Large-area wound first-aid dressing kit and use method thereof

Abstract

The invention belongs to the field of biological material preparation and biomedical application, and particularly relates to a large-area wound first-aid dressing kit and a use method thereof. Comprising a first container filled with a first ingredient containing dodecylated chitosan; the second container is filled with a second ingredient containing silk fibroin and catechol; the mass ratio of the dodecylated chitosan to the silk fibroin to the catechol is (5-15): 1: (5-15). The dodecylated chitosan, the silk fibroin and the catechol which are respectively and independently packaged are mixed during use, rapid hemostasis can be realized, and rapid hemostasis of large-area bleeding within 3 seconds can be realized according to a preferable scheme (refer to an embodiment 14).

Description

Technical Field

[0001] This invention belongs to the field of biomaterial preparation and biomedical applications, specifically relating to a large-area wound emergency dressing kit and its usage method. Background Technology

[0002] For many years, the treatment of acute and severe bleeding caused by large-area trauma or ruptured major arteries in scenarios such as battlefields, traffic accidents, and surgical procedures has always been a huge challenge for clinical medicine. This type of bleeding is rapid and involves a large amount of blood loss, and the injured person can die from hemorrhagic shock in a very short time. Therefore, the "golden treatment time" is often only a few minutes, which places extremely high demands on the immediacy and reliability of hemostatic materials.

[0003] Currently, the main emergency hemostatic materials on the market include zeolite-based, collagen-based, chitosan-based, fibrin adhesives, and α-cyanoacrylate tissue glues. However, these materials all have significant limitations in dealing with the aforementioned extreme situations:

[0004] 1. Traditional materials have poor adhesion to wet wounds: Zeolite releases heat when absorbing blood, which may cause tissue burns; materials such as collagen and alginate are easily washed away or dissolved under the impact of surging blood flow, and cannot form an effective seal on wet wounds.

[0005] 2. Insufficient response speed of pre-formed dressings: Many hemostatic dressings based on chitosan and collagen are made into sponges or pads through a pre-crosslinking and freeze-drying process. These materials need to absorb liquid and swell before they can exert their effects, and their hemostatic process is relatively slow, making it difficult to achieve rapid physical closure within seconds. This results in insufficient "first-wave" suppression for major arterial bleeding.

[0006] 3. Difficulty in achieving both strength and speed in in-situ gel formation: Although some techniques employ the idea of ​​forming hydrogels by mixing two components, most of them focus on wound filling or long-term healing rather than emergency hemostasis. These gels often have excessively long gelation times (tens of seconds to several minutes), or even if they form gels quickly, their mechanical strength is weak and their cohesion is insufficient, making them prone to detachment on wet, dynamic wound surfaces and failing to form a strong physical barrier.

[0007] 4. Insufficient biocompatibility and functionality: Fibrin adhesives pose a risk of viral contamination and are expensive; cyanoacrylate tissue adhesives may have cytotoxic degradation products and are extremely sensitive to moist environments, with adhesion decreasing sharply on bloody wounds.

[0008] In summary, existing technologies lack an emergency hemostatic product capable of instantly forming an artificial sealing layer with superior mechanical strength and adhesion within seconds on large-area, wet, and dynamically bleeding wounds, thereby achieving instantaneous physical closure and hemostasis, while also possessing good biocompatibility. This is precisely the core technological bottleneck that this invention aims to address. Summary of the Invention

[0009] The technical problem solved by this invention is to provide a novel large-area wound emergency dressing kit and its method of use. It uses separately packaged dodecyl chitosan, silk fibroin and catechin, which are mixed together during use to achieve rapid hemostasis. The preferred solution can achieve rapid hemostasis of large-area bleeding within 3 seconds (see Example 14).

[0010] To achieve the above-mentioned objectives, the present invention adopts the following technical solution:

[0011] In a first aspect, the present invention provides a large-area trauma emergency dressing kit, comprising:

[0012] The first container contains a first ingredient comprising dodecylated chitosan;

[0013] The second container contains a second ingredient including silk fibroin and catechins;

[0014] The mass ratio of the dodecyl chitosan, silk fibroin and catechin is 5-15:1:1-15.

[0015] The dressing of this invention can achieve hemostasis of major arteries within 3 seconds, which is a significant breakthrough in the field. The applicant has published relevant literature and won a gold medal in a project in this field, demonstrating the groundbreaking hemostatic effect of this invention from multiple perspectives.

[0016] Optionally, after the first and second ingredients are mixed on a wet wound, a hydrogel can be formed at the wound within 10 seconds, and the hemostasis time is ≤15 seconds.

[0017] Optionally, the tensile strength of the hydrogel is not less than 0.9 MPa.

[0018] Optionally, the mass ratio of the dodecyl chitosan, silk fibroin, and catechin is 10-15:1:10-15.

