CN103418023B - Multilayer composite hemostatic material and preparation method thereof - Google Patents

Abstract

The invention provides a multilayer composite hemostatic material, which is characterized in that the material is formed by compounding a PLGA hollow nano fiber layer, a gelatin hollow nano fiber layer, a chitosan hollow nano fiber layer and an alginate nano fiber layer through mould pressing at the periphery. The multilayer composite hemostatic material has the advantages that the defects that the fiber hemostatic material prepared through electrostatic spinning is thinner and softer are overcome, the thickness of the material is increased, the compression hemostasis effect is improved, the defects that the traditional hemostatic material has a single variety, is narrow in application range, and cannot be applied to various complex bleeding situations are overcome, and the application range is widened. In addition, the multilayer composite hemostatic material provided by the invention takes the material shape with the soft middle part and hard periphery, easy to take for use, and can be applied in an operation more conveniently.

Description

A kind of multi-layer composite hemostatic material and preparation method thereof

technical field

The invention relates to a multilayer composite hemostatic material and a preparation method thereof, belonging to the field of medicines. the

Background technique

During surgery and trauma, there is usually a lot of bleeding. Conventional treatment methods include manual pressing, cauterization, suture, etc., but they cannot always effectively control the loss of blood. Traditional hemostatic agents, such as gelatin-based powders, sponges, and fabrics, provide a grid for natural thrombus formation, but tend to stimulate the patient's endogenous hemostatic cascade, which can easily lead to embolism and local inflammation. sexual response. Therefore, there is a need for a bioabsorbable hemostatic material with excellent hemostatic effect that is suitable for various operations and field trauma treatment to replace traditional hemostatic materials, especially in heart surgery, spine surgery, liver transplantation or tumor removal, and neurosurgery. Among them, the hemostatic speed and hemostatic effect of the hemostatic material are particularly important. the

At present, a number of patents and a large number of literatures have reported the development of new hemostatic materials, and the types and preparation techniques of the developed materials are also varied. In these patents and documents, there are a lot of similarities, such as the selection of hemostatic materials is nothing more than several common hemostatic materials such as sodium carboxymethyl cellulose materials (ZL201210563634.9 sodium alginate and sodium carboxymethyl cellulose Blended fiber and its preparation and application; 201110328419.6 medical wet composite dressing and its manufacturing method), gelatin materials (ZL200910092911.0 a multi-purpose gelatin fiber and its preparation method; ZL200910217762.6 a tissue-engineered cornea with controllable degradation fiber scaffold and preparation method), chitosan material (ZL201010185135.1 hemostatic agent for local and internal use; ZL201020688022.9 a chitosan fiber medical dressing; ZL201110044474.2 chitosan emergency hemostatic material), alginate materials (ZL201110081468.4 spinning stock solution and method of manufacturing biomedical fiber; ZL200810100981.1 preparation method of pure sodium alginate nanofiber membrane material), and the uniform composite or blended material of these compounds combined with each other, the material mainly adopts Prepared by methods such as wet spinning, freeze-drying, and electrospinning. the

Among them, the electrospinning method is a simple, convenient and cheap method for producing nanofiber film materials, and the diameter of the prepared fibers can be adjusted and changed arbitrarily between tens of nanometers and hundreds of nanometers. Compared with traditional fiber fabrics, this kind of nanofiber has a larger specific surface area, so it has better adsorption performance. However, the fatal defect of electrospinning materials is that only very thin fiber membrane materials can be obtained. It cannot form an effective compression hemostatic effect, which limits its practical application in clinical medicine. the

Contents of the invention

The object of the present invention is to provide a multi-layer composite hemostatic material with outstanding hemostatic effect and a preparation method thereof. the

The multi-layer composite hemostatic material of the present invention is characterized in that it is composed of PLGA hollow nanofiber layer, gelatin hollow nanofiber layer, chitosan hollow nanofiber layer and alginate nanofiber layer through molding and compounding. the

In addition, hemostatic drugs, anti-infection drugs, analgesic drugs or tissue repair promoting factors can also be filled between the layers of the multi-layer composite hemostatic material according to needs. The thickness of the multi-layer composite hemostatic material of the present invention can be adjusted arbitrarily according to actual needs, generally between 0.3-3cm. the

The method for preparing the above-mentioned multilayer composite hemostatic material of the present invention is characterized in that it comprises the following steps:

(1) Preparation of PLGA hollow nanofiber web by coaxial spinning method

Use chloroform, DMF, THF or a mixture of DMF and THF as a solvent to prepare a PLGA spinning stock solution with a concentration of 0.5-1.5g/ml. This solution is the shell solution, and the core solution uses sesame oil, mineral oil, dimethyl Silicone oil or blended edible oil, the flow rate of the shell layer solution is 0.5-2.0ml/h, the flow rate of the core layer solution is 0.2-1.0ml/h, the solution flow rate is precisely controlled by the syringe pump, the spinning voltage is adjusted between 10-20kV, and the collected The distance is 5-15cm, the collection time is 8-72 hours, the thickness of the collected fiber web is continuously adjustable from several hundred microns to 3 mm, first put the collected fiber web into a high-speed centrifuge (for example, 30,000 rpm/ Part) centrifugal deoiling, then extract the oil component with cyclohexane or carbon tetrachloride, and finally vacuum dry to obtain pure PLGA hollow nanofiber network;

