Surgical anti-adhesion fluid

Abstract

The present application relates to the technical field of medical biomaterials, and particularly relates to a surgical anti-adhesion liquid, which comprises A component and B component; the A component comprises catecholized-sulfhydrylated oxidized sodium alginate; and the B component comprises N-succinyl chitosan. The surgical anti-adhesion liquid provided by the present application has good fluidity in the initial use stage, can realize uniform flushing and sufficient coverage in the operation, and can rapidly form in-situ gelation under physiological conditions after contacting with tissues to form a stable and continuous barrier layer; the formation of the gel network is completed by the synergistic regulation of three types of functional groups, the aldehyde group and the amino group form a basic crosslinking network through Schiff base reaction, the catechol group forms a stable adhesion interface on the tissue surface through hydrogen bonding, covalent bonding and coordination, and the sulfhydryl group participates in a dynamic crosslinking reaction and a network reconstruction process, thereby providing additional crosslinking sites and enhancing the stability of the system.

Description

Technical Field

[0001] This invention relates to the field of medical biomaterials technology, and in particular to a surgical anti-adhesion solution. Background Technology

[0002] Postoperative tissue adhesions are one of the most common complications in abdominal and pelvic surgery. Their mechanism is closely related to the inflammatory response following tissue injury, fibrin deposition, and excessive proliferation of fibroblasts. To reduce the incidence of adhesions, physical barrier materials are typically used clinically to isolate damaged tissues in the early postoperative period, thereby blocking the formation of abnormal fibrous junctions.

[0003] Currently, the following physical barrier materials are mainly used in clinical practice: Hydroxyethyl starch-based irrigation solutions, as one of the earlier anti-adhesion products, mainly form a temporary physical barrier by increasing the retention time of intraperitoneal fluid. However, these products essentially rely solely on "hydration and buoyancy," lack a stable structure, and are easily absorbed or diluted by body fluids, resulting in a short duration of barrier effect that cannot cover the critical window period for adhesion formation (approximately 3 to 7 days post-surgery).

[0004] Hyaluronic acid and its sodium salts form a high-viscosity gel that covers the tissue surface, providing lubrication and physical isolation. However, hyaluronic acid is rapidly degraded by hyaluronidase in vivo, and its barrier effect typically lasts less than 3 days, making it insufficient to meet the protective needs throughout the entire adhesion formation process.

[0005] As research progressed, poloxamer 407 thermosensitive gel was developed. It is liquid at low temperatures and undergoes a sol-gel transition at body temperature, thus forming an in-situ gel barrier in vivo. However, this type of material suffers from low mechanical strength, susceptibility to dilution by body fluids, and poor structural stability. Furthermore, its function primarily relies on physical barrier action and lacks biological activity. In addition, high concentrations of poloxamer may trigger local irritation, limiting its clinical application.

[0006] In summary, existing surgical anti-adhesion materials generally suffer from insufficient structural stability. Therefore, improving the structural stability of surgical anti-adhesion materials is a technical problem that urgently needs to be solved. Summary of the Invention

[0007] To address the problem of insufficient structural stability in existing surgical anti-adhesion materials, this invention provides a surgical anti-adhesion liquid that can still adhere stably in a dynamic liquid environment and possesses good mechanical properties and controllable gelation behavior, thus solving the problem of insufficient structural stability in existing surgical anti-adhesion materials.

[0008] The technical solution adopted by this invention to solve its technical problem is: A surgical anti-adhesion solution includes component A and component B; component A includes catechol-thiolated oxidized sodium alginate; and component B includes N-succinyl chitosan.

[0009] Optionally, the catechol-thiolized sodium alginate is prepared according to the following method: S1: Oxidized sodium alginate is prepared by using sodium alginate and sodium periodate as raw materials through an oxidative ring-opening reaction; S2: Using the aforementioned oxidized sodium alginate and catechol derivatives as raw materials, catecholized-oxidized sodium alginate is prepared by an amino coupling reaction; S3: Using the aforementioned catecholized-oxidized sodium alginate and small molecule thiols as raw materials, catecholized-oxidized sodium alginate is prepared by a thiolation reaction.

[0010] Optionally, the oxidative ring-opening reaction in step S1 is a gradient oxidative ring-opening reaction.

[0011] Optionally, the gradient oxidation ring-opening reaction is carried out according to the following process: S11: Under light-protected conditions at a temperature of 0-5℃, sodium periodate is added to sodium alginate to carry out the first stage of oxidation, and the first oxidation product is obtained. S12: Raise the temperature of the first oxidation product to 15-25°C, add the second sodium periodate, and carry out the second stage reaction to obtain the second oxidation product; S13: Add a terminator to the second oxidation product, and then proceed with dialysis purification and freeze drying to obtain sodium alginate oxide.

[0012] Optionally, the average oxidation degree of the oxidized sodium alginate is 15% to 35%.

[0013] Optionally, the catechol derivative is selected from at least one of dopamine, 3,4-dihydroxyaniline, and 3,4-dihydroxybenzoic acid derivatives.

[0014] Optionally, the small molecule thiol is selected from at least one of mercaptoacetic acid, mercaptoethylamine, 2-mercaptoethanol, and N-acetylcysteine.

[0015] Optionally, the mass-volume concentration of the catecholized-thiolized oxidized sodium alginate in component A is 1.0% to 3.5%.

[0016] Optionally, the mass-volume concentration of N-succinyl chitosan in component B is 0.8% to 3.0%.

[0017] Optionally, component B may further include an antibacterial component.

[0018] The beneficial effects of this invention are: The surgical anti-adhesion solution provided by this invention has good fluidity in the initial use stage, enabling uniform rinsing and full coverage during surgery. After contact with tissue, it can rapidly gel in situ under physiological conditions to form a stable and continuous barrier layer. The formation of the gel network is synergistically regulated by three types of functional groups: aldehyde and amino groups form a basic cross-linking network through Schiff base reaction; catechol groups form a stable adhesion interface on the tissue surface through hydrogen bonding, covalent bonding, and coordination; and thiol groups participate in dynamic cross-linking reaction and network reconstruction process, providing additional cross-linking sites and enhancing the stability of the system. Detailed Implementation

[0019] The present invention will now be described in further detail. The embodiments described below are exemplary and intended to explain the present invention, and should not be construed as limiting the present invention. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without inventive effort are within the scope of protection of the present invention.

