CN103736149B - A kind of injectable type biologically active artificial bone material and preparation method and application - Google Patents
A kind of injectable type biologically active artificial bone material and preparation method and application Download PDFInfo
- Publication number
- CN103736149B CN103736149B CN201310729315.5A CN201310729315A CN103736149B CN 103736149 B CN103736149 B CN 103736149B CN 201310729315 A CN201310729315 A CN 201310729315A CN 103736149 B CN103736149 B CN 103736149B
- Authority
- CN
- China
- Prior art keywords
- artificial bone
- biologically active
- phosphate
- bone material
- preparation
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
Landscapes
- Materials For Medical Uses (AREA)
- Prostheses (AREA)
Abstract
The invention discloses a kind of injectable type biologically active artificial bone material and preparation method, belong to medical art.Injectable type biologically active artificial bone material in the present invention is in harmonious proportion preparation by solid phase and liquid phase two parts by solid-to-liquid ratio (0.8 ~ 1.2) g:1ml; Wherein said solid phase comprises phosphate mixt and carbodiimide; Described phosphate mixt and the mass ratio of condensing agent are 10:1; Described phosphate mixt is tetracalcium phosphate and the calcium hydrogen phosphate of mol ratio 1:2 mixing.Described liquid phase comprises water, chitosan and citric acid, Qi Zhongshui: chitosan: the mass ratio of citric acid is 100:(1 ~ 2): (4 ~ 5).The present invention adopts carbodiimide type condensation reagent to realize the covalent coupling of bioactive molecule and chitosan derivatives.Extensive use in artificial bone prepared by above-mentioned injectable type biologically active artificial bone material.
Description
Technical field
The invention belongs to medical art, be specifically related to a kind of injectable type biologically active artificial bone material and preparation method and application.
Background technology
Calcium phosphate bone cement adopts maximum injectable bone repair materials at present, but as tissue engineering bracket, the greatest drawback of calcium phosphate bone cement is that hole is little, be unfavorable for that cell is grown into, and curing rate is slower.After solidification, granular calcium microcosmic salt easily runs off, is shifted, and cured strength is low.Another kind of self-curing bone cement PMMA, there is solidification process heat release, biologically inert, poor with autologous synosteosis, the defects such as residual monomer is harmful, it is doubted in clinical use.
The research and development of current injectable bone tissue engineering bracket material focuses mostly in cartilage, less in the research and development of bone tissue engineer.Injectable tissue engineering timbering material few in number is the biomaterials such as injectable calcium alginate, fibrin gel.Mostly be gel after being injected in vivo, being easy to run off, and not possessing certain mechanical strength, for the bone defect healing requiring some strength, is obviously unfavorable.But be fluidised form before the injection of injectable tissue engineering bracket material, be easy to load and the composite growth factor of seed cell.According to the free plastotype solidification of Cranial defect shape after injection, simple to operate, the intensity after solidification can meet or exceed autologous spongiosa bone, is convenient to Clinical practice.Therefore the research of new injectable tissue engineering bracket is by playing the advantage of tissue engineering technique and Minimally Invasive Surgical Technology, is extensive patients service better.
Summary of the invention
For defect and the deficiency of prior art, primary and foremost purpose of the present invention is to provide a kind of injectable type biologically active artificial bone material.
Another object of the present invention is to the preparation method that above-mentioned injectable type biologically active artificial bone material is provided.
Another object of the present invention is the application providing above-mentioned injectable type biologically active artificial bone material.
