Antibacterial hemostatic powder capable of rapid in-situ self-gelling and preparation method thereof

Abstract

The present invention discloses an antibacterial hemostatic powder that can self-gel in situ quickly and a preparation method thereof. The preparation method of the antibacterial hemostatic powder comprises the following steps: S1, injecting chitosan solution into polyacrylic acid solution by microinjection to obtain polyacrylic acid / chitosan hydrogel microspheres; S2, freeze-drying and grinding the polyacrylic acid / chitosan hydrogel microspheres. The self-gelling hemostatic powder of the present invention has the following advantages: it can adapt to deep and irregular wounds, and can quickly and massively absorb blood, tissue fluid, etc. exuded from the wound surface to activate the coagulation reaction; after absorbing the blood at the wound surface, it can quickly form a hydrogel physical barrier with good mechanical properties to seal the wound; after in situ self-gelation, it has flexibility and adhesion, and can fit closely with the tissue to adapt to tissue deformation; it has antibacterial effect and can resist infection at the wound surface; it has good biocompatibility and biodegradability; it is easy to store and simple to operate, which greatly improves the practicality in complex emergency scenarios.

Description

Technical Field

[0001] The invention relates to a hemostatic powder and a preparation method thereof, in particular to an antibacterial hemostatic powder capable of rapid in-situ self-gelling and a preparation method thereof, and belongs to the field of biomedical materials. Background Art

[0002] Post-traumatic hemorrhage is an important cause of traumatic death in humans. After trauma, the endogenous hemostatic mechanism is difficult to stop bleeding quickly and effectively. When the blood loss reaches more than 20% of the total blood volume, it will lead to serious complications such as hypotension, organ dysfunction, and even death. Therefore, there is an urgent need for efficient hemostatic materials to deal with post-traumatic hemorrhage in clinical practice.

[0003] At present, the hemostatic gauze and sponge commonly used in clinical practice mainly accelerate the endogenous coagulation mechanism, that is, absorb the liquid in the blood, aggregate blood cells and platelets, and concentrate the coagulation factors to accelerate hemostasis. However, gauze and sponges need to be pressed and bandaged during use, and lack biological adhesion, so there are limitations when used in irregular bleeding sites or incompressible bleeding sites. In addition, since most gauze and sponges have the problem of poor biodegradability, they are easy to adhere to tissues, and are easy to cause secondary damage to the wound site during replacement and removal. Hydrogel hemostatic materials show great potential in the field of post-traumatic hemostasis, but most of the current hydrogel hemostatic materials require pretreatment, and the complicated operation limits the practicality of emergency situations. In addition, due to the high water content of hydrogels, blood will greatly weaken the adhesion between hydrogels and tissues, resulting in reduced hemostatic effect. Moreover, hemostatic hydrogel materials also have problems with residual monomers, crosslinking agents, and initiator toxicity. The new hemostatic powder material is receiving more and more attention because it is easy to use and store, can absorb blood to increase the concentration of coagulation factors at the wound, and greatly accelerate the hemostasis process. However, the common hemostatic powder on the market has poor tissue adhesion, cannot form a stable scab, and is easily washed away, resulting in poor hemostatic effect, which greatly limits the practicality of hemostatic powder in actual scenarios.

[0004] Ideal hemostatic materials generally need to meet the following requirements: (1) be able to quickly and massively absorb blood from the wound surface to concentrate coagulation factors; (2) be able to adapt to deep and irregular wound surfaces; (3) have good mechanical properties to form a physical barrier to seal the wound; (4) have flexibility and adhesion, and be able to fit tightly with tissues to adapt to tissue deformation; (5) have antibacterial effects to resist infection at the wound surface; (6) have good biocompatibility and biodegradability; (7) be easy to store and use. Hemostatic powder that can self-gel in situ is a super absorbent material that combines the advantages of powder and hydrogel. It can quickly and massively absorb blood from the wound surface, and at the same time form a hydrogel in situ to achieve wound sealing and hemostasis. The present invention is intended to provide a hemostatic powder with good biocompatibility that can meet the hemostatic needs of wound surfaces in vivo. Summary of the invention

[0005] The purpose of the present invention is to provide an antibacterial hemostatic powder that can quickly self-gel in situ, so as to solve the problems of poor biocompatibility, complicated operation during use, low tissue adhesion, difficulty in forming a stable barrier and the like of existing hemostatic materials.