[0019] Optionally, the method for preparing the dodecylated chitosan includes the following steps:

[0020] Chitosan was dissolved in acetic acid solution, lauraldehyde and sodium dodecyl sulfonate phase transfer catalyst were added to react and generate Schiff base. The solution was adjusted to 4-5 with NaOH solution, and then NaBH4 solution was added dropwise. After heating and stirring to carry out the reduction reaction, lauraldehyde and NaBH4 were added again to carry out a secondary reduction reaction. The chitosan was then purified by utilizing the difference in solubility of chitosan with different degrees of alkylation in chloroform solvent.

[0021] Optionally, it also contains silver alginate, wherein the mass ratio of silver alginate to modified chitosan is 1:1 to 5.

[0022] Secondly, the present invention also provides a method for using the large-area trauma emergency dressing kit in any of the above-mentioned solutions, comprising the following steps:

[0023] Remove the first ingredient from the first container;

[0024] Remove the second ingredient from the second container;

[0025] Mix the first and second ingredients at the wound site at a mixing speed of 0.01-1 L / min to form a hydrogel dressing; the hemostasis time after mixing is ≤15 seconds.

[0026] Optionally, the mixing is carried out in a humid environment, which includes at least one of water, PBS buffer, or blood.

[0027] (III) Beneficial Effects

[0028] Compared with the prior art, the beneficial effects of the present invention are:

[0029] 1. The large-area trauma first aid dressing kit of the present invention, through its independent packaging, instant mixing design and ultra-fast cross-linking reaction between specific components, can form a hydrogel sealing layer at the wound within 10 seconds, achieving second-level physical closure of large-area trauma and major arterial bleeding (hemostasis time ≤15 seconds), or even hemostasis within 3 seconds.

[0030] 2. The present invention forms a double-network hydrogel with a specific ratio of dodecyl chitosan, silk fibroin and catechin, which has excellent tensile strength (up to 1.57 MPa) and adhesion strength, can firmly adhere to wet and dynamic wound surfaces, effectively resist blood flow erosion and is not easy to fall off.

[0031] 3. The specific combinations of the present invention (especially the use of catechins) produce a significant synergistic effect. Replacing catechins with other polyphenols (such as rhein) leads to a sharp decrease in hydrogel strength and a rapid decline in hemostatic effect, demonstrating that the component selection of the present invention is not obvious and the effects are unpredictable.

[0032] 4. This invention is provided in kit form, is stable in storage, and is easy to operate, making it very suitable for use by medical staff or even patients in emergency situations. Attached Figure Description

[0033] Figure 1 The results of the cytotoxicity test of the dressing obtained in Example 7 of this invention;

[0034] Figure 2 The results of the in vitro hemostasis test of the dressing experimental group obtained in Example 7 of the present invention;

[0035] Figure 3 The results of the in vitro hemostasis test of the dressing control group obtained in Example 7 of the present invention are shown. Detailed Implementation

[0036] To further understand the present invention, preferred embodiments of the present invention are described below in conjunction with examples. However, it should be understood that these descriptions are only for further illustrating the features and advantages of the present invention, and not for limiting the scope of the claims of the present invention.

[0037]

Implementation Method 1

[0038] This invention provides a large-area wound first aid dressing, which comprises the following components: modified chitosan, biomass protein fiber and polyphenol compound.

[0039] In this invention, modified chitosan, biomass protein fiber, and polyphenolic compounds with good biocompatibility are selected as the main raw materials. Through self-assembly and synergistic compounding, a novel hemostatic material that can achieve rapid adhesion and antibacterial properties in wet environments (including water, PBS buffer, and blood environment) is obtained.

[0040] Modified chitosan, such as dodecyl chitosan, is a natural mucopolysaccharide with good biocompatibility. Its cationic effect and three-dimensional molecular network structure in blood enable it to rapidly capture and activate erythrocytes and platelets to form blood clots. Furthermore, segments of modified chitosan molecules, such as dodecyl chitosan, can target the phospholipid bilayer of bacterial cell membranes, thereby enhancing its anti-inflammatory and bactericidal effects through the addition of polyphenolic compounds or other bactericidal substances, ultimately killing bacteria and achieving bactericidal function.

[0041] The self-assembly between polyphenolic compounds and natural biomass protein fiber (such as silk fibroin) molecules, as well as the chelation between modified chitosan and polyphenolic polymers, form a double network, thereby achieving rapid hemostasis.

[0042] This invention, on the one hand, leverages the function of modified chitosan in aggregating erythrocytes, promoting coagulation, and the ability of modified chitosan segments to target the phospholipid bilayer of bacterial cell membranes, thereby killing bacteria through bactericidal substances. On the other hand, it utilizes the superior wet tissue adhesion capabilities of functionalized biomass protein fibers to achieve rapid closure of injured tissues. The resulting adhesive exhibits high hemostatic efficiency in moist environments, rapid adhesion, wide applicability, strong targeted antibacterial properties, safety, non-toxicity, good biocompatibility, and ease of operation, providing a widely applicable solution for modern clinical medicine to overcome aortic bleeding.

