Self-healing pH-sensitive hydrogel as well as preparation method and application thereof

Abstract

The invention discloses a self-healing pH-sensitive hydrogel and a preparation method and application thereof, the self-healing pH-sensitive hydrogel comprises a carrier and a bioactive compound loaded on the carrier, the carrier is a cationic guar gum / hyaluronic acid / zinc ion compound, and the bioactive compound is a ferulic acid / beta-cyclodextrin compound; the synergistic antibacterial effect is achieved through FA, Zn and CG, meanwhile, FA and CG provide oxidation resistance, the hydrogel is endowed with self-repairing performance through electrostatic interaction of carboxyl in HA and quaternary ammonium salt groups in CG, and the wound healing effect of the hydrogel is comprehensively improved.

Description

Technical Field

[0001] The present invention relates to the technical field of tissue engineering, and in particular to a self-healing pH-sensitive hydrogel and a preparation method and application thereof. Background Art

[0002] The pH value of normal skin is acidic, maintained at around 4-6, and this acidic environment can effectively protect the skin from microbial infection. However, once the skin is injured, the wound will undergo an acid-base-acid process as it heals, and eventually return to a typical acidic environment. However, continued microbial invasion can seriously hinder or delay the healing process after injury. At the same time, various substances released by pathogens may cause excessive, long-term chronic inflammation, seriously delaying the transition of wound healing to the next stage, turning it into a chronic wound. The chronic wound microenvironment presents the following characteristics: the accumulation of M1 macrophages increases the level of reactive oxygen and produces oxidative stress, leading to cell damage and dysregulation of the wound microenvironment. In addition, impaired angiogenesis affects the transport of oxygen and nutrients to the defective wound, thereby delaying wound healing. Therefore, it is ideal to design a wound dressing with antibacterial, anti-inflammatory and angiogenic functions to promote the healing of infected wounds.

[0003] The hydrogel matrix has a natural porous network structure and a suitable swelling ratio, mimicking the extracellular matrix (ECM), which can absorb exudate and cover defective skin, providing a moist environment for wound recovery. Hydrogels derived from polysaccharides have attracted widespread attention for their biosafety and versatility. Related Technology Cationic guar gum (CG) obtained by grafting and modifying guar gum shows new properties, especially the ability to achieve gelation by adjusting pH without introducing a cross-linking agent. More importantly, pure CG hydrogel can capture and adsorb through electrostatic interactions, thereby killing bacteria. At the same time, hyaluronic acid (HA) is selected for the construction of polysaccharide hydrogels for tissue engineering because it can balance the physiological environment of ECM by providing hydration, lubrication and matrix homeostasis.

[0004] However, the above-mentioned polysaccharide hydrogel cannot treat infected wounds due to the insufficient antibacterial properties of CG, and the wound healing speed as a dressing is slow, and the wound healing rate still needs to be improved. Summary of the invention

[0005] In view of this, the present application provides a self-healing pH-sensitive hydrogel and a preparation method and application thereof, which are used to solve the problem of how to improve the wound healing effect of the hydrogel.

[0006] In order to achieve the above technical objectives, this application adopts the following technical solutions: In a first aspect, the present application provides a self-healing pH-sensitive hydrogel, comprising a carrier and a bioactive complex loaded on the carrier, wherein the carrier is a cationic guar gum / hyaluronic acid / zinc ion complex, and the bioactive complex is a ferulic acid / β-cyclodextrin complex.

[0007] In a second aspect, the present application provides a method for preparing a self-healing pH-sensitive hydrogel, comprising the following steps: S1. dispersing β-cyclodextrin and ferulic acid in water, stirring for reaction, filtering, and freeze-drying to obtain a bioactive complex; S2. Disperse cationic guar gum and hyaluronic acid in water to obtain a CG-HA solution, then add the aqueous solution of the bioactive complex and the zinc ion solution to the CG-HA solution, stir and react, and obtain a self-healing pH-sensitive hydrogel dressing.

[0008] Preferably, the molar ratio of β-cyclodextrin to ferulic acid is 1-1.5:3.

[0009] Preferably, in the CG-HA solution, the concentration of cationic guar gum is 3-4wt%, and the concentration of hyaluronic acid is 2-2.5%.

