Preparation method and application of multi-level pore structure compound traditional Chinese medicine drug-loaded hydrogel

Abstract

The application belongs to the technical field of functional materials, and relates to a preparation method and application of a compound traditional Chinese medicine drug-loaded hydrogel with a hierarchical pore structure. The compound traditional Chinese medicine drug-loaded hydrogel with the hierarchical pore structure comprises the following components in parts by weight: 10-30 parts of carboxymethyl chitosan, 5-20 parts of oxidized sodium alginate, 3-5 parts of tannic acid, 0.5-1 part of baicalin, and 200-500 parts of water. The compound traditional Chinese medicine drug-loaded hydrogel with the hierarchical pore structure has low toxicity and high biological safety; the gel has a porous structure and good drug release effect after being treated with sodium borohydride; the gel has multiple effects such as antibacterial effect, anti-inflammatory effect, antioxidant effect, antitumor effect and antiviral effect; and the gel can be prepared at room temperature, and the operation process is simple.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of functional materials, and relates to a preparation method and application of a compound traditional Chinese medicine drug-loaded hydrogel with a hierarchical porous structure. BACKGROUND

[0002] In modern society, people's fast-paced life and unhealthy lifestyle can easily cause a series of chronic diseases, such as cardiovascular disease, diabetes, cancer, etc. These diseases often require long-term treatment and management, which seriously affects the quality of life of patients. For the treatment of such diseases, western medicine has a quick effect, but long-term use undoubtedly causes harm to the body. Traditional Chinese medicine has small toxic and side effects, and also has good health care and preventive effects. Among them, the compound preparation of traditional Chinese medicine can make multiple components of traditional Chinese medicine synergistically enhance the efficacy. Some studies have shown that there are interaction modes between active ingredients of some medicinal materials. For example, coptis and scutellaria are a pair of representative herbs traditionally used to treat gastrointestinal diseases. Berberine in coptis and baicalin can assemble into nanostructures in water, showing excellent antibacterial activity. There is various evidence that scutellaria extract has a good therapeutic effect on colonic inflammation, which has been confirmed in various models and clinical trials. Tannic acid is a natural polyphenol, which exists in many traditional Chinese medicines such as red wine, black tea, green tea and rhubarb, and has a protective effect on acute or chronic intestinal inflammation. However, traditional Chinese medicine decoction often has the disadvantages of complex decoction process and unsuitable for long-term preservation. Therefore, the combination of hydrogel and compound preparation of traditional Chinese medicine can make up for the shortcomings of traditional Chinese medicine decoction to some extent.

[0003] Carboxymethyl chitosan is a water-soluble polysaccharide derivative obtained by chemical modification of chitosan. Compared with chitosan, it has better water solubility, ion exchange capacity and biological activity. The amino groups in its molecular structure provide active sites for Schiff base reaction, which can react with various compounds to form hydrogels. In addition, carboxymethyl chitosan has the advantages of biodegradability and biocompatibility, which makes it widely used in the field of biomedicine.

[0004] Compared with traditional hydrogels, porous structure hydrogels greatly improve the water absorption rate and water absorption ratio of hydrogels. The excellent performance of large pore structure, air permeability and short diffusion path enables it to have the ability of rapid transmission of liquid, gas and biomolecule fixation media. This is very advantageous for application occasions that require rapid material exchange, such as water treatment and biomedical applications.

[0005] Therefore, it is necessary to develop a new method to combine traditional Chinese medicine compound preparations with hydrogels and obtain a compound traditional Chinese medicine drug-loaded hydrogel material rich in hierarchical porous structure. SUMMARY

[0006] In view of the deficiencies of the prior art, the application provides a multi-level pore structure hydrogel compound traditional Chinese medicine drug delivery system and a preparation method and application thereof.The preparation method is simple, and the use of biological polysaccharide materials such as carboxymethyl chitosan, sodium alginate and tannic acid has the characteristics of low toxicity and high biological safety, wherein the tannic acid serves as a crosslinking agent and is also a natural medicine with excellent antibacterial capacity. The hydrogel is treated by NaBH4 to form a multi-level pore structure, thereby having excellent drug release capacity and excellent antibacterial effect.

