A medical tissue sealant with biological activity and its preparation method and application

Abstract

The application discloses a medical tissue sealant with biological activity and a preparation method and application thereof, and relates to the field of biomedical technology. The preparation raw material of the medical tissue sealant comprises component A and component B, wherein the component A comprises a polyethylene glycol derivative; and the component B comprises a blood product, and the blood product comprises a blood component and an anticoagulant. The medical tissue sealant can efficiently retain the biological active components inherent in the blood product, such as platelet-derived growth factor and fibrinogen, has the advantages of short gelation time (3-30 seconds), excellent mechanical properties (the compression strength is higher than 800 kPa, the shear strength is higher than 100 kPa, and the anti-burst pressure is higher than 700 mmHg), and moderate degradation time (which can be regulated within 15-30 days), and has a good application prospect in the fields of wound plugging, hemostasis and wound repair.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, and in particular to a bioactive medical tissue sealant, its preparation method, and its application. Background Technology

[0002] Wound repair and tissue regeneration are core directions in biomedicine, focusing on multiple scenarios such as acute trauma, organ defects, and chronic wounds. Through material intervention and tissue engineering, they aim to achieve wound healing and functional reconstruction, addressing critical medical needs such as emergency treatment and surgery. Currently, traditional biomaterials face a core bottleneck of insufficient adaptability to various scenarios. For example, in cases of acute bleeding in solid organs, the adhesion of traditional hemostatic materials easily weakens in a moist environment, making it difficult to withstand arterial pressure; in the repair of hollow organ defects, sealing materials have poor mechanical properties, easily leading to secondary leakage, and lack regenerative induction activity; in the repair of chronic and infected wounds, conventional gels can only moisturize and cannot regulate the inflammatory microenvironment or prolong the healing period. Furthermore, some materials suffer from common problems such as insufficient biocompatibility, mismatched degradation rates, and limited functionality, driving the field towards the research and development of novel biomaterials with multi-functional synergistic effects.

[0003] Among related technologies, blood products are highly valuable bioactive materials in the field of regenerative medicine. Their core advantage stems from their natural abundance of bioactive components such as hemoglobin, platelet-derived growth factor, fibrinogen, and thrombin. These components can form a biomimetic three-dimensional network structure through the dynamic polymerization of a fibrin matrix. This structure can both mimic the physiological hemostasis process to achieve rapid coagulation and sustainably release various cytokines, synergistically regulating key processes such as cell proliferation and matrix repair at the injury site. Based on differences in raw material composition, preparation processes, and clinical application scenarios, blood products can be divided into three main categories: plasma-derived, blood cell-derived, and composite products. Each category relies on the natural bioactivity of blood and forms differentiated application directions targeting different clinical needs. For example, platelet-rich plasma (PRP) is characterized by a platelet concentration 2-10 times that of whole blood. Upon activation, it sustainably releases various growth factors such as PDGF, EGF, and VEGF, while retaining the nutrients and biocompatible matrix of plasma. Therefore, it is widely used clinically in scenarios requiring the promotion of tissue repair and regeneration, such as bone defect repair, tendon and ligament injury healing, granulation tissue induction in chronic wounds (diabetic foot, pressure ulcers), and periodontal regeneration surgery in dentistry. However, current technologies still have some shortcomings. Common blood product loading platforms, such as fibrin glue, have weak mechanical properties, with a viscoelastic modulus typically below 5 kPa, making it difficult to adapt to dynamic wound stress changes and accelerating interfacial adhesion failure. Furthermore, insufficient wound adhesion stability limits long-term effects. More critically, the lack of controlled release of bioactive components leads to an imbalance between angiogenesis and cell migration. These issues have become key bottlenecks restricting clinical translation and efficacy improvement.

[0004] Therefore, there is an urgent need to develop a medical tissue sealant that can solve the defects of blood product-based sealing materials, such as easy deactivation of bioactive components, weak mechanical properties, poor interfacial adhesion, and poor sustained-release effect. Summary of the Invention

[0005] The first aspect of the present invention is to provide a bioactive medical tissue sealant.

[0006] The second objective of this invention is to provide a method for preparing a bioactive medical tissue sealant.

[0007] A third aspect of the present invention is to provide an injection device.

[0008] The fourth aspect of this invention aims to provide the use of a bioactive medical tissue sealant in the preparation of products for wound sealing, hemostasis, and / or wound repair.

[0009] To achieve the above objectives, the technical solution adopted by the present invention is as follows: In a first aspect, the present invention provides a bioactive medical tissue sealant, the raw materials for which are prepared include component A and component B, wherein component A comprises a polyethylene glycol derivative; and component B comprises a blood product. The polyethylene glycol derivative is selected from at least one of the compounds with structural formulas as shown in any one of Formulas 1 to 3; , , ; In the formula, m is taken from a positive integer between 2 and 2300, x is taken from a positive integer between 2 and 4, and R is selected from any group shown in R1 to R3; , , , Where n is a positive integer between 1 and 6.

[0010] The medical tissue sealant according to embodiments of the present invention has at least the following beneficial effects: The sealant of this invention uses blood products (such as platelet-rich plasma and coagulation factor preparations) as its core raw material. By compounding with polyethylene glycol derivatives, it efficiently retains the inherent natural bioactive components of blood products, such as platelet-derived growth factor (PDGF), transforming growth factor-β (TGF-β), hemoglobin, and fibrinogen, while preventing the inactivation of these active factors during preparation. Furthermore, the sealant's compressive strength is adjustable, exceeding 800 kPa, and its shear strength is adjustable, exceeding 100 kPa. This allows it to stably adapt to dynamic changes in the mechanical stress of wounds, making it effective for clinical applications such as acute bleeding control, organ / vascular anastomosis closure, chronic wound healing, and tissue regeneration induction, providing a new solution for complex wound repair.

