Hydrogel as well as preparation method and application thereof

Abstract

The invention relates to a hydrogel. The hydrogel comprises caffeic acid functionalized gelatin, polylysine and a drug-loaded liposome. The hydrogel can respond to an oxidative stress environment, accurately control drug release, prolong drug action time and provide enough physical strength and tissue adhesion.

Description

Technical Field

[0001] This invention relates to the field of hydrogel technology, and in particular to a hydrogel, its preparation method, and its applications. Background Technology

[0002] Traumatic bleeding after nephrectomy is a common and critical complication in renal surgery, which not only increases blood loss but also affects postoperative recovery. While mechanical suturing, cauterization, or local hemostatic materials can control bleeding to some extent in clinical management, these methods often fail to simultaneously achieve hemostasis and tissue repair, and may still lead to further tissue damage postoperatively.

[0003] More seriously, during surgeries such as partial nephrectomy, to reduce bleeding and facilitate the procedure, it is often necessary to temporarily occlude the renal artery or even the renal vein, leading to local or complete ischemia in the renal tissue. Although blood supply can be restored after the occlusion is lifted and blood is re-perfused, it triggers a series of pathological processes such as oxidative stress, inflammatory response, apoptosis, and ferroptosis, thereby causing renal ischemia-reperfusion injury (IRI), further aggravating nephron damage and declining renal function.

[0004] Current treatments for ischemia-reperfusion injury (IRI) primarily rely on antioxidants, anti-inflammatory drugs, and drug delivery systems. However, these methods typically only alleviate some symptoms and fail to significantly reduce the damage caused by ischemia-reperfusion. Even when drugs can reach the kidneys via delivery systems, issues such as inaccurate release, low solubility, and low bioavailability persist, hindering optimal efficacy at appropriate times and in suitable environments. Furthermore, while local administration can increase drug concentration to some extent, existing systems lack responsiveness to oxidative stress environments, resulting in insufficient targeted release and limited efficacy. Simultaneously, existing partially injectable hydrogels exhibit weak adhesion at surgical sites, insufficient mechanical properties and strength, making them ill-suited to the complex physiological environment of nephrectomy and ischemia-reperfusion procedures.

[0005] The shortcomings of existing technologies are mainly reflected in several aspects: First, the drug solubility is poor, especially water-insoluble drugs, which are difficult to effectively deliver to the target area, making it impossible to achieve rapid and precise drug release; second, existing drug delivery systems lack a response to oxidative stress environments, resulting in drugs not being released at the most appropriate time and failing to fully exert their therapeutic effects; third, while ensuring drug efficacy, local delivery systems often fail to take into account sufficient physical strength and tissue adhesion, affecting hemostasis and repair effects; finally, existing treatment methods often cannot simultaneously meet the needs of both hemostasis and treatment, thus failing to effectively reduce kidney damage caused by IRI and promote tissue repair.

[0006] Pemafibrate, a PPARα agonist, is a drug with significant anti-inflammatory, antioxidant, and renal protective effects. It can effectively reduce oxidative stress, inhibit inflammatory responses, and improve renal function recovery. However, because pemafibrate is water-insoluble, its bioavailability is low, and its duration of action in vivo is short, making it difficult to maintain effective drug concentrations in areas of renal injury, thus limiting its therapeutic efficacy. Furthermore, pemafibrate's delivery capacity through the kidneys is limited, especially during renal ischemia-reperfusion, where the drug cannot effectively target the damaged area, resulting in unsatisfactory therapeutic effects. To improve drug delivery, current research often uses various carrier materials. However, while traditional liposome systems can improve drug solubility and bioavailability, they lack the ability to respond to specific physiological conditions (such as oxidative stress), making it impossible to precisely control the timing and rate of drug release.

[0007] Furthermore, while natural polymers such as chitosan and polylysine possess good biocompatibility and biodegradability, their insufficient physical strength and tissue adhesion in the treatment of kidney injury affect their stability and efficacy at the surgical site. Current drug delivery systems still struggle to simultaneously achieve precise drug release, prolonged duration of action, and effective targeting, resulting in suboptimal therapeutic outcomes. Therefore, developing a multifunctional therapeutic system capable of responding to oxidative stress, precisely controlling drug release, prolonging drug action time, and providing sufficient physical strength and tissue adhesion remains a significant technological challenge. Summary of the Invention

[0008] To address the shortcomings of existing technologies, this invention provides a hydrogel comprising caffeic acid-functionalized gelatin, polylysine, and drug-loaded liposomes.

[0009] In one embodiment, the raw materials for preparing the caffeic acid functionalized gelatin include gelatin, adipic acid dihydrazide, and caffeic acid.

[0010] In one embodiment, the liposome preparation raw materials of the drug-loaded liposomes include soybean lecithin, cholesterol, and nanocomposite materials. The nanocomposite materials are mainly composed of distearate phosphatidylethanolamine (DSPE), ketethiocyanate (TK), and polyethylene glycol (PEG). The drug includes a PPARα agonist.

[0011] In one embodiment, the PPARα agonist includes Pemafibrat.

[0012] In one embodiment, the molecular weight of the nanocomposite material is 2000.

[0013] A second aspect of the present invention also provides a method for preparing the above-mentioned hydrogel, comprising the following steps:

[0014] Preparation of caffeic acid functionalized gelatin (Gel-ADH-CA): Gelatin (Gel) and adipic acid dihydrazide (ADH) were dissolved, mixed, reacted, purified by dialysis, and freeze-dried to obtain gelatin-adipic acid dihydrazide (Gel-ADH). Gelatin-adipic acid dihydrazide was dissolved, activated, and caffeic acid (CA) solution was added to react, purified by dialysis, and freeze-dried to obtain caffeic acid functionalized gelatin. Preparation of drug-loaded liposomes (Lip / DSPE@P): Soybean lecithin, cholesterol and nanocomposite material were dissolved, drugs were added, and lipid membranes were obtained by rotary evaporation. Solvents were added to hydrate the lipid membranes, sonicated on ice, filtered to obtain liposome emulsion suspensions, and free drugs were removed by centrifugation. Preparation of hydrogel: Dissolve caffeic acid-functionalized gelatin, add polylysine (ε-PL) and drug-loaded liposome solution, stir and mix evenly in an ice bath, and let stand to obtain hydrogel.

[0015] In one embodiment, the preparation of the caffeic acid-functionalized gelatin includes the following steps: dissolving gelatin in water, dissolving adipic acid dihydrazide in buffer solution, mixing the two and adding 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) for reaction, purifying by dialysis using a dialysis bag after the reaction, and lyophilizing to obtain gelatin-dipic acid dihydrazide; dissolving gelatin-dipic acid dihydrazide in buffer solution, adding 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide for activation in the dark, adding DMSO solution of caffeic acid for reaction, purifying by dialysis using a dialysis bag after the reaction, and lyophilizing to obtain caffeic acid-functionalized gelatin.

