CN109111591B

CN109111591B - Preparation method of drug-loaded hemostatic sponge and drug-loaded hemostatic sponge prepared by preparation method

Info

Publication number: CN109111591B
Application number: CN201810973621.6A
Authority: CN
Inventors: 汤克勇; 范夏莲; 李萌雅; 刘捷; 郑学晶; 裴莹; 李修敏; 王芳
Original assignee: Zhengzhou University
Current assignee: Zhengzhou University
Priority date: 2018-08-24
Filing date: 2018-08-24
Publication date: 2020-10-09
Anticipated expiration: 2038-08-24
Also published as: CN109111591A

Abstract

The invention discloses a preparation method of a medicine-carrying hemostatic sponge, which comprises the following steps: (1) preparing a lignocellulose sponge; (2) chitosan and hemostatic are loaded on the lignocellulose sponge in a spraying or dipping mode, and the drug-loaded hemostatic sponge is obtained by freeze-drying. The preparation process of the lignocellulose sponge comprises the following steps: 1) freezing the alkali urea solution to form ice slurry, and then adding lignocellulose into the ice slurry to prepare a lignocellulose solution; 2) adding a surfactant into the lignocellulose solution, and stirring to form a foam body; 3) freezing the foam body into foam ice slurry, adding a pore-foaming agent into the foam ice slurry, and freezing and aging the foam ice slurry after injection molding to obtain gel; 4) soaking to remove the surfactant and pore-forming agent in the gel, freezing, shaping, and freeze-drying to obtain the lignocellulose sponge. The medicine-carrying hemostatic sponge prepared by the invention can rapidly absorb blood on the surface of a wound, effectively promote the coagulation of the wound surface and rapidly stanch through the synergistic effect of chitosan and hemostatic medicines.

Description

Preparation method of drug-loaded hemostatic sponge and drug-loaded hemostatic sponge prepared by preparation method

Technical Field

The invention belongs to the field of medical high polymer materials, and particularly relates to a preparation method of a drug-loaded hemostatic sponge, and a drug-loaded hemostatic sponge product prepared by the method.

Background

Blood is an important carrier for oxygen delivery to the body. When the human body loses excessive blood due to injury, organs and cells can generate dysfunction and failure, and the life safety is influenced. Uncontrolled bleeding has always been the main cause of casualties. Although personal protection devices such as body armor, helmets and the like are used for protection, bleeding still is a main cause of death of battlefield trauma in modern war, and particularly when the trauma occurs in limb connection parts such as armpits, groin, neck and the like, the parts can not be stopped by tourniquets or artificial compression, and people are easy to suffer hemorrhagic shock and even death. Therefore, timely and effective hemostasis can greatly reduce the death rate of battlefield trauma. The 1000 necropsy reports from the army dying in the afghanistan and iraq war showed 85% of hemorrhage in salvageable deaths, with 69% of hemorrhage not normally compressible. Therefore, effective achievement of rapid hemostasis is critical to improving the survival rate of the injured in a battlefield or in a major accident.

The fast hemostatic materials reported at present mainly comprise hemostatic powder (zeolite, collagen powder and potato starch), porous sponge (gelatin sponge and collagen sponge), fibrin glue and the like, but all of them have certain disadvantages: the tissue adhesion of gelatin and collagen is poor, and both of them depend on sufficient platelets and coagulation factors for their hemostatic function; fibrin is derived from blood and may cause viral infection; porous zeolite and potato starch give off a large amount of heat after absorbing moisture in blood, and easily cause inflammation of wounds.

The Chinese patent application with the publication number of CN 107050502A introduces a super-absorbent polymer hydrogel xerogel sponge and a preparation method and application thereof, the super-absorbent polymer hydrogel xerogel sponge is prepared by using chitosan as a framework and super-absorbent polymers as branched chains, and a stereo mesh porous sponge with macromolecules or polymers with a flexible structure as a cross-linking agent is prepared, the sponge is of a porous structure and has super-absorbent characteristics, the integral structure and certain mechanical strength are kept after water absorption, but the used super-absorbent polymers such as polyacrylamide and the like and the cross-linking agent glutaraldehyde and the like can bring certain toxicity to the porous sponge, and potential danger exists; in addition, the material has low strength, is easy to break into tiny fragments, and can possibly cause starting venous thrombosis. Therefore, the safe, stable, quick and effective hemostatic sponge is imperative to be prepared.

Disclosure of Invention

The invention aims to provide a preparation method for preparing a medicine-carrying hemostatic sponge and a medicine-carrying hemostatic sponge product prepared by the method.

Based on the purpose, the invention adopts the following technical scheme:

the invention firstly provides a preparation method of a medicine-carrying hemostatic sponge, which comprises the following steps:

(1) preparing a lignocellulose sponge:

1) an alkali urea solution is prepared and frozen to form an ice slurry (i.e.: a solid-liquid two-phase solution containing suspended ice crystal particles); adding lignocellulose into the alkali urea solution in the ice slurry state by stirring to prepare a lignocellulose solution;

2) adding a surfactant into the lignocellulose solution prepared in the step 1), and quickly stirring to form a stable foam body;

3) freezing the foam prepared in the step 2) to form foam ice slurry, adding a pore-foaming agent into the foam ice slurry while stirring, and freezing and aging after injection molding to obtain gel;

4) soaking the gel prepared in the step 3) in water to remove the surfactant and the pore-forming agent, freezing and shaping, and freeze-drying to prepare the lignocellulose sponge;

(2) loading chitosan and hemostatic drug on the lignocellulose sponge:

a. preparing a chitosan solution, adding a hemostatic drug into the chitosan solution, and uniformly stirring and dispersing to obtain a chitosan-hemostatic drug composite solution;

b. loading the chitosan-hemostatic drug composite solution on the lignocellulose sponge by adopting a spraying or dipping method, then freezing and shaping, and then freeze-drying to obtain the drug-loaded hemostatic sponge.

