Anion-π-based zwitterionic supramolecular hydrogel and its preparation method and application

Abstract

The present invention discloses a zwitterionic supramolecular hydrogel based on anion-π and its preparation method and application. The hydrogel is self-assembled by a zwitterionic copolymer and a p-styrene sulfonic acid copolymer. In the present invention, the supramolecular hydrogel only needs to be rapidly formed by simply mechanically mixing the zwitterionic polymer copolymer and the p-styrene sulfonate copolymer solution, and the dynamic cross-linking characteristics give it excellent injectability and self-repairing properties. This hydrogel is composed of zwitterions with excellent immune compatibility and poly (p-styrene sulfonate) with heparin-like properties, so it has excellent immune compatibility and can play a role in various in vivo application scenarios. In addition, this dynamic cross-linking network based on anion-π interaction and the heparin-like structure are very beneficial to the preservation of proteins and have broad application prospects in the fields of protein delivery and cell encapsulation.

Description

Technical Field

[0001] The present invention relates to the technical field of medical materials, and in particular to an anion-π-based zwitterionic supramolecular hydrogel and a preparation method and application thereof. Background Art

[0002] Hydrogels are a class of polymer materials prepared through chemical or physical crosslinking. They possess a three-dimensional network structure that can firmly bind a large number of water molecules, forming a gel system with a high water content. This structure gives hydrogels excellent biocompatibility, thus holding enormous potential for application in the biomedical field.

[0003] Chemically cross-linked hydrogels exhibit good structural stability due to the stability of their chemical bonds. However, this high stability also brings inconvenience in application. Chemically cross-linked hydrogels with fixed shapes usually need to be implanted surgically, which not only increases the complexity of the operation but also limits the choice of implantation site. The surgical process is also prone to causing large areas of tissue damage, and lacks flexibility when filling irregularly shaped tissues, which seriously restricts its scope of application.

[0004] In recent years, supramolecular hydrogels based on non-covalent interactions have attracted widespread attention due to their unique dynamic reversible properties. These hydrogels construct dynamic cross-linked networks through non-covalent bonds such as hydrogen bonds and hydrophobic interactions, offering significant advantages such as injectability and self-healing properties.

[0005] CN119570070A discloses a citric acid-cyclodextrin derivative controlled-release composite supramolecular hydrogel and its preparation method. The method comprises mixing citric acid and cyclodextrin and dissolving them thoroughly through ultrasonic treatment; adjusting the pH of the solution to 0.5-2.0 to promote cross-linking, and conducting the cross-linking reaction at 40-60°C for 3-5 hours under optimized temperature conditions; cooling the resulting solution to room temperature, and adding an appropriate amount of a cross-linking agent to allow self-assembly to form a gel; and drying the gel to form the desired morphology while maintaining a stable structure. By optimizing the cross-linking reaction conditions between citric acid and cyclodextrin, a hydrogel with a unique network structure and strong mechanical strength is obtained. This material exhibits excellent water absorption, biocompatibility, and mechanical properties, making it particularly suitable for use in tissue engineering, biomedicine, and environmental protection. The preparation process includes multiple steps, including chemical treatment of the raw materials, control of reaction conditions, self-assembly and cross-linking reactions, molding, and drying.

[0006] CN110885391A discloses a supramolecular hydrogel based on double hydrogen bonds and its preparation method. Acryloyl alaninamide (NAAA) monomers are prepared by reacting alaninamide hydrochloride with acryloyl chloride. This monomer possesses a double hydrogen bond structure. This monomer is mixed with water to form a solution of a certain concentration and then polymerized by ultraviolet light to form supramolecular hydrogels based on double hydrogen bonds of varying solid contents. The comonomers with double hydrogen bonds are polymerized by free radicals to form a hydrogen-bonded physical hydrogel. Compared to gels prepared from acrylamide monomers, these double hydrogen bond monomers have a higher hydrogen bond crosslink density. Therefore, hydrogels can be formed using the double hydrogen bond monomers without the addition of any crosslinking agent, and the resulting supramolecular hydrogels are more stable than acrylamide hydrogels.