[0007] In order to achieve the above-mentioned application purposes, the application provides the following technical scheme:

[0008] A multi-level pore structure compound traditional Chinese medicine drug delivery hydrogel comprises the following components in parts by weight: 10-30 parts of carboxymethyl chitosan, 5-20 parts of oxidized sodium alginate, 3-5 parts of tannic acid, 0.5-1 part of baicalin and 200-500 parts of water.

[0009] Preferably, the multi-level pore structure compound traditional Chinese medicine drug delivery hydrogel is composed of the following components in parts by weight: 20 parts of carboxymethyl chitosan, 10 parts of carboxymethyl chitosan oxidized sodium alginate, 4 parts of tannic acid, 1 part of baicalin and 200 parts of water.

[0010] The preparation method of the multi-level pore structure compound traditional Chinese medicine drug delivery hydrogel comprises the following steps:

[0011] (1) In a solvent A, sodium alginate and a sodium periodate aqueous solution are reacted to prepare oxidized sodium alginate.

[0012] (2) Carboxymethyl chitosan, tannic acid and baicalin are dissolved in distilled water to obtain a mixed solution; the oxidized sodium alginate prepared in step (1) is dissolved in distilled water to obtain an oxidized sodium alginate solution; the mixed solution and the oxidized sodium alginate solution are uniformly mixed, and the mixture is placed at 4 DEG C to obtain a hydrogel.

[0013] (3) The hydrogel is soaked in a sodium borohydride solution for reaction to obtain a multi-level pore structure hydrogel.

[0014] Preferably, in step (1), the solvent A is methanol, ethanol or acetone; further preferably, the solvent A is ethanol; the sodium alginate is added into the solvent A to obtain a sodium alginate suspension, and the mass of the sodium alginate to the volume of the solvent A is 0.1-0.3 g / mL.

[0015] Preferably, in step (1), the mass of the sodium periodate in the sodium periodate aqueous solution to the volume of the distilled water is 0.1-0.3 g / mL.

[0016] Further preferably, in step (1), the volume ratio of the sodium alginate suspension to the sodium periodate aqueous solution is 1:0.5-1.

[0017] Preferably, in step (1), the reaction temperature is room temperature, the reaction condition is light shielding, and the reaction time is 6-18 h.

[0018] Preferably, in step (1), an ethylene glycol with the same molar amount as the sodium periodate is added when the reaction is terminated.

[0019] Preferably, in step (1), after the reaction is completed, the reaction mixture is poured into a large amount of anhydrous ethanol under vigorous stirring, and the volume ratio of the reaction mixture to the anhydrous ethanol is 1:5-8 to obtain a precipitate.

[0020] Further preferably, in step (1), the obtained precipitate is suction filtered and vacuum dried at 40℃, and the obtained solid powder is dialyzed with distilled water for 24 h.

[0021] Preferably, in step (2), the mass of the carboxymethyl chitosan and the volume of the distilled water in the mixed solution are in a ratio of 0.1-0.3 g / mL; the mass of the tannic acid and the volume of the distilled water in the mixed solution are in a ratio of 0.04-0.08 g / mL; and the mass of the baicalin and the volume of the distilled water in the mixed solution are in a ratio of 0.001-0.02 g / mL.

[0022] Preferably, in step (2), the mass fraction of the oxidized sodium alginate solution is 5-10%.

[0023] Preferably, in step (2), the volume ratio of the mixed solution to the oxidized sodium alginate solution is 1:0.5-1.5.

[0024] Preferably, in step (3), the mass fraction of the sodium borohydride solution is 0.1-2%.

[0025] Preferably, in step (3), the reaction time is 2-5 min.

[0026] The application further discloses an application of the compound traditional Chinese medicine drug-loaded hydrogel with a multi-level pore structure in antibacterial materials.

