Janus hydrogel material as well as preparation method and application thereof

Abstract

The invention discloses a Janus hydrogel material as well as a preparation method and application thereof, and belongs to the technical field of material preparation. A building group substance, gallic acid and polyvinyl alcohol are mixed to prepare a PVA (at) DES coating with anti-blocking performance, then the PVA (at) DES coating is combined with a DM (at) DES coating which contains magnesium salt and zwitterionic group substances and has blocking performance, and the obtained Janus hydrogel material has asymmetric adhesion and degradability. The anti-inflammatory and anti-oxidation medicine can be degraded and released along with the increase of time, the anti-inflammatory effect is achieved, and anastomotic leakage can be effectively prevented and treated; on the premise of ensuring complete healing of the surgical site, secondary surgical taking-out is not needed, and surgical risks are reduced.

Description

Technical Field

[0001] The present invention belongs to the technical field of material preparation, and particularly relates to a Janus hydrogel material, a preparation method thereof and an application thereof. Background Art

[0002] Anastomotic leakage is one of the serious complications after colorectal cancer surgery, especially the incidence of rectal anastomotic leakage is higher. The incidence of rectal anastomotic leakage (AL) is 3%-30%, and the mortality rate is 2%-16.4%. It will not only prolong the hospital stay, increase the rate of reoperation, but also be related to tumor recurrence and reduced long-term survival rate, increase medical risks, and seriously affect the health status and quality of life of patients. Therefore, it is of great significance to study the prevention and treatment measures for rectal AL.

[0003] With the rise of biomaterials, many scholars have tried to improve the safety of anastomosis through new biomaterials, among which gel patches are the most widely used. However, the existing tissue adhesive hydrogels either only consider their strong adhesiveness or only focus on anti-adhesion, which will cause serious postoperative adhesions; some are non-degradable, and patients need to undergo another operation to remove them after wound healing, which severely limits the application of gel materials in internal tissue repair. Summary of the Invention

[0004] The technical problem to be solved by the present invention is to provide a Janus hydrogel material, a preparation method thereof and an application thereof, so as to solve the technical problems that the existing hydrogels have poor prevention and treatment effects on anastomotic leakage and are prone to postoperative adhesions.

[0005] To achieve the above object, the technical solution adopted by the present invention is: to provide a preparation method of a Janus hydrogel material, comprising the following steps:

[0006] S1. Dissolve a building block substance and gallic acid (GA) in a solvent at a mass ratio of 500-550:0.25-0.3, and stir at 55-65°C for 5-10 minutes to obtain a DES; the building block substance is hydroxypropyl-β-cyclodextrin (HP-β-CD), sodium sulfobutyl-β-cyclodextrin (SBE-β-CD), hydroxyethyl-β-cyclodextrin (HE-β-CD), glucosyl cyclodextrin, maltosyl cyclodextrin, diglucosyl cyclodextrin or dimaltosyl cyclodextrin;

[0007] The hydrogen bond between the -OH group of the building group substance and the -COOH group of gallic acid (GA) forms a supramolecular polymer through complexation. Moreover, the building group substance disrupts the molecular hydrogen bond, increases the solubility of cyclodextrin, and reduces nephrotoxicity. The modified cyclodextrin is actually a mixture of different-site substitutions. When complexing with guest molecules, it is not easy to form inclusion compound crystals and precipitate, increasing stability. It has a special three-dimensional ring structure with "hydrophobic inside and hydrophilic outside". The hydrophobic cavity can encapsulate drugs and can form inclusion compounds with a variety of compounds, thereby improving the water solubility and stability of guest molecules. After the building group substance is complexed with the drug, its physical and chemical properties can be changed, which has the effects of reducing toxic and side effects, increasing solubility and stability, and can also promote the healing of intestinal wounds.

[0008] S2. Dissolve the building group substance and gallic acid in a solvent, then add polyvinyl alcohol (PVA) and DES, stir at 90 - 100 °C until the solids are completely dissolved, and finally repeatedly freeze-thaw the mixed solution at -25 to -15 °C to obtain a PVA@DES coating; the ratio of the building group substance, gallic acid, polyvinyl alcohol, and DES is 500 - 550 mg : 0.25 - 0.3 mg : 1 - 1.5 g : 7 - 10 mL;

[0009] The PVA in the PVA@DES layer contains many active hydroxyl groups, which can react with the carboxyl groups on GA to form a cross-linked structure with strong chemical bonds. And the abundant hydrogen bonds of PVA have high crystallinity, making the coating have unique ultra-low expansion performance. This structure helps to enhance the strength and durability of the material, thereby preventing adhesion to a certain extent.

