Hyaluronic acid / polyethylene oxide electrostatic spinning fiber membrane as well as preparation method and application thereof

Abstract

The invention relates to a hyaluronic acid / polyethylene oxide electrostatic spinning fiber membrane as well as a preparation method and application thereof. The preparation method comprises the following steps: step 1, preparing a hyaluronic acid / polyethylene oxide mixed solution by using hyaluronic acid (HA) and polyethylene oxide (PEO) raw materials, trichloromethane serving as a solvent and cocamide serving as an activating agent (DEA); 2, magnetically stirring the hyaluronic acid / polyethylene oxide mixed solution, and standing to eliminate bubbles, so as to obtain a hyaluronic acid / polyethylene oxide spinning solution; and step 3, placing the hyaluronic acid / polyethylene oxide spinning solution in electrostatic spinning equipment to prepare the hyaluronic acid / polyethylene oxide electrostatic spinning fiber membrane. The hyaluronic acid / polyethylene oxide fiber membrane disclosed by the invention has good mechanical property, hydrophilicity and biocompatibility and huge potential of promoting wound healing, and is a promising wound dressing for clinical wound treatment.

Description

Technical Field

[0001] The invention relates to an electrostatic spinning fiber membrane and a preparation method and application thereof, and in particular to a hyaluronic acid / polyethylene oxide electrostatic spinning fiber membrane and a preparation method and application thereof. Background Art

[0002] Skin tissue defect has always been one of the common clinical diseases. Electrospinning nanofiber membrane simulates the structure similar to extracellular matrix (ECM) and has good application prospects in skin defect repair.

[0003] However, the spun fiber membranes in the prior art are poor in mechanical properties, hydrophilicity, biocompatibility and promotion of wound healing when used for skin defect repair, and need to be improved. Summary of the invention

[0004] The present invention designs a hyaluronic acid / polyethylene oxide electrospun fiber membrane and its preparation method and application, and solves the technical problem that the spun fiber membrane in the prior art is poor in mechanical properties, hydrophilicity, biocompatibility and promotion of wound healing when used for skin defect repair, and needs to be improved.

[0005] In order to solve the above-mentioned technical problems, the present invention adopts the following solutions: A preparation method of a hyaluronic acid / polyethylene oxide electrostatic spinning fiber membrane comprises the following steps: step 1, using hyaluronic acid (HA) and polyethylene oxide (PEO) raw materials, solvent chloroform (TM) and activator cocamide (DEA) to prepare a hyaluronic acid / polyethylene oxide mixed solution; step 2, magnetically stirring the hyaluronic acid / polyethylene oxide mixed solution and letting it stand to eliminate bubbles, so as to obtain a hyaluronic acid / polyethylene oxide spinning solution; step 3, placing the hyaluronic acid / polyethylene oxide spinning solution in an electrostatic spinning device to prepare a hyaluronic acid / polyethylene oxide electrostatic spinning fiber membrane.

[0006] As a natural polymer and the main component of skin ECM, HA is highly hydrophilic due to the presence of a large number of carboxyl and hydroxyl groups in its structure. Combined with the production and release of pro-inflammatory cytokines and interleukins and the promotion of fibrin clot formation, HA promotes the proliferation of fibroblasts / keratinocytes and the differentiation of fibroblasts into myofibroblasts. Therefore, HA has good hydrophilicity, biocompatibility, biodegradability and chemical modification ability, and has certain anti-inflammatory and antioxidant effects. However, due to the unclear protein structure, it is impossible to electroform a simple film. Therefore, polyethylene oxide (PEO) with high water solubility, hydrophilicity, high viscosity, biocompatibility with other bioactive substances, low toxicity, semi-crystalline, bioadhesive and adhesive properties is selected as the raw material. The polymer chain entanglement in HA is an important factor affecting its electrospinning properties. Polyethylene oxide (PEO) with relatively high molecular weight and limited crystal formation forms extensive chain entanglements with hyaluronic acid (HA). Cocamide (DEA) as an activator has good thickening effect, is easily soluble in water, has low irritation and is easily biodegradable, and is more conducive to the formation of Taylor cones by electrospinning. Chloroform (TM) is an organic compound with a density greater than water and is easily volatile, which helps electrospinning to form fiber membranes.

[0007] Preferably, in step 1, the mass ratio of hyaluronic acid (HA) to polyethylene oxide (PEO) is 1:1.

[0008] Preferably, the mass fraction of the hyaluronic acid / polyethylene oxide mixed solution in step 1 is 2-10%, that is: (HA+PEO) / (HA+PEO+DEA+TM)=2-10%.

[0009] Preferably, the magnetic stirring time in step 2 is ≥ 12 h.