(2) Preparation of gelatin hollow nanofiber network by coaxial spinning method

Formic acid or water is used as a solvent to prepare a gelatin spinning stock solution with a concentration of 15-30wt%. This solution is a shell solution, and the core solution uses sesame oil, mineral oil, simethicone or blended edible oil. The flow rate of the shell solution is 0.5-1.0ml/h, the flow rate of the core layer solution is 0.2-0.5ml/h, the solution flow rate is precisely controlled by the syringe pump, the spinning voltage is adjusted between 10-30kV, the collection distance is 5-15cm, and the collection time is 4- For 72 hours, the ambient temperature during spinning is controlled above 40°C, and the thickness of the collected fiber web is continuously adjustable from several hundred microns to 3 mm. First, the collected fiber web is placed in a high-speed centrifuge (such as 30,000 rpm /min) to deoil by centrifugation, then extract the oil component with cyclohexane, and finally vacuum-dry to obtain pure gelatin hollow nanofiber network;

(3) Preparation of chitosan hollow nanofiber network by coaxial spinning method

Concentrated acetic acid or trifluoroethanol is used as a solvent to prepare a chitosan spinning stock solution with a concentration of 5-30wt%. This solution is the shell solution, and the core solution uses sesame oil, mineral oil, simethicone or blended edible oil. The flow rate of the shell solution is 10-40μl/h, the flow rate of the core layer solution is 5-20μl/h, the solution flow rate is precisely controlled by the syringe pump, the spinning voltage is adjusted between 10-30kV, the collection distance is 5-15cm, the collection time For 4-72 hours, the thickness of the collected fiber web is continuously adjustable from several hundred microns to 3 mm. The oil component is extracted with cyclohexane, and finally vacuum-dried to obtain pure chitosan hollow nanofiber network;

(4) Preparation of alginate nanofiber web by electrospinning method

It can be prepared by patent and literature methods. For details, please refer to the preparation method of Chinese patent ZL200810100981.1 pure sodium alginate nanofiber membrane material and the literature Biomacromolecules2008, 9, 1362-1365. The specific steps are: mix alginate and glycerin at a weight-to-volume ratio of 2-4g/ml, then add solvent water to the mixture, wherein the volume ratio of water to glycerin is 0.5-1:1, stir evenly, and stand still Remove air bubbles to obtain spinning stock solution, the flow rate of spinning stock solution is 50-200μl/h, the solution flow rate is precisely controlled by a syringe pump, the spinning voltage is adjusted between 10-40kV, the distance between the spinneret and the receiver is 5-15cm, The receiver is immersed in a coagulation bath composed of CaCl ₂ aqueous solution and absolute ethanol. The collection time is 36-72 hours. The thickness of the collected fiber web is continuously adjustable between several hundred microns and 2 mm. The net is vacuum-dried to obtain the alginate nanofiber net;

(5) Composite layers of materials by molding

Take the prepared fiber nets, superimpose them together, and fill hemostatic drugs, anti-infective drugs, pain-relieving drugs or factors promoting tissue repair between the layers according to the needs, and use a hollow mold to place them Press it into a shape with hard surroundings and soft middle of the specified size, and then cut it with a knife, where the molding pressure is 100-400N/cm ² (varies with the thickness of the sample), and the number of layers of each fiber web is 1 or more ;

(6) Disinfection treatment

After the obtained composite material is sterilized by gamma-rays, a multi-layer composite hemostatic material is obtained. the

In at least any one of the coaxial spinning shell solutions in steps (1)-(3), it is also preferred to further add single-wall or multi-wall carbon nanotubes to play the role of toughening and antibacterial. In step (5), various drugs can be added using electrostatic atomization technology. the

The multi-layer composite hemostatic material of the present invention overcomes the shortcomings of thinner and softer fiber hemostatic materials prepared by electrospinning, increases the thickness of the material, improves the hemostatic effect of compression, and improves the single variety and narrow application range of traditional hemostatic materials. , The shortcomings of being unable to deal with a variety of complex bleeding situations have expanded its application range. In addition, the multi-layer composite hemostatic material of the present invention is formed into a material shape with a soft center and a hard periphery, which is easy to take and can be used more conveniently in surgery. the

Description of drawings

Fig. 1 is a schematic diagram showing the appearance of the multilayer composite hemostatic material of the present invention. the

Fig. 2 is a schematic diagram showing the cross-sectional structure of the multilayer composite hemostatic material of the present invention. the

