Polyvinyl alcohol-based composite hydrogel, preparation method and application thereof

Abstract

The application provides a polyvinyl alcohol-based composite hydrogel, a preparation method and application thereof. The composite hydrogel is obtained by cross-linking of shikonin (SHK) and polyvinyl alcohol (PVA). In an alkaline environment, SHK and PVA are cross-linked, wherein the hydroxyl groups on the side chains of PVA and the phenolic hydroxyl groups in the SHK molecules can form stable and firm hydrogen bonds under alkaline conditions, so as to anchor the SHK on the PVA molecular chains on the surface of the hydrogel. When encountering a moist environment such as cells and tissues, the SHK / PVA hydrogel swells, the hydrogen bonds are opened, the SHK existing on the surface of the hydrogel is released to the surrounding environment in the form of a raw drug molecule, and the SHK plays a biological activity role of anti-inflammatory, antioxidant and repair, and can effectively achieve the purpose of promoting rapid wound healing.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of hydrogel preparation, in particular to a polyvinyl alcohol-based composite hydrogel, a preparation method and application thereof. BACKGROUND

[0002] Skin, as the first line of defense of the human body, has important functions such as protection, regulation and secretion. However, skin injuries caused by burns, trauma, infection and chronic diseases still face great challenges worldwide. Delayed wound healing increases the risk of wound infection or complications and results in high treatment costs, so it is urgent to develop a rapid and effective wound repair material (i.e., wound dressing).

[0003] In order to achieve rapid wound healing, wound dressings need to have multiple functions such as anti-inflammatory, antibacterial, angiogenesis, proliferation promotion and remodeling. However, traditional wound dressings are dry and cannot provide a moist environment for wound healing. Hydrogels can retain a large amount of water and create a moist healing environment, and are considered an ideal dressing for damage repair. Among them, polyvinyl alcohol (PVA) hydrogel is a non-toxic and non-immunogenic polymer, which is widely used in the field of tissue engineering due to its stable chemical properties, high water content, good permeability and biocompatibility. However, a single PVA hydrogel does not have biological activity and antibacterial properties, and cannot promote cell adhesion and wound healing.

[0004] In order to make the prepared wound dressing have antibacterial properties and promote cell adhesion and wound healing, most of the current wound dressings introduce antibacterial drugs (such as ciprofloxacin) and growth factors (such as recombinant human epidermal growth factor (rhEGF)) into the hydrogel dressing, which significantly increases the preparation cost of the dressing, and the preparation process of the dressing is complicated. The active ingredients of the growth factor have strict requirements for the storage and transportation of the drug-loaded dressing, resulting in a significant increase in transportation cost, and the addition of antibacterial drugs may cause the biocompatibility of the wound dressing to deteriorate. SUMMARY

[0005] Therefore, the purpose of the present application is to provide a polyvinyl alcohol-based composite hydrogel, a preparation method and application thereof. The polyvinyl alcohol-based composite hydrogel has low preparation cost and excellent biological activity and biocompatibility, and can promote cell adhesion and wound healing.

[0006] To achieve this purpose, the present application adopts the following technical solutions:

[0007] In a first aspect, the present application provides a polyvinyl alcohol-based composite hydrogel obtained by cross-linking of shikonin and polyvinyl alcohol.

[0008] Preferably, the mass ratio of shikonin to polyvinyl alcohol is (0-3):(97-100).

[0009] Preferably, the polyvinyl alcohol has a polymerization degree of 1000-3000.

[0010] Preferably, the polyvinyl alcohol has an alcoholysis degree of 98-100%.

[0011] In a second aspect, the present application further provides a preparation method of the polyvinyl alcohol-based composite hydrogel, comprising the following steps:

[0012] S1: drying the polyvinyl alcohol solution to obtain a polyvinyl alcohol hydrogel;

[0013] S2: crosslinking the baphalan with the polyvinyl alcohol hydrogel in an alkaline environment to obtain the polyvinyl alcohol-based composite hydrogel.