[0007] Anion-π interactions are an emerging non-covalent force, characterized by electrostatic attraction between electron-rich anions (such as sulfonates and carboxylates) and electron-deficient aromatic rings (such as hexafluorophenyl and triazine). Despite well-established mechanisms of anion-π interactions in biological processes such as enzyme catalysis and bioadhesion, their application in materials engineering remains relatively limited. Currently, no supramolecular hydrogels based on anion-π interactions have been reported, limiting our understanding and application of this interaction in materials design. Summary of the Invention

[0008] In view of the difficulty of supramolecular hydrogels to achieve both structural stability and injectability, the present invention provides a zwitterionic supramolecular hydrogel based on anion-π. The hydrogel is obtained by self-assembly of a zwitterionic copolymer and a styrene sulfonate copolymer. Its dynamic cross-linking properties give it excellent injectability and self-healing properties, and it has broad application prospects in the medical field.

[0009] To achieve the above object, the technical solution adopted by the present invention is:

[0010] A zwitterionic supramolecular hydrogel based on anion-π is self-assembled from a polyzwitterionic copolymer and a p-styrenesulfonic acid copolymer.

[0011] The supramolecular hydrogel of the present invention rapidly forms by simply mechanically mixing a zwitterionic copolymer with a p-styrene sulfonate copolymer solution. Its dynamic crosslinking properties impart excellent injectability and self-healing properties. Composed of zwitterionics with excellent immunocompatibility and p-styrene sulfonate copolymers with heparin-like properties, this hydrogel exhibits excellent immunocompatibility and is suitable for a variety of in vivo applications. Furthermore, this dynamic crosslinking network based on anion-π interactions and heparin-like structure are highly beneficial for protein preservation, promising broad applications in protein delivery and cell encapsulation.

[0012] The zwitterionic copolymer is obtained by copolymerizing raw materials including zwitterionic monomers, wherein the zwitterionic monomers include any one or more of sulfonyl betaine methacrylate (SBMA), carboxylic betaine methacrylate (CBMA), and methacryloxyethyl phosphorylcholine (MPC);

[0013] The p-styrene sulfonate copolymer is obtained by copolymerizing monomers containing p-styrene sulfonic acid, and the monomers containing p-styrene sulfonic acid include one or more of p-styrene sulfonic acid, sodium p-styrene sulfonate, potassium p-styrene sulfonate, and lithium p-styrene sulfonate.

[0014] The weight average molecular weight of the zwitterionic copolymer is 50,000 to 5,000,000; the weight average molecular weight of the p-styrenesulfonate copolymer is 50,000 to 5,000,000. The larger the molecular weight, the higher the gel strength and the better the elasticity.

[0015] The molar ratio of zwitterions to p-styrenesulfonic acid repeating units in the zwitterionic copolymer and p-styrenesulfonate copolymer is 1:0.1-10. Preferably, the molar ratio of zwitterions to p-styrenesulfonic acid repeating units in the zwitterionic copolymer and p-styrenesulfonate copolymer is 1:0.25-4; preferably, the molar ratio of zwitterions to p-styrenesulfonic acid repeating units in the zwitterionic copolymer and p-styrenesulfonate copolymer is 1:0.5-2; when the molar ratio of zwitterions to p-styrenesulfonic acid repeating units in the zwitterionic copolymer and p-styrenesulfonate is 1:1, the interaction is strongest, and the hydrogel has better elasticity and strength.

[0016] The polymerization monomer raw material of the zwitterionic copolymer further includes a first comonomer; the first comonomer includes one or more monomers selected from acrylates, methacrylates, and acrylamides;

[0017] Preferably, the zwitterionic monomer accounts for more than 50% by weight of the polymerizable monomer raw materials of the zwitterionic copolymer to effectively form a gel, and more preferably, the zwitterionic monomer accounts for more than 60%, more than 70%, or more than 80% by weight.