Object of the present invention is achieved through the following technical solutions: a kind of injectable type biologically active artificial bone material, is in harmonious proportion preparation by solid phase and liquid phase two parts by solid-to-liquid ratio (0.8 ~ 1.2) g:1ml;
Described solid phase comprises phosphate mixt and condensing agent; Described phosphate mixt and the mass ratio of condensing agent are preferably 10:1;
Described liquid phase comprises water, chitosan and citric acid, Qi Zhongshui: chitosan: the mass ratio of citric acid is 100:(1 ~ 2): (4 ~ 5);
Described phosphate mixt is tetracalcium phosphate and the calcium hydrogen phosphate of mol ratio 1:2 mixing;
Described condensing agent is preferably carbodiimide;
The preparation method of injectable type biologically active artificial bone material of the present invention, comprises the following steps:
(1) mixed according to mol ratio 1:2 with calcium hydrogen phosphate by tetracalcium phosphate, in anhydrous ethanol medium, ball milling mixing, dries at 80 DEG C, obtained phosphate mixt;
(2) phosphate mixt step (1) obtained and carbodiimide in mass ratio 10:1 are mixed homogeneously, obtained solid phase;
(3) by chitosan, citric acid and water according to mass ratio (1 ~ 2): (4 ~ 5): the ratio of 100 mixes, and fully stirs, obtained liquid phase;
(4) liquid phase prepare in the solid phase of preparation in step (2) and step (3) is pressed solid-to-liquid ratio (0.8 ~ 1.2) g:1ml to be in harmonious proportion and to prepare, obtain injectable type biologically active artificial bone material.
The deacetylation of the chitosan described in step (3) is 70 ~ 100%, and weight average molecular weight is 2000 ~ 1,000,000.
Above-mentioned injectable type biologically active artificial bone material is applied preparing in artificial bone.
The present invention adopts carbodiimide type condensation reagent to realize the covalent coupling of bioactive molecule and chitosan derivatives.
The present invention has following advantage and effect relative to prior art:
1. the present invention adopts tetracalcium phosphate and calcium hydrogen phosphate to be mixed with the compound prescription of the synthos of acquisition according to mol ratio 1:2, makes material system have suitable curing rate.
2. adopt biodegradable cross-linker carbodiimide in the present invention, covalent cross-linking chitosan; Adopt natural product hydrogen bond crosslinks chitosan, make the polymer frame shelf structure of formation better " fixing " calcium microcosmic salt, prevent body fluid erosion.
Detailed description of the invention
Below in conjunction with embodiment, the present invention is described in further detail, but embodiments of the present invention are not limited thereto.
Embodiment 1
(1) mixed according to mol ratio 1:2 with calcium hydrogen phosphate by tetracalcium phosphate, in anhydrous ethanol medium, ball milling mixing, dries at 80 DEG C, obtained phosphate mixt;
(2) phosphate mixt step (1) obtained and carbodiimide in mass ratio 10:1 are mixed homogeneously, obtained solid phase;
(3) chitosan, citric acid and the water ratio according to mass ratio 1:4:100 is mixed, and fully stir, obtained liquid phase;
(4) liquid phase prepare in the solid phase of preparation in step (2) and step (3) is pressed solid-to-liquid ratio 0.8g:1ml to be in harmonious proportion and to prepare, obtain injectable type biologically active artificial bone material.
Embodiment 2
(1) mixed according to mol ratio 1:2 with calcium hydrogen phosphate by tetracalcium phosphate, in anhydrous ethanol medium, ball milling mixing, dries at 80 DEG C, obtained phosphate mixt;
(2) phosphate mixt step (1) obtained and carbodiimide in mass ratio 10:1 are mixed homogeneously, obtained solid phase;
(3) chitosan, citric acid and the water ratio according to mass ratio 2:5:100 is mixed, and fully stir, obtained liquid phase;
(4) liquid phase prepare in the solid phase of preparation in step (2) and step (3) is pressed solid-to-liquid ratio 1g:1ml to be in harmonious proportion and to prepare, obtain injectable type biologically active artificial bone material.