[0020] To address the problem of insufficient structural stability in existing surgical anti-adhesion materials, this invention provides a surgical anti-adhesion solution. This surgical anti-adhesion solution is a top-component system, comprising component A and component B. Before use, components A and B are stored separately, and during use, they are mixed to form an in-situ gel system. Specifically, this invention preferably includes catechol-thiolated oxidized sodium alginate (CT-OSA) in component A and N-succinyl chitosan (NSC) in component B.

[0021] Among them, catecholized-thiolated oxidized sodium alginate (CT-OSA) is a polysaccharide material containing trifunctional groups obtained by multiple modifications of sodium alginate. The molecular chain of this catecholized-thiolated oxidized sodium alginate (CT-OSA) simultaneously contains aldehyde structure, catechol group and thiol group. After being mixed with component B containing N-succinyl chitosan, it can form a dynamic cross-linking network through the following triple mechanism: (1) The aldehyde group reacts with the amino group via a Schiff base reaction to form an imine bond (C=N) crosslinking; (2) The catechol group adheres to the amino, thiol or metal ions on the tissue surface; (3) The mercapto group reacts with the residual aldehyde group via a thiazide reaction or forms hydrogen bonds with the amino group to strengthen the network.

[0022] Using N-succinyl chitosan as a crosslinking component reduces cytotoxicity while ensuring reactivity. Furthermore, the system's osmotic pressure and stability can be adjusted through various excipients, improving the overall biocompatibility and application suitability of the system, making the product more suitable for clinical irrigation fluid applications.

[0023] Specifically, after component A and component B are mixed, they can gel in situ within 60 to 150 seconds in a physiological environment of 37°C, forming a three-dimensional hydrogel network with strong adhesion and self-healing properties. The formed hydrogel forms a continuous barrier layer on the tissue surface to inhibit postoperative tissue adhesion.

[0024] The surgical anti-adhesion solution provided by this invention has good fluidity in the initial use stage, enabling uniform rinsing and full coverage during surgery. After contact with tissue, it can rapidly gel in situ under physiological conditions to form a stable and continuous barrier layer. The formation of the gel network is synergistically regulated by three types of functional groups: aldehyde and amino groups form a basic cross-linking network through Schiff base reaction; catechol groups form a stable adhesion interface on the tissue surface through hydrogen bonding, covalent bonding, and coordination; and thiol groups participate in dynamic cross-linking reaction and network reconstruction process, providing additional cross-linking sites and enhancing the stability of the system.

[0025] In this invention, the three functional groups do not act independently, but rather synergistically regulate the formation, structural stability, and interfacial adhesion behavior of the gel on both temporal and spatial scales, thereby constructing a three-dimensional network structure with multiple dynamic characteristics.

[0026] The preferred method for preparing catechol-thiolized oxidized sodium alginate (CT-OSA) according to the present invention is as follows: S1: Oxidized sodium alginate is prepared by using sodium alginate and sodium periodate as raw materials through an oxidative ring-opening reaction; In this step, the vicinal diol structure of the sugar unit in the sodium alginate molecular chain undergoes selective oxidation ring-opening to form an aldehyde group; S2: Catecholized-oxidized sodium alginate (C-OSA) was prepared by amino coupling reaction using sodium oxidized alginate and catechol derivatives as raw materials. In this step, the catechol group is grafted onto the aldehyde or carboxyl site of the sodium alginate molecular chain via a coupling reaction; preferably, the grafting degree of the catechol group is 5% to 20%. This step can be performed as follows: Dissolve 3-8 g of sodium alginate in 200 mL of distilled water and adjust the pH to 5.5; add catechol derivative (molar ratio of aldehyde to catechol derivative 1:(0.1-0.5), react at room temperature for 5-10 hours; dialysis purification (MWCO 3000-8000, dialysis for 24-72 hours), freeze-dry at -40 to -80℃ for 24-72 hours to obtain C-OSA; S3: Using catecholized-oxidized sodium alginate and small molecule thiols as raw materials, catecholized-oxidized sodium alginate was prepared by thiolation reaction; In this step, thiol groups are introduced through a thiolization reaction; the grafting degree of the thiol groups is preferably 3% to 15%; the molecular weight of CT-OSA is preferably 80 to 250 kDa, and some complete molecular chain segments are retained to maintain mechanical properties. This step can be performed as follows: Dissolve 1-5 g of C-OSA in 100 mL of distilled water and adjust the pH to 8.0. Add a small molecule thiol (molar ratio of 1:(0.3-0.8) to the residual aldehyde group) and react at room temperature for 10-15 hours. Purify by dialysis (MWCO 3000-8000, dialysis for 24-72 hours) and freeze-dry at -40 to -80℃ for 24-72 hours to obtain CT-OSA.

[0027] This invention introduces aldehyde, thiol and catechol groups to construct a ternary synergistic dynamic cross-linking network, forming a multiple dynamic cross-linking mechanism, which makes the gel network structure more stable and has self-healing ability. It can maintain a continuous barrier structure when subjected to mechanical disturbance or tissue movement, which is significantly better than a single Schiff base system.

[0028] This invention enables the controllable transformation of irrigation fluid into a highly adhesive gel through the synergistic regulation of multifunctional groups. This allows the system to maintain low viscosity and fluidity at room temperature, facilitating intraoperative operations. Simultaneously, it can rapidly form a gel upon contact with tissue and bind to the tissue through the catechol group, thereby significantly improving adhesion and solving the problem of traditional systems being easily washed away by body fluids.

[0029] The gel provided by this invention can maintain a high residue rate under fluid flushing conditions, significantly improving the adhesion stability in dynamic environments. It can adapt to dynamic liquid environments such as the abdominal cavity, solving the key problem of unstable adhesion of existing anti-adhesion materials in practical applications.