[0043] Experiments have shown that:

[0044] 1. The dressing in this invention achieves hemostasis for large-area wounds within 15 seconds;

[0045] 2. The dressing of the present invention has a significantly shorter hemostasis time under the same hemostatic conditions than hemostatic materials containing modified chitosan and biomass protein fibers but not polyphenolic compounds.

[0046] 3. The dressing of the present invention has a significantly shorter hemostasis time under the same hemostatic conditions than hemostatic materials containing polyphenolic compounds and modified chitosan but not biomass protein fibers.

[0047] 4. The dressing of the present invention has a significantly shorter hemostasis time under the same hemostatic conditions than hemostatic materials containing polyphenolic compounds and biomass protein fibers but not modified chitosan.

[0048] The preferred weight ratios of the components in this invention are as follows: modified chitosan, biomass protein fiber, and polyphenolic compound are in the following mass ratios:

[0049] 1~1000∶1~1000∶1~1000;

[0050] The preferred composition is: modified chitosan, biomass protein fiber and polyphenol compound in a mass ratio of 1-15:1-15:1-15;

[0051] The preferred composition is: modified chitosan, biomass protein fiber, and polyphenolic compound in a mass ratio of 3-6:1-10:1-15;

[0052] More preferably, the modified chitosan, biomass protein fiber and polyphenol compound are in a mass ratio of 15:1:15.

[0053] The bio-protein fiber in this invention can be one or a combination of two or more of silk fibroin, keratin, collagen, and marine mussel protein; preferably silk fibroin.

[0054] The modified chitosan in this invention can be one or a combination of two or more of dodecyl chitosan, quaternary ammonium salt chitosan and carboxylated chitosan; preferably dodecyl chitosan.

[0055] The polyphenolic compounds in this invention are one or more of phenolic hydroxyl compounds, tannins, phenolic acids, and anthocyanins; preferably, they are one or more of flavonoids, tannic acid, gallic acid, dopamine, and catechols.

[0056] Using marine-derived dodecyl chitosan, natural high-molecular-weight silk fibroin, natural polyphenolic compounds from plants, and alginate extracted from marine brown algae as the main raw materials, a new hemostatic material with antibacterial properties is obtained through self-assembly and synergistic compounding, which can achieve rapid adhesion in wet environments (including water, PBS buffer, and blood environment).

[0057] Dodecyl chitosan, a natural mucopolysaccharide, possesses excellent biocompatibility. Its cationic effect and three-dimensional molecular network structure in blood enable it to rapidly capture and activate erythrocytes and platelets to form blood clots. Furthermore, segments of the dodecyl chitosan molecule can localize to the phospholipid bilayer of bacterial cell membranes, targeting bacteria and further killing them through silver ions, thus achieving a bactericidal function.

[0058] To improve the bactericidal and analgesic functions of the large-area wound first aid dressing of the present invention, the dressing also includes one or more of the following components: silver-containing compounds, antimicrobial peptides, aspirin, curcumin, celecoxib, or analgesics.

[0059] Preferably, it contains silver compounds, more preferably silver ion compounds, and even more preferably silver alginate; during the process of promoting coagulation, the modified chitosan molecules can be localized to the phospholipid bilayer of the bacterial cell membrane, targeting bacteria, and can kill bacteria through silver ions in silver alginate, and alginate ions participate in the formation of the double network, thereby improving hemostatic function.

[0060] To facilitate dressing use, the large-area wound emergency dressing includes a first ingredient and a second ingredient. The first ingredient is modified chitosan, and the second ingredient is a mixture of biomass protein fibers and polyphenolic compounds. During use, the first and second ingredients are mixed to form the dressing, which achieves hemostasis in a moist environment. The first and second ingredients must be stored separately before use.

[0061]

Implementation Method Two

[0062] This embodiment provides a method for preparing a large-area wound first aid dressing, which includes the following steps: adding a biomass protein fiber solution and a polyphenol compound solution to a modified chitosan solution and mixing them thoroughly to obtain the dressing.

[0063] In the modified chitosan solution, it is preferable to add the biomass protein fiber solution and the polyphenol compound solution in sequence. After adding the polyphenol compound solution, silver alginate can also be added. Using this addition method, the dressing has a better and more stable network structure.

[0064] The mixing speed is 0.01–1 L / min, and stirring is carried out during the mixing process.

[0065] In the modified chitosan solution, the concentration of modified chitosan is 0.001–10 g / mL; the modified chitosan solution is preferably a dodecylated chitosan solution, and its preparation method is as follows:

[0066] Chitosan was dissolved in acetic acid solution, lauraldehyde and sodium dodecyl sulfonate (a phase transfer catalyst) were added to react and generate Schiff base. The solution was then adjusted to a concentration of 4-5 with NaOH solution.

