CN104689365A - Compound biological dressing for accelerating wound healing and preparation method thereof - Google Patents

Abstract

The invention discloses a composite biological dressing for promoting wound healing and a preparation method thereof. The composite biological dressing is composed of medical gelatin with poor hydrophilicity and hyaluronic acid with strong hydrophilicity mixed in a certain proportion and added with cross It is made of combination agent and vacuum freeze-dried to make the dressing porous and breathable, so as to obtain the ideal water vapor transmission rate characteristics, which can fully absorb the exudate of the wound surface, avoid fluid accumulation, and maintain the best wound healing effect. A humid environment promotes mitosis of skin cells and increases the number of skin cells on the wound, thereby accelerating wound healing. The composite biological dressing of the present invention has the advantages of being extracted from animals and has the advantages of easy availability of raw materials, simple preparation process, degradability and convenient use, and is an ideal dressing for wound repair.

Description

A composite biological dressing for promoting wound healing and preparation method thereof

technical field

The invention belongs to the technical field of medical dressings, in particular to a degradable composite biological dressing for promoting skin wound healing and a preparation method thereof.

background field

Medical dressings after skin trauma can protect the wound surface and provide a good environment for wound repair. Commonly used clinical dressings, such as various gauzes and cotton pads, are low in cost and have a wide range of raw materials, but the surface of traditional dressings is rough and dry, and they are easy to rub and cause further damage to the wound. At the same time, the new granulation on the wound surface is easy to grow into the mesh of the dressing, and pulling the dressing when changing the dressing is likely to cause pain and cause bleeding to damage the wound surface. Traditional gauze has poor water absorption, and often needs to be replaced several times for patients with a lot of wound exudation, which increases the clinical workload. Especially with the approach of an aging society and the increasing number of diabetic patients, the repair of refractory wounds such as diabetic ulcers, venous ulcers, and decubitus ulcers has become a major problem for clinicians. Traditional dressings have been difficult to meet the requirements for promoting the healing of these wounds. Since the theory of moist wound healing was put forward, the effect of moist wound environment on promoting wound healing has been widely recognized and accepted.

As a good dressing, it must be able to provide the most suitable moist environment for wound growth, so as to promote the proliferation and differentiation of wound tissue cells, thereby promoting wound healing. Excessive moisture will lead to wound fluid, while insufficient wound surface moisture will lead to dryness and dehydration of the wound surface, which is not conducive to wound healing. Therefore, the ideal wound healing environment requires the degree of humidity to be within an appropriate range. An ideal wound dressing requires the ability to maintain the moisture of the wound surface at an optimal level. Previous experimental studies on wound dressings have shown that the water vapor transmission rate (WVTR) of a material can be used to judge its ability to absorb moisture and breathe. If the water vapor transmission rate If it is around 2000-2500g/m ² /day, it means that the material can keep the wound surface moist to an ideal state, and it is an ideal wound dressing. The water vapor of wound dressing mainly depends on the water absorption performance of the material and the air permeability pore size of the material. Through our research, we found that when medical gelatin and hyaluronic acid are coupled in a certain ratio and technical process, the best water vapor transmission rate can be achieved, and an ideal wound dressing can be produced.

Although both medical gelatin and hyaluronic acid have many biological advantages, neither medical gelatin nor hyaluronic acid alone can be the most ideal wound dressing. Medical gelatin is not hydrophilic enough to fully absorb the exudate of the wound, resulting in wound/wound effusion; while hyaluronic acid is too hydrophilic, which may lead to dehydration of the wound. If the two are combined in a certain ratio, they can achieve proper water absorption sex.

Through our research, we found that when the weight ratio of medical gelatin: hyaluronic acid = 99:1-50:50, and the porous composite material is formed by low-temperature freeze-drying under appropriate conditions, a good water vapor transmission rate can be achieved.

Contents of the invention

The object of the present invention is to provide a composite biological dressing which is porous, degradable, has good water vapor transmission rate and can effectively promote wound healing.

In order to achieve the above object, the technical solution provided by the present invention is: a composite biological dressing for promoting wound healing, its raw materials are composed by weight: 50-99 parts of medical gelatin, 1-50 parts of hyaluronic acid; in the above components 0.05%-2% of a cross-linking agent is added, and the cross-linking agent is one of carbodiimide, glutaraldehyde, formaldehyde, diepoxide, and divinyl sulfone. The pore diameter of the composite biological dressing is 50-500um.

Preferably, the raw materials are composed by weight: 80-90 parts of medical gelatin, 10-20 parts of hyaluronic acid; 0.05%-2% of a cross-linking agent is added to the above components, and the cross-linking agent is carbodiazepine One of amines, glutaraldehyde, formaldehyde, diepoxides, and divinylsulfone. The pore diameter of the composite biological dressing is 50-500um.

