CN107158452B

CN107158452B - Bone wound hemostatic composition and preparation method and application thereof

Info

Publication number: CN107158452B
Application number: CN201710309890.8A
Authority: CN
Inventors: 任芳; 李波; 王月月; 张林林; 刘长凤; 张春霞; 赵成如
Original assignee: Saikesaisi Biotechnology Co ltd
Current assignee: Saikesaisi Biotechnology Co ltd
Priority date: 2017-05-05
Filing date: 2017-05-05
Publication date: 2020-05-01
Anticipated expiration: 2037-05-05
Also published as: CN107158452A

Abstract

The invention discloses a bone wound hemostatic composition and a preparation method and application thereof. The hemostatic composition has good bone adhesion and plasticity, can mechanically fill fracture and blood sinus of a bone wound surface, can accelerate the natural coagulation process, and has good hemostatic effect; the hemostatic powder is easy to degrade in a human body or an animal body, can be absorbed and metabolized, has good biocompatibility, no stimulation to the organism, no toxic or side effect and high safety, does not influence wound healing, is convenient to use clinically, and has good application prospect in hemostasis of bone wounds of human beings or animals.

Description

Bone wound hemostatic composition and preparation method and application thereof

Technical Field

The invention relates to a bone wound hemostasis composition, a preparation method and application thereof, in particular to a bone wound hemostasis composition which is strong in plasticity, good in adhesion, absorbable and free of influence on wound healing, a preparation method and application thereof, and belongs to the technical field of bone hemostasis.

Background

Clinically, cancellous bone structure is loose, blood circulation is abundant, bleeding of wound surfaces is mostly blood seepage, cancellous bone wound surfaces bleed continuously or are difficult to stop bleeding thoroughly because bone wound surface tissue structure is loose, blood circulation is abundant to form densely distributed blood sinuses, vasoconstriction in tissues is poor, blood platelets are gathered, and blood clots are difficult to adhere to the bone wound surfaces. The bleeding of the bone wound surface is difficult to be stopped by vasoconstriction, and the bleeding is difficult to be stopped by conventional methods such as electric coagulation, clamping, hemostatic gauze, gelatin sponge filling and the like in the operation. Bone wax is commonly used for hemostasis of cancellous bone wound surfaces in clinic. The main components of the bone wax are beeswax, sesame oil and the like, the bone wax is a mixture of disinfected beeswax and vaseline, has good softening performance, can be molded after being kneaded and softened by hands, and is non-toxic. The bone wax has the action principle of blocking capillary blood seepage of marrow parts by a physical method, and can be used for stopping bleeding of various emergency patients during bone blood seepage. At present, the bone wax commonly used in clinic performs hemostasis by a mechanical stuffing principle of sealing bleeding passages among bones and controlling bone injury bleeding, has quick and effective hemostasis effect, but has poor biocompatibility and is difficult to degrade and absorb by organisms, and can be used as a foreign body to be remained in the body for a long time after being applied, so that on one hand, the anti-infection capacity of tissues is reduced, and the risk of postoperative infection is increased; on the other hand, the bone wound surface is also prevented from being repaired, and symptoms such as local pain, effusion and the like caused by foreign body granuloma are formed. Researchers at home and abroad have carried out research work on bone wax substitute products such as collagen fibers, gelatin sponges, oxidized cellulose, polylactic acid, polysaccharide and the like from the aspects of artificial synthesis of high polymer materials, biological agents and the like. Animal experiments and clinical application show that: although the substances have good biocompatibility and degradability, most of the substances are powdery and colloidal, have poor adhesion with wound surfaces, are difficult to operate in operation and are easy to wash away by blood flow. In addition, the polymer substitute also has the unsolved practical problems of expensive raw materials, difficult modification of physical properties, material safety and the like, so that the polymer substitute cannot be converted into an ideal material suitable for bone wound hemostasis at present, and further intensive research and improvement are still needed. There are also some patent disclosures on such products, as follows:

patent application No. 201210067344.5 discloses a material for completely degrading in human or animal body, replacing bone wax, stopping bleeding of bone wound surface without changing doctor's habit and promoting bone tissue healing, which comprises oligosaccharide, polysaccharide or mixture of oligosaccharide and polysaccharide, excipient is one or more of polyhydroxy alcohol, one or more of vegetable oil, and one or more of emulsifier. The material has high saccharide matrix content, and the paste formed by hydrophilic saccharide, polyhydric alcohol and hydrophobic vegetable oil has poor stability, and if the material is developed into a product on the market, the material has the defects of short effective period, poor stability and low water absorption. And the product is prepared by blending under vacuum condition and immediately cooling at 0-4 ℃. The hemostatic material disclosed in the patent is a bone hemostatic with relatively excellent performance in the existing degradable materials, but the preparation process of the hemostatic is relatively complex, the preparation process not only involves high-temperature preparation, but also vacuum reaction, and also involves low-temperature cooling, the paste composed of hydrophilic and hydrophobic media has poor stability, and the effective period is relatively short if the hemostatic material is developed into a product on the market. Meanwhile, the problems existing in the bone wax market at present can not be well solved in consideration of the problems of high difficulty in mass production, high production cost and difficulty in realizing mass production.

The patent of application No. 200410040076.3 discloses an absorbable bone hemostatic agent for surgery or orthopedic surgery, which comprises chitosan or modified chitosan and other substances, wherein chitosan is a common hemostatic material, and the hemostatic principle mainly utilizes the positive charge of chitosan itself to accelerate blood coagulation and enrich blood platelets to achieve hemostatic effect, and the material is degradable and absorbable as a biological material. However, the chitosan material belongs to animal-derived materials and has certain potential harm to human bodies. The cement formed by taking chitosan as a main component is used as a bone hemostatic, and other performances such as water absorption, adhesion, biocompatibility and the like are not reported. Generally, chitosan materials have high intradermal stimulation, and have high biological stimulation if the chitosan materials are not well treated. The material is easy to deteriorate under heat and radiation, and special design is required for sterilization, storage and transportation, so that the material cost is greatly increased.