[0067] After adding NaBH4 solution and heating and stirring to carry out the reduction reaction, lauraldehyde and NaBH4 were added again for a second reduction reaction. The chitosan was purified by taking advantage of the difference in solubility of chitosan with different degrees of alkylation in chloroform solvent. Specifically, the product precipitated by the second reduction was washed with distilled water until neutral, excess lauraldehyde and the reduction product lauryl alcohol were removed with acetone, and then purified with a Soxhlet extractor. The precipitate was dissolved in chloroform, acetone was added to obtain the precipitate, and the dodecyl chitosan was obtained by filtration. The obtained dodecyl chitosan was dissolved in aluminum hexachloride solution and stirred at a constant temperature of 10-60℃ for 0.5-6h to obtain a dodecyl chitosan solution.

[0068] In the biomass protein fiber solution, the concentration of biomass protein fiber is 0.001–10 g / mL;

[0069] The preferred biomass protein fiber solution is a silk fibroin solution, which is prepared as follows: deionized water is heated to boiling, and Na2CO3 is added to degumm the silk. After drying, the silk is dissolved in lithium bromide solution, calcium chloride, anhydrous ethanol or a ternary solution of water, and then dialyzed with deionized water. The dialyzed solution is centrifuged and filtered to obtain the silk fibroin solution.

[0070] The concentration of polyphenol compounds in the polyphenol compound solution is 0.001–10 g / mL;

[0071] Among them, silver alginate is mainly prepared by mixing a soluble silver salt solution with a concentration of 0.001-10 g / mL and an alginate solution with a concentration of 0.001-10 g / mL.

[0072] To better understand the above technical solutions, exemplary embodiments of the present invention will be described in more detail below with reference to the accompanying drawings. Although exemplary embodiments of the present invention are shown in the drawings, it should be understood that the present invention can be implemented in various forms and should not be limited to the embodiments set forth herein. Rather, these embodiments are provided so that the present invention can be understood more clearly and thoroughly, and that the scope of the present invention can be fully conveyed to those skilled in the art.

[0073] Example 1

[0074] This embodiment provides a method for preparing a dodecylated chitosan solution, the steps of which are as follows:

[0075] 4.0 g of chitosan was weighed into 120 mL of 5% acetic acid solution and stirred until completely dissolved. 12.0 g of lauraldehyde and 0.5 g of sodium dodecyl sulfonate, a phase transfer catalyst, were added to the solution, and the mixture was heated and stirred at 60 °C for 6 hours. After the reaction was complete, the pH was adjusted to 4.5 with 1 M NaOH solution, and then 1.5 times the excess of 10% NaBH4 solution was slowly added dropwise for reduction. The mixture was stirred at 60 °C for another 2.5 hours. Subsequently, 6.0 g of lauraldehyde and an equal proportion of NaBH4 solution were added for a second reduction reaction for 2 hours, yielding a precipitate. The precipitate was washed with distilled water until neutral, and then washed three times with acetone to remove excess lauraldehyde and its reduction product, lauryl alcohol. The precipitate was then purified using a Soxhlet extractor with chloroform as the solvent for 48 hours. The purified product was dissolved in chloroform, filtered to remove insoluble matter, and acetone was added to the filtrate to obtain a white precipitate. This precipitate was filtered and dried to obtain dodecylated chitosan solid.

[0076] Dissolve 15g of aluminum hexachloride in water, add 15g of the prepared dodecyl chitosan, heat to 40℃ and stir for 6 hours to obtain a homogeneous and clear dodecyl chitosan solution with a concentration of 40%, and store it in a sealed container away from light.

[0077] Example 2: Preparation of silk fibroin solution

[0078] Heat 5L of deionized water to boiling, add 8g of anhydrous sodium carbonate (Na2CO3) and dissolve thoroughly. Add 7g of raw silkworm silk to the solution and degumme at 100℃ for 30 minutes to remove sericin. After treatment, wash the degummed silk fibers thoroughly with deionized water until the washing solution is neutral, then dry in an 80℃ oven.

[0079] The dried, degummed silk fibroin fibers were placed in a 9.3M lithium bromide (LiBr) solution and stirred at 70°C for 5 hours to dissolve, yielding an orange-yellow silk fibroin-LiBr solution. This solution was transferred to a dialysis bag with a molecular weight cutoff of 12,000-14,000 and dialyzed with deionized water for 72 hours, changing the water every 8 hours to completely remove LiBr. After dialysis, the solution was centrifuged at 4°C and 10,000 rpm for 20 minutes. The supernatant was collected and filtered through a 0.22 μm microporous membrane to obtain a pure silk fibroin solution with a mass fraction of approximately 10% w / w.

[0080] Example 3: Preparation of Catechol Solution

[0081] Weigh 15g of catechin powder and dissolve it in 15g of deionized water. Stir magnetically for 30 minutes at room temperature until completely dissolved to obtain a 50% w / w catechin solution.

[0082] Example 4

[0083] Preparation of solid silver alginate: Mix 15g of silver nitrate and 15g of sodium alginate solution at room temperature, and dry the water-insoluble product to obtain solid silver alginate.

[0084] Example 5

[0085] The dodecyl alkylated chitosan solution obtained in Example 1, the silk fibroin solution obtained in Example 2, and the catechol solution obtained in Example 3 were prepared at a mass ratio of 1:5:1. The silk fibroin solution and the catechol solution were added to the dodecyl alkylated chitosan solution in sequence and mixed thoroughly to obtain a large-area wound emergency dressing with antibacterial effect that can achieve rapid hemostasis of aortas.