[0006] The antibacterial hemostatic powder of the present invention can absorb blood at the wound surface and quickly form a gel barrier in situ. The hemostatic powder forms a physical barrier with sufficient mechanical properties based on electrostatic effects and hydrogen bonding between components, and has adhesion and antibacterial properties, can fit closely to the wound surface and effectively resist infection. In addition, the hemostatic powder of the present invention also has good biocompatibility and can meet the hemostasis needs of the wound surface in vivo.

[0007] The method for preparing the antibacterial hemostatic powder capable of rapid in-situ self-gelling provided by the present invention comprises the following steps: S1, injecting the chitosan solution into the polyacrylic acid solution by microinjection to obtain polyacrylic acid / chitosan hydrogel microspheres; S2. The polyacrylic acid / chitosan hydrogel microspheres are freeze-dried and ground to obtain an antibacterial hemostatic powder that can quickly self-gel in situ.

[0008] In the preparation method of the present invention, in the chitosan solution, the mass fraction of chitosan is 1.0% to 2.0%, preferably 1.2 to 1.8%, and the deacetylation degree is ≥95%.

[0009] In the preparation method of the present invention, in the polyacrylic acid solution, the mass fraction of polyacrylic acid is 1% to 5%, preferably 5%, and the molecular weight of polyacrylic acid is 2000 Da to 5000 Da, preferably 2000 Da. In the preparation method of the present invention, the microinjection method uses a 5-20 mL syringe to inject the chitosan solution into the polyacrylic acid solution at a speed of 1-3 mL / min.

[0010] In the preparation method of the present invention, in step S2, the freeze-drying temperature is -20°C to -40°C.

[0011] When the antibacterial hemostatic powder capable of rapid in-situ self-gelling is used, it can be directly sprayed on the wound, absorbs the blood at the wound, and rapidly gels in-situ, adheres to the tissue surface to form a physical barrier, and has certain mechanical strength and antibacterial properties.

[0012] Compared with the prior art, the present invention has the following significant beneficial effects: 1. The self-gelling hemostatic powder of the present invention can adapt to deep and irregular wounds, and can quickly and massively absorb blood, tissue fluid, etc. exuded from the wound surface to activate the coagulation reaction; 2. After absorbing the blood at the wound surface, the self-gelling hemostatic powder of the present invention can quickly form a hydrogel physical barrier with good mechanical properties to seal the wound; 3. After in-situ self-gelation, the self-gelling hemostatic powder of the present invention has flexibility and adhesion, and can fit tightly with the tissue to adapt to tissue deformation; 4. The self-gelling hemostatic powder of the present invention is based on natural polymer chitosan, has antibacterial effects and can resist infection at the wound surface; 5. The self-gelling hemostatic powder of the present invention has good biocompatibility and biodegradability; 6. The hemostatic powder of the present invention is easy to store and simple to operate, which greatly improves the practicality in complex emergency scenarios. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Figure 1 This is a physical picture of the in-situ self-gelling of the hemostatic powder of Example 5 of the present invention.

[0014] Figure 2 These are SEM images of the hemostatic powder of Example 5 of the present invention before and after self-gelation.

[0015] Figure 3 It is a quantitative characterization of the time required for the self-gelling process of the hemostatic powder in Examples 1-6 of the present invention.

[0016] Figure 4 It is a quantitative characterization of the liquid absorption amount of the hemostatic powder in Examples 1-6 of the present invention.

[0017] Figure 5 The tissue adhesion performance of the hemostatic powder of Example 5 of the present invention is characterized.

[0018] Figure 6 It is a characterization of the Young's modulus of the hemostatic powder after self-gelation in Examples 1-6 of the present invention.