[0011] In some embodiments of the present invention, the molecular weight of the polyethylene glycol derivative is 2 to 200 kDa. For example, it can be 2, 4, 8, 10, 12, 15, 20, 25, 30, 50, 60, 80, 100, 120, 150, 180 or 200 kDa.

[0012] In some embodiments of the present invention, the blood product comprises blood components and an anticoagulant.

[0013] In some embodiments of the present invention, the blood component is human or animal blood.

[0014] In some embodiments of the present invention, the animal includes at least one of cattle, horses, donkeys, sheep, rabbits, dogs, chickens, rats, and pigs.

[0015] In some embodiments of the present invention, the blood components include one or more of blood cells, plasma, serum, plasma or serum-derived proteins, and their derivatives.

[0016] In some embodiments of the present invention, the blood cells include at least one of red blood cells, white blood cells, and platelets.

[0017] In some embodiments of the present invention, the protein components derived from plasma or serum include at least one of albumin, globulin, fibrinogen, coagulation-related proteins, and immunoglobulins.

[0018] In some embodiments of the present invention, the derivatives include lysates, extracts, enrichments, lyophilized products, detergents, concentrates, etc.

[0019] Specifically, the products may be selected from blood cell products or blood cell derivatives (such as red blood cell products, white blood cell products, platelet products and their lysates, extracts, lyophilized products, detergents, concentrates or enrichments), plasma / serum products or their derivatives (such as fresh plasma, fresh frozen plasma, cryoprecipitate, serum, serum protein preparations, plasma protein preparations, fibrinogen preparations, coagulation factor preparations, immunoglobulin preparations), and blood composite products (such as platelet-rich plasma (PRP), platelet-rich fibrinogen (PRF), concentrated growth factor (CGF), platelet-rich leukocyte plasma, and fibrin-rich clots).

[0020] In some embodiments of the present invention, the anticoagulant includes at least one of heparin, EDTA, and citrate.

[0021] In some embodiments of the present invention, the mass percentage of the anticoagulant in the blood product is 0.001% to 50%. For example, it can be 0.001%, 0.01%, 0.02%, 0.05%, 0.1%, 0.5%, 1%, 2%, 5%, 8%, 10%, 20%, 30%, 40%, or 50%, etc.

[0022] In some embodiments of the present invention, component A includes the polyethylene glycol derivative and buffer A.

[0023] In some embodiments of the present invention, the concentration of the polyethylene glycol derivative in component A is 100-500 mg / mL. In some embodiments of the present invention, component B includes the blood product and buffer B.

[0024] In some embodiments of the present invention, the concentration of the blood product in component B is 100-500 mg / mL.

[0025] Preferably, in component B, the concentration of the blood product is 100-400 mg / mL. For example, it can be 100, 120, 150, 180, 200, 250, 280, 300, 320, 350, 400 mg / mL, etc.

[0026] In some embodiments of the present invention, buffer A and buffer B are independently selected from at least one of phosphate buffer; borate buffer; Tris buffer; sodium carbonate-sodium bicarbonate buffer; N-(2-hydroxyethyl)piperazine-N'-2-ethanesulfonic acid buffer; 2-(N-morpholine)ethanesulfonic acid buffer; 3-(N-morpholine)propanesulfonic acid buffer; piperazine-3-propanesulfonic acid buffer; and N-(tris(hydroxymethyl)-3-aminopropanesulfonic acid buffer.

[0027] In some embodiments of the present invention, the pH values ​​of buffer A and buffer B are independently 6 to 10. For example, the pH values ​​can be 6, 6.5, 7, 7.2, 7.4, 7.5, 7.6, 7.8, 8.0, 8.2, 8.5, 9, 9.5, or 10, etc.

[0028] In some embodiments of the present invention, the volume ratio of component A to component B is 0.05 to 20:1.

[0029] In some embodiments of the present invention, the volume ratio of component A to component B is 0.5 to 10:1. Specifically, the volume ratio of component A to component B can be 0.5:1, 0.8:1, 1:1, 1.2:1, 1.5:1, 2:1, 5:1, 8:1, or 10:1, etc.

[0030] A second aspect of the present invention provides a method for preparing a bioactive medical tissue sealant as described in the first aspect, comprising mixing the components A and B and then subjecting the mixture to a crosslinking reaction.

[0031] A third aspect of the present invention provides an injection device comprising a first syringe and a second syringe, wherein one end of the first syringe is connected to one end of the second syringe; The first syringe contains component A as described in the first aspect; the second syringe contains component B as described in the first aspect.

[0032] A fourth aspect of the invention provides the use of the bioactive medical tissue sealant as described in the first aspect in the preparation of products for wound sealing, hemostasis, and / or wound repair. In some embodiments of the present invention, the wound sealing includes any one of the following: lung air leakage sealing, gastric perforation sealing, intestinal perforation sealing, dura mater sealing, spinal dura mater sealing, tooth extraction wound sealing, and pancreatic fistula sealing.

[0033] In some embodiments of the present invention, the hemostasis includes, but is not limited to, organs, skin, etc. The organs include, but are not limited to, the liver, spleen, heart, kidneys, stomach, intestines, and lungs.

[0034] In some embodiments of the present invention, the wound repair includes, but is not limited to, epidermal tissue wound repair, chronic diabetic wound repair, burn wound repair, or infected wound repair.

[0035] Other features and advantages of the present invention will be set forth in the following description. Attached Figure Description

[0036] The present invention will be further described below with reference to the accompanying drawings and embodiments, wherein: Figure 1 Images showing the sealant prepared in Example 1 of this invention subjected to various external forces such as folding, stretching, twisting, and pressing.

[0037] Figure 2 Images of the sealant prepared in Example 2 of this invention under various external forces such as folding, stretching, and torsion after being adhered to the surface of pigskin.

[0038] Figure 3 This is a diagram illustrating the burst pressure resistance test of the sealant prepared in Example 7 of the present invention.