[0086] The weight ratio of the 12-alkylated chitosan solution, silk fibroin solution, and catechin solution after conversion to dry weight at a mass ratio of 1:5:1 is 12:1:1.

[0087] Example 6

[0088] This embodiment provides a method for preparing and using a large-area trauma first aid dressing kit, the steps of which are as follows:

[0089] S1 Take 2 mL of the dodecyl chitosan solution (first ingredient) prepared in Example 1, fill it into a 2.5 mL medical syringe (first container), and seal it.

[0090] S2. The silk fibroin solution prepared in Example 2 and the catechin solution prepared in Example 3 are mixed at a mass ratio of 1:1 and gently shaken to form a second mixture.

[0091] S3 takes the above-mentioned second ingredient mixture, fills it into another medical syringe (second container), and seals it.

[0092] S4 comprises two syringes, a dual-syringe connector, and a mixing nozzle packaged together in a sterile packaging bag, thus constituting the large-area wound first aid dressing kit of the present invention. The dodecyl alkylated chitosan solution, silk fibroin solution, and catechol solution in the large-area wound first aid dressing kit are in a mass ratio of 1:5:1.

[0093] The weight ratio of the 12-alkylated chitosan solution, silk fibroin solution, and catechin solution after conversion to dry weight at a mass ratio of 1:5:1 is 12:1:1.

[0094] The method of using the large-area trauma emergency dressing kit in this embodiment is as follows:

[0095] When using, take out the kit, install the two syringes on both sides of the dual syringe connector, and install the mixing nozzle at the end.

[0096] After performing simple debridement on a large-area bleeding wound, aim the mixing nozzle at the wound and extrude the first and second ingredients simultaneously at a propulsion speed of approximately 0.5 L / min.

[0097] The two components are initially mixed as they flow through the mixing nozzle, and then rapidly cross-link upon contact with the wet wound (filled with blood) (<10 seconds), forming a hydrogel sealing layer that firmly covers the wound.

[0098] This hydrogel sealing layer can completely stop arterial blood from gushing out within 8 seconds, achieving instantaneous physical hemostasis. Tests have shown that the resulting hydrogel has a tensile strength exceeding 0.97 MPa, exhibits strong adhesion to wet tissues, and is not easily detached by blood flow.

[0099] Example 7

[0100] This embodiment provides a method for preparing and using a large-area trauma first aid dressing kit, the steps of which are as follows:

[0101] Steps S1 and S2 are the same as in Example 6;

[0102] S3 takes the above-mentioned second ingredient mixture, fills it into another medical syringe (second container), and seals it.

[0103] S4 grinds the silver alginate solid into powder and fills it into a specially designed 2.5mL vial (third container) with an inner stopper, then seals it.

[0104] The large-area trauma emergency dressing kit contains 12-alkylated chitosan solution, silk fibroin solution, catechol solution, and silver alginate solid in a mass ratio of 1:5:1:1.

[0105] S5. The first container (syringe), the second container (syringe), the third container (vial), a double syringe connector and a mixing nozzle are packaged together in a sterile, light-proof aluminum foil bag, which constitutes the trauma first aid dressing kit of the present invention.

[0106] The method of using the large-area trauma emergency dressing kit in this embodiment is as follows:

[0107] S1 Open the aseptic packaging and remove the kit.

[0108] Remove the needle from the second container (the syringe containing the second ingredient mixture) and align its outlet with the inner stopper of the third container (vial). Forcefully push all the second ingredient mixture (silk fibroin + catechins) from the syringe into the vial. Vigorously shake the vial for 10-15 seconds to fully dissolve and disperse the silver alginate solid powder in the vial into the second ingredient mixture, forming a homogeneous, enhanced second ingredient solution containing the three active ingredients (silk fibroin, catechins, and silver alginate).

[0109] S2 kit assembly

[0110] Open the vial cap and replace it with a dedicated double-ended extraction needle. Pull out the plunger of the first container (the syringe containing the dodecyl chitosan solution) and connect its barrel to one end of the double-ended extraction needle. By pulling the plunger, draw all the fortified second ingredient solution from the vial into the syringe barrel. Replace the plunger of the first container syringe; at this point, the syringe contains both the first ingredient and the fortified second ingredient solution, but they are not yet mixed (or only slightly mixed). Connect the first container syringe to one end of the dual-syringe connector. The other end of the connector can be connected to an empty syringe barrel to provide injection force; the mixing nozzle is attached to the other end.

[0111] After performing simple debridement on a large-area bleeding wound, the S3 nozzle is aimed at the wound. The syringe piston of the first container is pushed, expelling the mixture from the cylinder at a rate of 0.5 L / min. As the mixture flows through the static mixer of the nozzle, dodecyl chitosan is thoroughly mixed with the solution containing silk fibroin, catechol, and silver alginate. This mixture rapidly cross-links upon contact with the moist wound (<10 seconds), forming a firmly sealing hydrogel layer that covers the wound.

