm / min, and the centrifugation time is 5-15 min.

[0018] Preferably, in step 4, the concentrations of the OSA solution and CMC solution are 1-3 wt%, and the concentration of the PDA@Drug solution is 0.5-2 wt%.

[0019] Preferably, in step 4, the OSA solution, CMC solution and PDA@Drug solution are uniformly mixed at a volume ratio of 2:2:(0~1) to prepare a hydrogel.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

[0021] 1. The double-layer structure biomimetic pain-relieving medical dressing prepared by the present invention combines the advantages of electrospinning and hydrogel technology, which can simulate the ECM structure and the characteristics of the raw materials used are important components of the ECM. It can simulate biomimetic human skin and has practical application value in wound healing.

[0022] 2. This invention uses electrospinning technology to prepare the lower nanofiber membrane, which largely avoids the potential toxicity of chemical reagents. It retains the original mechanical properties of PCL and provides better hydrophilicity and cell affinity. The fibers can continuously release GT protein molecules, creating a favorable environment for cell attachment and proliferation.

[0023] 3. The present invention uses hydrogel technology to prepare the upper drug-loaded hydrogel layer without the need to add crosslinking agents. It has certain rheological properties, adhesion properties, photothermal properties and biocompatibility properties, and can also release drugs under near-infrared response.

[0024] 4. The processing method of this invention is simple and effective, and can be mass-produced. Attached Figure Description

[0025] Figure 1 The cross-sectional scanning electron microscope images of Embodiment 2 of the present invention are shown in (a) SEM image at 100x magnification; (b) SEM image at 150x magnification.

[0026] Figure 2 To demonstrate the hemostatic performance of Comparative Example 1, Example 1, Example 2, and Example 3 of the present invention ((a) total blood loss from mouse liver over 60 s; (b) photographs of liver hemorrhage over 60 s).

[0027] Figure 3 The antioxidant properties of Examples 1, 2, and 3 of this invention are shown in (a) DPPH free radical scavenging rate; (b) DPPH solution photograph.

[0028] Figure 4 The cell proliferation performance of Comparative Example 1, Example 1, Example 2, and Example 3 of the present invention is shown in ((a) OD value of L929 at 450 nm; (b) live / dead staining diagram of L929 proliferation).

[0029] Figure 5 The wound healing performance of Comparative Example 1, Example 1, Example 2, and Example 3 of the present invention is shown in (a) Wound healing effect diagram; (b) Wound size).

[0030] Figure 6 This is a schematic diagram showing the incidence of pain response in Comparative Example 1, Example 1, Example 2, and Example 3 of the present invention. Detailed Implementation

[0031] The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings, so that those skilled in the art can better understand the advantages and features of the present invention, thereby making a clearer definition of the scope of protection of the present invention. The embodiments described in this invention are only some embodiments of the present invention, not all embodiments. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention.

[0032] Example 1:

[0033] A method for preparing a double-layer biomimetic analgesic dressing includes the following steps:

[0034] Step 1: Polycaprolactone (PCL), silk fibroin (SF), and gelatin (GT) were blended and dissolved in hexafluoroisopropanol (HFIP). After stirring at room temperature for 12 h, a spinning solution with a total mass fraction of 20 wt% was obtained. Spinning was performed at room temperature with a spinning speed of 1 mL / h, a spinning voltage of 17 kV, a receiving distance of 15 cm, a spinning time of 5 h, and a roller speed of 2500 rpm. After vacuum drying, an oriented PCL / SF / GT electrospun fiber membrane was prepared.

[0035] Step 2: Dissolve sodium alginate (SA) in deionized water to a concentration of 1 wt%, and slowly add 0.5 mol / L sodium periodate (NaIO4) dissolved in deionized water to the SA solution dropwise. Stir and react for 48 h under light-protected conditions. Finally, add 3 mL of ethylene glycol and stir for 1 h to terminate the oxidation reaction. Place the solution in a 14000 Da dialysis bag and dialyze with deionized water for 3 days. After freeze-drying, obtain a white flocculent product of oxidized sodium alginate (OSA).

[0036] Step 3: Dissolve sodium oxidized alginate (OSA) and carboxymethyl chitosan (CMC) in deionized water to obtain 2 wt% OSA solution and 2 wt% CMC solution respectively. Then, mix them uniformly at a volume ratio of 2:2 to prepare OSA / CMC hydrogel.

[0037] Step 4: Directly cover the surface of the hydrogel prepared in Step 4 with the nanofiber membrane spun in Step 1. Prepare a biomimetic composite dressing with a bilayer structure of electrospun fiber membrane / hydrogel, with the lower layer being a fiber membrane and the upper layer being a hydrogel, by physically stacking the layers one by one.