[0039] Figure 4 The image shows the results of a liver hemostasis experiment on rats using the sealant prepared in Example 9 of this invention.

[0040] Figure 5 The image shows the results of a rat spleen hemostasis experiment using the sealant prepared in Example 4 of this invention.

[0041] Figure 6 The image shows the results of a rat kidney hemostasis experiment using the sealant prepared in Example 14 of this invention.

[0042] Figure 7 The image shows the results of a rat gastric perforation sealing experiment using the sealant prepared in Example 11 of this invention.

[0043] Figure 8 The image shows the results of an intestinal perforation sealing experiment in rats using the sealant prepared in Example 16 of this invention.

[0044] Figure 9 The image shows the results of a rat dura mater occlusion experiment using the sealant prepared in Example 8 of this invention.

[0045] Figure 10 The image shows the results of a rat femoral artery hemostasis experiment using the sealant prepared in Example 10 of this invention.

[0046] Figure 11 The image shows the results of a rat burn wound healing experiment using the sealant prepared in Example 12 of this invention.

[0047] Figure 12 The image shows the results of a rat experiment on the healing of diabetic wounds using the sealant prepared in Example 15 of this invention. Detailed Implementation

[0048] The following will describe the concept and technical effects of the present invention clearly and completely with reference to the embodiments, so as to fully understand the purpose, features and effects of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are all within the scope of protection of the present invention.

[0049] The term "preferred" in this invention refers to embodiments of the invention that provide certain beneficial effects under certain circumstances. However, other embodiments may also be preferred under the same or other circumstances. Furthermore, the description of one or more preferred embodiments does not imply that other embodiments are unavailable, nor is it intended to exclude other embodiments from the scope of the invention.

[0050] When a numerical range is disclosed herein, the range is considered continuous and includes the minimum and maximum values ​​of the range, as well as every value between the minimum and maximum values. Furthermore, when the range refers to integers, it includes every integer between the minimum and maximum values ​​of the range. Additionally, when multiple ranges are provided to describe a feature or characteristic, the ranges may be combined. In other words, unless otherwise specified, all ranges disclosed herein should be understood to include any and all subranges to which they are incorporated.

[0051] In the description of this invention, the reference term "and / or" includes all and any combination of one or more of the associated listed items. For example, A and / or B includes (A and B) and (A or B).

[0052] In an embodiment of the present invention, platelet-rich plasma was purchased from Zhengzhou Pingrui Biotechnology Co., Ltd.

[0053] Unless otherwise specified in the examples, the procedures should be performed under standard conditions or conditions recommended by the manufacturer. Reagents or instruments whose manufacturers are not specified are all commercially available products.

[0054] Example 1 This embodiment provides a bioactive medical tissue sealant, the preparation method of which includes the following steps: (1) Weigh 200 mg of tetra-arm polyethylene glycol succinimide succinate and dissolve it in 1 mL of pH 8.2 PBS buffer solution to prepare solution A with a mass-volume concentration of 200 mg / mL.

[0055] The structural formula of the aforementioned four-arm polyethylene glycol succinimide succinate is shown in Formula 3: , ; In the formula, x=4, m=222, R is R2, n=2, and the molecular weight is approximately 10 kDa.

[0056] (2) Weigh 200 mg of freshly extracted human platelet-rich plasma (anticoagulant is sodium citrate, concentration is 10.9 mmol / L) lyophilized powder, dissolve it in 1 mL of pH 8.2 PBS buffer to prepare solution B with a mass-volume concentration of 200 mg / mL.

[0057] (3) Fill the two solutions into the two chambers of the dual-channel syringe at a volume ratio of 1:1. After injection, inject the mixture into a cylindrical mold, let it stand to form a gel, and then take it out.

[0058] Figure 1 The morphological images of the bioactive medical tissue sealant prepared by this method under folding, stretching, torsion and pressure are shown, indicating that no obvious morphological damage was observed on its surface.

[0059] Example 2-16 Examples 2-16 provide bioactive medical tissue sealants with different component contents, the raw materials for which are prepared are shown in Table 1.

[0060] Table 1:

[0061] The structural formulas of Equations 2 and 3 are as follows: , .

[0062] The structural formulas for R1 to R3 are as follows: , , .

[0063] The preparation methods of Examples 2-16 above are the same as those in Example 1.

[0064] Comparative Example 1 This comparative example provides a medical tissue sealant, the specific preparation method of which is as follows: Weigh 1 mL of platelet-rich plasma, then add 80 μL of 2 wt% CaCl2 and 40 IU of thrombin, mix well, and the product is called platelet-rich plasma hydrogel.

[0065] Comparative Example 2 This comparative example provides a medical tissue sealant, the specific preparation method of which is as follows: Mix 5% ε-polylysine (EPL) and 20% PEG-NHS (purchased from Sinobond) in a 1:1 ratio to obtain the product, denoted as EPL + PEG-NHS.

[0066] Test Example 1: Determination of the gelation time of sealant This example measures the gelation time of the sealants from Examples 1-16 and Comparative Examples 1-2 as follows: First, a blood product solution was drawn using one syringe, and an equal volume of a polyethylene glycol derivative solution was drawn using another syringe. Then, both solutions were simultaneously injected into a transparent sample vial, and gently shaken to mix thoroughly. Next, the sample vial was inverted, and the time required for the liquid to stop flowing was defined as the gelation time. For Comparative Examples 1 and 2, the gelation time was measured after mixing the raw materials. The gelation time results for the sealant are detailed in Table 2.

[0067] Table 2

[0068] The gelation time test results of the sealant showed that the shortest gelation time in the examples was about 3 seconds and the longest was no more than 30 seconds. This indicates that the gelation time of the sealant is relatively rapid but there are certain differences. After selecting the type of blood product according to the needs of clinical treatment, the gelation time of the sealant can also be changed by changing the concentration or the type of polyethylene glycol derivative. It can be flexibly adapted to the needs of different scenarios and provides certain choices for diverse needs.