[0112] This hydrogel sealing layer can completely stop arterial blood from gushing out within 8 seconds, achieving instant hemostasis.

[0113] The dressing in this embodiment provides immediate physical closure: through the ultra-fast cross-linking of the chitosan-silk fibroin-catechin system, it achieves hemostasis within seconds.

[0114] Synergistic antibacterial effect: After the hydrogel is formed, dodecyl chitosan targets and destroys the bacterial cell membrane, while silver alginate continuously releases silver ions. The two work synergistically to achieve highly effective protection of the wound.

[0115] Stable structure: The hydrogel formed by this process has a tensile strength of up to 0.97 MPa, strong adhesion to wet tissues, and is not easily detached by blood flow.

[0116] In this embodiment, the dry weight ratio of dodecyl chitosan, silk fibroin, catechin, and silver alginate is 12:1:1:1.

[0117] Example 8

[0118] The rest is the same as in Example 7, except that in the dressing kit, the dodecyl alkylated chitosan solution, silk fibroin solution, catechol solution and silver alginate solid are prepared in a mass ratio of 1:15:1:1.

[0119] In this embodiment, the dry weight ratio of dodecyl chitosan, silk fibroin, catechin, and silver alginate is 12:15:1:1.

[0120] Example 9

[0121] The rest is the same as in Example 7, except that in the dressing kit, the dodecyl alkylated chitosan solution, silk fibroin solution, catechol solution and silver alginate solid are prepared in a mass ratio of 1:1:5:1.

[0122] In this embodiment, the dry weight ratio of dodecyl chitosan, silk fibroin, catechin, and silver alginate as active ingredients is 12:1:5:1.

[0123] Example 10

[0124] The rest is the same as in Example 7, except that in the dressing kit, the dodecyl alkylated chitosan solution, silk fibroin solution, catechol solution and silver alginate solid are prepared in a mass ratio of 1:1:15:1.

[0125] In this embodiment, the dry weight ratio of dodecyl chitosan, silk fibroin, catechin, and silver alginate as active ingredients is 2:1:15:1.

[0126] Example 11

[0127] The rest is the same as in Example 7, except that in the dressing kit, the dodecyl alkylated chitosan solution, silk fibroin solution, catechol solution and silver alginate solid are prepared in a mass ratio of 1:1:1:5.

[0128] In this embodiment, the dry weight ratio of dodecyl chitosan, silk fibroin, catechin, and silver alginate as active ingredients is 12:1:1:5.

[0129] Example 12

[0130] The rest is the same as in Example 7, except that in the dressing kit, the dodecyl alkylated chitosan solution, silk fibroin solution, catechol solution and silver alginate solid are prepared in a mass ratio of 1:1:1:15.

[0131] In this embodiment, the dry weight ratio of dodecyl chitosan, silk fibroin, catechin, and silver alginate as active ingredients is 12:1:1:15.

[0132] Example 13

[0133] The rest is the same as in Example 7, except that in the dressing kit, the dodecyl alkylated chitosan solution, silk fibroin solution, catechol solution and silver alginate solid are prepared in a mass ratio of 5:1:1:1.

[0134] In this embodiment, the dry weight ratio of dodecyl chitosan, silk fibroin, catechin, and silver alginate as active ingredients is 5:1:1:1.

[0135] Example 14

[0136] The rest is the same as in Example 7, except that in the dressing kit, the dodecyl alkylated chitosan solution, silk fibroin solution, catechol solution and silver alginate solid are prepared in a mass ratio of 15:1:1:1.

[0137] In this embodiment, the dry weight ratio of dodecyl chitosan, silk fibroin, catechin, and silver alginate as active ingredients is 15:1:1:1.

[0138] Comparative Example 1

[0139] The rest is the same as in Example 7, except that in step S5, the catechin solution and silver alginate solid are added sequentially to the dodecyl alkylated chitosan solution to obtain the dressing.

[0140] Comparative Example 2

[0141] The rest is the same as in Example 7, except that in step S5, silk fibroin solution and silver alginate solid are added sequentially to the dodecyl alkylated chitosan solution to obtain the dressing.

[0142] Comparative Example 3

[0143] The rest is the same as in Example 7, except that in step S5, silk fibroin solution, catechin solution and silver alginate solid are mixed to prepare excipients.

[0144] Comparative Example 4

[0145] The rest is the same as in Example 7, except that in step S5, silver alginate solid is added to the 12-alkylated chitosan solution to obtain the excipient.

[0146] Comparative Example 5

[0147] The rest is the same as in Example 7, except that in step S5, silk fibroin solution, catechin solution and silver alginate solid are sequentially added to the dodecyl alkylated chitosan solution to obtain excipients.

[0148] Comparative Example 6

[0149] The rest is the same as in Example 7, except that in step S5, silk fibroin solution, catechin solution and silver alginate solid are sequentially added to the dodecyl alkylated chitosan solution to obtain excipients.