[0018] The polymerization monomer raw material of the styrene sulfonate copolymer further includes a second comonomer; the second comonomer includes one or more of acrylic acid, methacrylic acid, maleic anhydride, styrene or its derivatives, acrylates, methacrylates, and acrylamides;

[0019] Preferably, the mass proportion of the p-styrene sulfonate monomer in the polymerization monomer raw material of the p-styrene sulfonate copolymer is 50% to effectively form a gel, and more preferably, the mass proportion is greater than 60%, greater than 70%, or greater than 80%.

[0020] The present invention also provides a method for preparing the anion-π based zwitterionic supramolecular hydrogel, comprising the steps of:

[0021] Step 1: copolymerizing a raw material containing a zwitterionic monomer to obtain a zwitterionic copolymer; copolymerizing a raw material containing a monomer containing p-styrenesulfonic acid to obtain a p-styrenesulfonate copolymer;

[0022] Step 2: dissolving the zwitterionic copolymer and the p-styrenesulfonate copolymer in an aqueous solution respectively, mixing them and self-assembling them to obtain the zwitterionic supramolecular hydrogel.

[0023] In step 2, the total concentration of the zwitterionic copolymer and the p-styrene sulfonate copolymer in the aqueous solution is 1 to 60 wt %. Preferably, the mass concentration of the zwitterionic copolymer and the p-styrene sulfonate copolymer in the aqueous solution is 0.1 to 50 wt %, more preferably 0.1 to 20 wt %, and even more preferably 0.1 to 15 wt %.

[0024] The aqueous phase solution includes one or more of water, PBS buffer, physiological saline, Hanks buffer, and Tris buffer.

[0025] In step 1, the copolymerization contains an initiator, the copolymerization temperature is 60-90° C., the reaction is carried out for 6-18 hours, and the product is dialyzed and freeze-dried to obtain a copolymer.

[0026] The initiator includes one or more of sodium persulfate, ammonium persulfate, and potassium persulfate, and the mass fraction of the initiator in the aqueous solution is 0.1-5%;

[0027] The present invention also provides the use of the anion-π zwitterionic supramolecular hydrogel in the preparation of immunocompatible drugs or medical materials. For example, it can be used in diabetic wound dressings. The supramolecular hydrogel based on anion-π interactions, composed of super-hydrophilic zwitterions and p-styrenesulfonate with a heparin-like structure, stabilizes basic fibroblast growth factor, which promotes wound healing. Furthermore, the zwitterionic structure can promote angiogenesis at the wound site. Therefore, the hydrogel can be used to load basic fibroblast growth factor and be applied to diabetic wound healing dressings. The data are as follows:

[0028] Another example is the use in cell culture scaffolds: the supramolecular hydrogel based on anion-π interaction is composed of zwitterionic polymers with excellent anti-fouling properties, so the hydrogel has excellent anti-cell and anti-bacterial adhesion properties. At the same time, the highly immune-compatible hydrogel based on this dynamic covalent bond structure can be used as a cell culture scaffold.

[0029] Compared with the prior art, the present invention has the following beneficial effects:

[0030] (1) The present invention prepares anion-π zwitterion supramolecular hydrogels, breaking through the traditional technical prejudice that "anion-π interaction is difficult to support the gel network" and broadening the preparation principle of supramolecular hydrogels;

[0031] (2) The prepared supramolecular hydrogel does not require sophisticated structural design and can be quickly and easily prepared using commercially available zwitterionic polymers and styrene sulfonate copolymers, avoiding the introduction of other chemical substances such as cross-linking agents. The process is simple and easy to promote.