[0016] In one embodiment, the gelatin is dissolved in water at a temperature of 48-52°C.

[0017] In one embodiment, the buffer solution comprises 2-morpholinoethanesulfonic acid (MES).

[0018] In one embodiment, the pH of the buffer solution is 5 to 6.

[0019] In one embodiment, the temperature of the reaction involving the addition of EDC and NHS is 40-50°C, and the reaction time is 3-5 hours.

[0020] In one embodiment, the dialysis purification using a dialysis bag and the freeze-drying of the gelatin-oxohydrazide obtained by the dialysis bag have a molecular weight cutoff of 7-9 kDa, and the dialysis time is 46-50 h.

[0021] In one embodiment, the light-shielded activation temperature is 23~28°C, and the time is 10~20 min.

[0022] In one embodiment, the reaction time of the DMSO solution with added caffeic acid is 3-5 hours.

[0023] In one embodiment, the dialysis bag used for purification and freeze-drying yields caffeic acid functionalized gelatin with a molecular weight cutoff of 7-9 kDa. The dialysis time is 10-38 h, and the dialysis includes first dialysis with a mixed solution of ethanol and water, followed by dialysis with water.

[0024] In one embodiment, the preparation of the drug-loaded liposomes includes the following steps: dissolving soybean lecithin, cholesterol, and nanocomposite materials in an organic solvent, adding a drug solution, rotary evaporating the mixture to obtain a lipid membrane, adding phosphate buffer solution (PBS) to hydrate the lipid membrane, sonicating on ice, filtering to obtain a liposome emulsion suspension, and centrifuging to remove free drugs.

[0025] In one embodiment, the rotary evaporation uses a rotary evaporator.

[0026] In one embodiment, the PBS contains Tween-80 and methylcellulose.

[0027] In one embodiment, the ultrasound time on ice is 8-12 minutes.

[0028] In one embodiment, the filtration membrane has a micropore size of 0.4~0.5μm.

[0029] In one embodiment, the centrifugation time for removing free drug is 25-35 minutes.

[0030] In one embodiment, the method for preparing the drug-loaded liposomes includes membrane hydration, and the organic solvent includes dichloromethane (CH3Cl2).

[0031] In one embodiment, the preparation of the hydrogel includes the following steps: dissolving caffeic acid-functionalized gelatin in a phosphate buffer solution, adding a phosphate buffer solution containing polylysine and drug-loaded liposomes dropwise to the caffeic acid-functionalized gelatin solution, stirring and mixing evenly in an ice bath, and allowing it to stand to obtain the hydrogel.

[0032] In one embodiment, the pH of the phosphate buffer solution in which the caffeine-functionalized gelatin is dissolved is 7-8.

[0033] In one embodiment, the ice bath stirring time is 3 to 8 minutes.

[0034] In one embodiment, the settling temperature is 36~38°C.

[0035] In one embodiment, in the preparation of the caffeic acid functionalized gelatin, the mass ratio of gelatin to adipate dihydrazide is (6~10):1; and the mass ratio of gelatin-adipate dihydrazide to caffeic acid is 1:(0.1~0.5). In the preparation of the drug-loaded liposomes, the mass ratio of soybean lecithin:cholesterol:nanocomposite material is (10~30):(5~15):(0.5~2). In the preparation of the hydrogel, the mass ratio of caffeic acid functionalized gelatin: polylysine: drug-loaded liposomes is (150~450): (150~250): (0.1~2).

[0036] A third aspect of the present invention also provides the use of the above-described hydrogel or the hydrogel obtained by the above-described preparation method in the preparation of products for the treatment of kidney injury.

[0037] Compared with the prior art, the present invention has the following beneficial effects: 1. Dual "targeting" mechanism for precise response (IRI): Existing technologies mostly use non-specific antioxidants or single-target drugs, which have limited efficacy. This invention innovatively combines ROS-responsive liposomes with the PPARα agonist Pemafibrate, using the local oxidative stress environment during IRI as a "bioswitch" to achieve precise drug release at the site of injury, avoiding the side effects of systemic administration. At the same time, it regulates inflammation, oxidative stress and apoptosis in multiple dimensions through the PPARα pathway, significantly improving efficacy. 2. Integrated design for synergistic hemostasis and damage prevention: Existing hemostatic materials (such as gelatin sponge and fibrin glue) only have physical sealing functions and cannot intervene in IRI. The caffeic acid modified gelatin / polylysine injectable gel designed in this invention achieves dual functions through the following innovations: (1) Enhanced dynamic cross-linking: Catechol groups interact with Na+ in the blood. + and Fe with increased IRI 3+ It forms ion chelates, and the gel strength adaptively increases with the bleeding environment, resulting in a hemostatic effect superior to static cross-linked materials; (2) Tissue adhesion: Catechol groups form covalent bonds with amino / thiol groups on the tissue surface, avoiding the shedding of traditional materials due to blood flushing; (3) Drug sustained release platform: The gel acts as a local reservoir for liposomes, prolonging the retention time of Pemafibrate at the site of injury and achieving a continuous effect of "hemostasis-anti-inflammation-repair". 3. Excellent biocompatibility and non-toxic degradation products: Traditional hemostatic materials (such as cyanoacrylates) may cause foreign body reactions or degradation toxicity. The gelatin, polylysine, and caffeic acid used in this invention are all FDA-approved safe materials, and their degradation products can be metabolized by renal tubular cells, making them especially suitable for patients with impaired kidney function and avoiding increased burden on the kidneys; 4. Strong clinical applicability and simplified surgical procedure: Existing IRI prevention and treatment strategies require additional intraoperative drug infusion or postoperative systemic drug administration, which is cumbersome. This invention uses an integrated "injectable gel-liposome" formulation to complete hemostasis and drug delivery in one step during surgery without changing the existing partial nephrectomy procedure, significantly reducing the technical threshold and time cost. Attached Figure Description

[0038] Figure 1 SEM images of the Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel; Figure 2 This is the standard release curve for Pemafibrat. Detailed Implementation

[0039] During partial nephrectomy, the renal artery (and sometimes the renal vein) is usually temporarily blocked, causing local or complete ischemia of the kidney. After the blockage is removed, blood is re-perfused into the kidney, resulting in ischemia-reperfusion (I / R). Although this reperfusion restores blood flow, it triggers oxidative stress, inflammatory responses, apoptosis, and ferroptosis, damaging the nephrons (tubules and glomeruli) and leading to renal ischemia-reperfusion injury (IRI), one of the main mechanisms of kidney function impairment.