[0030] Furthermore, the preferred oxidation ring-opening reaction in step S1 of this invention is a gradient oxidation ring-opening reaction.

[0031] Specifically, the preferred procedure for this gradient oxidation ring-opening reaction is as follows: S11: Under light-protected conditions at a temperature of 0-5℃, sodium periodate is added to sodium alginate, denoted as sodium periodate one, to carry out the first stage of oxidation and obtain the first oxidation product; This step can be performed as follows: Sodium alginate was dissolved in distilled water to form a solution with a concentration of 1% to 3% (w / v). Under light-protected conditions at a temperature of 0 to 5°C, an oxidant was slowly added at a molar ratio of sodium alginate sugar units to sodium periodate of 1:(0.15 to 0.25). The reaction was carried out for 2 to 4 hours, which caused selective oxidation and ring-opening of the vicinal diol structure of some sugar units to generate aldehyde groups, thus obtaining the first oxidation product. The preferred sodium alginate of this invention has a molecular weight of 200 kDa and a G / M ratio of 0.8. S12: Raise the temperature of the first oxidation product to 15-25°C, add sodium periodate (referred to as second sodium periodate), and carry out the second stage reaction to obtain the second oxidation product. This step can be performed as follows: The temperature of the first-stage reaction system is raised to 15-25°C, and sodium periodate is added to make the total molar ratio of sugar units to sodium periodate reach 1:(0.3-0.5). The reaction continues for 3-6 hours. In the second-stage oxidation process, the oxidation reaction continues but the oxidation rate is lower than that in the first stage, so that the molecular chains form an oxidation degree gradient distribution. S13: Add a terminator to the second oxidation product, and then proceed with dialysis purification and freeze drying to obtain sodium alginate oxide; This step can be performed as follows: Ethylene glycol was added to terminate the oxidation reaction, and after dialysis purification for 48–72 hours, the mixture was freeze-dried to obtain graded oxidized sodium alginate (G-OSA).

[0032] By controlling the distribution of aldehyde groups on the molecular chain through a gradient oxidation process, the material can maintain the activity of cross-linking reaction while avoiding molecular chain breakage caused by excessive oxidation, thereby achieving a balance between cross-linking density and mechanical properties.

[0033] Specifically, in this invention, the average oxidation degree of sodium alginate is 15% to 35%, and the oxidation degree distribution on the molecular chain exhibits a gradient characteristic, with lower oxidation degree of sugar units near the ends of the molecular chain and higher oxidation degree of sugar units near the middle of the molecular chain, so as to retain some complete chain segments to maintain the mechanical properties of the material.

[0034] Furthermore, the catechol derivatives of the present invention are preferably selected from at least one of dopamine, 3,4-dihydroxyaniline, and 3,4-dihydroxybenzoic acid derivatives; the small molecule thiols are preferably selected from at least one of mercaptoacetic acid, mercaptoethylamine, 2-mercaptoethanol, and N-acetylcysteine.

[0035] Furthermore, the present invention preferably contains a catechol-thiolated sodium alginate in component A with a mass-volume concentration of 1.0% to 3.5% (w / v); preferably contains N-succinyl chitosan (NSC) in component B with a degree of substitution of 0.4 to 0.8, and preferably contains N-succinyl chitosan (NSC) in component B with a mass-volume concentration of 0.8% to 3.0% (w / v); and further preferably contains a volume ratio of component A to component B of 1:1.

[0036] Furthermore, the preferred surgical anti-adhesion solution of the present invention may also contain the following functional excipients: (1) Antibacterial component: ε-polylysine, with a concentration of 0.01% to 0.2% (w / v), is used to provide broad-spectrum antibacterial function; (2) Osmotic pressure regulators: 1.0% to 3.0% (w / v) glycerol and 0.6% to 0.9% (w / v) sodium chloride, used to adjust the osmotic pressure of the system to be close to the physiological environment; (3) Buffer system: 5-20 mM phosphate buffer solution, used to maintain the pH of the mixed system at 7.2-7.6; (4) Antioxidant: Mannitol 0.5%~2.0% (w / v) is used to inhibit the non-specific oxidation of aldehyde groups and improve storage stability.

[0037] Specifically, component A of the present invention preferably further includes a buffer system, an osmotic pressure regulator, and an antioxidant; component B preferably further includes a buffer system, an osmotic pressure regulator, and an antibacterial component.

[0038] This invention introduces ε-polylysine as an antibacterial component, enabling the gel to form a physical barrier while possessing active antibacterial function, thereby achieving a synergistic effect of antibacterial and anti-adhesion and effectively reducing the risk of postoperative infection.

[0039] The surgical anti-adhesion solution provided by this invention has an apparent viscosity of 50–200 mPa·s, preferably 80–150 mPa·s, at 25°C, maintaining good fluidity to meet the requirements of intraoperative irrigation. The hydrogel has the following performance characteristics: (1) Gelation time: 60-150 seconds at 37℃, preferably 60-90 seconds; (2) Tissue adhesion strength: tested with fresh pigskin, adhesion strength ≥15 kPa, preferably ≥18 kPa; (3) Self-healing efficiency: The compression modulus recovery rate within 30 minutes after gel cutting is ≥80%, preferably ≥85%; (4) In vitro degradation cycle: In PBS buffer containing hyaluronidase (50 U / mL), the residual mass rate was 30% to 60% within 7 to 21 days; (5) Antibacterial properties: The inhibition rate against Escherichia coli and Staphylococcus aureus is ≥90%.

[0040] In recent years, injectable hydrogel systems constructed from oxidized polysaccharides and amino-containing polymers via Schiff base reactions have attracted attention due to their in-situ gelation and certain biodegradability. However, existing Schiff base-based crosslinked hydrogel systems still have several limitations, restricting their clinical application in irrigation fluid scenarios.