[0032] (3) The anionic-π supramolecular hydrogel prepared in the present invention has excellent immunocompatibility and shows broad application prospects in the biomedical field. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] In order to more clearly illustrate the technical solutions of the embodiments of the present application, the following is a brief introduction to the drawings required for use in the embodiments of the present application. It should be understood that the following drawings only show certain embodiments of the present application and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other relevant drawings can be obtained based on these drawings without paying any creative work.

[0034] Figure 1 The strain scan of the anionic-π supramolecular hydrogel prepared in Example 1 ( Figure 1 a) and angular frequency sweep test ( Figure 1 b) Results.

[0035] Figure 2 UV spectra of anionic-π supramolecular hydrogel and two copolymer raw materials.

[0036] Figure 3 The heat released by the titration of PSBMA solution with PSSNa solution measured by nano titration calorimeter and the Gibbs free energy, enthalpy change and entropy change obtained after fitting calculation.

[0037] Figure 4 Schematic diagram of the working principle of the anionic-π supramolecular hydrogel prepared in Example 1.

[0038] Figure 5 Visualization of the formation mechanism of anionic-π supramolecular hydrogels demonstrated by molecular simulation calculations: in the absence of sodium ions (left) and in the presence of sodium ions (right).

[0039] Figure 6 Statistical graph of elasticity and strength of the gel obtained by mixing PSBMA and PSSNa at different repeating unit molar ratios in Example 4.

[0040] Figure 7This is a comparison chart of the degree of fibrosis in Masson trichrome immunohistochemistry sections of the injectable PEGDA and sodium alginate hydrogels in Application Example 1 and the PSBMA-PSSNa supramolecular hydrogel implanted in Example 1, one month after subcutaneous implantation in the back of mice.

[0041] Figure 8 This is a control chart showing the use of the PSBMA-PSSNa anion-π supramolecular hydrogel prepared in Application Example 2 to preserve the activity of basic fibroblast growth factor (bFGF) protein.

[0042] Figure 9 This is a comparison chart of the wound healing areas on days 0, 4, 7, and 14 after the bFGF-loaded SSH hydrogel, the bFGF-free SSH hydrogel, the commercial sodium alginate wound dressing, and the gauze in Application Example 3 were used for wound dressing in diabetic mice.

[0043] Figure 10 This is a digital comparison of the wound healing conditions on days 0, 4, 7, and 14 after the bFGF-loaded SSH hydrogel, the bFGF-free SSH hydrogel, the commercial sodium alginate wound dressing, and the gauze were used for wound dressing in diabetic mice in Application Example 3. DETAILED DESCRIPTION

[0044] In order to make the purpose, technical solutions and advantages of the present invention clearer, the present invention is further described in detail below in conjunction with embodiment. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the present invention. Those skilled in the art will make modifications or equivalent substitutions based on understanding the technical solution of the present invention, without departing from the spirit and scope of the technical solution of the present invention, and all should be encompassed within the protection scope of the present invention.

[0045] The raw materials used in the following specific embodiments are all purchased from the market.

[0046] Example 1

[0047] Step 1: 10% mass fraction of sulfobetaine monomer (SBMA) was dissolved in water, and then 1 wt% of sodium persulfate by weight of the reaction monomer was added, and nitrogen was bubbled for 20 minutes. Then, it was placed in a 70°C oil bath and stirred for 12 hours, and then dialyzed and freeze-dried to obtain PSBMA polymer powder with a molecular weight of 170,000; 10% mass fraction of sodium p-styrenesulfonate (SSNa) was dissolved in water, and then sodium persulfate was added at 1 wt% of the reaction monomer mass, and nitrogen was bubbled for 20 minutes. Then, it was placed in a 70°C oil bath and stirred for 12 hours, and then dialyzed and freeze-dried to obtain PSSNa polymer powder with a molecular weight of 200,000;

[0048] Step 2: 10% mass fraction of PSBMA was dissolved in PBS buffer, and 30% mass fraction of PSSNa was dissolved in aqueous solution. Then, 1 mL of PSBMA solution was taken and thoroughly mixed with 0.25 mL of PSSNa solution (the molar ratio of zwitterion to p-styrenesulfonic acid repeating units in the copolymer was 1:1). The resulting hydrogel was placed in an 80°C oven for equilibration for 2 h to obtain the PSBMA-PSSNa anion-π supramolecular hydrogel.