[0038] Example 2:

[0039] A method for preparing a double-layer biomimetic analgesic dressing includes the following steps:

[0040] Step 1: Polycaprolactone (PCL), silk fibroin (SF), and gelatin (GT) were blended and dissolved in hexafluoroisopropanol (HFIP). After stirring at room temperature for 12 h, a spinning solution with a total mass fraction of 20 wt% was obtained. Spinning was performed at room temperature with a spinning speed of 1 mL / h, a spinning voltage of 17 kV, a receiving distance of 15 cm, a spinning time of 5 h, and a roller speed of 2500 rpm. After vacuum drying, an oriented PCL / SF / GT electrospun fiber membrane was prepared.

[0041] Step 2: Dissolve sodium alginate (SA) in deionized water to a concentration of 1 wt%, and slowly add 0.5 mol / L sodium periodate (NaIO4) dissolved in deionized water to the SA solution dropwise. Stir and react for 48 h under light-protected conditions. Finally, add 3 mL of ethylene glycol and stir for 1 h to terminate the oxidation reaction. Place the solution in a 14000 Da dialysis bag and dialyze with deionized water for 3 days. After freeze-drying, obtain a white flocculent product of oxidized sodium alginate (OSA).

[0042] Step 3: First, dissolve the pain reliever (drug) at a concentration of 2 mg / mL in DMSO solution, then add 300 mL of Tris-HCl buffer to dilute to a concentration of 1×10⁻⁶. -6 After stirring the M solution for 15 min, dopamine hydrochloride was added, and the pH was adjusted to 8.5 at room temperature. The solution was then magnetically stirred for 48 h. The resulting solution was then centrifuged at 7000 rpm for 10 min to obtain a polymer precipitate of polydopamine-encapsulated drug (PDA@Drug).

[0043] Step 4: Dissolve sodium oxidized alginate (OSA), carboxymethyl chitosan (CMC), and polydopamine-encapsulated drug (PDA@Drug) in deionized water to obtain 2 wt% OSA solution, 2 wt% CMC solution, and 1 wt% PDA@Drug solution, respectively. Then, mix them uniformly at a volume ratio of 2:2:1 to prepare OSA / CMC / PDA@Drug hydrogel.

[0044] Step 5: Directly cover the surface of the hydrogel prepared in Step 4 with the nanofiber membrane spun in Step 1. Prepare a biomimetic composite dressing with a bilayer structure of electrospun fiber membrane / hydrogel, with the lower layer being a fiber membrane and the upper layer being a hydrogel, by physically stacking the layers one by one.

[0045] Example 3:

[0046] A method for preparing a double-layer biomimetic analgesic dressing includes the following steps:

[0047] Step 1: Polycaprolactone (PCL), silk fibroin (SF), and gelatin (GT) were blended and dissolved in hexafluoroisopropanol (HFIP). After stirring at room temperature for 12 h, a spinning solution with a total mass fraction of 20 wt% was obtained. Spinning was performed at room temperature with a spinning speed of 1 mL / h, a spinning voltage of 17 kV, a receiving distance of 15 cm, a spinning time of 5 h, and a roller speed of 250 rpm. After vacuum drying, an oriented PCL / SF / GT electrospun fiber membrane was prepared.

[0048] Step 2: Dissolve sodium alginate (SA) in deionized water to a concentration of 1 wt%, and slowly add 0.5 mol / L sodium periodate (NaIO4) dissolved in deionized water to the SA solution dropwise. Stir and react for 48 h under light-protected conditions. Finally, add 3 mL of ethylene glycol and stir for 1 h to terminate the oxidation reaction. Place the solution in a 14000 Da dialysis bag and dialyze with deionized water for 3 days. After freeze-drying, obtain a white flocculent product of oxidized sodium alginate (OSA).

[0049] Step 3: First, dissolve the pain reliever (drug) at a concentration of 2 mg / mL in DMSO solution, then add 300 mL of Tris-HCl buffer to dilute to a concentration of 1×10⁻⁶. -6 After stirring the M solution for 15 min, dopamine hydrochloride was added, and the pH was adjusted to 8.5 at room temperature. The solution was then magnetically stirred for 48 h. The resulting solution was then centrifuged at 7000 rpm for 10 min to obtain a polymer precipitate of polydopamine-encapsulated drug (PDA@Drug).

