A Janus zwitterionic hydrogel anti-adhesion film and its preparation method and application

Abstract

The present invention discloses a Janus zwitterionic hydrogel anti-adhesion film and its preparation method and application, which is characterized by consisting of an adhesion layer and an anti-adhesion layer. The adhesion layer is a copolymer of a zwitterionic monomer, N-acryloylglycine amide, 3-methacryloyldopamine and a chemical cross-linking agent, and the anti-adhesion layer is a copolymer of a zwitterionic monomer, N-acryloylglycine amide and a chemical cross-linking agent. The adhesion layer of the Janus zwitterionic hydrogel anti-adhesion film has strong adhesion to wet tissues, can quickly seal the wounds of different organs and quickly stop bleeding; its anti-adhesion layer has good anti-bioadhesion performance and can prevent adhesion of viscera and tissues. The Janus zwitterionic hydrogel anti-adhesion film can be used for anti-adhesion in abdominal wall hernia, postoperative abdominal cavity, thoracic cavity, etc.

Description

Technical Field

[0001] The present invention relates to a Janus zwitterionic hydrogel anti-adhesion film, its preparation method and application. It is a double-layer film composed of an adhesion layer and an anti-adhesion layer formed by different zwitterionic copolymers, and is suitable for the development of anti-adhesion patches. Background Art

[0002] Surgical operation is an important method for treating various diseases. During the operation, it is usually inevitable to cause damage to tissues and organs, leaving wounds, which need to be healed by relying on the self-healing function of tissues after the operation. However, during the normal healing process, the internal tissues and organs are likely to come into contact with the surgical site wounds, which will lead to postoperative tissue adhesion. Severe postoperative adhesions can cause chronic pain and various complications, such as adjacent organ dysfunction, intestinal obstruction, female infertility, etc., seriously affecting the quality of life of patients, bringing a heavy burden to patients, and even endangering the lives of patients. Therefore, people have been committed to developing various anti-adhesion strategies to isolate the wounds caused during the operation from tissues and organs or weaken the inflammatory response of tissues and organs, in order to achieve the purpose of preventing postoperative adhesions.

[0003] The anti-adhesion film is a commonly used and mature method for clinically preventing postoperative adhesions. It can directly physically isolate the injured part from adjacent tissues and organs, thus playing an anti-adhesion role. An ideal anti-adhesion film should have good biodegradability, biocompatibility and appropriate tissue adhesion (without suture fixation), and can maintain a relatively complete physical form during the critical period of preventing adhesion formation, while not affecting the wound healing, or being able to promote wound healing. In recent years, the application of anti-adhesion films in surgical operations has become more and more extensive, and it can effectively prevent the occurrence of postoperative tissue adhesions. Currently, the commercially available anti-adhesion film materials are usually polylactic acid (PLA), polyethylene glycol (PEG), carboxymethyl cellulose, hyaluronic acid, etc. Their adhesion performance to tissues is poor, they need surgical sutures, causing great secondary damage to tissues, and generally the anti-adhesion film cannot avoid the adhesion and proliferation of cell tissues on its surface, thus affecting the anti-adhesion effect.

[0004] Therefore, enhancing the adhesion performance to the tissue to be healed while also making it have an anti-bioadhesion effect with other tissues is an important direction for optimizing the performance of the anti-adhesion film. Summary of the Invention

[0005] The purpose of the present invention is to design and prepare a Janus zwitterionic hydrogel anti-adhesion film, which is characterized by a double-layer film composed of an adhesion layer with wet tissue adhesion and an anti-adhesion layer with anti-bioadhesion, which can quickly seal the wounds of different organs and has a hemostatic function; the anti-adhesion layer has good anti-bioadhesion performance and is used for anti-adhesion treatment after abdominal wall hernia, abdominal cavity, thoracic cavity, etc.

[0006] To achieve the above object, the present invention adopts the following technical solutions:

[0007] A Janus zwitterionic hydrogel anti-adhesion film, the anti-adhesion film is composed of an adhesion layer and an anti-adhesion layer;

[0008] The adhesion layer is a copolymer of a zwitterionic monomer, N-acryloylglycine amide (NAGA), 3-methacryloyldopamine (DMA), and a chemical crosslinking agent; among them, N-acryloylglycine amide can improve the gel strength of the adhesion layer; the catechol group in 3-methacryloyldopamine can bind to various nucleophilic groups (such as amino, mercapto, and imidazole groups) in polypeptides and proteins on the tissue surface, thereby adhering to the tissue surface; the zwitterionic hydrogel has excellent hydrophilicity and can quickly absorb water molecules on the tissue surface, thereby enhancing the adhesion force between 3-methacryloyldopamine and the tissue surface; the chemical crosslinking agent can prevent the adhesion layer from swelling.

[0009] The anti-adhesion layer is a copolymer of a zwitterionic monomer, N-acryloylglycine amide, and a chemical crosslinking agent. Among them, the zwitterionic hydrogel formed by the polymerization of the zwitterionic monomer has good hydrophilicity, and a stable hydration layer can be formed on the surface, thereby effectively preventing the occurrence of postoperative tissue adhesion; N-acryloylglycine amide can improve the strength of the hydrogel; the chemical crosslinking agent can prevent the anti-adhesion layer from swelling.

[0010] The adhesion layer of the Janus zwitterionic hydrogel anti-adhesion film of the present invention has strong adhesion to wet tissues, can quickly seal the wounds of different organs and quickly stop bleeding; the anti-adhesion layer has good anti-bioadhesion performance, can inhibit the surface adhesion of biological components such as proteins and cells, and can prevent the adhesion of organs and tissues.

[0011] Preferably, the adhesion layer is a copolymer formed by a zwitterionic monomer, N-acryloylglycine amide, 3-methacryloyldopamine, and a chemical crosslinking agent under the initiation of an initiator; the anti-adhesion layer is a copolymer formed by a zwitterionic monomer, N-acryloylglycine amide, and a chemical crosslinking agent under the initiation of an initiator.