[0049] The anionic-π supramolecular hydrogel prepared in Example 1 was subjected to rheological characterization. All rheological tests were performed on a rotational rheometer (TA Instruments, Discovery HR-20) using a 20 mm diameter, 1500 μm gap plate fixture at 37°C. The sample was allowed to rest for 1 minute before testing, and then subjected to an oscillatory strain sweep (strain range: 0.1–2000%, fixed frequency: 10 rad / s) and an oscillatory frequency sweep (frequency range: 0.1–100 rad / s, fixed strain: 1%). The results are shown in Table 1. Figure 1 As shown, these curves show that the elastic and viscous properties of the material change with strain ( Figure 1 a) and angular frequency ( Figure 1 The elastic properties dominate, further indicating that the material exhibits solid-like behavior.

[0050] To confirm the gelation mechanism of the hydrogel, UV spectroscopy was used to analyze the PSSNa solution, PSBMA solution, and the mixed diluted solution of PSSNa and PSBMA. Figure 2 As shown in the figure, in the presence of PSBMA, the characteristic UV absorption peak of the benzene ring in the PSSNa molecule undergoes a red shift, indicating that its electron cloud distribution is affected. Based on this, it can be inferred that non-covalent interactions related to the aromatic ring occur in the system.

[0051] Further, such as Figure 3 As shown in the figure, the thermodynamic properties of PSSNa and PSBMA were characterized using chemical isothermal titration. The results showed that under dilution conditions, the binding process of PSBMA and PSSNa presented an equivalent ratio (1:1). At the same time, the titration results showed that the Gibbs free energy during the mixing process of the two was less than 0, and the entropy of the system increased significantly, proving that the process was a spontaneous process driven by entropy, providing a thermodynamic basis for the proposed theoretical model.

[0052] Based on the above experimental evidence, the following Figure 4The anion-π interaction model shown in Figure 2 shows two primary interactions between PSBMA and PSSNa. First, the sulfonate anions in the PSBMA molecules interact with the benzene rings on the PSSNa side chains and the cations in the system through electron cloud interactions, forming anion-π-cation interactions (hereafter referred to as anion-π interactions). Second, ionic interactions occur between the quaternary ammonium cations in PSBMA and the sulfonate anions of PSSNa. The synergistic effect of these two interactions leads to a tightly interlocked structure between the side chains of PSBMA and PSSNa, significantly influencing the microstructure and macroscopic properties of the hybrid system.

[0053] Figure 5 The molecular simulation results shown further confirm the rationality of the anion-π interaction model at the molecular level. Visual analysis shows that in the presence of cations, a non-covalent interaction (anion-π interaction) exists between the sulfonate group and the benzene ring, while in the absence of cations, this interaction completely disappears.

[0054] Example 2

[0055] Step 1: 10% mass fraction of carboxybetaine monomer (CBAA) was dissolved in water, and then 1 wt% of sodium persulfate was added to the reaction monomer mass, and nitrogen was bubbled for 20 minutes, and then it was placed in a 70°C oil bath and stirred for 12 hours, and then dialyzed and freeze-dried to obtain PCBAA polymer powder; 10% mass fraction of sodium p-styrenesulfonate (SSNa) was dissolved in water, and then added, and then 1 wt% of sodium persulfate was added to the reaction monomer mass, and nitrogen was bubbled for 20 minutes, and then it was placed in a 70°C oil bath and stirred for 12 hours, and then dialyzed and freeze-dried to obtain PSSNa polymer powder;

[0056] Step 2: 10% by mass of PCBMA was dissolved in PBS buffer, and 30% by mass of PSSNa was dissolved in aqueous solution. Subsequently, 1 mL of PCBAA solution was thoroughly mixed with 0.25 mL of PSSNa solution (the molar ratio of zwitterions to p-styrenesulfonic acid repeating units in the copolymer was 1.2:1). The resulting hydrogel was placed in an 80°C oven for equilibrium for 2 h to obtain the CBAA-PSSNa anion-π supramolecular hydrogel.