[0069] Test Example 2: Determination of the Adhesion Strength of Sealants This example demonstrates the adhesive strength test of the sealants used in Examples 1-16 and Comparative Examples 1-2. The specific steps are as follows: First, the blood product solution (solution B) and polyethylene glycol derivative solution (solution A) required in Examples 1-16 were respectively filled into the two channels of a dual-channel syringe. Then, solutions A and B were mixed using the dual-channel syringe and injected onto the surface of a strip of pigskin. Immediately, another strip of pigskin was attached. At room temperature, a 200 g weight was applied to the attachment site for 60 seconds, and then the pigskin was stretched using a universal tensile testing machine at a speed of 10 mm / min until the pigskin adhesive broke. The maximum shear strength was recorded as the shear adhesion strength of the sealant. The adhesion strength results are detailed in Table 3.

[0070] Table 3

[0071] Note: Compared with the comparative example, P<0.05.

[0072] According to the data in Table 3, the tissue adhesion strength of the sealants prepared in Examples 1-16 of this invention is significantly better than that of the comparative example (platelet-rich plasma hydrogel). The tissue adhesion ability of the sealant also changes when conditions such as the type and concentration of blood products and polyethylene glycol derivatives, the type of buffer, and pH are altered. Conversely, when the types of blood products and polyethylene glycol derivatives are fixed, their concentrations can be varied to control tissue adhesion. The adhesion strength, reaching up to 109 kPa, is sufficient to meet the adhesion requirements of various tissue wounds and effectively prevents post-adhesion detachment.

[0073] Figure 2 The adhesion strength test results of the sealant of Example 1 are shown. The sealant prepared in Example 1 was injected onto the surface of pigskin using a dual-channel syringe and left to stand for 60 seconds to stabilize the adhesion. After bending, twisting, stretching and other treatments, the sealant was still able to adhere firmly to the surface of pigskin without falling off; and the strength of the sealant itself kept its structure intact without any defects.

[0074] Test Example 3: Determination of the compressive strength of sealant This example demonstrates the compressive strength determination of the sealants from Examples 1-16 and Comparative Examples 1-2. The specific steps are as follows: First, the blood product solution (solution B) and polyethylene glycol derivative solution (solution A) required in Examples 1-16 were respectively filled into the two channels of a dual-channel syringe. Then, solutions A and B were mixed using the dual-channel syringe and injected into a mold with a diameter of 7 mm and a height of 4 mm to obtain a cylindrical sealant material. Subsequently, the material was compressed using a universal testing machine at a speed of 10 mm / min until the material deformation reached 90%. The maximum compressive strength was recorded as the compressive strength of the sealant. The compressive strength results of the sealants in Examples 1-16 and Comparative Examples 1-2 are detailed in Table 4.

[0075] Table 4

[0076] Note: Compared with the comparative example, P<0.05.

[0077] The results show that the sealant prepared in Examples 1-16 of this invention has a compressive strength of up to about 850 kPa, which can be further controlled by changing the concentration and the type of polyethylene glycol derivative. Its excellent mechanical properties are sufficient to meet the sealing requirements of various wounds. When the wound deforms, the sealant can maintain its structural integrity to maintain adhesion to the wound.

[0078] Test Example 4: Determination of the burst pressure resistance of sealant This example demonstrates the burst pressure resistance of the sealants from Examples 1-16 and Comparative Examples 1-2 as follows: First, take fresh pigskin and clean its surface, then cut it into a circle with a diameter of 5 cm. Use a punch to create a 3 mm diameter hole in the center of the pigskin. Fill the hole with petroleum jelly to prevent sealant from entering, and place a cylindrical mold with a diameter of 1 cm and a height of 5 mm above the hole. Next, fill the two channels of a dual-channel syringe with the blood product solution (solution B) and polyethylene glycol derivative solution (solution A) required in Examples 1-16, respectively. Then, use the dual-channel syringe to mix solutions A and B and inject them into the mold on the hole in the pigskin. After standing for 60 seconds, remove the mold and remove the petroleum jelly from the hole, leaving it hollow. Then, fix the pigskin on a burst pressure testing instrument. Increase the water flow to increase the pressure on the sealant. When the sealant fails to adhere or its structure breaks, causing water leakage and a pressure drop, the maximum pressure before the pressure drop is taken as the burst pressure value of the sealant sample. The burst pressure resistance results of the sealants in Examples 1-16 and Comparative Examples 1-2 are detailed in Table 5.

[0079] Table 5

[0080] Test results show that the sealant of the present invention has a burst pressure resistance of over 600 mmHg, and the sealant of Example 1 has a burst pressure resistance of over 700 mmHg (e.g., Figure 3 As shown in the figure, the blood pressure is much higher than the normal human arterial blood pressure (about 120 mmHg). It can be seen that the sealant material of the present invention can meet the needs of hemostasis for high-pressure arterial bleeding, and has certain application value in the sealing of solid organs (such as lungs and heart).

[0081] Test Example 5: In vitro degradation test of sealant This example demonstrates the in vitro degradation determination of the sealants from Examples 1-16 and Comparative Examples 1-2. The specific steps are as follows: The blood product solution (solution B) and polyethylene glycol derivative solution (solution A) required in Examples 1-16 were respectively filled into the two channels of a dual-channel syringe. Solution A and Solution B were then mixed using the dual-channel syringe and injected into a mold with a diameter of 7 mm and a height of 4 mm to obtain a cylindrical sealant material. Subsequently, the sealant was placed in a centrifuge tube containing at least 10 times its weight of PBS buffer at pH 7.4 and placed in a constant-temperature shaker at 37°C and a shaking speed of 100 r / min. The volume change of the sample in the buffer was observed. The in vitro degradation time of the sealant material was recorded when the sample could not be observed with the naked eye and could not be removed in solid form. The relevant results are detailed in Table 6.