[0027] Compared with the prior art, the application has the following beneficial effects:

[0028] (1) The multi-level pore structure compound traditional Chinese medicine drug-loaded hydrogel has low toxicity and high biological safety. The carboxymethyl chitosan, oxidized sodium alginate and tannic acid used in the application all belong to biological materials, and have the advantages of non-toxicity and high biological compatibility. The hydrogel extract solution is co-cultured with mouse fibroblasts (L929 cells) for 24 hours, and the survival rate of the cells is tested by CKK-8 experiment. The results show that the multi-level pore structure compound traditional Chinese medicine drug-loaded hydrogel has low biological toxicity.

[0029] (2) The multi-level pore structure compound traditional Chinese medicine drug-loaded hydrogel has good drug release effect. The gel has a porous structure after being treated by sodium borohydride, and has better drug release effect. The gel is placed in a PBS solution, and the drug release amount is measured after different time. The results show that the drug release rate of the hydrogel treated by sodium borohydride is about twice that of the hydrogel without sodium borohydride treatment, and the release rate is also significantly higher than that of the untreated hydrogel.

[0030] (3) The multi-level pore structure compound traditional Chinese medicine drug-loaded hydrogel has good antibacterial effect. The tannic acid / baicalin compound traditional Chinese medicine coated by the hydrogel has antibacterial, anti-inflammatory, antioxidant, antitumor and antiviral effects, and has wide pharmacological activity and clinical application potential. Staphylococcus aureus is uniformly coated on LB solid culture medium, and the sterilized sample is attached to the surface of the culture medium and pressed gently with sterile tweezers to make the sample fully contact with the culture medium. The culture dish is placed in a constant temperature incubator at 37 DEG C and cultured for 24 hours. The antibacterial test results show that the inhibition ring diameter is about 28 mm, and has obvious antibacterial effect.

[0031] (4) The preparation method of the multi-level pore structure compound traditional Chinese medicine drug-loaded hydrogel is simple. The hydrogel is prepared by Schiff base reaction between carboxymethyl chitosan and oxidized sodium alginate, and hydrogen bond action between carboxymethyl chitosan and tannic acid. The hydrogel can be prepared in a few seconds at room temperature, and the reaction condition is mild and the operation process is simple. BRIEF DESCRIPTION OF DRAWINGS

[0032] Figure 1 The scanning electron microscope pictures of the hydrogels prepared in example 1 and comparative examples 1, 2 and 3 are shown in the following table:

[0033] Figure 2 The electron microscope pictures of the hydrogels prepared in example 1 and comparative examples 1, 2 and 3 are shown in the following table:

[0034] Figure 3 The drug release rate comparison chart of the hydrogels in example 1 and comparative example 1 in different pH simulated body fluids is shown in the following table:

[0035] Figure 4Figure for comparison of drug release rate of hydrogel in different pH simulated body fluid in Comparative Example 2 and Comparative Example 3;

[0036] Figure 5 Figure for cell survival rate of hydrogel prepared in Example 1 and Comparative Examples 1-3;

[0037] Figure 6 Figure for antibacterial experiment results of hydrogel in Example 1, Comparative Example 1, Comparative Example 2 and Comparative Example 3. DETAILED DESCRIPTION

[0038] The application will be further described in connection with the specific embodiments, but not limited thereto.

[0039] Meanwhile, the experimental methods described in the following examples are all conventional methods unless otherwise specified; and the reagents and materials, unless otherwise specified, can be obtained from commercial channels.

[0040] Example 1

[0041] A drug delivery system of a multi-level porous structure hydrogel compound traditional Chinese medicine and a preparation method thereof, the preparation method comprising the following steps:

[0042] (1) Preparation of aldehyde-modified sodium alginate (OSA):

[0043] 10 g of sodium alginate was dispersed in 50 mL of anhydrous ethanol to prepare a suspension I. 10 g of sodium periodate was dissolved in 50 mL of water to prepare a solution II. The solution II was added to the suspension I, and the mixture was magnetically stirred at room temperature overnight in the dark. An equal molar amount of ethylene glycol was added to terminate the reaction. The reaction mixture was poured into a large amount of anhydrous ethanol under vigorous stirring (V (reaction mixture): V (ethanol) = 1:5) to precipitate the product. The precipitate was filtered and vacuum dried at 40°C. The obtained solid powder was dialyzed against distilled water for 24 h to remove unreacted sodium periodate and ethylene glycol and other small molecular impurities. The liquid in the dialysis bag was freeze-dried to obtain a white flocculent product, which was aldehyde-modified sodium alginate.