[0012] Preferably, in the copolymer forming the adhesion layer, the ratio of the total mass of N-acryloylglycine amide monomers to the total mass of zwitterionic monomers is 0.125 to 2, the ratio of the total mass of 3-methacryloyldopamine monomers to the total mass of zwitterionic monomers is 0.125 to 2, and the percentage of the chemical crosslinking agent in the total mass of zwitterionic monomers is 0.5 w% to 15 w%;

[0013] The ratio of the total mass of N - acryloylglycine amide monomers to the total mass of zwitterionic monomers in the copolymer forming the anti - adhesion layer is 0.125 - 0.75, and the mass percentage of the chemical cross - linker in the zwitterionic monomers is 0.5w% - 15w%.

[0014] Preferably, the zwitterionic monomer is selected from any one of methacryloylethyl sulfobetaine (SBMA), 2 - methacryloyloxyethyl phosphorylcholine (MPC), and carboxybetaine methacrylate (CBMA).

[0015] Preferably, the chemical cross - linker is selected from one or more of N,N - methylene bisacrylamide (MBA), N,N - bis(acryloyl)cystamine (MSBA), ethylene glycol dimethacrylate (EBA), bis - methacryloylethyl carboxybetaine (CBBA), and poly(ethylene glycol) diacrylate (PEGDA).

[0016] Preferably, the initiator is selected from photo - initiator 2959.

[0017] The present invention also discloses a preparation method of the above - mentioned Janus zwitterionic hydrogel anti - adhesion film, which includes the following steps:

[0018] (1) Prepare an adhesion layer precursor solution, which is an aqueous mixed solution of a zwitterionic monomer, N - acryloylglycine amide, 3 - methacryloyldopamine, an initiator, and a chemical cross - linker.

[0019] (2) Prepare an anti - adhesion layer precursor solution, which is a mixed solution of a zwitterionic monomer, N - acryloylglycine amide, an initiator, and a cross - linker.

[0020] (3) Inject the adhesion layer precursor solution into a mold, polymerize it with an ultraviolet lamp at 10 - 60 °C. After molding, then inject the anti - adhesion layer precursor solution into the mold and polymerize it with an ultraviolet lamp at 10 - 60 °C to obtain a Janus zwitterionic hydrogel anti - adhesion film.

[0021] Preferably, in the adhesion layer precursor solution, the concentration of the zwitterionic monomer is 10w% - 40w%, the concentration of the N - acryloylglycine amide monomer is 5w% - 20w%, the concentration of the 3 - methacryloyldopamine monomer is 5w% - 20w%, the mass percentage of the chemical cross - linker in the zwitterionic monomer is 0.5w% - 15w%, and the mass percentage of the initiator in the zwitterionic monomer is 0.5% - 20%;

[0022] The concentration of zwitterionic monomer in the anti-adhesion layer precursor solution is 10w%-40w%, the concentration of N-acryloylglycine amide monomer is 5w%-13w%, the dosage of crosslinking agent accounts for 0.5w%-15w% of the mass of zwitterionic, and the dosage of initiator accounts for 0.5w%-20w% of the mass of zwitterionic.

[0023] Preferably, ultraviolet polymerization is carried out under a 365nm ultraviolet lamp for 30-60min.

[0024] The Janus zwitterionic hydrogel anti-adhesion film provided by the present invention can be used alone as a medical anti-adhesion patch, or can be formulated with bioactive substances such as drugs, active factors, polypeptides, and electrodes to form anti-adhesion products for the anti-adhesion treatment of diseases such as abdominal wall hernia, postoperative peritoneal adhesion, and thoracic adhesion.

[0025] The beneficial effects of the present invention are as follows:

[0026] The preparation technology of the Janus zwitterionic hydrogel anti-adhesion film of the present invention is simple, the preparation conditions are mild, the raw materials used are not only convenient to prepare but also have low cost, have good biocompatibility, and have both long-lasting wet tissue adhesiveness and anti-adhesion effects. Its adhesion layer solves the problems of the need for suturing or stapling fixation of existing anti-adhesion films or hernia repair patches and the resulting traumatic injuries, and the anti-adhesion layer solves the problems of poor biocompatibility and postoperative organ adhesion of existing anti-adhesion films or hernia repair patches. As a hernia repair patch and postoperative anti-adhesion film, it has application prospects in hernia, abdominal cavity and other surgeries. Description of the Drawings

[0027] Figure 1 : Cross-section and surface SEM images of the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1.

[0028] Figure 2 : 180° peel strength between each interface of the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1.

[0029] Figure 3 : Water contact angle of each layer of the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1.

[0030] Figure 4 : Interface toughness and shear strength between the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1 and the surface of wet porcine skin.

[0031] Figure 5 : Adhesion images and shear strength between the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1 and different wet tissues.

[0032] Figure 6: Anti-burst pressure test diagram of the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1 on different tissue surfaces.

[0033] Figure 7 : Commercial anti-adhesion film (Hongjian polylactic acid anti-adhesion film), commercial PP patch (Fu Lite polypropylene flat sheet and pre-cut flat sheet, Beijing Tianzhu Changyun Medical Technology Co., Ltd.) and the front and back anti-protein, anti-platelet, anti-bacterial and anti-cell adhesion performance diagrams of the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1.

[0034] Figure 8 : Schematic diagram of in vitro wound sealing of different organs by the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1.

[0035] Figure 9 : Rabbit carotid artery hemostasis performance diagram of the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1. (a) Physical diagram of the Janus zwitterionic hydrogel anti-adhesion film for rabbit carotid artery hemostasis, (b) Masson staining photos at different time points after rabbit carotid artery hemostasis, (c) Carotid artery wound width at different time points after rabbit carotid artery hemostasis.

[0036] Figure 10 : Cytotoxicity test results of the Janus zwitterionic hydrogel anti-adhesion film of Example 1.