[0040] To achieve rapid hemostasis and reduce kidney damage caused by intrarenal infarction (IRI) after partial nephrectomy, this invention utilizes ROS-responsive liposomes to deliver the PPARα agonist pemafibrate (a water-insoluble drug). Liposomes enhance the solubility of pemafibrate and rapidly release it in response to the oxidative stress environment induced by IRI. Simultaneously, a weakly ionic cross-linked injectable gel formed by caffeic acid-modified gelatin and polylysine through electrostatic and hydrogen bonding interactions is injected into the post-resection cavity. The catechol groups chelate with sodium ions in the blood and iron ions elevated by the oxidative stress environment, enhancing the strength of the hydrogel and ensuring strong adhesion to the tissue.

[0041] To facilitate understanding of the present invention, a more complete description will be given below with reference to the accompanying drawings. Preferred embodiments of the invention are shown in the drawings. However, the invention can be implemented in many different forms and is not limited to the embodiments described herein. Rather, these embodiments are provided to provide a thorough and complete understanding of the disclosure of the invention.

[0042] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. The terminology used herein in the description of the invention is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. The term "and / or" as used herein includes any and all combinations of one or more of the associated listed items.

[0043] Unless otherwise specified, all reagents, materials, and equipment used in this embodiment are commercially available; unless otherwise specified, all test methods are conventional test methods in this field.

[0044] Example I. This invention provides a method for preparing a hydrogel, comprising the following steps: 1. Preparation of Gelatin-Oxydihydrazide (Gel-ADH) Dissolve 2.0 g of gelatin in 100 mL of deionized water at 50 °C. Separately, dissolve 0.25 g of adipic dihydrazide (ADH) in 20 mL of LME S buffer (0.1 M, pH = 5.5). Then mix the two solutions and add 0.4 g of EDC and 0.3 g of NHS. React at 45 °C for 4 h. After the reaction is complete, dialyze the solution against deionized water for 48 h using a dialysis bag with a molecular weight cutoff of 8 kDa (changing the water every 8 h). Freeze-dry to obtain white flocculent gel-ADH.

[0045] In this embodiment, the mass ratio of gelatin to adipic dihydrazide is 8:1.

[0046] 2. Preparation of coffee acid-functionalized gelatin (Gel-ADH-CA) 0.5 g of Gel-ADH was redissolved in 50 mL of MES buffer (0.1 M, pH 5.5), and 0.25 g of EDC and 0.20 g of NHS were added. The mixture was activated at 25 °C in the dark for 15 min. Then, 10 mL of DMSO solution containing 0.15 g of caffeic acid was added dropwise, and the reaction was continued for 4 h. After the reaction was completed, the reaction solution was transferred to an 8 kDa dialysis bag and dialyzed with 20% ethanol-water for 12 h, followed by dialyzed with pure water for 24 h. Finally, the solution was lyophilized to obtain brownish-yellow porous sponge-like Gel-ADH-CA.

[0047] In this embodiment, the mass ratio of gelatin-oxaloacetic acid dihydrazide to caffeic acid is 1:0.3.

[0048] 3. Preparation of drug-loaded liposomes (Lip / DSPE@P) Liposomes were prepared using a membrane hydration method: 200 mg of soybean lecithin, 100 mg of cholesterol, and 10 mg of DSPE-TK-PEG2000 were dissolved in 5 mL of CH3Cl2. A 1 mg / mL solution of Pemafibrat was added to the mixture, and the solution was transferred to a round-bottom flask. The organic solvent was evaporated using a rotary evaporator, forming a lipid membrane at the bottom of the flask. Next, 3 mL of PBS solution containing 2% Tween-80 and 0.5% methylcellulose was added to the flask to hydrate the lipid membrane. The mixture was sonicated on ice for 10 min and filtered through a 0.45 μm microporous membrane to obtain a nanoliposome emulsion suspension. Centrifugation at 13000 rpm for 30 min effectively removed free Pemafibrat.

[0049] In this embodiment, the mass ratio of soybean lecithin:cholesterol:nanocomposite material is 20:10:1.

[0050] 4. Preparation of Gel-ADH-CA / ε-PL hydrogels loaded with Lip / DSPE@P Dissolve 2g of Gel-ADH-CA in 5mL of PBS (pH=7.4). Then, under continuous magnetic stirring, slowly add 5mL of PBS solution containing 1.0g of ε-PL and 1mg / mL of Lip / DSPE@P suspension to the above Gel-ADH-CA solution dropwise. After the addition is complete, stir in an ice bath for 5min to ensure uniform mixing. Finally, transfer the mixture to 37°C and let it stand. The system can spontaneously form a hydrogel loaded with liposomes Lip / DSPE@P through electrostatic interactions and hydrogen bonds.

[0051] In this embodiment, the mass ratio of caffeic acid-functionalized gelatin: polylysine: drug-loaded liposomes is 400:200:1.

[0052] The material ratios for preparing hydrogels in the embodiments and comparative examples of this invention are shown in Table 1: Table 1 Material composition of different embodiments and comparative examples

[0053] Example 1 Preparation of Gel-ADH-CA: 2.0 g gelatin, 0.25 g ADH, 0.4 g EDC and 0.3 g NHS were dissolved in 100 mL of deionized water at 50 °C and reacted at 45 °C for 4 h. After the reaction was completed, the mixture was dialyzed and lyophilized to obtain white flocculent Gel-ADH. 0.5 g Gel-ADH was redissolved, and 0.25 g EDC and 0.20 g NHS were added. Then 10 mL of a solution containing 0.15 g CA was added dropwise, and the reaction was continued for 4 h. After the reaction was completed, the mixture was dialyzed. First, it was dialyzed with 20% ethanol-water for 12 h, and then dialyzed with pure water for 24 h. Finally, it was lyophilized to obtain brownish-yellow porous sponge-like Gel-ADH-CA.

[0054] Preparation of Lip / DSPE@P liposomes: 0.2g soybean lecithin, 0.1g cholesterol and 0.01g DSPE-TK-PEG2000 were dissolved in 5mL CH3Cl2, and then 1mg / mL Pemafibrat solution was added. The lipid membrane was obtained by rotary evaporation. Then, 3mL of PBS solution containing 2% Tween-80 and 0.5% methylcellulose was added dropwise. After sonication, the mixture was filtered through a 0.45μm microporous membrane to obtain a nanoliposome emulsion suspension. The suspension was centrifuged at 13000rpm for 30min to obtain Lip / DSPE@P.