[0041] First, existing systems primarily rely on physical coverage to form a barrier, lacking effective adhesion to the tissue interface. In the continuously flowing environment of the abdominal cavity or wound, they are easily washed away and detached, making it difficult to maintain stable coverage and resulting in insufficient barrier duration. Second, the single Schiff base crosslinking mechanism leads to a relatively simple gel network structure and limited crosslinking density, making it difficult to simultaneously achieve both mechanical properties and dynamic stability. This makes them prone to structural damage in complex physiological environments. Furthermore, traditional oxidative modification methods are mostly homogeneous oxidation processes, making precise control of the oxidation degree difficult. Excessive oxidation often leads to polysaccharide chain breakage, reducing the material's structural integrity and long-term stability, while insufficient oxidation affects crosslinking efficiency, making it difficult to achieve a performance balance. Simultaneously, for the specific application of irrigation fluids, existing gel systems generally suffer from high initial viscosity and insufficient fluidity, hindering uniform spreading and operational control during surgery.

[0042] To address the aforementioned issues, this invention develops a novel hydrogel system that combines low viscosity flowability, strong tissue adhesion, and multiple dynamic cross-linking capabilities. This invention constructs a biomimetic adhesion interface by introducing catechol groups onto an oxidized polysaccharide backbone, while simultaneously introducing thiol groups to provide additional dynamic cross-linking sites. Furthermore, a gradient oxidation strategy is employed to achieve synergistic regulation of molecular chain structure and reactivity. This results in a hydrogel system for anti-adhesion irrigation that maintains stable adhesion in dynamic liquid environments and possesses excellent mechanical properties and controllable gel behavior. The gel formed by this invention not only exhibits excellent tissue adhesion but also maintains continuous adhesion in dynamic body fluid environments. It can maintain barrier integrity under tissue movement or mechanical disturbance conditions, effectively isolating tissue contact and preventing postoperative adhesions.

[0043] This invention introduces catechol-thiolized sodium alginate, wherein the catechol groups adhere to the tissue surface via the following mechanism: (1) Hydrogen bonding: The hydroxyl groups in the catechol group form hydrogen bonds with the amino and hydroxyl groups in the protein molecules on the tissue surface, contributing 20% ​​to 35% to the adhesion strength; (2) Covalent bond effect: under weakly alkaline conditions, the catechol group can be oxidized to a quinone structure. The quinone structure forms a covalent bond with the thiol or amino groups on the tissue surface through Michael addition reaction or Schiff base reaction, contributing 40% to 60% to the adhesion strength; (3) Metal chelation: The catechol group forms chelate bonds with calcium and iron ions on the tissue surface or in the extracellular matrix, contributing 10% to 20% to the adhesion strength; The three adhesion mechanisms work synergistically to achieve a tissue adhesion strength of 15–25 kPa for CT-OSA-based hydrogels.

[0044] The cross-linking enhancement mechanism of the thiol-thiolized oxidized sodium alginate includes: (1) Thiane reaction: The mercapto group reacts with the residual aldehyde group on the CT-OSA molecular chain to form a thiane bond (CSC), providing additional covalent crosslinking sites; (2) Hydrogen bond enhancement: The thiol group forms a strong hydrogen bond with the amino group on the N-succinyl chitosan molecular chain, which enhances the stability of the cross-linking network; (3) Dynamic reversibility: The thiol-aldehyde reaction is reversible, enabling the crosslinked network to have self-healing ability. The modulus recovery rate is not less than 80% within 30 minutes after cutting.

[0045] This invention provides a highly adhesive, in-situ gelling anti-adhesion rinsing solution constructed based on a multiple dynamic cross-linking mechanism, which is suitable for the prevention of tissue adhesions after abdominal and pelvic surgeries in general surgery, obstetrics and gynecology, orthopedics, and other procedures.

[0046] In summary, this invention provides a two-component in-situ gelling anti-adhesion rinsing solution constructed based on a catechol-mercapto-aldehyde ternary synergistic dynamic crosslinking mechanism. The anti-adhesion rinsing solution adopts a two-component separation and storage system, including solution A and solution B.

[0047] Solution A is a modified sodium alginate aqueous solution containing multifunctional groups, specifically a sodium alginate derivative (CT-OSA) obtained through gradient oxidation, catecholization, and thiolation modification; Solution B is an amino-containing N-succinyl chitosan aqueous solution and a functional excipient system.

[0048] The two components are mixed in a predetermined ratio before use to form a low-viscosity fluid system that maintains good fluidity at room temperature to meet the needs of intraoperative irrigation. After contact with human tissue, a three-dimensional cross-linked hydrogel barrier is rapidly formed within about 60 to 120 seconds through multiple dynamic reactions at physiological temperature (37°C) and in a neutral environment (pH 7.2 to 7.6).

[0049] in: The mass concentration of CT-OSA in solution A is 1.5% to 3.5% (w / v), and its molecular weight is 100 to 300 kDa; sodium alginate is prepared by a gradient oxidation process with an oxidation degree of 15% to 40%, preferably 20% to 30%, and catechol groups and thiol groups are introduced into the molecular chain. Solution B contains N-succinyl chitosan at a mass concentration of 1.0%–3.0% (w / v) and a degree of substitution of 0.4–0.8. It also contains the following excipient system: glycerol (1.0%–3.0%), sodium chloride (0.6%–0.9%), phosphate buffer system (5–20 mM), ε-polylysine (0.01%–0.1%), and mannitol (0.5%–2.0%).

[0050] To make the above-mentioned objects, features and advantages of the present invention more apparent and understandable, specific embodiments of the present invention will be described in detail below. Example 1

[0051] Step 1: Gradient oxidation to prepare G-OSA 10 g of sodium alginate (molecular weight 200 kDa, G / M=0.8) was dissolved in 500 mL of distilled water (2% concentration). The solution was placed in an ice bath (0–4°C) and, under dark conditions, sodium periodate solution (molar ratio 1:0.2) was slowly added, and the reaction was allowed to proceed for 3 hours. The system temperature was then raised to 20°C, and sodium periodate was added again (total molar ratio 1:0.35), and the reaction was continued for 4 hours. The reaction was terminated by adding ethylene glycol, dialyzed (MWCO 3500) for 48 hours, and then freeze-dried to obtain G-OSA.