According to the preparation method of the drug-loaded hemostatic sponge, preferably, in the step 1), the weight of the added lignocellulose is 1wt% -3 wt% of the alkali urea solution; in the step 2), the weight of the added surfactant is 0.1-8 wt% of the weight of the lignocellulose solution; in the step 3), the weight of the added pore-foaming agent is 40 to 80 weight percent of the weight of the lignocellulose solution

According to the preparation method of the drug-loaded hemostatic sponge, preferably, the surfactant is one or more of sodium dodecyl sulfate, sodium dodecyl benzene sulfonate, alkyl glycoside, sodium fatty alcohol-polyoxyethylene ether sulfate and fatty acid methyl ester sulfonate. The surfactants are wide in source and rich in foam, and are added into the lignocellulose solution and stirred to easily form a stable foam body, so that the using amount of the pore-forming agent can be effectively reduced, the porosity and the water absorption of the sponge are greatly improved, and the softness of the sponge is improved.

According to the preparation method of the drug-loaded hemostatic sponge, the pore-forming agent is preferably sodium sulfate decahydrate (Na)₂SO₄·10H₂O), anhydrous sodium sulfate (Na)₂SO₄) Anhydrous magnesium sulfate (MgSO)₄) One or more of (a).

According to the preparation method of the drug-loaded hemostatic sponge, preferably, the freezing temperature in the step 3) is-18 to-24 ℃, and experiments prove that the freezing in the temperature range can meet the requirements of the invention.

According to the preparation method of the drug-loaded hemostatic sponge, preferably, the lignocellulose sponge prepared in the step 4) has the porosity of 96.10-98.77% and the density of 18.7-59.6 mg/cm³。

According to the preparation method of the drug-loaded hemostatic sponge, preferably, the temperature of the freezing and shaping in the step 4) is-18 to-24 ℃, and experiments prove that the freezing and shaping in the temperature range can meet the requirements of the invention.

According to the preparation method of the drug-loaded hemostatic sponge, preferably, in the step a, the chitosan is acid-soluble chitosan with a deacetylation degree of more than 90%, and the acid-soluble chitosan is prepared into a chitosan solution by using 0.01-0.5 mol/L of dilute acetic acid as a solvent. More preferably, the acid-soluble chitosan is prepared into a chitosan solution by using 0.1 mol/L diluted acetic acid as a solvent.

According to the preparation method of the drug-loaded hemostatic sponge, preferably, in the step a, the chitosan is water-soluble chitosan, and the water-soluble chitosan is one or more of carboxymethyl chitosan, hydroxypropyl chitosan and 2-hydroxypropyl trimethyl ammonium chloride chitosan.

According to the preparation method of the drug-loaded hemostatic sponge, preferably, in the step a, the mass fraction of chitosan in the chitosan solution is 0.1wt% -1 wt%; the mass fraction of the hemostatic medicament in the chitosan-hemostatic medicament composite solution is 0.1-1 wt%.

According to the preparation method of the drug-loaded hemostatic sponge, preferably, in the step a, the hemostatic drug is one or more of vitamin K1, vitamin K3, vitamin K4, acetic glycine ethylenediamine, carbachol, carbazochrome sodium sulfonate, Yunnan white drug powder and thrombin.

According to the preparation method of the drug-loaded hemostatic sponge, preferably, the spraying in the step b is to add the chitosan-hemostatic drug composite solution into a spray gun and uniformly spray the chitosan-hemostatic drug composite solution on the surface of the lignocellulose sponge.

According to the preparation method of the drug-loaded hemostatic sponge, preferably, the soaking in the step b is to soak the lignocellulose sponge in the chitosan-hemostatic drug composite solution for 2-5 hours.

According to the preparation method of the drug-loaded hemostatic sponge, preferably, the freezing and shaping temperature in the step b is-18 to-24 ℃, and experiments prove that the freezing and shaping within the temperature range can meet the requirements of the invention.

In the hemostatic drug, the carbachol can directly act on blood vessels, has no adrenomimetic effect, thus not influencing the blood pressure and the heart rate, and can enhance the resistance of capillary vessels to injury, stabilize acidic mucopolysaccharide in the blood vessels and surrounding tissues, reduce the permeability of the capillary vessels, enhance the retraction effect of the end of the injured capillary vessels, ensure that blood clots are not easy to fall off from the vessel walls, thereby shortening the hemostasis time without influencing the blood coagulation process. Meanwhile, the synthesis and release of prostaglandin E1(PGE1) can be inhibited, so that the permeability of capillary vessels is reduced, and the exudation of pyrogenic substances is prevented. The product can be absorbed by oral administration, but can be rapidly destroyed and discharged in gastrointestinal tract.