[0019] Figure 7 The antibacterial properties of the hemostatic powder in Examples 1-6 of the present invention are characterized. Figure 8 The cytocompatibility characterization of the hemostatic powders of Examples 1, 3, and 5 of the present invention.

[0020] Fig. 9 This is the in vivo hemostatic performance characterization of the hemostatic powder of Example 5 of the present invention. DETAILED DESCRIPTION

[0021] Unless otherwise specified, the experimental methods and processing techniques used in the following examples are conventional methods.

[0022] Unless otherwise specified, the raw materials, reagents, etc. used in the following examples can be obtained through commercial channels.

[0023] The quantitative experiments in the following examples were all repeated three times, and the results were averaged.

[0024] Example 1: Preparation of in-situ rapid self-gelling hemostatic powder (1) The raw materials used in the self-gelling hemostatic powder of this embodiment are composed of the following, in terms of mass percentage: 1.0% chitosan solution (the solvent is 1% dilute acetic acid, the degree of deacetylation is ≥95%), 5% polyacrylic acid solution (the solvent is deionized water, the molecular weight is 2000 Da).

[0025] (2) The specific steps of the preparation are as follows: S1. Inject the chitosan solution into the polyacrylic acid solution by microinjection (syringe volume is 10 mL) at a rate of 2 mL / min to prepare polyacrylic acid / chitosan hydrogel microspheres.

[0026] S2. freeze-dry the polyacrylic acid / chitosan microspheres obtained in S1 at -40°C and grind them. Example 2: Preparation of in-situ rapid self-gelling hemostatic powder (1) The raw materials used in the self-gelling hemostatic powder of this embodiment are composed of the following components, by mass percentage: 1.2% chitosan solution (the solvent is 1% dilute acetic acid, the degree of deacetylation is ≥95%), 5% polyacrylic acid solution (the solvent is deionized water, the molecular weight is 2000 Da).

[0027] (2) The specific steps of the preparation are as follows: S1. Inject the chitosan solution into the polyacrylic acid solution by microinjection (syringe volume is 10 mL) at a rate of 2 mL / min to prepare polyacrylic acid / chitosan hydrogel microspheres.

[0028] S2. freeze-dry the polyacrylic acid / chitosan microspheres obtained in S1 at -40°C and grind them.

[0029] Example 3: Preparation of in-situ rapid self-gelling hemostatic powder (1) The raw materials used in the self-gelling hemostatic powder of this embodiment are composed of the following, in terms of mass percentage: 1.4% chitosan solution (the solvent is 1% dilute acetic acid, the degree of deacetylation is ≥95%), 5% polyacrylic acid solution (the solvent is deionized water, the molecular weight is 2000 Da).

[0030] (2) The specific steps of the preparation are as follows: S1 The chitosan solution was injected into the polyacrylic acid solution by microinjection (the syringe volume was 10 mL) at a rate of 2 mL / min to prepare polyacrylic acid / chitosan hydrogel microspheres.

[0031] S2 The polyacrylic acid / chitosan microspheres obtained in S1 were freeze-dried at -40°C and ground. Example 4: Preparation of in-situ rapid self-gelling hemostatic powder (1) The raw materials used in the self-gelling hemostatic powder of this embodiment are composed of the following, in terms of mass percentage: 1.6% chitosan solution (the solvent is 1% dilute acetic acid, the degree of deacetylation is ≥95%), 5% polyacrylic acid solution (the solvent is deionized water, the molecular weight is 2000 Da).

[0032] (2) The specific steps of the preparation are as follows: S1. Inject the chitosan solution into the polyacrylic acid solution by microinjection (syringe volume is 10 mL) at a rate of 2 mL / min to prepare polyacrylic acid / chitosan hydrogel microspheres.

[0033] S2. freeze-dry the polyacrylic acid / chitosan microspheres obtained in S1 at -40°C and grind them. Example 5: Preparation of in-situ rapid self-gelling hemostatic powder (1) The raw materials used in the self-gelling hemostatic powder of this embodiment are composed of the following, in terms of mass percentage: 1.8% chitosan solution (the solvent is 1% dilute acetic acid, the degree of deacetylation is ≥95%), 5% polyacrylic acid solution (the solvent is deionized water, the molecular weight is 2000 Da).