[0052] Step 2: Catechinization to prepare C-OSA Dissolve 5 g of G-OSA in 200 mL of distilled water and adjust the pH to 5.5. Add dopamine (aldehyde to dopamine molar ratio 1:0.3) and react at room temperature for 8 hours. Purify by dialyzing and freeze-dry to obtain C-OSA.

[0053] Step 3: Thiolization to prepare CT-OSA Dissolve 3 g of C-OSA in 100 mL of distilled water and adjust the pH to 8.0. Add mercaptoethylamine (molar ratio of 1:0.5 to residual aldehyde group) and react at room temperature for 12 hours. Purify by dialyzing and freeze-dry to obtain CT-OSA.

[0054] Preparation of solution A: Weigh 2.0 g of CT-OSA and add it to 100 mL of phosphate buffer (10 mM, pH 7.4), stirring to dissolve. Add 0.7 g of sodium chloride and 1.0 g of mannitol, then filter to sterilize.

[0055] Preparation of solution B: Weigh 1.5 g of NSC (degree of substitution 0.6), add 100 mL of phosphate buffer, and stir to dissolve. Add 2.0 g of glycerol and 0.05 g of ε-polylysine, and filter to sterilize.

[0056] How to use: Mix liquid A and liquid B at a volume ratio of 1:1, gently invert to mix thoroughly, and obtain an in-situ gelation system. Example 2

[0057] Step 1: Gradient oxidation to prepare G-OSA 10 g of sodium alginate (molecular weight 200 kDa, G / M=0.8) was dissolved in 500 mL of distilled water (2% concentration). The solution was placed in an ice bath (0–4°C) and, under dark conditions, sodium periodate solution (molar ratio 1:0.2) was slowly added, and the reaction was allowed to proceed for 3 hours. The system temperature was then raised to 20°C, and sodium periodate was added again (total molar ratio 1:0.35), and the reaction was continued for 4 hours. The reaction was terminated by adding ethylene glycol, dialyzed (MWCO 3500) for 48 hours, and then freeze-dried to obtain G-OSA.

[0058] Step 2: Catechinization to prepare C-OSA Dissolve 5 g of G-OSA in 200 mL of distilled water and adjust the pH to 5.5. Add dopamine (aldehyde to dopamine molar ratio 1:0.3) and react at room temperature for 8 hours. Purify by dialyzing and freeze-dry to obtain C-OSA.

[0059] Step 3: Thiolization to prepare CT-OSA Dissolve 3 g of C-OSA in 100 mL of distilled water and adjust the pH to 8.0. Add mercaptoethylamine (molar ratio of 1:0.5 to residual aldehyde group) and react at room temperature for 12 hours. Purify by dialyzing and freeze-dry to obtain CT-OSA.

[0060] Preparation of solution A: Weigh 3.0 g of CT-OSA and add 100 mL of phosphate buffer (10 mM, pH 7.4), stirring to dissolve. Add 0.7 g of sodium chloride and 1.0 g of mannitol, then filter to sterilize.

[0061] Preparation of solution B: Weigh 2.5 g of NSC (degree of substitution 0.6), add 100 mL of phosphate buffer, and stir to dissolve. Add 2.0 g of glycerol and 0.05 g of ε-polylysine, and filter to sterilize.

[0062] How to use: Mix liquid A and liquid B at a volume ratio of 1:1, gently invert to mix thoroughly, and obtain an in-situ gelation system.

[0063] This embodiment is based on Example 1, but the CT-OSA concentration is increased to 3.0% (w / v), the NSC concentration is increased to 2.5% (w / v), and the other conditions remain unchanged. Example 3

[0064] Step 1: Gradient oxidation to prepare G-OSA 10 g of sodium alginate (molecular weight 200 kDa, G / M=0.8) was dissolved in 500 mL of distilled water (2% concentration). The solution was placed in an ice bath (0–4°C) and, under dark conditions, sodium periodate solution (molar ratio 1:0.2) was slowly added, and the reaction was allowed to proceed for 3 hours. The system temperature was then raised to 20°C, and sodium periodate was added again (total molar ratio 1:0.35), and the reaction was continued for 4 hours. The reaction was terminated by adding ethylene glycol, dialyzed (MWCO 3500) for 48 hours, and then freeze-dried to obtain G-OSA.

[0065] Step 2: Catechinization to prepare C-OSA Dissolve 5 g of G-OSA in 200 mL of distilled water and adjust the pH to 5.5. Add dopamine (aldehyde to dopamine molar ratio 1:0.5) and react at room temperature for 8 hours. Purify by dialyzing and freeze-dry to obtain C-OSA.

[0066] Step 3: Thiolization to prepare CT-OSA Dissolve 3 g of C-OSA in 100 mL of distilled water and adjust the pH to 8.0. Add mercaptoethylamine (molar ratio of 1:0.8 to residual aldehyde) and react at room temperature for 12 hours. Purify by dialyzing and freeze-dry to obtain CT-OSA.

[0067] Preparation of solution A: Weigh 2.0 g of CT-OSA and add it to 100 mL of phosphate buffer (10 mM, pH 7.4), stirring to dissolve. Add 0.7 g of sodium chloride and 1.0 g of mannitol, then filter to sterilize.

[0068] Preparation of solution B: Weigh 1.5 g of NSC (degree of substitution 0.6), add 100 mL of phosphate buffer, and stir to dissolve. Add 2.0 g of glycerol and 0.05 g of ε-polylysine, and filter to sterilize.

[0069] How to use: Mix liquid A and liquid B at a volume ratio of 1:1, gently invert to mix thoroughly, and obtain an in-situ gelation system.

[0070] This embodiment is based on Example 1, but the molar ratio of aldehyde group to dopamine in oxidized sodium alginate is increased to 1:0.5; in the thiolation step, the molar ratio of mercaptoethylamine to residual aldehyde group is increased to 1:0.8, while the other reaction conditions remain unchanged.

[0071] Each comparative example in this invention is compared with Example 1.