[0055] Preparation of Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel: Dissolve 2.0g Gel-ADH-CA in 5mL PBS, then add dropwise 5mL PBS solution containing 1.0g ε-PL and 1mg / mL Lip / DSPE@P suspension. React the mixture at room temperature for 5h to obtain Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel.

[0056] Example 2 Preparation of Gel-ADH: Dissolve 2.0g gelatin, 0.25g ADH, 0.4g EDC and 0.3g NHS in 100mL of 50℃ deionized water, react at 45℃ for 4h, dialyze after the reaction is completed, and freeze dry to obtain white flocculent Gel-ADH.

[0057] Preparation of Lip / DSPE@P liposomes: 0.2g soybean lecithin, 0.1g cholesterol and 0.01g DSPE-TK-PEG2000 were dissolved in 5mL CH3Cl2, and then 1mg / mL Pemafibrat solution was added. The lipid membrane was obtained by rotary evaporation. Then, 3mL of PBS solution containing 2% Tween-80 and 0.5% methylcellulose was added dropwise. After sonication, the mixture was filtered through a 0.45μm microporous membrane to obtain a nanoliposome emulsion suspension. The suspension was centrifuged at 13000rpm for 30min to obtain Lip / DSPE@P.

[0058] Preparation of Gel-ADH / ε-PL / Lip / DSPE@P hydrogel: 2.0 g Gel-ADH-CA was dissolved in 5 mL PBS, and then 5 mL PBS solution containing 1.0 g ε-PL and 1 mg / mL Lip / DSPE@P suspension was added dropwise. The mixture was reacted at room temperature for 5 h to obtain Gel-ADH / ε-PL / Lip / DSPE@P hydrogel.

[0059] Example 3 Preparation of Gel-ADH-CA: 2.0 g gelatin, 0.25 g ADH, 0.4 g EDC and 0.3 g NHS were dissolved in 100 mL of deionized water at 50 °C and reacted at 45 °C for 4 h. After the reaction was completed, the mixture was dialyzed and lyophilized to obtain white flocculent Gel-ADH. 0.5 g Gel-ADH was redissolved, and 0.25 g EDC and 0.20 g NHS were added. Then 10 mL of a solution containing 0.10 g CA was added dropwise, and the reaction was continued for 4 h. After the reaction was completed, the mixture was dialyzed. First, it was dialyzed with 20% ethanol-water for 12 h, and then dialyzed with pure water for 24 h. Finally, it was lyophilized to obtain brownish-yellow porous sponge-like Gel-ADH-CA.

[0060] Preparation of Lip / DSPE@P liposomes: 0.2 g soybean lecithin, 0.1 mg cholesterol and 0.01 g DSPE-TK-PEG2000 were dissolved in 5 mL CH3Cl2, and then 1 mg / mL Pemafibrat solution was added. The lipid membrane was obtained by rotary evaporation. Then, 3 mL of PBS solution containing 2% Tween-80 and 0.5% methylcellulose was added dropwise. After sonication, the mixture was filtered through a 0.45 μm microporous membrane to obtain a nanoliposome emulsion suspension. The suspension was centrifuged at 13000 rpm for 30 min to obtain Lip / DSPE@P.

[0061] Preparation of Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel: 1.0 g Gel-ADH-CA was dissolved in 5 mL PBS, and then 5 mL PBS solution containing 1.0 g ε-PL and 1 mg / mL Lip / DSPE@P suspension was added dropwise. The mixture was reacted at room temperature for 5 h to obtain Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel.

[0062] Example 4 Preparation of Gel-ADH-CA: 2.0 g gelatin, 0.25 g ADH, 0.4 g EDC and 0.3 g NHS were dissolved in 100 mL of deionized water at 50 °C and reacted at 45 °C for 4 h. After the reaction was completed, the mixture was dialyzed and lyophilized to obtain white flocculent Gel-ADH. 0.5 g Gel-ADH was redissolved, and 0.25 g EDC and 0.20 g NHS were added. Then 10 mL of a solution containing 0.15 g CA was added dropwise, and the reaction was continued for 4 h. After the reaction was completed, the mixture was dialyzed. First, it was dialyzed with 20% ethanol-water for 12 h, and then dialyzed with pure water for 24 h. Finally, it was lyophilized to obtain brownish-yellow porous sponge-like Gel-ADH-CA.

[0063] Preparation of Lip / DSPE@P liposomes: 0.2g soybean lecithin, 0.1g cholesterol and 0.01g DSPE-TK-PEG2000 were dissolved in 5mL CH3Cl2, and then 1mg / mL Pemafibrat solution was added. The lipid membrane was obtained by rotary evaporation. Then, 3mL of PBS solution containing 2% Tween-80 and 0.5% methylcellulose was added dropwise. After sonication, the mixture was filtered through a 0.45μm microporous membrane to obtain a nanoliposome emulsion suspension. The suspension was centrifuged at 13000rpm for 30min to obtain Lip / DSPE@P.

[0064] Preparation of Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel: 1.0 g Gel-ADH-CA was dissolved in 5 mL PBS, and then 5 mL PBS solution containing 0.5 g ε-PL and 1 mg / mL Lip / DSPE@P suspension was added dropwise. The mixture was reacted at room temperature for 5 h to obtain Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel.

[0065] Example 5 Preparation of Gel-ADH-CA: 2.0 g gelatin, 0.25 g ADH, 0.4 g EDC and 0.3 g NHS were dissolved in 100 mL of deionized water at 50 °C and reacted at 45 °C for 4 h. After the reaction was completed, the mixture was dialyzed and lyophilized to obtain white flocculent Gel-ADH. 0.5 g Gel-ADH was redissolved, and 0.25 g EDC and 0.20 g NHS were added. Then 10 mL of a solution containing 0.15 g CA was added dropwise, and the reaction was continued for 4 h. After the reaction was completed, the mixture was dialyzed. First, it was dialyzed with 20% ethanol-water for 12 h, and then dialyzed with pure water for 24 h. Finally, it was lyophilized to obtain brownish-yellow porous sponge-like Gel-ADH-CA.

[0066] Preparation of Lip / DSPE@P liposomes: 0.2g soybean lecithin, 0.1g cholesterol and 0.01g DSPE-TK-PEG2000 were dissolved in 5mL CH3Cl2, and then 1mg / mL Pemafibrat solution was added. The lipid membrane was obtained by rotary evaporation. Then, 3mL of PBS solution containing 2% Tween-80 and 0.5% methylcellulose was added dropwise. After sonication, the mixture was filtered through a 0.45μm microporous membrane to obtain a nanoliposome emulsion suspension. The suspension was centrifuged at 13000rpm for 30min to obtain Lip / DSPE@P.

