CN101954122A - Preparation method of natural bone repairing material with pre-plasticity - Google Patents
Preparation method of natural bone repairing material with pre-plasticity Download PDFInfo
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- CN101954122A CN101954122A CN2010102803069A CN201010280306A CN101954122A CN 101954122 A CN101954122 A CN 101954122A CN 2010102803069 A CN2010102803069 A CN 2010102803069A CN 201010280306 A CN201010280306 A CN 201010280306A CN 101954122 A CN101954122 A CN 101954122A
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Abstract
The invention relates to a preparation method of a natural bone repairing material with pre-plasticity. The natural bone repairing material is prepared by the steps of uniformly mixing forged bone particles and a bone collagen solution and freeze-drying at low temperature; protein and a growth factor for promoting bone formation are added into the bone collagen solution. The bone repairing material prepared by the method comprises natural inorganic components, the collagen component and the natural bone component are tightly combined, so that the stability of the material in a transplantation area is ensured and new bone can be rapidly formed. The prepared material can continuously release the active factor for bone formation for a long time, has certain elasticity and tenacity, can be cut and formed freely as required, thereby being convenient for clinic operations. Animal experiments verify that the bone repairing speed of the material is obviously higher than that of other artificial bone repairing materials. The material has simple preparation method and wide and cheap raw material resources, and the material is applicable to large-scale industrial production, can reduce medical cost and can relieve the economical burden of patients.
Description
Technical field
The invention belongs to the tissue engineering technical field of biomedical materials, be specifically related to a kind of preparation method with nature bone repair materials of pre-plasticity.
Background technology
Disease such as wound, bone tumor can cause each position bone in various degree damaged, and the bone transplanting is one of damaged main method of treatment bone, and the effect of autologous bone transplanting is best usually.But because limited, and can increase patient's misery and postoperative complication, therefore adopt some artificial bone grafting materials to be applied to clinical treatment from the source of body bone.For example macromolecule polymer, bioceramic, calcium phosphate bone cement, and metal material etc., and these materials all have different shortcomings, have limited their application clinically.
The macromolecule polymer can not adapt with the reconstruction speed of bone, and can produce the acid degradation product owing to too fast at the intravital degradation speed of people, causes inflammatory reaction, is unfavorable for the osseous tissue healing.
Metal material is the most traditional bone renovating material, with its superior mechanical performance, is extensive use of in the bone defect repair of big section.Its drawback also is conspicuous, because material itself is not degraded, is present in the body as foreign body for a long time, influences patient's quality of life; The metal ion that material dissolves goes out can cause the disorder of human body physiological function; And must carry out the individual character treatment, industrialization production on a large scale by the damaged clinical preparation sample of situation of patient.
Bioceramic class material mainly contains hydroxyapatite, tricalcium phosphate; This class material is to utilize self can provide to organize the necessary calcium ion of mineralising to stimulate the reconstruction of osseous tissue.Because differ greatly (people's bone inorganic constituents is the abundant weak crystalline hydroxy apatite of carbonate) of material composition and normal person's bone component, process of reconstruction that can not the interior normal bone of analogue body influences knitting; And this class material is graininess or bulk product, and granular material can only be filled into defect by loosely, and the instability of damaged site material can cause graft failure; Bulk product needs to cut into the shape of defect when clinical use, because material itself does not have elasticity, can not coincide well with plant bed, and the bulk material centre is not smooth because of the body fluid circulation, the possibility that forms downright bad chamber is arranged, along with the raising of clinical requirement, this class material will be eliminated gradually.
For graft materials can be coincide well with plant bed, bioceramic class material mixes the calcium phosphate bone cement material that forms with bone cement, can be moulding in clinical use by the shape of patient's bone defect, made things convenient for clinical use, have the transplanted sites material settling out, can with the fine identical advantage of plant bed.But such material does not change the composition of calcium phosphate ceramic, has also introduced bone cement, and its degraded can produce tart microenvironment, produces the non-bacterial inflammatory reaction, is unfavorable for the osseous tissue reconstruction; Experiment confirm, the bone reconstruction speed that such material participates in is not significantly improved.
