CN102085392A - Nano-apatite/collagen compound stent and preparation method and application thereof - Google Patents
Nano-apatite/collagen compound stent and preparation method and application thereof Download PDFInfo
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- CN102085392A CN102085392A CN2011100311810A CN201110031181A CN102085392A CN 102085392 A CN102085392 A CN 102085392A CN 2011100311810 A CN2011100311810 A CN 2011100311810A CN 201110031181 A CN201110031181 A CN 201110031181A CN 102085392 A CN102085392 A CN 102085392A
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Abstract
The invention discloses a nano-apatite/collagen compound stent and a preparation method and an application thereof. The method comprises the steps of firmly distributing nano-apatite in a collage sponge with high porosity, then shaping, freeze-drying and cross-linking for preparation. The nano-apatite/collagen compound stent prepared by the method can be applied as a repair material in orthopaedics and dentistry. The porosity of the nano-apatite/collagen compound stent prepared by the method can be above 98%, the size is distributed in the range of 5-200 mu m, and micropores are mutually communicated; nano-apatite in the nano-apatite/collagen compound stent is distributed in collagen fiber and less prone to shedding; and the original shape can be kept after being immersed in water, and the nano-apatite/collagen compound stent can be restored to the original shape after being extruded.
Description
Technical field
The invention belongs to technical field of biological material, specifically, relate to a kind of nanometer calcium microcosmic salt/collagen compound rest and preparation method thereof and application.
Background technology
Contingencies such as daily vehicle accident, industrial accident and some orthopaedic diseases all can cause the disappearance of skeleton.It is estimated that annual potential artificial bone of China and joint prosthesis replace case load about 3,000,000 examples.Nowadays, the potential market of global bone grafting material has reached 6.5 hundred million dollars.Have every year 4000000 routine patients to carry out the bone transplant operation approximately, employed graft materials 45% is to be synthetic material from body bone, 40% for homogeneous allogenic bone and decalcified bone matrix, 15%.
Bone is transplanted has become the graft that has the call that is only second to blood transfusion.The filling material of bone that has used at present mainly contains several big classes of metal, pottery and artificial-synthetic copolymer, rises biomimetic material recently again, and bionical filling material of bone also claims the nanometer filling material of bone, and such material generally is to be substrate with collagen, chitosan, agarose etc.Though the mechanical strength of such filling material of bone is lower, osteoinductive that it is good and biodegradable, absorbent properties are being well worth doing it aspect the skeleton defect repair.
Structure and function by the natural imitation bone prepare bone renovating material from bionic angle design.At present existing a spot of bionical bone product has entered market, but exists fillibility bad, and biocompatibility is high and to defectives such as osteoblastic inducibility are low.
US 5532217 has reported and a kind of the collagen protein spinning has been prepared into collagen fiber, obtains the mineralized collagen fiber through mineralising and be used for bone repair, but this product pore structure is bad, and the collagen fiber easily degraded rapidly in vivo that obtains by the collagen protein spinning; Chinese patent: 200610013009.1 have reported and a kind of the mineralized collagen extrusion modling have been obtained the method for filling material of bone, but the porosity of this product is lower, is unfavorable for the diffusion and the growth of osteocyte; Chinese patent: 200510107942.0 obtain filling material of bone with collagen protein with the lyophilization of nanometer hydroxyapatite mixing final vacuum, but hydroapatite particles and collagen protein adhesion in the product for preparing with this method are very weak, come off easily, cause biocompatibility poor.
Summary of the invention
Technical problem to be solved by this invention provides a kind of nanometer calcium microcosmic salt/collagen compound rest and preparation method thereof and application.The product of this method preparation is made of type i collagen with good biocompatibility and calcium microcosmic salt, and has good pore structure, and the diffusion of pair cell, adhesion and growth are all highly beneficial.In addition, this product has higher suppleness under hygrometric state, but can restore rapidly after laying down external force.