[0034] (2) The specific steps of the preparation are as follows: S1. Inject the chitosan solution into the polyacrylic acid solution by microinjection (syringe volume is 10 mL) at a rate of 2 mL / min to prepare polyacrylic acid / chitosan hydrogel microspheres.

[0035] S2. freeze-dry the polyacrylic acid / chitosan microspheres obtained in S1 at -40°C and grind them. Example 6: Preparation of in-situ rapid self-gelling hemostatic powder (1) The raw materials used in the self-gelling hemostatic powder of this embodiment are composed of the following, in terms of mass percentage: 2.0% chitosan solution (the solvent is 1% dilute acetic acid, the degree of deacetylation is ≥95%), 5% polyacrylic acid solution (the solvent is deionized water, the molecular weight is 2000 Da).

[0036] (2) The specific steps of the preparation are as follows: S1. Inject the chitosan solution into the polyacrylic acid solution by microinjection (syringe volume is 10 mL) at a rate of 2 mL / min to prepare polyacrylic acid / chitosan hydrogel microspheres.

[0037] S2. freeze-dry the polyacrylic acid / chitosan microspheres obtained in S1 at -40°C and grind them.

[0038] The changes of the hemostatic powder before and after the gelation of the above-mentioned Example 5 are as follows: Figure 1-2 As shown, Figure 1 This is a physical picture of the hemostatic powder self-gelling from powder state to gel state. Figure 2 From the SEM pictures of the hemostatic powder before and after self-gelling, it can be seen that the powder particles of the hemostatic powder of the present invention are interconnected to form a whole after absorbing the liquid.

[0039] Example 7: Quantitative characterization of the time required for the gelation process of the in-situ rapid self-gelling hemostatic powder The gel transition time of the hemostatic powder of Examples 1-6 of the present invention was tested using a rheometer. Figure 3 As shown in the figure, as the chitosan concentration increases from 1.0% to 1.8%, the gelation time decreases from 6.12±6.43 s to 1.96±1.94 s. When the chitosan concentration reaches 2%, the excess chitosan will cause the mechanical properties of the whole system to decrease rapidly and the gelation transition time will increase significantly.

[0040] Example 8: Characterization of liquid absorption capacity of in-situ rapid self-gelling hemostatic powder The liquid absorption capacity of the hemostatic powder of Examples 1-6 of the present invention is tested by weighing. The mass change before and after water absorption is weighed using a precision balance to calculate the liquid absorption rate of Examples 1-6 (liquid absorption rate = (m-m0) / m0×100%, where m0 is the weight of the hemostatic powder before water absorption and m is the weight of the hemostatic powder after water absorption). Figure 4 As shown, as the chitosan concentration increases from 1.0% to 2.0%, the hydrophilic groups such as hydroxyl and amino groups increase accordingly, and the liquid absorption capacity of the hemostatic powder of the present invention increases from 154.56% to 234.68%.

[0041] Example 9: Characterization of tissue adhesion effect of in situ rapid self-gelling hemostatic powder The tissue adhesion effect of the hemostatic powder of Example 5 of the present invention was characterized using ex vivo animal tissue. Figure 5 As shown, the hemostatic powder has excellent adhesion effect on pig skin, pig heart and chicken skin, and can resist the erosion of water flow.

[0042] Example 10: Mechanical property test of in-situ rapid self-gelling hemostatic powder The hemostatic powders of Examples 1-6 of the present invention were prepared into dumbbell-shaped hydrogel samples with a length of 50 mm and a thickness of 5 mm, and their tensile mechanical properties were tested. Figure 6As shown in the figure, as the chitosan concentration increases from 1.0% to 1.8%, some non-covalent bonds such as hydrogen bonds increase, which increases the Young's modulus of the hemostatic powder from 59.60±1.28 kPa to 158.40±5.80 kPa, which has sufficient mechanical strength as a physical barrier. When the chitosan concentration reaches 2%, the excess chitosan causes the Young's modulus of the entire system to drop rapidly to 73.57+4.4 kPa.

