CN114904045B

CN114904045B - Hydrogel medical dressing containing traditional Chinese medicine extract and preparation method thereof

Info

Publication number: CN114904045B
Application number: CN202210607307.2A
Authority: CN
Inventors: 满佳; 李宗耀; 张永琪; 王晓杰; 李建勇; 李剑峰
Original assignee: Shandong University
Current assignee: Shandong University
Priority date: 2022-05-31
Filing date: 2022-05-31
Publication date: 2023-01-17
Anticipated expiration: 2042-05-31
Also published as: CN114904045A

Abstract

The invention relates to a hydrogel medical dressing containing traditional Chinese medicine extracts and a preparation method thereof, and the hydrogel medical dressing containing traditional Chinese medicine extracts has the functions of promoting blood coagulation and healing wounds. The preparation method provided by the invention takes sodium alginate and carrageenan as a composite matrix, takes cellulose as a reinforcement, takes Chinese herbal medicine extracts as bioactive components, and prepares the composite hydrogel medical dressing through the plasticizing effect of glycerol. The hydrogel medical dressing has better cell affinity and full degradation performance, the elongation at break can reach 80%, and the problem of rapid blood coagulation and hemostasis is solved. The Chinese herbal medicine extract has obvious bacteriostatic effect on staphylococcus aureus and escherichia coli, and does not contain antibiotics. The hydrogel medical dressing can promote the expression of f-actin of cells. The expression level of the f-actin of the cells co-cultured with the medical dressing is 70 percent higher than that of the control group.

Description

Hydrogel medical dressing containing traditional Chinese medicine extract and preparation method thereof

Technical Field

The invention belongs to the technical field of composite hydrogel medical dressings, and particularly relates to a hydrogel medical dressing containing traditional Chinese medicine extracts and a preparation method thereof.

Background

The information in this background section is only for enhancement of understanding of the general background of the invention and is not necessarily to be construed as an admission or any form of suggestion that this information forms the prior art that is already known to a person of ordinary skill in the art.

As the aging population increases, the number of elderly patients who rely on oral anticoagulant medication will continue to increase, and these medications are prone to traumatic, prolonged bleeding. At the same time, the problem of how soldiers perform effective coagulation of wounds during training and combat and after completion of many surgical procedures has plagued humans. In many cases, excessive bleeding from a wound can be a serious threat to the health of the injured person and can even be life threatening. Therefore, dressings that accelerate the clotting process and thereby reduce bleeding time are well suited for these patients. In summary, accelerating coagulation is an important issue, both in the treatment of major wounds and in the treatment of surface wounds.

The evolution of superbacteria with antibiotic resistance has become a major clinical challenge in the 21 st century. Especially, chronic wounds often need frequent replacement of antibacterial dressings, which easily leads to the evolution of antibiotic-resistant pathogens. The proliferation of bacteria at the wound site is liable to cause inflammation of the wound infection, which leads to stagnation of the wound in the inflammatory phase, resulting in difficulties in subsequent wound treatment and healing. The antibacterial principle of Chinese herbal medicine relates to the complicated processes of instability of cytoplasmic membranes, inhibition of extracellular microbial enzymes, deprivation of substrates required by microbial growth or direct action on microbial metabolism by inhibiting oxidative phosphorylation. Compared with the antibiotic with single action mode, the Chinese herbal medicine can reduce the generation of antibiotic resistance. This is an advantage over chemical antibiotics.

Disclosure of Invention

In order to solve the problems of procoagulant hemostasis, bacteriostasis, antioxidation and promotion of wound healing in wound care, the invention aims to provide a hydrogel medical dressing containing a traditional Chinese medicine extract and a preparation method thereof. The preparation method provided by the invention takes sodium alginate and carrageenan as a composite matrix, takes cellulose as a reinforcement, takes Chinese herbal medicine extracts as bioactive components, and prepares the composite hydrogel medical dressing through the plasticizing effect of glycerol.