[0050] Step 4: Dissolve sodium oxidized alginate (OSA), carboxymethyl chitosan (CMC), and polydopamine-encapsulated drug (PDA@Drug) in deionized water to obtain 2 wt% OSA solution, 2 wt% CMC solution, and 1 wt% PDA@Drug solution, respectively. Then, mix them uniformly at a volume ratio of 2:2:1 to prepare OSA / CMC / PDA@Drug hydrogel.

[0051] Step 5: Directly cover the surface of the hydrogel prepared in Step 4 with the nanofiber membrane spun in Step 1. Prepare a biomimetic composite dressing with a bilayer structure of electrospun fiber membrane / hydrogel, with the lower layer being a fiber membrane and the upper layer being a hydrogel, by physically stacking the layers one by one.

[0052] Comparative Example 1

[0053] Using no sample treatment as Control Example 1, subsequent tests were conducted.

[0054] Test Experiment

[0055] 1. Hemostatic Performance Experiment: The in vivo hemostatic performance of the composite dressing was tested using an 8-week-old ICR male mouse model of liver bleeding. Mice were anesthetized by intraperitoneal injection of 2 wt% tribromoethanol (0.35 mL / 20 g), abdominal hair was removed, and the liver was exposed through a skin incision at the corresponding location on the abdomen. Surrounding tissue fluid was carefully wiped away. The liver was then placed on pre-weighed quantitative filter paper, and the liver was punctured with a needle to initiate bleeding. Different samples from each group were applied to the bleeding points on the liver; the control group received no treatment after puncture. Real-time photographs of liver bleeding were taken at 5, 10, 15, 30, and 60 seconds. After 1 minute, the weight of the blood-stained filter paper was measured to calculate the final total blood loss, thus evaluating the hemostatic performance of the sample.

[0056] 2. Antioxidant Performance Experiment: The antioxidant performance of the samples was evaluated by measuring their ability to eliminate 1,1-diphenyl-2-trinitrophenylhydrazine (DPPH) free radicals. A 0.1 mM DPPH solution was prepared using ethanol as a solvent in the dark. The DPPH solution served as the negative control group, while ascorbic acid (VC), which has excellent antioxidant properties, was added to the DPPH solution as the positive control group. Different samples were added to the DPPH solution as example groups. After standing, the supernatant was collected, and the absorption peaks in the 450-650 nm range and the absorbance at 517 nm of each group of samples were detected using a UV spectrophotometer. The DPPH free radical scavenging rate was calculated.

[0057] 3. Antibacterial Performance Test: The antibacterial performance of the samples was tested according to the shaking method in the national standard GB / T20944.3-2008. The test bacteria were Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli. All samples were sterilized with ultraviolet light for 30 min beforehand, with the bacterial suspension of the blank sample serving as the control group. First, the bacterial suspension was incubated at 37℃ for 24 h and then inoculated. The bacterial suspension was then diluted and inoculated onto 0.1 g of different composite dressings, and incubated with shaking in a shaker at 24℃ for 18 h. Finally, the bacterial suspension, after being fully incubated with the samples, was diluted to a gradient concentration, and 50 μL was aspirated each time and dispersed in droplets in agar-filled petri dishes, and incubated at 37℃ for 24 h. After incubation, the number of colonies per drop of bacterial suspension on each petri dish was photographed and counted, and the antibacterial rate of the sample was calculated.

[0058] 4. Cell proliferation assay: L929 cells were seeded at a density of 3 × 10³ cells / well in 24-well plates (500 μl per well) and cultured at 37°C for 24 h. Cell proliferation was assessed on days 1, 3, and 5 after seeding with the cell and sample extracts. The culture medium was aspirated from the wells, and the cells were rinsed three times with PBS buffer. Then, 100 μL of CCK-8 assay solution was added to each well, and the plates were incubated at 37°C for 2 h. Finally, the absorbance at 450 nm was measured using a microplate reader. Alternatively, cells and sample extracts were seeded together in wells and cultured for a fixed time, followed by rinsing three times with PBS buffer. Live / dead staining working solution was prepared according to the Calcein / PI assay kit instructions and added to each well for staining. The wells were incubated in the dark for 30 min, and the cell status was observed using a fluorescence microscope.

[0059] 5. Wound Healing Performance Experiment: First, a mouse burn model was established: Eight-week-old male ICR mice were used as experimental animals. They were anesthetized by an abdominal injection of 2 wt% tribromoethanol at a dose of 0.35 mL / 20 g, and their back hair was shaved. A preheated copper platform (10 mm in diameter) was used to vertically press on the rat's back for approximately 30 seconds to establish a deep burn wound. The wound was left in place for 24 hours after the burn, and the formed eschar tissue was removed. Subsequently, samples that had been pre-sterilized by ultraviolet light were applied to the wound. The dressings were changed every 3 days, with at least 5 mice in each group. Wound changes were observed and recorded on days 1, 4, 7, 10, and 15 after the burn. The wound area was measured using ImageJ software, and the wound healing rate was calculated to evaluate the wound healing effect.