[0057] Example 3

[0058] Step 1: 8% mass fraction of sulfobetaine monomer (SBMA) and 2% mass fraction of acrylamide (AAM) were dissolved in water, and then 1 wt% of sodium persulfate (based on the mass of the reaction monomer) was added and nitrogen was bubbled for 20 minutes. Then, it was placed in a 70°C oil bath and stirred for 12 hours, then dialyzed and freeze-dried to obtain PSBMA-AAM polymer powder with a molecular weight of 210,000; 10% mass fraction of sodium p-styrenesulfonate (SSNa) was dissolved in water, and then added, and then 1 wt% of sodium persulfate (based on the mass of the reaction monomer) was added and nitrogen was bubbled for 20 minutes. Then, it was placed in a 70°C oil bath and stirred for 12 hours, then dialyzed and freeze-dried to obtain PSSNa polymer powder with a molecular weight of 200,000;

[0059] Step 2: 10% by mass of PCBMA was dissolved in PBS buffer, and 30% by mass of PSSNa was dissolved in aqueous solution. Subsequently, 1 mL of PCBAA solution was thoroughly mixed with 0.25 mL of PSSNa solution. The resulting hydrogel was placed in an 80°C oven for equilibration for 2 h to obtain the PSBMA-AAM-PSSNa anion-π supramolecular hydrogel.

[0060] Example 4

[0061] According to the preparation process of Example 1, the mixing ratio of the PSBMA solution and the PSSNa solution in step 2 was changed to make the molar ratio of the p-styrenesulfonic acid and zwitterionic repeating units in PSSNa and PSBMA 1:4, 1:2, 2:1, and 3:1 to obtain different hydrogels. The elasticity and strength test results of the gels obtained by mixing polysulfobetaine (PSBMA) with a molecular weight of 170,000 and sodium poly(p-styrenesulfonate) with a molecular weight of 200,000 at different repeating unit molar ratios are shown as follows: Figure 6 As shown, it can be seen that the ratio of the two has an impact on the mechanical properties of the hydrogel. When the ratio of the repeating units of zwitterion and styrenesulfonic acid is in the range of 1:0.5-2, the hydrogel has better mechanical strength and elasticity.

[0062] Application Example 1: Fibrotic response of representative anionic-π injectable hydrogels in vivo

[0063] 1. Animal surgery process

[0064] This application example describes the fibrotic response of a PSBMA-PSSNa anionic-π supramolecular hydrogel in a mouse subcutaneous implantation model. The PSBMA-PSSNa hydrogel (SSH) prepared in Example 1, along with PEGDA and sodium alginate hydrogel (Alg), was injected subcutaneously into the backs of C57BL / 6 female mice. 100 μL of the solution was injected into each mouse, and the fibrotic response was evaluated on day 28.

[0065] 2. H&E and Masson trichrome staining immunohistochemical sections

[0066] Mice were sacrificed at week 4, and the implants and surrounding tissues were excised, fixed with 10% formaldehyde, embedded in paraffin, and sectioned for hematoxylin and eosin (H&E) / Masson's trichrome staining and immunohistochemical staining. Images were acquired using a Nikon intensilight CHGFI microscope equipped with NIS-Elements AR software and an Olympus virtual slide microscope (VS120-S6-W). Collagen density was measured by calculating the percentage coverage of blue pixels at 50 μm intervals in Masson's trichrome-stained tissue images.