[0010] S3. Co-dissolve the building group substance, gallic acid, and magnesium salt in a solvent at a mass ratio of 500 - 550 : 0.25 - 0.3 : 0.3 - 0.4, stir at 55 - 65 °C for 5 - 10 min to obtain an intermediate solution. Mix the intermediate solution, the substance containing zwitterionic groups, and a photoinitiator at 0.5 - 1.5 : 0.5 - 1.5 : 0.001 - 0.003, then cover it on the surface of the PVA@DES coating, and finally irradiate it with ultraviolet light for 5 - 15 min to obtain a Janus hydrogel material (PVA-DM@DES).

[0011] Mg 2+ It is used to enhance the complexation of the material to improve the mechanical properties, and also has antibacterial activity and the effect of promoting wound healing. Since the zwitterionic group can form ion-dipole or dipole-dipole interactions with other charged groups or polar groups, the prepared hydrogel layer can adhere to the surfaces of various materials, further enhancing the adhesion performance, and finally obtaining a Janus hydrogel material with obvious adhesion and non-adhesion properties on both sides.

[0012] On the basis of the above technical solutions, the present invention can also be improved as follows:

[0013] Further, the ratio of the building group substance, gallic acid, polyvinyl alcohol and DES in S2 is 539.5 mg: 0.282 mg: 1.2 g: 8.8 mL.

[0014] Further, the magnesium salt is magnesium sulfate.

[0015] Further, the zwitterionic group-containing substance is 2-[(methacryloyloxy)ethyl]dimethyl-(3-sulfopropyl) ammonium hydroxide (DMAPS).

[0016] Further, the photoinitiator is I2959. I2959 is a compound that can absorb energy of a certain wavelength in the ultraviolet region (250 - 420 nm) or the visible light region (400 - 800 nm) to generate free radicals or cations, and is used to initiate the UV polymerization reaction of unsaturated prepolymer systems.

[0017] The present invention also discloses a Janus hydrogel material prepared by the above preparation method.

[0018] The present invention also discloses the application of the Janus hydrogel material in the preparation of anastomotic leakage patches.

[0019] The beneficial effects of the present invention are as follows:

[0020] 1. The present invention uses the green and non-toxic deep eutectic solvent DES to replace the ionic liquid as the solvent of the gel, which has low cost and is easy to prepare.

[0021] 2. The Janus hydrogel material has asymmetric adhesiveness. One side can firmly adhere to the surface of wet tissues to prevent liquid leakage, and the other side has anti-adhesion property to prevent postoperative tissue adhesion, thereby reducing the incidence of intestinal obstruction.

[0022] 3. The Janus hydrogel material has degradability and can degrade over time and release anti-inflammatory and antioxidant drugs to exert an anti-inflammatory effect, which can effectively prevent and treat the occurrence of anastomotic leakage; without the need for a second operation to remove it under the premise of ensuring complete healing of the surgical site, reducing the surgical risk. Description of the Drawings

[0023] Figure 1 It is the electron microscope scanning image of the PVA-DM@DES hydrogel; among them, Figure (a) is the electron microscope scanning image of the PVA@DES coating with anti-adhesion property on the upper surface of the PVA-DM@DES hydrogel, Figure (b) is the electron microscope scanning image of the DM@DES coating with adhesiveness on the lower surface of the PVA-DM@DES hydrogel, and Figure (c) is the electron microscope scanning image of the cross-section of the PVA-DM@DES hydrogel;

[0024] Figure 2 Water contact angle analysis results for PVA@DES and DM@DES;

[0025] Figure 3 Comparison of the prevention and treatment effects of anastomotic leakage under different treatments;

[0026] Figure 4 Comparison results of the concentration of inflammatory factor TNF-α;

[0027] Figure 5 Comparison results of the concentration of anti-inflammatory factor IL-10. Specific embodiments

[0028] The following describes the specific embodiments of the present invention to facilitate those skilled in the art of the present technology to understand the present invention. For those not specified in the embodiments, they are carried out according to conventional conditions or the conditions recommended by the manufacturer. For reagents or instruments not specified by the manufacturer, they are all conventional products that can be obtained through commercial purchase. However, it should be clear that the present invention is not limited to the scope of the specific embodiments. For those of ordinary skill in the art of the present technology, as long as various changes are within the spirit and scope of the present invention defined and determined by the appended claims, these changes are obvious, and all inventions and creations using the concept of the present invention are within the scope of protection.