[0037] Figure 11 : Commercial anti-adhesion film and the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1 were used in the anti-adhesion model of SD rat cecum-abdominal wall injury. (a) Physical diagrams of the commercial anti-adhesion film and the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1 at different time points after SD rat cecum-abdominal wall injury, (b) HE and Masson staining photos of the commercial anti-adhesion film and the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1 after SD rat cecum-abdominal wall injury.

[0038] Figure 12 : Commercial PP patch and the Janus zwitterionic hydrogel anti-adhesion film of Example 1 were used to repair the rabbit abdominal wall defect model. (a) Physical diagrams of the commercial PP patch and the Janus zwitterionic hydrogel anti-adhesion film of Example 1 at different time points after repairing the rabbit abdominal wall defect, (b) HE and Masson staining photos of the commercial PP patch and the Janus zwitterionic hydrogel anti-adhesion film of Example 1 after repairing the rabbit abdominal wall defect.

[0039] Figure 1-7 , In 10, the front is the adhesion layer; the back is the anti-adhesion layer. Detailed implementation mode

[0040] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the protection scope of the present invention.

[0041] Example 1

[0042] (1) Preparation of the adhesion layer hydrogel precursor solution: Take 2 mL of distilled water, and add 0.4 g of SBMA (concentration 14 w%), 0.2 g of NAGA (concentration 7 w%), 0.2 g of DMA (concentration 7 w%), 0.02 g of MBA (5 w% of SBMA), and 0.008 g of photoinitiator I-2959 (2 w% of SBMA).

[0043] (2) Preparation of the anti-adhesion layer hydrogel precursor solution: Take 2 mL of distilled water, and add 0.4 g of SBMA (concentration 15.2 w%), 0.2 g of NAGA (concentration 7.6 w%), 0.02 g of MBA (5 w% of SBMA), and 0.008 g of photoinitiator I-2959 (2 w% of SBMA).

[0044] (3) Preparation of the Janus zwitterionic hydrogel anti-adhesion film: Inject the adhesion layer precursor solution into the groove of a 2 cm × 2 cm × 2 mm silicone mold, and polymerize it under a 365 nm ultraviolet lamp at 25 °C for 30 min. After molding, inject the anti-adhesion layer precursor solution into the mold, and polymerize it under a 365 nm ultraviolet lamp at 25 °C for 30 min to obtain the Janus zwitterionic hydrogel anti-adhesion film.

[0045] The structural properties of the anti-adhesion film prepared in Example 1 are as Figure 1-12 and Table 2.

[0046] Figure 1 are the cross-section and surface SEM images of the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1. It can be seen from Figure 1 that the Janus zwitterionic hydrogel anti-adhesion film has a bilayer structure, and the two layers are firmly combined. It can be seen from Figure 2 that there is a strong binding force between the two layers of the Janus zwitterionic hydrogel anti-adhesion film. It can be seen from Figure 3 the water contact angle of each layer of the anti-adhesion film that there is a significant difference in hydrophilicity between the two layers of the Janus zwitterionic hydrogel anti-adhesion film, and the anti-adhesion layer on the reverse side has super hydrophilicity. Figure 4 It can be seen that the adhesion layer on the front side of the Janus zwitterionic hydrogel anti-adhesion film has strong adhesion on the surface of pigskin. Figure 5It is demonstrated that the Janus zwitterionic hydrogel anti-adhesion film has good wet tissue adhesion performance on different tissue surfaces. Figure 6 The data show that the wet tissue sealed by the Janus zwitterionic hydrogel anti-adhesion film has strong anti-burst pressure ability. Figure 7 It is shown that compared with the commercial anti-adhesion film (Hongjian polylactic acid anti-adhesion film) and the commercial PP patch ( Lightweight polypropylene flat sheet and pre-cut flat sheet, Beijing Tianzhuchangyun Medical Technology Co., Ltd.), the anti-adhesion layer of the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1 has better anti-bioadhesion performance than the above products.

[0047] When the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1 is used in vitro for sealing wounds of different organs, such as Figure 8 shown, the Janus zwitterionic hydrogel anti-adhesion film can quickly seal wounds of different organs, and the sealed organ wounds have good sealing performance.

[0048] Such as Figure 9 , the rabbit carotid artery hemostasis performance test of the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1 shows that: the Janus zwitterionic hydrogel anti-adhesion film can quickly seal the rabbit carotid artery and has a rapid hemostasis function. And Figure 10 the results of the cytotoxicity test prove that: the Janus zwitterionic hydrogel anti-adhesion film has good cytocompatibility.

[0049] Such as Figure 11 , taking the commercial anti-adhesion film as a control, the application performance of the Janus zwitterionic hydrogel anti-adhesion film prepared in Example 1 was evaluated on the SD rat cecum-abdominal wall injury adhesion model. From Figure 11 it can be seen that: no visceral adhesion occurred in the animals using the Janus zwitterionic hydrogel anti-adhesion film. And for the test results of repairing the abdominal wall defect of rabbits, such as Figure 12 , it shows that: the hernia of the animals using the Janus zwitterionic hydrogel anti-adhesion film was basically repaired after 28 days, and no organ adhesion occurred.

[0050] Examples 2 - 5

[0051] According to the method of Example 1, the operation is the same as that of Example 1, the reaction temperature and reaction time are the same as those of Example 1, and according to the composition in Table 1, the amounts of each reactant are adjusted to prepare the Janus zwitterionic hydrogel anti-adhesion film.

[0052] Examples 6 - 8

[0053] According to the method of Example 1, the operation is the same as that of Example 1, and according to the composition in Table 1, different zwitterionic monomers and different chemical cross-linking agents are used, and the amounts of each reactant are adjusted to prepare the precursor solutions of the adhesion layer and the anti-adhesion layer respectively.