[0060] 6. Pain Response Incidence Experiment: This experiment employed a modified paradigm of the mechanical withdrawal reflex threshold test to specifically assess the mechanosensitive response of burned mice on the back of the injured skin. A Von-Frey fiber optic analgesia device was used, employing a mechanical stimulation needle with a stimulation intensity of 0.04 g / pair, applied vertically to the injured skin area of ​​the mouse. The mechanical stimulation was repeated 10 times, with each stimulation lasting 1-2 seconds and an interval of at least 5 seconds between stimulations to avoid adaptation. The number of times the mice exhibited clearly pain-related behaviors (i.e., evasion of the stimulus) under stimulation was observed and recorded. The percentage of pain responses at each intensity out of the total number of stimulations (pain percentage) was calculated. A higher pain percentage indicates a stronger pain response, and vice versa.

[0061] Test Results Explanation

[0062] 1. Hemostatic performance test: such as Figure 2As shown, compared with the comparative example, all three groups of examples exhibited significant acute hemostatic effects. After 60 seconds of bleeding, the total blood loss in the livers of mice treated with each example was significantly reduced to 0.078 g, 0.078 g, and 0.077 g, respectively, with no significant difference among the three groups. The experimental results indicate that the hemostatic function of the composite dressing mainly depends on the hydrogel layer; the hydrogel can rapidly absorb blood and effectively cover the bleeding site through its physical sealing effect.

[0063] Table 1 Results of hemostatic performance test

[0064]

[0065] 2. Antioxidant performance test: such as Figure 3 As shown, compared with the negative control, the DPPH radical scavenging rate of Examples 2 and 3 was higher than 70%, similar to that of the positive control, and the solution was reduced to a pale yellow color, indicating that the introduction of PDA@Drug endowed Examples 2 and 3 with excellent antioxidant activity.

[0066] Table 2 Antioxidant performance test results

[0067]

[0068] 3. Antibacterial Performance Experiment: As shown in Table 3, all three sets of examples exhibited excellent antibacterial performance, with inhibition rates of over 95% against *S. aureus* and *E. coli*. Compared with the comparative examples, Example 1 showed a weaker antibacterial effect, while Examples 2 and 3 both demonstrated more prominent antibacterial effects, indicating that PDA@Drug can enhance the antibacterial performance of the examples.

[0069] Table 3 Antibacterial performance test results

[0070] Example 1 Example 2 Example 3 S. aureus antibacterial rate (%) 99.97 99.99 99.99 E. coli inhibition rate (%) 95.86 99.47 99.57

[0071] 4. Cell proliferation performance experiments: such as Figure 4 As shown, the OD value of L929 cells continued to increase with the increase of culture time, and the cell viability of each group was above 94% until day 5. Figure 4 (b) shows more clearly that L929 cells showed a steady growth trend as the number of days increased, indicating that the composite dressing not only has good cell compatibility, but can also promote cell proliferation.

[0072] Table 4 Results of cell proliferation performance test

[0073]

[0074] 5. Wound healing performance test: such as Figure 5As shown, compared with the control example, the wounds treated in the examples healed better, and all showed near-complete closure by day 14. Furthermore, comparing the wound shape on day 14, it can be seen that the wounds treated in Examples 1 and 2 were more regular and aesthetically pleasing than those treated in Example 3, indicating that the oriented fibrous structure can induce the direction of wound healing. The wound healing rate of the comparative example was 77.22%, while the wound healing rates of the examples were all above 90%. The wound size and healing rate of the comparative example were significantly different from those of the examples, indicating that the treatment using the examples can promote wound healing.

[0075] Table 5. Results of wound healing performance test

[0076]

[0077] 6. Pain response rate experiment: such as Figure 6 As shown, compared with the control example, the percentage of pain in Examples 2 and 3 was significantly lower, indicating that the treatment with Examples 2 and 3 can reduce the pain response.

[0078] In summary, this invention prepares an inner electrospun fiber membrane using electrospinning technology, and then prepares an outer drug-loaded hydrogel layer through a Schiff base reaction. The two layers are then physically combined to obtain a novel biocompatible biomimetic composite dressing with a bilayer structure. By combining the advantages of both electrospinning and hydrogel technologies, and introducing analgesic drugs that can be released in response to near-infrared radiation, this invention provides a new approach for developing a multifunctional dressing for clinical wound treatment, offering pain relief, hemostasis, antibacterial properties, and tissue growth promotion.