[0014] 5. The preparation method of claim 4, wherein the drying temperature is 20-80℃.

[0015] Preferably, the alkaline environment is provided by an alkaline solution.

[0016] Preferably, the alkaline solution is selected from any one or more of a sodium hydroxide solution, a potassium hydroxide solution, ammonia water or a sodium ethoxide solution.

[0017] Preferably, the concentration of OH- in the alkaline solution is 0-6 mol / L.

[0018] Preferably, the crosslinking time is 10-60 min.

[0019] Preferably, the crosslinking is followed by a washing step.

[0020] Preferably, the washing reagent is water.

[0021] Preferably, the washing is completed when the washing solution is neutral.

[0022] In a third aspect, the present application provides a wound dressing comprising the polyvinyl alcohol-based composite hydrogel involved in the technical solutions described above.

[0023] Compared with the prior art, the present application has the following advantages:

[0024] The present application obtains a polyvinyl alcohol-based composite hydrogel by cross-linking a natural Chinese herbal medicine component, shikonin (SHK), with polyvinyl alcohol, wherein the SHK has multiple pharmacological effects, including anti-inflammatory, antioxidant, anti-tumor, antibacterial, antiviral and repair effects, and the PVA has stable chemical properties, high water content, good permeability and biocompatibility. The present application cross-links SHK and PVA in an alkaline environment, wherein the hydroxyl groups on the side chains of PVA and the phenolic hydroxyl groups in the SHK molecules can form stable and firm hydrogen bonds under alkaline conditions, thereby anchoring the SHK on the PVA molecular chains on the surface of the hydrogel. When encountering a moist environment such as cells and tissues, the SHK / PVA hydrogel swells, the hydrogen bonds open, and the SHK present on the surface of the hydrogel is released into the surrounding environment in the form of a raw drug molecule, thereby exerting its biological activity effects of anti-inflammatory, antioxidant and repair, and effectively achieving the purpose of promoting rapid wound healing, and having good biocompatibility. BRIEF DESCRIPTION OF DRAWINGS

[0025] Figure 1 It is a schematic diagram of the hydrogen bond combination of SHK and PVA in the SHK / PVA hydrogel;

[0026] Figure 2 It is a body microscope image of the SHK / PVA hydrogel obtained in Examples 1-3 and the PVA hydrogel obtained in Comparative Example 1;

[0027] Figure 3 It is a schematic diagram of the chemical structure of SHK and Que;

[0028] Figure 4 (a) is a comparison diagram of the drug content of the SHK / PVA hydrogel obtained in Examples 1-3 in DMSO calculated by the standard dissolution curve;

[0029] Figure 4 (b) is a comparison diagram of the drug content of the Que / PVA hydrogel obtained in Comparative Examples 2-4 in DMSO calculated by the standard dissolution curve;

[0030] Figure 5 It is a comparison diagram of the proliferation state of the NIH3T3 cells to the SHK / PVA hydrogel obtained in Examples 1-3 and the PVA hydrogel obtained in Comparative Example 1;

[0031] Figure 6 It is a comparison diagram of the activity of the NIH3T3 cells to the SHK / PVA hydrogel obtained in Examples 1-3 and the PVA hydrogel obtained in Comparative Example 1;

[0032] Figure 7A comparison chart of wound healing of rats from day 0 to day 14 after the rats' full-thickness wounds are treated with the SHK / PVA hydrogels obtained in Examples 1-3 and the PVA hydrogel obtained in Comparative Example 1;

[0033] Figure 8 A comparison chart of relative wound area of rats from day 0 to day 14 after the rats' full-thickness wounds are treated with the SHK / PVA hydrogels obtained in Examples 1-3 and the PVA hydrogel obtained in Comparative Example 1;

[0034] Figure 9 A comparison chart of fluorescence of inflammatory factors IL-6 and TNF-α after the tissue of defects is treated with the SHK / PVA hydrogels obtained in Examples 1-3 and the PVA hydrogel obtained in Comparative Example 1. DETAILED DESCRIPTION

[0035] The technical solutions of the present application will be described clearly and completely below in combination with the embodiments of the present application. Obviously, the described embodiments are only some of the embodiments of the present application, but not all the embodiments. Based on the embodiments in the present application, all the other embodiments obtained by those skilled in the art without creative labor fall within the scope of protection of the present application.