[0067] Preparation of Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel: Dissolve 2.0 g of Gel-ADH-CA in 5 mL of PBS, then add dropwise 5 mL of PBS solution containing 2.0 g of ε-PL and 1 mg / mL of Lip / DSPE@P suspension. The mixture is reacted at room temperature for 5 h to obtain Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel.

[0068] Comparative Example 1 Preparation of Gel-ADH-CA: 2.0 g gelatin, 0.25 g ADH, 0.4 g EDC and 0.3 g NHS were dissolved in 100 mL of deionized water at 50 °C and reacted at 45 °C for 4 h. After the reaction was completed, the mixture was dialyzed and lyophilized to obtain white flocculent Gel-ADH. 0.5 g Gel-ADH was redissolved, and 0.25 g EDC and 0.20 g NHS were added. Then 10 mL of a solution containing 0.15 g CA was added dropwise, and the reaction was continued for 4 h. After the reaction was completed, the mixture was dialyzed. First, it was dialyzed with 20% ethanol-water for 12 h, and then dialyzed with pure water for 24 h. Finally, it was lyophilized to obtain brownish-yellow porous sponge-like Gel-ADH-CA.

[0069] Preparation of Lip@P liposomes: 0.2g soybean lecithin and 0.1mg cholesterol were dissolved in 5mL CH3Cl2, and then 1mg / mL Pemafibrat solution was added. The lipid membrane was obtained by rotary evaporation. Then, 3mL of PBS solution containing 2% Tween-80 and 0.5% methylcellulose was added dropwise. After sonication, the mixture was filtered through a 0.45μm microporous membrane to obtain a nanoliposome emulsion suspension. The suspension was centrifuged at 13000rpm for 30min to obtain Lip@P.

[0070] Preparation of Gel-ADH-CA / ε-PL / Lip@P hydrogel: Dissolve 1.0 g Gel-ADH-CA in 5 mL PBS, then add dropwise 5 mL PBS solution containing 1.0 g ε-PL and 1 mg / mL Lip@P suspension. React the mixture at room temperature for 5 h to obtain Gel-ADH-CA / ε-PL / Lip@P hydrogel.

[0071] Comparative Example 2 Preparation of Gel-ADH-CA: 2.0 g gelatin, 0.25 g ADH, 0.4 g EDC and 0.3 g NHS were dissolved in 100 mL of deionized water at 50 °C and reacted at 45 °C for 4 h. After the reaction was completed, the mixture was dialyzed and lyophilized to obtain white flocculent Gel-ADH. 0.5 g Gel-ADH was redissolved, and 0.25 g EDC and 0.20 g NHS were added. Then 10 mL of a solution containing 0.15 g CA was added dropwise, and the reaction was continued for 4 h. After the reaction was completed, the mixture was dialyzed. First, it was dialyzed with 20% ethanol-water for 12 h, and then dialyzed with pure water for 24 h. Finally, it was lyophilized to obtain brownish-yellow porous sponge-like Gel-ADH-CA.

[0072] Preparation of Lip / DSPE@P liposomes: 0.2g soybean lecithin, 0.1g cholesterol and 0.005g DSPE-TK-PEG2000 were dissolved in 5mL CH3Cl2, and then 1mg / mL Pemafibrat solution was added. The lipid membrane was obtained by rotary evaporation. Then, 3mL of PBS solution containing 2% Tween-80 and 0.5% methylcellulose was added dropwise. After sonication, the mixture was filtered through a 0.45μm microporous membrane to obtain a nanoliposome emulsion suspension. The suspension was centrifuged at 13000rpm for 30min to obtain Lip / DSPE@P.

[0073] Preparation of Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel: 1.0 g Gel-ADH-CA was dissolved in 5 mL PBS, and then 5 mL PBS solution containing 1.0 g ε-PL and 1 mg / mL Lip / DSPE@P suspension was added dropwise. The mixture was reacted at room temperature for 5 h to obtain Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel.

[0074] Comparative Example 3 Preparation of Gel-ADH-CA: 2.0 g gelatin, 0.25 g ADH, 0.4 g EDC and 0.3 g NHS were dissolved in 100 mL of 50 °C deionized water and reacted at 45 °C for 4 h. After the reaction was completed, the mixture was dialyzed and lyophilized to obtain white flocculent Gel-ADH. 0.5 g Gel-ADH was redissolved, and 0.25 g EDC and 0.20 g NHS were added. Then 10 mL of a solution containing 0.05 g CA was added dropwise, and the reaction was continued for 4 h. After the reaction was completed, the mixture was dialyzed. First, it was dialyzed with 20% ethanol-water for 12 h, and then dialyzed with pure water for 24 h. Finally, it was lyophilized to obtain brownish-yellow porous sponge-like Gel-ADH-CA.

[0075] Preparation of Lip / DSPE@P liposomes: 0.2g soybean lecithin, 0.1g cholesterol and 0.015g DSPE-TK-PEG2000 were dissolved in 5mL CH3Cl2, and then 1mg / mL Pemafibrat solution was added. The lipid membrane was obtained by rotary evaporation. Then, 3mL of PBS solution containing 2% Tween-80 and 0.5% methylcellulose was added dropwise. After sonication, the mixture was filtered through a 0.45μm microporous membrane to obtain a nanoliposome emulsion suspension. The suspension was centrifuged at 13000rpm for 30min to obtain Lip / DSPE@P.

[0076] Preparation of Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel: 1.0 g Gel-ADH-CA was dissolved in 5 mL PBS, and then 5 mL PBS solution containing 1.0 g ε-PL and 1 mg / mL Lip / DSPE@P suspension was added dropwise. The mixture was reacted at room temperature for 5 h to obtain Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel.

[0077] Comparative Example 4 Preparation of Gel-ADH-CA: 2.0 g gelatin, 0.25 g ADH, 0.4 g EDC and 0.3 g NHS were dissolved in 100 mL of deionized water at 50 °C and reacted at 45 °C for 4 h. After the reaction was completed, the mixture was dialyzed and lyophilized to obtain white flocculent Gel-ADH. 0.5 g Gel-ADH was redissolved, and 0.25 g EDC and 0.20 g NHS were added. Then 10 mL of a solution containing 0.15 g CA was added dropwise, and the reaction was continued for 4 h. After the reaction was completed, the mixture was dialyzed. First, it was dialyzed with 20% ethanol-water for 12 h, and then dialyzed with pure water for 24 h. Finally, it was lyophilized to obtain brownish-yellow porous sponge-like Gel-ADH-CA.

[0078] Preparation of Lip / DSPE@P liposomes: 0.2g soybean lecithin, 0.1g cholesterol and 0.01g DSPE-TK-PEG2000 were dissolved in 5mL CH3Cl2, and then 1mg / mL Pemafibrat solution was added. The lipid membrane was obtained by rotary evaporation. Then, 3mL of PBS solution containing 2% Tween-80 and 0.5% methylcellulose was added dropwise. After sonication, the mixture was filtered through a 0.45μm microporous membrane to obtain a nanoliposome emulsion suspension. The suspension was centrifuged at 13000rpm for 30min to obtain Lip / DSPE@P.