The inventor discovers that ideal bone grafting material should be consistent with the natural human bone component, and degradation in vivo speed should be complementary with the bone reconstruction speed; And can have good hydrophilic at the defect Selfstabilizing, can adsorb peripheral blood and form blood clot (blood clot is the osteoplastic first step), accelerated bone forms; Can be moulding fast in clinical use by the true form of patient's defect, be convenient to operation; And induce new osteoplastic activated protein molecule.
Summary of the invention
At above the deficiencies in the prior art, the purpose of this invention is to provide a kind of preparation method with nature bone repair materials of pre-plasticity; Prepared bone renovating material is made up of natural component, the degradation speed of material can be complementary with the formation speed of new bone, and can induce new bone formation, improves skeletonization efficient; Prepared material structure loose can cut mouldingly as required arbitrarily in the use, can adsorb the blood of surrounding tissue after implanting damaged site, forms blood clot, quickens skeletonization; Its stability helps forming fast new bone.
Preparation method with pre-plasticity nature bone repair materials proposed by the invention is characterized in that, is with behind forging bone granule and the ossein solution mixing, is prepared from through low-temperature freeze drying; Be added with in the described ossein solution and promote osteoplastic albumen and somatomedin; Change the intensity of bone renovating material by the mass-energy density of adjusting osseous granules and ossein.Its concrete grammar may further comprise the steps:
Step 1, forging bone preparation of granules
1) remove organic matter: the spongy bone of animal femur or vertebra is removed surperficial soft tissue, and the broken bone piece that mechanical activation comminution becomes particle diameter 1～5mm size cleans with pure water; Diethyl ether solution concussion with 2 times of volumes of broken bone piece was cleaned 20 minutes, reuse concentration is that the pancreatin solution of 0.5～2g/L digested 1～3 day, clean back stirring and washing 6 hours in the 1M urea liquid with pure water, then ultrasonic Treatment 2～4 hours in the NaOH of 1M solution, changed NaOH solution in per 0.5～1 hour, pure water cleans to neutral, 37 ℃ of dryings 12 hours.
2) calcining and preparation osseous granules: dried broken bone piece is placed Muffle furnace, (calcining heat and time were being fired to white fully with the bone piece in 2～6 hours for calcining at constant temperature after being warmed up to 500～900 ℃, select calcining heat and time according to incinerating amount, the general low temperature calcination time is long, high-temperature time is short), naturally cooling back mechanical activation comminution is the granule of particle diameter 0.1～0.5mm, and is standby;
Step 2, preparation ossein slow release solution: ossein is become the ossein solution of 5～40mg/ml with the acetic acid solutions of 1% concentration, add bone morphogenetic protein 50～200 μ g/ml therein, platelet derived growth factor 5～20ng/ml;
Step 3, ossein and forging bone granule are compound: the pH value to 7.4 of adjusting ossein solution; 's 9～2: 1 ratio in the mass/volume of osseous granules and ossein solution than (g/ml), the forging bone granule for preparing is added in the ossein solution, abundant mixing, under 37 ℃ of conditions, place, after treating the ossein solution solidifies, put into-80 ℃ of environment pre-freeze half an hour,, obtain block nature bone repair materials again through cryogenic vacuum freeze drying bone collagen and forging bone composite.
The nature bone repair materials of the present invention's preparation is by the inorganic mineralising composition of nature bone, natural ossein composition and can promote osteoplastic activated protein to become to be grouped into; Natural mineralising composition wherein is to handle the back calcining by the animal spongy bone through physics and chemistry to form, and the natural ossein composition is to be obtained by animal long bone tissue extraction, adds commercial bone source property activated protein in the collagen.Collagen composition in the nature bone repair materials of the present invention's preparation is interconnected to form space network, and the natural inorganic osseous granules closely is wrapped in this stereoscopic graticule, and this space structure is given this material certain compression strength and spatial stability.The 3 D stereo web frame that collagen forms has bigger water absorption, presents sponge sample form after material is moisture, can be according to the true form of clinical patient bone defect area, moulding arbitrarily, make things convenient for the doctor to operate, also make material can be fully and plant bed integrate, help osseous tissue and rebuild.
