Preparation method of multifunctional composite hemostatic powder capable of realizing rapid in-situ self-gelling

Abstract

The invention provides a preparation method of quick in-situ self-gelling multifunctional composite hemostatic powder, which comprises the following steps: S1, reacting natural polysaccharide with chloroacetic acid to prepare anionic natural polysaccharide, washing, drying and crushing to obtain anionic natural polysaccharide powder; s2, the natural polysaccharide reacts with 3-chloro-2-hydroxypropyl trimethyl ammonium chloride to prepare cationic natural polysaccharide, and cationic natural polysaccharide powder is obtained through washing, drying and smashing; s3, mixing the anionic natural polysaccharide powder with the cationic natural polysaccharide powder to obtain the multifunctional composite styptic powder capable of realizing rapid in-situ self-gelling. The composite styptic powder has good cell compatibility and blood compatibility, and good antibacterial effects on gram-negative escherichia coli and positive staphylococcus aureus.

Description

Technical Field

[0001] The invention relates to the field of medical polymer materials, in particular to a method for preparing multifunctional composite hemostatic powder capable of rapid in-situ self-gelation. Background Art

[0002] With the increasing frequency of natural disasters and traffic accidents, uncontrolled heavy bleeding remains the leading cause of death among people under 45 years old, especially for severe trauma to soft tissues such as the skin, liver, lungs and heart, where it is particularly difficult to stop bleeding.

[0003] In recent years, scientists have paid more and more attention to hemostatic materials used to control bleeding from incompressible wounds in pre-hospital treatment or surgery, and proposed different hemostatic strategies. Although these materials have different degrees of hemostatic effects in the hemostatic process, they all show limitations that cannot be ignored for wounds of varying depths and irregular shapes. For example, sponges / foams mainly rely on absorbing excess blood from damaged tissues to expand to seal wounds, but this usually causes tissue pain and exerts local pressure on nerves, which is not conducive to the health of patients; traditional hemostatic particles / powders are conducive to hemostasis of incompressible wounds, by aggregating coagulation factors in the blood to form thrombi, but due to their weak mechanical strength and poor tissue adhesion, they are easily dispersed by blood, leading to hemostasis failure. In addition, wound infection during the hemostasis process is an inevitable problem, which may lead to shock, disability or even death if not handled in time. Therefore, in order to ensure the rapid healing of patients' wounds, it is particularly important to develop a dressing that has both hemostatic and antibacterial functions.

[0004] Natural polysaccharides are abundant in sources, biocompatible, and biodegradable, and are therefore widely used in hemostatic medical materials. For example, chitosan (CS) is a linear biopolymer obtained by deacetylation of chitin, which is widely used in the biomedical field due to its good biosafety and antibacterial properties. The insolubility of chitosan in water limits its scope of application. Quaternary ammonium chitosan (QCS) obtained after CS modification is a cationic polysaccharide, which not only enhances water solubility, but also enhances the antibacterial activity of QCS due to the introduction of positive charges. At the same time, QCS can also establish electrostatic interactions with anionic polysaccharides with opposite charges to form polyelectrolyte complexes. It is worth noting that powder hemostatic agents are suitable for hemostasis of irregularly shaped, non-pressable wounds due to their shape adjustability. As a typical powder hemostatic agent, the U.S. Army Institute of Surgery recommends the use of aluminosilicate-based products (QuikClot) and their application on the battlefield. However, although QuikClot can quickly absorb plasma to concentrate red blood cells and coagulation factors to promote wound coagulation, it may be accompanied by the risk of thermal injury.

[0005] Therefore, constructing hemostatic materials with biosafety, efficient hemostatic performance and antibacterial activity will play a significant role in the development of advanced hemostatic dressings. Summary of the invention

[0006] In view of this, the object of the present invention is to provide a method for preparing a multifunctional composite hemostatic powder with rapid in-situ self-gelation, so that the hemostatic powder has biological safety, high-efficiency hemostatic performance and antibacterial activity.