The invention also provides a medicine-carrying hemostatic sponge product prepared by the preparation method of the medicine-carrying hemostatic sponge. Furthermore, according to different wound positions and wound shapes, the medicine-carrying hemostatic sponge product prepared by the invention can be placed into molds of different shapes for compression molding treatment, vacuum packaging is carried out after compression, and then the compressed medicine-carrying hemostatic sponge with a specific shape is obtained so as to meet the hemostatic requirements of different wounds, and meanwhile, the compressed medicine-carrying hemostatic sponge after compression treatment is convenient to store, transport and carry.

Compared with the prior art, the invention has the following advantages:

1) the preparation method of the invention selects lignocellulose as the base material, and utilizes the surfactant and the inorganic salt pore-forming agent to jointly foam, so that the prepared product is light, soft and elastic. Although the surfactant is rich in foam, the produced foam is unstable, and the foam is gradually reduced along with the increase of the standing time of the solution; the proper amount of inorganic salt pore-forming agent is added, the inorganic salt particles stabilize the foam by occupying vacant sites in the cellulose solution, and simultaneously, the porosity and the fluffiness of the lignocellulose sponge can be increased, and the inorganic salt particle and the lignocellulose sponge supplement each other to jointly promote the formation of the lignocellulose sponge with high porosity and high fluffiness.

In addition, the combined action of the pore-forming agent and the surfactant also helps to form fiber bundles in the lignocellulose sponge, the fiber bundle structure can show excellent capillary effect, the water absorption rate and the water absorption capacity of the lignocellulose sponge are greatly improved, the lignocellulose sponge shows faster water absorption reaction, the redundant blood on the surface of a wound can be quickly absorbed, and the purpose of quick hemostasis is achieved.

2) Dissolving the lignocellulose in the presence of ice crystals, which helps to better disperse the lignocellulose; and a pore-foaming agent is further added into the foam containing the ice crystals, so that the powdery pore-foaming agent can be effectively prevented from eliminating foam in the adding process, and the stability of the foam is kept.

3) The preparation method provided by the invention has mild conditions, no cross-linking agent is used in the preparation process, the lignocellulose sponge is prepared by combining the surfactant and the pore-forming agent, both the surfactant and the pore-forming agent are easy to elute by water immersion, no harmful substance residue exists, and the safety is high.

4) The lignocellulose sponge prepared by the invention has high porosity (the porosity range is 96.10-98.77%), high water absorption capacity and a special microstructure of a sponge body, and has the characteristics of rapid water absorption and high water absorption capacity. In addition, the inventor finds in the research process that after the chitosan and the hemostatic drug are loaded on the lignocellulose sponge, the loading of the chitosan and the hemostatic drug has little influence on the porosity of the lignocellulose sponge, and the porosity of the sponge is not changed basically. Therefore, by virtue of the high porosity and high water absorption capacity of the lignocellulose sponge and the special microstructure of the sponge body, the medicine-carrying hemostatic sponge product prepared by the invention also has the characteristics of rapid water absorption, strong water absorption capacity and the like, and can rapidly absorb blood on the surface of a wound. Meanwhile, by loading chitosan and the hemostatic sponge, on one hand, chitosan has the effects of promoting blood coagulation (the chitosan surface has a large number of amino groups, and the positively charged amino groups and negatively charged red blood cells act to make the red blood cells adhere and aggregate, so that blood is coagulated), sterilizing, promoting wound healing, absorbing wound exudate, being not easy to dehydrate and shrink and the like, and on the other hand, the hemostatic drug has the effects of hemostasis, blood coagulation, inflammation diminishing and the like. Therefore, the chitosan, the hemostatic medicine and the hemostatic sponge are matched and have synergistic effect, so that the blood on the surface of a wound can be rapidly absorbed, the coagulation of the wound surface is effectively promoted, and the aim of rapidly stopping bleeding is fulfilled.

5) According to different shapes of wound parts and wounds, the medicine-carrying hemostatic sponge prepared by the invention can be put into molds of different shapes for compression molding treatment, and vacuum packaging is carried out after compression, so that the compressed medicine-carrying hemostatic sponge with a specific shape (such as wedge shape, nail shape and the like) is obtained, and the hemostatic requirements of different wounds are met. Meanwhile, the compression-molded hemostatic sponge can basically realize adsorption balance within 60s, and the expansion volume reaches more than 6 times of the self volume, so that the compression-molded compression drug-loaded hemostatic sponge can quickly absorb blood and expand when being plugged into a wound, occupies and accumulates the space of a large-area wound, particularly a penetrating wound, and can play an effective compression hemostasis role; moreover, the compressed drug-loaded hemostatic sponge can absorb blood and swell, can reduce the possibility of bacteria invading into wounds, and is particularly suitable for the limbs and limb connection parts such as armpits, groins, necks and other parts which cannot utilize tourniquets or artificially press for hemostasis, thereby improving the survival rate of the casualties which can be saved in battlefields or major accident sites. In addition, the hemostatic sponge after compression molding is convenient to store, transport and carry, and can further facilitate emergency operation.

6) The hemostatic sponge material is light and soft in texture, is rich in elasticity, and cannot stimulate and rub wounds.