[0044] (2) Preparation of the drug delivery system of the hydrogel:

[0045] 0.1 g of carboxymethyl chitosan, 0.02 g of tannic acid and 0.005 g of baicalin were dissolved in 0.5 mL of distilled water to prepare a solution IV. 1 g of aldehyde-modified sodium alginate prepared in step (1) was dissolved in 9 g of distilled water to prepare a 10% aldehyde-modified sodium alginate solution. The solution IV and the 10% aldehyde-modified sodium alginate solution were mixed in equal proportions and uniformly, and then the mixture was allowed to stand at 4°C to obtain the hydrogel.

[0046] (3) The hydrogel prepared in step (2) was soaked in 1% NaBH4 for 5 min to obtain a multi-level porous structure hydrogel.

[0047] The multi-level porous hydrogel is composed of the following components in parts by weight: 20 parts of carboxymethyl chitosan, 10 parts of oxidized sodium alginate, 4 parts of tannic acid, 1 part of baicalin, and 200 parts of water.

[0048] Example 2

[0049] A multi-level porous hydrogel compound traditional Chinese medicine drug delivery system and a preparation method thereof, the preparation method comprising the following steps:

[0050] (1) Preparation of aldehyde-modified sodium alginate (OSA):

[0051] 10 g of sodium alginate was dispersed in 50 mL of anhydrous ethanol to prepare a suspension I. 10 g of sodium periodate was dissolved in 50 mL of water to prepare a solution II. Solution II was added to suspension I, and the mixture was magnetically stirred at room temperature overnight in the dark. An equimolar amount of ethylene glycol was added to terminate the reaction. The reaction mixture was poured into a large amount of vigorously stirred anhydrous ethanol (V (reaction mixture): V (ethanol) = 1:5) to precipitate the product. The precipitate was filtered and dried at 40°C under vacuum. The resulting solid powder was dialyzed against distilled water for 24 hours to remove unreacted sodium periodate and ethylene glycol and other small molecular impurities. The liquid in the dialysis bag was freeze-dried to obtain a white flocculent product, which was aldehyde-modified sodium alginate.

[0052] (2) Preparation of the hydrogel drug delivery system:

[0053] 0.1 g of carboxymethyl chitosan, 0.04 g of tannic acid, and 0.005 g of baicalin were dissolved in 0.5 mL of distilled water to prepare solution IV. 1 g of aldehyde-modified sodium alginate prepared in step (1) was dissolved in 9 g of distilled water to prepare a 10% aldehyde-modified sodium alginate solution. Solution IV was mixed with the 10% aldehyde-modified sodium alginate solution in equal proportions and stirred until homogeneous. The mixture was then allowed to stand at 4°C to obtain the hydrogel.

[0054] (3) The hydrogel prepared in step (2) was soaked in 1% NaBH4 for 5 minutes to obtain a multi-level porous hydrogel.

[0055] The multi-level porous hydrogel is composed of the following components in parts by weight: 20 parts of carboxymethyl chitosan, 10 parts of oxidized sodium alginate, 4 parts of tannic acid, 1 part of baicalin, and 200 parts of water.

[0056] Example 3

[0057] A multi-level porous hydrogel compound traditional Chinese medicine drug delivery system and a preparation method thereof, the preparation method comprising the following steps:

[0058] (1) Preparation of aldehyde-modified sodium alginate (OSA):

[0059] A suspension I was prepared by dispersing 10 g of sodium alginate in 50 mL of absolute ethanol. A solution II was prepared by dissolving 10 g of sodium periodate in 50 mL of water. The solution II was added to the suspension I, and the mixture was magnetically stirred at room temperature overnight in the dark. The reaction was terminated by adding ethylene glycol in an amount equivalent to the amount of sodium periodate. The reaction mixture was poured into a large amount of absolute ethanol (V (reaction mixture) : V (ethanol) = 1:5) under vigorous stirring to precipitate a solid. The precipitate was suction filtered and dried under vacuum at 40°C. The obtained solid powder was dialyzed against distilled water for 24 h to remove unreacted sodium periodate and ethylene glycol and other small molecular impurities. The liquid in the dialysis bag was freeze-dried to obtain a white flocculent product, which was aldehyde-modified sodium alginate.