[0007] In order to solve the above technical problems, the technical solution of the present invention is: a method for preparing a multifunctional composite hemostatic powder with rapid in-situ self-gelling, comprising the following steps: S1: natural polysaccharide reacts with chloroacetic acid to prepare anionic natural polysaccharide, and the anionic natural polysaccharide is washed, dried and crushed to obtain anionic natural polysaccharide powder; S2: reacting natural polysaccharide with 3-chloro-2-hydroxypropyltrimethylammonium chloride to prepare cationic natural polysaccharide, and washing, drying and crushing the cationic natural polysaccharide to obtain cationic natural polysaccharide powder; S3: Mix anionic natural polysaccharide powder with cationic natural polysaccharide powder to obtain a rapid in-situ self-gelling multifunctional composite hemostatic powder.

[0008] As a preferred embodiment of the present invention, the natural polysaccharide in steps S1 and S2 is one of cellulose, chitosan and starch.

[0009] As a preferred embodiment of the present invention, the anionic natural polysaccharide in step S1 is one of carboxymethyl cellulose, carboxymethyl chitosan and carboxymethyl starch.

[0010] As a preferred embodiment of the present invention, the cationic natural polysaccharide in step S2 is one of quaternary ammoniated cellulose, quaternary ammoniated chitosan and quaternary ammoniated starch.

[0011] As a preferred embodiment of the present invention, the mixing ratio of the anionic natural polysaccharide powder and the cationic natural polysaccharide powder in step S3 is determined by the contents of carboxymethyl and quaternary amino groups, and the molar amounts of the carboxymethyl and quaternary amino groups are equal.

[0012] As a preferred embodiment of the present invention, the degree of carboxymethyl substitution of the anionic natural polysaccharide is 0.4-1.2.

[0013] As a preferred embodiment of the present invention, the degree of substitution of the quaternary amino group of the cationic natural polysaccharide is 20%-98%.

[0014] In summary, the present invention has the following beneficial effects: After the anionic natural polysaccharide powder and cationic natural polysaccharide powder carrying opposite charges come into contact with blood, they swell and dissolve. The electrostatic interaction and hydrogen bonding between the molecular chains cause the two polysaccharides to quickly physically crosslink in situ to form a hydrogel within 10 seconds. At the same time, the red blood cells and platelets in the blood are concentrated and locked in the hydrogel network to form a thrombus, leading to rapid hemostasis. In addition, the quaternary amino groups on the polysaccharide molecular chains have a good antibacterial effect on Gram-negative and Gram-positive bacteria. Therefore, the composite hemostatic powder has the dual functions of hemostasis and antibacterial. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 The scanning electron micrographs of the hydrogels formed by induction of three composite hemostatic powders prepared in the present invention, namely, Example 1 quaternized chitosan / carboxymethyl chitosan, Example 2 quaternized chitosan / carboxymethyl starch, and Example 3 quaternized chitosan / carboxymethyl cellulose, and water; Figure 2 The in vitro coagulation index statistical graph of three composite hemostatic powders prepared by the present invention, namely, quaternized chitosan, carboxymethyl chitosan, carboxymethyl starch, carboxymethyl cellulose, quaternized chitosan / carboxymethyl chitosan of Example 1, quaternized chitosan / carboxymethyl starch of Example 2, quaternized chitosan / carboxymethyl cellulose of Example 3, and CeloxTM hemostatic powder of the control group; Figure 3 Statistical graph of shear adhesion strength of three composite hemostatic powders prepared by the present invention, namely quaternized chitosan, carboxymethyl chitosan, carboxymethyl starch, carboxymethyl cellulose, quaternized chitosan / carboxymethyl chitosan of Example 1, quaternized chitosan / carboxymethyl starch of Example 2, quaternized chitosan / carboxymethyl cellulose of Example 3, and CeloxTM hemostatic powder of the control group.