[0082] Table 6

[0083] The results showed that, due to the different types of blood products and polyethylene glycol derivatives selected for the sealant in each embodiment, as well as the different concentrations of buffer solution and solids used to prepare the sealant, the in vitro degradation time ranged from 15 to 30 days. Moreover, the in vitro degradation rate of the sealant accelerated only when the concentrations of blood products and polyethylene glycol derivatives decreased due to the decrease in crosslinking density. Therefore, the degradation time of the sealant can be adjusted according to the needs of various scenarios.

[0084] Test Example 6: In Vitro Release Test of Sealant The application of blood products typically relies on the release of their small molecules. For example, platelet-rich plasma (PRP) continuously releases multiple growth factors such as PDGF, EGF, and VEGF after activation. Coagulation factor preparations selectively enrich single or multiple coagulation factors (such as factors VIII, IX, and XIII) from plasma, exhibiting high purity and stable activity, and can precisely supplement specific coagulation factors lacking in the body. Therefore, this example uses the sealant prepared in Example 1 as an example, with the sealant prepared in Comparative Example 1 serving as a control group, to characterize its in vitro release capability of small molecules.

[0085] The specific method is as follows: Weigh the sealant material prepared in Example 1 or Comparative Example 1, place it in 10 times its weight of pH 7.4 PBS buffer, and incubate it in a constant temperature shaker at 37°C with a shaking speed of 100 r / min. At 1, 2, 4, 8, 12, 24, 48, and 72 hours, aspirate 100 μL of the solution and replenish with an equal volume of pH 7.4 PBS buffer. The concentration of PDGF in the solution is tested using an ELISA kit. The relevant results are detailed in Table 7.

[0086] Table 7

[0087] The results showed that, because the sealant in Example 1 had undergone cell disruption treatment of platelet-rich plasma, the release rate and total amount of PDGF were much higher than those of the platelet-rich plasma hydrogel comparison material which used platelet-rich plasma as a loading material. Moreover, it could release a large amount of growth factors in the early stage of wound healing, regulate cell function and microenvironment at the molecular level, and ultimately accelerate healing, improve healing quality and reduce complications.

[0088] Test Example 7: Hemostasis Test of Sealant in Rat Hearts In this example, the sealant prepared in Example 1 above was used to conduct a cardiac hemostasis experiment in rats. The specific method is as follows: Solutions A and B from Example 1 were mixed at a volume ratio of 1:1 and separately filled into the two chambers of a dual-channel syringe for later use. During the experiment, rats were first anesthetized by endotracheal intubation, and their thoracic position was fixed after the anesthesia took effect. Then, surgical scissors were used to longitudinally incise the skin and subcutaneous tissue along the midline of the sternum, layer by layer, to fully expose the sternum and superficial intrathoracic structures. Next, a rib retractor was used to slowly open the bilateral ribs, fully exposing the cardiac surgical area. Finally, an incision was prepared by puncturing the anterior wall of the left ventricle of the rat's heart with an 18G needle, and blood was observed to spurt out immediately.

[0089] Five seconds after bleeding occurs, the sealant is injected into the wound in situ using a dual-channel syringe, and the bleeding stops immediately.

[0090] Test Example 8: Hemostasis Test of Sealant on Rat Liver In this example, the sealant prepared in Example 9 above was used to conduct a liver hemostasis experiment in rats. The specific method is as follows: Solutions A and B from Example 9 were mixed at a volume ratio of 1:1 and separately filled into the two chambers of a dual-channel syringe for later use. During the experiment, rats were first anesthetized with an anesthetic. After the anesthesia took effect, a small transverse incision was made below the sternum using surgical scissors to expose the liver tissue. After aspirating surrounding blood, tissue fluid, and other fluids, a piece of filter paper was placed under the liver. A 3 mm deep incision was prepared on the liver surface using a 5 mm diameter punch. Liver tissue was removed from the incision using surgical scissors, forming a circular defect.

[0091] like Figure 4 As shown, 5 seconds after bleeding occurred, the sealant was injected into the wound in situ using a dual-channel syringe. The bleeding stopped immediately, and the liver tissue defect was filled with the sealant.

[0092] Test Example 9: Hemostasis Test of Sealant on Rat Spleen In this example, the sealant prepared in Example 4 above was used to conduct a spleen hemostasis experiment in rats. The specific method is as follows: Solutions A and B from Example 4 were mixed at a volume ratio of 1:1 and separately filled into the two chambers of a dual-channel syringe for later use. During the experiment, rats were first anesthetized with an anesthetic. After the anesthesia took effect, a small transverse incision was made below the sternum using surgical scissors to expose the spleen tissue. After aspirating surrounding blood, tissue fluid, and other fluids, a piece of filter paper was placed under the spleen. A 3 mm deep incision was prepared on the surface of the spleen using a 5 mm diameter punch. Liver tissue was removed from the incision using surgical scissors, creating a circular defect.

[0093] like Figure 5As shown, 5 seconds after bleeding occurred, the sealant was injected into the wound in situ using a dual-channel syringe. The bleeding stopped immediately, and the splenic tissue defect was filled with the sealant.

[0094] Test Example 10: Sealant test for sealing air leaks in the lungs of a beagle dog This example uses the sealant prepared in Example 5 above to conduct a lung leak sealing experiment in beagle dogs. The specific method is as follows: Solutions A and B from Example 5 were mixed at a volume ratio of 1:1 and separately filled into the two chambers of a dual-channel syringe for later use. In the experiment, after induction of anesthesia in beagles, intravenous access to the forelimbs was established. The right chest was prepared and disinfected, followed by endotracheal intubation and connection to a respiratory anesthesia machine. Isoflurane was used to maintain anesthesia, and intraoperative ECG monitoring, blood oxygen saturation monitoring, and circulating assistance were performed simultaneously. An incision was made along the right fourth and fifth rib space, and the skin, subcutaneous tissue, and chest wall muscles were dissected layer by layer to expose the pleural cavity. A rib retractor was used to fully expose the target lung lobe. A 1.0 cm full-thickness incision was made on the surface of the lung lobe, and gas accompanied by a small amount of blood was observed continuously leaking from the wound, confirming the successful establishment of the air leakage model.