[0036] Since the single PVA hydrogel does not have biological activity and antibacterial performance, the existing technology mostly introduces antibacterial drugs and growth factors into the hydrogel dressing, which will lead to high preparation cost and transportation cost of the hydrogel dressing, and the introduction of antibacterial drugs will have the risk of reducing the biocompatibility of the hydrogel dressing. Based on this, the present application provides a polyvinyl alcohol-based composite hydrogel (also referred to as "SHK / PVA hydrogel") obtained by cross-linking shikonin and polyvinyl alcohol. The shikonin (SHK) is a soluble naphthoquinone compound extracted from traditional Chinese herbal medicine shikonin, and the molecular formula is C 16 H 16O5, which has multiple pharmacological effects, including anti-inflammatory, antioxidant, antitumor, antibacterial, antiviral and repair effects, and is gradually applied in the field of tissue engineering. The polyvinyl alcohol (PVA) is a non-toxic, non-immunogenic polymer with stable chemical properties, high water content, good permeability and biocompatibility, and is also widely used in the field of tissue engineering. The source of the shikonin and the polyvinyl alcohol in the present application is not particularly limited and can be a general commercially available product. Since the degree of polymerization and the degree of alcoholysis of the polyvinyl alcohol can affect the mechanical properties of the hydrogel finally prepared, in some embodiments of the present application, the degree of polymerization of the polyvinyl alcohol is 1000-3000, and can be 1000, 1200, 1500, 1700, 2000, 2200, 2500, 2700 or 3000, etc.; the degree of alcoholysis is 98-100%, and can be 98%, 98.5%, 99%, 99.5% or 100%, etc. In some embodiments of the present application, the mass ratio of shikonin to polyvinyl alcohol is (0-3):(97-100), which can be 0.1:99.9, 0.5:99.5, 1:99, 1.5:98.5, 2:98, 2.3:97.7, 2.5:97.5 or 3:97, etc. It should be noted that the amount of shikonin is much less than that of polyvinyl alcohol, but the amount of shikonin cannot be 0. First, the surface of the PVA hydrogel has limited hydroxyl groups, and the loading capacity of shikonin is limited. In addition, the method of the present application fixes shikonin to the surface of the PVA hydrogel, and when the PVA hydrogel is applied as a wound dressing to tissue damage, the shikonin molecules on the surface of the PVA hydrogel efficiently act on the wound, thereby exerting their biochemical effects and promoting tissue damage. Considering the cost, the amount of shikonin added should not be too high. Therefore, the grid structure in the SHK / PVA hydrogel provided by the present application is mainly the hydrogen bond structure formed by the crosslinking of the hydroxyl groups of polyvinyl alcohol itself.

[0037] In the present application, SHK and PVA are crosslinked, and the hydroxyl groups on the side chain of PVA and the phenolic hydroxyl groups in the SHK molecule can form stable and firm hydrogen bonds under alkaline conditions, thereby anchoring the SHK on the PVA molecular chain on the surface of the hydrogel (as shown in the schematic diagram Figure 1 When encountering a wet environment such as cells and tissues, the SHK / PVA hydrogel will swell by absorbing water, the hydrogen bonds will open, and the SHK present on the surface of the hydrogel will be released to the surrounding environment in the form of the original drug molecules, exerting its biological activities such as anti-inflammatory, antioxidant and repair effects, and effectively achieving the purpose of promoting rapid wound healing.