Drawings

FIG. 1 is a graph of the effect of lignocellulosic sponges prepared with different amounts of cellulose;

FIG. 2 is a scanning electron microscope image of a lignocellulosic sponge prepared with different amounts of cellulose;

FIG. 3 is a scanning electron microscope image of a lignocellulosic sponge prepared with different amounts of porogen;

FIG. 4 is a comparison of a lignocellulosic sponge before and after water absorption;

FIG. 5 is a graph of the response of a lignocellulosic sponge to water swelling versus time;

FIG. 6 is a stress-strain diagram of a drug loaded hemostatic sponge;

fig. 7 is a bacteriostatic experimental picture of the drug-loaded hemostatic sponge.

Detailed Description

The technical solution of the present invention will be further described with reference to the following specific examples.

The first embodiment is as follows: discussion experiment for determining dosage of lignocellulose

Preparation of lignocellulosic sponge

1) Taking sodium hydroxide and urea, and preparing an alkali urea solution with the alkali concentration of 7 wt% and the urea concentration of 12 wt%; placing the alkali urea solution in a cold trap for quick freezing for 10min to form ice slurry containing suspended ice crystal particles; adding lignocellulose into the alkali urea solution in an ice slurry state while stirring to prepare a lignocellulose solution;

2) placing the lignocellulose solution prepared in the step 1) in a cold trap for quick freezing for 10min to form an ice layer, then adding a pore-forming agent while stirring, performing injection molding, and then placing at-18 ℃ for freezing and aging for 48h to obtain gel;

3) soaking and washing the gel prepared in the step 2) in water for 2 days, changing water for 5-6 times during soaking to remove pore-forming agents, freezing at-18 ℃ for 24 hours for shaping, and freeze-drying to obtain the lignocellulose sponge.

To investigate the effect of the amount of lignocellulose on the properties of the prepared lignocellulose sponge, the inventors prepared lignocellulose according to the lignocellulose preparation method described in this example and made the following experiments (see table 1) for example 1-1, example 1-2, example 1-3, example 1-4 and example 1-5, respectively, with the amounts of lignocellulose corresponding to 1wt%, 2 wt%, 3wt%, 4 wt% and 5 wt%, respectively, based on the weight of the alkali urea solution.

TABLE 1 investigation experiment of lignocellulose amount

When the quantity of contrast lignocellulose is to the influence of lignocellulose sponge performance, mainly press stability from the compliance and the absorption of water of sponge (the absorption presses stability to be surveyed through the absorption of water press experiment, the specific operation of absorption press experiment is put into the sponge, press the sponge after the saturation of absorbing water, observe sponge shape recovery ability and press the process sponge and whether appear collapsing and breaking, if the sponge is pressed through absorbing water repeatedly, the shape is recovered well, and do not appear collapsing and breaking, explain sponge stable in structure) and synthesize and consider selecting the optimal lignocellulose quantity.

FIG. 1 is a graph of the effect of lignocellulosic sponges produced at different dosages of lignocellulose; wherein A is a lignocellulose sponge effect graph prepared by using different lignocellulose amounts, and the corresponding lignocellulose amounts from left to right are 1wt%, 2 wt%, 3wt%, 4 wt% and 5 wt%, respectively; b is an effect graph of pressing after the lignocellulose sponges prepared by different lignocellulose dosages absorb water and are saturated, and the corresponding lignocellulose dosages from left to right are 1wt%, 2 wt% and 3 wt%. From the softness of the prepared lignocellulose sponge, when the dosage of the lignocellulose is 1wt%, 2 wt% and 3wt% respectively, the softness of the prepared lignocellulose sponge is reduced sequentially along with the increase of the dosage of the lignocellulose, but the prepared lignocellulose sponge shows good elasticity and water absorption capacity, when the dosage of the lignocellulose is increased to 4 wt%, the concentration of a lignocellulose solution is obviously thickened, the lignocellulose is not easy to uniformly disperse, the obtained lignocellulose sponge is hard, and the water absorption reaction is slow. The lignocellulose sponges prepared according to the lignocellulose use amounts of 1wt%, 2 wt% and 3wt% are placed into water to absorb water and are saturated, and then the sponges are pressed, so that the lignocellulose sponges prepared according to the lignocellulose use amounts of 1wt% and 2 wt% all collapse and break, and the lignocellulose sponges prepared according to the lignocellulose use amounts of 3wt% are repeatedly pressed, so that the shape of the sponges is recovered well, good elasticity is shown, the structure of the sponges is stable, and the optimum lignocellulose addition amount is 3 wt%.

The sectional morphology of the lignocellulose sponge samples prepared in examples 1-1, 1-2 and 1-3 is observed by a Quanta200 scanning electron microscope, and the scanning electron microscope image is shown in fig. 2 (in fig. 2, the amounts of lignocellulose corresponding to A, B, C are 1wt%, 2 wt% and 3wt%, and D, E, F is an enlarged view of A, B, C). As can be seen from fig. 2, the pore size of the lignocellulose sponge gradually decreases with the increase of the amount of the lignocellulose, the structure becomes more compact, and the cellulose gradually changes from a sheet structure to a single fiber structure.