The patent of application No. 200910076033.3 discloses a degradable hemostatic material in bone cavity and a preparation method thereof, wherein the material comprises sodium alginate, medical starch and inorganic particles, and the adhesion of sodium alginate hydrogel is poor. The material has no reports on other performances such as water absorption, adhesion, biocompatibility, plasticity, hemostatic effect, degradation characteristic and the like.

The patent of application No. 201410780581.5 discloses a chitosan hemostatic gauze and a preparation method thereof, the hemostatic gauze contains chitosan, sodium alginate, sodium carboxymethyl cellulose, hydroxyethyl starch, squalane, traditional Chinese medicine components and water, and is spun into fibers by an electrostatic spinning method. In the invention, firstly, the chitosan is an animal-derived material and has certain potential harm to human bodies. The gauze formed by taking chitosan as a main component is used as a hemostatic material for the bone wound surface, and other performances such as water absorption, adhesion, biocompatibility and the like are not reported. Generally, chitosan materials have high intradermal stimulation, and have high biological stimulation if not well treated, and in addition, the hemostatic materials in the form of gauze are difficult to firmly adhere to the wound surface of bleeding bone. The material is easy to deteriorate under heat and radiation, and special design is required for sterilization, storage and transportation, so that the material cost is greatly increased.

The patent of application No. 200510117740.4 discloses a biodegradable and absorbable polymer nano-fiber membrane material, its preparation method and application, the product is a nano-fiber membrane containing one or more of gelatin, hyaluronic acid, chitosan, collagen, heparin, polyglycolide, polyvinylpyrrolidone, polyvinyl alcohol, cellulose, starch and other high molecular materials, and inorganic reinforcing agent components such as calcium carbonate, potassium chromate, hydrogen phosphate and the like. When the invention is used as a hemostatic material, the bone wound surface blood sinuses; secondly, the plasticity is poor, and the bone wound surfaces in different shapes cannot be packed and smeared differently; in addition, many components in the invention such as gelatin, chitosan and collagen are animal-derived materials, and have potential biological hazard; finally, many high molecular components in the invention have longer degradation time when being used for hemostasis of bone wound surfaces, and can block the healing of bone tissues.

In addition, many reports disclose that polymers such as polyoxyethylene, polyoxypropylene, polylactic acid and the like are utilized to prepare bone hemostatic agents, and the materials are high molecular polymers with certain molecular weight, can be degraded and absorbed, but have slow degradation speed and long degradation period and can hinder bone healing; in addition, the water absorption is poor, the blood can not be well absorbed, the adhesive force and the plasticity are poor, and the clinical operation is inconvenient.

The ideal bone hemostatic material has the advantages of soft texture and strong plasticity of ①, capability of rapidly filling cracks and blood sinuses of a wound surface, capability of firmly adhering to the wound surface without falling off within a certain time due to bone adhesion property of ②, good biocompatibility of ③, no toxic or side effect, small tissue reaction, capability of gradually degrading after the completion of the hemostatic action of ④, absorption by organism tissues and no influence on wound healing, and from the understanding of the prior art, no good ideal bone hemostatic material exists at present, and further research and improvement are needed.

Disclosure of Invention

Aiming at the defects of the existing bone hemostatic material, the invention provides the bone wound hemostatic composition which has good bone adhesion and plasticity, can mechanically fill fracture and blood sinus of the bone wound, can accelerate the natural blood coagulation process and has good hemostatic effect. The composition is easily degraded in human body or animal body, can be absorbed and metabolized, has good biocompatibility, no irritation to organism, no toxic or side effect, high safety, and no influence on wound healing.

The invention also provides a preparation method of the bone wound hemostasis composition, the preparation method is simple, the implementation is easy, the cost is low, the obtained product has ideal hardness, toughness and viscosity, the plasticity can be well coated, the hemostasis effect is good, the toxic and side effects are avoided, the tissue reaction is small, the wound healing is not influenced, and the clinical use is convenient.

The invention also provides the application of the bone wound hemostasis composition in preparing a bone wound hemostasis product for human or animals, and the product can be used for hemostasis of all bone wound bleeding of human or animals in operation.

Further, the bone wound hemostasis composition provided by the invention comprises a solvent matrix, a thickening agent, an excipient and an ionic salt, wherein the solvent matrix is polyhydroxy alcohol, and the ionic salt is one or more of inorganic calcium salt and inorganic iron salt.

In the bone wound hemostasis composition, the solvent matrix is polyhydroxy alcohol, and the solvent matrix is used for providing a basic environment for the thickening agent and the excipient, so that the powder or the liquid can form an ointment or a gel with stronger adhesion and better plasticity. In addition, changes in the solvent base content can cause changes in properties such as plasticity, adhesion, water absorption characteristics, and degradation time. The polyhydric alcohol is one or two of glycerol, polyethylene glycol-400, polyethylene glycol-300 and propylene glycol, preferably one or two of glycerol and polyethylene glycol-400.

In the bone wound hemostasis composition, the weight percentage of the solvent matrix in the bone wound hemostasis composition is 30-80%, and preferably 50-70%.