[0038] The present application also provides a preparation method of the above-mentioned polyvinyl alcohol-based composite hydrogel, comprising the following steps:

[0039] S1: drying the polyvinyl alcohol solution to obtain a polyvinyl alcohol hydrogel;

[0040] S2: crosslinking the shikonin with the polyvinyl alcohol hydrogel in an alkaline environment to obtain a polyvinyl alcohol-based composite hydrogel.

[0041] According to the present application, the polyvinyl alcohol solution is first subjected to drying treatment to obtain a polyvinyl alcohol hydrogel. The polyvinyl alcohol solution is obtained by mixing polyvinyl alcohol and water, and the water can be deionized water, distilled water or ultrapure water. The mixing temperature is preferably 80-100°C, preferably 90°C, and the time is 8-15h, preferably 10-12h. The mixing is preferably carried out under stirring. After obtaining the polyvinyl alcohol solution, in some embodiments of the present application, the polyvinyl alcohol solution is subjected to drying treatment at 20-80°C, preferably 30-60°C, more preferably 40-60°C, and the drying is stopped when there is no obvious moisture on the surface of the product, i.e. the polyvinyl alcohol hydrogel is obtained. In some embodiments of the present application, the polyvinyl alcohol solution is preferably poured or coated in a mold and dried at 20-80°C to obtain a polyvinyl alcohol film in the form of a hydrogel.

[0042] According to the present application, the shikonin is crosslinked with the polyvinyl alcohol hydrogel in an alkaline environment to obtain a polyvinyl alcohol-based composite hydrogel. In the present application, the alkaline environment is provided by an alkaline solution, and the alkaline solution is selected from any one or more of sodium hydroxide solution, potassium hydroxide solution, ammonia or sodium ethoxide solution. The alkaline solution in the present application is used to dissolve the shikonin on the one hand, and to reinforce the hydrogen bond network structure in the polyvinyl alcohol hydrogel on the other hand. Since in a too strong alkaline environment, the shikonin will undergo self-polymerization, the present application preferably controls the concentration of OH in the alkaline solution to 0.01-1mol / L, preferably 0.1-0.5mol / L. -The concentration of the basic solution is 0-6 mol / L, and 0 mol / L is excluded. Specifically, the concentration can be 0.1 mol / L, 0.5 mol / L, 1 mol / L, 2 mol / L, 3 mol / L, 4 mol / L, 5 mol / L or 6 mol / L, and preferably 2-5 mol / L. In some embodiments of the present application, the shikonin is first dissolved in a basic solution to obtain a shikonin basic solution, and the concentration of the shikonin basic solution in the basic solution is 0-3200 μmol / L, and 0 is excluded. Specifically, the concentration can be 50 μmol / L, 80 μmol / L, 100 μmol / L, 150 μmol / L, 200 μmol / L, 400 μmol / L, 800 μmol / L, 1000 μmol / L, 1200 μmol / L, 1500 μmol / L, 2000 μmol / L, 2500 μmol / L, 3000 μmol / L or 3200 μmol / L, etc. Then, the shikonin basic solution is soaked in the polyvinyl alcohol hydrogel or polyvinyl alcohol film to cross-link the phenolic hydroxyl group of shikonin with the hydroxyl group in the polyvinyl alcohol hydrogel / film, thereby obtaining a polyvinyl alcohol-based composite hydrogel. The soaking can be performed at room temperature, and the soaking time is 10-60 min, preferably 20-40 min, and more preferably 25-30 min. In some embodiments of the present application, the shikonin basic solution is added to a mold containing dried polyvinyl alcohol film, and soaked for 10-60 min. Since the cross-linking of shikonin and polyvinyl alcohol is performed in an alkaline environment, in some embodiments of the present application, a cleaning step is further included after the soaking is completed. Specifically, the soaked film is cleaned with water until the washing liquid is neutral. The purpose of the cleaning is to remove unreacted shikonin and residual alkali solution, so as to facilitate the subsequent practical application of the prepared polyvinyl alcohol-based composite hydrogel.