[0010] Preferably, in step S2, the concentration of the aqueous solution of the bioactive complex is 1-2 wt %, and the volume of the aqueous solution of the bioactive complex added is 1-2 ml.

[0011] Preferably, in step S2, the concentration of the zinc ion solution is 2-2.5 mg / ml, and the added volume of the zinc ion solution is 1-2 ml.

[0012] Preferably, in step S1, the stirring reaction temperature is 20-35° C., and the stirring reaction time is 1-2 h.

[0013] Preferably, in step S2, the stirring reaction temperature is 20-35° C., and the stirring reaction time is 12-18 h.

[0014] Preferably, in step S1, the filtering method is: filtering using a 0.45 μm microporous filter membrane.

[0015] In a third aspect, the present application provides an application of a self-healing pH-sensitive hydrogel in the preparation of a wound dressing or an antibacterial agent.

[0016] The beneficial effects of this application are as follows: The hydrogel of the present application has self-healing, injectability, adaptability, pH sensitivity, antibacterial and antioxidant capabilities and good biocompatibility, so it can be used to close irregular wounds and eliminate bacteria present in the wound, which is beneficial to provide a good repair environment for the wound.

[0017] In the self-healing pH-sensitive hydrogel (FA / β-CD@CG-HA-Zn) prepared in the present application, FA, Zn and CG achieve synergistic antibacterial effects. At the same time, FA and CG provide antioxidant properties. The electrostatic interaction between the carboxyl group in HA and the quaternary ammonium salt group in CG gives the hydrogel self-healing properties, thereby improving the wound healing effect.

[0018] The self-healing pH-sensitive hydrogel of the present application has a bioactive complex FA / β-CD, and uses CG-HA-Zn hydrogel as a stable carrier to improve the bioavailability of FA / β-CD, thereby improving the antibacterial and wound healing effects. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 is a scanning electron micrograph of the self-healing pH-sensitive hydrogel of Example 1; Figure 2 This is a diagram showing the effect of the self-healing pH-sensitive hydrogel of Example 1 on Escherichia coli and Staphylococcus aureus; Figure 3 This is a bar graph showing the efficiency of the self-healing pH-sensitive hydrogel in Example 1 in promoting healing of infected wounds; Figure 4 This is an injectable diagram of the self-healing pH-sensitive hydrogel of Example 1; Figure 5 This is the adaptive diagram of the self-healing pH-sensitive hydrogel of Example 1. DETAILED DESCRIPTION

[0020] In order to make the purpose, technical solution and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with the embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0021] Passive drug delivery may lead to a burst release of bioactive substances, causing unnecessary cytotoxicity and affecting wound healing. During the healing process, many factors (such as temperature and pH) affect wound healing. Advanced hydrogel wound dressings should respond to these changes and have a positive impact on the wound environment. The microenvironment of chronic infected wounds is alkaline. However, most current pH-responsive hydrogel wound dressings release more bioactive substances under acidic conditions, and there are relatively few reports on release in the alkaline chronic wound pH microenvironment. Therefore, it is necessary to construct a self-healing, injectable and adaptive pH-responsive hydrogel to address the problems that ordinary hydrogel wound dressings are difficult to combine multiple therapeutic properties (antibacterial, antioxidant) and lack sustained-release drugs in response to the alkaline microenvironment of chronic wounds.

[0022] Based on this, this application was created.

[0023] The present application provides a self-healing pH-sensitive hydrogel, comprising a carrier and a bioactive complex loaded on the carrier, wherein the carrier is a cationic guar gum / hyaluronic acid / zinc ion complex, and the bioactive complex is a ferulic acid / β-cyclodextrin complex.

[0024] The self-healing pH-sensitive hydrogel of the present application can release ferulic acid and Zn according to the pH response of the microenvironment at the site of chronic wounds, thereby removing bacteria, inhibiting inflammation, promoting angiogenesis and re-epithelialization, and finally achieving rapid healing of chronic wounds; wherein, Zn 2+ It can bind to the negatively charged bacterial surface through electrostatic force, causing damage to the bacterial cell membrane. In addition, the complexation of zinc ions with hydroxyl and carboxyl groups can enhance the crosslinking density of the hydrogel and accelerate the gelation process. Ferulic acid has antioxidant and antibacterial biological activities, and regulates macrophage polarization, reduces the level of oxidative stress, and thus promotes tissue regeneration. The active substances in this application (zinc ions, ferulic acid, cationic guar gum, hyaluronic acid) are synergistically encapsulated in the hydrogel system for sustained release, thereby extending the retention time of the active substances in the wound and enhancing their therapeutic effects.