In the bone wound hemostasis composition, the thickening agent is used for improving the adhesion and plasticity of the composition, and is one or two of sodium carboxymethylcellulose, hydroxypropyl methylcellulose, hydroxyethyl cellulose, carrageenan and carbomer. When the thickener is chosen to have a particular viscosity, the resulting product properties are better. The sodium carboxymethylcellulose is divided into the following components according to viscosity: the high-viscosity sodium carboxymethyl cellulose, the medium-viscosity sodium carboxymethyl cellulose and the low-viscosity sodium carboxymethyl cellulose are preferably medical high-viscosity sodium carboxymethyl cellulose, medium-viscosity sodium carboxymethyl cellulose or low-viscosity sodium carboxymethyl cellulose. The hydroxypropyl methylcellulose is preferably hydroxypropyl methylcellulose with nominal viscosity of 6-15000 cps, more preferably 50-1500 cps. The hydroxyethyl cellulose preferably adopts hydroxyethyl cellulose with nominal viscosity of 250-400cps, 1500-2500cps, 3500-5500cps or 4500-6500cps, and more preferably adopts one of the hydroxyethyl cellulose with nominal viscosity of 1500-2500cps and 3500-5500 cps; the carrageenan is preferably of medical grade. The carbomer is preferably carbomer 910, carbomer 934, carbomer 940 or carbomer 941, and more preferably one of carbomer 934 and carbomer 940. The nominal viscosity is the viscosity obtained by adopting a rotational viscosity meter.

In the bone wound hemostatic composition, the weight percentage of the thickening agent in the bone wound hemostatic composition is 0.5-10%, preferably 5-7%.

In the bone wound hemostasis composition, the excipient has the function of improving the plasticity, adhesion and water absorption of the product, and is one or more of sodium carboxymethyl starch, potato starch, soluble starch, sodium alginate, dextrin and dextrin derivatives.

In the bone wound hemostatic composition, the weight percentage of the excipient in the bone wound hemostatic composition is 10-55%, preferably 20-45%.

In the bone wound hemostasis composition, the ionic salt is used as a coagulant to accelerate and assist hemostasis. The ionic salt is inorganic calcium salt, inorganic ferric salt, or a mixture of inorganic calcium salt and inorganic ferric salt. The inorganic calcium salt is calcium sulfate, calcium chloride, calcium nitrate, the inorganic ferric salt is ferric chloride, ferric nitrate, ferric sulfate, the ionic salt can be selected from these inorganic calcium salt and inorganic ferric salt, for example, one or more of inorganic calcium salt can be selected, one or more of inorganic ferric salt can be selected, one of inorganic calcium salt and inorganic ferric salt can be selected, multiple of inorganic ferric salt can be selected, and multiple of inorganic calcium salt can be selected.

In the bone wound hemostatic composition, the weight percentage of the ionic salt in the bone wound hemostatic composition is 0.05-5%, preferably 1-3%.

The preparation method of the bone wound hemostasis composition comprises the following steps:

a. mixing the solvent matrix and the thickening agent, stirring at 20-75 ℃ until a viscous solvent matrix is formed, and swelling overnight;

b. slowly adding the excipient into a viscous solvent matrix which is completely swelled under the conditions of stirring, blending or kneading, and then continuously stirring, blending or kneading until uniform paste or gum is formed;

c. slowly adding the ionic salt into the paste or the colloid under the conditions of stirring, blending or kneading, and continuously stirring, blending or kneading after the ionic salt is added until uniform paste or the colloid is formed, namely the bone wound hemostatic composition.

The bone wound hemostasis composition is a uniform viscous paste or gel, has excellent smearing operability, plasticity and adhesiveness and excellent blood absorption characteristic, can be absorbed and degraded, does not influence wound healing, has no toxic or side stimulation to organisms, and can be prepared into preparations such as paste or smearing preparation and the like. In actual use, the uniform paste or the gel can be made into different shapes in order to be suitable for different wound shapes and parts, for example, the uniform paste can be directly put into a closed paste box, vacuum-sealed in an aluminum foil bag, sterilized and stored at normal temperature. Or making into sheet, column or other shapes with different molds, packaging with laminating paper, vacuum sealing in aluminum foil bag, sterilizing, and storing at room temperature. Or packaging in various medical operating tools, such as medical applicator, medical hose, medical injector, etc., vacuum sealing in aluminum foil bag, sterilizing, and storing at room temperature. However, the paste or gel has good plasticity no matter what shape the paste or gel is made into and stored in any container.

In the step a, the temperature is raised to 20-75 ℃, and stirring is continued until a viscous solvent matrix is formed. It takes about 30 min.

In the above steps b and c, the mixing is carried out by stirring, blending or kneading, and these operations may be carried out in a vessel with a stirring device, an emulsion mixer, a blender or a kneader.

In the step b, the time for forming the uniform paste is about 1-2 hours; in the step c, the time for forming the uniform paste is about 0.5-1 h.

The bone wound hemostasis composition has the advantages of good and random shaping characteristics, can be shaped by a clinician according to the requirements according to the characteristics of the bone wound, can be smeared and filled, and can be used for hemostasis of various bone wound bleeding with good clinical application effect. Based on the bone wound hemostatic composition, the invention also provides application of the bone wound hemostatic composition in preparation of a human or animal bone wound hemostatic product. The bone wound surface can be a wound surface formed by bone wounds caused by various reasons.

The invention has the following beneficial effects:

1. the composition has good arbitrary shaping characteristics, can be shaped by a clinician according to the requirements according to the characteristics of a bone wound surface, can be smeared and filled, has good bone adhesion, can be firmly adhered to the bone wound surface by depending on the specific bonding strength, can fill bleeding parts (blood sinuses and cracks) of the bone wound surface, and has good hemostatic effect. In addition, the composition has high water absorption property, and liquid absorbability of not less than 30 times, and can absorb water in blood exuded from bone wound surface, concentrate blood, collect blood platelet, erythrocyte and blood coagulation protein in blood, and accelerate natural blood coagulation process, thereby improving hemostatic effect of bone wound surface.

2. The pH value of the composition is neutral, all indexes are strictly controlled, the adopted solvent matrix, the thickening agent and the excipient are easy to degrade in a human body or an animal body, can be absorbed and metabolized by the organism after degradation, have no residue in the human body or the animal body, have no stimulation to the organism, have no toxic or side effect, high safety and good biocompatibility, avoid the occurrence of complications such as foreign body reaction, tissue inflammation and the like of a user, and do not influence the healing of a wound surface.