[0043] Example 11: Antibacterial performance test of in-situ rapid self-gelling hemostatic powder The hemostatic powder of the above-mentioned Examples 1-6 of the present invention was prepared into a disc sample with a diameter of 10 mm, and its antibacterial property was tested by using the inhibition zone test. 200 μL of Escherichia coli suspension (1×10 5 CFU / g) and 200 μL of Staphylococcus aureus suspension (1×10 5 CFU / g) were spread on LB agar medium. Then the hydrogel disc was inoculated on the LB agar medium coated with bacterial solution and incubated at 37℃ for 24 h before counting the diameter of the inhibition zone. Figure 7 As shown, as the chitosan concentration increased from 1.0% to 2.0%, the diameter of the Escherichia coli inhibition zone increased from 11.32±0.33 cm to 25.10±0.42 cm, and the diameter of the Staphylococcus aureus inhibition zone increased from 16.94±0.98 cm to 24.71±0.83 cm, indicating that the hemostatic powder of the present invention has excellent antibacterial effect.

[0044] Example 12: Characterization of the Cytocompatibility of In-situ Rapid Self-gelling Hemostatic Powder The hemostatic powders of Examples 1, 3, and 5 of the present invention were co-cultured with cells, and the cell compatibility was tested by CCK-8 method and live / dead cell staining method, respectively. Figure 8 As shown, after the hemostatic powder was co-cultured with cells for 24 hours, the cell morphology was normal and the cell viability was above 100%, indicating excellent cell compatibility. Example 13: In-situ rapid self-gelling hemostatic powder in vivo hemostatic performance test The in vivo hemostatic performance of the hemostatic powder of Example 5 of the present invention was tested using a mouse liver bleeding model. Fig. 9As shown, after inducing liver bleeding in mice, the hemostasis time of the blank group without intervention was 65.33±4.73 s, and the blood loss was 106.67±11.55 mg. The hemostasis time of the control group treated with commercial hemostatic powder was 55.67±5.13 s, and the blood loss was 84.67±13.61 mg. The hemostasis time of the experimental group treated with in situ rapid self-gelling hemostatic powder was 35.67±3.21 s, and the blood loss was 30.33±10.79 mg. It can be seen that the amount of bleeding of the in situ rapid self-gelling hemostatic powder was significantly reduced, and the hemostasis time was significantly shortened. In vivo experiments have proved that the self-gelling hemostatic powder of the present invention has excellent hemostatic properties.

Claims

1. A method for preparing an antibacterial hemostatic powder capable of rapid in-situ self-gelling, comprising the following steps: S1, injecting the chitosan solution into the polyacrylic acid solution by microinjection to obtain polyacrylic acid / chitosan hydrogel microspheres; S2. The polyacrylic acid / chitosan hydrogel microspheres are freeze-dried and ground to obtain an antibacterial hemostatic powder that can quickly self-gel in situ.

2. The preparation method according to claim 1, characterized in that: In the chitosan solution, the mass fraction of chitosan is 1.0% to 2.0%, and the deacetylation degree is ≥95%.

3. The preparation method according to claim 1, characterized in that: In the polyacrylic acid solution, the mass fraction of polyacrylic acid is 1% to 5%, and the molecular weight of polyacrylic acid is 2000 Da to 5000 Da.

4. The preparation method according to claim 1, characterized in that: The microinjection method uses a 5-20 mL syringe to inject the chitosan solution into the polyacrylic acid solution at a speed of 1-3 mL / min.

5. The preparation method according to any one of claims 1 to 4, characterized in that: In step S2, the freeze-drying temperature is -20°C to -40°C.

6. The antibacterial hemostatic powder prepared by the method according to any one of claims 1 to 5.

7. Use of the antibacterial hemostatic powder according to claim 6 in the preparation of hemostatic materials.

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