3 is a SEM image of the gelatin hollow nanofibrous web material prepared in Example 1. the

Figure 4 is the PLGA hollow nanofibrous web material prepared in Example 1, where (a) is a physical photo, (b) and (c) are SEM images of the front and section, respectively. the

Fig. 5 is the SEM image of the chitosan hollow nanofibrous web material prepared in Example 1. the

Detailed ways

The multi-layer composite hemostatic material of the present invention is characterized in that it is composed of PLGA hollow nanofiber layer, gelatin hollow nanofiber layer, chitosan hollow nanofiber layer and alginate nanofiber layer through molding and compounding. That is, the multi-layer composite hemostatic material of the present invention is a three-dimensional structure in which the middle layer is formed by molding technology and the surrounding layers are not layered. the

Fig. 1 is a schematic diagram showing the appearance of the multilayer composite hemostatic material of the present invention, and Fig. 2 is a schematic diagram showing the cross-sectional structure of the multilayer composite hemostatic material of the present invention. As shown in Figures 1 and 2, the multilayer composite hemostatic material of the present invention has a hard part A (that is, the part that has been molded around) and a soft part B (that is, the part that has not been molded in the middle), and layers 1-4 respectively represent the parts of the present invention. Each layer of the multi-layer composite hemostatic material, that is, the PLGA hollow nanofiber layer, the gelatin hollow nanofiber layer, the chitosan hollow nanofiber layer and the alginate nanofiber layer, has no special restrictions on the order of the layers, and can be customized according to the needs. It can be adjusted arbitrarily, but it is preferable to arrange the layers in the order of gelatin, alginate, chitosan, and PLGA. This is because gelatin fibers and alginate fibers have strong hygroscopicity and can be adsorbed on the wound surface in an instant to form a coagulate. The gel state is conducive to the rapid hemostasis of the wound, and the network structure of chitosan fibers and PLGA fibers (especially PLGA fibers) has a special support effect on the wound, which can reduce the secondary tearing and wound adhesion of the wound. the

The multi-layer composite hemostatic material of the present invention has at least the above-mentioned 4-layer structure, and the number and ratio of each fiber layer can be adjusted according to the mechanical properties required for practical application and the required hemostatic effect. the

In addition, as shown in Figure 2, hemostatic drugs, anti-infective drugs, analgesic drugs or tissue repair promoting factors can also be filled between the layers of the multi-layer composite hemostatic material of the present invention as required, that is, this The inventive multi-layer composite hemostatic material can further contain a drug layer 5 . Examples of hemostatic drugs include batrotine, carbosulfonate, spinyne, hemostatin, and hemostatic aromatic acid; examples of anti-infective drugs include gentamicin, lincomycin, and erythromycin; Lidocaine hydrochloride etc. are mentioned as an analgesic drug. In addition, the thickness of the multi-layer composite hemostatic material of the present invention can be adjusted arbitrarily according to actual needs, generally between 0.3-3 cm. the

The method for preparing the above-mentioned multilayer composite hemostatic material of the present invention is characterized in that it comprises the following steps:

(1) Preparation of PLGA hollow nanofiber web by coaxial spinning method

Use chloroform, DMF, THF or a mixture of DMF and THF as a solvent to prepare a PLGA spinning stock solution with a concentration of 0.5-1.5g/ml. This solution is the shell solution, and the core solution uses sesame oil, mineral oil, dimethyl Silicone oil or blended edible oil, the flow rate of the shell layer solution is 0.5-2.0ml/h, the flow rate of the core layer solution is 0.2-1.0ml/h, the solution flow rate is precisely controlled by the syringe pump, the spinning voltage is adjusted between 10-20kV, and the collected The distance is 5-15cm, the collection time is 8-72 hours, the thickness of the collected fiber web is continuously adjustable from several hundred microns to 3 mm, first put the collected fiber web into a high-speed centrifuge (for example, 30,000 rpm/ Centrifugal deoiling, then extract the oil component with cyclohexane or carbon tetrachloride, and finally vacuum dry to obtain pure PLGA hollow nanofiber network;

(2) Preparation of gelatin hollow nanofiber network by coaxial spinning method

Formic acid or water is used as a solvent to prepare a gelatin spinning stock solution with a concentration of 15-30wt%. This solution is a shell solution, and the core solution uses sesame oil, mineral oil, simethicone or blended edible oil. The flow rate of the shell solution is 0.5-1.0ml/h, the flow rate of the core layer solution is 0.2-0.5ml/h, the solution flow rate is precisely controlled by the syringe pump, the spinning voltage is adjusted between 10-30kV, the collection distance is 5-15cm, and the collection time is 4- For 72 hours, the ambient temperature during spinning is controlled above 40°C, and the thickness of the collected fiber web is continuously adjustable from several hundred microns to 3 mm. First, the collected fiber web is placed in a high-speed centrifuge (such as 30,000 rpm /min) to deoil by centrifugation, then extract the oil component with cyclohexane, and finally vacuum-dry to obtain pure gelatin hollow nanofiber network;