[0054] Preparation of Janus zwitterionic hydrogel anti-adhesion film: Inject the precursor solution of the adhesion layer into the groove of a 2 cm × 2 cm × 2 mm silicone mold, and polymerize it under a 365 nm ultraviolet lamp at 40 °C for 40 min. After molding, inject the precursor solution of the anti-adhesion layer into the mold, and polymerize it under a 365 nm ultraviolet lamp at 40 °C for 40 min to obtain the Janus zwitterionic hydrogel anti-adhesion film.

[0055] Examples 9 - 13

[0056] According to the method of Example 1, the operation is the same as that of Example 1. According to the composition in Table 1, different zwitterionic monomers and different cross-linking agents are used, and the amounts of each reactant are adjusted to prepare the precursor solutions of the adhesion layer and the anti-adhesion layer respectively.

[0057] Preparation of Janus zwitterionic hydrogel anti-adhesion film: Inject the precursor solution of the adhesion layer into the groove of a 2 cm × 2 cm × 2 mm silicone mold, and polymerize it under a 365 nm ultraviolet lamp at 30 °C for 50 min. After molding, inject the precursor solution of the anti-adhesion layer into the mold, and polymerize it under a 365 nm ultraviolet lamp at 30 °C for 50 min to obtain the Janus zwitterionic hydrogel anti-adhesion film.

[0058] Example 14

[0059] According to the method of Example 1, the operation is the same as that of Example 1. According to the composition in Table 1, different zwitterionic monomers and different cross-linking agents are used, and the amounts of each reactant are adjusted to prepare the precursor solutions of the adhesion layer and the anti-adhesion layer respectively.

[0060] Preparation of Janus zwitterionic hydrogel anti-adhesion film: Inject the precursor solution of the adhesion layer into the groove of a 2 cm × 2 cm × 2 mm silicone mold, and polymerize it under a 365 nm ultraviolet lamp at 10 °C for 60 min. After molding, inject the precursor solution of the anti-adhesion layer into the mold, and polymerize it under a 365 nm ultraviolet lamp at 10 °C for 60 min to obtain the Janus zwitterionic hydrogel anti-adhesion film.

[0061] Example 15

[0062] According to the method of Example 1, the operation is the same as that of Example 1. According to the composition in Table 1, different zwitterionic monomers and different cross-linking agents are used, and the amounts of each reactant are adjusted to prepare the precursor solutions of the adhesion layer and the anti-adhesion layer respectively.

[0063] Preparation of Janus zwitterionic hydrogel anti-adhesion film: Inject the precursor solution of the adhesion layer into the groove of a 2 cm×2 cm×2 mm silicone mold, and polymerize it under a 365 nm ultraviolet lamp at 60 °C for 45 min. After molding, inject the precursor solution of the anti-adhesion layer into the mold, and polymerize it under a 365 nm ultraviolet lamp at 60 °C for 45 min to obtain the Janus zwitterionic hydrogel anti-adhesion film.

[0064] The properties of the anti-adhesion layer and adhesion layer of the Janus zwitterionic hydrogel anti-adhesion films prepared in Examples 1 to 15 are shown in Tables 2 and 3.

[0065] Table 1 Composition of precursor solutions for preparing Janus zwitterionic hydrogel anti-adhesion films in Examples 1 to 15

[0066]

[0067]

[0068] a Refers to the concentration of the monomer in the precursor solution. b Refers to the percentage of the cross-linking agent or initiator in the mass of the zwitterionic monomer.

[0069] Zm: Zwitterionic monomer; Cm: Cross-linking agent.

[0070] Table 2 Anti-adhesion performance of the anti-adhesion layer of the Janus zwitterionic hydrogel anti-adhesion films prepared in Examples 1 to 15

[0071]

[0072] MPC: 2-methacryloyloxyethyl phosphorylcholine; CBMA: Carboxybetaine methacrylate; Ad pro : Protein adhesion amount; Ad pla : Platelet adhesion amount; Ad bac : Bacterial adhesion amount; Ad cel : Cell adhesion amount.

[0073] As can be seen from the data in Table 2, compared with the zwitterionic monomer SBMA, the swelling ratios of the anti-adhesion layers formed by the other two zwitterionic monomers (MPC, CBMA) are almost unchanged, and they also exhibit very excellent superhydrophilic, anti-protein adhesion, anti-platelet adhesion, anti-bacterial adhesion and anti-cell adhesion properties. This benefits from the high hydrophilicity of the zwitterionic polymer and its extremely low interactions with proteins, platelets, bacteria, cells, etc. With the increase in the concentration of the SBMA monomer, both the hydrophilicity and anti-bioadhesion properties of the anti-adhesion layer are improved, but the swelling ratio also increases; with the increase in the concentration of the NAGA monomer, the swelling ratio of the anti-adhesion layer decreases, but both its hydrophilicity and anti-bioadhesion properties decrease. With the increase in the concentration of the cross-linking agent MBA, the anti-swelling property of the anti-adhesion layer is improved, but both its hydrophilicity and anti-bioadhesion properties decrease. With the change in the dosage of the photoinitiator I-2959, the hydrophilicity and anti-bioadhesion properties of the anti-adhesion layer are almost unchanged. This is because with the increase in the cross-linking agent content, the polymer chain segments are more tightly entangled with each other, its water absorption decreases, and the surface hydration ability decreases. Therefore, its swelling ratio decreases and the anti-bioadhesion property decreases. The overall performance of the hydrogel is almost independent of the initiator content.

[0074] The adhesion time, adhesion strength, bursting pressure, hemostasis time and swelling ratio of the adhesion layer of the Janus zwitterionic hydrogel anti-adhesion film prepared with different types of reactants and different dosages of reactants were measured, as shown in Table 3.