3. The composition has the advantages of simple preparation method, convenient use, ideal hardness, toughness and viscosity, plasticity, spreadability, degradability, absorbability, good hemostatic effect, convenient clinical use and good application prospect in hemostasis of bone wound surfaces of people or animals.

4. The composition of the invention can be developed into bone hemostat for promoting bone healing after being added with growth factors, calcium phosphate, hydroxyapatite, calcium carbonate, calcium sulfate, bioactive glass and other substances for promoting bone growth; the compositions of the invention, when supplemented with antibiotics or other therapeutically functional agents, can be developed into bone hemostats with anti-infective or other therapeutic effects.

Detailed Description

The present invention will be described in further detail with reference to specific examples.

Example 1

The formula of the bone wound hemostasis composition is shown in the following table 1:

TABLE 1 formulation Table of example 1

1. weighing glycerol and medical-grade high-viscosity sodium carboxymethylcellulose according to a formula table, and stirring to obtain a uniform solution. The solution was slowly warmed and stirred at 30 ℃ for 30min to allow the homogeneous solution to become a viscous solvent matrix which was allowed to swell overnight.

2. Weighing sodium carboxymethyl starch according to a formula table, slowly adding the sodium carboxymethyl starch into a viscous solvent matrix which is completely swelled, and strongly stirring the mixture while adding the sodium carboxymethyl starch until a uniform paste is formed for about 2 hours;

3. and weighing calcium sulfate according to a formula table, slowly adding the calcium sulfate into the paste, and continuously stirring until uniform paste is formed for about 1 hour.

4. Filling the paste obtained in the step (3) into a sealing paste box, sealing the sealing paste box in an aluminum foil bag in vacuum, and preserving the sealing paste box at normal temperature after sterilization; or tabletting to obtain tablet and column, packaging with laminating paper, vacuum sealing in aluminum foil bag, sterilizing, and storing at room temperature; or packaging in medical injector, vacuum sealing in aluminum foil bag, sterilizing, and storing at room temperature; or placing into gel feeder, vacuum sealing in aluminum foil bag, sterilizing, and storing at room temperature.

Example 2

The formula of the bone wound hemostasis composition is shown in the following table 2:

TABLE 2 formulation table of example 2

1. weighing glycerol, polyethylene glycol-400, hydroxypropyl methyl cellulose with the nominal viscosity of 50-1500 cps and hydroxyethyl cellulose with the nominal viscosity of 1500-2500cps according to a formula table, and stirring to obtain a uniform solution. The solution was slowly warmed and stirred at 75 ℃ for 30min to allow the homogeneous solution to become a viscous solvent matrix which was allowed to swell overnight.

2. Putting the obtained viscous solvent matrix into a kneader, starting the kneader to stir, weighing sodium alginate and dextrin according to a formula table, uniformly mixing the sodium alginate and the dextrin, slowly adding the mixture into the kneader, kneading the mixture while adding the mixture until a uniform paste is formed, and keeping the uniform paste for about 1 hour;

3. and weighing calcium chloride and ferric sulfate according to a formula table, sequentially and slowly adding the calcium chloride and the ferric sulfate into the paste, and continuously kneading until uniform paste is formed for about 0.5 h.

Example 3

The formula of the bone wound hemostasis composition is shown in the following table 3:

TABLE 3 formulation Table of example 3

1. weighing propylene glycol, polyethylene glycol-300, medical grade carrageenan, hydroxyethyl cellulose with the nominal viscosity of 250-400cps and hydroxypropyl methyl cellulose with the nominal viscosity of 4000-9000 cps according to a formula table, and stirring to obtain a uniform solution. The solution was slowly warmed and stirred at 50 ℃ for 30min to allow the homogeneous solution to become a viscous solvent matrix which was allowed to swell overnight.

2. Putting the obtained viscous solvent matrix into a kneader, starting the kneader to stir, weighing the potato starch and the sodium carboxymethyl starch according to a formula table, uniformly mixing the potato starch and the sodium carboxymethyl starch, slowly adding the mixture into the kneader while kneading until a uniform paste is formed for about 1 hour;

3. weighing ferric nitrate according to the formula table, slowly adding into the paste, and continuously kneading until uniform paste is formed for about 0.5 h.

Example 4

The formula of the bone wound hemostasis composition is shown in the following table 4:

TABLE 4 formulation table of example 4

1. weighing polyethylene glycol-400 and carbomer 934 according to a formula table, putting into an emulsion stirrer, fully stirring at room temperature until a viscous solvent matrix is formed, and swelling overnight.

2. Putting the obtained viscous solvent matrix into a kneader, starting the kneader to stir, weighing the soluble starch, the sodium carboxymethyl starch and the dextrin according to a formula table, uniformly mixing the soluble starch, the sodium carboxymethyl starch and the dextrin, slowly adding the mixture into the kneader, kneading the mixture while adding the mixture until a uniform paste is formed, and keeping the time for about 1.5 hours;

3. and weighing calcium nitrate according to a formula table, slowly adding the calcium nitrate into the paste, and continuously kneading until uniform paste is formed for about 0.5 h.

Example 5

The formula of the bone wound hemostasis composition is shown in the following table 5:

TABLE 5 formulation table of example 5

1. according to the formula, propylene glycol and hydroxyethyl cellulose with nominal viscosity of 4500-6500cps are weighed, and uniform solution is obtained under stirring condition. The solution was slowly warmed and stirred at 40 ℃ for 30min to allow the homogeneous solution to become a viscous solvent matrix which was allowed to swell overnight.

2. Putting the obtained viscous solvent matrix into a kneader, starting the kneader to stir, weighing dextrin according to a formula table, slowly adding the dextrin into the kneader while kneading until uniform paste is formed, and keeping for about 1.5 hours;

3. weighing ferric chloride according to the formula table, slowly adding into the paste, and kneading continuously until uniform paste is formed for about 0.5 h.