(3) Preparation of chitosan hollow nanofiber network by coaxial spinning method

Concentrated acetic acid or trifluoroethanol is used as a solvent to prepare a chitosan spinning stock solution with a concentration of 5-30wt%. This solution is the shell solution, and the core solution uses sesame oil, mineral oil, simethicone or blended edible oil. The flow rate of the shell solution is 10-40μl/h, the flow rate of the core layer solution is 5-20μl/h, the solution flow rate is precisely controlled by the syringe pump, the spinning voltage is adjusted between 10-30kV, the collection distance is 5-15cm, the collection time For 4-72 hours, the thickness of the collected fiber web is continuously adjustable from several hundred microns to 3 mm. The oil component is extracted with cyclohexane, and finally vacuum-dried to obtain pure chitosan hollow nanofiber network;

(4) Preparation of alginate nanofiber web by electrospinning method

It can be prepared by patent and literature methods. For details, please refer to the preparation method of Chinese patent ZL200810100981.1 pure sodium alginate nanofiber membrane material and the literature Biomacromolecules2008, 9, 1362-1365. The specific steps are: mix alginate and glycerin at a weight-to-volume ratio of 2-4g/ml, then add solvent water to the mixture, wherein the volume ratio of water to glycerin is 0.5-1:1, stir evenly, and stand still Remove air bubbles to obtain spinning stock solution, the flow rate of spinning stock solution is 50-200μl/h, the solution flow rate is precisely controlled by a syringe pump, the spinning voltage is adjusted between 10-40kV, the distance between the spinneret and the receiver is 5-15cm, The receiver is immersed in a coagulation bath composed of CaCl ₂ aqueous solution and absolute ethanol. The collection time is 36-72 hours. The thickness of the collected fiber web is continuously adjustable between several hundred microns and 2 mm. The net is vacuum-dried to obtain the alginate nanofiber net;

(5) Composite layers of materials by molding

Take the prepared fiber nets, superimpose them together, and fill hemostatic drugs, anti-infective drugs, pain-relieving drugs or factors promoting tissue repair between the layers according to the needs, and use a hollow mold to place them Press it into a shape with hard surroundings and soft middle of the specified size, and then cut it with a knife, where the molding pressure is 100-400N/cm ² (varies with the thickness of the sample), and the number of layers of each fiber web is 1 or more ;

(6) Disinfection treatment

After the obtained composite material is sterilized by gamma-rays, a multi-layer composite hemostatic material is obtained. the

In at least any one of the coaxial spinning shell solutions in steps (1)-(3), it is also preferred to further add single-wall or multi-wall carbon nanotubes to play the role of toughening and antibacterial. In step (5), various drugs can be added using electrostatic atomization technology. the

The multilayer composite hemostatic material of the present invention has the following beneficial effects:

(1) Combining with multi-layer molding technology, it overcomes the shortcomings of thin (thickness <1mm) and soft hemostatic fiber material prepared by electrospinning, increases the thickness of the material, and improves the hemostatic effect of compression. the

(2) Various drugs of different varieties and different doses are added between layers according to needs, which improves the shortcomings of traditional hemostatic materials such as single variety, narrow application range, and inability to deal with various complicated bleeding situations, expands its application range, and reduces Anaphylaxis and rejection of patients during surgery. the

(3) Through mold pressing, it forms a material shape with a soft middle and a hard periphery, which is easy to access and can be used more conveniently in surgery. the

(4) PLGA fiber has good mechanical properties. After gelatin fiber, chitosan fiber and alginate fiber absorb water to form a gel, it can still maintain the original fiber network structure in the dressing, maintain a wet environment, and The formed gel film can be placed on the wound site for a long time without being easily damaged. When the wound enters the stage of granulation growth, PLGA fibers can serve as a skeleton, providing space and nutrient exchange environment for granulation growth. In the last stage of wound healing, that is, the epithelialization and regeneration stage, the network structure of PLGA fibers is very conducive to the growth and adhesion of epithelial tissue and promotes wound healing. For wounds with severe exudation, the network structure of PLGA fibers has special effects on some wounds that need support, and can also reduce secondary tearing of wounds. the

(5) The use of nanofibers, especially hollow nanofibers, increases the specific surface area of fibers and improves the adsorption performance and drug loading capacity of hemostatic materials. the

(6) Degradable biomedical materials are used, which can be automatically degraded after surgery, and can prevent adhesion between tissues and speed up wound repair. the

Example

The present invention is further illustrated below by way of examples. However, the present invention is not limited by this embodiment, and appropriate changes can be made within the range consistent with the purpose of the present invention, and these are all included in the technical scope of the present invention. the