[0075] Table 3 Adhesion layer properties of the Janus zwitterionic hydrogel anti-adhesion films prepared in Examples 1 to 15

[0076]

[0077]

[0078] As can be seen from the data in Table 3, compared with the zwitterionic monomer SBMA, the swelling rate of the adhesion layer, the adhesion time with tissue, the bonding strength and the hemostasis time formed by the other two zwitterionic monomers (MPC, CBMA) showed no significant changes. Compared with the crosslinking agent MBA, the adhesion time with tissue, the bonding strength, the hemostasis time and the swelling rate of the adhesion layer formed by the other four crosslinking agents (MSBA, EBA, CBBA, PEGDA) showed no significant changes. With the increase in the concentration of DMA monomer, the adhesion performance of the front adhesion layer of the hydrogel anti-adhesion film was improved, but when the concentration of DMA monomer increased to a certain range, its front adhesion performance no longer changed; with the increase in the concentration of NAGA monomer, the swelling rate of the front adhesion layer of the hydrogel anti-adhesion film decreased, but its adhesion performance decreased; with the increase in the concentration of SBMA monomer, both the adhesion time and the hemostasis time decreased, but the overall adhesion strength decreased and the swelling rate increased. This is because the zwitterionic hydrogel can quickly adsorb the water on the tissue surface, making the front adhesion layer of the hydrogel anti-adhesion film quickly adhere to the tissue surface. However, due to its own swelling effect, the adhesion strength decreased. With the increase in the concentration of crosslinking agent MBA, the anti-swelling performance of the adhesion layer was improved, the adhesion time and the hemostasis time were both prolonged, and the shear strength and the burst pressure decreased. With the change in the dosage of photoinitiator I-2959, the adhesion performance of the adhesion layer remained almost unchanged. This is because with the increase in the crosslinking agent content, the polymer chain segments are more tightly entangled with each other, its water absorption decreases, the water absorption rate slows down, and the time taken to absorb the water molecules on the surface of the wet tissue is prolonged. Therefore, the adhesion time and the hemostasis time are prolonged and the swelling rate decreases. Due to the tight entanglement of the polymer chain segments with each other, the number of catechol groups with high affinity in direct contact with the tissue surface decreases. Therefore, the adhesion performance decreases.

[0079] Comparative Example 1

[0080] According to the method of Example 1, the operation was the same as that of Example 1, the reaction temperature and reaction time were the same as those of Example 1, except that the anti-adhesion layer had no zwitterionic monomer and the concentrations of other monomers remained unchanged.

[0081] Comparative Example 2

[0082] According to the method of Example 1, the operation was the same as that of Example 1, the reaction temperature and reaction time were the same as those of Example 1, except that the adhesion layer had no DMA monomer and the concentrations of other monomers remained unchanged.

[0083] Table 4 Anti-adhesion performance of the anti-adhesion layer of the Janus zwitterionic hydrogel anti-adhesion film prepared in Comparative Example 1

[0084]

[0085] As can be seen from the data in Table 4, when there is no zwitterionic monomer in the anti-adhesion layer, the swelling rate of the anti-adhesion layer decreases significantly, but the hydrophilicity of the material surface decreases, and the anti-protein adhesion, anti-platelet adhesion, anti-bacterial adhesion and anti-cell adhesion decrease. This is because zwitterionic polymers have high hydrophilicity and can form a hydration layer on the surface of the anti-adhesion layer, reducing the occurrence of biological adhesion.

[0086] Table 5 Adhesion layer properties of the Janus zwitterionic hydrogel anti-adhesion film prepared in Comparative Example 2

[0087]

[0088] As can be seen from the data in Table 5, when there is no DMA monomer in the adhesion layer, the adhesion layer of the Janus zwitterionic hydrogel anti-adhesion film has no adhesion performance on the tissue surface. This is because the catechol group in DMA can bind to various nucleophilic groups (such as amino, mercapto and imidazole groups) in polypeptides and proteins on the tissue surface, thus adhering to the tissue surface.

[0089] Figure 1-Figure 12 The test method is as follows:

[0090] Characterization method of Janus zwitterionic hydrogel anti-adhesion film:

[0091] 1. Morphology test of Janus zwitterionic hydrogel anti-adhesion film under scanning electron microscope:

[0092] Use a field emission scanning electron microscope (S-4800, HITACHI) to analyze the surface and cross-sectional morphology of the Janus zwitterionic hydrogel anti-adhesion film. First, immerse the sample in PBS solution, perform ultrasonic cleaning, and then perform freeze-drying. Cut the sample into 1×1 cm 2 size, sputter gold on the surface, and fix it on the observation platform with conductive glue. During the observation, set the acceleration voltage to 15 eV.

[0093] 2. 180° peel test of Janus zwitterionic hydrogel:

[0094] Measure the interfacial bonding strength between the two layers of the Janus patch through a 180° peel experiment. First, prepare a Janus patch with a width of 20 mm, a length of 200 mm, and a thickness of 2 mm, and fix the adhesion layer and the anti-adhesion layer on the tensile machine respectively. Stretch at a strain rate of 5 mm / min and record the tensile force during the peeling process. The peel strength is the ratio of the tensile force to the contact width between the two layers. According to the test results, the bonding strength between the hydrogel and the substrate material can be calculated, in units of N / m.

[0095] 3. Contact angle test of Janus zwitterionic hydrogel anti-adhesion film:

[0096] The hydrophilicity of both sides of the Janus zwitterionic hydrogel anti-adhesion film was characterized using a DSA100 (Kruss GmbH, Germany) contact angle measuring instrument. The dried Janus zwitterionic hydrogel anti-adhesion film was attached to a glass slide and mounted on the goniometer. In the static contact angle measurement, a total of 3 μL of double-distilled water was dropped onto the air-side surface of the sample at room temperature and a relative humidity of 80%, and the water contact angle of the surface was measured. Five measurements were performed for each group, and the average value was taken.

[0097] 4. Adhesion performance test of Janus zwitterionic hydrogel anti-adhesion film:

[0098] The adhesion performance tests in this study were divided into the following two categories: lap shear test and T-peel test, which characterized the shear strength (SS) and T-peel strength (TPS) of the Janus zwitterionic hydrogel anti-adhesion film, respectively.