In order to achieve the technical purpose, the technical scheme adopted by the invention is as follows:

a preparation method of hydrogel medical dressing containing traditional Chinese medicine extract comprises the following steps:

(1) Extracting effective components of the Chinese herbal medicines: cleaning the Chinese herbal medicines, drying, and then crushing into powder; soaking the powder with deionized water; grinding by using colloid mill equipment, and filtering the obtained fine particle turbid liquid; adding methanol into the filter residue, and stirring and extracting to obtain an extract; filtering to obtain filtrate, and lyophilizing the filtrate to obtain Chinese medicinal extract;

(2) Preparation of the medical dressing: taking carrageenin and sodium alginate as a composite matrix, adding microcrystalline cellulose and water, heating and stirring, then adding glycerol and continuing stirring, and finally adding Chinese herbal medicine extracts to obtain a film forming solution;

(3) And (4) carrying out suction filtration on the film forming solution, removing bubbles in the film forming solution, and then putting the film forming solution into an air drying box for air drying at room temperature to obtain a final finished product.

Further, the Chinese herbal medicine is madder or amaranthus spinosus, and is preferably madder.

Further, the soaking time is 1-3 h; the weight of the deionized water is 30-50 times of that of the powder during soaking.

Further, the grinding method comprises the following steps: the distance between the grinding discs of the colloid mill is gradually reduced from +800 mu m to-50 mu m, and the fine particle turbid liquid is obtained after repeated grinding for 10 times by keeping the distance of-50 mu m.

Further, the extraction method comprises the following steps: adding 20-30 times of methanol into the filter residue, wherein the concentration of the methanol is 60-85%, and stirring and extracting for 36-72 h.

Further, the filtering is performed by adopting a 600-mesh screen.

Further, the mass ratio of carrageenan to sodium alginate is 2:1; carrageenin and sodium alginate are used as a composite matrix, and the dosage of microcrystalline cellulose is 2-10% of the weight of the matrix; the amount of water is 30-50 times of the weight of the matrix.

Further, heating and stirring the mixture in a water bath kettle at the temperature of between 70 and 80 ℃ for 15 to 30min.

Furthermore, the using amount of the glycerol is 0.05-2% of the weight of the matrix; the dosage of the Chinese herbal medicine extract is 0.1-30% of the weight of the matrix.

Further, the air drying time is 24-72h.

The hydrogel medical dressing containing the traditional Chinese medicine extract is prepared according to the preparation method.

The invention has the beneficial effects that:

the invention provides a novel hydrogel medical dressing which has better cell affinity and full degradation performance. The breaking elongation of the hydrogel medical dressing can reach 80 percent, and the hydrogel medical dressing can be used for sports wounds of parts (such as elbows, buttocks and knees) with telescopic body surfaces, which are difficult to adapt to traditional wound dressings. The hydrogel medical dressing can reduce the blood coagulation time by 75 percent, can achieve the extracorporeal whole Blood Coagulation Index (BCI) of 11 percent, and can solve the problem of rapid blood coagulation and hemostasis. The hydrogel medical dressing adopts Chinese herbal medicine extracts, has obvious bacteriostatic effect on staphylococcus aureus and escherichia coli, and does not contain antibiotics. Has important significance for treating chronic wounds and reducing the generation of drug-resistant bacteria. The hydrogel medical dressing can promote the expression of cell f-actin. The expression level of the f-actin of the cells co-cultured with the medical dressing is 70 percent higher than that of the control group.

Drawings

The accompanying drawings, which are incorporated in and constitute a part of this specification, are included to provide a further understanding of the invention, and are incorporated in and constitute a part of this specification, illustrate exemplary embodiments of the invention and together with the description serve to explain the invention and not to limit the invention.

Fig. 1 is a schematic flow chart of the preparation of the composite hydrogel medical dressing.

FIG. 2 is a graph showing the results of the clotting time test performed in accordance with the present invention.

FIG. 3 is an image of the zone of inhibition experiment performed by the present invention.

FIG. 4 shows the results of DPPH scavenging experiments performed in accordance with the present invention.