[0150] Comparative Example 7

[0151] The dressing was prepared according to patent CN201610006406.X.

[0152] Comparative Example 8

[0153] The rest is the same as in Example 7, except that the catechin solution is replaced with the same number of moles of rhein.

[0154] The following experiments demonstrate the effectiveness of specific embodiments of the present invention:

[0155] Experiment 1

[0156] In vitro coagulation experiments showed that the dressing in Example 7 had the best hemostatic effect. Cytotoxicity experiments were then conducted on the dressing obtained in Example 7, and the results are shown below. Figure 1 .Depend on Figure 1 It is evident that the dressing obtained by this invention has good biocompatibility.

[0157] Experiment 2

[0158] The dressings prepared in Examples 7 to 14 of this invention were subjected to performance tests, and the results are shown in Table 1.

[0159] Table 1. Performance tests of dressings prepared in Examples 7-14 of the present invention

[0160]

[0161]

[0162] As shown in Table 1, the composite dressings prepared in Examples 7-14 of this invention have antibacterial effects, can achieve rapid hemostasis of large arteries, have good mechanical properties, fast hemostatic efficiency, and excellent overall performance, making them suitable for various clinical conditions. The dressings rapidly capture and activate erythrocytes and platelets and accelerate the release of coagulation factors through the cationic effect of chitosan and the three-dimensional molecular network structure of natural alginate fibers in blood. Utilizing the self-assembly between polyphenolic compounds and silk fibroin biomolecules, as well as the chelation between natural mucopolysaccharides and polyphenolic polymers, a double network is formed within the silk fibroin / chitosan / tannic acid adhesive, thereby achieving rapid hemostasis. The main components of the material of this invention—chitosan, silk fibroin, tannic acid, and silver alginate—are all natural extracts, and have been verified through in vivo and in vitro experiments to have good tissue and blood biocompatibility. Meanwhile, by constructing various animal trauma models, we will study the efficacy and biosafety of this novel hemostatic material in hemostasis, wound closure, antibacterial activity, promoting healing, and inhibiting scar hyperplasia, and then systematically evaluate its military and clinical application value.

[0163] In Example 14 of this application, a dressing prepared by mixing dodecyl chitosan solution, silk fibroin solution, catechol solution, and silver alginate solid in a mass ratio of 15:1:1:1 exhibits a hemostasis time of 3 seconds and a relatively long tensile length. It should be noted that the actual hemostasis time is less than 3 seconds. Applying the dressing of Example 14 effectively achieves the unexpected technical effect of immediate hemostasis.

[0164] As those skilled in the art will know, proteins belong to a broad class of substances, and different proteins have different biological functions, sharing the same structure of amino acids. However, differences in multilevel structures and the types of amino acids lead to vastly different effects of proteins. In Example 14 of this application, silk fibroin, due to its unique structure, is a natural high-molecular-weight fibrous protein that can form a double-network structure with other substances, thereby blocking blood vessel openings and achieving rapid hemostasis.

[0165] Furthermore, the tensile length (MPa) and hemostasis time (s) of Comparative Example 8 of this invention are significantly lower than those of Example 7. Moreover, the replacement with emodin does not result in a linear decrease, but rather a precipitous drop in performance (hemostasis time changes from <10 seconds to >30 seconds, i.e., from "effective" to "virtually ineffective"). This substitution result, leading to the failure of the technical solution, is something that those skilled in the art could not have predicted. The significant difference in performance between Example 7 and Comparative Example 8 stems from the overall molecular structure rather than the number of hydroxyl functional groups in emodin alone. Those skilled in the art, based on the simple theory of "hydroxyl number," could not have predicted that the two natural polyphenols would exhibit such drastically different behaviors in the specific system of this invention (dodecyl chitosan / silk fibroin).

[0166] Experiment 3: The dressings prepared in Examples 7 to 14 of this invention were subjected to medical performance tests, and the results are shown in Table 2.

[0167] Table 2. Performance tests of dressings prepared in Examples 7-14 of the present invention

[0168]

[0169] As shown in Table 2, the dressing prepared by this invention has good medical properties.

[0170] Experiment 4: In vitro hemostasis test

[0171] The large-area wound first aid dressing prepared in Example 7 was subjected to a femoral artery puncture hemostasis experiment in mice. The experimental results are shown in [Figure 7]. Figure 2 And a blank experiment was used as a control, and the results were as follows: Figure 3 As shown. By Figure 2 The results of a femoral vein puncture hemostasis experiment in mice showed that no blood flowed out after the application of the emergency dressing, demonstrating its rapid hemostatic ability. This is mainly due to the cationic effect of dodecyl chitosan and the three-dimensional molecular network structure of natural alginate fibers in blood, which rapidly captures and activates erythrocytes and platelets, and accelerates the release of coagulation factors. Furthermore, a double network is formed within the adhesive through the self-assembly of polyphenolic compounds and silk fibroin biomolecules, as well as the chelation between dodecyl chitosan and polyphenolic polymers. Simultaneously, the silver ions released from silver alginate effectively combine with the bacterial targeting function of dodecyl chitosan, further enhancing the wound healing function of the dressing based on its bactericidal and anti-inflammatory effects. Therefore, this novel, multifunctional, large-area wound emergency dressing shows great clinical potential.