The present invention addresses the above problem the technical scheme that is adopted: the preparation method of nanometer calcium microcosmic salt/collagen compound rest comprises the steps:
Step 1: the mineralising of collagen fiber
(1) collagen fiber are dispersed in the saturated aqua calcis, stirring makes its abundant swelling obtain the collagen fiber dispersion liquid;
(2) phosphate solution of preparation 0.001～0.1M;
(3) the prepared phosphoric acid saline solution is added drop-wise in the collagen fiber dispersion liquid, is 9～13 at pH, temperature is to carry out mineralising under 0 ℃～40 ℃ the condition, obtain the mineralized collagen fiber after finishing mineralising;
Step 2: porous calcium phosphorus salt/collagen support preparation
(1) collagen protein is joined in the mineralized collagen fiber, stir under 0 ℃～10 ℃ environment, until the dope that obtains homogeneous, wherein collagen protein is 1:9～9:1 with mineralized collagen quality of fiber ratio;
(2) dope mixed well is injected mould, put into that pre-freeze with the vacuum freeze drier lyophilizing of the product after the pre-freeze, obtains sponge-like porous material more than 0.5 hour then under-20 ℃～-80 ℃ the environment;
Step 3: crosslinked
(1) sponge-like porous material that obtains was placed 0.005%～0.03% glutaraldehyde solution crosslinked 15～120 minutes;
(2) material that takes out after crosslinked cleans with distilled water;
(3) material after cleaning is put under-20 ℃～-80 ℃ the environment freezingly more than 0.5 hour, then with the vacuum freeze drier lyophilizing of the product after crosslinked, obtains nanometer calcium microcosmic salt-collagen compound rest.
Particularly, described phosphate comprises sodium ascorbyl phosphate, ammonium phosphate salt, potassium phosphate.
In (3) step of described step 1, count Ca:PO in molar ratio behind the adding phosphate
4 3-Be 1.2～2, the time of whole dropping process is 4～12 hours, and the mass ratio of calcium microcosmic salt and collagen fiber is 1:9～1:1 after the mineralising.
Should place under the mineralising condition ageing more than 12 hours after finishing mineralising, clean with distilled water at last, 0 ℃～10 ℃ preservations get final product.
The porosity of described nanometer calcium microcosmic salt/collagen compound rest reach more than 98% and pore size distribution in 5～200 μ m, intercommunication between the hole.
A kind of nanometer calcium microcosmic salt/collagen compound rest by method for preparing.
Nanometer calcium microcosmic salt/collagen the compound rest of method for preparing is in the application as orthopaedics and dental prosthetic material
The invention has the beneficial effects as follows:
1, the porosity of the prepared nanometer calcium microcosmic salt/collagen compound rest of the inventive method reach (record) more than 95% by the liquid-soaked method and distribution of sizes in 5～200 μ m, intercommunication between the micropore.
2, the nano-calcium-phosphor salt distribution in the resulting nanometer calcium microcosmic salt of the present invention/collagen compound rest is in the inside of collagen fiber, and the calcium ion in the calcium microcosmic salt can form compact complex, difficult drop-off with the carboxylate radical complexation in the collagen.
3, the resulting nanometer calcium microcosmic salt of the present invention/collagen compound rest can keep original form after immersion, and can be returned to original form after the extruding.
Description of drawings
Fig. 1 is nanometer hydroxyapatite, collagen scaffold material internal structural representation.
The specific embodiment
Below in conjunction with embodiment and accompanying drawing, the present invention is described in further detail, but embodiments of the present invention are not limited thereto.
The preparation method of present embodiment nanometer calcium microcosmic salt/collagen compound rest is for being distributed in the nanometer calcium microcosmic salt securely among the collagen sponge of high porosity, and molding then, lyophilizing, crosslinked and get specifically may further comprise the steps and process conditions:
Collagen fiber, collagen protein are laboratory self-control and conformance with standard.
1, the 2.0g collagen fiber is distributed in the saturated aqua calcis of 100ml, stirs and make the abundant swelling of collagen fiber.
2, with 46ml, the sodium radio-phosphate,P-32 solution of 0.03M is added drop-wise in the collagen fiber swelling thing with peristaltic pump, and rate of addition is 0.1ml/min, and whole mineralization process is finished at ambient temperature.
3, finish collagen fiber after the mineralising and be statically placed under the room temperature condition ageing 24 hours.
4, with the collagen fiber after the distilled water cleaning mineralising, be neutral until the liquid that washes out.