[0072] Comparative Example 1 Step 1: Gradient oxidation to prepare G-OSA 10 g of sodium alginate (molecular weight 200 kDa, G / M=0.8) was dissolved in 500 mL of distilled water (2% concentration). The solution was placed in an ice bath (0–4°C) and, under dark conditions, sodium periodate solution (molar ratio 1:0.2) was slowly added, and the reaction was allowed to proceed for 3 hours. The system temperature was then raised to 20°C, and sodium periodate was added again (total molar ratio 1:0.35), and the reaction was continued for 4 hours. The reaction was terminated by adding ethylene glycol, dialyzed (MWCO 3500) for 48 hours, and then freeze-dried to obtain G-OSA.

[0073] Step 2: Catechinization to prepare C-OSA Dissolve 5 g of G-OSA in 200 mL of distilled water and adjust the pH to 5.5. Add dopamine (aldehyde to dopamine molar ratio 1:0.3) and react at room temperature for 8 hours. Purify by dialyzing and freeze-dry to obtain C-OSA.

[0074] Preparation of solution A: Weigh 2.0 g of C-OSA and add it to 100 mL of phosphate buffer (10 mM, pH 7.4), stirring to dissolve. Add 0.7 g of sodium chloride and 1.0 g of mannitol, then filter to sterilize.

[0075] Preparation of solution B: Weigh 1.5 g of NSC (degree of substitution 0.6), add 100 mL of phosphate buffer, and stir to dissolve. Add 2.0 g of glycerol and 0.05 g of ε-polylysine, and filter to sterilize.

[0076] How to use: Mix liquid A and liquid B at a volume ratio of 1:1, gently invert to mix thoroughly, and obtain an in-situ gelation system.

[0077] In this comparative example, catecholized sodium alginate (C-OSA) was used instead of CT-OSA, and the other conditions were the same as in Example 1.

[0078] Comparative Example 2 Preparation of OSA-SH by thiolation Dissolve 3 g of sodium alginate (molecular weight 200 kDa, G / M=0.8) in 100 mL of distilled water and adjust the pH to 8.0. Add mercaptoethylamine (molar ratio of 1:0.5 to aldehyde) and react at room temperature for 12 hours. Purify by dialysis and freeze-dry to obtain OSA-SH.

[0079] Preparation of solution A: Weigh 2.0 g of OSA-SH and add it to 100 mL of phosphate buffer (10 mM, pH 7.4), stirring to dissolve. Add 0.7 g of sodium chloride and 1.0 g of mannitol, then filter to sterilize.

[0080] Preparation of solution B: Weigh 1.5 g of NSC (degree of substitution 0.6), add 100 mL of phosphate buffer, and stir to dissolve. Add 2.0 g of glycerol and 0.05 g of ε-polylysine, and filter to sterilize.

[0081] How to use: Mix liquid A and liquid B at a volume ratio of 1:1, gently invert to mix thoroughly, and obtain an in-situ gelation system.

[0082] In this comparative example, sodium alginate oxidized without catechol modification (OSA-SH, only thiolized) was used instead of CT-OSA, and the other conditions were the same as in Example 1.

[0083] Comparative Example 3 Preparation of solution A: Weigh 2.0 g of sodium alginate (molecular weight 200 kDa, G / M=0.8), add 100 mL of phosphate buffer (10 mM, pH 7.4), and stir to dissolve. Add 0.7 g of sodium chloride and 1.0 g of mannitol, and filter to sterilize.

[0084] Preparation of solution B: Weigh 1.5 g of NSC (degree of substitution 0.6), add 100 mL of phosphate buffer, and stir to dissolve. Add 2.0 g of glycerol, and filter to sterilize.

[0085] How to use: Mix liquid A and liquid B at a volume ratio of 1:1, gently invert to mix thoroughly, and obtain an in-situ gelation system.

[0086] In this comparative example, ordinary oxidized sodium alginate (OSA) was used instead of CT-OSA, and catechol and thiol groups were not introduced. The other conditions were the same as in Example 1.

[0087] The performance of the surgical anti-adhesion solutions prepared in the above embodiments and comparative examples was tested using the following methods: 1. Gelation time test Methods: Examples 1-3 and Comparative Examples 1-3 were mixed with liquid A and liquid B at a volume ratio of 1:1. To evaluate the operational performance and gelation behavior of the materials in actual clinical use, tests were conducted at room temperature (25°C) and physiological temperature (37°C).

[0088] First, room temperature viscosity was measured. The mixed sample was immediately placed in a rotational rheometer at 25°C, using a rotor for low-viscosity LV-2 fluid, and the shear rate was set to 50 s⁻¹. -1 The sample was continuously measured for 30 seconds, and its apparent viscosity (unit: mPa·s) under steady-state conditions was recorded. Each group of samples was tested in parallel three times, and the average value was taken as the final result.

[0089] The gelation time was then determined. The mixed samples from the same batch were quickly transferred to a 37°C constant temperature water bath, and the inverted test tube method was used for detection. Specifically, the sample was added to a transparent test tube, and the tube was slowly inverted every 5 seconds. The time required for the system to stop flowing in the inverted state was recorded as the gelation time. Each group underwent three parallel experiments, and the average value was taken.

[0090] The test results are shown in Table 1: Table 1 The experimental results show that all systems in each embodiment exhibited low apparent viscosity (approximately 120–210 mPa·s) at room temperature (25°C), falling within the typical low-viscosity fluid range, which meets the requirements for flowability and operability during intraoperative irrigation. Specifically, Example 1 had a viscosity of 120 mPa·s, within the optimal range, ensuring good flowability while achieving uniform spreading. Example 2, due to increased concentrations of CT-OSA and NSC, experienced enhanced polymer chain entanglement, leading to a viscosity increase to 210 mPa·s. Although the flowability decreased slightly, it remained within acceptable limits. Example 3, by increasing the grafting degree of catechol and thiol groups, maintained a low viscosity without significantly increasing the polymer concentration, indicating that increasing the density of functional groups does not significantly affect rheological properties.