[0029] Example 1

[0030] A preparation method of a Janus hydrogel material, comprising the following steps:

[0031] S1. Dissolve 500 mg of sodium sulfobutyl-β-cyclodextrin and 0.3 mg of gallic acid in 10 mL of distilled water, and stir at 55 °C for 10 min to obtain DES;

[0032] S2. Dissolve 550 mg of sodium sulfobutyl-β-cyclodextrin and 0.25 mg of gallic acid in 10 mL of distilled water, then add 1 g of polyvinyl alcohol and 10 mL of DES, and stir at 90 °C until the solid is completely dissolved to obtain an anti-adhesion precursor solution with a volume fraction of 13 wt%. Finally, the anti-adhesion precursor solution is repeatedly frozen and thawed at -25 °C to obtain a PVA@DES coating;

[0033] S3. Dissolve 0.4 mg of magnesium sulfate in 10 mL of distilled water, then add 500 mg of sulfobutyl-β-cyclodextrin sodium and 0.3 mg of gallic acid, and stir at 55 °C for 10 min to obtain an intermediate solution; mix the intermediate solution and 2-[(methacryloyloxy)ethyl]dimethyl-(3-sulfopropyl)ammonium hydroxide (DMAPS) in a mass ratio of 1:1, then add 0.1 wt% of I2959 and dissolve it by ultrasonic treatment. Then cover the mixed solution on the surface of the PVA@DES coating and irradiate it under an ultraviolet lamp with a wavelength of 365 nm for 5 min to obtain the Janus hydrogel material (PVA-DM@DES).

[0034] Example 2

[0035] A preparation method of a Janus hydrogel material, comprising the following steps:

[0036] S1. Dissolve 550 mg of glucosyl cyclodextrin and 0.25 mg of gallic acid in 10 mL of distilled water, and stir at 65 °C for 5 min to obtain DES;

[0037] S2. Dissolve 500 mg of glucosyl cyclodextrin and 0.3 mg of gallic acid in 10 mL of distilled water, then add 1.5 g of polyvinyl alcohol and 7 mL of DES, and stir at 100 °C until the solids are completely dissolved to obtain an anti-adhesion precursor solution with a volume fraction of 11 wt%; finally, repeatedly freeze-thaw the anti-adhesion precursor solution at -15 °C to obtain the PVA@DES coating;

[0038] S3. Dissolve 0.3 mg of magnesium sulfate in 10 mL of distilled water, then add 550 mg of glucosyl cyclodextrin and 0.25 mg of gallic acid, and stir at 65 °C for 5 min to obtain an intermediate solution; mix the intermediate solution and 2-[(methacryloyloxy)ethyl]dimethyl-(3-sulfopropyl)ammonium hydroxide (DMAPS) in a mass ratio of 1:1, then add 0.1 wt% of I2959 and dissolve it by ultrasonic treatment. Then cover the mixed solution on the surface of the PVA@DES coating and irradiate it under an ultraviolet lamp with a wavelength of 365 nm for 15 min to obtain the Janus hydrogel material (PVA-DM@DES).

[0039] Example 3

[0040] A preparation method of a Janus hydrogel material, comprising the following steps:

[0041] S1. Dissolve 539.5 mg of hydroxypropyl-β-cyclodextrin and 0.282 mg of gallic acid in 10 mL of distilled water, and stir at 60 °C for 8 min to obtain DES;

[0042] S2. Dissolve 539.5 mg of hydroxypropyl-β-cyclodextrin and 0.282 mg of gallic acid in 10 mL of distilled water. Subsequently, add 1.2 g of polyvinyl alcohol and 8.8 mL of DES, and stir at 95 °C until the solids are completely dissolved to obtain an anti-adhesion precursor solution with a volume fraction of 12 wt%. Finally, subject the anti-adhesion precursor solution to repeated freezing and thawing at -20 °C to obtain a PVA@DES coating;

[0043] S3. Dissolve 0.36 mg of magnesium sulfate in 10 mL of distilled water. Subsequently, add 539.5 mg of hydroxypropyl-β-cyclodextrin and 0.282 mg of gallic acid, and stir at 60 °C for 8 min to obtain an intermediate solution. Mix the intermediate solution and 2-[(methacryloyloxy)ethyl]dimethyl-(3-sulfopropyl)ammonium hydroxide (DMAPS) in a mass ratio of 1:1, then add 0.1 wt% of I2959 and dissolve it by ultrasonic treatment. Then cover the mixed solution on the surface of the PVA@DES coating and irradiate it under ultraviolet light with a wavelength of 365 nm for 10 min to obtain a Janus hydrogel material (PVA-DM@DES).