5, collagen protein is joined in the collagen fiber after the mineralising according to 30% of dry weight, and under cryogenic condition, stir 5h, obtain uniform pastel.
6, above-mentioned uniform mixture is injected the mould of 10cm * 10cm * 0.5cm, place-80 ℃ freezing 3 hours of cryogenic refrigerator, will freeze real sample vacuum freezing drying method drying then, obtain sponge-like porous material.
7, above-mentioned spongy material is soaked in 0.03% the glutaraldehyde solution crosslinked 1 hour.
8, the sample after crosslinked fully cleans with distilled water, and then vacuum lyophilization.
9, dried sample advanced packing, sterilization promptly can be used as tissue engineering bracket.
Picture such as Fig. 1 that the sample that obtains arrives by scanning electron microscopic observation, its porosity is very high, and the nano-calcium-phosphor salinity is distributed in collagen fiber inside (can prove that can't see the surface in the scanning electron microscope picture has mineral by Fig. 1, be distributed in collagen inside substantially).
By the nanometer calcium microcosmic salt/collagen compound rest of method for preparing, material calcium microcosmic salt content that records with the method for burning and the porosity measurement data that record with the toluene soaking method are as follows:
With a slice specification is the surface that the sample of 50mm * 50mm * 5m is put into normal saline, and sample will can permeate in 5 seconds fully, and relays resilience rapidly in the entry after the extruding of the sample after will soaking, and good resilience is arranged.
As mentioned above, just can realize the present invention preferably.
Claims (7)
1. the preparation method of nanometer calcium microcosmic salt/collagen compound rest is characterized in that, comprises the steps:
Step 1: the mineralising of collagen fiber
(1) collagen fiber are dispersed in the saturated aqua calcis, stirring makes its abundant swelling obtain the collagen fiber dispersion liquid;
(2) phosphate solution of preparation 0.001～0.1M;
(3) the prepared phosphoric acid saline solution is added drop-wise in the collagen fiber dispersion liquid, is 9～13 at pH, temperature is to carry out mineralising under 0 ℃～40 ℃ the condition, obtain the mineralized collagen fiber after finishing mineralising;
Step 2: porous calcium phosphorus salt/collagen support preparation
(1) collagen protein is joined in the mineralized collagen fiber, stir under 0 ℃～10 ℃ environment, until the dope that obtains homogeneous, wherein collagen protein is 1:9～9:1 with mineralized collagen quality of fiber ratio;
(2) dope mixed well is injected mould, put into that pre-freeze with the vacuum freeze drier lyophilizing of the product after the pre-freeze, obtains sponge-like porous material more than 0.5 hour then under-20 ℃～-80 ℃ the environment;
Step 3: crosslinked
(1) sponge-like porous material that obtains was placed 0.005%～0.03% glutaraldehyde solution crosslinked 15～120 minutes;
(2) material that takes out after crosslinked cleans with distilled water;
(3) material after cleaning is put under-20 ℃～-80 ℃ the environment freezingly more than 0.5 hour, then with the vacuum freeze drier lyophilizing of the product after crosslinked, obtains nanometer calcium microcosmic salt-collagen compound rest.
2. the preparation method of nanometer calcium microcosmic salt according to claim 1/collagen compound rest is characterized in that described phosphate comprises sodium ascorbyl phosphate, ammonium phosphate salt, potassium phosphate.
3. the preparation method of nanometer calcium microcosmic salt according to claim 1/collagen compound rest is characterized in that, in (3) step of described step 1, counts Ca:PO in molar ratio behind the adding phosphate
4 3-Be 1.2～2, the time of whole dropping process is 4～12 hours, and the mass ratio of calcium microcosmic salt and collagen fiber is 1:9～1:1 after the mineralising.
4. according to the preparation method of claim 1 or 3 described nanometer calcium microcosmic salt/collagen compound rests, it is characterized in that, should place under the mineralising condition ageing more than 12 hours after finishing mineralising, clean with distilled water at last, 0 ℃～10 ℃ preservations get final product.
5. according to the preparation method of claim 1 or 3 described nanometer calcium microcosmic salt/collagen compound rests, it is characterized in that, the porosity of described nanometer calcium microcosmic salt/collagen compound rest reach more than 98% and pore size distribution in 5～200 μ m, intercommunication between the hole.