Another object of the present invention is to provide the preparation method of the above-mentioned composite biological dressing for promoting wound healing, comprising the following steps:

1) Dissolving the medical gelatin with an acetic acid solution of pH=4.7 into a solution with a concentration of 5 mg/mL;

2) Dissolving hyaluronic acid in acetic acid solution with pH=4.7 into a solution with a concentration of 5 mg/mL;

3) The above two solutions are mixed according to a certain ratio, so that the mixed solution includes 50-99 parts of medical gelatin and 1-50 parts of hyaluronic acid.

4) Add a cross-linking agent with a mass fraction of 0.05%-2% to the mixed solution, and after rapid stirring, freeze the mixed solution at -10°C to 45°C for 12-48h, and then freeze-dry it in a vacuum freeze dryer for 48h , to obtain a composite hydrogel, which can be sterilized with cobalt ⁶⁰ .

The composite biological dressing for promoting wound healing of the present invention has the following characteristics: In addition to the biological effects of both medical gelatin and hyaluronic acid, the dressing has suitable hydrophilicity through the proper proportion of the two raw materials As well as the appropriate pore size and ideal water vapor transmission rate, it can well protect wounds and wounds and promote wound healing.

Description of drawings

The present invention will be further described below in conjunction with the accompanying drawings.

Fig. 1 is the SEM topography diagram of the surface and section of the composite biological dressing of the present invention.

Figure 2(1)-(2) is a comparison chart of water vapor transmission rate of composite biological dressings with different proportions.

Figure 3 is a morphological comparison of the cytotoxicity experiments between the composite biological dressings with different proportions and the blank group.

Fig. 4 is a comparison chart of cytotoxicity test data analysis of composite biological dressings with different proportions.

Fig. 5 is an observation and comparison diagram of different ratios of composite biological dressings applied to mouse wounds.

Specific implementation method:

Example 1:

A certain amount of medical gelatin and hyaluronic acid were respectively added to an acetic acid solution with a pH of 4.7 to prepare two solutions with a concentration of 5 mg/mL. The prepared medical gelatin solution and the hyaluronic acid solution were uniformly mixed at a mass ratio of 9:1, and 0.1% EDC cross-linking agent was added to the mixed solution, and stirred rapidly for 4 hours. The mixture was frozen at -45°C for 12 hours, and then freeze-dried in a vacuum freeze dryer for 48 hours to obtain a medical gelatin/hyaluronic acid composite hydrogel. Cobalt ⁶⁰ disinfection can obtain the SEM morphology of the surface and section of the medical gelatin/hyaluronic acid (9:1) composite biological dressing shown in Figure 1.

Example 2:

A certain amount of medical gelatin and hyaluronic acid were respectively added to the acetic acid solution with pH = 4.7 to prepare two solutions with a concentration of 5 mg/mL. The prepared medical gelatin solution and the hyaluronic acid solution were uniformly mixed at a mass ratio of 8:2, and 0.1% EDC cross-linking agent was added to the mixed solution, and stirred rapidly for 4 hours. The mixture was frozen at -45°C for 12 hours, and then freeze-dried in a vacuum freeze dryer for 48 hours to obtain a medical gelatin/hyaluronic acid composite hydrogel. Cobalt ⁶⁰ can be used for disinfection.

Example 3: Water vapor transmission rate experiment of medical gelatin/hyaluronic acid composite biological dressing.

The water vapor transmission rate (WVTR) of the hydrogel determines the moisture absorption and breathability of the material, and the water vapor transmission rate of the material is measured according to the medical gelatin STM standard. The material obtained in Example 1 and Example 2 is placed in a container containing 10ml of water In a cylindrical plastic cup (diameter 34mm). Teflon was used around the material to prevent any moisture from evaporating through the boundaries of the material, placed in an incubator at 37°C and 35% relative humidity. Intervals are weighed and the material is trended over time. The water vapor passing rate is calculated by the following formula:

A refers to the area of the sample to be measured. Each group of materials with different proportions was repeated three times. The water vapor transmission rate results of different proportions of medical gelatin/hyaluronic acid composite biological dressings are shown in Figure 2. Wherein the water vapor transmission rate of medical gelatin: hyaluronic acid=9:1 is 2247g/ ^m2 /d gelatin y, the water vapor transmission rate of medical gelatin: hyaluronic acid=8: ² is 2263g/m2/day, they The water vapor transmission rate is just in the range of 2000-2500g/m ² /day.