[0044] Comparative Example 1

[0045] The difference between this comparative example and Example 3 is that: all steps omit the use of gallic acid and magnesium sulfate, and the remaining implementation conditions are the same as those in Example 3, to prepare a hydrogel material (PVA-DH@DES).

[0046] Comparative Example 2

[0047] The difference between this comparative example and Example 3 is that: magnesium sulfate is omitted, and the remaining implementation conditions are the same as those in Example 3, to prepare a hydrogel material (PVA-D@DES).

[0048] Experimental Example 1 Gelation Performance and Adhesion Performance

[0049] Take the hydrogel prepared in Example 3 as an example for microscopic characterization. Figure 1 It is the electron microscope scanning image of the PVA-DM@DES hydrogel. Among them, Figure (a) is the electron microscope scanning image of the PVA@DES coating with anti-adhesion property on the upper surface of the PVA-DM@DES hydrogel, Figure (b) is the electron microscope scanning image of the DM@DES coating with adhesion property on the lower surface of the PVA-DM@DES hydrogel, and Figure (c) is the electron microscope scanning image of the cross-section of the PVA-DM@DES hydrogel. The gradient structure of the PVA-DM@DES hydrogel can be intuitively reflected by the pore size change from the upper surface to the lower surface. It can be clearly seen that the pore size on the cross-section gradually increases from the upper surface to the lower surface. There are no pores formed on the upper surface of the PVA-DM@DES hydrogel, and a pore network structure exists on the lower surface. The upper surface shows a denser structure than the lower surface, which fully demonstrates that the PVA-DM@DES hydrogel has an asymmetric microscopic structure.

[0050] Example 2: Hydrophobicity and hydrophilicity

[0051] The mixed solution prepared in step S3 of Example 3 was placed under an ultraviolet lamp with a wavelength of 365 nm for 10 min to obtain the DM@DES sample. The PVA@DES coating prepared in step S2 and the DM@DES sample were placed on a horizontal platform, water droplets were added, and the images of the water droplets were captured by a camera, and then the water contact angle system was used to evaluate their hydrophobicity and hydrophilicity.

[0052] Figure 2 Figure shows the water contact angle analysis results of PVA@DES and DM@DES. The water contact angle (WCA, 92.2°) of the PVA@DES layer and that of the DM@DES layer (WCA, 30.3°) indicate the presence of hydrophobicity and hydrophilicity in the PVA@DES layer and the DM@DES layer, respectively. In the PVA@DES layer, chemical bonds are formed between the PVA molecular chains and GA to form a three-dimensional network structure, which may lead to its hydrophobic properties; secondly, heat treatment of PVA during the preparation process can make its molecular chains more closely arranged, reducing free volume and surface defects, thereby improving its hydrophobic properties. In the DM@DES layer, zwitterionic polymers produce a strong hydration effect through eutectic solventization to form a stable hydration layer, which not only enhances the hydrophilicity of DM@DES, but also promotes the adhesion performance to a certain extent. And chain-interchain dipole-dipole associations can be formed between zwitterionic polymers, and this physical cross-linking interaction helps to improve the adhesion strength and stability of DM@DES.

[0053] Example 3: In vivo effect detection

[0054] The materials (PVA-DH@DES, PVA-D@DES, and PVA-DM@DES) prepared in Example 3, Comparative Example 1, and Comparative Example 2 were used as samples for experiments. Male SD rats aged 6 - 8 weeks (purchased from the Experimental Animal Center of Daping Hospital, Chongqing Army Medical Center) were divided into 4 groups, with 8 rats in each group. First, the SD rats were given an adaptation period of one week. The rats were anesthetized with isoflurane using an anesthetic machine air pump, the abdomen was shaved, the shaved area was wiped and disinfected with iodophor, the skin layer and muscle layer were cut open with sterilized scissors, after opening the abdominal cavity, a sterile gauze was laid and an expander was placed into the abdominal cavity, the cecum and other organs were dragged to one side, the rectal position was found, a 2 / 3 incision was made in the rectum with sterilized scissors, and the wound was sparsely and appropriately sutured to establish a rectal anastomotic leakage rat model. Then different material samples (PVA-DH@DES, PVA-D@DES, and PVA-DM@DES) were applied to the wound. At the same time, a control group (without any patch material) was set up. The expander was removed and the organs were put back into the abdominal cavity, and the abdominal cavity was closed and sutured. Samples were collected and observed on the 3rd day and the 7th day respectively, and the results are as Figure 3as shown

[0055] From Figure 3 It can be seen that compared with the other 3 groups, the wounds of the rats in the PVA-DM@DES group recovered well, and the prevention and treatment effect of anastomotic leakage was the most obvious, without obvious abscesses and tissue adhesions. In contrast, there were abscesses and dense adhesions to varying degrees in the other 3 groups, indicating that the Janus hydrogel material prepared by the present invention has an obvious effect on preventing and treating rectal anastomotic leakage.