[0095] The sealant was injected in situ into the wound using a dual-channel syringe to seal it and check for leaks. The results showed that the sealant could quickly close the wound in a short time, and the bleeding and gas leakage from the lung wound stopped immediately.

[0096] Test Example 11: Hemostasis Test of Sealant in Rat Kidneys In this example, the sealant prepared in Example 14 above was used for a kidney hemostasis experiment in rats. The specific method is as follows: Solutions A and B from Example 4 were mixed at a volume ratio of 1:1 and separately filled into the two chambers of a dual-channel syringe for later use. In the experiment, the rat was first fixed on the operating table and anesthetized by intraperitoneal injection. A longitudinal incision of about 1.5-2 cm was made 1.5 cm to the left of the midline of the abdomen with a scalpel, and the skin and subcutaneous connective tissue were cut open. After cutting, the perirenal fat capsule and fascia tissue were gently separated with curved hemostats to gradually expose the left kidney. The exudate on the surface and around the kidney was gently aspirated with sterile gauze soaked in physiological saline to ensure a clear view of the wound. A wedge-shaped incision with a diameter of 3 mm and a depth of 2 mm was precisely made on the surface of the renal cortex with a sterile scalpel. Continuous bleeding and a small amount of renal tissue fluid were immediately observed at the incision site.

[0097] like Figure 6 As shown, 5 seconds after bleeding occurred, the sealant was injected onto the wound surface using a dual-channel syringe. Bleeding stopped immediately, and the sealant evenly covered the entire wound surface and remained adhered.

[0098] Test Example 12: Sealant-based gastric perforation closure experiment in rats This example uses the sealant prepared in Example 11 above to perform a gastric perforation closure experiment in rats. The specific method is as follows: Solutions A and B from Example 11 were mixed at a volume ratio of 1:1 and separately filled into the two chambers of a dual-channel syringe for later use. During the experiment, rats were first anesthetized with an anesthetic. After the anesthesia took effect, a small incision was made in the abdomen using surgical scissors to expose the stomach tissue. Sterile gauze soaked in physiological saline was used to gently absorb the exudate from the stomach surface, ensuring a clear operating field. Next, the anterior wall tissue of the stomach was gently grasped with toothless forceps, and a 5 mm diameter section of tissue was removed with scissors. A small amount of stomach contents were observed to spill out, successfully constructing a gastric perforation model.

[0099] like Figure 7 As shown, a sealant was injected into the gastric defect to seal it, and it was observed that the gastric defect was quickly sealed and the contents no longer leaked.

[0100] Test Example 13: Sealant-induced intestinal perforation closure experiment in rats In this example, the sealant prepared in Example 16 above was used to perform an intestinal perforation closure experiment in rats. The specific method is as follows: Solutions A and B from Example 16 were mixed at a volume ratio of 1:1 and separately filled into the two chambers of a dual-channel syringe for later use. In the experiment, rats were anesthetized with isoflurane using an anesthesia machine, followed by routine disinfection of the abdominal surgical area with iodine. Then, a 1.5-2 cm longitudinal incision was made at the midline of the abdomen with a scalpel, and the skin, subcutaneous fat, and peritoneum were cut layer by layer. The connective tissue and mesentery in the abdomen were gently separated with hemostatic forceps. After cutting, the intestine was gently pulled with toothless forceps to locate the middle section of the small intestine (about 10 cm from the suspensory ligament of the duodenum). A 3-5 mm full-thickness perforation was precisely cut at the opposite side of the small intestinal mesentery with sterile surgical scissors. A small amount of intestinal contents were observed to spill out, and the intestinal perforation model was successfully constructed.

[0101] like Figure 8 As shown, the injected sealant completely covered the wound, and it was observed that the sealant quickly closed the perforation without leakage of intestinal contents.

[0102] Test Example 14: Sealant Occlusion Experiment in Rats In this example, the sealant prepared in Example 13 above was used for a rat dura mater occlusion experiment. The specific method is as follows: Solutions A and B from Example 13 were mixed at a volume ratio of 1:1 and separately filled into the two chambers of a dual-channel syringe for later use. In the experiment, rats were first anesthetized with isoflurane using an anesthesia machine. The skin of the surgical area on the frontal side was then prepared and routinely disinfected with iodine. Next, an incision of about 3 cm was made along the midline of the skull with a scalpel, extending posteriorly to the occipital protuberance. The subcutaneous tissue and galea aponeurotica were gently separated with hemostatic forceps to expose the skull surface. A circular bone defect with a diameter of 5 mm was prepared 2 mm posterior to the coronal suture and 3 mm to the left of the sagittal suture using an electric grinder. During the operation, physiological saline was continuously dripped to cool the bone, and the lower cortical bone was carefully dissected to avoid damaging the underlying dura mater. After the dura mater was fully exposed, a 3 mm long linear defect was precisely cut in the center of the dura mater with microscissors. A small amount of cerebrospinal fluid was observed to slowly seep out, confirming the successful construction of the dural defect model.

[0103] The sealant was injected to completely cover the wound, and it was observed that the sealant quickly closed the defect without any leakage of cerebrospinal fluid.