Example two: preparation of lignocellulosic sponge

The preparation method of the lignocellulose sponge comprises the following steps:

3) placing the foam prepared in the step 2) in a cold trap for quick freezing for 10min to form foam ice slurry, adding a pore-foaming agent into the foam ice slurry while stirring, performing injection molding, and performing freezing aging at-18 ℃ for 48h to obtain gel;

4) soaking and washing the gel prepared in the step 3) in water for 2 days, changing water for 5-6 times during soaking to remove the surfactant and the pore-forming agent, freezing at-18 ℃ for 24 hours for shaping, and freeze-drying to prepare the lignocellulose sponge.

Discussion experiment of surfactant dosage

To investigate the effect of the amount of surfactant on the properties of the prepared lignocellulosic sponges, the inventors prepared lignocellulosic sponges according to the method of preparation of lignocellulosic sponges described in this example, and made the following experiments (see table 2) with amounts of surfactant of 0, 0.1wt%, 0.5 wt%, 1wt%, 2 wt%, 3wt%, 4 wt%, 5 wt%, 6 wt%, 7 wt%, 8wt%, respectively. The results are shown in Table 3.

TABLE 2 investigation of surfactant amounts

TABLE 3 Density and porosity of lignocellulosic sponges made with different surfactant loadings

The amount of the surfactant (wt%)	0	0.1	0.5	1	2	4	6	8
									Density (mg/cm)³)	63.2	59.6	41.9	36.2	30.9	28.7	24.3	20.1
Porosity (%)	95.86	96.10	97.26	97.63	97.98	98.12	98.41	98.68

As can be seen from Table 3, the density of the lignocellulosic sponge increased from 63.2mg/cm with increasing amounts of surfactant (sodium lauryl sulfate)³Quickly reduced to 20.1mg/cm³The porosity is improved from 95.86 percent to 98.68 percent; the density of the lignocellulose sponge is reduced to below 1/3, which shows that the addition of the surfactant can rapidly improve the foamability of the cellulose solution, greatly increase the fluffiness and porosity of the cellulose sponge, reduce the density and obtain the cellulose spongeCan be more excellent lignocellulose sponge.

(II) investigation experiment of the amount of porogen used

To investigate the effect of the amount of the pore-forming agent on the properties of the prepared lignocellulose sponge, the inventors prepared lignocellulose sponges according to the preparation method of the lignocellulose sponge described in this example, and made the following experiments (see table 4) with the amount of the pore-forming agent of 40wt%, 50 wt%, 60 wt%, 70 wt%, and 80wt%, respectively. The results are shown in Table 5.

TABLE 4 investigation experiment of pore-foaming agent dosage

TABLE 5 Density and porosity of lignocellulosic sponges prepared with different amounts of porogen

The amount of surfactant used	40wt％	50wt％	60wt％	70wt％	80wt％
						Density (mg/cm)³)	42.7	37.8	30.9	24.6	18.7
Porosity (%)	97.21	97.53	97.98	98.39	98.77

As can be seen from Table 5, it is accompanied by a porogen (Na)₂SO₄) The dosage is gradually increased, and the density of the lignocellulose sponge is 42.7mg/cm³Reduced to 18.7mg/cm³The porosity increased from 97.21% to 98.77%. Surfactants increase the porosity and fluffiness of lignocellulosic sponges by increasing the foamability of the lignocellulosic solution, while porogens (Na)₂SO₄) The porosity and the fluffiness of the lignocellulose sponge are increased by occupying vacant sites in the lignocellulose solution by inorganic salt particles, and the two principles are different and supplement each other. The foam generated by the surfactant is unstable, the foam is gradually reduced along with the increase of the standing time of the solution, and a proper amount of Na is added into the solution₂SO₄Can occupy foam interstitial space rapidly, stabilize the foam, increase lignocellulose sponge's porosity and fluffiness simultaneously, the synergistic effect of mutually supporting of both can prepare the better lignocellulose sponge of performance.

FIG. 3 is a scanning electron microscope image of the lignocellulose sponges prepared by different amounts of pore-forming agent, wherein the pore-forming agent (Na) corresponds to a, b, c, d and e₂SO₄) Amounts are 40wt%, 50 wt%, 60 wt%, 70 wt%, 80wt%, respectively, f is a partial enlargement of e. As can be seen from fig. 3, with increasing use amount of the pore-forming agent, the lignocellulose composite sponge gradually changes into a fluffy form of interweaving single fibers, so that the pores are larger, the capillary effect is more obvious, and the water absorption capacity and the rapid reaction capacity of the lignocellulose composite sponge can be improved.

(III) investigation experiment for surfactant type and pore-forming agent type

In order to investigate the influence of the type and amount of the pore-forming agent on the properties of the prepared lignocellulose sponge, the inventors prepared the lignocellulose sponge according to the preparation method of the lignocellulose sponge described in this example, and performed the following experiments (see table 6).

TABLE 6 discussion of surfactant types and amounts

In the research process, several surfactants (sodium dodecyl sulfate, sodium dodecyl benzene sulfonate, alkyl glycoside, sodium fatty alcohol polyoxyethylene ether sulfate and fatty acid methyl ester sulfonate) in table 6 are added into the lignocellulose solution to show good foaming effect, and the foaming effect comparison is shown through observation: the foaming agent has certain influence on the density and porosity of the cellulose sponge, the better the foaming effect, the higher the porosity of the prepared lignocellulose sponge is, the lower the density is, and the higher the water absorption capacity and the water absorption rate are. The type of the pore-foaming agent has little influence on the prepared lignocellulose sponge. However, when sodium dodecyl benzene sulfonate is used as a surfactant, the foaming rate is high, so that the prepared lignocellulose sponge has low density, poor structural stability and easy cracking.