The prepared bone renovating material of the present invention comes from the animal bone tissue, has the nature bone composition consistent with people's bone, and the degradation speed of material agrees with mutually with the speed that people's bone is rebuild; Tridimensional network and natural inorganic bone that ossein forms are combined closely, and have guaranteed the spatial stability of material, can Selfstabilizing after the implantation defect; The tridimensional network of ossein is given the certain elasticity of this material, toughness, can be moulding as required, make things convenient for clinical manipulation; Contain bone morphogenetic protein, two kinds of activated proteins with the effect of promotion bone formation of platelet derived growth factor in the prepared material, ossein provides the three-D space structure of the slow release osteogenic activity factor, can continue to be released into the bone active factor for a long time.
The inventive method is passed through the forging bone granule mass/volume different with the ossein solution Mechanical Characters of Composite Ground of control material recently, and greater than 5: 1 o'clock, material had higher mechanical strength at mass ratio, can support damaged site, character similar sponge in hydrophilic back can be cut mouldingly arbitrarily, has elasticity; Be lower than at 5: 1 o'clock at mass ratio, the similar ooze of material hydrophilic back character can directly be clogged to damaged site, and is easy to operate.
In the material of the present invention's preparation, collagen composition and natural inorganic bone component are combined closely, and have guaranteed that material in the stability of transplanting the zone, helps forming fast new bone.Confirm that by a large amount of zooperies the bone of the prepared material of the present invention is repaired speed obviously faster than other artificial bone renovating materials.
Preparation method of the present invention is simple, and raw material sources are extensive and cheap, is suitable for large-scale industrialization production, reduces medical treatment cost, alleviates patient's financial burden.
Accompanying drawing and explanation thereof
Accompanying drawing 1 is the white block nature bone repair materials outward appearance of 1 * 1 * 0.5cm specification in the photo for the cardinal principle photo of the nature bone repair materials of employing the inventive method preparation.Accompanying drawing 2 is the ultramicroscope micrograph of material partial sweep shown in Figure 1, and visible flaky ossein becomes branch to form the three-dimensional netted integral body that connects, and blocky natural inorganic bone is wrapped in wherein.Accompanying drawing 3 for the rat skull each side the round defect of 8mm diameter repair back 6 months microCT and observe photo; The left side is the blank group, promptly makes the 8mm diameter and does not carry out any way reparation after damaged; The right side is a material reparation group, and the nature bone repair materials that promptly adopts the present invention to prepare is repaired; The damaged all healings of the bone of visible material reparation group, and the blank group do not see tangible knitting.
The specific embodiment
Below in conjunction with example technical solution of the present invention is described in further detail.
Embodiment 1,
1. remove organic: get bulls bone epiphysis end spongy bone, behind the soft tissue on removal surface, be machined into the broken bone piece of particle diameter 1～5mm size, pure water stirring and washing 30 minutes; Diethyl ether solution concussion with 2 times of volumes of bone piece was cleaned 20 minutes, broken bone piece being inserted concentration is digestion 70h under the 0.5g/L pancreatin solution room temperature again, the pure water stirring and washing was placed in the 1M urea liquid stirring and washing in 10 minutes 6 hours, then ultrasonic waves for cleaning 2 hours in the NaOH of 1M solution, change solution once per half an hour, the pure water stirring and washing is to neutral, in 37 ℃ of drying baker 12 hours.
2. calcine and the preparation osseous granules: dried broken bone piece is put into crucible, in Muffle furnace, calcine; Calcining at constant temperature is 3 hours after being warmed up to 800 ℃, and temperature in the stove is reduced to room temperature, takes out forging bone, and mechanical activation comminution is the granule of particle diameter 0.1～0.25mm, and is standby.
3. prepare ossein solution: ossein is dissolved in the acetic acid solution of 1% concentration (V/V), is mixed with 10mg/ml ossein solution, in ossein solution, add bone morphogenetic protein 50 μ g/ml, platelet derived growth factor 10ng/ml.