[0010] Preferably, the hyaluronic acid / polyethylene oxide spinning solution is poured into a disposable 10 ml syringe, the spinning parameters are set as follows: the distance between the spinneret and the roller is 15 cm, the injection pump propulsion speed is 1 ml / h, the voltage is set to 20 kv, the speed is set to 300 r / min, and the lead foil is fixed on the roller to collect the spinning.

[0011] Preferably, the spinning time is 10 h at room temperature, and the prepared hyaluronic acid / polyethylene oxide electrospun fiber membrane is placed in an oven at 37° C. for drying for later use.

[0012] A hyaluronic acid / polyethylene oxide electrostatic spinning fiber membrane, characterized in that it is prepared using the above preparation method.

[0013] Application of the hyaluronic acid / polyethylene oxide electrospun fiber membrane prepared by the above preparation method in skin defect repair.

[0014] The hyaluronic acid / polyethylene oxide electrospinning fiber membrane and its preparation method and application have the following beneficial effects: The hyaluronic acid / polyethylene oxide fiber membrane of the present invention has good mechanical properties, hydrophilicity, biocompatibility and great potential for promoting wound healing, and is a promising wound dressing for clinical wound treatment. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 : Flow chart of preparation of hyaluronic acid / polyethylene oxide electrospinning fiber membrane of the present invention; Figure 2 : A schematic diagram of the structure of the present invention under an electron microscope; Figure 3 : Schematic diagram of the differences in water absorption, porosity and water vapor permeability between the HA / PEO fiber membrane of the present invention and the comparative material; Figure 4 : Schematic diagram of the healing of full-thickness skin defects constructed on the back of male SD rats in the present invention; Figure 5 : Comparison diagram of Masson staining results of in vivo experiments of the present invention. DETAILED DESCRIPTION

[0016] Combine the following Figures 1 to 5 , the present invention is further described: like Figure 1 As shown, hyaluronic acid HA and polyethylene oxide PEO were weighed in a ratio of 1:1 (0.2g PEO and 0.2g HA), chloroform was used as a solvent, and cocamide DEA activator was added, and they were placed in a 20 ml vial in turn to prepare a hyaluronic acid / polyethylene oxide mixed solution with a mass fraction of 2%. Magnetic stirring was performed for 12 hours, and the bubbles were eliminated by standing to obtain a hyaluronic acid / polyethylene oxide spinning solution. The hyaluronic acid / polyethylene oxide spinning solution was poured into a disposable 10 ml syringe, and the spinning parameters were set to a distance of 15 cm between the spinneret and the roller, a propulsion speed of 1 ml / h for the injection pump, a voltage of 20 kv, and a speed of 300 r / min. The lead foil was fixed on the roller to collect the spinning. The spinning time was 10 hours at room temperature, and the prepared spinning membrane was placed in an oven at 37 ° C for drying for standby use.

[0017] like Figure 2 It can be seen that thinner fibers appear in the HA / PEO fiber membrane, and there is no bead-like structure, showing a dense network structure with an average diameter of 280.7±10.24nm. The average diameter of polyethylene oxide PEO is about 480nm.

[0018] Figure 2a is the scanning electron microscope image of PEO and HA / PEO, Figure 2 b is the distribution frequency of the diameter of 50 PEO fibers randomly selected. Figure 2 c is the distribution frequency of the diameters of 50 randomly selected HA / PEO fibers, Figure 2 d is the statistical analysis of the diameter size of HA / PEO and PEO. The diameter of electrospun fibers is mainly in the range of 20-1000 nm. Fibers with smaller diameters increase the surface roughness and the fraction of the droplet contact area with the air trapped between the fibers, which can produce high-surface-area membranes and increase the synthesis of collagen and proteoglycans in fibroblasts. The smaller the fiber diameter, the larger the surface area (for fibers of the same unit mass, the smaller the fiber diameter, the more fibers there are than when the diameter is large, so the sum of the surface areas of all fibers with smaller fiber diameters is greater than the sum of the surface areas of fibers with larger diameters but fewer numbers), the better the tensile properties, and the better the proliferation of dermal fibroblasts. HA / PEO fiber membranes contain finer fibers than pure PEO fiber membranes, and there is no beaded structure presenting a dense mesh structure. At the same time, due to its high porosity and water vapor permeability, it can promote gas-liquid exchange of tissue cells and avoid excessive dehydration and drying of wounds.

[0019] as follows Figure 3 It can be seen that in vitro characterization shows that the material has good water absorption, porosity and water vapor permeability.

[0020] Specifically, a is the dry weight and wet weight of HA / PEO fiber membrane and gauze before and after soaking in PBS for 24 hours, b is the water absorption analysis; c is the dry weight and wet weight of HA / PEO fiber membrane, PEO fiber membrane and gauze before and after soaking in 95% anhydrous ethanol, d is the statistical analysis of the porosity of HA / PEO fiber membrane, PEO fiber membrane and gauze; e is the dry weight and wet weight of HA / PEO fiber membrane, PEO fiber membrane and gauze before and after soaking in physiological saline, f is the statistical analysis of the water vapor permeability of HA / PEO fiber membrane, PEO fiber membrane and gauze. From b, d and f, it can be seen that HA / PEO fiber membrane has the best water absorption, porosity and water vapor permeability.