[0025] The present application provides a method for preparing a self-healing pH-sensitive hydrogel, comprising the following steps: S1. dispersing β-cyclodextrin and ferulic acid in water, stirring for reaction, filtering, and freeze-drying to obtain a bioactive complex; S2. Disperse cationic guar gum and hyaluronic acid in water to obtain a CG-HA solution, then add the aqueous solution of the bioactive complex and the zinc ion solution to the CG-HA solution, stir and react, and obtain a self-healing pH-sensitive hydrogel dressing.

[0026] In some embodiments, the molar ratio of β-cyclodextrin to ferulic acid is 1-1.5:3.

[0027] In some embodiments, in the CG-HA solution, the concentration of cationic guar gum is 3-4 wt %, and the concentration of hyaluronic acid is 2-2.5%.

[0028] In some embodiments, in step S2, the concentration of the aqueous solution of the bioactive complex is 1-2 wt %, and the volume of the aqueous solution of the bioactive complex added is 1-2 ml.

[0029] In some embodiments, in step S2, the concentration of the zinc ion solution is 2-2.5 mg / ml, and the added volume of the zinc ion solution is 1-2 ml.

[0030] In some embodiments, in step S1, the stirring reaction temperature is 20-35° C., and the stirring reaction time is 1-2 h.

[0031] In some embodiments, in step S2, the stirring reaction temperature is 20-35° C., and the stirring reaction time is 12-18 h.

[0032] In some embodiments, in step S1, the filtering method is: filtering using a 0.45 μm microporous filter membrane.

[0033] Specifically, the preparation method of the self-healing pH sensitive hydrogel comprises the following steps: S1. Dissolve β-cyclodextrin (β-CD) and ferulic acid (FA) in 100 mL of deionized water at a molar ratio of 1-1.5:3, and then stir at room temperature of 20-35°C for 1-2 hours. After the reaction, filter the reaction solution with a 0.45 μm microporous filter membrane and freeze-dry it to obtain a white solid, i.e., the bioactive complex (FA / β-CD). S2. Dissolve FA / β-CD in deionized water to obtain a 1-2wt% aqueous solution of the bioactive complex (FA / β-CD solution); disperse cationic guar gum (CG) and hyaluronic acid (HA) in deionized water, and stir at room temperature for 12 hours to obtain a CG-HA solution (wherein the concentration of CG is 3-4 wt% and the concentration of HA is 2-2.5 wt%); add 1-2ml of the 1-2wt% FA / β-CD solution and 1-2ml of the 2-2.5 mg / mL zinc ion solution to the above CG-HA solution, mix into a gel at room temperature, and obtain a self-healing pH-sensitive hydrogel dressing (FA / β-CD@CG-HA-Zn).

[0034] The present application provides an application of a self-healing pH-sensitive hydrogel in the preparation of a wound dressing or an antibacterial agent.

[0035] The self-healing pH-sensitive hydrogel of the present application has excellent self-healing properties, injectability, adaptability, pH sensitivity, significant antibacterial properties, excellent antioxidant capacity, and good biocompatibility. The hydrogel of the present application can provide a good repair environment for wounds, accelerate the rapid healing of bacterial-infected wounds, and can be used in the treatment and management of wounds.

[0036] The present solution is further described below through specific embodiments.

[0037] Example 1 A method for preparing a self-healing pH-sensitive hydrogel comprises the following steps: S1. β-CD and FA were dissolved in 100 mL of deionized water at a mass ratio of 1:3, and then stirred at 25°C for 1 hour. After the reaction, the reaction solution was filtered with a 0.45 μm microporous filter membrane and freeze-dried to obtain a bioactive complex (FA / β-CD); S2. 0.6 g of cationic guar gum (CG) and 0.4 g of hyaluronic acid (HA) were dispersed in 20 g of deionized water and stirred at room temperature for 1 hour to obtain a CG-HA solution (wherein the concentration of CG was 3 wt% and the concentration of HA was 2 wt%). 1 ml of a 1 wt% FA / β-CD solution and 1 ml of a 2 mg / mL zinc ion solution were added to the CG-HA solution (wherein the concentration of CG was 3 wt% and the concentration of HA was 2 wt%), stirred at 25°C for 12 hours, and mixed into a gel to obtain a self-healing pH-sensitive hydrogel dressing (FA / β-CD@CG-HA-Zn).