[0060] (2) Preparation of a hydrogel drug delivery system:

[0061] A solution IV was prepared by dissolving 0.1 g of carboxymethyl chitosan, 0.02 g of tannic acid and 0.005 g of baicalin in 0.5 mL of distilled water. A 5% aldehyde-modified sodium alginate solution was prepared by dissolving 0.5 g of the aldehyde-modified sodium alginate prepared in step (1) in 9.5 g of distilled water. The solution IV and the 5% aldehyde-modified sodium alginate solution were mixed in equal proportions and uniformly mixed, and the mixture was allowed to stand at 4°C to obtain a hydrogel.

[0062] (3) The hydrogel prepared in step (2) was soaked in 1% NaBH4 for 5 min to obtain a hydrogel with a hierarchical porous structure.

[0063] The hydrogel with a hierarchical porous structure comprises the following components in parts by weight: 20 parts of carboxymethyl chitosan, 5 parts of oxidized sodium alginate, 4 parts of tannic acid, 1 part of baicalin and 205 parts of water.

[0064] Comparative Example 1

[0065] A drug delivery system of a hydrogel compound traditional Chinese medicine with a hierarchical porous structure and a preparation method thereof, comprising the following steps:

[0066] (1) Preparation of aldehyde-modified sodium alginate (OSA):

[0067] A suspension Ⅰ was prepared by dispersing 10 g of sodium alginate in 50 mL of absolute ethanol. A solution Ⅱ was prepared by dissolving 10 g of sodium periodate in 50 mL of water. The solution Ⅱ was added to the suspension Ⅰ, and the mixture was stirred magnetically overnight at room temperature in the dark. The reaction was terminated by adding ethylene glycol in an amount equivalent to that of sodium periodate. The reaction mixture was poured into a large amount of absolute ethanol (V (reaction mixture) : V (ethanol) = 1 : 5) stirred vigorously to precipitate the product. The precipitate was filtered and dried at 40°C under vacuum. The solid powder obtained was dialyzed against distilled water for 24 h to remove unreacted sodium periodate and ethylene glycol and other small molecular impurities. The liquid in the dialysis bag was freeze-dried to obtain a white flocculent product, which was aldehyde-modified sodium alginate.

[0068] (2) Preparation of the hydrogel drug delivery system:

[0069] A solution Ⅲ was prepared by dissolving 0.1 g of carboxymethyl chitosan, 0.02 g of tannic acid and 0.005 g of baicalin in 0.5 mL of distilled water. A 10% aldehyde-modified sodium alginate solution was prepared by dissolving 1 g of the aldehyde-modified sodium alginate prepared in step (1) in 9 g of distilled water. The solution Ⅲ and the 10% aldehyde-modified sodium alginate solution were mixed in equal proportions and allowed to stand at 4°C to obtain a hydrogel.

[0070] Comparative Example 2

[0071] A hydrogel compound traditional Chinese medicine drug delivery system and a preparation method thereof, comprising the steps of:

[0072] (1) Preparation of aldehyde-modified sodium alginate (OSA):

[0073] A suspension Ⅰ was prepared by dispersing 10 g of sodium alginate in 50 mL of absolute ethanol. A solution Ⅱ was prepared by dissolving 10 g of sodium periodate in 50 mL of water. The solution Ⅱ was added to the suspension Ⅰ, and the mixture was stirred magnetically overnight at room temperature in the dark. The reaction was terminated by adding ethylene glycol in an amount equivalent to that of sodium periodate. The reaction mixture was poured into a large amount of absolute ethanol (V (reaction mixture) : V (ethanol) = 1 : 5) stirred vigorously to precipitate the product. The precipitate was filtered and dried at 40°C under vacuum. The solid powder obtained was dialyzed against distilled water for 24 h to remove unreacted sodium periodate and ethylene glycol and other small molecular impurities. The liquid in the dialysis bag was freeze-dried to obtain a white flocculent product, which was aldehyde-modified sodium alginate.