[0091] Regarding gelation time, Example 2 exhibited the fastest gelation speed (approximately 60 seconds), primarily attributed to the higher density of crosslinking sites accelerating the Schiff base reaction and assisted crosslinking process. Examples 1 and 3 had gel times of 80 seconds and 75 seconds, respectively, ensuring sufficient processing time while rapidly forming stable structures in vivo. In contrast, Comparative Example 3 (traditional OSA system) had the longest gelation time (approximately 90 seconds), indicating relatively low crosslinking efficiency of the single Schiff base reaction. While Comparative Examples 1 and 2 could form gels, the overall crosslinking process remained relatively simple due to the lack of synergistic effects from thiol or catechol, and the reaction rate was not significantly improved.

[0092] In summary, this invention, by constructing a ternary synergistic system of aldehyde-mercapto-catechol, achieves a good balance between gelation rate and operating window without significantly increasing the system viscosity, demonstrating superior rheological and gelling performance matching compared to traditional systems.

[0093] 2. Self-healing performance test Methods: Samples from each group were mixed to form a gel, which was then used to prepare cylindrical hydrogels with a diameter of 20 mm and a thickness of 5 mm. The gels were cut in half using a blade, and the two halves were immediately reconnected and allowed to stand at 37°C for 30 minutes. Compression tests were then performed using a universal testing machine to compare the compressive modulus recovery rates of the intact gel and the self-healing gel. Simultaneously, macroscopic observation was used to record whether the repair interface disappeared.

[0094] The test results are shown in Table 2: Table 2 The self-healing performance test results showed that the modulus recovery rates of Examples 1 and 3 reached 88% and 90%, respectively, significantly higher than that of Comparative Example 3 (60%), indicating that the ternary synergistic system has excellent structural reconstruction capabilities. This performance mainly stems from the synergistic mechanism of multiple dynamic interactions within the system: First, the Schiff base bonds themselves possess a certain degree of reversibility, allowing them to reform after breakage; second, reversible reactions or exchange reactions can occur between thiol groups or between thiol and aldehyde groups, providing the network with additional dynamic recombination capabilities; furthermore, the catechol groups can further enhance local structural stability through hydrogen bonding and weak interactions. This multi-dynamic interaction enables the gel to rapidly recover its network structure after mechanical damage.

[0095] The modulus recovery rate of Example 2 was 72%, significantly lower than that of Examples 1 and 3. This is because the higher crosslinking density limited the migration ability of the molecular chains, making it difficult for the chain segments at the fracture interface to rearrange, thus affecting the self-healing efficiency. The recovery rates of Comparative Examples 1 and 2 were 75% and 78%, respectively. Although these were higher than the traditional system, they were still lower than the ternary system, indicating that a single functional group cannot provide sufficient dynamic recombination ability. Comparative Example 3, due to its reliance solely on the Schiff base reaction, had a relatively rigid network structure and lacked multiple reversible mechanisms, resulting in the worst self-healing performance.

[0096] 3. In vitro degradation test Methods: Hydrogel samples (initial mass W0) were placed in centrifuge tubes containing 50 U / mL PBS buffer (pH 7.4) and incubated at 37°C with shaking. Samples were removed every 1, 3, 7, and 14 days, surface moisture was removed, and the samples were weighed (W0). t ), calculate the residual mass rate: Residual rate (%) = W t / W0 × 100% Three parallel samples were set up for each group of samples.

[0097] The test results are shown in Table 3.

[0098] Table 3 In vitro degradation experiments showed that the residual rates of Examples 1, 2, and 3 at 14 days were approximately 42%, 62%, and 50%, respectively, significantly higher than that of Comparative Example 3 (20%). This result indicates that the system of the present invention has superior structural stability and controllable degradation characteristics. Example 2 exhibited the slowest degradation rate due to its highest crosslinking density, which is closely related to its denser network structure. Examples 1 and 3 demonstrated moderate degradation rates, maintaining an effective barrier during the critical anti-adhesion window of 7–14 days post-surgery, while gradually degrading in the later stages to avoid long-term residue.

[0099] In contrast, Comparative Example 3 degraded faster, with a residual rate of only 45% after 7 days, indicating that the traditional single Schiff base cross-linking system has poor stability in the body fluid environment and is difficult to maintain a long-term barrier effect. Although Comparative Examples 1 and 2 showed slight improvement over Comparative Example 3, their degradation behavior was still difficult to precisely control due to the lack of a ternary synergistic cross-linking structure.

[0100] 4. Antibacterial performance test Methods: The antibacterial properties were tested using the plate coating method. *Escherichia coli* (Gram-negative) and *Staphylococcus aureus* (Gram-positive) were selected as representative strains. After preparing gels from each group of samples, they were placed in a bacterial suspension (10⁻⁶ m² / 400 μL). 6 The culture was co-cultured for 24 hours with CFU / mL. Subsequently, the culture was serially diluted and plated onto LB agar plates, incubated at 37°C for 24 hours, and the colony count (CFU) was calculated to determine the inhibition rate. Inhibition rate (%) = (Number of colonies in control group - Number of colonies in experimental group) / Number of colonies in control group × 100% The test results are shown in Table 4.

[0101] Table 4 Antibacterial test results showed that the inhibition rates of the system in the examples against *Escherichia coli* and *Staphylococcus aureus* both reached over 94%, significantly better than Comparative Example 3 (approximately 40%–45%). This result is mainly attributed to the introduction of ε-polylysine into the system, which, as a natural cationic antimicrobial peptide, can achieve broad-spectrum antibacterial activity by disrupting the bacterial cell membrane structure. The antibacterial performance differences between Examples 1, 2, and 3 were minimal, indicating that ternary modification does not affect the effectiveness of the antibacterial components.