[0043] The preparation method of the polyvinyl alcohol-based composite hydrogel provided by the present application is simple and convenient, and the cost is low. After the polyvinyl alcohol solution is dried, the obtained product is cross-linked with shikonin in an alkaline environment, which is efficient, fast and easy to realize large-scale production.

[0044] In order to explore the universality of the above preparation method for plant polyphenolic drugs, Que is used to replace shikonin (SHK) to obtain Que / PVA hydrogel. It is found through research that only when a relatively high content of Que is used, a certain drug loading amount of Que in the obtained hydrogel can be achieved. When the amount of Que added is small, the content of Que in the obtained hydrogel is too low to be detected. The results show that compared with Que, the PVA hydrogel provided by the present application is more suitable for carrying SHK, and SHK is the most preferred drug model for the drug loading method of the present application.

[0045] The polyvinyl alcohol-based composite hydrogel involved in the technical scheme in the present application has good abilities of promoting cell adhesion and proliferation through in-vitro experimental research. Meanwhile, the polyvinyl alcohol-based composite hydrogel prepared through a rat skin defect repair experiment further proves to have the abilities of promoting skin tissue regeneration and accelerating wound healing. Moreover, the polyvinyl alcohol-based composite hydrogel has good biocompatibility. Therefore, the present application further provides a wound dressing comprising the polyvinyl alcohol-based composite hydrogel involved in the technical scheme.

[0046] In order to further illustrate the present application, the following examples are used for detailed description. The experimental raw materials used in the following examples of the present application can be purchased from the market or prepared according to the conventional preparation method well known to those skilled in the art. Shikonin is purchased from Aladdin Bio-Chem Technology Co., Ltd., and the model number is S115193.

[0047] The cross-linking schematic diagram between SHK and PVA in the SHK / PVA hydrogel involved in the following examples is shown in Figure 1 It can be seen that the phenolic hydroxyl group of SHK is cross-linked with the hydroxyl group of PVA, so that SHK is carried on PVA.

[0048] Example 1

[0049] The present example provides a SHK / PVA hydrogel, and the preparation method is as follows:

[0050] S1: PVA (polymerization degree 1700, alcoholysis degree greater than 99%) is dissolved in deionized water, and magnetic stirring is performed at 90℃ overnight to obtain a PVA solution of 100mg / mL. -1 Then the PVA mixed solution is poured or coated in a mold, and is completely dried at 37℃;

[0051] S2: A certain amount of SHK is dissolved in a 6M sodium hydroxide solution to configure a cross-linking solution with a SHK concentration of 80uM, and the SHK cross-linking solution is added to the mold containing the dried PVA film, soaked for 30min, and then washed once with water until the pH of the washing liquid is neutral, so as to wash away the residual alkali solution, and obtain 80 SHK / PVA hydrogel (80 refers to the molar concentration of SHK in the alkaline solution).

[0052] Example 2

[0053] The present example provides a SHK / PVA hydrogel, and the preparation method is as follows:

[0054] S1: PVA (polymerization degree 1700, alcoholysis degree greater than 99%) is dissolved in deionized water, and magnetic stirring is performed at 90℃ overnight to obtain a PVA solution of 100mg / mL. -1PVA solution. Then the PVA mixed solution is poured or coated in a mold and allowed to dry completely at 35°C;

[0055] S2: A certain amount of SHK is dissolved in 5M potassium hydroxide solution, a cross-linking solution with SHK concentration of 160 uM is configured, and the SHK cross-linking solution is added to the mold containing the dried PVA film, soaked for 25 min, then washed with water once until the washing solution is neutral, to wash away the residual alkali solution, to obtain 160 SHK / PVA hydrogel (160 refers to the molar concentration of SHK in the alkaline solution).