[0074] (2) Preparation of the hydrogel drug delivery system:

[0075] 0.1 g carboxymethyl chitosan, 0.02 g tannic acid is dissolved in 0.5 mL distilled water, recorded as solution III. 1 g of aldehyde sodium alginate prepared in step (1) is dissolved in 9 g distilled water to prepare a 10% aldehyde sodium alginate solution. Solution III and 10% aldehyde sodium alginate solution are mixed in equal proportions and uniformly placed at 4°C to obtain a hydrogel.

[0076] (3) The hydrogel prepared in step (2) is soaked in 1% NaBH4 for 5 min to obtain a hierarchical porous hydrogel.

[0077] Comparative Example 3

[0078] A drug delivery system of a hydrogel compound traditional Chinese medicine and a preparation method thereof, comprising the steps of:

[0079] (1) Preparation of aldehyde sodium alginate (OSA):

[0080] 10 g sodium alginate is dispersed in 50 mL anhydrous ethanol to prepare a suspension I. 10 g sodium periodate is dissolved in 50 mL water to prepare a solution II. Solution II is added to suspension I, and the reaction is terminated by adding ethylene glycol in an amount equal to that of sodium periodate under magnetic stirring at room temperature overnight. The reaction mixture is poured into a large amount of anhydrous ethanol under vigorous stirring (V (reaction mixture): V (ethanol) = 1:5) to precipitate the product. The precipitate is filtered and vacuum dried at 40°C. The obtained solid powder is dialyzed against distilled water for 24 h to remove unreacted sodium periodate and ethylene glycol and other small molecular impurities. The liquid in the dialysis bag is freeze-dried to obtain a white flocculent product, which is aldehyde sodium alginate.

[0081] (2) Preparation of a hydrogel drug delivery system:

[0082] 0.1 g carboxymethyl chitosan, 0.02 g tannic acid is dissolved in 0.5 mL distilled water, recorded as solution IV. 1 g of aldehyde sodium alginate prepared in step (1) is dissolved in 9 g distilled water to prepare a 10% aldehyde sodium alginate solution. Solution IV and 10% aldehyde sodium alginate solution are mixed in equal proportions and uniformly placed at 4°C to obtain a hydrogel.

[0083] The porous structure material has the advantages of high specific surface area, strong permeability and adsorption in daily life and production. In order to verify the internal structure of the hydrogel sample, SEM and optical microscope tests are carried out.

[0084] Test Example:

[0085] Hydrogel micro-morphology

[0086] As Figure 1As shown in, (a), (b), (c), and (d) correspond to the microscopic SEM images of the hydrogels in Example 1 and Comparative Examples 1-3, respectively. Figure 1 (a)-(d) show the scanning electron microscopy images of COT@BC@NaBH4 (Example 1), COT@NaBH4 (Comparative Example 2), COT@BC (Comparative Example 1), and COT (Comparative Example 3) hydrogel samples. COT is carboxymethyl chitosan / oxidized sodium alginate / tannic acid hydrogel, and BC is baicalin. All four hydrogels have a porous network structure. However, it can be observed that Figure 1 In (a) and (b), the number of pores in COT@BC@NaBH4 and COT@NaBH4 is significantly greater than that in COT@BC and COT, and they are denser and have smaller pore sizes. Therefore, it can be inferred that the treatment with NaBH4 will affect the porous structure of the hydrogel, making it have a denser pore structure. Figure 1 Comparison between (a) and (b), and (c) and (d) shows that the baicalin-loaded hydrogel exhibits a smaller internal pore structure, indicating that baicalin has been successfully dispersed in the hydrogel.