[0099] For the lap shear test, one surface of the Janus zwitterionic hydrogel anti-adhesion film and the wet tissue sample were respectively bonded to a rigid polyester backing with cyanoacrylate, and then a pressure of 500 g was applied to each of the two pads to make the patch fit with the tissue sample for 10 min. Then, the universal testing machine was loaded in the lap shear test mode, and the loading rate was set to 5 mm·min -1 , and the test was carried out until the Janus zwitterionic hydrogel anti-adhesion film was pulled off from the wet tissue. The shear strength (SS) of the Janus zwitterionic hydrogel anti-adhesion film was calculated, and the definition formula is as follows:

[0100]

[0101] In the formula, F max represents the maximum pull-off force, and L and W represent the length and width of the initial contact area between the Janus zwitterionic hydrogel anti-adhesion film and the wet tissue sample, respectively.

[0102] For the T-peel test, two L-shaped backing materials were prepared in advance, and one side of the Janus zwitterionic hydrogel anti-adhesion film and the wet tissue was bonded to a rigid polyester backing with cyanoacrylate. Then, the exposed sides of the Janus zwitterionic hydrogel anti-adhesion film and the wet tissue were brought into contact with each other to form a T-shape. After applying a pressure of 500 g to make the Janus zwitterionic hydrogel anti-adhesion film fit with the tissue for 10 min, the universal testing machine was loaded in the T-peel test mode, and the loading rate was set to 5 mm·min -1 and the test was carried out until the Janus zwitterionic hydrogel anti-adhesion film was separated from the wet tissue. The T-peel strength (TPS) of the Janus zwitterionic hydrogel anti-adhesion film was calculated, and the definition formula is as follows:

[0103]

[0104] In the formula, F max represents the maximum peeling force, and W represents the width of the initial contact area between the Janus zwitterionic hydrogel adhesion prevention film and the wet tissue sample.

[0105] 5. Blasting pressure test of the Janus zwitterionic hydrogel adhesion prevention film:

[0106] First, cut the prepared wet tissue sample into circular slices and make a circular hole with a diameter of 8 mm in its center. Then fix the wet tissue to the opening of a cylindrical stainless steel container with an open top, place one side of the circular Janus zwitterionic hydrogel adhesion prevention film with a diameter of 15 mm on the circular hole of the tissue sample, and apply a force of 500 g for 10 min to bond the two. Introduce air into the container at a speed of 10 mL·min -1 to generate a stable increase in internal pressure, and record the magnitude of the internal pressure when the Janus zwitterionic hydrogel adhesion prevention film separates from the wet tissue.

[0107] 6. Anti-bioadhesion test of the adhesion prevention layer of the Janus zwitterionic hydrogel adhesion prevention film:

[0108] Protein adhesion test: The test method used in the protein adsorption experiment is the BCA protein kit method. The adhesion behavior of a mixed solution of three proteins, bovine serum albumin, fibrinogen, and γ-GL, on the sample is tested and characterized. First, cut the sample into a size of 1×1 cm 2 , soak it in PBS solution at 37°C for 6 h, then group the cut samples and immerse them in the mixed solution of proteins (2.0 mg·mL -1 bovine serum albumin solution, 0.3 mg·mL -1 fibrinogen solution, 1 mg·mL -1 γ-GL protein solution) and soak for 2 h at 37°C. Subsequently, rinse the sample with fresh PBS solution and transfer it to a 96-well plate containing 1.0 wt% sodium dodecyl sulfate (SDS) solution, and ultrasonically treat it for 20 min at room temperature to separate the adsorbed proteins. Finally, use a BCA protein analysis kit to measure its absorbance at 562 nm to obtain the protein concentration.

[0109] Platelet adhesion test: First, separate whole blood at a centrifugal speed of 1500 r / min for 15 min using a high-speed centrifuge, and aspirate the supernatant as platelet-rich plasma (PRP) for standby. Prepare the sample (1×1 cm 2) After sterilization, place the samples in a 12-well plate. Use a pipette to aspirate 100 μL of PRP and evenly drop it onto the surface of the samples. Then place the samples in a constant temperature water bath at 37 °C and incubate them with shaking for 1 h. After taking out the samples, wash the samples 5 times with PBS, then add a 2.5 wt% glutaraldehyde solution to submerge the samples, and fix them overnight at 4 °C. Take out the samples from the glutaraldehyde solution, and prepare anhydrous ethanol solutions with concentrations of 50%, 75%, 90%, and 100% respectively. Immerse the dried samples into the above gradient anhydrous ethanol solutions for dehydration, and the dehydration time for each time is 15 min. Use a scanning electron microscope to observe the number of platelets adhered to the surface of the samples.

[0110] Cell adhesion test: Cut the samples into 5×5 mm 2 size, and sterilize them by ultraviolet irradiation at room temperature for 4 h. Then, incubate the samples with a mouse embryonic fibroblast (3T3-L1) cell suspension (5×10 5 cells / mL, 1 mL / well) at 37 °C for 12 h. Separate the non-adherent cells with PBS, fix the adherent cells with paraformaldehyde, and then soak them in Triton X-100 for 5 min. Then, stain with phalloidin and DAPI respectively to show the actin cytoskeleton and cell nuclei. Use a laser scanning confocal microscope to observe the adhesion of cells on the surface of the samples to determine the anti-cell adhesion performance of the samples.

[0111] Bacterial adhesion test: Select Staphylococcus aureus and Escherichia coli to evaluate the anti-bacterial adhesion ability of the samples. Cut the samples into 5×5 mm 2 size, and after sterilization, soak them in sterile PBS buffer at room temperature for 6 h. Subsequently, transfer the samples to a 24-well plate, and soak them with a single bacterial suspension (1 mL, 1.0×10 8 CFU·mL -1 ) for 6 h, and then rinse them three times with sterile PBS to remove weakly adherent or non-adherent bacteria. Fix the samples in a 4% glutaraldehyde solution for 12 h, and then dehydrate them with a series of ethanol with increasing concentrations (50%, 60%, 70%, 80%, 90%, and 100% respectively, 30 min each time). For the Escherichia coli group, observe by SEM; for the Staphylococcus aureus group, first stain with DAPI and then observe with a fluorescence microscope.