[0172] Experiment 5: Comparative Experiment

[0173] Comparative experiments showed that the hemostatic and bactericidal effects of the large-area wound emergency dressing of Example 7 of the present invention were significantly better than those of Comparative Examples 1-3.

[0174] In the hemostasis experiment, experimental animals with the same growth condition were selected and designated as Group 7 (Example 7), Group 1 (Comparative Example 1), and Group 2 (Comparative Example 2). The same wounds were created at the same locations in each of these three groups of animals. The dressings from Example 7 and Comparative Examples 1 and 2 were then applied to the wound sites, respectively.

[0175] The area covered by the dressing and the blood flow after hemostasis were measured. The larger the coverage area, the worse the hemostasis effect.

[0176] The total number of bacteria at the wound site was determined by placing 1 mL of hydrogel in a bacterial incubation environment at 37°C and measuring the total number of bacterial colonies that grew around the hydrogel.

[0177] The results are shown in Table 1. The data in Table 3 are the average values ​​within the allowable error range of multiple valid experimental data.

[0178] Table 3 Evaluation of the hemostatic and antibacterial properties of dressings for large-area wound emergency treatment

[0179] <![CDATA[Bleeding area (cm 2 )]]> Total bacterial count (CFU / mL) Comparative Example 1 2.45±0.80 4500±550 Comparative Example 2 1.20±0.45 2700±420 Example 7 0.18±0.05 350±50

[0180] As can be seen from the data in Table 3, the bleeding area and total bacterial count of the dressing in Example 7 are one order of magnitude different from those in Comparative Examples 1 and 2, indicating that the silk fibroin solution and catechol solution have a great synergistic effect on the dressing of the present invention in terms of hemostasis and antibacterial properties, and have unexpected technical effects.

[0181] Compared with Comparative Examples 1 and 2, the dressing of Example 7 of the present invention has a more significant antibacterial effect and meets the product application requirements of less than 500 CFU / mL.

[0182] Furthermore, experiments have shown that, although specific values ​​will not be exhaustive, Example 7 of this invention differs from Comparative Examples 3-8 by more than an order of magnitude in terms of bleeding area and total bacterial count, indicating that the hemostatic and antibacterial effects of Example 7 are significantly better than those of Comparative Examples 3-8. The dodecyl alkylated chitosan, silk fibroin, catechol, and silver alginate have a great synergistic effect on the hemostatic and antibacterial properties of the dressing of this invention, resulting in unexpected technical effects.

[0183] The above description of the disclosed embodiments enables those skilled in the art to make or use the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A large area wound first aid dressing kit characterized in that, The kit comprises: a first container containing a first ingredient comprising dodecylated chitosan; a second container containing a second ingredient comprising silk fibroin and catechol; the mass ratio of the dodecylated chitosan, silk fibroin and catechol is 5-15:1:1-15.

2. The kit of claim 1, wherein: the first ingredient and the second ingredient are mixed on a wet wound surface to form the emergency dressing.

3. The kit of claim 1, wherein: the first ingredient and the second ingredient, after being mixed on a wet wound surface, can form a hydrogel within 10 seconds at the wound site, and the hemostatic time is less than or equal to 15 seconds.

4. The kit of claim 3, wherein: the tensile strength of the hydrogel is not less than 0.9 MPa.

5. The large area wound first aid dressing kit of claim 1, wherein: the mass ratio of the dodecylated chitosan, silk fibroin and catechol is 15:1:

15.

6. The large area wound care dressing kit of claim 1, wherein, the method for preparing the dodecylated chitosan comprises the following steps: dissolving chitosan in an acetic acid solution, adding lauryl aldehyde and a phase transfer catalyst sodium dodecyl sulfonate to generate a Schiff base, adjusting to 4-5 with a NaOH solution, adding a NaBH4 solution dropwise, heating and stirring to perform a reduction reaction, adding lauryl aldehyde and NaBH4 again to perform a secondary reduction reaction, and purifying by using the solubility difference of chitosan with different degrees of alkylation substitution in chloroform solvent.

7. The large area wound care dressing kit of claim 1, wherein, further comprising silver alginate, wherein the mass ratio of the silver alginate to the modified chitosan is 1:1-5.

8. A method of using the large area trauma dressing kit according to any one of claims 1 to 7, characterized in that, the method comprises the following steps: removing the first ingredient from the first container; removing the second ingredient from the second container; mixing the first ingredient and the second ingredient on a wet wound surface at a mixing speed of 0.01-1 L / min to form a hydrogel dressing; and 9. The method of using a large area trauma dressing kit as defined in claim 8, wherein, the mixing is performed in a wet environment, and the wet environment comprises at least one of water, a PBS buffer or blood.

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