Embodiment 3
(1) mixed according to mol ratio 1:2 with calcium hydrogen phosphate by tetracalcium phosphate, in anhydrous ethanol medium, ball milling mixing, dries at 80 DEG C, obtained phosphate mixt;
(2) phosphate mixt step (1) obtained and carbodiimide in mass ratio 10:1 are mixed homogeneously, obtained solid phase;
(3) chitosan, citric acid and the water ratio according to mass ratio 1:5:100 is mixed, and fully stir, obtained liquid phase;
(4) liquid phase prepare in the solid phase of preparation in step (2) and step (3) is pressed solid-to-liquid ratio 1.2g:1ml to be in harmonious proportion and to prepare, obtain injectable type biologically active artificial bone material.
The extraction of embodiment 4 human umbilical cord mesenchymal stem cells, cultivation and induction
(1) acquisition of healthy umbilical cord: entrust No.1 Hospital Attached to Jinan Univ.'s department of obstetrics and gynecology to set up puerpera's health check-up health account, obtain after I and family members agree to and sign contract, the puerpera of planned production is shifted to an earlier date half a year and regularly provided and comprises routine blood test, biochemistry, the infectious diseases such as HIV, syphilis, each Hepatitis virus, the detections such as tumor markers, the puerpera that screening is healthy, obtains umbilical cord its point of puerperium, umbilical cord and neonate are filed, and set up donor follow-up system.
(2) extraction of stem cell: peel off the arteriovenous on umbilical cord, shred after wash buffer, the collagenase II, the mass fraction that are 0.1% through mass fraction after elder generation are the trypsinization 10 ~ 20min of 0.25%, collect suspension cell, be inoculated in the plastic culture bottle containing F-12 complete culture solution, be placed in 37 DEG C, volume fraction 5%CO
2incubator is cultivated, and regularly changes liquid.
(3) cultivation in bioreactor: when observation of cell 80% merges, mass fraction 0.25% trypsinization is rinsed.The Cytodex-3 type microcarrier of the cell and prehydration of getting some is mixed in proportion in F-12 complete medium, be placed in bioreactor, initial volume of culture accounts for the half of reactor volume, and after cell attachment, volume of culture is increased to reactor volume gradually.Pulsed power increases with volume of culture, until constant.Thereafter regularly change liquid, add microcarrier implementation continuous culture.Period sampling stem cell labeling quality testing survey.
(4) osteogenic induction of stem cell: after the sampling cell majority observed on microcarrier merges, use the F-12 culture fluid of the osteogenic induction containing dexamethasone 1 × 10-8mol/L, sodium β-glycerophosphate 10mmol/L, vitamin C 50ug/ml instead, after continuous culture 8-10 days, obtain osteoblast, can use with Material cladding.Period sampling is done osteoblast marker quality testing and is surveyed.
Embodiment 5 animal experiment
Get 3 ~ 4 monthly age new zealand rabbits of the same race, Cranial defect model is made at proximal ends of tibia, defect is 10 × 5 × 5mm about, to bone cortex, deeply reach medullary cavity, do not rinse, adding embodiment 4 before the injectable type biologically active artificial bone material cured prepared in embodiment 1 induces the osteoblast of preparation to inject in (both ratios are mass ratio 10:1) defect in the lump, material does not exceed normal bone cortex, sew up wound, carries out animal feeding; Within 1,2,4,6,8,12,18,24 months, carry out x-ray respectively at Post operation, ECT detects and bone densitometry, and histology, pathology detect, bone component detects.What found that the proximal ends of tibia of defect in postoperative 24 months obtains recovery, and the problems such as distortion do not appear in artificial bone.
Embodiment 6 animal experiment
Get 3 ~ 4 monthly age new zealand rabbits of the same race, set up rabbit lumbar intervertebral Fusion Model, take out disc tissue, strike off upper and lower vertebral plate, with the micro-oozing of blood of sclerotin for degree, add embodiment 4 before the injectable type biologically active artificial bone material cured prepared in embodiment 1 and induce the osteoblast of preparation to inject (both ratios are mass ratio 10:1) interbody space in the lump, make material constraints in gap, until material solidify completely hard after sew up wound, carry out animal feeding; Within 1,2,4,6,8,12,18,24 months, carry out x-ray respectively at Post operation, ECT detects and bone densitometry, and histology, pathology detect, bone component detects.Found that in postoperative 24 months, rabbit lumbar intervertebral merges.