[0079] The descriptions and practices disclosed in this invention are readily apparent and understandable to those skilled in the art, and various modifications and refinements can be made without departing from the principles of this invention. Therefore, any modifications or improvements made without departing from the spirit of this invention should also be considered within the scope of protection of this invention.

Claims

1. A method for preparing a double-layer biomimetic analgesic dressing, characterized in that, Includes the following steps: Step 1: Polycaprolactone (PCL), silk fibroin (SF), and gelatin (GT) were blended and dissolved in hexafluoroisopropanol (HFIP). After stirring at room temperature for 12 h, the mixture was spun at room temperature and dried under vacuum to prepare electrospun PCL / SF / GT fiber membranes with different orientations. Step 2: Dissolve sodium alginate (SA) in deionized water, and slowly add sodium periodate (NaIO4) dissolved in deionized water to the SA solution dropwise. Stir the reaction under light-protected conditions. Finally, add ethylene glycol and stir to terminate the oxidation reaction. Place the solution in a dialysis bag and dialyze with deionized water for 3 days. After freeze-drying, obtain a white flocculent product of oxidized sodium alginate (OSA). Step 3: First, dissolve the pain-relieving drug Drug in DMSO solution, then add it to Tris-HCl buffer and stir to disperse. Add dopamine hydrochloride and adjust the pH to 8.5-9.5 at room temperature, and then magnetically stir the reaction. After centrifugation, the solution after reaction is obtained as a polymer precipitate of drug encapsulated by polydopamine (PDA@Drug). Step 4: Dissolve sodium alginate (OSA), carboxymethyl chitosan (CMC), and polydopamine-encapsulated drug PDA@Drug in deionized water to obtain OSA solution, CMC solution, and PDA@Drug solution, respectively. Then, mix them uniformly in a certain volume ratio to prepare a hydrogel. Step 5: Directly cover the surface of the hydrogel prepared in Step 4 with the nanofiber membrane spun in Step 1. Prepare a biomimetic composite dressing with a bilayer structure of electrospun fiber membrane / hydrogel, with the lower layer being a fiber membrane and the upper layer being a hydrogel, by physically stacking the layers one by one.

2. The method for preparing a double-layer biomimetic analgesic dressing according to claim 1, characterized in that, In step 1, PCL, SF and GT are blended and dissolved in HFIP at a mass ratio of 5:3:2-8:1:1 to obtain a spinning solution with a total mass fraction of 10-20 wt%. The roller speed is 250-2500 rpm, thereby obtaining fiber membranes with different orientations.

3. The method for preparing a double-layer biomimetic analgesic dressing according to claim 1, characterized in that, In step 1, the spinning needle size is 23 G, the spinning speed is 1-2 mL / h, the spinning voltage is 15-20 KV, the receiving distance is 10-20 cm, and the spinning time is 2-8 h.

4. The method for preparing a double-layer biomimetic analgesic dressing according to claim 1, characterized in that, In step 2, the concentration of the SA solution is 0.5-2 wt%, the concentration of the NaIO4 solution is 0.5 mol / L, the reaction time is 24-48 h, 3 mL of ethylene glycol is added to terminate the oxidation reaction, the termination time is 0.5-2 h, and the dialysis bag is a molecular weight cutoff of 14000 Da.

5. The method for preparing a double-layer biomimetic analgesic dressing according to claim 1, characterized in that, In step 3, the analgesic drug is first dissolved in DMSO solution at a concentration of 1-2 mg / mL, and then diluted to a concentration of 0.5 × 10⁻⁶ mg / mL. -6 - 1.5×10 -6 Solution M reacts with dopamine hydrochloride.

6. The method for preparing a double-layer biomimetic analgesic dressing according to claim 1, characterized in that, In step 3, the magnetic stirring reaction time is 24-48 h, the centrifuge speed is 5000-7000 rpm / min, and the centrifugation time is 5-15 min.

7. The method for preparing a double-layer biomimetic analgesic dressing according to claim 1, characterized in that, In step 4, the concentrations of the OSA solution and CMC solution are 1-3 wt%, and the concentration of the PDA@Drug solution is 0.5-2 wt%.

8. The method for preparing a double-layer biomimetic analgesic dressing according to claim 1, characterized in that, In step 4, the OSA solution, CMC solution and PDA@Drug solution are uniformly mixed at a volume ratio of 2:2:(0~1) to prepare a hydrogel.

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