[0056] Example 4 Anti-inflammatory Effect Evaluation

[0057] PVA-DH@DES, PVA-D@DES, and PVA-DM@DES were placed under ultraviolet light overnight for material disinfection and extracted with high-glucose medium without serum to obtain an extract of 0.1 g / mL for subsequent experiments. Five experimental groups were set up, and macrophages were seeded in 24-well plates at an inoculation density of 1×10 5 and placed in a 5% CO 2 and 37 °C cell culture incubator for 4 h. Then, LPS (lipopolysaccharide) solution with a concentration of 10 μg / mL was added to 4 of the experimental groups to induce macrophage polarization for 12 h. After successful polarization induction, the extracts of PVA-DH@DES, PVA-D@DES, and PVA-DM@DES were added to 3 of the experimental groups respectively, and then the plates were placed in a 5% CO 2 and 37 °C cell culture incubator for 3 d. The culture medium was collected and centrifuged at 5000 rpm for 10 min to remove cell debris. The supernatant of the collected culture medium was used to detect the concentrations of TNF-α and IL-10 in the culture medium using the corresponding ELISA kits. The specific steps refer to the instructions of the ELISA kits (Abclonal Technology).

[0058] Figure 4 and Figure 5 For the in vitro anti-inflammatory effect evaluation of PVA-DM@DES, it can be seen from the figure that the PVA-DM@DES patch can effectively reduce the secretion of inflammatory factor (TNF-α) and increase the secretion of anti-inflammatory factor (IL-10). After co-culture, the concentration of TNF-α in the PVA-DM@DES group was only 61.32% of that in the control group (Control group), indicating that the PVA-DM@DES patch prepared by the present invention can significantly reduce the concentration of inflammatory factors and increase the concentration of anti-inflammatory factors, and has anti-pressure and antioxidant effects.

Claims

1. A method for preparing a Janus hydrogel material, characterized in that: The following steps are involved: S1. Dissolve the building group substance and gallic acid in a solvent at a mass ratio of 500-550:0.25-0.3, and stir at 55-65°C for 5-10 minutes to obtain DES; the building group substance is hydroxypropyl-β-cyclodextrin, sulfobutyl-β-cyclodextrin sodium, hydroxyethyl-β-cyclodextrin, glucosyl cyclodextrin, maltosyl cyclodextrin, diglucosyl cyclodextrin or dimaltosyl cyclodextrin; S2, dissolving the building group substance and gallic acid in a solvent, then adding polyvinyl alcohol and DES, stirring at 90-100°C until the solid is completely dissolved, and finally freezing and thawing the mixed solution repeatedly at -25 to -15°C to obtain a PVA@DES coating; The ratio of the building group substance, gallic acid, polyvinyl alcohol and DES is 500-550 mg: 0.25-0.3 mg: 1-1.5 g: 7-10 mL; S3. Dissolve the building group substance, gallic acid and magnesium salt in a solvent at a mass ratio of 500-550:0.25-0.3:0.3-0.4, stir at 55-65°C for 5-10 minutes to obtain an intermediate solution, mix the intermediate solution, the substance containing zwitterionic groups and the photoinitiator at a ratio of 0.5-1.5:0.5-1.5:0.001-0.003, and then cover the surface of the PVA@DES coating, and finally irradiate under ultraviolet light for 5-15 minutes to obtain a Janus hydrogel material.

2. The method for preparing the Janus hydrogel material according to claim 1, characterized in that: The ratio of the building block substance, gallic acid, polyvinyl alcohol and DES in the S2 is 539.5 mg: 0.282 mg: 1.2 g: 8.8 mL.

3. The method for preparing the Janus hydrogel material according to claim 1, characterized in that: The magnesium salt is magnesium sulfate.

4. The method for preparing the Janus hydrogel material according to claim 1, characterized in that: The substance containing zwitterionic groups is 2-(methacryloyloxy)ethyl]dimethyl-(3-sulfonic acid propyl)ammonium hydroxide.

5. The method for preparing the Janus hydrogel material according to claim 1, characterized in that: The photoinitiator is I2959.

6. A Janus hydrogel material, characterized in that: The method is prepared by the preparation method according to any one of claims 1 to 5.

7. Use of the Janus hydrogel material according to claim 6 in preparing anastomotic leakage patches.

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