Example 6

The formula of the bone wound hemostasis composition is shown in the following table 6:

TABLE 6 formulation table of example 6

1. according to a formula table, weighing glycerol, carbomer 934, medical-grade low-viscosity sodium carboxymethylcellulose and hydroxyethyl cellulose with the nominal viscosity of 3500-5500cps, and stirring to obtain a uniform solution. Stirring at room temperature for 30min to make the homogeneous solution become a viscous solvent matrix, and swelling overnight.

2. Putting the obtained viscous solvent matrix into a kneader, starting the kneader to stir, weighing dextrin and sodium carboxymethyl starch according to a formula table, uniformly mixing the dextrin and the sodium carboxymethyl starch, slowly adding the mixture into the kneader, kneading the mixture while adding the mixture until a uniform paste is formed, and keeping the time for 1 hour;

3. weighing ferric chloride according to a formula table, slowly adding the ferric chloride into the paste, and continuously kneading until uniform paste is formed for about 1 hour.

Example 7

The formula of the bone wound hemostasis composition is shown in the following table 7:

TABLE 7 formulation Table of example 7

1. weighing glycerol, hydroxypropyl methylcellulose with nominal viscosity of 10000-15000 and carbomer 910 according to a formula table, and stirring to obtain a uniform solution. The solution was slowly warmed and stirred at 30 ℃ for 30min to allow the homogeneous solution to become a viscous solvent matrix which was allowed to swell overnight.

2. Weighing sodium carboxymethyl starch, potato starch and dextrin according to a formula table, slowly adding the sodium carboxymethyl starch, the potato starch and the dextrin into a viscous solvent matrix which is completely swelled in sequence under strong stirring, and stirring while adding until a uniform paste is formed for about 2 hours;

3. weighing ferric chloride and calcium chloride according to a formula table, slowly adding into the paste, and continuously stirring until uniform paste is formed for about 1 h.

Comparative example 1

The formula comprises (% w/w): solvent matrix: 35% of glycerol, 30% of mannitol, and a thickening agent: PVP 5.5%, xanthan gum 1.5%, excipient: 26% soluble starch, ionic salt: 2 percent of calcium dihydrogen phosphate.

Glycerol, mannitol, PVP, and xanthan gum were mixed according to the method of example 2, and the resulting homogeneous solution was stirred. The solution was slowly warmed and stirred at 75 ℃ for 30min to allow the homogeneous solution to become a viscous solvent matrix which was allowed to swell overnight. And (3) putting the obtained viscous solvent matrix into a kneader, starting the kneader to stir, slowly adding soluble starch while kneading until the soluble starch is uniformly mixed, slowly adding calcium dihydrogen phosphate, and continuously kneading until the soluble starch is uniformly mixed. Putting the obtained mixture into a sealing paste box, vacuum sealing in an aluminum foil bag, sterilizing, and storing at normal temperature; or processing into sheet, column or other shapes, packaging with laminating paper, vacuum sealing in aluminum foil bag, sterilizing, and storing at room temperature.

Comparative example 2

The formula comprises (% w/w): maltose 10%, carboxymethyl cellulose 20%, hydroxypropyl starch 15%, chitosan 10%, gelatin 10%, glycerol 30% and tween 5%.

Mixing glycerol and tween, heating to 40 deg.C, stirring under sealed condition for 1 hr, adding maltose, carboxymethyl cellulose, hydroxypropyl starch, chitosan and gelatin, and stirring under vacuum at 60 deg.C for 4-5 hr. Putting the obtained mixture into a sealing paste box, vacuum sealing in an aluminum foil bag, sterilizing, and storing at normal temperature; or packaging with laminating paper, vacuum sealing in aluminum-plastic bag, sterilizing, and storing at room temperature.

The excellent performances of the product such as effectiveness, stability, safety, biocompatibility, hemostaticity and the like can be proved by the following tests.

Verification example 1: test for effectiveness and stability

According to YY/T0681.1-2009, sterile medical device packaging test method part 1: according to the method specified in the accelerated aging test guideline, the sample after sterilization is subjected to an accelerated aging test, and whether the performance of the sample after accelerated aging is stable or not and whether the performance is good or not are observed so as to evaluate the stability of the sample. The evaluation indexes of the accelerated aging test mainly comprise:

1. pH value: placing the sterilized sample in a constant temperature and humidity box at 60 ℃, taking out after 0 month and 3 months respectively, mixing with purified water according to a certain proportion, stirring uniformly, and leaching for 24 hours to obtain the pH value;

2. coating operability: placing the sterilized sample in a constant temperature and humidity box at 60 ℃, taking out after 0 month and 3 months respectively, taking the sample in the hand, smearing the sample on a sample plate soaked by water in advance, and observing whether the sample is easy to be smeared uniformly and can be firmly adhered to the sample plate;

3. and (3) plasticity: placing the sterilized sample in a constant temperature and humidity box at 60 ℃, taking out after 0 month and 3 months respectively, kneading and shaping randomly by hands, evaluating the difficulty degree in the shaping process, and observing whether the state after shaping is good or not;

4. water absorption: and placing the sterilized sample in a constant temperature and humidity box at 60 ℃, taking out after 0 month and 3 months respectively, soaking in sufficient water, separating out water after the sample is completely and uniformly dispersed, and calculating the weight of the water absorbed by the sample to be multiple of the weight of the sample.

5. Strength: the sterilized sample is placed in a constant temperature and humidity box at 60 ℃, taken out after 0 month and 3 months respectively, and the strength of the sample is tested by using a Bloom system of a texture analyzer according to the provisions of the industry standard BS757:1975, wherein the index influences the smearing operability and the plasticity of the product and is also a main parameter for the stable performance of the sample.