[0016] Figure 4 Statistical graphs of bursting pressures of three composite hemostatic powders prepared by the present invention: quaternized chitosan, carboxymethyl chitosan, carboxymethyl starch, carboxymethyl cellulose, Example 1 quaternized chitosan / carboxymethyl chitosan, Example 2 quaternized chitosan / carboxymethyl starch, Example 3 quaternized chitosan / carboxymethyl cellulose, and the control group CeloxTM hemostatic powder; Figure 5 Scanning electron micrographs of the adhesion of red blood cells to three composite hemostatic powders prepared in accordance with the present invention, namely, Example 1 quaternized chitosan / carboxymethyl chitosan, Example 2 quaternized chitosan / carboxymethyl starch, Example 3 quaternized chitosan / carboxymethyl cellulose, and the control group CeloxTM hemostatic powder; Figure 6 The hemostatic conditions and bleeding volume of three composite hemostatic powders prepared in Example 1, quaternized chitosan / carboxymethyl chitosan, Example 2, quaternized chitosan / carboxymethyl starch, Example 3, quaternized chitosan / carboxymethyl cellulose and the control group CeloxTM hemostatic powder; Figure 7 Diagram of the gelation process of hemostatic powder. DETAILED DESCRIPTION

[0017] The specific implementation modes of the present invention are further described below in conjunction with the accompanying drawings to make the technical solutions of the present invention easier to understand and grasp. Example

[0018] A method for preparing a carboxymethyl chitosan / quaternized chitosan composite hemostatic powder comprises the following steps: S1. Preparation of carboxymethyl chitosan powder. The chitosan powder was dispersed in a sodium hydroxide solution (50%, w / v), a certain amount of isopropanol was added, and the mixed solution was stirred at 50°C for 1h. Subsequently, chloroacetic acid isopropanol solution was added dropwise and reacted at 50°C for 4h. Acetic acid was then added, the pH of the reaction solution was adjusted to 7, the precipitate was collected, and the precipitate was washed with methanol aqueous solution. Finally, it was dried in an oven at 80°C for 48h, and the carboxymethyl chitosan powder was obtained by grinding and sieving. The degree of carboxymethyl substitution was 0.9.

[0019] S2. Preparation of quaternized chitosan powder. Dissolve chitosan powder in an alkali-urea system mixed solution to obtain a 4wt% chitosan solution. Then, drop 60wt% 3-chloro-2-hydroxypropyltrimethylammonium chloride solution and react at 30°C for 24h. Dialyze the product in a hydrochloric acid aqueous solution for 1 week. Finally, freeze-dry, grind and sieve to obtain quaternized chitosan powder. The degree of substitution of the quaternary ammonium group is 92%. 3. Mechanically blend the carboxymethyl chitosan powder obtained in step 1 and the quaternized chitosan powder obtained in step 2 in a mass ratio of 1:1 to obtain carboxymethyl chitosan / quaternized chitosan. Recorded as composite hemostatic powder-1. Example

[0020] S1. Starch (8 wt%) and NaOH (4 wt%) were added to an ethanol solution, and then mechanically stirred in a 50 ℃ water bath for 1 h to swell and alkalize the starch. Then a certain amount of chloroacetic acid was added dropwise, and the mixture was reacted at 50 ℃ for 4 h. The pH of the mixture was adjusted to 7 with a hydrochloric acid aqueous solution to obtain a precipitate. Carboxymethyl starch powder was obtained after washing, drying, grinding and sieving.

[0021] S2. Mechanically blend the carboxymethyl starch powder obtained in step 1 with the quaternary ammonium chitosan powder obtained in step 2 of Example 1 to obtain carboxymethyl starch / quaternary ammonium chitosan, which is referred to as composite hemostatic powder-2. Example

[0022] S1. Disperse cellulose powder in sodium hydroxide solution (35%, w / v), add a certain amount of isopropanol, and stir the mixed solution at 300 rpm at 30°C for 1 h. Then, add chloroacetic acid solution dropwise and react at 55°C for 4 h. Filter the mixture and collect the residue, stir and disperse the residue in methanol for 15 min, and then neutralize the dispersion with acetic acid. Then filter the dispersion and wash it repeatedly with ethanol for 5 times. Then dry it at 60°C for 48 h. Grind and sieve to obtain carboxymethyl cellulose powder.