FIG. 5 is a confocal image of the f-actin staining experiment performed in accordance with the present invention.

Fig. 6 is a graph of the results of mechanical performance testing of two dressings of the present invention.

Detailed Description

It should be noted that the following detailed description is exemplary and is intended to provide further explanation of the disclosure. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs.

The invention will now be further described with reference to the accompanying drawings and detailed description.

Example 1

A preparation method of hydrogel medical dressing containing traditional Chinese medicine extract comprises the following steps (the preparation process can be shown in figure 1):

(1) Extracting active ingredients of madder: cleaning radix Rubiae, and drying. Pulverizing dried radix Rubiae with small pulverizer. Putting a certain amount of the powder into a container, adding thirty times of deionized water by weight, and soaking for 1h. Grinding with colloid mill, gradually decreasing the distance between the grinding discs of colloid mill from +800 μm to-50 μm, repeatedly grinding for 10 times while maintaining the distance between-50 μm to obtain fine-particle turbid liquid, and filtering. Then twenty times of methanol by weight is added into the filter residue, the concentration of the methanol is 60 percent, and the mixture is stirred and extracted for 36 hours. Filtering with 600 mesh sieve, and lyophilizing the filtrate to obtain radix Rubiae extract.

(2) Preparation of the medical dressing: taking a certain amount of carrageenan and sodium alginate (2:1) as a composite matrix, adding microcrystalline cellulose accounting for 2% of the weight of the matrix, adding water accounting for thirty times of the weight of the matrix, heating and stirring in a 70 ℃ water bath kettle for 15min, then adding 2wt% of glycerol into the mixed solution, continuing stirring for 20min, finally adding Chinese herbal medicine extract accounting for 30wt% of the matrix into the mixed solution, continuing stirring for 2min, and uniformly mixing the solution to obtain a film forming solution. And (3) carrying out suction filtration on the film forming solution prepared in the steps, removing bubbles in the film forming solution, and then placing the film forming solution into an air drying box to carry out air drying for 48 hours at room temperature to obtain a final finished product.

Example 2

This example is substantially the same as example 1, except that:

in this embodiment, the step (1) specifically includes: cleaning the Chinese herbal medicine acalypha spinosa, and drying for later use. Pulverizing dried Acalypha spinosa into powder with a small pulverizer. Putting a certain amount of the powder into a container, adding thirty times of deionized water by weight, and soaking for 1h. Grinding with colloid mill equipment, gradually decreasing the distance between the grinding discs of the colloid mill from +800 μm to-50 μm, repeatedly grinding for 10 times while maintaining the distance between-50 μm to obtain fine particle turbid solution, and filtering. Then twenty times of methanol by weight is added into the filter residue, the concentration of the methanol is 60 percent, and the mixture is stirred and extracted for 36 hours. Filtering with 600 mesh sieve, and lyophilizing the filtrate to obtain Acalypha australis extract.

Comparative example: medical dressings in other research

The hydrogel medical dressing and the comparative product obtained in the examples 1 to 2 are tested by the following specific testing method:

and (3) testing mechanical properties: and (3) performing a tensile test on the hydrogel medical dressing by using a general electronic testing machine to obtain the Tensile Strength (TS) and the Elongation At Break (EAB) of the dressing. The tensile specimen has a measured length of 25mm, a width of 5mm and a moving jig speed of 5mm/min. The samples were measured in parallel at least 5 times and the results were averaged.

In vitro coagulation test: first, 5ml of anticoagulated blood with sodium citrate and 150ul of CaCl were added ₂ The solutions (0.2 mol/L) were mixed well and a hydrogel wound dressing 8mm in diameter was placed in the bottom of a test tube with a capacity of 50 ml. Then, 100ul of the above mixed blood was added to the tube, and the tube was incubated at 37 ℃ for 5min. 25ml of deionized water was then carefully added to each tube to resuspend the unset blood without breaking the original clot. Finally, the supernatant in the tube was carefully removed and its absorbance at 540nm was measured with an ultraviolet spectrophotometer. The blood clotting properties of plain gauze were used as a control and untreated dicalcium whole blood was used as a blank. Zero is indicated by the absorbance of deionized water at 540 nm. The whole blood coagulation performance (BCI) of the hydrogel was evaluated by the following formula:

“Abs _{sample (I)} "refers to the absorbance of the supernatant in the sample set.