Example 4: Cell morphology observation of medical gelatin/hyaluronic acid composite biological dressing

According to the ISO10993 standard, the materials obtained in Example 1 and Example 2 (composite medical gelatin: hyaluronic acid = 9: 1 and medical gelatin: hyaluronic acid = 8: 2) were mixed at 6 cm ² /ml The standard leached in 1640 culture solution, placed in a constant temperature incubator at 37° C., 5% CO ₂ , and 100% humidity, and cultivated for 24 hours to obtain an extract. In a 24-well culture plate, 1ml of L929 cell suspension with a cell density of 5× ¹⁰³ cells/ml was seeded. After 24 hours of culture, the original culture medium was replaced, washed twice with PBS, and then the material extract was added. After culturing for 1 day, the cell morphology was observed under an inverted phase-contrast optical microscope, so as to evaluate the growth of the cells. The test showed ( FIG. 3 ) that the L929 fibroblasts grown by adding the material extract had no difference in cell morphology from the normal fibroblasts. However, the number of cells in the medical gelatin: hyaluronic acid = 9:1 group grew more than other groups.

Embodiment 5: the cytotoxicity of medical gelatin/hyaluronic acid composite biological dressing

In order to further quantify the cytotoxicity of the materials obtained in Example 1 and Example 2, that is, the growth and quantity of cells after adding different proportions of material extracts. We took the MTT approach for measurement. According to the ISO10993 standard, the materials obtained in Example 1, Example 2, and Example 3 were leached into 1640 culture solution at a standard of 6 cm ² /ml, and placed in a room with 37°C, 5% CO ₂ , and a humidity of 100%. Cultivate in a constant temperature incubator for 24 hours to obtain an extract. Plant 100ul of L929 cell suspension with a cell density of ¹⁰⁴ /ml in a 96-well culture plate, culture for 1 day, replace the original culture medium, wash twice with PBS, and add 100ul of medical gelatin in different proportions to each well and hyaluronic acid compound biological dressing material extract. After culturing for 1 day, the cell viability was detected by MTT assay. Add 20ul of MTT solution (5mg/ml, ie 0.5% MTT) to each well and continue culturing for 4h. To terminate the culture, carefully aspirate the culture medium in the well. Add 150ul of DMSO to each well, shake on a shaker at low speed for 10min, and fully dissolve the crystals. The absorbance of each well was measured at OD490nm in an enzyme-linked immunosorbent assay instrument. When the leachate method is used to test the cytotoxicity of biomedical materials, the relative proliferation degree of cells is often used as the standard to divide the safety level.

The results are shown in the figure ( FIG. 4 ). The cytotoxicity of the composite materials of Chinese medicine gelatin: hyaluronic acid = 9: 1 and medical gelatin: hyaluronic acid = 8: 2 is zero order, that is, there is no cytotoxicity.

Embodiment 6: Live animal experiment of medical gelatin/hyaluronic acid composite biological dressing

A full-thickness wound model was established on the back of C57BL/6J mice. The experiment is divided into an experimental group and a blank control group. After the wound is set up, the experimental group adopts the dressings obtained in Example 1 and Example 2 to change the wound (composite Chinese medicine gelatin: hyaluronic acid=9: 1 and medical gelatin: hyaluronic acid=8:2 two groups), and the blank control group was allowed to heal naturally. Under the condition of fixed focal length and pixels, the back wounds of mice were continuously photographed, and the wound healing conditions of mice in different material groups were recorded. The experiment showed that the healing time of the experimental group was significantly shorter than that of the blank control group (see Figure 5).

The specific implementation above is only a preferred embodiment of the present invention, and is not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present invention should be included in the present invention within the scope of protection.

Claims (5)

1. A composite biological dressing for promoting wound healing, characterized in that the raw materials of the composite biological dressing consist of 50-99 parts of medical gelatin and 1-50 parts of hyaluronic acid by weight. 2. The composite biological dressing for promoting wound healing according to claim 1, characterized in that the raw materials of the composite biological dressing are preferably composed of 80-90 parts of medical gelatin and 10-20 parts of hyaluronic acid. 3. according to the composite biological dressing of promoting wound healing according to claim 1, it is characterized in that: add the linking agent of 0.05%-2% in above-mentioned components, and described linking agent is carbodiimide, glutaraldehyde, One of formaldehyde, diepoxide, and divinyl sulfone. 4. The composite biological dressing for promoting wound healing according to claim 1, characterized in that: the composite biological dressing has a pore size of 50-500um. 5. the preparation method of the composite biological dressing that promotes wound healing described in one of claims 1-4, is characterized in that: described preparation method comprises the following steps: 1) Dissolving the medical gelatin with an acetic acid solution of pH=4.7 into a solution with a concentration of 5 mg/mL; 2) Dissolve hyaluronic acid with pH: 4.7 acetic acid solution to a solution with a concentration of 5 mg/mL; 3) The above two solutions are mixed according to a certain ratio, so that the mixed solution accounts for 50-99 parts of medicinal gelatin and 1-50 parts of hyaluronic acid; 4) Add a cross-linking agent with a mass fraction of 0.05%-2% to the mixed solution, and after rapid stirring, freeze the mixed solution at -10°C to 45°C for 12-48h, and then freeze-dry it in a vacuum freeze dryer for 48h After obtaining the composite hydrogel, it can be sterilized with cobalt ⁶⁰ .