[0079] Preparation of Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel: 1.0 g of Gel-ADH-CA was dissolved in 5 mL of PBS, and then 5 mL of PBS solution containing 1.5 g of ε-PL and 1 mg / mL of Lip / DSPE@P suspension was added dropwise. The mixture was reacted at room temperature for 5 h to obtain Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel.

[0080] Comparative Example 5 Preparation of Gel-ADH-CA: 2.0 g gelatin, 0.25 g ADH, 0.4 g EDC and 0.3 g NHS were dissolved in 100 mL of deionized water at 50 °C and reacted at 45 °C for 4 h. After the reaction was completed, the mixture was dialyzed and lyophilized to obtain white flocculent Gel-ADH. 0.5 g Gel-ADH was redissolved, and 0.25 g EDC and 0.20 g NHS were added. Then 10 mL of a solution containing 0.15 g CA was added dropwise, and the reaction was continued for 4 h. After the reaction was completed, the mixture was dialyzed. First, it was dialyzed with 20% ethanol-water for 12 h, and then dialyzed with pure water for 24 h. Finally, it was lyophilized to obtain brownish-yellow porous sponge-like Gel-ADH-CA.

[0081] Preparation of Lip / DSPE@P liposomes: 0.2 g soybean lecithin, 0.1 mg cholesterol and 0.01 g DSPE-TK-PEG2000 were dissolved in 5 mL CH3Cl2, and then 1 mg / mL Pemafibrat solution was added. The lipid membrane was obtained by rotary evaporation. Then, 3 mL of PBS solution containing 2% Tween-80 and 0.5% methylcellulose was added dropwise. After sonication, the mixture was filtered through a 0.45 μm microporous membrane to obtain a nanoliposome emulsion suspension. The suspension was centrifuged at 13000 rpm for 30 min to obtain Lip / DSPE@P.

[0082] Preparation of Gel-ADH-CA / Lip / DSPE@P hydrogel: Dissolve 1.0 g of Gel-ADH-CA in 5 mL of PBS, then add 5 mL of PBS solution containing 1 mg / mL Lip / DSPE@P suspension. React the mixture at room temperature for 5 h to obtain Gel-ADH-CA / Lip / DSPE@P hydrogel.

[0083] Implementation effect evaluation 1. SEM image of the hydrogel Figure 1SEM images of the Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel are shown (accelerating voltage 5.00 kV, working distance 10.0 mm, magnification 150×). The images reveal a highly interconnected three-dimensional porous network structure with uniform pore size distribution, approximately 40–60 µm (scale bar 50 µm). The pore walls are thin and continuous, with no obvious collapse or dense regions, indicating sufficient cross-linking and good structural stability. The abundant porosity contributes to improved swelling properties and drug loading and release capabilities, providing a structural basis for its use as a ROS-responsive drug delivery carrier.

[0084] 2. Drug release test First, standard curves were established using pemafibrat at different concentrations (500, 250, 125, 62.5, 31.75 μg / mL). Then, H2O2 solutions of different concentrations (0 mM, 10 mM, 100 mM) were prepared. 400 μL of the Gel-ADH-CA / ε-PL / Lip / DSPE@P microgel assembly was gelled and placed in 1 mL of H2O2 solution of a specific concentration. The mixture was then incubated in a constant temperature shaking incubator at 37℃ and 100 rpm. Samples were taken at different time points (0.5, 1, 2, 4, 6, 8, 12, 16, 24 h). After taking 1 mL of culture medium with different H2O2 concentrations, the samples were replaced with an equal volume of fresh PBS. The samples were centrifuged at 4000 rpm for 5 min, and the absorbance of pemafibrat in PBS solution was measured using UV spectrophotometry at an excitation wavelength of 272 nm.

[0085] Figure 2 The linear relationship between the absorbance and concentration of pemafibrate at a specific wavelength was demonstrated to establish a standard curve for drug release. The results show that the fitted equation, y = 0.0359x – 0.03434, has a small standard error in both the slope and intercept, indicating a reliable fit. Correlation analysis shows that R0... 2 The value of 0.9994 indicates a good linear correlation between absorbance and concentration, and the sum of squared residuals of 0.02133 further demonstrates a very small fitting error. Overall, the standard curve shows a significant linear relationship and can be accurately used for quantitative analysis in subsequent Pemafibrate release experiments.

[0086] Table 2. Cumulative Pemafibrat release from Gel-ADH-CA / ε-PL / Lip / DSPE@P hydrogel at different H2O2 concentrations.

[0087] The drug release experiment results are shown in Table 2. Liposomes with different DSPE contents exhibited significantly different release behaviors under ROS conditions. Comparative Example 1 (without DSPE) showed a cumulative release rate of only 24.65% in 24 hours, indicating a slow release rate and a lack of ROS responsiveness. Comparative Example 2 (0.005g DSPE) showed a certain degree of accelerated release, with a cumulative release rate of 31.83% in 24 hours, but still below the ROS sensitivity requirement. Conversely, when the DSPE content was increased to 0.01g (Example 1), drug release accelerated significantly, reaching 21.82% in 2 hours and 85.56% in 24 hours, significantly higher than Comparative Examples 1 and 2, indicating that the liposomes at this ratio have excellent ROS-sensitive response characteristics. When the DSPE content was further increased to 0.015g (Comparative Example 3), the release rate was further improved, with a cumulative release rate of 88.13% in 24 hours, but the release was too rapid in the early stage (0.5~8 hours), showing a certain burst release phenomenon.

[0088] In summary, the introduction of DSPE endows liposomes with significant ROS responsiveness, enabling accelerated release in ROS environments. An appropriate DSPE content (e.g., 0.01 g) can ensure a good release rate while avoiding excessively rapid burst release, thus achieving a relatively ideal drug delivery effect. This fully demonstrates that the liposome system constructed in this study possesses ROS-sensitive release characteristics, and that the DSPE concentration has a direct regulatory effect on release efficiency. This invention shows that 0.005–0.01 g is the optimal concentration of DSPE, which can balance release efficiency and system stability, reflecting the excellent ROS-sensitive release performance of this liposome system.