[0087] like Figure 2 As shown in Figure 1, (a), (b), (c), and (d) correspond to optical microscopic images of the hydrogels in Example 1 and Comparative Examples 1-3, respectively. Comparison of the gel images under an optical microscope reveals that sodium borohydride treatment creates numerous large pores within the gel, forming a multi-level porous structure based on the inherent porous structure of the hydrogel.

[0088] Performance test of hydrogel adsorbent:

[0089] (1) In vitro drug release performance test of hydrogel

[0090] The hydrogels prepared in Example 1 and Comparative Example 1 were placed in PBS solutions with pH values ​​of 3, 5, and 7, respectively. The PBS solutions were taken out at intervals to test their UV absorbance. A standard curve was established to calculate the drug release rate. The test results are shown in Table 1. Figure 3 .

[0091] Depend on Figure 3 It can be seen that the release rate and release ratio of the hydrogel prepared in Example 1 are higher than those of the hydrogel prepared in Comparative Example 1. Figure 3 It can be seen that the prepared hydrogel has pH-responsive properties, and the release rate under neutral conditions is higher than that under acidic conditions.

[0092] Figure 4 This is a comparison of the drug release rates of the hydrogels in Comparative Examples 2 and 3 in simulated body fluids with different pH values, indicating that the drug release rate of the hydrogel treated with sodium borohydride is significantly higher than that of the hydrogel not treated with sodium borohydride.

[0093] (2) Biotoxicity test of the hydrogel

[0094] The hydrogel extract was co-cultured with mouse fibroblasts (L929 cells) for 24 h, and the survival rate of the cells was tested by CKK-8 experiment. As shown in Table 2, the survival rate of the cells was the ratio of the experimental group to the blank sample: the compound traditional Chinese medicine drug-loaded hydrogel with multi-level pore structure described in the application had lower biotoxicity. The blank sample was a mouse fibroblast culture sample without the addition of the hydrogel extract. Figure 5

[0095] (3) Antimicrobial performance test of the hydrogel

[0096] LB liquid medium - 100 mL of distilled water was measured by a measuring cylinder and poured into a 250 mL reagent bottle, 2.5 g of LB broth medium was weighed by an analytical electronic balance and added for mixing, and then sterilized in a high-temperature and high-pressure steam sterilization pot at 121°C for 15 min, and then used after sterilization.

[0097] LB solid medium - 100 mL of distilled water was measured by a measuring cylinder and poured into a 250 mL reagent bottle, 2.5 g of LB broth medium and 1.5 g of agar powder were weighed by an analytical electronic balance and added for mixing, and then sterilized in a high-temperature and high-pressure steam sterilization pot at 121°C for 15 min. When the medium cooled to about 40-50°C, 15 mL of the medium was sucked into a disposable sterile flat dish by an electric pipettor.

[0098] Two 12 mL bacterial culture tubes were taken, 3 mL of LB liquid medium was added to each, a single colony was picked from the Staphylococcus aureus solid medium and added to the liquid medium, and the other was used as a blank control. Place in a constant temperature shaker (37°C, 200 rpm) and shake culture overnight (15 h).

[0099] The Staphylococcus aureus bacterial solution was diluted to 10 6 CFU / mL with sterile PBS solution, and then 100 μL of the solution was evenly coated on the LB solid medium. The sterilized samples (Example 1 and Comparative Example 1) were attached to the surface of the medium, and a sterile forceps was used to press gently to ensure that the sample was in full contact with the medium. The culture dish was placed in a constant temperature incubator at 37°C and cultured for 24 h. After the culture was completed, a normal camera was used to take a photo and measure the size of the inhibition zone, and the test results are shown in Table 2. Figure 6 , wherein (a), (b), (c), and (d) correspond to the inhibition ring photos of Example 1, Comparative Example 1, Comparative Example 2, and Comparative Example 3, respectively.

[0100] From the above test results, it can be seen that the compound traditional Chinese medicine drug-loaded hydrogel with multi-level pore structure described in the application has good antibacterial performance. Figure 6 ​It can be seen that the diameter of the bacteriostatic circle of Example 1 is obviously higher than that of Comparative Example 1, proving that it has a higher antibacterial effect.