Above-described embodiment is the present invention's preferably embodiment; but embodiments of the present invention are not restricted to the described embodiments; change, the modification done under other any does not deviate from spirit of the present invention and principle, substitute, combine, simplify; all should be the substitute mode of equivalence, be included within protection scope of the present invention.
Claims (4)
1. an injectable type biologically active artificial bone material, is characterized in that being in harmonious proportion preparation by solid phase and liquid phase two parts by solid-to-liquid ratio (0.8 ~ 1.2) g:1ml;
Described solid phase comprises phosphate mixt and condensing agent; Described phosphate mixt and the mass ratio of condensing agent are 10:1;
Described liquid phase comprises water, chitosan and citric acid, Qi Zhongshui: chitosan: the mass ratio of citric acid is 100:(1 ~ 2): (4 ~ 5);
Described phosphate mixt is tetracalcium phosphate and the calcium hydrogen phosphate of mol ratio 1:2 mixing;
Described condensing agent is carbodiimide.
2. the preparation method of injectable type biologically active artificial bone material according to claim 1, is characterized in that comprising the following steps:
(1) mixed according to mol ratio 1:2 with calcium hydrogen phosphate by tetracalcium phosphate, in anhydrous ethanol medium, ball milling mixing, dries at 80 DEG C, obtained phosphate mixt;
(2) phosphate mixt step (1) obtained and carbodiimide in mass ratio 10:1 are mixed homogeneously, obtained solid phase;
(3) by chitosan, citric acid and water according to mass ratio (1 ~ 2): (4 ~ 5): the ratio of 100 mixes, and fully stirs, obtained liquid phase;
(4) liquid phase prepare in the solid phase of preparation in step (2) and step (3) is pressed solid-to-liquid ratio (0.8 ~ 1.2) g:1ml to be in harmonious proportion and to prepare, obtain injectable type biologically active artificial bone material.
3. the preparation method of injectable type biologically active artificial bone material according to claim 2, is characterized in that: the deacetylation of the chitosan described in step (3) is 70 ~ 100%, and weight average molecular weight is 2000 ~ 1,000,000.
4. injectable type biologically active artificial bone material according to claim 1 is applied preparing in artificial bone.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310729315.5A CN103736149B (en) | 2013-12-25 | 2013-12-25 | A kind of injectable type biologically active artificial bone material and preparation method and application |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310729315.5A CN103736149B (en) | 2013-12-25 | 2013-12-25 | A kind of injectable type biologically active artificial bone material and preparation method and application |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103736149A CN103736149A (en) | 2014-04-23 |
| CN103736149B true CN103736149B (en) | 2015-12-30 |
Family
ID=50493261
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310729315.5A Expired - Fee Related CN103736149B (en) | 2013-12-25 | 2013-12-25 | A kind of injectable type biologically active artificial bone material and preparation method and application |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103736149B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106512086B (en) * | 2016-11-11 | 2019-09-17 | 江西理工大学 | A kind of doped yttrium calcium phosphate bone cement and preparation method thereof |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP1803442A1 (en) * | 2005-10-28 | 2007-07-04 | Zimmer Inc. | Mineralized hydrogels and methods of making and using mineralized hydrogels |
| CN101057979A (en) * | 2007-04-03 | 2007-10-24 | 暨南大学 | Injectable self-curable calcium phosphate bone tissue repairing material and its preparation method and application |
| CN101496909A (en) * | 2008-02-01 | 2009-08-05 | 华东理工大学 | Polysaccharide/calcium orthophosphate composite bone cement and preparation method thereof |
| CN101716379A (en) * | 2009-12-25 | 2010-06-02 | 武汉嘉特利佰联创科技有限公司 | Double-solidified composite calcium phosphate cement (CPC) and preparation method as well as application thereof |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2009029734A2 (en) * | 2007-08-28 | 2009-03-05 | Pioneer Surgical Technology, Inc. | Cement products and methods of making and using the same |