“Abs _{Blank space} "refers to the absorbance of the supernatant in the blank.

At the same time, the actual clotting time of the dressing was determined. First, a wound dressing 8mm in diameter was placed in a centrifuge tube. Then, 0.2ml of potassium oxalate (50 mg/ml) was previously added to a 10ml syringe. 10ml of fresh rabbit blood was drawn using a syringe and 0.5ml of blood sample was added to each tube. Then 0.1ml of CaCl is added into each centrifuge tube ₂ Solution (3 mg/ml), blood clotting time was observed. Blank groups were as above. Experiment ofThis was repeated 3 times and the average was taken.

And (3) testing antibacterial performance: the antibacterial performance of the wound dressing on escherichia coli and staphylococcus aureus is measured by adopting an antibacterial ring method. First, an LB liquid medium and an LB solid medium were prepared. Then 3 tubes of 12mL bacterial culture tubes are taken, 3mL LB liquid culture medium is added, single colony is selected from solid culture medium of escherichia coli and staphylococcus aureus respectively and added into the liquid culture medium, and the other tube is used as a blank control group. The cells were cultured with shaking in a constant temperature shaker (37 ℃ C., 200 rpm) for 15 hours. Respectively preparing the amaranthus spinosus wound dressing and the madder wound dressing into round pieces with the diameter of 8mm by using a biopsy puncher, and sterilizing the two sides of the round pieces for 30min by ultraviolet irradiation for later use. Respectively diluting escherichia coli and staphylococcus aureus liquid to CFU/mL concentration by using sterile PBS solution, and uniformly coating 100 mu L of each diluted liquid on an LB solid culture medium. The sample holder was placed in the corresponding plate and gently pressed to bring the sample into full contact with the surface of the medium. And (3) placing the culture dish in a constant-temperature incubator at 37 ℃ for standing culture for 24h, taking out and taking a picture, and measuring the size of a bacteriostatic zone. The experiment was repeated three times.

And (3) testing the oxidation resistance: the antioxidant efficiency of the wound dressing was determined in terms of DPPH free radical scavenging capacity. First, DPPH ethanol solutions at concentrations of 0.01, 0.02, 0.03, and 0.04mg/mL were carefully prepared as standard solutions. These standard solutions were then scanned at 517nm using an ultraviolet-visible spectrometer. Within the selected concentration range (0.01-0.04 mg/mL), a linear calibration curve (concentration at 517nm versus absorbance) was obtained with a correlation coefficient R =0.9997. 0.5g of the cut wound dressing was placed in DPPH ethanol solution (4 mL, 0.04mg/mL) and left in the dark at 25 ℃ for 1h. The absorbance of the solution was then scanned by UV-visible spectroscopy at 517nm to obtain the remaining DPPH concentration from the calibration curve.

Cell f-actin expression test: the phenotype of HaCaT cells co-cultured with different medical dressings was observed using a scanning laser confocal microscope. Cells were washed with PBS and then fixed with 4% paraformaldehyde at room temperature for 30-45min. Fixed cells were stained for f-actin with phalloidin, nuclei with Hoechst, and for 1h at room temperature. Cells in the control group were cultured normally without soaking the dressing in the medium.

The result of the blood coagulation time test performed by the method is shown in fig. 2, and compared with the blank group, the madder dressing can shorten the blood coagulation time by 75%, and the acalypha australis dressing can shorten the blood coagulation time by 40%.

Fig. 3 is an image of the inhibition zone experiment performed in the present invention, wherein fig. 3a is the experimental result of the inhibition zone of escherichia coli (the left acalypha spinosa and the right madder), and fig. 3b is the experimental result of the inhibition zone of staphylococcus aureus (the left acalypha spinosa and the right madder). As can be seen from the figure, the madder dressing has better bacteriostatic effect.