1. Preparation of multilayer composite hemostatic material

Embodiment 1 (preparation of No. 1 sample)

(1) Preparation of gelatin hollow nanofiber web material

A coaxial spinning needle was used in the spinning experiment, and its specification was that the inner diameter of the inner needle was 0.3 mm, and the inner diameter of the outer needle was 1.2 mm. Gelatin is used as raw material and formic acid is used as solvent. Dissolve gelatin in formic acid at room temperature. The stirred solution is uniform and transparent. The mass fraction of gelatin is 20%. This solution is the shell solution of coaxial spinning. Nucleus solution uses sesame oil. The above solution was left to remove air bubbles as the spinning stock solution, and injected into the syringe. The flow rate of the shell layer solution was 0.6ml/h, and the flow rate of the core layer solution was 0.3ml/h. The solution flow rate was precisely controlled by a syringe pump. Under the conditions of 22kV voltage, the distance between the spinneret and the receiver (aluminum film) was 12cm, the electrospinning was carried out, the collection time was 48 hours, and the ambient temperature during the spinning was controlled above 40°C. The collected hollow fiber web materials The thickness is about 2mm. First put the collected hollow fiber web material into a high-speed centrifuge at 30,000 rpm to deoil, then extract the oil component with cyclohexane, and finally vacuum dry for 8 hours to obtain pure gelatin hollow nanofibrous web material . the

(2) Preparation of sodium alginate nanofiber web material

Sodium alginate is used as raw material, and the modifier is glycerin. Sodium alginate is dispersed in glycerin at room temperature, wherein the ratio of sodium alginate to glycerin is 4g/ml by weight and volume, and then water is added to the mixture, of which the water The volume ratio to glycerin is 1:1, stir evenly, remove air bubbles at rest, and obtain spinning stock solution. When the voltage is 22kV, the flow rate is 92μl/h, the distance between the spinneret and the receiver (copper mesh) is 10cm, and the receiver is soaked in a coagulation bath (the coagulation bath is composed of a CaCl ₂ aqueous solution with a mass percentage concentration of 6% and a mixed solution In a mixed solution composed of 80% absolute ethanol by mass percentage), electrospinning was carried out, the collection time was 48 hours, and the thickness of the collected fiber web material was about 1 mm. The obtained material was vacuum-dried for 8 hours to obtain a sodium alginate nanofibrous web material.

(3) Preparation of PLGA hollow nanofiber web material

A coaxial spinning needle was used in the spinning experiment, and its specification was that the inner diameter of the inner needle was 0.3 mm, and the inner diameter of the outer needle was 1.2 mm. Using PLGA (50:50, M _w =120,000) as the raw material, using the mixture of DMF and THF as the solvent (the volume ratio of DMF and THF is 1:1), dissolve PLGA in the mixture of DMF and THF at room temperature In the process, the concentration of PLGA was prepared to be 1.0g/ml, stirred evenly, and left to remove air bubbles to obtain the spinning stock solution, which was the shell solution of coaxial spinning. Nucleus solution uses sesame oil. The above solution was injected into the syringe, and the flow rate of the solution was precisely controlled by a syringe pump, the flow rate of the shell layer solution was 1.0 ml/h, and the flow rate of the core layer solution was 0.6 ml/h. The spinning voltage was 15kV, the distance between the spinneret and the receiver (aluminum film) was 15cm, the collection time was 48 hours, and the thickness of the collected hollow fiber web material was about 1.5mm. First put the obtained material into a high-speed centrifuge at 30,000 rpm to deoil, then extract the oil component with cyclohexane, and finally vacuum-dry for 8 hours to obtain a pure PLGA hollow nanofiber web material.

(4) Preparation of chitosan hollow nanofiber mesh material

Using chitosan (molecular weight 106,000g/mol) as raw material and concentrated acetic acid (aqueous solution with a mass fraction of 90% acetic acid) as a solvent, dissolve chitosan in concentrated acetic acid at room temperature to prepare a mass fraction of 6%. Chitosan solution, stirred evenly, and left to stand at room temperature to remove air bubbles, this solution is the shell solution of coaxial spinning. Nucleus solution uses sesame oil. The solution flow rate is precisely controlled by a syringe pump, the shell solution flow rate is 20 μl/h, and the core layer solution flow rate is 10 μl/h. The spinning voltage is 30kV, the distance between the spinneret and the receiver (aluminum film) is 10cm, the collection time is 48 hours, and the thickness of the collected hollow fiber web material is about 1.5mm. First put the material into a high-speed centrifuge at 30,000 rpm for deoiling, then extract the oil component with cyclohexane, and finally vacuum-dry for 8 hours to obtain a pure chitosan hollow nanofiber net material. the

(5) Molding of composite materials

Take one portion of the fiber mesh materials prepared above, stack the layers together in the order of gelatin, alginate, chitosan, and PLGA, and use a hollow mold (outer diameter 1.0cm×1.0cm, inner diameter 0.6cm ×0.6cm), under the pressure of 180N/ ^cm2 , press on the tablet machine for 3 minutes, take it off, cut the excess part with a knife, and form a shape with hard surrounding and soft middle, of which the soft part is 0.6cm×0.6cm , the hard part is around the soft part, and the width of each side is 0.2cm. The thickness of the soft part after pressing is about 7mm, and the thickness of the hard part is about 4mm.