[0102] Comparative Examples 1 and 2, while also containing antibacterial components, exhibited slightly lower antibacterial rates than the system in the Example, possibly due to their weaker gel network structure. In a looser structure, antibacterial components diffuse and dissipate more easily, reducing local antibacterial concentration. In contrast, the dense network structure formed by the ternary synergistic system helps limit the rapid release of antibacterial molecules, maintaining a higher concentration in the local environment and thus enhancing the antibacterial effect. Comparative Example 3, lacking antibacterial components, showed significantly poor antibacterial effect, exhibiting only a slight inhibitory effect. This indicates that the material itself does not possess significant antibacterial capabilities, and the antibacterial performance primarily stems from the introduction of functional components.

[0103] 5. Dynamic scouring adhesion performance test Methods: Fresh porcine peritoneum or porcine small intestinal serosa tissue was selected as the substrate material and cut into tissue slices approximately 2 cm × 2 cm in size. The slices were repeatedly rinsed with physiological saline to remove surface blood and impurities, and then placed in phosphate-buffered saline (PBS, pH 7.4) for later use. Before the experiment, the tissue surface was gently blotted dry to ensure uniform adhesion of the material.

[0104] Mix solution A and solution B at a volume ratio of 1:1, gently invert to mix, and immediately drop the mixture onto the tissue surface. The amount added to each sample should be controlled at 0.5 mL, ensuring even coverage of the entire tissue area. Incubate at a constant temperature of 37°C for approximately 2 minutes to allow the system to undergo initial gelation and form a stable adhesion layer.

[0105] Subsequently, the processed tissue samples were fixed in a flow apparatus or shaker culture plate with the gel layer facing upwards, exposed to the fluid environment. Preheated PBS buffer (approximately 50 mL) to 37°C was added to the system, and the shaker speed was set to 100 rpm, maintaining a constant temperature of 37°C to simulate the in vivo fluid flow and shear stress environment. Samples were removed at set time points (e.g., 2 hours, 4 hours, 6 hours), and the surface was gently rinsed with PBS to remove any adhering substances. The change in sample mass was measured by weighing, and the residual rate (residual gel mass) was used as the evaluation index. To improve the reliability of the experiment, at least three parallel samples were prepared for each group, and the average value was used for statistical analysis.

[0106] The test results are shown in Table 5.

[0107] Table 5 Dynamic scouring experiments showed that Examples 1 and 3 retained approximately 78% and 82% of their residue after 6 hours, respectively, while Comparative Example 3 only retained 20%, a highly significant difference. This is because the catechol groups can form strong interactions with the tissue surface through various mechanisms such as hydrogen bonding, covalent bonding, and metal ion coordination, thereby establishing a stable interfacial bonding layer; the thiol groups improve the overall structure's resistance to damage by participating in network reconstruction and enhancing cross-linking density; and the aldehyde groups provide the basic cross-linking framework, enabling the gel to rapidly form a continuous structure. The synergistic effect of these three elements gives the system excellent adhesion stability in a dynamic liquid environment, which is also a key advantage that distinguishes it from existing technologies.

[0108] Comparative Example 1 (without thiol) and Comparative Example 2 (without catechol) both showed significantly reduced residue rates, at 45% and 35%, respectively, indicating that a single functional group cannot provide sufficient erosion resistance. Comparative Example 2, in particular, was extremely prone to detachment in dynamic environments due to the lack of catechol adhesion. Comparative Example 3, lacking both an adhesive structure and multiple cross-linking, performed the worst.

[0109] Based on the above-described preferred embodiments of the present invention, and through the foregoing description, those skilled in the art can make various changes and modifications without departing from the inventive concept. The technical scope of this invention is not limited to the contents of the specification, but must be determined according to the scope of the claims.

Claims

1. A surgical anti-adhesion fluid, characterized in that, It includes component A and component B; component A includes catecholized-thiolated oxidized sodium alginate; component B includes N-succinyl chitosan.

2. The surgical anti-adhesion solution as described in claim 1, characterized in that, The catecholized-thiolized oxidized sodium alginate was prepared according to the following method: S1: Oxidized sodium alginate is prepared by using sodium alginate and sodium periodate as raw materials through an oxidative ring-opening reaction; S2: Using the aforementioned oxidized sodium alginate and catechol derivatives as raw materials, catecholized-oxidized sodium alginate is prepared by an amino coupling reaction; S3: Using the aforementioned catecholized-oxidized sodium alginate and small molecule thiols as raw materials, catecholized-oxidized sodium alginate is prepared by a thiolation reaction.

3. The surgical anti-adhesion solution as described in claim 2, characterized in that, The oxidative ring-opening reaction in step S1 is a gradient oxidative ring-opening reaction.

4. The surgical anti-adhesion solution as described in claim 3, characterized in that, The gradient oxidation ring-opening reaction is carried out according to the following process: S11: Under light-protected conditions at a temperature of 0-5℃, sodium periodate is added to sodium alginate to carry out the first stage of oxidation, and the first oxidation product is obtained. S12: Raise the temperature of the first oxidation product to 15-25°C, add the second sodium periodate, and carry out the second stage reaction to obtain the second oxidation product; S13: Add a terminator to the second oxidation product, and then proceed with dialysis purification and freeze drying to obtain sodium alginate oxide.

5. The surgical anti-adhesion solution as described in claim 4, characterized in that, The average oxidation degree of the oxidized sodium alginate is 15% to 35%.

6. The surgical anti-adhesion solution as described in claim 2, characterized in that, The catechol derivative is selected from at least one of dopamine, 3,4-dihydroxyaniline, and 3,4-dihydroxybenzoic acid derivatives.

7. The surgical anti-adhesion solution as described in claim 2, characterized in that, The small molecule thiol is selected from at least one of thioglycolic acid, mercaptoethylamine, 2-mercaptoethanol, and N-acetylcysteine.

8. The surgical anti-adhesion solution according to any one of claims 1-7, characterized in that, The mass-volume concentration of the catecholized-thiolized oxidized sodium alginate in component A is 1.0% to 3.5%.

9. The surgical anti-adhesion solution according to any one of claims 1-7, characterized in that, The N-succinyl chitosan in component B has a mass-volume concentration of 0.8% to 3.0%.

10. The surgical anti-adhesion solution as described in claim 9, characterized in that, Component B also includes an antibacterial component.

Images