6. nanometer calcium microcosmic salt/collagen compound rest by method for preparing.
7. the nanometer calcium of method for preparing microcosmic salt/collagen compound rest is in the application as orthopaedics and dental prosthetic material.
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103705976A (en) * | 2014-01-09 | 2014-04-09 | 山东省淡水渔业研究院 | Composite bone repair material and preparation method thereof |
| CN104096268A (en) * | 2014-06-19 | 2014-10-15 | 北京奥精医药科技有限公司 | Mineralized collagen artificial periosteum and preparation method thereof |
| CN104436308A (en) * | 2013-09-23 | 2015-03-25 | 杨述华 | Mineralized collagen-base supporting apparatus for femur head |
| CN105536061A (en) * | 2015-09-30 | 2016-05-04 | 王松灵 | Biological tooth root scaffold/cell/diaphragm complex containing bionic periodontal membrane, and tissue engineering construction method thereof |
| CN106512104A (en) * | 2016-10-31 | 2017-03-22 | 北京奥精医药科技有限公司 | Mineralized collagen-based alveolar bone restoration material and preparation method thereof |
| CN106943629A (en) * | 2017-05-10 | 2017-07-14 | 张展 | Carry safe energy hydroxyapatite enzymolysis bone collagen nano-artificial bone |
| CN115869467A (en) * | 2022-12-12 | 2023-03-31 | 上海典范医疗科技有限公司 | Preparation method of mineralized collagen for bone repair material |
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| JP2000350778A (en) * | 1999-04-20 | 2000-12-19 | L'oreal Sa | Aged skin equivalent, preparation method thereof and application thereof |
| CN1513806A (en) * | 2003-04-30 | 2004-07-21 | 东南大学 | Calcium phosphorus kind ceramic bone structure engineering porous support material |
| CN101081314A (en) * | 2007-06-27 | 2007-12-05 | 浙江大学 | Method for preparation of collagen / chitose supporting stand of composite hydroxylapatite |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2000350778A (en) * | 1999-04-20 | 2000-12-19 | L'oreal Sa | Aged skin equivalent, preparation method thereof and application thereof |
| CN1513806A (en) * | 2003-04-30 | 2004-07-21 | 东南大学 | Calcium phosphorus kind ceramic bone structure engineering porous support material |
| CN101081314A (en) * | 2007-06-27 | 2007-12-05 | 浙江大学 | Method for preparation of collagen / chitose supporting stand of composite hydroxylapatite |
Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104436308A (en) * | 2013-09-23 | 2015-03-25 | 杨述华 | Mineralized collagen-base supporting apparatus for femur head |
| CN104436308B (en) * | 2013-09-23 | 2017-06-27 | 杨述华 | Mineralized collagen base support device for head of femur |
| CN103705976A (en) * | 2014-01-09 | 2014-04-09 | 山东省淡水渔业研究院 | Composite bone repair material and preparation method thereof |
| CN104096268A (en) * | 2014-06-19 | 2014-10-15 | 北京奥精医药科技有限公司 | Mineralized collagen artificial periosteum and preparation method thereof |
| CN104096268B (en) * | 2014-06-19 | 2016-11-23 | 北京奥精医药科技有限公司 | A kind of mineralized collagen artificial periosteum and preparation method thereof |
| CN105536061A (en) * | 2015-09-30 | 2016-05-04 | 王松灵 | Biological tooth root scaffold/cell/diaphragm complex containing bionic periodontal membrane, and tissue engineering construction method thereof |
| WO2017054335A1 (en) * | 2015-09-30 | 2017-04-06 | 首都医科大学 | Biological root complex containing bionic periodontal membrane and preparation method thereof |
| CN106512104A (en) * | 2016-10-31 | 2017-03-22 | 北京奥精医药科技有限公司 | Mineralized collagen-based alveolar bone restoration material and preparation method thereof |
| CN106943629A (en) * | 2017-05-10 | 2017-07-14 | 张展 | Carry safe energy hydroxyapatite enzymolysis bone collagen nano-artificial bone |
| CN115869467A (en) * | 2022-12-12 | 2023-03-31 | 上海典范医疗科技有限公司 | Preparation method of mineralized collagen for bone repair material |
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