Example three: loading chitosan and hemostatic on lignocellulose sponge

Chitosan and hemostatic are loaded on the lignocellulose sponge by adopting an impregnation method, and the operation steps are as follows:

a. taking acid-soluble chitosan with deacetylation degree of more than 90%, dissolving the acid-soluble chitosan into 0.1 mol/L diluted acetic acid, and preparing a chitosan solution; adding a hemostatic drug into the chitosan solution, and uniformly stirring and dispersing to obtain a chitosan-hemostatic drug composite solution; wherein the mass fraction of chitosan in the chitosan solution is 0.1-2 wt%, and the mass fraction of hemostatic in the chitosan-hemostatic composite solution is 0.1-1 wt%;

b. soaking the lignocellulose sponges prepared in the examples 2-4 in the chitosan-hemostatic drug composite solution for 2 hours, taking out, freezing and shaping at-18 ℃, and then freezing and drying to obtain the drug-loaded hemostatic sponge.

In the method for preparing the drug-loaded hemostatic sponge by loading chitosan and hemostatic drugs on the lignocellulose sponge, specific setting conditions of various parameters are shown in each specific example in table 7.

TABLE 7 specific examples of loading chitosan and hemostatic drugs on lignocellulosic sponges by dipping

Example four: loading chitosan and hemostatic on lignocellulose sponge

Chitosan and hemostatic are loaded on the lignocellulose sponge by a spraying method, and the operation steps are as follows:

a. preparing water-soluble chitosan into a chitosan solution by using water-soluble chitosan; adding a hemostatic drug into the chitosan solution, and uniformly stirring and dispersing to obtain a chitosan-hemostatic drug composite solution; wherein the mass fraction of chitosan in the chitosan solution is 0.1-2 wt%, and the mass fraction of hemostatic in the chitosan-hemostatic composite solution is 0.1-1 wt%;

b. the surface of the lignocellulose sponge prepared in the embodiment 2-4 is sprayed with chitosan-hemostatic drug composite solution, and then the sprayed lignocellulose sponge is frozen and shaped at-18 ℃, and then is frozen and dried to obtain the drug-loaded hemostatic sponge.

In the method for preparing the drug-loaded hemostatic sponge by loading chitosan and hemostatic drugs on the lignocellulose sponge, specific setting conditions of various parameters are shown in each specific example in table 8.

TABLE 8 specific examples of chitosan and hemostatic drug loading on lignocellulosic sponges by spray coating

Performance test experiment

(first) Water absorption test

1. Water absorption swelling test

The water absorption properties of the lignocellulosic sponges prepared in examples 2-4 were investigated. The specific operation is as follows: the lignocellulose sponges prepared in examples 2 to 4 (cylindrical lignocellulose sponges with a diameter of 2cm and a height of 6 cm) were compressed into cylindrical compressed lignocellulose sponges with a diameter of 2cm and a height of 1cm, then the compressed lignocellulose sponges were placed in deionized water, and the water-swelling capacity of the compressed lignocellulose sponges was observed, and the results are shown in fig. 4. In fig. 4, a is compressed lignocellulose sponge that has not been soaked in water, and B is compressed lignocellulose sponge that has been placed in deionized water and has been soaked for 60 seconds to absorb water sufficiently. As can be seen from fig. 4, the height of the compressed lignocellulose sponge which is not absorbed with water is 1cm, and after 60 seconds of sufficient water absorption, the length of the compressed lignocellulose sponge becomes about 6cm, and the compressed lignocellulose sponge is basically saturated, which indicates that the compressed sponge can be rapidly expanded to about 6 times of the volume of the compressed sponge in a short time. Therefore, when the compressed lignocellulose sponge sheet is placed in a large artery wound, the compressed lignocellulose sponge sheet not only can rapidly absorb moisture in blood and enable the blood to be rapidly coagulated, but also can provide enough compression force effect for the sponge with the rapid expansion volume, so that the wound is blocked and bacterial infection is avoided, and meanwhile, the artery blood vessel is compressed to prevent secondary bleeding.

2. Water absorption swelling response-time determination experiment

The water absorption properties of the lignocellulosic sponges prepared in examples 2-4 were investigated. The specific operation is as follows: the lignocellulosic sponges prepared in examples 2-4 (cylindrical lignocellulosic sponges with a diameter of 2cm and a height of 6 cm) were compressed into cylindrical compressed lignocellulosic sponges with a diameter of 2cm and a height of 1 cm. Taking a compressed lignocellulose sponge with the diameter of 2cm and the height of 1cm, accurately weighing the compressed lignocellulose sponge by an analytical balance, and recording the weight as m₁The compressed lignocellulosic sponge was then immersed in deionized water and the compressed wood was measured at 10s, 20s, 30s, 60s, 90s, 120s, respectivelyWeight of cellulose sponge, noted m_tAnd then calculating the water absorption times of the lignocellulose composite sponge in each time period according to the following formula, and performing at least three groups of parallel tests on each group of samples. The results are shown in FIG. 5.