[0056] Example 3

[0057] This example provides a SHK / PVA hydrogel, which is prepared as follows:

[0058] S1: PVA (polymerization degree 1700, alcoholysis degree greater than 99%) is dissolved in deionized water, magnetically stirred at 90°C overnight, to obtain a 100 mg / mL PVA solution. Then the PVA mixed solution is poured or coated in a mold and allowed to dry completely at 40°C; -1

[0059] S2: A certain amount of SHK is dissolved in 6M sodium hydroxide solution, a cross-linking solution with SHK concentration of 320 uM is configured, and the SHK cross-linking solution is added to the mold containing the dried PVA film, soaked for 35 min, then washed with water once until the washing solution is neutral, to wash away the residual alkali solution, to obtain 320 SHK / PVA hydrogel (320 refers to the molar concentration of SHK in the alkaline solution).

[0060] Comparative Example 1

[0061] S1: PVA (polymerization degree 1700, alcoholysis degree greater than 99%) is dissolved in deionized water, magnetically stirred at 90°C overnight, to obtain a 100 mg / mL PVA solution. Then the PVA mixed solution is poured or coated in a mold and allowed to dry completely at 37°C; -1

[0062] S2: A 6M sodium hydroxide solution is configured as a cross-linking solution, and the cross-linking solution is added to the mold containing the dried PVA film, soaked for 30 min, then washed with water once until the washing solution is neutral, to wash away the residual alkali solution, to obtain PVA hydrogel.

[0063] Comparative Example 2

[0064] S1: PVA (polymerization degree 1700, alcoholysis degree greater than 99%) is dissolved in deionized water, magnetically stirred at 90°C overnight, to obtain a 100 mg / mL PVA solution. Then the PVA mixed solution is poured or coated in a mold and allowed to dry completely at 37°C;​​-1 PVA solution. Then the PVA mixed solution was poured or coated in the mold and allowed to dry completely at 37℃;

[0065] S2: Quercetin (Que) was dissolved in 6M sodium hydroxide solution, a cross-linking solution with Que concentration of 1600uM was configured, and the Que cross-linking solution was added to the mold containing the dried PVA film, soaked for 30min, then washed with water once until the washing liquid was neutral, to wash away the residual alkali solution, to obtain 1600Que / PVA hydrogel (1600 refers to the molar concentration of Que in the alkaline solution).

[0066] Comparative Example 3

[0067] S1: PVA (polymerization degree 1700, alcoholysis degree greater than 99%) was dissolved in deionized water, and stirred magnetically at 90℃ overnight to obtain a PVA solution of 100mg mL -1 . Then the PVA mixed solution was poured or coated in the mold and allowed to dry completely at 37℃;

[0068] S2: Quercetin (Que) was dissolved in 6M sodium hydroxide solution, a cross-linking solution with Que concentration of 320uM was configured, and the Que cross-linking solution was added to the mold containing the dried PVA film, soaked for 30min, then washed with water once until the washing liquid was neutral, to wash away the residual alkali solution, to obtain 320Que / PVA hydrogel (320 refers to the molar concentration of Que in the alkaline solution).

[0069] Comparative Example 4

[0070] S1: PVA (polymerization degree 1700, alcoholysis degree greater than 99%) was dissolved in deionized water, and stirred magnetically at 90℃ overnight to obtain a PVA solution of 100mg mL -1 . Then the PVA mixed solution was poured or coated in the mold and allowed to dry completely at 37℃;

[0071] S2: Quercetin (Que) was dissolved in 6M sodium hydroxide solution, a cross-linking solution with Que concentration of 160uM was configured, and the Que cross-linking solution was added to the mold containing the dried PVA film, soaked for 30min, then washed with water once until the washing liquid was neutral, to wash away the residual alkali solution, to obtain 160Que / PVA hydrogel (160 refers to the molar concentration of Que in the alkaline solution).

[0072] Morphology characterization and drug loading test

[0073] The hydrogels obtained in Examples 1-3 and Comparative Example 1 were subjected to morphology characterization by using a body microscope, and the results were as follows: Figure 2As shown, compared with Comparative Example 1, it can be seen that the products obtained in Examples 1-3 have different contents of SHK loaded in the PVA hydrogel.