The above examples and comparative examples were subjected to accelerated aging tests, respectively, and the results are shown in the following tables 8 and 9:

TABLE 8 Performance Table of samples before accelerated ageing test, i.e. 0 month

From the results, the sample prepared by the formula and the process has excellent performance, can be arbitrarily shaped according to needs, is easy to coat when used for hemostasis of the bone wound surface, is firmly adhered, has excellent liquid absorbability and good hemostasis effect, and is very suitable for clinical operation of clinicians.

TABLE 9 sample Performance Table after 3 months of accelerated aging test

From the above results, it can be seen that after the aging test, the sample has stable performance and still meets the requirements of the clinician for hemostasis of the bone wound. According to the standard, the sample is accelerated and aged for 3 months at 60 ℃, which is equivalent to the time of storing at room temperature for about two years, and the experimental result shows that the sample obtained by the process formula disclosed by the invention is not only easy to realize batch production and meets the clinical operation requirement, but also meets the requirement of excellent performance of the general products on the market within 24-month validity period, and has relatively great practical significance.

Verification example 2: safety and biocompatibility testing

To ensure safety for clinical use, samples were evaluated for biocompatibility. The evaluation method is as follows:

1. cytotoxicity assays

1.1 preparation of test solution:

the sample is extracted in a cell culture medium containing serum under the extraction conditions of 37 +/-1 ℃ and 24 +/-2 hours.

1.2 preparation of cell suspension

Digesting the cells cultured for 48-72 h and growing vigorously by using a digestive juice, adding a cell culture solution, blowing and uniformly mixing by using a suction tube, counting under a microscope by using a blood counting chamber, and calculating the cell density according to the following formula:

C＝10⁴×n/4

in the formula:

c- -cell density in units of individual per milliliter (per mL);

n- - -counting the total number of cells in four big grids at four corners of the plate, wherein the unit is one.

According to the actually measured cell density, a proper amount of cell culture solution is added to prepare cell suspension with the density required by the test for later use.

1.3 the prepared 1X 10⁴The cell suspension is inoculated in a 96-well culture plate, blank control, negative control, positive control and test article groups are arranged, each group is provided with at least 6 wells, and each well is inoculated with 100 mu L of cell suspension. Culturing at 37 deg.C for 24 hr in carbon dioxide incubator (containing 5 vol% carbon dioxide gas), and discarding the stock culture solution. Adding fresh cell culture solution into the blank control group, adding negative control leaching liquor into the negative control group, adding positive control solution or positive control leaching liquor into the positive control group, adding test sample leaching liquor into the test sample group, placing the test sample group in a carbon dioxide gas incubator for continuous culture for 72h, wherein each well is 100 mu L.

After the culture medium was replaced, the cell morphology was observed under a microscope. Adding 20 μ L of tetrazolium salt solution with mass concentration of 5g/L into each well, culturing for 4h, discarding the liquid in the well, adding 150 μ L of DMSO, placing on an oscillator, shaking for 10min, measuring absorbance at 570nm and 630nm wavelength of an enzyme-labeling instrument, and calculating relative increment rate (RGR) according to the following formula.

RGR＝A/A₀×100％

In the formula:

RGR- -relative increment rate,%;

a- - - -absorbance of the test sample group (negative, positive group);

A₀-absorbance of blank control.

Judged according to RGR according to the following table classification criteria. The negative control group should have no more than grade 1 response, and the positive control group should have at least grade 3 response. If the reaction of the negative control group and the positive control group is not established, the test is carried out again.

Grading table of cytotoxicity response

Rank of	Relative increase rate/%)
		0	≥100
1	80～99
		2	50～79
3	30～49
		4	0～29

And (4) judging a result: in the case where the negative control and the positive control produce the expected reaction, the degree of the test sample cytotoxic reaction is determined by analysis.

2. Intradermal reaction

2.1 preparation of test solution:

2.2 test procedure

Thoroughly removing hair on two sides of the spine of the animal 4-18 hours before the test to prepare for injecting the leaching liquor.

0.2mL of the leachate prepared with polar solvent was injected intradermally into 5 points on one side of the spine of each rabbit. Also, 0.2mL of a polar solvent was injected intradermally toward the control solution in the last five spots on the same side of the spine of each rabbit. The other side of the spine of each rabbit was injected with the leach solution prepared with the non-polar solvent and the control with the non-polar solvent, and the procedure was as above.

The condition of each injection site was recorded immediately after injection and observed at 24h, 48h and 72 h. Tissue response scores were performed according to the scoring system in the table below and the results of the test were recorded.

3. Hemolysis test

A sample is extracted at 37 +/-1 ℃ for 72 +/-2 hours, the extraction medium is normal saline, the normal saline is used as a negative control, and sterilized distilled water is used as a positive control. Diluted fresh rabbit blood was added to each, and the supernatant was subjected to determination of the respective Optical Density (OD) values with an ultraviolet spectrophotometer at 545 nm. The hemolysis rate was calculated as follows:

hemolysis rate (%) - (D)_t-D_nc)/(D_pc-D_nc)

In the formula, D_tThe absorbance of the test sample is obtained; d_ncAbsorbance of negative control; d_pcAbsorbance was positive control.

The hemolysis rate of the test sample is within 5 percent, which is in accordance with the regulation.

4. Pyrogen test

The sample is leached in a cell culture medium containing serum, and the leaching condition is 37 +/-1 ℃ and 72 +/-2 h, so as to obtain a test solution.

Within 15 minutes after the normal body temperature of the applicable rabbit 3 is measured, a specified dose of a sample solution which is warmed to about 38 ℃ is slowly injected from an ear vein, then the body temperature of the rabbit is measured 1 time and 6 times in total at intervals of 30 minutes according to the previous method, and the normal body temperature is subtracted from the highest one of the 6 body temperatures, so that the raising temperature of the body temperature of the rabbit is obtained.