[0089] 3. Antioxidant properties Test Method: The antioxidant performance of different examples and comparative examples was evaluated by scavenging 1,1-diphenyl-2-pyridylhydrazide (DPPH) free radicals. Mixtures of the different examples and comparative examples with DPPH reagent were incubated in the dark for 30 min with stirring. The remaining DPPH was analyzed using UV-Vis spectroscopy. The formula for determining the DPPH scavenging rate is: D (%) = {[A blank - (A assay - A control)] / A blank} × 100% Table 3 Free radical scavenging rate of the gel

[0090] The free radical scavenging experiment results showed that the caffeic acid content directly affected the antioxidant performance. Example 2 (caffeic acid content of 0) showed only about 5% scavenging rate, with almost no antioxidant effect; while Comparative Example 3 (caffeic acid content of 0.05g) showed an increased scavenging rate of about 47%, indicating that even a low caffeic acid content can produce a certain free radical scavenging effect. Further increasing the caffeic acid content to Example 3 (caffeic acid content of 0.10g), the scavenging rate reached about 67%, showing a significantly enhanced antioxidant effect. When the caffeic acid content was 0.15g (Examples 1, 4, 5 and Comparative Examples 1, 2, 4, 5), the free radical scavenging rate remained at a high level of about 83%~89%, indicating that the saturation effect was close at this content. Comparing the examples and comparative examples at the same content, it can be seen that the samples prepared by liposomes (Examples 1, 4, 5) and free caffeic acid (Comparative Examples 1, 2, 4, 5) all showed comparable or even slightly better scavenging rates, indicating that liposomes not only retain the antioxidant capacity of caffeic acid, but may also improve its stability and effectiveness.

[0091] In summary, the experimental results show that the caffeic acid content is concentration-dependent on the free radical scavenging rate, and the appropriate content range is 0.10–0.15 g, with 0.15 g being optimal. Furthermore, the liposome carrier system can maintain or even enhance the antioxidant effect of caffeic acid, demonstrating that the liposome possesses good antioxidant capacity.

[0092] 4. Gel mechanical strength test Table 4 Mechanical strength test of gel

[0093] According to the mechanical strength test results in Table 4, different ε-PL contents have a significant impact on the compressive modulus of the hydrogel. Examples 1, 2, and 3 (ε-PL 1.0 g) and Comparative Examples 1, 2, and 3 (ε-PL 1.0 g) show that the compressive modulus of the hydrogel is approximately 20.4~20.6 kPa, exhibiting relatively stable and moderate mechanical strength. In Example 4 (ε-PL 0.5 g), the modulus drops to 12.8 kPa, and the hydrogel is relatively soft, which may make it difficult to form a stable gel, thus affecting the hemostatic effect. In Example 5 (ε-PL 2.0 g), the modulus increases to 28.3 kPa, while that of Comparative Example 5 (ε-PL 0 g) is only 5.2 kPa, showing extreme cases of being too hard and too soft, respectively. The former may hinder drug release, while the latter is unlikely to achieve hemostasis.

[0094] Comprehensive analysis shows that the appropriate content of ε-PL should be around 1.0g. At this concentration, the hydrogel has moderate mechanical strength, which can form a stable gel to achieve hemostasis without being too hard to affect the timely release of drugs, thus reflecting a balance between mechanical properties and functionality.

[0095] 5. Adhesion strength rating Table 5. Viscosity rating of gels

[0096] Experimental results showed significant differences in tissue adhesion strength among hydrogels with different formulations. Comparative Example 3, with a caffeic acid content of 0.05 g, exhibited the lowest adhesion strength, only about 12 kPa, demonstrating weak tissue adhesion. When the CA content increased to 0.10 g (Example 3), the adhesion strength significantly improved to about 28 kPa, indicating that increasing the number of catechol groups effectively enhanced interfacial forces. In the examples and comparative examples with a caffeic acid content of 0.15 g, the adhesion strength reached approximately 40-44 kPa, showing the best and most stable performance overall, indicating that this content range can achieve strong adhesion performance while ensuring gel injectability. Therefore, a CA content range of 0.10-0.15 g can be considered the optimal choice for this system, significantly improving tissue adhesion strength while avoiding insufficient hemostatic ability due to excessively low concentrations.

[0097] 6. Coagulation Index Rating Whole blood coagulation index (WBCI) assay method: Pre-warmed sample discs at 37°C were placed at the bottom of a transparent plate (washed 3 times with PBS to remove surface free liquid). 200 μL of sodium citrate-anticoagulated whole blood and 20 μL of 0.2M CaCl2 were added to each well to initiate recalcification. After incubation at the preset time points (5 / 10 / 20 min), 2.0 mL of deionized water was quickly added and gently shaken for 3 min to dissolve red blood cells not embedded in the clot. The supernatant was collected and the absorbance was measured at 540 nm. The WBCI (%) was calculated based on the blank wells (without sample surface). The lower the value, the more complete the coagulation at that time point (the less residual soluble hemoglobin).

[0098] Table 9 Whole blood coagulation index of each group

[0099] The coagulation index (WBCI) of Examples 1-5 at 5 / 10 / 20 min was significantly better than that of Comparative Example 3 (p<0.05), indicating that the system of the present invention can initiate and stabilize thrombus formation to embed red blood cells more quickly. Among them, Examples 1, 4, and 5 reached a high plateau phase at 10-20 min, and the difference between them and Comparative Examples 1, 2, 4, and 5 was not significant (p>0.05), indicating that these formulations achieved a good and stable balance between initiation rate and final coagulation strength. Example 3 had a low WBCI at 5 min, but it rapidly increased from 10 to 20 min, approaching the "fully coagulated" plateau, suggesting that its initial chemical cross-linking / catechol-metal coordination and subsequent secondary solidification or liposome drug release (GW7647) synergistically accelerated the formation of fibrin network and improved thrombus density. Conversely, Comparative Example 3 (low CA) was significantly lower at all time points, suggesting that insufficient catechol sites weaken the hemostasis initiation rate and final clot strength. Overall, the hemostatic properties of this system are consistent with the synergistic effects of metal chelation and interfacial adhesion provided by catechol and drug release by liposomes under oxidative stress, supporting the optimal caffeic acid content range of 0.10~0.15g in existing formulations.

[0100] 7. Cytotoxicity Test method: According to GB / T 16886.5-2017 Biological evaluation of medical devices - Part 5: In vitro cytotoxicity test. After the colloid was prepared and completely cured according to the implementation method, it was extracted with culture medium for 24 hours according to the standard method; the extract was then used to treat L929 cells for 24 hours, and the absorbance was detected by CCK-8 assay and the viability was calculated. Normal cultured cells were used as controls; if the viability was <70%, it was considered to have potential cytotoxicity (n=3).

[0101] Table 10 Viscosity rating of gels

[0102] The cell survival rate of each group was ≥90%, with no potential cytotoxicity, proving that the system of the present invention has good cell compatibility.