[0112] 7. In vitro organ adhesion and sealing ability test of Janus zwitterionic hydrogel anti-adhesion film:

[0113] The adhesion performance of the Janus zwitterionic hydrogel anti-adhesion film to different organs was evaluated through in vitro organ adhesion experiments. Tissue specimens such as porcine heart, lung, muscle, kidney, trachea, and aorta were taken for adhesion tests. For the adhesion test of the isolated porcine heart, a 1-cm long wound was first made with a scalpel, and pig blood was sprayed to simulate tissue injury. Then, the Janus zwitterionic hydrogel anti-adhesion film was adhered to the wound, and after pressing for 15 seconds, the wound was sealed. After 8 hours, the adhesion situation was checked, and the Janus zwitterionic hydrogel anti-adhesion film was torn off. To further test the sealing effect of the Janus zwitterionic hydrogel anti-adhesion film on the injured small intestine, two 5-mm long incisions were made on the surface of the small intestine with a scalpel, and then one of the incisions was sealed with the Janus zwitterionic hydrogel anti-adhesion film. After being placed underwater for 12 hours, one side of the isolated small intestine was sealed at one end, and air was injected from the other side. The generation of air bubbles at the wound was observed underwater to check the airtightness of the incision sealed by the Janus zwitterionic hydrogel anti-adhesion film. To further test the sealing effect of the Janus zwitterionic hydrogel anti-adhesion film on the injured trachea and lung lobe, 5-mm long incisions were made on the trachea and lung lobe of the pig with a scalpel respectively. Air was injected into the trachea to inflate the pig lung, and the generation of air bubbles at the wound was observed underwater. Then, the Janus zwitterionic hydrogel anti-adhesion film was used to seal the incision, and the generation of air bubbles at the wound was observed again after sealing to determine the sealing effect.

[0114] 8. Test on the hemostatic ability of the Janus zwitterionic hydrogel anti-adhesion film in the carotid artery of rabbits in vivo:

[0115] New Zealand white rabbits used in the experiment were weighed and recorded in sequence. Ketamine at 20 mg / kg and xylazine at 3 mg / kg were selected to anesthetize them. After anesthesia, they were fixed on the operating table, and continuous anesthesia was performed using a respiratory anesthetic machine. After skin preparation and disinfection, the carotid artery was exposed. After both ends were clamped with hemostatic clips, a carotid artery injury model was established with a needle with a diameter of 2 mm. After the hemostatic clip was loosened to confirm blood flow, the hemostatic clip was closed, and the injured area was sealed with the Janus zwitterionic hydrogel anti-adhesion film. After pressing for 15 seconds, the hemostatic clip was opened, and it was observed whether the Janus zwitterionic hydrogel anti-adhesion film sealed the injured area and whether there was blood flow. On the 3rd, 5th, and 7th days after the experiment, the rabbits were euthanized by injecting an overdose of pentobarbital sodium, and their carotid arteries were taken out. The adhesion situation of the wound surface was observed, and HE and Masson staining were performed to observe the wound repair situation.

[0116] 9. In vitro cytotoxicity test of the Janus zwitterionic hydrogel anti-adhesion film:

[0117] The sample (2×2 mm 2 ) was sterilized by ultraviolet irradiation at room temperature for 4 hours and then placed in a 96-well plate. The cells were diluted to 1×10 5The concentration of cells was x cells / mL. 100 μL was added to each well of a 96-well plate, and after 24 h, the cell proliferation rate was detected using the Cell Counting Kit-8 (CCK8).

[0118] 10. Testing of the Janus zwitterionic hydrogel anti-adhesion membrane in a cecum-abdominal wall adhesion model in rats:

[0119] Fifteen healthy rats were selected for the experiment and divided into a control group (without any anti-adhesion treatment), a commercial anti-adhesion membrane treatment group, and a Janus zwitterionic hydrogel anti-adhesion membrane treatment group. The entire experiment was carried out under sterile conditions. The rats were first anesthetized with 10% chloral hydrate, placed in the supine position, then the abdomen was shaved and disinfected with 0.5% iodine. The abdomen was carefully incised with a scalpel to form a wound about 3 cm long, and the internal abdominal organs were exposed. The serosa surface of the cecum was carefully rubbed with medical sandpaper until slight damage and bleeding appeared on the serosa surface but without perforation. Subsequently, the peritoneum was gently scraped with medical sandpaper to expose the muscular layer of the right abdominal wall. For the control group, the abdominal wall and cecal wound surfaces were slightly sutured to make the two wound surfaces contact each other. For the CF group, a polylactic acid film was placed between the abdominal wall wound surface and the cecal wound surface, and the three were sutured. For the Janus zwitterionic hydrogel anti-adhesion membrane group, the adhesive surface of the Janus zwitterionic hydrogel anti-adhesion membrane was directly attached to the wound abdominal wall, completely covering the wound surface. On the 14th and 28th days after the experiment, each rat was laparotomized, and the wound adhesion situation was observed and photographed. All experimental rats were euthanized by injecting an overdose of pentobarbital sodium on the 28th day.

[0120] 11. Testing of the Janus zwitterionic hydrogel anti-adhesion membrane in a rabbit abdominal wall hernia repair model:

[0121] New Zealand white rabbits used in the experiment were weighed and recorded in sequence. They were anesthetized with 20 mg / kg ketamine and 3 mg / kg xylazine. After anesthesia, they were placed in the supine position. After hair removal and skin preparation, the skin was disinfected with 0.5% iodophor. An incision about 3 cm long was made in the midline of the lower abdomen, and the skin and subcutaneous tissues were incised layer by layer. The space between the skin and the abdominal wall muscles was dissected to expose the bilateral rectus abdominis muscles. Two complete muscle defects (retaining the peritoneum) with a diameter of 1 cm were excised from the flat abdominal muscles to make an abdominal wall hernia model in this way. The pre-prepared commercial PP patch and the Janus zwitterionic hydrogel anti-adhesion membrane of Example 1 were implanted into the abdominal cavity. The commercial PP patch was fixed along the edge in sequence, and the Janus zwitterionic hydrogel anti-adhesion membrane was fixed using its own adhesion property. Subsequently, the abdomen was closed layer by layer with sutures. On the 14th and 28th days after the operation, 6 experimental rabbits were taken out at each time point. After abdominal skin preparation, the adhesion, healing and other situations were observed and photographed, and then they were stored in formalin solution for later use.