In all 3 rabbits, the body temperature was raised below 0.6 ℃ and the sum of the body temperature rises of 3 rabbits was below 1.3 ℃. Judging that the pyrogen examination of the test sample meets the regulation.

5. Acute systemic toxicity test

The sample is extracted in a cell culture medium containing serum under the extraction conditions of 37 +/-1 ℃ and 72 +/-2 hours to obtain a test solution. 5 mice qualified for health quarantine are selected, and the test solution is injected into the tail vein/abdominal cavity, wherein the sample dosage is 50 ml/kg. The mice were observed for immediate response after injection and the weight gain status as well as general status, toxicity performance and number of dead animals were observed and recorded at 4h, 24h, 48h, 72h after administration for each group of animals. The weight is obviously increased, no death, no abnormal condition of general behavior examination and autopsy and no toxic reaction are caused, and the product is qualified.

6. Bone implant test

The test samples were as received, bone implanted for 1 week, 4 weeks, 12 weeks, and histological observations should be good. The sterilized samples were prepared into cylinders of 2cm diameter and 6cm length and filled into bone cavities drilled in rabbit femurs according to the bone implantation method specified in GB/T16886.6-1997.

The safety and biocompatibility tests were conducted on the above examples and comparative examples, respectively, and the results are shown in the following table 10:

TABLE 10 evaluation results of safety and biocompatibility

Repeated biological evaluation tests show that strict operation is performed every time, and the final result shows that the sample obtained by the process formula is safe, non-irritant, stable in performance and safe for clinical application.

Verification example 3: hemostasis test

Healthy adult common-grade New Zealand white rabbits are selected, and the weight of the rabbits is 2.5-3.0 Kg without limitation to male and female. Raising temperature: 20-26 ℃; relative humidity: 40-70%; illumination: 15-20 Lx; feeding complete granular rabbit feed, and freely drinking water.

1. Ilium hemostasis

The ilium cancellous bone has a large area, rich bone pore network structures and large pore diameters, is the site of the organism with the most bleeding from the skeletal bone wound surface, and has a representative hemostatic effect when the bone hemostatic composition is observed at the site.

Taking 5 experimental animals, carrying out ear intravenous injection anesthesia by using 2% pentobarbital sodium, unhairing and skin preparation, sterilizing an operation area, paving a sterile operation hole towel, respectively making bilateral external iliac crest incisions, stripping periosteum, leaking external iliac plates, drilling a wound surface with the depth of about 4mm by using a bone drill with the diameter of 2.5mm, and after the wound surface bleeds, smearing hemostasis by using the sample of the embodiment 2 of the invention to observe the hemostasis effect. After the sample is smeared on the wound surface, the sample can be firmly adhered to the bone wound surface, no bleeding exists around the wound surface, and the bleeding is stopped after the sample is smeared, so that the bleeding is quickly stopped. In addition, the surface bone part with the thickness of about 1mm of the external iliac plate is cut off by using bone forceps, the wound surface with the diameter of about 1cm of the surface of the iliac is exposed, the surface bleeding is rapid, the bleeding amount is large, the sample of the embodiment 7 of the invention is used for smearing hemostasis, and the hemostasis effect is observed. Because the area of the bone wound surface is large, the bone wound surface has slight bleeding after being coated with a layer of the sample, and has no bleeding after the sample coating amount is supplemented. The hemostasis time of both parts is not more than 5 min. The whole process is convenient to operate, the smearing is simple, and the hemostatic effect is good.

3 experimental animals were taken, wound surfaces of about 4mm depth were drilled on the external iliac plate in the same manner as above, and bleeding was stopped by applying the sample of comparative example 1 after bleeding on the wound surfaces, and the hemostatic effect was observed. After the sample of the comparative example 1 is coated on the wound surface, the sample can not be adhered to the bleeding bone wound surface and can not stop bleeding effectively. Meanwhile, the surface bone part with the thickness of about 1mm of the external iliac plate is cut off by using a rongeur by the same method, the surface of the iliac is exposed, the sample of the comparative example 1 is used for smearing for hemostasis, and the hemostasis effect is observed. After the sample of the comparative example 1 is coated on the wound surface, the sample can not be adhered to the bleeding and bleeding bone wound surface and can not stop bleeding effectively.

2. Skull hemostasis

The skull part has higher intracranial pressure, and if the skull part has bone defect and bleeds, the bleeding amount is larger because the pressure is too large, and the bleeding is more urgent and difficult to stop bleeding.

Taking 5 experimental animals, anesthetizing by ear intravenous injection with 2% pentobarbital sodium, depilating and preparing skin, sterilizing an operation area, paving a sterile operation hole towel, making an incision about 3cm long in the middle of the head, stripping periosteum, leaking parietal bones of the head, prying the parietal bones open irregular-shaped wound surfaces by using bone drills and rongeurs, after the wound surfaces bleed, smearing the sample of the embodiment 4 of the invention for hemostasis, and observing the hemostasis effect. After the sample is smeared on the wound surface, the sample can be firmly adhered to the bone wound surface, the sample is changed into red from white after being soaked by blood, trace amount of blood seeps around the sample, the amount of the blood seeps is not increased within five minutes, and the sample can stop bleeding quickly. For irregular bone wound surfaces, the composition is also convenient to operate and has a good hemostatic effect.

Taking 3 experimental animals, cutting irregular wound surfaces at parietal bones by the same method, smearing the sample of the comparative example 1 for hemostasis, and observing the hemostasis effect. After the sample of comparative example 1 is coated on the wound surface, the sample can not be adhered to the bone wound surface, and the bleeding is not effectively stopped.