[0104] Test Example 15: Sealant Occlusion Experiment in Rats In this example, the sealant prepared in Example 8 above was used for a rat dura mater occlusion experiment. The specific method is as follows: Solutions A and B from Example 8 were mixed at a volume ratio of 1:1 and separately filled into the two chambers of a dual-channel syringe for later use. In the experiment, after the rats were anesthetized with isoflurane via an anesthesia machine, they were fixed to the operating table. The skin on their backs was prepared and routinely disinfected three times with povidone-iodine. The skin, subcutaneous fat, and paravertebral muscles of the lumbar spine were sequentially incised along the midline of the back. The muscle attachment points were gently separated using hemostatic forceps to fully expose the spinous processes and lamina. Subsequently, the spinous processes and part of the lamina were removed using single-joint bone shears, forming a bone window of approximately 0.8 cm × 0.5 cm. Bone fragments were carefully removed to expose the underlying dura mater sac. Under direct vision, slight pulsation of the dura mater with respiration was visible. Next, the dura mater was lifted with micro-hooked forceps, and the dura mater and the underlying arachnoid membrane were longitudinally incised with micro-scissors, with an incision length of approximately 1 cm. At this point, clear cerebrospinal fluid continuously flowed out, confirming the successful preparation of the dural defect model.

[0105] like Figure 9 As shown, when sealant was injected into the wound to completely cover it, the dural defect was completely closed and no cerebrospinal fluid leakage was observed, indicating that the sealant effectively sealed the linear incision of the dural tract.

[0106] Test Example 16: Sealing of Tooth Extraction Wounds in Rats with Sealant In this example, the sealant prepared in Example 7 above was used to perform a tooth extraction wound sealing experiment in rats. The specific method is as follows: Solutions A and B from Example 7 were mixed at a volume ratio of 1:1 and separately filled into the two chambers of a dual-channel syringe for later use. In the experiment, after anesthetizing the rats, they were fixed supine on the operating table with their heads slightly elevated to expose the oral cavity. The oral cavity was cleaned with sterile saline, and the left mandibular region was disinfected three times with povidone-iodine solution. Using an oral retractor, the left corner of the mouth was gently opened to fully expose the left mandibular first molar, and the eruption status of the tooth and the absence of redness and inflammation of the surrounding gingiva were observed. Using miniature animal extraction forceps, the angle of the beaks was adjusted to precisely grasp the middle third of the crown of the left mandibular first molar. A moderate biting force was slowly applied for fixation, and the tooth was gently rotated along its long axis and pulled upwards at a uniform speed. After the periodontal ligament separated, the tooth was completely extracted. After extraction, blood accumulated in the extraction socket was gently absorbed with sterile filter paper to fully expose the extraction wound, which was approximately 1.5 mm in diameter. Continuous bleeding from the wound confirmed the successful preparation of the extraction wound model.

[0107] A sealant was injected into the extraction wound, and it was observed that the extraction wound was completely filled, the sealant cured and adhered tightly to the surrounding gingival tissue, and no bleeding or leakage occurred.

[0108] Test Example 17: Hemostasis Experiment of Sealant on Rat Femoral Artery In this example, the sealant prepared in Example 10 above was used to conduct a femoral artery hemostasis experiment in rats. The specific method is as follows: Solutions A and B from Example 10 were mixed at a volume ratio of 1:1 and separately filled into the two chambers of a dual-channel syringe for later use. In the experiment, rats were anesthetized with isoflurane via an anesthesia machine, and fixed supine on the operating table with elastic bands securing the limbs to maintain surgical site stability. Subsequently, the groin area was prepared and routinely disinfected three times with povidone-iodine. A 1.5 cm longitudinal incision was made along the femoral artery, and the skin, subcutaneous fat, and superficial fascia were sequentially cut layer by layer. The connective tissue surrounding the femoral artery was gently separated with hemostatic forceps. After clearly identifying the accompanying femoral vein and femoral nerve, proximal blood flow was temporarily blocked with vascular clamps to control subsequent bleeding. The femoral artery was then completely freed for approximately 1 cm. After the femoral artery was fully freed, the vascular clamps were removed, and it was quickly cut off with microsurgical scissors. Jetting bleeding was observed from the severed end of the femoral artery, successfully establishing a femoral artery hemorrhage model.

[0109] like Figure 10 As shown, after the blood spurting from the wound was suctioned away, a sealant was immediately injected to completely fill the wound. It was observed that the sealant completely sealed the femoral artery defect and adhered tightly to the surrounding tissue, and the bleeding from the femoral artery stopped immediately.

[0110] Test Example 18: Sealant's Effect on Rat Burn Wound Healing This example uses the sealant prepared in Example 12 above to conduct a burn wound healing experiment in rats. The specific method is as follows: Solutions A and B from Example 12 were mixed at a volume ratio of 1:1 and separately filled into the two chambers of a dual-channel syringe for later use. In the experiment, rats were anesthetized and fixed prone on the operating table. Long hair on the back was thoroughly removed using an electric shaver. The hair-removed area was repeatedly rinsed with sterile saline and, after drying, routinely disinfected three times with povidone-iodine. Subsequently, a constant-temperature burn model was created: the instrument was adjusted so that the circular metal head (approximately 1.5 cm in diameter and approximately 2 cm² in area) was heated to 100°C and continuously contacted the disinfected skin area on the rat's back for 10 seconds, maintaining a vertical position and uniform pressure during contact. The wound was visibly pale and hard to the touch, indicating a deep second-degree burn. After the wound has dried, use sterile surgical scissors to cut away the necrotic epidermal tissue along the wound edge to expose the fresh wound. Then, gently rinse the wound with sterile saline to remove exudate and necrotic debris. Subsequently, wound treatment (such as applying therapeutic drugs or covering with repair materials) is performed according to the experimental groups, and the wound healing is observed regularly.

[0111] like Figure 11 As shown, a sealant was injected into the wound until it was completely filled. A control group served as a blank control. Wound images were taken and the wound area was calculated on days 0, 7, 14, and 28 after wound modeling. Analysis of wound healing images from day 0 to day 28 revealed severe inflammation and bacterial pus in the control group, resulting in poor healing. In contrast, after sealant treatment, almost no inflammatory cells were observed, and the wound showed minimal scab formation and scarring over time. Compared to the control group, the sealant group exhibited significantly faster wound closure and higher closure efficiency, reaching 96% after 14 days.