[0067] like Figure 7 As shown in the images, it can be clearly seen that a dense collagen capsule was formed in the tissue around the PEG hydrogel and the alginate hydrogel. Notably, almost no fibrotic reaction was observed around the SSH hydrogel, indicating its excellent ability to resist foreign body reaction.

[0068] Application Example 2: Representative anion-π injectable hydrogels for protein encapsulation and maintenance of protein activity

[0069] 1. Experimental Procedure

[0070] This experiment evaluated the ability of anionic-π injectable hydrogels to preserve protein activity. First, 50 μL of a 10% solution of PSBMA (molecular weight approximately 1,210,000) was thoroughly mixed with 150 μL of a PBS solution containing 2.5 mg / mL basic fibroblast growth factor (bFGF). Next, the resulting solution was mixed with a 3.75% solution of PSSNa (molecular weight approximately 200,000) at a volume ratio of 4:1. After gentle stirring, a gel was formed, thereby preparing a bFGF-loaded gel (bFGF@SSH).

[0071] Subsequently, the bFGF@SSH gel and free bFGF solution were treated separately: one portion was incubated at 56°C for 30 minutes, while the other portion was stored at 37°C for 3 days. This resulted in four experimental sample groups: bFGF@SSH (heated), bFGF (heated), bFGF@SSH (stored), and bFGF (stored). Before the experiment began, fresh bFGF@SSH and free bFGF solutions were prepared, along with a protein-free SSH gel prepared according to the above procedure as a control. 4 μL of each sample was added to the wells seeded with cells, and the plates were incubated under appropriate conditions for 24 hours. After incubation, cell viability was determined using the MTT assay. A PBS solution group was used as a blank control for subsequent data comparison and analysis.

[0072] 2. Results Analysis

[0073] The results of the cell proliferation experiment were as follows Figure 8 As shown in the figure. After treatment, free bFGF essentially lost its activity. In contrast, bFGF@SSH retained approximately 80% of its activity after heat treatment and approximately 60% after storage at 37°C compared to a fresh bFGF solution. Furthermore, there was no significant difference in the efficiency of promoting cell proliferation between fresh free bFGF solution and fresh bFGF@SSH solution. These results demonstrate that encapsulating bFGF in SSH does not result in loss of its activity. Furthermore, because SSH has a stabilizing effect on proteins, it can effectively maintain protein activity in the face of environmental challenges.

[0074] Application Example 3

[0075] Basic fibroblast growth factor (bFGF)-loaded SSH hydrogels for promoting wound healing

[0076] 1. Animal Surgical Procedure: To induce a diabetic model, male C57BL / 6 mice (weighing approximately 20 g) were intraperitoneally injected with 150 mg / kg of streptozotocin (STZ) after a 16-hour fast. Successful diabetic modeling was considered achieved when blood glucose levels reached ≥13.88 mmol / L (250 mg / dL). Following successful modeling, mice exhibited typical diabetic symptoms, including polydipsia, polyphagia, polyuria, and significant weight loss. Before wound modeling, mice were anesthetized, shaved, and disinfected. Full-thickness skin was then excised from the back and buttocks to create standardized wounds. Mice were randomly divided into four groups: a control group (medical gauze), an alginate dressing group (AD), a bFGF@SSH hydrogel application group (prepared in Application Example 1), and an SSH hydrogel application group. Wounds were photographed on days 0, 2, 4, 7, and 14 to assess changes in wound area. On day 14, wound tissue was collected for histological analysis, including hematoxylin and eosin (H&E) staining, Masson's trichrome staining, and immunofluorescence staining. Dermal and epidermal thickness, wound healing area, and collagen volume fraction were quantitatively analyzed using ImageJ and Adobe Photoshop 2025.