[0023] S2. Mechanically blend the carboxymethyl cellulose powder obtained in step 1 with the quaternary ammonium chitosan powder obtained in step 2 of Example 1 to obtain carboxymethyl cellulose / quaternary ammonium chitosan, which is referred to as composite hemostatic powder-3. Example

[0024] Example 4 relates to the hemostatic effect test of hemostatic powder, comprising the following steps: S1. Inject 1 mL of 10% chloral hydrate intraperitoneally to anesthetize the rat. S2, making a 0.3 cm × 0.3 cm × 0.3 cm puncture wound on the liver; S3. Immediately after blood seeps out, 40 mg of the composite hemostatic powder sample to be tested is injected into the liver wound, and the time is recorded immediately. When the blood no longer seeps out from the liver wound, the recording is stopped. The mass of the gauze before and after hemostasis is weighed to calculate the amount of bleeding. Each sample is repeated 5 times during the experiment. Celox TM The hemostatic effect of hemostatic powder on the liver was used as a sample and served as a blank control group; S4, control group Celox TM The bleeding amount of the hemostatic powder and the composite hemostatic powder used in Examples 1-3 is as follows: Figure 5 Compared with the control group Celox TM Compared with the hemostatic powder, the composite hemostatic powder used in Examples 1-3 has a greatly improved hemostatic effect in the rat liver puncture model. TM When the hemostatic powder was injected into the cylindrical wound on the liver of a rat, the powder was easily dispersed by the flowing blood. The blood quickly spread to the surface of the liver and flowed from the gap to the gauze pad placed under the liver. The clotting time was as long as 4.50±0.057min, and the blood loss was as high as 0.819±0.033g. The composite hemostatic powder showed excellent hemostatic effect. For example, when the composite hemostatic powder was injected into the liver puncture injury wound, the powder would quickly absorb the excess blood on the wound and quickly self-gel, forming a physical barrier to tightly seal the bleeding point. Its clotting time and blood loss were much lower than Celox. TMIn particular, the composite hemostatic powder-1 achieved hemostasis at 0.52±0.015min of wound bleeding, and the blood loss was as low as 0.05±0.006g, accounting for only 5.70% of the CeloxTM group.

[0025] Of course, the above are only typical examples of the present invention. In addition, the present invention may also have many other specific implementations. All technical solutions formed by equivalent replacement or equivalent transformation fall within the scope of protection required by the present invention.

Claims

1. A method for preparing a multifunctional composite hemostatic powder with rapid in-situ self-gelling, characterized in that: The steps include: S1: natural polysaccharide reacts with chloroacetic acid to prepare anionic natural polysaccharide, and the anionic natural polysaccharide is washed, dried and crushed to obtain anionic natural polysaccharide powder; S2: reacting natural polysaccharide with 3-chloro-2-hydroxypropyltrimethylammonium chloride to prepare cationic natural polysaccharide, and washing, drying and crushing the cationic natural polysaccharide to obtain cationic natural polysaccharide powder; S3: Mix anionic natural polysaccharide powder with cationic natural polysaccharide powder to obtain a rapid in-situ self-gelling multifunctional composite hemostatic powder.

2. The method for preparing a multifunctional composite hemostatic powder with rapid in-situ self-gelling according to claim 1, characterized in that: The natural polysaccharide in steps S1 and S2 is one of cellulose, chitosan and starch.

3. The method for preparing a multifunctional composite hemostatic powder with rapid in-situ self-gelling according to claim 1, characterized in that: The anionic natural polysaccharide in step S1 is one of carboxymethyl cellulose, carboxymethyl chitosan and carboxymethyl starch.

4. The method for preparing a multifunctional composite hemostatic powder with rapid in-situ self-gelling according to claim 3, characterized in that: The cationic natural polysaccharide in step S2 is one of quaternary ammoniated cellulose, quaternary ammoniated chitosan and quaternary ammoniated starch.

5. The method for preparing a multifunctional composite hemostatic powder with rapid in-situ self-gelling according to claim 4, characterized in that: The mixing ratio of the anionic natural polysaccharide powder and the cationic natural polysaccharide powder in step S3 is determined by the content of carboxymethyl and quaternary amino groups, and the molar amounts of the carboxymethyl and quaternary amino groups are equal.

6. The method for preparing a multifunctional composite hemostatic powder with rapid in-situ self-gelling according to claim 5, characterized in that: The carboxymethyl substitution degree of the anionic natural polysaccharide is 0.4-1.

2.

7. The method for preparing a multifunctional composite hemostatic powder with rapid in-situ self-gelling according to claim 6, characterized in that: The degree of substitution of the quaternary amino group of the cationic natural polysaccharide is 20%-98%.

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