[0112] Test Example 19: Sealant-induced wound healing in diabetic rats This example uses the sealant prepared in Example 15 above to conduct a rat diabetic wound healing experiment. The specific method is as follows: Solutions A and B from Example 15 were mixed at a volume ratio of 1:1 and separately filled into the two chambers of a dual-channel syringe for later use. In the experiment, type 1 diabetic rats were anesthetized and fixed prone on the operating table. Long hair on their backs was thoroughly removed using an electric shaver. The hair-removed area was repeatedly rinsed with sterile saline solution, and after drying, the area was routinely disinfected three times with povidone-iodine. A circular metal tip (approximately 1.5 cm in diameter and 2 cm² in area) was used to mark the skin on the rat's back. Sterile surgical scissors were used to cut away the epidermal tissue along the mark to expose the fresh wound. The wound was then gently rinsed with sterile saline solution to remove exudate. Subsequently, wound treatment (such as applying therapeutic drugs or covering with repair materials) was performed according to the experimental groups, and the wound healing was observed regularly.

[0113] like Figure 12As shown, a sealant was injected into the wound until it was completely filled, with a control group serving as a blank control. Wound images were taken on days 0, 4, 7, and 14 after wound modeling, and the wound area was calculated. Analysis of wound healing images from day 0 to day 28 showed a decrease in wound area over time in all groups. Calculation of the healed area further quantified the wound healing rate. Compared to the control group, the sealant group showed a significantly faster wound closure rate and higher closure efficiency, reaching 98% after 14 days, indicating basic wound healing.

[0114] In summary, this invention provides a bioactive medical tissue sealant, its preparation method, and its applications. This sealant is formed by a cross-linking reaction between polyethylene glycol derivatives and blood products (such as platelet-rich plasma and coagulation factor preparations) at room temperature or under physiological conditions, creating a stable three-dimensional gel framework structure. This structure effectively retains the inherent bioactive components of blood products, such as platelet-derived growth factor and fibrinogen. It boasts advantages such as short gelation time (3-30 seconds), excellent mechanical properties (compressive strength exceeding 800 kPa, shear strength exceeding 100 kPa, and burst pressure exceeding 700 mmHg), and moderate degradation time (adjustable within 15-30 days). Simultaneously, it enables long-term, controllable release of bioactive factors. Furthermore, this sealant can be used in various clinical scenarios, including acute bleeding control, closure of solid organ and vascular injuries, repair of chronic wounds (such as diabetic ulcers and burns), and tissue regeneration. It effectively overcomes the shortcomings of traditional blood product-based materials, such as weak mechanical properties, poor adhesion, and easy inactivation of active factors, demonstrating promising application prospects.

[0115] The embodiments of the present invention have been described in detail above. However, the present invention is not limited to the above embodiments. Within the scope of knowledge possessed by those skilled in the art, various changes can be made without departing from the spirit of the present invention. Furthermore, the embodiments of the present invention and the features thereof can be combined with each other unless otherwise specified.

Claims

1. A bioactive medical tissue sealant, characterized in that, The raw materials for preparation include component A and component B, wherein component A contains a polyethylene glycol derivative; and component B contains blood products. The polyethylene glycol derivative is selected from at least one of the compounds with structural formulas as shown in any one of Formulas 1 to 3; 、 、 ； In the formula, m is taken from a positive integer between 2 and 2300, x is taken from a positive integer between 2 and 4, and R is selected from any group shown in R1 to R3; 、 、 ， Where n is a positive integer between 1 and 6.

2. The medical tissue sealant according to claim 1, characterized in that, The blood products contain blood components and anticoagulants.

3. The medical tissue sealant according to claim 2, characterized in that, The blood components include any one or more of blood cells, plasma, serum, plasma or serum-derived proteins, and their derivatives; And / or, the anticoagulant includes at least one of heparin, EDTA, and citrate.

4. The medical tissue sealant according to claim 1 or 2, characterized in that, Component A includes the polyethylene glycol derivative and buffer A; And / or, the B component includes the blood product and buffer B.

5. The medical tissue sealant according to claim 4, characterized in that, In component A, the concentration of the polyethylene glycol derivative is 100~500 mg / mL; And / or, in component B, the concentration of the blood product is 100~500 mg / mL; And / or, buffer A and buffer B are independently selected from at least one of phosphate buffer; borate buffer; Tris buffer; sodium carbonate-sodium bicarbonate buffer; N-(2-hydroxyethyl)piperazine-N'-2-ethanesulfonic acid buffer; 2-(N-morpholine)ethanesulfonic acid buffer; 3-(N-morpholine)propanesulfonic acid buffer; piperazine-3-propanesulfonic acid buffer; and N-(tris(hydroxymethyl)-3-aminopropanesulfonic acid buffer.

6. The medical tissue sealant according to claim 5, characterized in that, The volume ratio of component A to component B is 0.05 to 20:

1.

7. A method for preparing a bioactive medical tissue sealant as described in any one of claims 1 to 6, characterized in that, This includes mixing component A and component B, followed by a cross-linking reaction to obtain the final product.

8. An injection device, characterized in that, It includes a first syringe and a second syringe, with one end of the first syringe connected to one end of the second syringe; The first syringe contains component A as described in any one of claims 1 to 6; the second syringe contains component B as described in any one of claims 1 to 6.

9. The use of the bioactive medical tissue sealant as described in any one of claims 1 to 6 in the preparation of products for wound sealing, hemostasis, and / or wound repair.

10. The application according to claim 9, characterized in that, The wound sealing includes any one of the following: lung air leakage sealing, gastric perforation sealing, intestinal perforation sealing, dura mater sealing, spinal dura mater sealing, and tooth extraction wound sealing.

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