Claims

1. A compound traditional Chinese medicine loaded hydrogel with a multi-level pore structure, characterized in that: The invention comprises the following components in parts by weight: 10-30 parts of carboxymethyl chitosan, 5-20 parts of oxidized sodium alginate, 3-5 parts of tannic acid, 0.5-1 part of baicalin, and 200-500 parts of water; The preparation method of the compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure comprises the following steps: (1) In solvent A, sodium alginate and an aqueous solution of sodium periodate are reacted to prepare oxidized sodium alginate; (2) dissolving carboxymethyl chitosan, tannic acid and baicalin in distilled water to obtain a mixed solution; dissolving the oxidized sodium alginate prepared in step (1) in distilled water to obtain an oxidized sodium alginate solution; mixing the mixed solution and the oxidized sodium alginate solution uniformly, and allowing to stand at 4°C to obtain a hydrogel; (3) The hydrogel is immersed in a sodium borohydride solution to react and obtain a compound Chinese medicine-loaded hydrogel with a multi-level pore structure.

2. The multi-level porous structured compound Chinese medicine-loaded hydrogel according to claim 1, characterized in that: The invention is composed of the following components in parts by weight: 20 parts of carboxymethyl chitosan, 10 parts of carboxymethyl chitosan oxidized sodium alginate, 4 parts of tannic acid, 1 part of baicalin, and 200 parts of water.

3. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 1, characterized in that: In step (1), the solvent A is methanol, ethanol or acetone.

4. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 3, characterized in that: In step (1), the solvent A is ethanol; the sodium alginate is added to the solvent A to obtain a sodium alginate suspension, and the mass ratio of the sodium alginate to the solvent A is 0.1-0.3 g / mL.

5. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 4, characterized in that: In step (1), the volume ratio of the mass of sodium periodate to distilled water in the sodium periodate aqueous solution is 0.1-0.3 g / mL.

6. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 5, characterized in that: In step (1), the volume ratio of the sodium alginate suspension to the sodium periodate aqueous solution is 1:(0.5-1).

7. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 1, characterized in that: In step (1), the reaction temperature is room temperature, the reaction condition is to avoid light; and the reaction time is 6-18 h.

8. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 7, characterized in that: The reaction time is 10-12 h.

9. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 1, characterized in that: In step (1), when the reaction is terminated, ethylene glycol in an amount equimolar to that of sodium periodate should be added.

10. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 1, characterized in that: In step (1), after the reaction is completed, the reaction mixture should be poured into vigorously stirred anhydrous ethanol, wherein the volume ratio of the reaction mixture to anhydrous ethanol is 1:(5-8) to obtain a precipitate.

11. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 10, characterized in that: In step (1), the obtained precipitate is filtered and vacuum-dried at 40° C., and the obtained solid powder is dialyzed with distilled water for 24 h.

12. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 1, characterized in that: In step (2), the mass ratio of carboxymethyl chitosan in the mixed solution to the volume ratio of distilled water is 0.1-0.3 g / mL; the mass ratio of tannic acid in the mixed solution to the volume ratio of distilled water is 0.04-0.08 g / mL; and the mass ratio of baicalin in the mixed solution to the volume ratio of distilled water is 0.001-0.02 g / mL.

13. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 1, characterized in that: In step (2), the mass fraction of the oxidized sodium alginate solution is 5-10%.

14. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 13, characterized in that: In step (2), the volume ratio of the mixed solution to the oxidized sodium alginate solution is 1:0.5-1.

5.

15. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 1, characterized in that: In step (3), the mass fraction of the sodium borohydride solution is 0.1-2%.

16. The compound traditional Chinese medicine-loaded hydrogel with a multi-level pore structure according to claim 1, characterized in that: In step (3), the reaction time is 2-5 min.

17. Use of the compound traditional Chinese medicine-loaded hydrogel with a multi-level porous structure according to any one of claims 1 to 16 in the preparation of antibacterial materials.

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