[0021] In vivo experiments, we used male SD rats weighing 200-250g to construct a full-thickness skin defect model on their backs. HE staining showed that the material was the best wound healing dressing, as follows Figure 4 It can be seen that on the 7th day, the wound healing rate of the HA / PEO group was 61.89%, and on the 14th day, the wound healing rate of the HA / PEO group was 90.78%. On the 21st day, the wound was close to closure, and the new skin and hair were close to normal, which helped to accelerate wound healing, showing better epithelialization, and the number of sebaceous glands and hair follicles was the largest.

[0022] The results of Masson staining in the in vivo experiment are as follows Figure 5It can be seen that the collagen area of ​​the HA / PEO group of the material is the largest (30.12±3.2%), and more obvious collagen deposition and wavy collagen fibers can be seen.

[0023] The present invention adopts electrospinning technology to prepare a soft, thin and breathable fiber membrane wound dressing with hyaluronic acid and polyethylene oxide as substrates for the treatment of skin defect repair. The average diameter was measured by scanning electron microscopy to be 280.7±10.24nm, and the absorbance, hydrophilicity, porosity, water vapor permeability, etc. of the fiber membrane were studied. The wound healing and histological examination were evaluated by in vivo experiments. The hyaluronic acid / polyethylene oxide (HA / PEO) combination showed a good wound healing rate of 90.78% and the thickest connective tissue thickness of 1498.20±234.44μm on the 14th day, and showed the most collagen area deposition (30.12±3.2%). All the results show that the hyaluronic acid / polyethylene oxide fiber membrane has good mechanical properties, hydrophilicity, biocompatibility and great potential for promoting wound healing.

[0024] The present invention is described above by way of example in conjunction with the accompanying drawings. It is obvious that the implementation of the present invention is not limited to the above-mentioned method. As long as various improvements are made using the method concept and technical solution of the present invention, or the concept and technical solution of the present invention are directly applied to other occasions without improvement, they are all within the protection scope of the present invention.

Claims

1. A method for preparing a hyaluronic acid / polyethylene oxide electrospinning fiber membrane, comprising the following steps: Step 1: Prepare a hyaluronic acid / polyethylene oxide mixed solution using hyaluronic acid (HA) and polyethylene oxide (PEO) raw materials, solvent chloroform (TM) and activator cocamide (DEA); Step 2, magnetically stirring the hyaluronic acid / polyethylene oxide mixed solution and then standing it to eliminate bubbles, thereby obtaining a hyaluronic acid / polyethylene oxide spinning solution; Step 3: Place the hyaluronic acid / polyethylene oxide spinning solution in an electrospinning device to prepare a hyaluronic acid / polyethylene oxide electrospinning fiber membrane.

2. The method for preparing the hyaluronic acid / polyethylene oxide electrospinning fiber membrane according to claim 1, characterized in that: In step 1, the mass ratio of hyaluronic acid (HA) to polyethylene oxide (PEO) is 1:

1.

3. The method for preparing the hyaluronic acid / polyethylene oxide electrospinning fiber membrane according to claim 2, characterized in that: In step 1, the mass fraction of the hyaluronic acid / polyethylene oxide mixed solution is 2-10%, that is: (HA+PEO) / (HA+PEO+DEA+TM)=2-10%.

4. The method for preparing the hyaluronic acid / polyethylene oxide electrospinning fiber membrane according to claim 1, characterized in that: The magnetic stirring time in step 2 is ≥ 12 h.

5. The method for preparing the hyaluronic acid / polyethylene oxide electrospinning fiber membrane according to claim 1, characterized in that: Pour the hyaluronic acid / polyethylene oxide spinning solution into a disposable 10 ml syringe, set the spinning parameters as follows: the distance between the spinneret and the roller is 15 cm, the injection pump propulsion speed is 1 ml / h, the voltage is set to 20 kv, the speed is set to 300 r / min, and fix the lead foil on the roller to collect the spinning.

6. The method for preparing the hyaluronic acid / polyethylene oxide electrospinning fiber membrane according to claim 1, characterized in that: The spinning time was 10 h at room temperature, and the prepared hyaluronic acid / polyethylene oxide electrospun fiber membrane was placed in an oven at 37°C for drying.

7. A hyaluronic acid / polyethylene oxide electrospinning fiber membrane, characterized in that: It is prepared by the preparation method described in any one of claims 1 to 6.

8. Use of the hyaluronic acid / polyethylene oxide electrospun fiber membrane prepared by the preparation method according to any one of claims 1 to 6 in repairing skin defects.

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