[0074] The SHK / PVA hydrogels obtained in Examples 1-3 and the Que / PVA hydrogels obtained in Comparative Examples 2-4 were dissolved using dimethyl sulfoxide (DMSO), and the content of shikonin and quercetin was measured at 520 nm using a multifunctional enzyme label meter. The drug loading amount of SHK / PVA hydrogels for SHK and Que was calculated by comparing the standard curve of shikonin and quercetin in DMSO solution.

[0075] The chemical structural formula of SHK and Que is as shown in Figure 3 The test results are as shown in Figure 4 (* represents p < 0.05), wherein Figure 4 (a) is a comparison diagram of the drug content calculated by the standard dissolution curve of SHK / PVA hydrogels obtained in Examples 1-3 in DMSO; Figure 4 (b) is a comparison diagram of the drug content calculated by the standard dissolution curve of Que / PVA hydrogels obtained in Comparative Examples 2-4 in DMSO.

[0076] As shown in Figure 4 (a), it can be seen that different contents of shikonin provided in Examples 1-3 are loaded in PVA, and the drug loading amount is 1.2-2.3 wt%. As shown in Figure 4 (b), when 1600 μM of Que base crosslinking solution is used for treatment, the drug content of 1600 Que / PVA hydrogel is close to 6%, which indicates that the drug loading method of the present application is also applicable to Que. However, compared with shikonin, the drug loading dose of quercetin is relatively low. When PVA hydrogel is treated with 160 μM of quercetin base solution, the content of quercetin in the hydrogel is too low to be detected. When the PVA hydrogel is treated with 320 μM of quercetin base solution, the drug loading amount of the hydrogel is about 1.3%. It is indicated that compared with Que, the PVA hydrogel provided by the present application is more suitable for carrying SHK, which indicates that SHK is the most preferred drug model for the drug loading method of the present application.

[0077] Test of proliferation state and activity of NIH3T3 cells

[0078] NIH3T3 cells were selected to evaluate the adhesion and proliferation state of cells in single PVA hydrogel and different SHK / PVA hydrogels, and the cell activity adhered to single PVA hydrogel and different SHK / PVA hydrogels was detected by cck-8 method.

[0079] The test method is as follows:

[0080] NIH3T3 cells (2 x 10 4The samples were seeded into 24-well cell culture plates containing hydrogel samples obtained in Examples 1-3 and Comparative Example 1. Meanwhile, untreated NIH3T3 cells were used as a blank group (i.e., Blank group). After culturing for 3 days in a sterile cell culture incubator with CO2 content of 5% and temperature of 37°C, the cell compatibility of the hydrogel was observed by live cell staining using a fluorescence inverted microscope.

[0081] Test results are as follows Figure 5 As shown, the SHK / PVA hydrogel dressing prepared in this invention is beneficial for cell adhesion, and the higher the SHK concentration, the greater the number of adherent cells. Simultaneously, on a single PVA hydrogel surface, cells exhibit an aggregated, adherent state. Cells cultured on an 80 SHK / PVA surface still appear in clusters, but the number and size of cell clusters adhering to the 80 SHK / PVA surface are significantly increased. In contrast, on the 160 SHK / PVA and 320 SHK / PVA hydrogel surfaces, cells exhibit a typical monodisperse, spindle-shaped morphology, indicating that the cells maintain good activity on the material surface. Furthermore, the cell proliferation efficiency on different SHK / PVA hydrogels was detected using the CCK-8 assay, and the results are as follows... Figure 6 As shown (* indicates p<0.05), the absorbance of the SHK / PVA hydrogel at 450 nm was significantly higher than that of the single PVA hydrogel and close to that of the blank group, indicating that the SHK / PVA hydrogel dressing prepared in this invention significantly promotes the proliferation of NIH3T3 cells, and the higher the SHK content, the stronger the NIH3T3 cell viability. The TCP group consisted of NIH3T3 cells cultured on TC-treated cell culture plates.