[0077] 2. Results Analysis

[0078] The results showed that the wounds treated with bFGF@SSH healed significantly faster than those in all other groups. Figure 9 On day 4, the wound closure rate in the bFGF@SSH group was approximately 55%, while that in the medical gauze group and the commercial dressing group was only 25% and 35%, respectively. It is noteworthy that the healing effect of the SSH hydrogel group alone was comparable to that of the commercial sodium alginate dressing. By day 14, the wounds in the bFGF@SSH group were almost completely healed, while the wounds in the other groups still had significant unclosed areas ( Figure 10 ). This demonstrates the potential application of bFGF-loaded SSH hydrogel in promoting diabetic wound healing models.

Claims

1. A zwitterionic supramolecular hydrogel based on anion-π, characterized in that It is self-assembled from a zwitterionic copolymer and a p-styrene sulfonate copolymer; The zwitterionic copolymer is obtained by copolymerizing raw materials including zwitterionic monomers, and the p-styrenesulfonate copolymer is obtained by copolymerizing monomers containing p-styrenesulfonic acid; The zwitterionic monomer is selected from any one or more of sulfonate betaine methacrylate, carboxylate betaine methacrylate, and methacryloyloxyethyl phosphorylcholine; The molar ratio of the zwitterion in the zwitterionic copolymer to the p-styrenesulfonic acid repeating unit in the p-styrenesulfonate copolymer is 1:0.1-10.

2. The zwitterionic supramolecular hydrogel based on anion-π according to claim 1, characterized in that The p-styrenesulfonic acid-containing monomer includes one or more of p-styrenesulfonic acid, sodium p-styrenesulfonate, potassium p-styrenesulfonate, and lithium p-styrenesulfonate.

3. The zwitterionic supramolecular hydrogel based on anion-π according to claim 1, characterized in that The weight average molecular weight of the zwitterionic copolymer is 50,000-5,000,000; the weight average molecular weight of the p-styrene sulfonate copolymer is 50,000-5,000,000.

4. The zwitterionic supramolecular hydrogel based on anion-π according to claim 1, characterized in that The polymerization monomer raw material of the zwitterionic copolymer also includes a first comonomer; the first comonomer includes one or more monomers of acrylates and acrylamides; the mass proportion of the zwitterionic monomer in the polymerization monomer raw material of the zwitterionic copolymer is more than 50%.

5. The anion-π based zwitterionic supramolecular hydrogel according to claim 1, characterized in that The polymerization monomer raw material of the styrene sulfonate copolymer further includes a second comonomer; the second comonomer includes one or more of acrylic acid, methacrylic acid, maleic anhydride, styrene or its derivatives, acrylates, methacrylates, and acrylamides; The weight percentage of monomers containing p-styrenesulfonic acid in the polymerization monomer raw materials of the p-styrenesulfonate copolymer is more than 50%.

6. A method for preparing an anion-π based zwitterionic supramolecular hydrogel according to any one of claims 1 to 5, characterized in that: Including steps: Step 1: copolymerizing a raw material containing a zwitterionic monomer to obtain a zwitterionic copolymer; copolymerizing a raw material containing a monomer containing p-styrenesulfonic acid to obtain a p-styrenesulfonate copolymer; Step 2: dissolving the zwitterionic copolymer and the p-styrenesulfonate copolymer in an aqueous solution respectively, mixing them and self-assembling them to obtain the zwitterionic supramolecular hydrogel.

7. The method for preparing anion-π based zwitterionic supramolecular hydrogel according to claim 6, characterized in that: In step 2, the total concentration of the zwitterionic copolymer and the p-styrene sulfonate copolymer in the aqueous solution is 1 to 60 wt %.

8. The method for preparing anion-π based zwitterionic supramolecular hydrogel according to claim 6, characterized in that: The copolymerization in step 1 contains an initiator, the copolymerization temperature is 60-90° C., the reaction is carried out for 6-18 hours, and the product is dialyzed and freeze-dried to obtain a copolymer.

9. Use of the anion-π based zwitterionic supramolecular hydrogel according to any one of claims 1 to 5 in the preparation of immunocompatible drugs or medical materials.

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