Verification example 4: degradation test

The experimental animals are divided into 4 groups, 24 animals are anesthetized by 2% sodium pentobarbital through ear intravenous injection, depilated and prepared into skin, the operation area is sterilized, a sterile operation hole towel is laid, an incision is made at the femoral part, periosteum is stripped, the femoral is exposed, a hole is drilled on the bone intermittently at a low rotating speed, the diameter of a drill bit is 2mm, and the operation is carried out by fully irrigating with physiological saline to avoid local tissue overheating. 3 holes were drilled per femur, and each group implanted the samples of examples 3, 6, 7 and comparative example 2, respectively, into the left leg bone hole and the right leg bone hole of a new zealand rabbit within the group, respectively. After surgery, 2 animals were randomly selected and sacrificed at 1 week, 4 weeks, and 12 weeks, respectively, and the operative area was exposed in the same manner, and the healing condition and the remaining condition of the sample at the left leg implantation site of the new zealand rabbit were observed and compared with the right leg of the same rabbit. The femoral implant was cut out, muscle tissue was removed, fixed by formalin immersion, and subjected to pathological observation.

In the test, the bone holes of the examples 3, 6 and 7 disappear at 1 week, the growth conditions of the callus of the right leg and the bone holes implanted in the examples 3, 6 and 7 are not greatly different after 4 weeks, most of the bone holes are covered, the growth conditions of the callus of the left leg and the right leg are not different after 12 weeks, the callus covers all the bone holes, and the bone is well healed. The sample of comparative example 2 was clearly visible at the bone hole at 1 week, after 4 weeks the sample was invisible to the naked eye, about one quarter to one half of the bone hole was masked by callus, but the right leg callus of the same rabbit had mostly masked the bone hole. After 12 weeks, no macroscopic sample was present at the implantation site in comparative example 2, and the bone openings were mostly covered by callus, while the bone openings were completely covered by callus from the right leg of the same rabbit.

Claims

1. A bone wound hemostatic composition, which is characterized in that: the ionic salt is composed of a solvent matrix, a thickening agent, an excipient and an ionic salt, wherein the solvent matrix is polyhydroxy alcohol, and the ionic salt is one or more of inorganic calcium salt and inorganic iron salt;

the thickening agent is one or two of sodium carboxymethylcellulose, hydroxypropyl methylcellulose, hydroxyethyl cellulose, carrageenan and carbomer;

the excipient is one or more of sodium carboxymethyl starch, potato starch, soluble starch, sodium alginate, dextrin and dextrin derivatives;

the weight percentage of the solvent matrix in the bone wound hemostasis composition is 30-80%; the weight percentage of the thickening agent in the bone wound hemostasis composition is 0.5-10%; the weight percentage of the excipient in the bone wound hemostasis composition is 10-55 percent; the weight percentage of the ionic salt in the bone wound hemostasis composition is 0.05-5%.

2. The orthopedic wound hemostatic composition of claim 1, wherein: the solvent matrix is one or two of glycerol, polyethylene glycol-400, polyethylene glycol-300 and propylene glycol; the inorganic calcium salt is calcium sulfate, calcium chloride and calcium nitrate, the inorganic ferric salt is ferric chloride, ferric nitrate and ferric sulfate, and the ionic salt is selected from one or more of the inorganic calcium salts, or selected from one or more of the inorganic ferric salts, or selected from one or more of the inorganic calcium salts and the inorganic ferric salts.

3. The orthopedic wound hemostatic composition of claim 1, wherein: the solvent matrix is one or two of glycerol and polyethylene glycol-400.

4. The orthopedic wound hemostatic composition of claim 1, wherein: the sodium carboxymethyl cellulose is medical high-viscosity sodium carboxymethyl cellulose, medium-viscosity sodium carboxymethyl cellulose or low-viscosity sodium carboxymethyl cellulose; the hydroxypropyl methyl cellulose is hydroxypropyl methyl cellulose with the nominal viscosity of 6-15000 cps; the hydroxyethyl cellulose is hydroxyethyl cellulose with nominal viscosity of 250-400cps, 1500-2500cps, 3500-5500cps or 4500-6500 cps; the carrageenan is medical grade carrageenan; the carbomer is carbomer 910, carbomer 934, carbomer 940 or carbomer 941.

5. The orthopedic wound hemostatic composition of claim 4, wherein: the hydroxypropyl methyl cellulose is hydroxypropyl methyl cellulose with the nominal viscosity of 50-1500 cps.

6. The orthopedic wound hemostatic composition of claim 4, wherein: the hydroxyethyl cellulose is hydroxyethyl cellulose with the nominal viscosity of 1500-2500cps or 3500-5500 cps.

7. The orthopedic wound hemostatic composition of claim 4, wherein: the carbomer is carbomer 934 or carbomer 940.

8. The orthopedic wound hemostatic composition of claim 1, wherein: the weight percentage of the solvent matrix in the bone wound hemostasis composition is 50-70%; the weight percentage of the thickening agent in the bone wound hemostasis composition is 5-7%; the weight percentage of the excipient in the bone wound hemostasis composition is 20-45%; the weight percentage of the ionic salt in the bone wound hemostasis composition is 1-3%.

9. A process for the preparation of a bone wound haemostatic composition according to any of claims 1-8, characterised in that it comprises the steps of:

b. slowly adding the excipient into the viscous solvent matrix which is completely swelled under the conditions of stirring, blending or kneading, and then continuously stirring, blending or kneading until uniform paste or gum is formed;

c. slowly adding the ionic salt into the paste or the colloid under the conditions of stirring, blending or kneading, and continuously stirring, blending or kneading after the ionic salt is added until uniform paste is formed, namely the bone wound hemostatic composition.

10. The method of claim 9, wherein: also comprises the step of preparing the bone wound hemostatic composition into a sheet shape, a column shape or other shapes; or the bone wound hemostasis composition is prepared into paste or smearing preparation.

11. Use of a bone wound haemostatic composition according to any of claims 1-8 in the manufacture of a human or animal bone wound haemostatic product.

12. Use according to claim 11, characterized in that: the bone wound surface is formed by bone wounds caused by various reasons.