-
2013
- 2013-12-25 CN CN201310729315.5A patent/CN103736149B/en not_active Expired - Fee Related
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP1803442A1 (en) * | 2005-10-28 | 2007-07-04 | Zimmer Inc. | Mineralized hydrogels and methods of making and using mineralized hydrogels |
| CN101057979A (en) * | 2007-04-03 | 2007-10-24 | 暨南大学 | Injectable self-curable calcium phosphate bone tissue repairing material and its preparation method and application |
| CN101496909A (en) * | 2008-02-01 | 2009-08-05 | 华东理工大学 | Polysaccharide/calcium orthophosphate composite bone cement and preparation method thereof |
| CN101716379A (en) * | 2009-12-25 | 2010-06-02 | 武汉嘉特利佰联创科技有限公司 | Double-solidified composite calcium phosphate cement (CPC) and preparation method as well as application thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| CN103736149A (en) | 2014-04-23 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Sims et al. | Tissue engineered neocartilage using plasma derived polymer substrates and chondrocytes | |
| Zhao et al. | Osteogenic media and rhBMP-2-induced differentiation of umbilical cord mesenchymal stem cells encapsulated in alginate microbeads and integrated in an injectable calcium phosphate-chitosan fibrous scaffold | |
| Bueno et al. | Cell-free and cell-based approaches for bone regeneration | |
| CN101478934B (en) | Bioengineered intervertebral discs and methods for their preparation | |
| Xu et al. | Umbilical cord stem cell seeding on fast-resorbable calcium phosphate bone cement | |
| CN104353124B (en) | A kind of 3D of composite magnetic nano material prints porous metals support and preparation method thereof | |
| CN106061438A (en) | Tissue grafts and methods of making and using the same | |
| Liu et al. | Fast-degradable microbeads encapsulating human umbilical cord stem cells in alginate for muscle tissue engineering | |
| CN108939151B (en) | Application of the nanoporous micro rack in regeneration and restoration | |
| CN104906637A (en) | Injectable-porous-drug loaded polymethyl methacrylate-based composite scaffold bone transplant material and preparation method thereof | |
| CN105944144A (en) | Bone tissue repair material based on shape memory composite material as well as preparation method and application method of bone tissue repair structure | |
| CN100536935C (en) | Composite porous calcium phosphate bone cement and method for making same | |
| CN109320970A (en) | A kind of temperature-sensitive hydrogel and the preparation method and application thereof for repair of cartilage | |
| CN109152864A (en) | A method of being used to prepare three-dimensional cartilage organoid block | |
| CN110947031B (en) | Bone tissue engineering scaffold material with high biological activity and preparation method and application thereof | |
| Gao et al. | Sr-HA-graft-poly (γ-benzyl-l-glutamate) nanocomposite microcarriers: controllable Sr2+ release for accelerating osteogenenisis and bony nonunion repair | |
| Chen et al. | Multi-level customized 3D printing for autogenous implants in skull tissue engineering | |
| Macías et al. | Cutting edge endogenous promoting and exogenous driven strategies for bone regeneration | |
| Li et al. | Collagen-based bioinks for regenerative medicine: Fabrication, application and prospective | |
| CN101993853B (en) | Injection type vascularized adipose tissue and construction method thereof | |
| CN102100925A (en) | Preparation method of novel injectable polypeptide hydrogel | |
| Hara et al. | Efficacy of treating segmental bone defects through endochondral ossification: 3D printed designs and bone metabolic activities | |
| Liao et al. | Injectable adipose-derived stem cells-embedded alginate-gelatin microspheres prepared by electrospray for cartilage tissue regeneration | |
| CN109182249B (en) | Preparation method of scaffold material for cell transplantation for in vivo repair | |
| CN103736149B (en) | A kind of injectable type biologically active artificial bone material and preparation method and application |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20151230 Termination date: 20181225 |