[0103] In summary, this invention provides a novel therapeutic material that, through an innovative drug carrier design, effectively improves drug solubility and responds to the oxidative stress environment generated during renal ischemia-reperfusion injury (IRI), achieving precise drug release. Utilizing liposome drug delivery technology, this material effectively protects the activity of water-insoluble drugs (such as the PPARα agonist pemafibrate), ensuring that the drug is released to the kidney injury area at the necessary time. Simultaneously, the material also incorporates an injectable hydrogel system with sustained-release function, enabling continuous drug release after renal ischemia-reperfusion, prolonging the effective duration of drug action, reducing dosing frequency, and lowering the risk of drug side effects. This hydrogel enhances its mechanical strength and improves tissue adhesion through ion chelation with sodium ions in the blood and elevated iron ions in the oxidative stress environment, thereby promoting rapid hemostasis and effectively reducing kidney damage caused by IRI.

[0104] The technical solution of this invention solves several problems in the prior art. First, by using liposomes to deliver the PPARα agonist Pemafibrate, not only is the drug's solubility improved, but it also responds to the oxidative stress environment during renal ischemia-reperfusion, precisely releasing the drug and ensuring it exerts its effect at the appropriate time, thereby significantly improving therapeutic efficacy. Second, by utilizing a composite hydrogel system constructed from caffeic acid-modified gelatin and polylysine, controllable dynamic cross-linking of the hydrogel network structure can be achieved during ischemia-reperfusion. This is achieved through hydrogen bonding between the phenolic hydroxyl groups of caffeic acid and polylysine, and reversible electrostatic interactions between the carboxyl groups of gelatin and polylysine. This cross-linking not only significantly enhances the mechanical strength and tissue adhesion of the hydrogel, effectively promoting hemostasis, but also enables precise drug delivery to the surgical site through the encapsulated Pemafibrate liposomes. Pemafibrate, as an anti-inflammatory, antioxidant, and renal-protective drug, can be continuously released in this system, synergistically mitigating kidney damage caused by ischemia-reperfusion in conjunction with the physical barrier effect of the hydrogel. Furthermore, the cross-linked structure of the hydrogel ensures its stability at the kidney surgical site, providing an ideal environment for kidney recovery. Therefore, this invention provides an innovative drug delivery and hemostasis method that can precisely control drug release, enhance efficacy, and effectively reduce renal ischemia-reperfusion injury.

[0105] The technical features of the above embodiments can be combined in any way. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0106] The embodiments described above are merely illustrative of several implementations of the present invention, and while the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the invention patent. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these all fall within the protection scope of the present invention. Therefore, the protection scope of this invention patent should be determined by the appended claims.

Claims

1. A hydrogel, characterized in that, The hydrogel comprises caffeic acid-functionalized gelatin, polylysine, and drug-loaded liposomes.

2. The hydrogel according to claim 1, characterized in that, The raw materials for preparing the caffeic acid functionalized gelatin include gelatin, adipic acid dihydrazide, and caffeic acid.

3. The hydrogel according to claim 1, characterized in that, The liposome preparation materials for the drug-loaded liposomes include soybean lecithin, cholesterol, and nanocomposite materials. The nanocomposite materials are mainly composed of distearate phosphatidylethanolamine, ketethiocyanate, and polyethylene glycol. The drug includes a PPARα agonist.

4. The method for preparing the hydrogel according to claims 1-3, characterized in that, Includes the following steps: Preparation of caffeic acid functionalized gelatin: gelatin and adipic acid dihydrazide were dissolved, mixed, reacted, purified by dialysis, and freeze-dried to obtain gelatin-dipic acid dihydrazide. Gelatin-dipic acid dihydrazide was dissolved, activated, reacted with caffeic acid solution, purified by dialysis, and freeze-dried to obtain caffeic acid functionalized gelatin. Preparation of drug-loaded liposomes: Soybean lecithin, cholesterol and nanocomposite material were dissolved separately, drugs were added, and lipid membranes were obtained by rotary evaporation. Solvents were added to hydrate the lipid membranes, and the membranes were sonicated on ice. The liposome emulsion suspension was obtained by filtration and centrifugation to remove free drugs. Preparation of hydrogel: Dissolve caffeic acid-functionalized gelatin, add polylysine and drug-loaded liposome solution, stir and mix evenly in an ice bath, and let stand to obtain hydrogel.

5. The preparation method according to claim 4, characterized in that, The preparation of the caffeic acid-functionalized gelatin includes the following steps: dissolving gelatin in water, dissolving adipate dihydrazide in buffer solution, mixing the two and adding 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide for reaction, dialysis purification using a dialysis bag after the reaction, and lyophilizing to obtain gelatin-adipate dihydrazide; dissolving gelatin-adipate dihydrazide in buffer solution, adding 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide for activation in the dark, adding DMSO solution of caffeic acid for reaction, dialysis purification using a dialysis bag after the reaction, and lyophilizing to obtain caffeic acid-functionalized gelatin.

6. The preparation method according to claim 4, characterized in that, The preparation of the drug-loaded liposomes includes the following steps: dissolving soybean lecithin, cholesterol, and nanocomposite materials in an organic solvent, adding a drug solution, rotary evaporating the mixture to obtain a lipid membrane, adding a phosphate buffer solution to hydrate the lipid membrane, sonicating on ice, filtering to obtain a liposome emulsion suspension, and centrifuging to remove free drugs.

7. The preparation method according to claim 6, wherein the preparation method of the drug-loaded liposomes includes membrane hydration, and the organic solvent includes dichloromethane.

8. The preparation method according to claim 4, characterized in that, The preparation of the hydrogel includes the following steps: dissolving caffeic acid-functionalized gelatin in a phosphate buffer solution, adding a phosphate buffer solution containing polylysine and drug-loaded liposomes dropwise to the caffeic acid-functionalized gelatin solution, stirring and mixing evenly in an ice bath, and allowing it to stand to obtain the hydrogel.

9. The preparation method according to any one of claims 4-8, characterized in that, In the preparation of the caffeic acid functionalized gelatin, the mass ratio of gelatin to adipate dihydrazide is (6~10):1; and the mass ratio of gelatin-adipate dihydrazide to caffeic acid is 1:(0.1~0.5). In the preparation of the drug-loaded liposomes, the mass ratio of soybean lecithin:cholesterol:nanocomposite material is (10~30):(5~15):(0.5~2). In the preparation of the hydrogel, the mass ratio of caffeic acid functionalized gelatin: polylysine: drug-loaded liposomes is (150~450): (150~250): (0.1~2).

10. The hydrogel according to claims 1-3, or the hydrogel obtained by the preparation method according to claims 4-9, in the preparation of products for the treatment of kidney injury.

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