[0038] The scanning electron microscope image of the product obtained in this example is shown in Figure 1 , indicating that a hydrogel with a three-dimensional network structure was obtained.

[0039] Example 2 A method for preparing a self-healing pH-sensitive hydrogel, the other contents are the same as those in Example 1, except that 2 ml of a 1 wt % FA / β-CD solution and 2 ml of a 2 mg / mL zinc ion solution are added to a CG-HA solution (wherein the CG concentration is 3 wt % and the HA is 2 wt %).

[0040] Example 3 A method for preparing a self-healing pH-sensitive hydrogel, the other contents are the same as those in Example 1, except that 1 ml of a 1 wt % FA / β-CD solution and 1 ml of a 2 mg / mL zinc ion solution are added to a CG-HA solution (wherein the CG concentration is 4 wt % and the HA concentration is 2 wt %).

[0041] Example 4 A method for preparing a self-healing pH-sensitive hydrogel, the other contents are the same as those in Example 1, except that 2 ml of a 1 wt % FA / β-CD solution and 2 ml of a 2 mg / mL zinc ion solution are added to a CG-HA solution (wherein the CG concentration is 4 wt % and the HA is 2 wt %).

[0042] Comparative Example 1 A method for preparing a hydrogel comprises the following steps: dispersing 0.6 g of cationic guar gum (CG) and 0.4 g of hyaluronic acid (HA) in 20 g of deionized water, stirring at room temperature for 1 hour to obtain a CG-HA solution (wherein the concentration of CG is 3 wt % and the concentration of HA is 2 wt %), and then allowing the solution to stand to obtain a hydrogel (CG-HA).

[0043] Comparative Example 2 A method for preparing a self-healing pH-sensitive hydrogel, the other contents of which are the same as those of Example 1, except that the method does not include adding a zinc ion solution.

[0044] Comparative Example 3 A method for preparing a self-healing pH-sensitive hydrogel, the other contents of which are the same as those of Example 1, except that ferulic acid is not added.

[0045] Test and Evaluation The hydrogel of Example 1 was used as the test object to conduct an antibacterial experiment: The hydrogel prepared in Example 1 was added to a 24-well cell culture plate. Sterilized with ultraviolet light for 24 hours, 1 mL of the diluted bacterial suspension (10 8 CFU / mL) was added to the sterilized hydrogel, and then two groups were selected as the control group. 1 mL of diluted bacterial suspension was added to the 24-cell culture plate without hydrogel in the control group one, and 1 mL of bacterial suspension was added to the 24-cell culture plate without hydrogel in the control group two; then, the three groups of samples were cultured in an oscillating incubator at 100 r / min and 37°C for 2 hours, each group of materials was washed with 1ml of sterile PBS, and then 100 μL of bacterial suspension was evenly dispersed on the agar plate and cultured in a constant temperature oscillating incubator for 12 hours. Finally, the antibacterial properties of the material were analyzed by taking pictures and counting the colonies on the agar plate. The results are shown in Figure 2 shown.

[0046] Depend on Figure 2 It can be seen that the hydrogel prepared in the present application has excellent antibacterial properties.

[0047] The hydrogel of Example 3 was used as the test object, and the comparative example 1 and the blank group were used as controls to test the wound healing effect: First, an infected wound model was established. Mice were anesthetized with 3% sodium pentobarbital, and the hair on the back of the anesthetized mice was shaved. Then, a full-thickness wound with a diameter of about 10 mm was prepared on the back of the mouse, and 40 μL of Staphylococcus aureus solution was added. After culturing for 24 hours, the bacterial infected wound model was successfully established. The experiment was divided into three groups. The first group was a control group without any treatment, the second group used CG-HA hydrogel to cover the wound, and the third group used FA / β-CD@CG-HA-Zn composite hydrogel in Example 3 to cover the wound. The wound healing rate was calculated on the 3rd, 7th and 14th days. The results are shown in Figure 3 shown.