(6) Disinfection treatment

After sterilizing with γ-rays, sample No. 1 was obtained. the

Embodiment 2 (preparation of No. 2 sample)

(1) Preparation of gelatin hollow nanofiber web material

In the preparation of the spinning stock solution, the mass fraction of the gelatin solution was set to 16%; during coaxial spinning, the flow rate of the shell solution was set to 0.5ml/h, the flow rate of the core layer solution was set to 0.2ml/h, and the spinning voltage Set it to 15kV, set the collection time to 36 hours, and the others are the same as in Example 1. The thickness of the collected web material was about 1.5 mm. the

(2) Preparation of sodium alginate nanofiber web material

In the preparation of the spinning stock solution, the weight-to-volume ratio of sodium alginate and glycerin was set to 3g/ml, and the volume ratio of water to glycerin was set to 0.5:1; during electrospinning, the voltage was set to 28kV, and the flow rate was set to 100μl/ml. h, the spinneret and the receiver distance are set to 12cm, and the coagulation bath is a CaCl _aqueous solution of 10% by mass percentage concentration and a mixed solution of 80% dehydrated alcohol in the mixed solution, and the others are the same as in Example 1 . The thickness of the collected web material was about 1 mm.

(3) Preparation of PLGA hollow nanofiber web material

In the preparation of the spinning stock solution, the concentration of the PLGA solution was set to 0.5g/ml; during coaxial spinning, the shell solution flow rate was set to 0.6ml/h, the core layer solution flow rate was set to 0.3ml/h, and the spinning voltage was set to Be 10kV, other with embodiment 1. The thickness of the collected fibrous web material was about 1 mm. the

(4) Preparation of chitosan hollow nanofiber mesh material

The shell solution is prepared as a chitosan solution with a mass fraction of 7%, and the others are the same as in Example 1. The thickness of the collected web material was about 1.5 mm. the

(5) Molding of composite nanomaterials

The molding pressure is set at 100 N/cm ² , and the thickness of the soft part is about 6 mm, and the thickness of the hard part is about 4 mm. Others are the same as in Embodiment 1.

(6) Disinfection treatment

With embodiment 1. the

Embodiment 3 (preparation of No. 3 sample)

(1) Preparation of gelatin hollow nanofiber web material

In the preparation of the spinning stock solution, the mass fraction of the gelatin solution was set to 18%; during coaxial spinning, the flow rate of the shell layer solution was set to 0.5ml/h, the flow rate of the core layer solution was set to 0.2ml/h, and the spinning voltage Set as 20kV, others are the same as embodiment 1. The thickness of the collected web material was about 2 mm. the

(2) Preparation of sodium alginate nanofiber web material

In the preparation of the spinning stock solution, the volume ratio of water and glycerin was set to 0.5:1; during electrospinning, the voltage was set to 28kV, the flow rate was set to 100μl/h, and the coagulation bath was composed of a CaCl ₂ aqueous solution with a mass percentage concentration of 10%. With the mixed solution that mass percentage is 80% dehydrated alcohol in the mixed solution, other is with embodiment 1. The thickness of the collected web material was about 1.2 mm.

(3) Preparation of PLGA hollow nanofiber web material

In the preparation of the spinning stock solution, the concentration of the PLGA solution was set to 1.5g/ml, and the volume ratio of DMF to THF was set to 3:1; during coaxial spinning, the flow rate of the shell solution was set to 1.2ml/h, and the core solution The flow rate was set at 0.5ml/h, the spinning voltage was set at 10kV, and the others were the same as in Example 1. The thickness of the collected web material was about 2 mm. the

(4) Preparation of chitosan hollow nanofiber mesh material

The shell solution is prepared as a chitosan solution with a mass fraction of 7.5%, and the others are the same as in Example 1. The thickness of the collected web material was about 1.5 mm. the

(5) Molding of composite materials

The molding pressure is set at 150 N/cm ² , and the thickness of the soft part is about 7 mm, and the thickness of the hard part is about 4.5 mm. Others are the same as in Embodiment 1.