The water absorption multiple calculation formula is as follows:

as can be seen from FIG. 5, the compressed lignocellulose sponge has absorbed about 22 times of water by its own weight in about 10s, and is substantially saturated in about 60s and reaches about 25 times of its own weight, which indicates that the lignocellulose sponge has good rapid water absorption reaction capability, and when being applied to a wound, the compressed lignocellulose sponge can rapidly absorb the water in the excessive blood flowing out from the wound to rapidly coagulate the blood.

(II) stress-Strain test of drug-loaded hemostatic sponge

The stress-strain performance of the drug loaded hemostatic sponge was studied using the drug loaded hemostatic sponges prepared in examples 3-3. The specific operation is as follows:

compressing the drug-loaded hemostatic sponge prepared in example 3-3 (cylindrical drug-loaded hemostatic sponge with diameter of 2cm and height of 6 cm) into a cylindrical compressed drug-loaded hemostatic sponge with diameter of 2cm and height of 1 cm; the method comprises the steps of immersing a cylindrical compressed medicine-carrying hemostatic sponge with the diameter of 2cm and the height of 1cm into deionized water for 90 seconds to fully absorb water to reach water saturation, taking out the medicine-carrying hemostatic sponge after water saturation, and testing the compression strength of the medicine-carrying hemostatic sponge by using a TA-XT PLUS type texture analyzer, wherein the compression rate is 0.6mm/min, and the test result is shown in figure 6.

As can be seen from fig. 6, the drug loaded hemostatic sponge can provide an acting force of about 5kPa when being deformed by 10%, an acting force of about 10kPa when being deformed by 30%, an acting force of about 20kPa when being deformed by 50%, and an acting force of about 46kPa when being deformed by 70%. The internal pressure of the human artery blood vessel is about 10-15 kPa, when the artery is damaged, the loss of the blood in the body can be accelerated under the action of the pressure in the blood vessel, and at the moment, the blood vessel needs to be pressed by external acting force to resist the action of the pressure in the blood vessel, so that the loss of the blood is reduced. When the lignocellulose composite sponge absorbs the water in the blood and expands, enough acting force can be effectively provided to act on the artery blood vessel, and the blood loss rate is effectively reduced.

(III) measurement of blood coagulation time

The hemostatic ability of the drug-loaded hemostatic sponge is judged by adopting the blood coagulation time of a test tube method.

The specific experimental operation is as follows: respectively taking the same mass of the hemostatic sponges prepared by the example 2-4, the example 3-1, the example 3-2, the example 3-3, the example 3-4 and the example 3-5, and then compressing the hemostatic sponges according to the same compression ratio to obtain compressed hemostatic sponges; 0.05g of each compressed hemostatic sponge is weighed respectively and sent to the bottom of a glass test tube with the diameter of 2cm, and three parallel samples are arranged for each compressed hemostatic sponge. Collecting 3mL of fresh blood, slowly injecting the blood into the test tube along the wall of the test tube, wherein each tube is 1mL, gently mixing for 3s, then placing the test tube into an electric heating constant-temperature water bath kettle at 37 ℃, inclining the test tube every 5s until the blood in the test tube does not flow, and recording the whole blood coagulation time. The specific results are shown in Table 9.

TABLE 9 clotting times of different sponges

As can be clearly seen from table 9, the hemostasis time of the common lignocellulose sponge is 270 ± 5s, because a large number of hydroxyl groups are distributed on the surface of the lignocellulose sponge, no aggregation is caused on platelets, red blood cells and the like, and the hemostasis capability of the common lignocellulose sponge mainly depends on the water absorption capability of the cellulose sponge, so that the common lignocellulose sponge can quickly absorb water in blood to play a role in coagulating platelets and red blood cells, thereby achieving the effect of blood coagulation and having long hemostasis time; the blood coagulation time of the lignocellulose sponge loaded with 0.5 wt% of hemostatic drugs is reduced to 165 +/-15 s, and the hemostatic time is shortened mainly by the hemostatic drugs; the hemostatic time of the drug-loaded hemostatic sponge is gradually shortened along with the increase of the mass fraction of chitosan, when the mass fraction of a chitosan solution is 2 wt%, the hemostatic time is 30 +/-5 s, which is mainly attributed to the better hemostatic capacity of chitosan, the surface of chitosan is rich in positive charge groups, and more prominently amino groups, so that the chitosan can fully act with negative charges on the surfaces of erythrocytes and platelets, and the erythrocytes, the platelets and the like are aggregated, and the aim of quickly stopping bleeding is fulfilled; although the blood coagulation time can be effectively reduced by increasing the chitosan concentration, the viscosity of the solution gradually increases with the increase of the chitosan concentration, when the mass fraction of the chitosan solution is 1wt%, the solution is relatively uniform and has good fluidity, and when the chitosan concentration is 2 wt%, the viscosity of the chitosan solution is sharply increased, the fluidity of the solution is poor, the subsequent spraying or soaking is not facilitated, and therefore the mass fraction of the chitosan solution is optimal to be 1 wt%.