4, ossein and forging bone granule are compound: adopt the pH value to 7.4 of the NaOH solution adjustment ossein solution of 1M, and in the forging bone granule adding ossein solution with preparation, abundant mixing.Osseous granules is 3: 1 with the mass/volume ratio of ossein solution, places 20 minutes at 37 ℃, treat the ossein solution solidifies after, put into-80 ℃ of cryogenic refrigerator pre-freezes; After half an hour, again through cryogenic vacuum freeze drying bone collagen and forging bone composite, behind the sterilization, promptly obtaining can be at the bone renovating material with pre-plasticity of clinical use.
The bone renovating material of the method preparation, hydrophilic after, character is like mud, can be used for the defect repair that causes by bone tumor, material directly is filled to damaged site, make material and plant bed fully identical by pushing, easy to operate, can absorb peripheral blood rapidly after material is implanted and form blood clot.
Embodiment 2,
1. remove organic: get bulls bone epiphysis end spongy bone, remove surperficial soft tissue after, be machined into the broken bone piece of particle diameter 1～3mm size, pure water cleaned 30 minutes; Diethyl ether solution concussion with 2 times of volumes of bone piece was cleaned 20 minutes, broken bone piece being inserted concentration is digestion 46h under the 1g/L pancreatin solution room temperature again, pure water cleans and to be placed in the 1M urea liquid stirring and washing in 10 minutes 6 hours, then ultrasonic waves for cleaning 3 hours in the NaOH of 1M solution, change solution once per half an hour, the pure water stirring and washing is to neutral, 37 ℃ of drying baker inner dryings 12 hours.
2. calcine and the preparation osseous granules: dried broken bone piece is put into crucible, in Muffle furnace, calcine; Calcining at constant temperature is 5 hours after being warmed up to 600 ℃, and temperature in the stove is reduced to room temperature naturally, takes out forging bone, and mechanical activation comminution is the granule of particle diameter 0.25～0.5mm, and is standby.
3. prepare ossein solution: ossein is dissolved in the acetic acid solution of 1% concentration (V/V), is mixed with 20mg/ml ossein solution, in ossein solution, add bone morphogenetic protein 100 μ g/ml, platelet derived growth factor 15ng/ml.
4, ossein and forging bone granule are compound: the pH value to 7.4 that adopts the NaOH solution adjustment ossein solution of 1M, the forging bone granule for preparing is added in the ossein solution, abundant mixing osseous granules and ossein solution, osseous granules is 7: 1 with the mass/volume ratio of ossein solution, under 37 ℃ of conditions, placed 15 minutes, after treating the ossein solution solidifies, put into-80 ℃ of cryogenic refrigerator pre-freezes; After half an hour, again through cryogenic vacuum freeze drying bone collagen and forging bone composite, behind the sterilization, promptly obtaining can be at the bone renovating material with pre-plasticity of clinical use.
The bone renovating material of the method preparation, hydrophilic after, character is like sponge, elasticity is better, has certain compression strength, the nest that can be used for having tooth pulled out is filled, and prunes material according to the shape of exodontia nest during use; The elasticity of material can make it fully contact with surrounding tissue, and the absorption peripheral blood forms blood clot, accelerated bone tissue reconstruction; Operation back 4～6 months, exodontia nest position forms a large amount of osseous tissues, at this moment, can implant implantation body in exodontia nest position, equips artificial tooth then.And bone absorption can take place in the exodontia nest of filling without material, and the implantation body that has a strong impact on the later stage implants.
Claims (2)
1. preparation method with pre-plasticity nature bone repair materials, adopting has the forging bone granule, it is characterized in that, be with behind forging bone granule and the ossein solution mixing, be prepared from through low-temperature freeze drying; Be added with in the described ossein solution and promote osteoplastic albumen and somatomedin; Change the intensity of bone renovating material by the mass ratio of adjusting osseous granules and ossein.