(6) Disinfection treatment

With embodiment 1. the

Embodiment 4 (preparation of No. 4 sample)

(1) Preparation of gelatin hollow nanofiber web material

In the preparation of the spinning stock solution, the mass fraction of the gelatin solution is set to 16%; during coaxial spinning, the flow rate of the shell layer solution is set to 0.5ml/h, and the flow rate of the core layer solution is set to 0.2ml/h, and other implementations are the same example 1. The thickness of the collected web material was about 1.5 mm. the

(2) Preparation of sodium alginate nanofiber web material

In the preparation of the spinning stock solution, the volume ratio of water and glycerin was set to 0.5:1; during electrospinning, the voltage was set to 28kV, the flow rate was set to 100μl/h, and the coagulation bath was composed of a CaCl ₂ aqueous solution with a mass percentage concentration of 10%. With the mixed solution that mass percentage is 80% dehydrated alcohol in the mixed solution, other is with embodiment 1. The thickness of the collected web material was about 1.5 mm.

(3) Preparation of PLGA hollow nanofiber web material

During coaxial spinning, the flow rate of the core layer solution was set to 0.5ml/h, the spinning voltage was set to 10kV, and the others were the same as in Example 1. The thickness of the collected web material was about 2 mm. the

(4) Preparation of chitosan hollow nanofiber mesh material

The shell solution is prepared as a chitosan solution with a mass fraction of 6.5%, and the others are the same as in Example 1. The thickness of the collected web material was about 1.5 mm. the

(5) Molding of composite materials

The molding pressure is set at 150 N/cm ² , and the thickness of the soft part is about 7 mm, and the thickness of the hard part is about 4.5 mm. Others are the same as in Embodiment 1.

(6) Disinfection treatment

With embodiment 1. the

Embodiment 5 (preparation of No. 5 sample)

(1) Preparation of gelatin hollow nanofiber web material

In the preparation of the spinning stock solution, the mass fraction of the gelatin solution was set to 14%; during coaxial spinning, the flow rate of the shell solution was set to 0.5ml/h, and the flow rate of the core layer solution was set to 0.2ml/h, and the others were implemented in the same way example 1. The thickness of the collected web material was about 1.5 mm. the

(2) Preparation of sodium alginate nanofiber web material

In the preparation of the spinning dope, the weight-to-volume ratio of sodium alginate to glycerin was set at 3 g/ml, and the others were the same as in Example 1. The thickness of the collected web material was about 1.5 mm. the

(3) Preparation of PLGA hollow nanofiber web material

During coaxial spinning, the flow rate of the core layer solution was set at 0.5ml/h, and the others were the same as in Example 1. The thickness of the collected web material was about 2 mm. the

(4) Preparation of chitosan hollow nanofiber mesh material

The shell solution is prepared as a chitosan solution with a mass fraction of 7%, and the others are the same as in Example 1. The thickness of the collected web material was about 1.5 mm. the

(5) Molding of composite materials

The outer diameter of the mold is 8cm×8cm, the inner diameter is 6cm×6cm, each side of the hard part is 1cm, the molding pressure is set to 150N/cm ² , the thickness of the soft part is about 7mm, and the thickness of the hard part is about 4.5mm. Others are the same Example 1.

(6) Disinfection treatment

With embodiment 1. the

2. Mouse hemostasis test

Experimental Kunming white mice: SPF grade, weighing 18-22 g, purchased from the Experimental Animal Center of Dalian Medical University (animal license number: SCXK (Liao) 2002-0002). the

The multi-layer composite hemostatic material is sample No. 1-5 prepared in the above examples; Tailing absorbable hemostatic gauze and ordinary gauze are provided by the Second Affiliated Hospital of Dalian Medical University. the

24 mice, half male and half male, were randomly divided into 4 groups, 6 mice in each group, respectively: multi-layer composite hemostatic material group (sample group No. 1-5), Tailing absorbable hemostatic gauze group (positive control group) ), ordinary gauze group and blank control group. After being anesthetized by intraperitoneal injection of 2% pentobarbital sodium (0.002mL/g), 1cm of the mouse tail was cut off with sharp surgical scissors, and the timing was started. Each experimental group covered the wound with corresponding materials or gauze (1cm×1cm Double-layer), and use the index finger to press slightly, so that the material or gauze is close to the wound, until the bleeding stops, and the hemostasis time is recorded. The blank control group was not covered with any hemostatic dressing or medicine, and the hemostatic time was recorded in the same way. the

Statistical software SPSS10.0 was used for data processing, and the results were expressed as x±s. The differences between groups were compared using t test, and P<0.05 was considered statistically significant. The experimental results are shown in Table 1. the

Experimental results: five new multi-layer composite hemostatic materials can significantly shorten the bleeding time, and the hemostatic effect of No. 1 and No. 2 materials is obviously better than that of the positive control group, and both are significantly better than the ordinary gauze group, which proves that the multi-layer composite hemostatic material of the present invention Has excellent hemostatic effect. the

Table 1 The effect of each group of materials on the hemostatic effect of docked mice (n=6)

Note: Compared with the blank control group, ^* p<0.05, ^** p<0.01; compared with the ordinary gauze group, #p<0.05, ##p<0.01; compared with the positive control group, △p<0.05.