[0048] Depend on Figure 3 It can be seen that the FA / β-CD@CG-HA-Zn composite hydrogel prepared in the present application can effectively promote the healing of bacterial-infected wounds.

[0049] Phenol red dye was added to FA / β-CD@CG-HA-Zn hydrogel to obtain red-dyed hydrogel. The original hydrogel and phenol red-dyed hydrogel were cut into hydrogel strips of the same size. The prepared red-dyed hydrogel strips were fully contacted with the original hydrogel strips, and the self-healing behavior within 1 hour was observed. Figure 4 This is an injectable image of the self-healing pH-sensitive hydrogel of the present application obtained in Example 1.

[0050] Depend on Figure 4 It can be seen that the FA / β-CD@CG-HA-Zn composite hydrogel prepared in the present application has good self-healing behavior.

[0051] Place 2 mL of FA / β-CD@CG-HA-Zn hydrogel into a specific gravity bottle to observe whether the hydrogel can fill the gaps in the glass spheres. Figure 5 This is the adaptive graph of the self-healing pH-sensitive hydrogel of the present application obtained in Example 1.

[0052] Depend on Figure 5 It can be seen that the FA / β-CD@CG-HA-Zn composite hydrogel prepared in the present application can completely fill the gaps in the glass spheres, proving that it has good adaptive ability.

[0053] The above are only preferred specific implementations of the present invention, but the protection scope of the present invention is not limited thereto. Any changes or substitutions that can be easily conceived by any technician familiar with the technical field within the technical scope disclosed by the present invention should be covered within the protection scope of the present invention.

Claims

1. A self-healing pH-sensitive hydrogel, characterized in that: The invention comprises a carrier and a bioactive complex loaded on the carrier. The carrier is a cationic guar gum / hyaluronic acid / zinc ion complex, and the bioactive complex is a ferulic acid / β-cyclodextrin complex.

2. A method for preparing the self-healing pH sensitive hydrogel according to claim 1, characterized in that: The following steps are involved: S1. dispersing β-cyclodextrin and ferulic acid in water, stirring for reaction, filtering, and freeze-drying to obtain the bioactive complex; S2. Disperse cationic guar gum and hyaluronic acid in water to obtain a CG-HA solution, then add the aqueous solution of the bioactive complex and the zinc ion solution to the CG-HA solution, stir and react, and obtain the self-healing pH-sensitive hydrogel dressing.

3. The method for preparing the self-healing pH sensitive hydrogel according to claim 2, characterized in that: The molar ratio of the beta-cyclodextrin to ferulic acid is 1-1.5:

3.

4. The method for preparing the self-healing pH sensitive hydrogel according to claim 2, characterized in that: In the CG-HA solution, the concentration of cationic guar gum is 3-4wt%, and the concentration of hyaluronic acid is 2-2.5%.

5. The method for preparing the self-healing pH sensitive hydrogel according to claim 2, characterized in that: In step S2, the concentration of the aqueous solution of the bioactive complex is 1-2 wt %, and the volume of the aqueous solution of the bioactive complex added is 1-2 ml.

6. The method for preparing the self-healing pH sensitive hydrogel according to claim 2, characterized in that: In step S2, the concentration of the zinc ion solution is 2-2.5 mg / ml, and the added volume of the zinc ion solution is 1-2 ml.

7. The method for preparing the self-healing pH sensitive hydrogel according to claim 2, characterized in that: In step S1, the stirring reaction temperature is 20-35° C., and the stirring reaction time is 1-2 h.

8. The method for preparing the self-healing pH sensitive hydrogel according to claim 2, characterized in that: In step S2, the stirring reaction temperature is 20-35° C., and the stirring reaction time is 12-18 h.

9. The method for preparing the self-healing pH sensitive hydrogel according to claim 2, characterized in that: In step S1, the filtering method is: filtering using a 0.45 μm microporous filter membrane.

10. Use of the self-healing pH-sensitive hydrogel obtained by the preparation method according to any one of claims 2 to 9 in the preparation of wound dressings or antibacterial agents.

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