[0122] The various embodiments in this specification are described in a progressive manner. Each embodiment focuses on the differences from other embodiments. For the same or similar parts among the various embodiments, reference can be made to each other.

[0123] The above description of the disclosed embodiments enables those skilled in the art to implement or use the present invention. Various modifications to these embodiments will be obvious to those skilled in the art. The general principles defined herein can be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention will not be limited to these embodiments shown herein, but rather will be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A Janus zwitterionic hydrogel anti-adhesion film, characterized in that, the anti-adhesion film is composed of an adhesion layer and an anti-adhesion layer; the adhesion layer is a copolymer of a zwitterionic monomer, N-acryloylglycine amide, 3-methacryloyldopamine and a chemical cross-linking agent; the anti-adhesion layer is a copolymer of a zwitterionic monomer, N-acryloylglycine amide and a chemical cross-linking agent.

2. The Janus zwitterionic hydrogel anti-adhesion film according to claim 1, characterized in that, the adhesion layer is a copolymer formed by a zwitterionic monomer, N-acryloylglycine amide, 3-methacryloyldopamine and a chemical cross-linking agent under the initiation of an initiator; the anti-adhesion layer is a copolymer formed by a zwitterionic monomer, N-acryloylglycine amide and a chemical cross-linking agent under the initiation of an initiator.

3. The Janus zwitterionic hydrogel anti-adhesion film according to claim 1, characterized in that, in the copolymer forming the adhesion layer, the ratio of the total mass of N-acryloylglycine amide monomer to the total mass of the zwitterionic monomer is 0.125 to 2, the ratio of the total mass of 3-methacryloyldopamine monomer to the total mass of the zwitterionic monomer is 0.125 to 2, and the percentage of the chemical cross-linking agent in the total mass of the zwitterionic monomer is 0.5 w% to 15 w%; in the copolymer forming the anti-adhesion layer, the ratio of the total mass of N-acryloylglycine amide monomer to the total mass of the zwitterionic monomer is 0.125 to 0.75, and the mass percentage of the chemical cross-linking agent in the zwitterionic monomer is 0.5 w% - 15 w%.

4. The Janus zwitterionic hydrogel anti-adhesion film according to claim 1, characterized in that, the zwitterionic monomer is selected from any one of methacryloylethyl sulfobetaine, 2-methacryloyloxyethyl phosphorylcholine, and carboxybetaine methacrylate.

5. The Janus zwitterionic hydrogel anti-adhesion film according to claim 1, characterized in that, the chemical cross-linking agent is selected from one or more of N,N-methylenebisacrylamide, N,N-bis(acryloyl)cystamine, ethylene glycol dimethacrylate, bis-methacryloylethyl carboxybetaine, and poly(ethylene glycol) diacrylate.

6. The Janus zwitterionic hydrogel anti-adhesion film according to claim 2, characterized in that, the initiator is selected from photoinitiator 2959.

7. A preparation method of the Janus zwitterionic hydrogel anti-adhesion film according to any one of claims 1 to 6, characterized in that, comprises the following steps: (1) Prepare an adhesion layer precursor solution, and the adhesion layer precursor solution is a mixed aqueous solution of a zwitterionic monomer, N-acryloylglycine amide, 3-methacryloyldopamine, an initiator and a chemical cross-linking agent; (2) Prepare an anti-adhesion layer precursor solution, and the anti-adhesion layer precursor solution is a mixed solution of a zwitterionic monomer, N-acryloylglycine amide, an initiator and a cross-linking agent. (3) Inject the precursor solution of the adhesion layer into the mold, and polymerize it with an ultraviolet lamp at 10 - 60 °C. After molding, inject the precursor solution of the anti-adhesion layer into the mold and polymerize it with an ultraviolet lamp at 10 - 60 °C to obtain the Janus zwitterionic hydrogel anti-adhesion film.

8. A method for preparing a Janus zwitterionic hydrogel anti-adhesion film according to claim 7, wherein, in the precursor solution of the adhesion layer, the concentration of the zwitterionic monomer is 10 w% - 40 w%, the concentration of the N-acryloylglycine amide monomer is 5 w% - 20 w%, the concentration of the 3-methacryloyldopamine monomer is 5 w% - 20 w%, the dosage of the chemical cross-linking agent accounts for 0.5 w% - 15 w% of the mass of the zwitterionic monomer, and the dosage of the initiator accounts for 0.5 w% - 20 w% of the mass of the zwitterionic monomer; in the precursor solution of the anti-adhesion layer, the concentration of the zwitterionic monomer is 10 w% - 40 w%, the concentration of the N-acryloylglycine amide monomer is 5 w% - 13 w%, the dosage of the cross-linking agent accounts for 0.5 w% - 15 w% of the mass of the zwitterionic monomer, and the dosage of the initiator accounts for 0.5 w% - 20 w% of the mass of the zwitterionic monomer.

9. An application of a Janus zwitterionic hydrogel anti-adhesion film according to any one of claims 1 - 6 in the preparation of a medical anti-adhesion patch.

10. An application of a Janus zwitterionic hydrogel anti-adhesion film according to claim 9 in the preparation of a medical anti-adhesion patch, wherein, the Janus zwitterionic hydrogel anti-adhesion film is used alone or in combination with any one or more of drugs, active factors, polypeptide bioactive substances, and electrodes to prepare a medical anti-adhesion patch.

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