Claims (5)

1. A multi-layer composite hemostatic material is characterized in that it is formed through molding compounding by PLGA hollow nanofiber layer, gelatin hollow nanofiber layer, chitosan hollow nanofiber layer and alginate nanofiber layer around. 2. The multi-layer composite hemostatic material according to claim 1, characterized in that hemostatic drugs, anti-infection drugs, analgesic drugs or tissue repair promoting factors are filled between the layers. 3. The multilayer composite hemostatic material according to claim 1, wherein the thickness of the multilayer composite hemostatic material is 0.3-3 cm. 4. A method for preparing the multilayer composite hemostatic material according to any one of claims 1-3, comprising the steps of: (1) Preparation of PLGA hollow nanofiber web by coaxial spinning method Use chloroform, DMF, THF or a mixture of DMF and THF as a solvent to prepare a PLGA spinning stock solution with a concentration of 0.5-1.5g/ml. This solution is the shell solution, and the core solution uses sesame oil, mineral oil, dimethyl Silicone oil or blended edible oil, the flow rate of the shell layer solution is 0.5-2.0ml/h, the flow rate of the core layer solution is 0.2-1.0ml/h, the solution flow rate is precisely controlled by the syringe pump, the spinning voltage is adjusted between 10-20kV, and the collected The distance is 5-15cm, the collection time is 8-72 hours, the thickness of the collected fiber web is continuously adjustable from several hundred microns to 3 mm, first put the collected fiber web into a high-speed centrifuge for deoiling, and then The oil component is extracted with cyclohexane or carbon tetrachloride, and finally vacuum-dried to obtain pure PLGA hollow nanofiber network; (2) Preparation of gelatin hollow nanofiber network by coaxial spinning method Formic acid or water is used as a solvent to prepare a gelatin spinning stock solution with a concentration of 15-30wt%. This solution is a shell solution, and the core solution uses sesame oil, mineral oil, simethicone or blended edible oil. The flow rate of the shell solution is 0.5-1.0ml/h, the flow rate of the core layer solution is 0.2-0.5ml/h, the solution flow rate is precisely controlled by the syringe pump, the spinning voltage is adjusted between 10-30kV, the collection distance is 5-15cm, and the collection time is 4- For 72 hours, the ambient temperature during spinning is controlled above 40°C, and the thickness of the collected fiber web is continuously adjustable from several hundred microns to 3 mm. Then extract the oil component with cyclohexane, and finally vacuum-dry to obtain pure gelatin hollow nanofiber network; (3) Preparation of chitosan hollow nanofiber network by coaxial spinning method Concentrated acetic acid or trifluoroethanol is used as a solvent to prepare a chitosan spinning stock solution with a concentration of 5-30wt%. This solution is a shell solution, and the core solution uses sesame oil, mineral oil, simethicone or blended edible oil. The flow rate of the shell solution is 10-40μl/h, the flow rate of the core layer solution is 5-20μl/h, the solution flow rate is precisely controlled by the syringe pump, the spinning voltage is adjusted between 10-30kV, the collection distance is 5-15cm, the collection time For 4-72 hours, the thickness of the collected fiber web is continuously adjustable from several hundred microns to 3 mm. First, put the collected fiber web into a high-speed centrifuge for centrifugal deoiling, and then extract the oil with cyclohexane Components, finally vacuum-dried to obtain pure chitosan hollow nanofibrous network; (4) Preparation of alginate nanofiber webs by electrospinning Mix alginate and glycerin at a weight-to-volume ratio of 2-4g/ml, then add solvent water to the mixture, wherein the volume ratio of water to glycerin is 0.5-1:1, stir evenly, and remove air bubbles at rest to obtain Spinning stock solution, the flow rate of spinning stock solution is 50-200μl/h, the solution flow rate is precisely controlled by a syringe pump, the spinning voltage is adjusted between 10-40kV, the distance between the spinneret and the receiver is 5-15cm, and the receiver is soaked in In the coagulation bath composed of CaCl ₂ aqueous solution and absolute ethanol, the collection time is 36-72 hours, the thickness of the collected fiber web is continuously adjustable between several hundred microns and 2 mm, and the obtained fiber web is vacuum-dried, Obtain alginate nanofibrous web; (5) Composite layers of materials by molding Take the prepared fiber nets, superimpose them together, and fill hemostatic drugs, anti-infective drugs, pain-relieving drugs or factors promoting tissue repair between the layers according to the needs, and use a hollow mold to place them Press it into a shape with hard sides and soft middle of the specified size, and then cut it with a knife. The molding pressure is 100-400N/cm ² , and the number of layers of each fiber web is 1 or more layers; (6) Disinfection treatment After the obtained composite material is sterilized by gamma-rays, a multi-layer composite hemostatic material is obtained. 5. The preparation method according to claim 4, characterized in that, in at least any one of the coaxial spinning shell solutions in steps (1)-(3), single-walled or multi-walled nano carbon tube.