Meanwhile, the results of the inhibition zone experiments of the madder dressing and the amaranthus spinosus dressing are shown in tables 1 and 2. As can be seen from the results in tables 1 and 2, the madder dressing in example 1 has a good bacteriostatic effect.

TABLE 1 Escherichia coli zone of inhibition experiment results

TABLE 2 Staphylococcus aureus circles test results

Fig. 4 shows the results of DPPH removal experiments performed in accordance with the present invention, where greater DPPH consumption indicates greater antioxidant capacity of the dressing, and the madder dressing is sufficient to reduce DPPH by 80%, while the acalypha dressing is capable of reducing DPPH by 40%.

Fig. 5 is a confocal imaging diagram of an f-actin staining experiment performed in the present invention, in which f-actin is stained red, compared to the control group, the red fluorescence intensity of the cells in the amaranthus spinosus dressing group is increased by about 60%, and the red fluorescence intensity of the cells in the madder dressing group is increased by about 75%.

Fig. 6 shows the mechanical properties of the two dressings of the invention, the elongation at break of the madder dressing can reach 72.43%, and the elongation at break of the amaranthus spinosus dressing can reach 81.21%. For the tensile strength, the madder dressing is obviously superior to the amaranthus spinosus dressing, the tensile strength of the madder dressing can reach 7Mpa, and the tensile strength of the amaranthus spinosus dressing is only about 3.4 Mpa.

The above description is only a preferred embodiment of the present application and is not intended to limit the present application, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, improvement and the like made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. A preparation method of hydrogel medical dressing containing traditional Chinese medicine extract is characterized by comprising the following steps:

(1) Extracting effective components of the Chinese herbal medicines: cleaning the Chinese herbal medicines, drying, and then crushing into powder; soaking the powder in deionized water; grinding by using colloid mill equipment, and filtering the obtained fine particle turbid liquid; adding methanol into the filter residue, and stirring and extracting to obtain an extract; filtering to obtain filtrate, and lyophilizing the filtrate to obtain Chinese medicinal extract;

(3) Filtering the film forming solution to remove air bubbles in the film forming solution, and then putting the film forming solution into an air drying box for air drying at room temperature to obtain a final finished product;

the Chinese herbal medicine is madder or amaranthus spinosus;

the mass ratio of carrageenan to sodium alginate is 2:1; carragheenan and sodium alginate are used as a composite matrix, and the dosage of microcrystalline cellulose is 2-10% of the weight of the composite matrix; the amount of water is 30-50 times of the weight of the composite matrix; the dosage of the glycerol is 0.05 to 2 percent of the weight of the composite matrix; the dosage of the Chinese herbal medicine extract is 0.1 to 30 percent of the weight of the composite matrix.

2. The method of claim 1, wherein the herb is madder.

3. The preparation method according to claim 1, wherein the soaking time is 1 to 3 hours; the weight of the deionized water is 30-50 times of that of the powder during soaking.

4. The method according to claim 1, wherein the grinding method is: the distance between the grinding discs of the colloid mill is gradually reduced from +800 mu m to-50 mu m, and then the distance between the grinding discs of the colloid mill and-50 mu m is kept for repeated grinding for 10 times to obtain fine particle turbid liquid.

5. The method of claim 1, wherein the extraction process is: adding 20-30 times of methanol into the filter residue, wherein the concentration of the methanol is 60-85%, and stirring and extracting for 36-72h.

6. The method of claim 1, wherein the filtering is performed with a 600 mesh screen.

7. The method according to claim 1, wherein the mixture is heated and stirred in a water bath at 70 to 80 ℃ for 15 to 30min.

8. The method of claim 1, wherein the air drying time is 24-72 hours.

9. The hydrogel medical dressing containing the Chinese medicinal extract prepared by the preparation method according to any one of the above claims 1-8.