(IV) antibacterial property test

The antibacterial property of the drug-loaded hemostatic sponge against staphylococcus aureus (s.aureus), pseudomonas aeruginosa (p.aeruginosa) and escherichia coli (e.coli) was tested by using a plate colony counting method on the drug-loaded hemostatic sponges prepared in examples 3-1, 3-2, 3-3 and 3-4.

Taking staphylococcus aureus as an example, the specific experimental operations are as follows: the drug-loaded hemostatic sponges prepared in the examples 3-1, 3-2, 3-3 and 3-4 were cut into round pieces with the thickness of 1mm and the diameter of 10mm, and then irradiated under an ultraviolet lamp for sterilization for 5 min. 10 μ L of a suspension of Staphylococcus aureus (. about.10)⁶CFU/mL) was gently applied to the surface of each drug loaded hemostatic sponge, and each disc was immersed in 10mL of PBS (pH 7.4) solution and incubated at 37 ℃ for 24 h. Then, gently taking out each wafer, respectively putting the wafer into 1mL of PBS buffer solution, and then carrying out vortex rotation for 2min to fully transfer bacteria in the wafer into the PBS buffer solution; PBS buffer was performed 10⁵Diluting, uniformly coating 100 mu L of diluted PBS buffer solution in an agar culture medium, and culturing for 24h at 37 ℃; the control group used 1mm thick, 10mm diameter circular paper sheets. Each set of experiments was performed in 3 parallel sets. The results are shown in FIG. 7.

The experimental operation for detecting the antibacterial property of the drug-loaded hemostatic sponge to pseudomonas aeruginosa and escherichia coli is the same as that of staphylococcus aureus, and is not described herein again.

As can be seen from FIG. 7, when chitosan was not added to the cellulose sponge, the bacterial colonies of Staphylococcus aureus, Pseudomonas aeruginosa and Escherichia coli were measured to be 5.5 × 10⁶About one, basically has no antibacterial effect, when 0.1wt% of chitosan is added, the number of bacterial colonies is greatly reduced, and the number of bacterial colonies of staphylococcus aureus is 2 × 10⁶About one, the number of colonies of Pseudomonas aeruginosa was 1.5 × 10⁶About, the number of Escherichia coli colonies was 1 × 10⁶About, along with the increase of the chitosan concentration, the antibacterial capacity of the drug-loaded hemostatic sponge is obviously improved, and when the mass fraction of the chitosan is 2 wt%, the colony number of the three bacteria is 3 × 10³About one, the bacteriostatic ability reaches more than 99 percent, which shows that the composite sponge has better antibacterial ability.

The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the present invention, but rather as the following description is intended to cover all modifications, equivalents and improvements falling within the spirit and scope of the present invention.

Claims

1. The preparation method of the medicine-carrying hemostatic sponge is characterized by comprising the following steps:

(1) preparing a lignocellulose sponge:

1) preparing an alkali urea solution, and freezing to form ice slurry; adding lignocellulose into the alkali urea solution in the ice slurry state by stirring to prepare a lignocellulose solution;

(2) loading chitosan and hemostatic drug on the lignocellulose sponge:

2. The preparation method of the drug-loaded hemostatic sponge according to claim 1, wherein in step 1), the weight of the added lignocellulose is 1-3 wt% of the alkali urea solution; in the step 2), the weight of the added surfactant is 0.1-8 wt% of the weight of the lignocellulose solution; in the step 3), the weight of the added pore-foaming agent is 40-80 wt% of the weight of the lignocellulose solution.

3. The preparation method of the drug-loaded hemostatic sponge according to claim 1, wherein the surfactant is one or more of sodium dodecyl sulfate, sodium dodecyl benzene sulfonate, alkyl glycoside, sodium fatty alcohol-polyoxyethylene ether sulfate, and fatty acid methyl ester sulfonate.

4. The method for preparing the drug-loaded hemostatic sponge according to claim 1, wherein the pore-forming agent is one or more of sodium sulfate decahydrate, anhydrous sodium sulfate and anhydrous magnesium sulfate.

5. The method for preparing a drug-loaded hemostatic sponge according to claim 1, wherein in step a, the chitosan is acid-soluble chitosan with a deacetylation degree of more than 90%, and the acid-soluble chitosan is prepared into a chitosan solution by using 0.01-0.5 mol// L of dilute acetic acid as a solvent.

6. The method for preparing a drug-loaded hemostatic sponge according to claim 1, wherein in step a, the chitosan is water-soluble chitosan, and the water-soluble chitosan is one or more of carboxymethyl chitosan, hydroxypropyl chitosan, and 2-hydroxypropyl trimethyl ammonium chloride chitosan.

7. The preparation method of the drug-loaded hemostatic sponge according to claim 1, wherein in step a, the mass fraction of chitosan in the chitosan solution is 0.1wt% to 1 wt%; the mass fraction of the hemostatic medicament in the chitosan-hemostatic medicament composite solution is 0.1-1 wt%.

8. The preparation method of the drug-loaded hemostatic sponge according to claim 1, wherein in step a, the hemostatic drug is one or more of vitamin K1, vitamin K3, vitamin K4, acetic glycine ethylenediamine, carbachol, carbazochrome sodium sulfonate, Yunnan white drug powder, and thrombin.

9. A drug-loaded hemostatic sponge product made by the method of making a drug-loaded hemostatic sponge of any one of claims 1-8.