2. preparation method according to claim 1 is characterized in that, specifically may further comprise the steps:
Step 1, forging bone preparation of granules
1) remove organic matter: the spongy bone of animal femur or vertebra is removed surperficial soft tissue, and the broken bone piece that mechanical activation comminution becomes particle diameter 1～5mm size cleans with pure water; Diethyl ether solution concussion with 2 times of volumes of broken bone piece was cleaned 20 minutes, reuse concentration is that the pancreatin solution of 0.5～2g/L digested 1～3 day, clean back stirring and washing 6 hours in the 1M urea liquid with pure water, then ultrasonic Treatment 2～4 hours in the NaOH of 1M solution, changed NaOH solution in per 0.5～1 hour, pure water cleans to neutral, 37 ℃ of dryings 12 hours;
2) calcining and preparation osseous granules: dried broken bone piece is placed Muffle furnace, be warmed up to 500～900 ℃ after calcining at constant temperature 2～6 hours, pulverizing is the granule of particle diameter 0.1～0.5mm after the cooling naturally;
Step 2, preparation ossein slow release solution: ossein is become the ossein solution of 5～40mg/ml with the acetic acid solutions of 1% concentration, add bone morphogenetic protein 50～200 μ g/ml therein, platelet derived growth factor 5～20ng/ml;
Step 3, ossein and forging bone granule are compound: the pH value to 7.4 of adjusting ossein solution; It in the mass/volume ratio of osseous granules and ossein solution 9～2: 1 ratio, the forging bone granule for preparing is added in the ossein solution, abundant mixing, under 37 ℃ of conditions, place, after treating the ossein solution solidifies, put into-80 ℃ of environment pre-freeze half an hour,, obtain block nature bone repair materials again through cryogenic vacuum freeze drying bone collagen and forging bone composite.
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104874021A (en) * | 2015-06-11 | 2015-09-02 | 北京奥斯特赛医学科技有限公司 | Method for preparing forging bone for surgical implantation |
| CN106344214A (en) * | 2016-08-29 | 2017-01-25 | 中国人民解放军第三军医大学第三附属医院 | Method for preparing implantable bone grafts by de-organification and 3D printing of animal bones |
| CN106943623A (en) * | 2017-05-14 | 2017-07-14 | 常州力纯数码科技有限公司 | A kind of preparation method of promotion osteanagenesis natural bone repairing material |
| CN112316212A (en) * | 2019-08-31 | 2021-02-05 | 深圳市立心科学有限公司 | Artificial bone composite material capable of being freely shaped and preparation method thereof |
| CN114848913A (en) * | 2022-05-23 | 2022-08-05 | 海南宏正生物科技有限公司 | Preparation method of induced bone regeneration repair material, product and application thereof |
| WO2022199051A1 (en) * | 2021-03-23 | 2022-09-29 | 潍坊奥精医学研究有限公司 | Method for preparing bone repair material |
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| CN1325734A (en) * | 2001-06-29 | 2001-12-12 | 清华大学 | Process for preparing nm-phase calcium-phosphorus salt/collagen/high-molecular bone compounded porous material |
| CN101366973A (en) * | 2007-08-13 | 2009-02-18 | 北京大清生物技术有限公司 | Alveolar bone repairing material, preparation method and uses thereof |
| CN101564553A (en) * | 2009-06-05 | 2009-10-28 | 陕西瑞盛生物科技有限公司 | Humanization active forging bone and preparation method thereof |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104874021A (en) * | 2015-06-11 | 2015-09-02 | 北京奥斯特赛医学科技有限公司 | Method for preparing forging bone for surgical implantation |
| CN104874021B (en) * | 2015-06-11 | 2018-03-16 | 北京奥斯特赛医学科技有限公司 | A kind of preparation method of Srgery grafting forging bone |
| CN106344214A (en) * | 2016-08-29 | 2017-01-25 | 中国人民解放军第三军医大学第三附属医院 | Method for preparing implantable bone grafts by de-organification and 3D printing of animal bones |
| CN106943623A (en) * | 2017-05-14 | 2017-07-14 | 常州力纯数码科技有限公司 | A kind of preparation method of promotion osteanagenesis natural bone repairing material |
| CN112316212A (en) * | 2019-08-31 | 2021-02-05 | 深圳市立心科学有限公司 | Artificial bone composite material capable of being freely shaped and preparation method thereof |
| WO2022199051A1 (en) * | 2021-03-23 | 2022-09-29 | 潍坊奥精医学研究有限公司 | Method for preparing bone repair material |
| CN114848913A (en) * | 2022-05-23 | 2022-08-05 | 海南宏正生物科技有限公司 | Preparation method of induced bone regeneration repair material, product and application thereof |
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Application publication date: 20110126 |