[0082] In vivo rat wound healing experiment

[0083] The testing method is as follows:

[0084] The wound-healing performance of the drug-loaded PVA hydrogel dressing prepared in this invention was evaluated using a rat dorsal full-thickness skin defect model. All animal experiments were approved by the Ethics Committee of the Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, and complied with the "Guidelines for the Care and Use of Laboratory Animals".

[0085] Experimental SD rats (purchased from Liaoning Changsheng Biotechnology Co., Ltd., 220-250g) were anesthetized with 1% sodium pentobarbital, and circular skin defects with a diameter of 14mm were created on the back of the rats. The SD rats were randomly divided into 5 groups according to the following treatment: blank group (i.e., Blank group), PVA group, 80SHK / PVA group, 160SHK / PVA group, and 320SHK / PVA group. The wounds of the four groups were covered with PVA hydrogel with a diameter of 14mm, 80SHK / PVA hydrogel, 160SHK / PVA hydrogel, and 320SHK / PVA hydrogel, respectively. The wounds of the blank group were bandaged with sterile gauze. The rats were housed individually after surgery, and their activity, diet, and health status were observed daily. The wound material was changed every 2 days. Subsequently, the wounds were photographed with a digital camera at 4, 7, 9, 12, and 14 days after surgery to observe wound healing and perform quantitative analysis.

[0086] Test results are as follows Figures 7-8 As shown, when SHK / PVA hydrogel is used as a dressing, the relative wound area of ​​rats gradually decreases from day 0 to day 14. The results indicate that the SHK / PVA hydrogel dressing prepared in this invention can significantly accelerate the wound healing process.

[0087] Biocompatibility testing

[0088] The testing method is as follows:

[0089] Rats in each group were sacrificed on days 3 and 7, and tissue specimens from their wounds were collected. Paraffin-embedded specimens were sectioned into sections approximately 5 μm thick, and immunohistochemical analysis of the expression of inflammatory factors IL-6 and TNF-α was performed according to previous research methods.

[0090] Test results are as follows Figure 9 As shown, the SHK / PVA hydrogel dressing prepared by this invention can effectively release SHK and significantly downregulate the immune response of tissues, indicating that the SHK / PVA hydrogel prepared by this invention has good biocompatibility.

[0091] The above description of the disclosed embodiments enables those skilled in the art to make or use the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A polyvinyl alcohol-based composite hydrogel, characterized by, is obtained by cross-linking shikonin and polyvinyl alcohol in an alkaline environment; the shikonin and polyvinyl alcohol are connected by hydrogen bonds, and the composite hydrogel releases shikonin when the hydrogen bonds are opened in a humid environment; the mass ratio of the shikonin to the polyvinyl alcohol is (0~3):(97~100); the polyvinyl alcohol has a polymerization degree of 1000~3000; the polyvinyl alcohol has an alcoholysis degree of 98~100%.

2. The method for preparing polyvinyl alcohol-based composite hydrogel according to claim 1, characterized in that, comprising the following steps: S1: drying polyvinyl alcohol solution to obtain polyvinyl alcohol hydrogel; S2: cross-linking shikonin and polyvinyl alcohol hydrogel in an alkaline environment to obtain polyvinyl alcohol-based composite hydrogel.

3. The preparation method according to claim 2, characterized in that, the drying temperature is 20~80℃.

4. The preparation method according to claim 2, characterized in that, the alkaline environment is provided by an alkaline solution; the alkaline solution is selected from any one or more of sodium hydroxide solution, potassium hydroxide solution, ammonia water or sodium ethoxide solution; The alkaline solution contains OH - The concentration is 0~6 mol / L.

5. The preparation method according to claim 2, characterized in that, the cross-linking time is 10~60 min.

6. The preparation method according to claim 2, characterized in that, the cross-linking is followed by a washing step.

7. The preparation method according to claim 6, characterized in that, the washing reagent is water; the washing is completed when the washing liquid is neutral.

8. A wound dressing, characterized in that, the polyvinyl alcohol-based composite hydrogel of claim 1 or prepared according to any one of claims 2~7.

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